Multi-part receptors and signaling complexes

Abstract

The present disclosure provides adoptive T-cell therapies having an improved architecture for targeting antigens and recruiting multimeric immune signaling complexes for treating, preventing, or ameliorating at least one symptom of cancer, infectious diseases, autoimmune diseases, inflammatory diseases, and immunodeficiency or conditions associated therewith.

Description

[0001] Cross-reference to related applications

[0002] This application claims the benefit of U.S. Provisional Application No. 63 / 329,003, filed on April 8, 2022, under 35 U.S.C. §119(e), which is incorporated herein by reference in its entirety. Technical Field

[0003] The present disclosure relates to improved adoptive cell therapy. More specifically, the present disclosure relates to improved immune receptor signaling molecules, cells, and methods of using the same for regulating spatial and / or temporal control of immune receptor signaling initiation and downstream responses during adoptive immunotherapy. Background Art

[0004] The global cancer burden doubled between 1975 and 2000. Cancer is the second leading cause of morbidity and mortality worldwide, with approximately 14.1 million new cases and 8.2 million cancer-related deaths in 2012. The number of new cancer cases is expected to increase to 22 million over the next two decades.

[0005] While chemotherapy and subsequently biologics have been the standard of care for decades, adoptive cell therapy is emerging as a powerful paradigm for delivering complex biological signals to treat cancer. Compared to small molecules and biologic drug compositions, adoptive cell therapy has the potential to perform unique therapeutic tasks due to its extremely large number of sensory and response programs and increasingly well-defined genetic control mechanisms. To realize this therapeutic value, cells need to be equipped with mechanisms for sensing and integrating chemical and / or biological information associated with the local physiological environment.

[0006] In recent years, assembling cells with chimeric antigen receptors (CARs) or transgenic T cell receptors (TCRs) has been shown to be an effective way to target immune cells to specific antigens (e.g., tumor antigens), stimulate T cells to activate signal transduction, and ultimately attack and kill antigen-related cells (e.g., cancer cells). Despite these successes, there are still fundamental differences and limitations between the two architectures, including i) sensitivity, ii) antigen recognition, iii) antigen-independent signaling activity, and iv) lack of adjustability. Therefore, there is still a need for improved targeting and signaling mechanisms that more effectively sense and respond to target antigens and related cells. Summary of the invention

[0007] The present disclosure generally relates in part to engineered immune receptor complexes that can both recognize target antigens and recruit and activate natural or transgenic immune receptor signaling complexes (e.g., T cell receptor (TCR) signaling complexes), polynucleotides and polypeptides encoding the same, compositions thereof, and methods of making and using the same to treat conditions or diseases (e.g., cancer or autoimmunity).

[0008] In one aspect, disclosed herein is a non-natural cell comprising a signaling component including a first multimerization domain and an actuator domain; and a targeting component comprising an extracellular domain, a second multimerization domain, and a transmembrane domain.

[0009] In one embodiment, the second multimerization domain and the transmembrane domain are separated by a hinge domain. In some embodiments, the hinge domain is selected from the group consisting of a CD4 hinge, a CD8 hinge, a CD28 hinge, an IgG4 hinge, and any fragment or variant or combination thereof. In some embodiments, the hinge domain is a CD4 hinge. In some embodiments, the CD4 hinge comprises an amino acid sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO: 41. In one embodiment, the CD4 hinge comprises an amino acid sequence as shown in SEQ ID NO: 41. In some embodiments, the hinge domain is a CD28 hinge. In some embodiments, the CD28 hinge comprises an amino acid sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO: 42. In one embodiment, the CD28 hinge comprises an amino acid sequence as shown in SEQ ID NO: 42. In some embodiments, the hinge domain is a CD8 hinge. In some embodiments, the CD8 hinge comprises an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98%, or 99% identical to SEQ ID NO: 44. In one embodiment, the CD8 hinge comprises an amino acid sequence as shown in SEQ ID NO: 44. In some embodiments, the hinge domain is an IgG4 hinge. In some embodiments, the IgG4 hinge comprises an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98%, or 99% identical to SEQ ID NO: 43. In one embodiment, the IgG4 hinge comprises an amino acid sequence as shown in SEQ ID NO: 43.

[0010] In some embodiments, the actuator domain is a CD3 polypeptide, an FcεR1γ polypeptide, an Igα / CD79a polypeptide, an Igβ / CD79b polypeptide, a DAP10 polypeptide, or a DAP12 polypeptide. In some embodiments, the CD3 polypeptide is a CD3ε (CD3epsilon) polypeptide or a variant thereof, a CD3γ (CD3gamma) or a variant thereof, or a CD3δ (CD3 delta) or a variant thereof. In some embodiments, the actuator domain is a CD3ε polypeptide or a variant thereof. In some embodiments, the actuator domain is a CD3ε polypeptide comprising an amino acid sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO: 32. In one embodiment, the actuator domain is a CD3ε polypeptide comprising an amino acid sequence as shown in SEQ ID NO: 32. In some embodiments, the actuator domain is a CD3γ polypeptide or a variant thereof. In some embodiments, the actuator domain is a CD3γ polypeptide comprising an amino acid sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO: 33. In one embodiment, the actuator domain is a CD3γ polypeptide comprising an amino acid sequence as shown in SEQ ID NO: 33. In some embodiments, the actuator domain is a CD3δ polypeptide or a variant thereof. In some embodiments, the actuator domain is a CD3δ polypeptide comprising an amino acid sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO: 34. In one embodiment, the actuator domain is a CD3δ polypeptide comprising an amino acid sequence as shown in SEQ ID NO: 34. In some embodiments, the actuator domain is an FcεR1γ polypeptide or a variant thereof. In some embodiments, the actuator domain is an FcεR1γ polypeptide comprising an amino acid sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO: 35. In one embodiment, the actuator domain is an FcεR1γ polypeptide comprising an amino acid sequence as shown in SEQ ID NO: 35. In some embodiments, the actuator domain is an FcεR1γ polypeptide comprising an amino acid sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO: 36. In one embodiment, the actuator domain is an FcεR1γ polypeptide comprising an amino acid sequence as shown in SEQ ID NO: 36. In some embodiments, the actuator domain is an Igα / CD79a polypeptide or a variant thereof.In some embodiments, the actuator domain is an Igα / CD79a polypeptide comprising an amino acid sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO: 37. In one embodiment, the actuator domain is an Igα / CD79a polypeptide comprising an amino acid sequence as shown in SEQ ID NO: 37. In some embodiments, the actuator domain is an Igβ / CD79b polypeptide or a variant thereof. In some embodiments, the actuator domain is an Igβ / CD79b polypeptide comprising an amino acid sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO: 38. In one embodiment, the actuator domain is an Igβ / CD79b polypeptide comprising an amino acid sequence as shown in SEQ ID NO: 38. In some embodiments, the actuator domain is a DAP10 polypeptide or a variant thereof. In some embodiments, the actuator domain is a DAP10 polypeptide comprising an amino acid sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO:39. In one embodiment, the actuator domain is a DAP10 polypeptide comprising an amino acid sequence as shown in SEQ ID NO:39. In some embodiments, the actuator domain is a DAP12 polypeptide or a variant thereof. In some embodiments, the actuator domain is a DAP12 polypeptide comprising an amino acid sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO:40. In one embodiment, the actuator domain is a DAP12 polypeptide comprising an amino acid sequence as shown in SEQ ID NO:40. In one embodiment, the actuator domain comprises both an extracellular portion and an intracellular portion.

[0011] In one embodiment, when the signaling component and the targeting component are expressed, the first multimerization domain and the second multimerization domain are located extracellularly. In one embodiment, the first multimerization domain is different from the second multimerization domain. In some embodiments, the multimerization domains of the signaling component and the targeting component are associated with a bridging factor selected from the group consisting of: rapamycin or its rapamycin analogs, gibberellin or its derivatives, abscisic acid (ABA) or its derivatives, methotrexate or its derivatives, cyclosporin A or its derivatives, FK506 / cyclosporin A or its derivatives, trimethoprim (Tmp) synthetic ligand (SLF) or its derivatives for FK506 binding protein (FKBP), wherein the bridging factor promotes the formation of a polypeptide complex, wherein the bridging factor is associated with the multimerization domains of the signaling component and the targeting component and is disposed between the multimerization domains. In some embodiments, the first multimerization domain and the second multimerization domain are a pair selected from the group consisting of: FK506 binding protein 1A (FKBP12) and FKBP12-rapamycin binding (FRB), FKBP12 and calcineurin, FKBP and cyclophilin A or any other member of the peptidyl-prolyl cis-trans isomerase (PPIase) family, FKBP and dihydrofolate reductase (DHFR), calcineurin and cyclophilin A or any other member of the peptidyl-prolyl cis-trans isomerase (PPIase) family, and PYR1-like 1 (PYL1) and abscisic acid insensitive 1 (ABI1). In one embodiment, the first multimerization domain comprises a first FRB polypeptide or a variant thereof, and the second multimerization domain comprises a first FKBP12 polypeptide or a variant thereof. In one embodiment, the first multimerization domain comprises a first FKBP12 polypeptide or a variant thereof, and the second multimerization domain comprises a first FRB polypeptide or a variant thereof. In one embodiment, the FRB polypeptide is a FRB T2098L variant. In some embodiments, the FRB polypeptide comprises the amino acid sequence as shown in SEQ ID NO: 1 or SEQ ID NO: 2. In some embodiments, the FKBP12 polypeptide comprises the amino acid sequence as shown in SEQ ID NO: 3 or SEQ ID NO: 4.

[0012] In some embodiments, the bridging factor is AP1903, AP20187, AP21967 (also known as C16-(S)-7-methylindorapamycin), everolimus, novolimus, pimecrolimus, ridaforolimus, sirolimus, tacrolimus, temsirolimus, umirolimus, zotarolimus or BPC015. In some embodiments, the first multimerization domain and the second multimerization domain are a pair of antibody-derived heterodimerization domains.

[0013] In some embodiments, the first multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO:5, or comprises the amino acid sequence as shown in SEQ ID NO:5; and the second multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO:6, or comprises the amino acid sequence as shown in SEQ ID NO:6. In some embodiments, the first multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO:6, or comprises the amino acid sequence as shown in SEQ ID NO:6; and the second multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO:5, or comprises the amino acid sequence as shown in SEQ ID NO:5. In some embodiments, the first multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98%, or 99% identity to SEQ ID NO:7, or comprises the amino acid sequence as shown in SEQ ID NO:7; and the second multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98%, or 99% identity to SEQ ID NO:8, or comprises the amino acid sequence as shown in SEQ ID NO:8. In some embodiments, the first multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98%, or 99% identity to SEQ ID NO:8, or comprises the amino acid sequence as shown in SEQ ID NO:8; and the second multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98%, or 99% identity to SEQ ID NO:7, or comprises the amino acid sequence as shown in SEQ ID NO:7. In some embodiments, the first multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO:9, or comprises the amino acid sequence as shown in SEQ ID NO:9; and the second multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO:10, or comprises the amino acid sequence as shown in SEQ ID NO:10.In some embodiments, the first multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO:10, or comprises the amino acid sequence as shown in SEQ ID NO:10; and the second multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO:9, or comprises the amino acid sequence as shown in SEQ ID NO:9.

[0014] In some embodiments, the first multimerization domain and the actuator domain are separated by a first polypeptide linker having a length of 2 to 40 amino acids. In some embodiments, the first polypeptide linker is selected from the group consisting of: GG, GS, SG, SS, GSS, SSG, GSG, SGS, SGG, G4S, 2xG4S, 3xG4S, 4xG4S, 5xG4S, and any combination thereof. In one embodiment, the first polypeptide linker is a 3xG4S linker. In some embodiments, the first polypeptide linker comprises a polypeptide comprising an amino acid sequence as shown in any one of SEQ ID NOs: 16-31. In some embodiments, the extracellular domain and the second multimerization domain are separated by a second polypeptide linker having a length of 2 to 40 amino acids. In some embodiments, the second polypeptide linker is selected from the group consisting of: GG, GS, SG, SS, GSS, SSG, GSG, SGS, SGG, G4S, 2x G4S, 3xG4s, 4xG4S, and any combination thereof. In one embodiment, the second polypeptide linker is a G4S linker. In some embodiments, the second polypeptide linker comprises a polypeptide comprising an amino acid sequence as shown in any one of SEQ ID NOs: 16-31.

[0015] In some embodiments, the transmembrane domain is a CD4 transmembrane domain. In some embodiments, the CD4 transmembrane domain comprises an amino acid sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity with SEQ ID NO: 45. In one embodiment, the CD4 transmembrane domain comprises an amino acid sequence as shown in SEQ ID NO: 45. In some embodiments, the transmembrane domain is a CD28 transmembrane domain. In some embodiments, the CD28 transmembrane domain comprises an amino acid sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity with SEQ ID NO: 46. In one embodiment, the CD28 transmembrane domain comprises an amino acid sequence as shown in SEQ ID NO: 46. In some embodiments, the transmembrane domain is a CD8 transmembrane domain. In some embodiments, the CD8 transmembrane domain comprises an amino acid sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity with SEQ ID NO: 47. In one embodiment, the CD8 transmembrane domain comprises the amino acid sequence shown in SEQ ID NO:47.

[0016] In some embodiments, the targeting component further comprises an intracellular signaling domain or a co-stimulatory domain derived from a protein selected from the group consisting of: an antigen receptor, a co-stimulatory receptor, a growth receptor, a cytokine receptor, an adaptor signaling protein, an intracellular signaling protein, or any fragment or variant thereof. In some embodiments, the intracellular signaling domain or the co-stimulatory domain on the targeting component is selected from the group consisting of: Toll-like receptor 1 (TLR1), TLR2, TLR3, TLR4, TLR5, TLR6, TLR7, TLR8, TLR9, TLR10, caspase recruitment domain family member 11 (CARD11), CD2, CD3ε, CD3γ, CD3δ, CD4, CD7, CD8, CD27, CD28, CD30, CD40, CD54 (ICAM), CD83, CD94, CD134 (OX40), CD137 (4-1B B), CD278 (ICOS), common gamma chain cytokine, DNAX-activating protein 10 (DAP10), linker for activating T cell family member 1 (LAT), interleukin 2 receptor (IL-2R), IL-4R, IL-7R, IL-9R, IL-12R, IL-13R, IL-15R, IL-21R, 76kD leukocyte protein containing SH2 domain (SLP76), T cell receptor associated transmembrane adaptor 1 (TRAT1), TNFR2, TNFRS14, TNFRS18, TNRFS25 and T cell receptor ζ chain associated protein kinase 70 (ZAP70). In some embodiments, the intracellular signaling domain or the costimulatory domain on the targeting component is selected from the group consisting of 4-1BB, CD28, TNFR2, OX40, ICOS and DAP10 costimulatory domains. In one embodiment, the costimulatory domain on the targeting component is a 4-1BB costimulatory domain, optionally wherein the 4-1BB costimulatory domain comprises an amino acid sequence as shown in SEQ ID NO: 98. In one embodiment, the costimulatory domain on the targeting component is a CD28 costimulatory domain, optionally wherein the CD28 costimulatory domain comprises an amino acid sequence as shown in SEQ ID NO: 99. In some embodiments, the intracellular signaling domain or the costimulatory domain is one or more cytokine receptor intracellular signaling domains. In some embodiments, the one or more cytokine receptor intracellular signaling domains are selected from the group consisting of: an IL7Rα intracellular signaling domain, an IL2Rβ intracellular signaling domain, a common γ chain intracellular signaling domain, and both IL2Rβ and common γ chain intracellular signaling domains.In some embodiments, the IL7Rα intracellular signaling domain comprises an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98%, or 99% identical to SEQ ID NO: 103. In one embodiment, the IL7Rα intracellular signaling domain comprises an amino acid sequence as shown in SEQ ID NO: 103. In some embodiments, the IL2Rβ intracellular signaling domain comprises an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98%, or 99% identical to SEQ ID NO: 104 or SEQ ID NO: 105. In one embodiment, the IL2Rβ intracellular signaling domain comprises an amino acid sequence as shown in SEQ ID NO: 104 or SEQ ID NO: 105. In some embodiments, the common gamma chain intracellular signaling domain comprises an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98%, or 99% identical to SEQ ID NO: 106. In one embodiment, the common gamma chain intracellular signaling domain comprises the amino acid sequence as shown in SEQ ID NO: 106. In some embodiments, the intracellular signaling domain or the costimulatory domain is a LAT domain. In some embodiments, the LAT domain comprises an amino acid sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity to the amino acid sequence as shown in SEQ ID NO: 102. In one embodiment, the LAT domain comprises an amino acid sequence as shown in SEQ ID NO: 102. In some embodiments, the intracellular signaling domain or the costimulatory domain is a CD4 co-receptor domain. In some embodiments, the CD4 co-receptor domain comprises an amino acid sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO: 100. In one embodiment, the CD4 co-receptor domain comprises an amino acid sequence as shown in SEQ ID NO: 100. In some embodiments, the intracellular signaling domain or the costimulatory domain is a CD8 co-receptor domain. In some embodiments, the CD8 co-receptor domain comprises an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO: 101. In one embodiment, the CD8 co-receptor domain comprises an amino acid sequence as shown in SEQ ID NO: 101.

[0017] In some embodiments, the targeting component does not include a functional intracellular signaling domain or a co-stimulatory domain. In some embodiments, the targeting component further includes a truncated intracellular CD4 polypeptide. In some embodiments, the truncated intracellular CD4 polypeptide includes an amino acid sequence as shown in SEQ ID NO:48 or SEQ ID NO:49. In some embodiments, the truncated intracellular CD4 polypeptide includes an amino acid sequence as shown in SEQ ID NO:48 or SEQ ID NO:49. In some embodiments, the extracellular domain includes a first targeting domain. In some embodiments, the first targeting domain includes a single-chain variable fragment (scFv) or a single domain antibody (sdAb). In some embodiments, the sdAb is a camel VHH, a nanobody, or a heavy chain antibody (HcAb) only. In one embodiment, the sdAb is a camel VHH. In some embodiments, the scFv or the sdAb is human or humanized. In some embodiments, the first targeting domain includes a sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity to any one of SEQ ID NO:50-83. In some embodiments, the first targeting domain comprises a sequence as shown in any one of SEQ ID NOs: 50-83. In some embodiments, the first targeting domain comprises a PD1 extracellular domain, a human A proliferation-inducing ligand (APRIL), a trimerized human APRIL, or a NKG2D membrane protein. In some embodiments, the PD1 extracellular domain comprises a sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO: 84 or SEQ ID NO: 85. In one embodiment, the PD1 extracellular domain comprises a sequence as shown in SEQ ID NO: 85. In some embodiments, the APRIL comprises a sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO: 86, or the trimerized human APRIL comprises a sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO: 87. In one embodiment, the APRIL comprises the sequence as shown in SEQ ID NO: 86, or the trimerized human APRIL comprises the sequence as shown in SEQ ID NO: 87. In some embodiments, the NKG2D membrane protein comprises a sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO: 88. In one embodiment, the NKG2D membrane protein comprises the sequence as shown in SEQ ID NO: 88.

[0018] In some embodiments, the extracellular domain further comprises a second targeting domain. In some embodiments, the second targeting domain comprises a second single-chain variable fragment (scFv) or a second single domain antibody (sdAb). In some embodiments, the second sdAb is a camel VHH, a nano antibody or a heavy chain antibody only (HcAb). In one embodiment, the second sdAb is a camel VHH. In some embodiments, the second scFv or the second sdAb is human or humanized. In some embodiments, the second targeting domain comprises a sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity with any one of SEQ ID NO:50-83. In some embodiments, the second targeting domain comprises a sequence as shown in any one of SEQ ID NO:50-83. In some embodiments, the second targeting domain comprises a PD1 extracellular domain, a human A proliferation-inducing ligand (APRIL), a trimerized human APRIL or a NKG2D membrane protein. In some embodiments, the PD1 extracellular domain comprises a sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO: 84 or SEQ ID NO: 85. In one embodiment, the PD1 extracellular domain comprises a sequence as shown in SEQ ID NO: 85. In some embodiments, the APRIL comprises a sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO: 86, or the trimerized human APRIL comprises a sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO: 87. In one embodiment, the APRIL comprises a sequence as shown in SEQ ID NO: 86, or the trimerized human APRIL comprises a sequence as shown in SEQ ID NO: 87. In some embodiments, the NKG2D membrane protein comprises a sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO: 88. In one embodiment, the NKG2D membrane protein comprises the sequence shown in SEQ ID NO:88.

[0019] In some embodiments, the targeting domain and the second targeting domain bind to the same antigen or different antigens. In some embodiments, the targeting domain and the second targeting domain are separated by a third polypeptide linker having a length of 2 to 40 amino acids. In some embodiments, the third polypeptide linker is selected from the group consisting of: GG, GS, SG, SS, GSS, SSG, GSG, SGS, SGG, G4S, 2x G4S, 3xG4s, 4xG4S, 5xG4S, any one of SEQ ID NO: 16-31 and any combination thereof. In some embodiments, the signaling component comprises a sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO: 107, SEQ ID NO: 108, SEQ ID NO: 109, SEQ ID NO: 110, SEQ ID NO: 111, SEQ ID NO: 112, SEQ ID NO: 113, SEQ NO: 114 or SEQ ID NO: 115. In some embodiments, the signaling component comprises a sequence as shown in SEQ ID NO:107, SEQ ID NO:108, SEQ ID NO:109, SEQ ID NO:110, SEQ ID NO:111, SEQ ID NO:112, SEQ ID NO:113, SEQ NO:114, or SEQ ID NO:115.

[0020] In some embodiments, the targeting component comprises a sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to the sequence shown in SEQ ID NO:116, SEQ ID NO:117, SEQ ID NO:118, SEQ ID NO:119, SEQ ID NO:120, SEQ ID NO:121, SEQ ID NO:122, SEQ ID NO:123, SEQ ID NO:124, SEQ ID NO:125, SEQ ID NO:126, SEQ ID NO:127, SEQ ID NO:128, SEQ ID NO:129, SEQ ID NO:130, SEQ ID NO:131, SEQ ID NO:132, SEQ ID NO:133, SEQ ID NO:134, SEQ ID NO:135 or SEQ ID NO:136. In some embodiments, the targeting component comprises a sequence as shown in SEQ ID NO:116, SEQ ID NO:117, SEQ ID NO:118, SEQ ID NO:119, SEQ ID NO:120, SEQ ID NO:121, SEQ ID NO:122, SEQ ID NO:123, SEQ ID NO:124, SEQ ID NO:125, SEQ ID NO:126, SEQ ID NO:127, SEQ ID NO:128, SEQID NO:129, SEQ ID NO:130, SEQ ID NO:131, SEQ ID NO:132, SEQ ID NO:133, SEQ ID NO:134, SEQ ID NO:135 or SEQ ID NO:136.

[0021] In some embodiments, the signaling component and the targeting component further comprise a signal sequence, optionally wherein the signal sequence is a CD8, IgK or PD1 signal sequence.

[0022] In some embodiments, the signaling component further comprises a signal sequence, optionally wherein the signal sequence is at least 90%, 95%, 96%, 97%, 98% or 99% identical to or comprises SEQ ID NO:95, SEQ ID NO:96 or SEQ ID NO:97.

[0023] In some embodiments, the targeting component further comprises a signal sequence, optionally wherein the signal sequence is at least 90%, 95%, 96%, 97%, 98% or 99% identical to or comprises SEQ ID NO:95, SEQ ID NO:96 or SEQ ID NO:97.

[0024] In one embodiment, the signaling component further comprises a signal sequence. In one embodiment, the signal sequence is a CD8 signal sequence. In one embodiment, the CD8 signal sequence comprises an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO: 96. In another embodiment, the CD8 signal sequence comprises an amino acid sequence as shown in SEQ ID NO: 96.

[0025] In another embodiment, the targeting component further comprises a signal sequence. In one embodiment, the signal sequence is an IgK signal sequence. In one embodiment, the IgK signal sequence comprises an amino acid sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO: 95. In one embodiment, the IgK signal sequence comprises an amino acid sequence as shown in SEQ ID NO: 95.

[0026] In one embodiment, the non-native cell comprises a fusion polypeptide comprising the targeting component and the signaling component. In some embodiments, the fusion polypeptide comprises SEQ ID NO: 139, SEQ ID NO: 140, SEQ ID NO: 141, SEQ ID NO: 142, SEQ ID NO: 143, SEQ ID NO: 144, SEQ ID NO: 145, SEQ ID NO: 146, SEQ ID NO: 147, SEQ ID NO: 148, SEQ ID NO: 149, SEQ ID NO: 150, SEQ ID NO: 151, SEQ ID NO: 152, SEQ ID NO: 153, SEQ ID NO: 154, SEQ ID NO: 155, SEQ ID NO: 156, SEQ ID NO: 157, SEQ ID NO: 158, SEQ ID NO: 159, SEQ ID NO: 160, SEQ ID NO: 161, SEQ ID NO: 162, SEQ ID NO: 163, SEQ ID NO: 164, SEQ ID NO: 165, SEQ ID NO: 166, SEQ ID NO: 167, SEQ ID NO: NO: 168, SEQ ID NO: 169 or SEQ ID NO: 170 having a sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical.In some embodiments, the fusion polypeptide comprises SEQ ID NO: 139, SEQ ID NO: 140, SEQ ID NO: 141, SEQ ID NO: 142, SEQ ID NO: 143, SEQ ID NO: 144, SEQ ID NO: 145, SEQ ID NO: 146, SEQ ID NO: 147, SEQ ID NO: 148, SEQ ID NO: 149, SEQ ID NO: 150, SEQ ID NO: 151, SEQ ID NO: 152, SEQ ID NO: 153, SEQ ID NO: 154, SEQ ID NO: 155, SEQ ID NO: 156, SEQ ID NO: 157, SEQ ID NO: 158, SEQ ID NO: 159, SEQ ID NO: 160, SEQ ID NO: 161, SEQ ID NO: 162, SEQ ID NO: 163, SEQ ID NO: 164, SEQ ID NO: 165, SEQ ID NO: 166, SEQ ID NO: 167, SEQ ID NO: 168, The sequence shown in NO:168, SEQ ID NO:169 or SEQ ID NO:170.

[0027] In some embodiments, the cell comprises a first nucleic acid molecule encoding the signaling component. In some embodiments, the cell comprises a second nucleic acid molecule encoding the targeting component. In some embodiments, the cell comprises a nucleic acid molecule encoding both the signaling component and the targeting component. In some embodiments, the cell further expresses an exogenous co-stimulatory factor, an immunomodulatory factor, an agonist of a co-stimulatory factor, an antagonist of an immunosuppressive factor, an immune cell engager, a flip receptor, or any combination thereof.

[0028] In some embodiments, the cell further expresses an exogenous lymphocyte receptor or co-receptor. In some embodiments, the exogenous lymphocyte receptor or co-receptor is selected from the group consisting of: TCRα (TCR alpha), TCRβ (TCR beta), TCRγ (TCR gamma), TCRδ (TCR delta), CD4, CD8, pre-T cell receptor alpha (pTα), Fc receptor alpha (FcRα), Fc receptor beta (FcRβ), Fc receptor gamma (FcRγ), natural killer group 2 member D (NKG2D), CD79A, CD79B and any combination thereof. In one embodiment, the cell further expresses an exogenous TCR.In some embodiments, the exogenous TCR binds to a target antigen selected from the group consisting of alpha-fetoprotein (AFP), B melanoma antigen (BAGE) family members, brothers of regulatory factors at imprinted sites (BORIS), cancer-testis antigen, cancer-testis antigen 83 (CT-83), carbonic anhydrase IX (CA1X), carcinoembryonic antigen (CEA), cytomegalovirus (CMV) antigen, antigen recognized by cytotoxic T cells (CTL) on melanoma (CAMEL), Epstein-Barr virus (Epstein-Barr virus, EBV) antigen, G antigen 1 (GAGE-1), GAGE-2, GAGE-3, GAGE-4, GAGE-5, GAGE-6, GAGE-7B, GAGE-8, glycoprotein 100 (GP100), hepatitis B virus (HBV) antigen, hepatitis C virus (HCV) nonstructural protein 3 (NS3), human epidermal growth factor receptor 2 (HER-2), human papillomavirus (HPV)-E6, HPV-E7, human telomerase reverse transcriptase (hTERT), IGF2BP3 / A3, K-Ras, K-Ras G12C, K-Ras G12D, K-Ras G12V, latent membrane protein 2 (LMP2), melanoma antigen family A, 1 (MAGE-A1), MAGE-A2, MAGE-A3, MAGE-A4, MAGE-A6, MAGE-A10, MAGE-A12, melanoma antigen recognized by T cells (MART-1), mesothelin (MSLN), mucin 1 (MUC1), mucin 16 (MUC16), New York esophageal squamous cell carcinoma-1 (NYESO-1), P53, P antigen (PAGE) family member, placenta-specific 1 (PLAC1), antigen preferentially expressed in melanoma (PRAME), survivin, synovial sarcoma X1 (SSX1), synovial sarcoma X2 (SSX2), synovial sarcoma X3 (SSX3), synovial sarcoma X4 (SSX4), synovial sarcoma X5 (SSX5), synovial sarcoma X 8 (SSX8), thyroglobulin, tyrosinase, tyrosinase-related protein (TRP) 1, TRP2, Wilms tumor protein (WT-1), X antigen family member 1 (XAGE1) and X antigen family member 2 (XAGE2). In some embodiments, the exogenous TCR is an αβ-TCR or a γδ-TCR.

[0029] In some embodiments, the cell further expresses a CAR, CCR, or flip receptor. In some embodiments, the cell further expresses a zetakine, an immune cell engager, or a BiTE. In one embodiment, the cell is a hematopoietic cell. In some embodiments, the cell is a T cell, an αβ-T cell, or a γδ-T cell. In some embodiments, the cell is a CD3 + cells, CD4 + cells and / or CD8 + Cell. In one embodiment, the cell is an immune effector cell. In some embodiments, the cell is a cytotoxic T lymphocyte (CTL), a tumor infiltrating lymphocyte (TIL) or a helper T cell. In some embodiments, the cell is a natural killer (NK) cell or a natural killer T (NKT) cell. In some embodiments, the source of the cell is a peripheral blood mononuclear cell, bone marrow, lymph node tissue, umbilical cord blood, thymus tissue, tissue from an infected site, ascites, pleural effusion, spleen tissue or a tumor. In one embodiment, the non-natural cell is an isolated non-natural cell. In one embodiment, the non-natural cell is obtained from a subject. In one embodiment, the non-natural cell is a human cell. In some embodiments, the cell further comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO: 137. In one embodiment, the cell further comprises an amino acid sequence as shown in SEQ ID NO: 137. In some embodiments, the cell further comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO: 138. In one embodiment, the cell further comprises an amino acid sequence as shown in SEQ ID NO:138.

[0030] In one aspect, a fusion polypeptide is disclosed herein, comprising a signaling component including a first multimerization domain and an actuator domain; a polypeptide cleavage signal; and a targeting component comprising an extracellular domain, a second multimerization domain, and a transmembrane domain. In one embodiment, when the signaling component and the targeting component are expressed, the first multimerization domain and the second multimerization domain are located extracellularly.

[0031] In one embodiment, the second multimerization domain and the transmembrane domain are separated by a hinge domain. In some embodiments, the hinge domain is selected from the group consisting of a CD4 hinge, a CD8 hinge, a CD28 hinge, an IgG4 hinge, and any fragment or variant or combination thereof. In some embodiments, the hinge domain is a CD4 hinge. In some embodiments, the CD4 hinge comprises an amino acid sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO: 41. In one embodiment, the CD4 hinge comprises an amino acid sequence as shown in SEQ ID NO: 41. In some embodiments, the hinge domain is a CD28 hinge. In some embodiments, the CD28 hinge comprises an amino acid sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO: 42. In one embodiment, the CD28 hinge comprises an amino acid sequence as shown in SEQ ID NO: 42. In some embodiments, the hinge domain is a CD8 hinge. In some embodiments, the CD8 hinge comprises an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98%, or 99% identical to SEQ ID NO: 44. In one embodiment, the CD8 hinge comprises an amino acid sequence as shown in SEQ ID NO: 44. In some embodiments, the hinge domain is an IgG4 hinge. In some embodiments, the IgG4 hinge comprises an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98%, or 99% identical to SEQ ID NO: 43. In one embodiment, the IgG4 hinge comprises an amino acid sequence as shown in SEQ ID NO: 43.

[0032] In some embodiments, the actuator domain is a CD3 polypeptide, an FcεR1γ polypeptide, an Igα / CD79a polypeptide, an Igβ / CD79b polypeptide, a DAP10 polypeptide, or a DAP12 polypeptide. In some embodiments, the CD3 polypeptide is a CD3ε (CD3epsilon) polypeptide or a fragment or variant thereof, a CD3γ (CD3 gamma) or a fragment or variant thereof, or a CD3δ (CD3 delta) or a fragment or variant thereof. In some embodiments, the actuator domain is a CD3ε polypeptide comprising an amino acid sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO: 32. In one embodiment, the actuator domain is a CD3ε polypeptide comprising an amino acid sequence as shown in SEQ ID NO: 32. In some embodiments, the actuator domain is a CD3γ polypeptide or a variant thereof. In some embodiments, the actuator domain is a CD3γ polypeptide comprising an amino acid sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity with SEQ ID NO: 33. In one embodiment, the actuator domain is a CD3γ polypeptide comprising an amino acid sequence as shown in SEQ ID NO: 33. In some embodiments, the actuator domain is a CD3δ polypeptide or a variant thereof. In some embodiments, the actuator domain is a CD3δ polypeptide comprising an amino acid sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity with SEQ ID NO: 34. In one embodiment, the actuator domain is a CD3δ polypeptide comprising an amino acid sequence as shown in SEQ ID NO: 34. In some embodiments, the actuator domain is an FcεR1γ polypeptide or a variant thereof. In some embodiments, the actuator domain is an FcεR1γ polypeptide comprising an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:35. In one embodiment, the actuator domain is an FcεR1γ polypeptide comprising an amino acid sequence as shown in SEQ ID NO:35. In some embodiments, the actuator domain is an FcεR1γ polypeptide comprising an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:36. In one embodiment, the actuator domain is an FcεR1γ polypeptide comprising an amino acid sequence as shown in SEQ ID NO:36. In some embodiments, the actuator domain is an Igα / CD79a polypeptide or a variant thereof. In some embodiments, the actuator domain is an Igα / CD79a polypeptide comprising an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:37.In one embodiment, the actuator domain is an Igα / CD79a polypeptide comprising an amino acid sequence as shown in SEQ ID NO:37. In some embodiments, the actuator domain is an Igβ / CD79b polypeptide or a variant thereof. In some embodiments, the actuator domain is an Igβ / CD79b polypeptide comprising an amino acid sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity with SEQ ID NO:38. In one embodiment, the actuator domain is an Igβ / CD79b polypeptide comprising an amino acid sequence as shown in SEQ ID NO:38. In some embodiments, the actuator domain is a DAP10 polypeptide or a variant thereof. In some embodiments, the actuator domain is a DAP10 polypeptide comprising an amino acid sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity with SEQ ID NO:39. In one embodiment, the actuator domain is a DAP10 polypeptide comprising an amino acid sequence as shown in SEQ ID NO:39. In some embodiments, the actuator domain is a DAP12 polypeptide or a variant thereof. In some embodiments, the actuator domain is a DAP12 polypeptide comprising an amino acid sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO: 40. In one embodiment, the actuator domain is a DAP12 polypeptide comprising an amino acid sequence as shown in SEQ ID NO: 40. In some embodiments, the actuator domain comprises both an extracellular portion and an intracellular portion.

[0033] In some embodiments, the fusion polypeptide further comprises a signal sequence, optionally wherein the signal sequence is at least 90%, 95%, 96%, 97%, 98% or 99% identical to or comprises SEQ ID NO:95, SEQ ID NO:96 or SEQ ID NO:97.

[0034] In some embodiments, the first multimerization domain is different from the second multimerization domain. In some embodiments, the multimerization domains of the signaling component and the targeting component are associated with a bridging factor selected from the group consisting of: rapamycin or a rapamycin analog thereof, gibberellins or derivatives thereof, abscisic acid (ABA) or derivatives thereof, methotrexate or derivatives thereof, cyclosporin A or derivatives thereof, FK506 / cyclosporin A or derivatives thereof, trimethoprim (Tmp) synthetic ligand (SLF) for FK506 binding protein (FKBP) or derivatives thereof, wherein the bridging factor promotes the formation of a polypeptide complex, wherein the bridging factor is associated with the multimerization domains of the signaling component and the targeting component and is positioned between the multimerization domains. In some embodiments, the first multimerization domain and the second multimerization domain are a pair selected from the group consisting of: FK506 binding protein 1A (FKBP12) and FKBP12-rapamycin binding (FRB), FKBP12 and calcineurin, FKBP and cyclophilin A or any other member of the peptidyl-prolyl cis-trans isomerase (PPIase) family, FKBP and dihydrofolate reductase (DHFR), calcineurin and cyclophilin A or any other member of the peptidyl-prolyl cis-trans isomerase (PPIase) family, and PYR1-like 1 (PYL1) and abscisic acid insensitive 1 (ABI1). In some embodiments, the first multimerization domain comprises a first FRB polypeptide or a variant thereof, and the second multimerization domain comprises a first FKBP12 polypeptide or a variant thereof. In some embodiments, the first multimerization domain comprises a first FKBP12 polypeptide or a variant thereof, and the second multimerization domain comprises a first FRB polypeptide or a variant thereof. In one embodiment, the FRB polypeptide is a FRB T2098L variant. In some embodiments, the FRB polypeptide comprises the amino acid sequence as shown in SEQ ID NO: 1 or SEQ ID NO: 2. In some embodiments, the FKBP12 polypeptide comprises the amino acid sequence as shown in SEQ ID NO: 3 or SEQ ID NO: 4.

[0035] In some embodiments, the bridging factor is AP1903, AP20187, AP21967 (also known as C16-(S)-7-methylindorapamycin), everolimus, novolimus, pimecrolimus, ridaforolimus, sirolimus, tacrolimus, temsirolimus, umirolimus, zotarolimus or BPC015. In some embodiments, the first multimerization domain and the second multimerization domain are a pair of antibody-derived heterodimerization domains. In some embodiments, the first multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO:5, or comprises the amino acid sequence as shown in SEQ ID NO:5; and the second multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO:6, or comprises the amino acid sequence as shown in SEQ ID NO:6. In some embodiments, the first multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO:6, or comprises the amino acid sequence as shown in SEQ ID NO:6; and the second multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO:5, or comprises the amino acid sequence as shown in SEQ ID NO:5. In some embodiments, the first multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98%, or 99% identity to SEQ ID NO: 7, or comprises the amino acid sequence as shown in SEQ ID NO: 7; and the second multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98%, or 99% identity to SEQ ID NO: 8, or comprises the amino acid sequence as shown in SEQ ID NO: 8. In some embodiments, the first multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98%, or 99% identity to SEQ ID NO: 8, or comprises the amino acid sequence as shown in SEQ ID NO: 8; and the second multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98%, or 99% identity to SEQ ID NO: 7, or comprises the amino acid sequence as shown in SEQ ID NO: 7.In some embodiments, the first multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98%, or 99% identity to SEQ ID NO: 9, or comprises the amino acid sequence as shown in SEQ ID NO: 9; and the second multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98%, or 99% identity to SEQ ID NO: 10, or comprises the amino acid sequence as shown in SEQ ID NO: 10. In some embodiments, the first multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98%, or 99% identity to SEQ ID NO: 10, or comprises the amino acid sequence as shown in SEQ ID NO: 10; and the second multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98%, or 99% identity to SEQ ID NO: 9, or comprises the amino acid sequence as shown in SEQ ID NO: 9.

[0036] In some embodiments, the first multimerization domain and the actuator domain are separated by a first polypeptide linker having a length of 2 to 40 amino acids. In some embodiments, the first polypeptide linker is selected from the group consisting of: GG, GS, SG, SS, GSS, SSG, GSG, SGS, SGG, G4S, 2xG4S, 3xG4S, 4xG4S, 5xG4S, and any combination thereof. In one embodiment, the first polypeptide linker is a 3xG4S linker. In some embodiments, the first polypeptide linker comprises a polypeptide comprising an amino acid sequence as shown in any one of SEQ ID NOs: 16-31. In some embodiments, the extracellular domain and the second multimerization domain are separated by a second polypeptide linker having a length of 2 to 40 amino acids. In some embodiments, the second polypeptide linker is selected from the group consisting of: GG, GS, SG, SS, GSS, SSG, GSG, SGS, SGG, G4S, 2x G4S, 3xG4s, 4xG4S, and any combination thereof. In one embodiment, the second polypeptide linker is a G4S linker. In some embodiments, the second polypeptide linker comprises a polypeptide comprising an amino acid sequence as shown in any one of SEQ ID NOs: 16-31.

[0037] In some embodiments, the transmembrane domain is a CD4 transmembrane domain. In some embodiments, the CD4 transmembrane domain comprises an amino acid sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity with SEQ ID NO: 45. In one embodiment, the CD4 transmembrane domain comprises an amino acid sequence as shown in SEQ ID NO: 45. In some embodiments, the transmembrane domain is a CD28 transmembrane domain. In some embodiments, the CD28 transmembrane domain comprises an amino acid sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity with SEQ ID NO: 46. In one embodiment, the CD28 transmembrane domain comprises an amino acid sequence as shown in SEQ ID NO: 46. In some embodiments, the transmembrane domain is a CD8 transmembrane domain. In some embodiments, the CD8 transmembrane domain comprises an amino acid sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity with SEQ ID NO: 47. In one embodiment, the CD8 transmembrane domain comprises the amino acid sequence shown in SEQ ID NO:47.

[0038] In some embodiments, the targeting component further comprises an intracellular signaling domain or a co-stimulatory domain derived from a protein selected from the group consisting of: an antigen receptor, a co-stimulatory receptor, a growth receptor, a cytokine receptor, an adaptor signaling protein, an intracellular signaling protein, or any fragment or variant thereof. In some embodiments, the intracellular signaling domain or the co-stimulatory domain on the targeting component is selected from the group consisting of: Toll-like receptor 1 (TLR1), TLR2, TLR3, TLR4, TLR5, TLR6, TLR7, TLR8, TLR9, TLR10, caspase recruitment domain family member 11 (CARD11), CD2, CD3ε, CD3γ, CD3δ, CD4, CD7, CD8, CD27, CD28, CD30, CD40, CD54 (ICAM), CD83, CD94, CD134 (OX40), CD137 (4-1B B), CD278 (ICOS), common gamma chain cytokine, DNAX-activating protein 10 (DAP10), linker for activating T cell family member 1 (LAT), interleukin 2 receptor (IL-2R), IL-4R, IL-7R, IL-9R, IL-12R, IL-13R, IL-15R, IL-21R, 76kD leukocyte protein containing SH2 domain (SLP76), T cell receptor associated transmembrane adaptor 1 (TRAT1), TNFR2, TNFRS14, TNFRS18, TNRFS25 and T cell receptor ζ chain associated protein kinase 70 (ZAP70). In some embodiments, the intracellular signaling domain or the costimulatory domain on the targeting component is selected from the group consisting of 4-1BB, CD28, TNFR2, OX40, ICOS and DAP10 costimulatory domains. In some embodiments, the costimulatory domain on the targeting component is a 4-1BB costimulatory domain, optionally wherein the 4-1BB costimulatory domain comprises the amino acid sequence as shown in SEQ ID NO: 98. In some embodiments, the costimulatory domain on the targeting component is a CD28 costimulatory domain, optionally wherein the CD28 costimulatory domain comprises the amino acid sequence as shown in SEQ ID NO: 99.

[0039] In some embodiments, the intracellular signaling domain or the co-stimulatory domain is one or more cytokine receptor intracellular signaling domains. In some embodiments, the one or more cytokine receptor intracellular signaling domains are selected from the group consisting of: an IL7Rα intracellular signaling domain, an IL2Rβ intracellular signaling domain, a common γ chain intracellular signaling domain, and both IL2Rβ and common γ chain intracellular signaling domains. In some embodiments, the IL7Rα intracellular signaling domain comprises an amino acid sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO: 103. In one embodiment, the IL7Rα intracellular signaling domain comprises an amino acid sequence as shown in SEQ ID NO: 103. In some embodiments, the IL2Rβ intracellular signaling domain comprises an amino acid sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO: 104 or SEQ ID NO: 105. In some embodiments, the IL2Rβ intracellular signaling domain comprises an amino acid sequence as shown in SEQ ID NO: 104 or SEQ ID NO: 105. In some embodiments, the common gamma chain intracellular signaling domain comprises an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO: 106. In one embodiment, the common gamma chain intracellular signaling domain comprises an amino acid sequence as shown in SEQ ID NO: 106. In some embodiments, the intracellular signaling domain or the costimulatory domain is a LAT domain. In some embodiments, the LAT domain comprises an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to the amino acid sequence as shown in SEQ ID NO: 102. In one embodiment, the LAT domain comprises an amino acid sequence as shown in SEQ ID NO: 102.

[0040] In some embodiments, the intracellular signaling domain or the co-stimulatory domain is a CD4 co-receptor domain. In some embodiments, the CD4 co-receptor domain comprises an amino acid sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO: 100. In one embodiment, the CD4 co-receptor domain comprises an amino acid sequence as shown in SEQ ID NO: 100. In some embodiments, the intracellular signaling domain or the co-stimulatory domain is a CD8 co-receptor domain. In some embodiments, the CD8 co-receptor domain comprises an amino acid sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO: 101. In one embodiment, the CD8 co-receptor domain comprises an amino acid sequence as shown in SEQ ID NO: 101.

[0041] In some embodiments, the targeting component does not include a functional intracellular signaling domain or a co-stimulatory domain. In some embodiments, the targeting component further includes a truncated intracellular CD4 polypeptide. In some embodiments, the truncated intracellular CD4 polypeptide includes an amino acid sequence as shown in SEQ ID NO:48 or SEQ ID NO:49. In some embodiments, the truncated intracellular CD4 polypeptide includes an amino acid sequence as shown in SEQ ID NO:48 or SEQ ID NO:49. In some embodiments, the extracellular domain includes a first targeting domain. In some embodiments, the first targeting domain includes a single-chain variable fragment (scFv) or a single domain antibody (sdAb). In some embodiments, the sdAb is a camel VHH, a nanobody, or a heavy chain antibody (HcAb) only. In one embodiment, the sdAb is a camel VHH. In some embodiments, the scFv or the sdAb is human or humanized. In some embodiments, the first targeting domain includes a sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity to any one of SEQ ID NO:50-83. In some embodiments, the first targeting domain comprises a sequence as shown in any one of SEQ ID NOs: 50-83. In some embodiments, the first targeting domain comprises a PD1 extracellular domain, a human A proliferation-inducing ligand (APRIL), a trimerized human APRIL, or a NKG2D membrane protein. In some embodiments, the PD1 extracellular domain comprises a sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO: 84 or SEQ ID NO: 85. In one embodiment, the PD1 extracellular domain comprises a sequence as shown in SEQ ID NO: 85. In some embodiments, the APRIL comprises a sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO: 86, or the trimerized human APRIL comprises a sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO: 87. In some embodiments, the APRIL comprises the sequence as shown in SEQ ID NO: 86, or the trimerized human APRIL comprises the sequence as shown in SEQ ID NO: 87. In some embodiments, the NKG2D membrane protein comprises a sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO: 88. In one embodiment, the NKG2D membrane protein comprises the sequence as shown in SEQ ID NO: 88.

[0042] In some embodiments, the extracellular domain further comprises a second targeting domain. In some embodiments, the second targeting domain comprises a second single-chain variable fragment (scFv) or a second single domain antibody (sdAb). In some embodiments, the second sdAb is a camel VHH, a nano antibody or a heavy chain antibody only (HcAb). In one embodiment, the second sdAb is a camel VHH. In some embodiments, the second scFv or the second sdAb is human or humanized. In some embodiments, the second targeting domain comprises a sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity with any one of SEQ ID NO:50-83. In some embodiments, the second targeting domain comprises a sequence as shown in any one of SEQ ID NO:50-83. In some embodiments, the second targeting domain comprises a PD1 extracellular domain, a human A proliferation-inducing ligand (APRIL), a trimerized human APRIL or a NKG2D membrane protein. In some embodiments, the PD1 extracellular domain comprises a sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO: 84 or SEQ ID NO: 85. In one embodiment, the PD1 extracellular domain comprises a sequence as shown in SEQ ID NO: 85. In some embodiments, the APRIL comprises a sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO: 86, or the trimerized human APRIL comprises a sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO: 87. In some embodiments, the APRIL comprises a sequence as shown in SEQ ID NO: 86, or the trimerized human APRIL comprises a sequence as shown in SEQ ID NO: 87. In some embodiments, the NKG2D membrane protein comprises a sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO: 88. In one embodiment, the NKG2D membrane protein comprises a sequence as shown in SEQ ID NO: 88. In some embodiments, the first targeting domain and the second targeting domain bind to the same antigen or different antigens. In some embodiments, the first targeting domain and the second targeting domain are separated by a third polypeptide linker having a length of 2 to 40 amino acids. In some embodiments, the third polypeptide linker is selected from the group consisting of: GG, GS, SG, SS, GSS, SSG, GSG, SGS, SGG, G4S, 2x G4S, 3xG4s, 4xG4S, 5xG4S, any one of SEQ ID NO: 16-31 and any combination thereof.

[0043] In some embodiments, the polypeptide cleavage signal is a viral self-cleaving polypeptide. In some embodiments, the polypeptide cleavage signal is a viral self-cleaving 2A polypeptide. In some embodiments, wherein the polypeptide cleavage signal is a viral self-cleaving polypeptide, the viral self-cleaving polypeptide is selected from the group consisting of: foot-and-mouth disease virus (FMDV) (F2A) peptide, equine rhinitis virus type A (ERAV) (E2A) peptide, Thosea asigna virus, TaV (T2A) peptide, porcine Teschovirus-1 (PTV-1) (P2A) peptide, Theilovirus 2A peptide and encephalomyocarditis virus 2A peptide.

[0044] In one embodiment, the signaling component further comprises a signal sequence. In one embodiment, the signal sequence is a CD8 signal sequence. In one embodiment, the CD8 signal sequence comprises an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO: 96. In another embodiment, the CD8 signal sequence comprises an amino acid sequence as shown in SEQ ID NO: 96.

[0045] In another embodiment, the targeting component further comprises a signal sequence. In one embodiment, the signal sequence is an IgK signal sequence. In one embodiment, the IgK signal sequence comprises an amino acid sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO: 95. In one embodiment, the IgK signal sequence comprises an amino acid sequence as shown in SEQ ID NO: 95.

[0046] In some embodiments, the signaling component comprises a sequence that is at least 90%, 95%, 96%, 97%, 98%, or 99% identical to SEQ ID NO: 107, SEQ ID NO: 108, SEQ ID NO: 109, SEQ ID NO: 110, SEQ ID NO: 111, SEQ ID NO: 112, SEQ ID NO: 113, SEQ NO: 114, or SEQ ID NO: 115. In some embodiments, the signaling component comprises a sequence as set forth in SEQ ID NO: 107, SEQ ID NO: 108, SEQ ID NO: 109, SEQ ID NO: 110, SEQ ID NO: 111, SEQ ID NO: 112, SEQ ID NO: 113, SEQ NO: 114, or SEQ ID NO: 115.

[0047] In some embodiments, the targeting component comprises a sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to the sequence shown in SEQ ID NO:116, SEQ ID NO:117, SEQ ID NO:118, SEQ ID NO:119, SEQ ID NO:120, SEQ ID NO:121, SEQ ID NO:122, SEQ ID NO:123, SEQ ID NO:124, SEQ ID NO:125, SEQ ID NO:126, SEQ ID NO:127, SEQ ID NO:128, SEQ ID NO:129, SEQ ID NO:130, SEQ ID NO:131, SEQ ID NO:132, SEQ ID NO:133, SEQ ID NO:134, SEQ ID NO:135 or SEQ ID NO:136. In some embodiments, the targeting component comprises a sequence as shown in SEQ ID NO: 116, SEQ ID NO: 117, SEQ ID NO: 118, SEQ ID NO: 119, SEQ ID NO: 120, SEQ ID NO: 121, SEQ ID NO: 122, SEQ ID NO: 123, SEQ ID NO: 124, SEQ ID NO: 125, SEQ ID NO: 126, SEQ ID NO: 127, SEQ ID NO: 128, SEQ ID NO: 129, SEQ ID NO: 130, SEQ ID NO: 131, SEQ ID NO: 132, SEQ ID NO: 133, SEQ ID NO: 134, SEQ ID NO: 135, or SEQ ID NO: 136. In some embodiments, the fusion polypeptide comprises the targeting component and the signaling component.

[0048] In some embodiments, the fusion polypeptide comprises SEQ ID NO: 139, SEQ ID NO: 140, SEQ ID NO: 141, SEQ ID NO: 142, SEQ ID NO: 143, SEQ ID NO: 144, SEQ ID NO: 145, SEQ ID NO: 146, SEQ ID NO: 147, SEQ ID NO: 148, SEQ ID NO: 149, SEQ ID NO: 150, SEQ ID NO: 151, SEQ ID NO: 152, SEQ ID NO: 153, SEQ ID NO: 154, SEQ ID NO: 155, SEQ ID NO: 156, SEQ ID NO: 157, SEQ ID NO: 158, SEQ ID NO: 159, SEQ ID NO: 160, SEQ ID NO: 161, SEQ ID NO: 162, SEQ ID NO: 163, SEQ ID NO: 164, SEQ ID NO: 165, SEQ ID NO: 166, SEQ ID NO: 167, SEQ ID NO: NO: 168, SEQ ID NO: 169 or SEQ ID NO: 170 having a sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical. In some embodiments, the fusion polypeptide comprises SEQ ID NO: 139, SEQ ID NO: 140, SEQ ID NO: 141, SEQ ID NO: 142, SEQ ID NO: 143, SEQ ID NO: 144, SEQ ID NO: 145, SEQ ID NO: 146, SEQ ID NO: 147, SEQ ID NO: 148, SEQ ID NO: 149, SEQ ID NO: 150, SEQ ID NO: 151, SEQ ID NO: 152, SEQ ID NO: 153, SEQ ID NO: 154, SEQ ID NO: 155, SEQ ID NO: 156, SEQ ID NO: 157, SEQ ID NO: 158, SEQ ID NO: 159, SEQ ID NO: 160, SEQ ID NO: 161, SEQ ID NO: 162, SEQ ID NO: 163, SEQ ID NO: 164, SEQ ID NO: 165, SEQ ID NO: 166, SEQ ID NO: 167, SEQ ID NO: 168, The sequence shown in NO:168, SEQ ID NO:169 or SEQ ID NO:170.

[0049] In one aspect, the present disclosure provides a nucleic acid molecule encoding any one of the fusion polypeptides disclosed herein.

[0050] In another aspect, the present disclosure provides a cell comprising any of the fusion polypeptides disclosed herein.

[0051] In another aspect, the present disclosure provides a cell comprising any of the nucleic acid molecules disclosed herein.

[0052] In some embodiments, the cell further expresses exogenous co-stimulatory factors, immunomodulatory factors, agonists of co-stimulatory factors, antagonists of immunosuppressive factors, immune cell engagers, flip receptors or any combination thereof. In some embodiments, the cell further expresses exogenous lymphocyte receptors or auxiliary receptors. In some embodiments, the exogenous lymphocyte receptors or auxiliary receptors are selected from the group consisting of: TCRα (TCR alpha), TCRβ (TCR beta), TCRγ (TCR gamma), TCRδ (TCR delta), CD4, CD8, pre-T cell receptor α (pTα), Fc receptor α (FcRα), Fc receptor β (FcRβ), Fc receptor γ (FcRγ), natural killer group 2 member D (NKG2D), CD79A, CD79B and any combination thereof. In one embodiment, the cell further expresses exogenous TCR.In some embodiments, the exogenous TCR binds to a target antigen selected from the group consisting of alpha-fetoprotein (AFP), B melanoma antigen (BAGE) family members, brothers of regulatory factors at imprinted sites (BORIS), cancer-testis antigen, cancer-testis antigen 83 (CT-83), carbonic anhydrase IX (CA1X), carcinoembryonic antigen (CEA), cytomegalovirus (CMV) antigen, antigen recognized by cytotoxic T cells (CTL) on melanoma (CAMEL), Epstein-Barr virus (Epstein-Barr virus, EBV) antigen, G antigen 1 (GAGE-1), GAGE-2, GAGE-3, GAGE-4, GAGE-5, GAGE-6, GAGE-7B, GAGE-8, glycoprotein 100 (GP100), hepatitis B virus (HBV) antigen, hepatitis C virus (HCV) nonstructural protein 3 (NS3), human epidermal growth factor receptor 2 (HER-2), human papillomavirus (HPV)-E6, HPV-E7, human telomerase reverse transcriptase (hTERT), IGF2BP3 / A3, K-Ras, K-Ras G12C, K-Ras G12D, K-Ras G12V, latent membrane protein 2 (LMP2), melanoma antigen family A, 1 (MAGE-A1), MAGE-A2, MAGE-A3, MAGE-A4, MAGE-A6, MAGE-A10, MAGE-A12, melanoma antigen recognized by T cells (MART-1), mesothelin (MSLN), mucin 1 (MUC1), mucin 16 (MUC16), New York esophageal squamous cell carcinoma-1 (NYESO-1), P53, P antigen (PAGE) family member, placenta-specific 1 (PLAC1), antigen preferentially expressed in melanoma (PRAME), survivin, synovial sarcoma X1 (SSX1), synovial sarcoma X2 (SSX2), synovial sarcoma X3 (SSX3), synovial sarcoma X4 (SSX4), synovial sarcoma X5 (SSX5), synovial sarcoma X 8 (SSX8), thyroglobulin, tyrosinase, tyrosinase-related protein (TRP) 1, TRP2, Wilms tumor protein (WT-1), X antigen family member 1 (XAGE1) and X antigen family member 2 (XAGE2). In some embodiments, the exogenous TCR is an αβ-TCR or a γδ-TCR. In some embodiments, the cell further expresses a CAR, a CCR or a flip receptor. In some embodiments, the cell further expresses a zetakine, an immune cell engager or a BiTE.

[0053] In one embodiment, the cell is a hematopoietic cell. In some embodiments, the cell is a T cell, an αβ-T cell, or a γδ-T cell. In some embodiments, the cell is a CD3 + cells, CD4 + cells and / or CD8 + Cell. In one embodiment, the cell is an immune effector cell. In some embodiments, the cell is a cytotoxic T lymphocyte (CTL), a tumor infiltrating lymphocyte (TIL) or a helper T cell. In some embodiments, the cell is a natural killer (NK) cell or a natural killer T (NKT) cell. In some embodiments, the source of the cell is a peripheral blood mononuclear cell, bone marrow, lymph node tissue, umbilical cord blood, thymus tissue, tissue from an infection site, ascites, pleural effusion, spleen tissue or a tumor. In one embodiment, the cell is an isolated cell. In one embodiment, the cell is obtained from a subject. In one embodiment, the cell is a human cell.

[0054] On the one hand, the present invention discloses a polypeptide complex comprising a signaling component including a first multimerization domain and an actuator domain; and a targeting component comprising an extracellular domain, a second multimerization domain and a transmembrane domain. In some embodiments, when the signaling component and the targeting component are expressed, the first multimerization domain and the second multimerization domain are located extracellularly.

[0055] In some embodiments, the second multimerization domain and the transmembrane domain are separated by a hinge domain. In some embodiments, the hinge domain is selected from the group consisting of a CD4 hinge, a CD8 hinge, a CD28 hinge, an IgG4 hinge, and any fragment or variant or combination thereof. In some embodiments, the hinge domain is a CD4 hinge. In some embodiments, the CD4 hinge comprises an amino acid sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO: 41. In one embodiment, the CD4 hinge comprises an amino acid sequence as shown in SEQ ID NO: 41. In some embodiments, the hinge domain is a CD28 hinge. In some embodiments, the CD28 hinge comprises an amino acid sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO: 42. In one embodiment, the CD28 hinge comprises an amino acid sequence as shown in SEQ ID NO: 42. In some embodiments, the hinge domain is a CD8 hinge. In some embodiments, the CD8 hinge comprises an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO: 44. In one embodiment, the CD8 hinge comprises an amino acid sequence as shown in SEQ ID NO: 44. In some embodiments, the hinge domain is an IgG4 hinge. In some embodiments, the IgG4 hinge comprises an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO: 43. In one embodiment, the IgG4 hinge comprises an amino acid sequence as shown in SEQ ID NO: 43. In some embodiments, the actuator domain is a CD3 polypeptide, an FcεR1γ polypeptide, an Igα / CD79a polypeptide, an Igβ / CD79b polypeptide, a DAP10 polypeptide or a DAP12 polypeptide. In some embodiments, the CD3 polypeptide is a CD3ε (CD3 epsilon) polypeptide or a fragment or variant thereof, CD3γ (CD3 gamma) or a fragment or variant thereof, or CD3δ (CD3 delta) or a fragment or variant thereof. In some embodiments, the actuator domain is a CD3ε polypeptide comprising an amino acid sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO: 32. In one embodiment, the actuator domain is a CD3ε polypeptide comprising an amino acid sequence as shown in SEQ ID NO: 32. In some embodiments, the actuator domain is a CD3γ polypeptide or a variant thereof.In some embodiments, the actuator domain is a CD3γ polypeptide comprising an amino acid sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity with SEQ ID NO: 33. In one embodiment, the actuator domain is a CD3γ polypeptide comprising an amino acid sequence as shown in SEQ ID NO: 33. In some embodiments, the actuator domain is a CD3δ polypeptide or a variant thereof. In some embodiments, the actuator domain is a CD3δ polypeptide comprising an amino acid sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity with SEQ ID NO: 34. In one embodiment, the actuator domain is a CD3δ polypeptide comprising an amino acid sequence as shown in SEQ ID NO: 34. In some embodiments, the actuator domain is an FcεR1γ polypeptide or a variant thereof. In some embodiments, the actuator domain is an FcεR1γ polypeptide comprising an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:35. In one embodiment, the actuator domain is an FcεR1γ polypeptide comprising an amino acid sequence as shown in SEQ ID NO:35. In some embodiments, the actuator domain is an FcεR1γ polypeptide comprising an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:36. In one embodiment, the actuator domain is an FcεR1γ polypeptide comprising an amino acid sequence as shown in SEQ ID NO:36. In some embodiments, the actuator domain is an Igα / CD79a polypeptide or a variant thereof. In some embodiments, the actuator domain is an Igα / CD79a polypeptide comprising an amino acid sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO:37. In one embodiment, the actuator domain is an Igα / CD79a polypeptide comprising an amino acid sequence as shown in SEQ ID NO:37. In some embodiments, the actuator domain is an Igβ / CD79b polypeptide or a variant thereof. In one embodiment, the actuator domain is an Igβ / CD79b polypeptide comprising an amino acid sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO:38. In one embodiment, the actuator domain is an Igβ / CD79b polypeptide comprising an amino acid sequence as shown in SEQ ID NO:38. In some embodiments, the actuator domain is a DAP10 polypeptide or a variant thereof.In some embodiments, the actuator domain is a DAP10 polypeptide comprising an amino acid sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO:39. In one embodiment, the actuator domain is a DAP10 polypeptide comprising an amino acid sequence as shown in SEQ ID NO:39. In some embodiments, the actuator domain is a DAP12 polypeptide or a variant thereof. In some embodiments, the actuator domain is a DAP12 polypeptide comprising an amino acid sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO:40. In one embodiment, the actuator domain is a DAP12 polypeptide comprising an amino acid sequence as shown in SEQ ID NO:40. In some embodiments, the actuator domain comprises both an extracellular portion and an intracellular portion.

[0056] In some embodiments, the polypeptide further comprises a signal sequence, optionally wherein the signal sequence is at least 90%, 95%, 96%, 97%, 98% or 99% identical to or comprises SEQ ID NO:95, SEQ ID NO:96 or SEQ ID NO:97.

[0057] In some embodiments, the first multimerization domain is different from the second multimerization domain. In some embodiments, the multimerization domains of the signaling component and the targeting component are associated with a bridging factor selected from the group consisting of: rapamycin or a rapamycin analog thereof, gibberellins or derivatives thereof, abscisic acid (ABA) or derivatives thereof, methotrexate or derivatives thereof, cyclosporin A or derivatives thereof, FK506 / cyclosporin A or derivatives thereof, trimethoprim (Tmp) synthetic ligand (SLF) for FK506 binding protein (FKBP) or derivatives thereof, wherein the bridging factor promotes the formation of a polypeptide complex, wherein the bridging factor is associated with the multimerization domains of the signaling component and the targeting component and is positioned between the multimerization domains. In some embodiments, the first multimerization domain and the second multimerization domain are a pair selected from the group consisting of: FK506 binding protein 1A (FKBP12) and FKBP12-rapamycin binding (FRB), FKBP12 and calcineurin, FKBP and cyclophilin A or any other member of the peptidyl-prolyl cis-trans isomerase (PPIase) family, FKBP and dihydrofolate reductase (DHFR), calcineurin and cyclophilin A or any other member of the peptidyl-prolyl cis-trans isomerase (PPIase) family, and PYR1-like 1 (PYL1) and abscisic acid insensitive 1 (ABI1). In some embodiments, the first multimerization domain comprises a first FRB polypeptide or a variant thereof, and the second multimerization domain comprises a first FKBP12 polypeptide or a variant thereof. In some embodiments, the first multimerization domain comprises a first FKBP12 polypeptide or a variant thereof, and the second multimerization domain comprises a first FRB polypeptide or a variant thereof. In one embodiment, the FRB polypeptide is a FRB T2098L variant. In some embodiments, the FRB polypeptide comprises the amino acid sequence as shown in SEQ ID NO: 1 or SEQ ID NO: 2. In some embodiments, the FKBP12 polypeptide comprises the amino acid sequence as shown in SEQ ID NO: 3 or SEQ ID NO: 4.

[0058] In some embodiments, the bridging factor is AP1903, AP20187, AP21967 (also known as C16-(S)-7-methylindorapamycin), everolimus, novolimus, pimecrolimus, ridaforolimus, sirolimus, tacrolimus, temsirolimus, umirolimus, zotarolimus or BPC015. In some embodiments, the first multimerization domain and the second multimerization domain are a pair of antibody-derived heterodimerization domains. In some embodiments, the first multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO:5, or comprises the amino acid sequence as shown in SEQ ID NO:5; and the second multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO:6, or comprises the amino acid sequence as shown in SEQ ID NO:6. In some embodiments, the first multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO:6, or comprises the amino acid sequence as shown in SEQ ID NO:6; and the second multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO:5, or comprises the amino acid sequence as shown in SEQ ID NO:5. In some embodiments, the first multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98%, or 99% identity to SEQ ID NO: 7, or comprises the amino acid sequence as shown in SEQ ID NO: 7; and the second multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98%, or 99% identity to SEQ ID NO: 8, or comprises the amino acid sequence as shown in SEQ ID NO: 8. In some embodiments, the first multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98%, or 99% identity to SEQ ID NO: 8, or comprises the amino acid sequence as shown in SEQ ID NO: 8; and the second multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98%, or 99% identity to SEQ ID NO: 7, or comprises the amino acid sequence as shown in SEQ ID NO: 7.In some embodiments, the first multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98%, or 99% identity to SEQ ID NO: 9, or comprises the amino acid sequence as shown in SEQ ID NO: 9; and the second multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98%, or 99% identity to SEQ ID NO: 10, or comprises the amino acid sequence as shown in SEQ ID NO: 10. In some embodiments, the first multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98%, or 99% identity to SEQ ID NO: 10, or comprises the amino acid sequence as shown in SEQ ID NO: 10; and the second multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98%, or 99% identity to SEQ ID NO: 9, or comprises the amino acid sequence as shown in SEQ ID NO: 9.

[0059] In some embodiments, the first multimerization domain and the actuator domain are separated by a first polypeptide linker having a length of 2 to 40 amino acids. In some embodiments, the first polypeptide linker is selected from the group consisting of: GG, GS, SG, SS, GSS, SSG, GSG, SGS, SGG, G4S, 2xG4S, 3xG4S, 4xG4S, 5xG4S, and any combination thereof. In one embodiment, the first polypeptide linker is a 3xG4S linker. In some embodiments, the first polypeptide linker comprises a polypeptide comprising an amino acid sequence as shown in any one of SEQ ID NOs: 16-31. In some embodiments, the extracellular domain and the second multimerization domain are separated by a second polypeptide linker having a length of 2 to 40 amino acids. In some embodiments, the second polypeptide linker is selected from the group consisting of: GG, GS, SG, SS, GSS, SSG, GSG, SGS, SGG, G4S, 2x G4S, 3xG4s, 4xG4S, and any combination thereof. In one embodiment, the second polypeptide linker is a G4S linker. In some embodiments, the second polypeptide linker comprises a polypeptide comprising an amino acid sequence as shown in any one of SEQ ID NOs: 16-31.

[0060] In some embodiments, the transmembrane domain is a CD4 transmembrane domain. In some embodiments, the CD4 transmembrane domain comprises an amino acid sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity with SEQ ID NO: 45. In one embodiment, the CD4 transmembrane domain comprises an amino acid sequence as shown in SEQ ID NO: 45. In some embodiments, the transmembrane domain is a CD28 transmembrane domain. In some embodiments, the CD28 transmembrane domain comprises an amino acid sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity with SEQ ID NO: 46. In one embodiment, the CD28 transmembrane domain comprises an amino acid sequence as shown in SEQ ID NO: 46. In some embodiments, the transmembrane domain is a CD8 transmembrane domain. In some embodiments, the CD8 transmembrane domain comprises an amino acid sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity with SEQ ID NO: 47. In one embodiment, the CD8 transmembrane domain comprises the amino acid sequence shown in SEQ ID NO:47.

[0061] In some embodiments, the targeting component further comprises an intracellular signaling domain or a co-stimulatory domain derived from a protein selected from the group consisting of: an antigen receptor, a co-stimulatory receptor, a growth receptor, a cytokine receptor, an adaptor signaling protein, an intracellular signaling protein, or any fragment or variant thereof. In some embodiments, the intracellular signaling domain or the co-stimulatory domain on the targeting component is selected from the group consisting of: Toll-like receptor 1 (TLR1), TLR2, TLR3, TLR4, TLR5, TLR6, TLR7, TLR8, TLR9, TLR10, caspase recruitment domain family member 11 (CARD11), CD2, CD3ε, CD3γ, CD3δ, CD4, CD7, CD8, CD27, CD28, CD30, CD40, CD54 (ICAM), CD83, CD94, CD134 (OX40), CD137 (4-1B B), CD278 (ICOS), common gamma chain cytokine, DNAX-activating protein 10 (DAP10), linker for activating T cell family member 1 (LAT), interleukin 2 receptor (IL-2R), IL-4R, IL-7R, IL-9R, IL-12R, IL-13R, IL-15R, IL-21R, 76kD leukocyte protein containing SH2 domain (SLP76), T cell receptor associated transmembrane adaptor 1 (TRAT1), TNFR2, TNFRS14, TNFRS18, TNRFS25 and T cell receptor ζ chain associated protein kinase 70 (ZAP70). In some embodiments, the intracellular signaling domain or the costimulatory domain on the targeting component is selected from the group consisting of 4-1BB, CD28, TNFR2, OX40, ICOS and DAP10 costimulatory domains. In some embodiments, the costimulatory domain on the targeting component is a 4-1BB costimulatory domain, optionally wherein the 4-1BB costimulatory domain comprises an amino acid sequence as shown in SEQ ID NO: 98. In some embodiments, the costimulatory domain on the targeting component is a CD28 costimulatory domain, optionally wherein the CD28 costimulatory domain comprises an amino acid sequence as shown in SEQ ID NO: 99. In some embodiments, the intracellular signaling domain or the costimulatory domain is one or more cytokine receptor intracellular signaling domains. In some embodiments, the one or more cytokine receptor intracellular signaling domains are selected from the group consisting of: an IL7Rα intracellular signaling domain, an IL2Rβ intracellular signaling domain, a common γ chain intracellular signaling domain, and both IL2Rβ and common γ chain intracellular signaling domains.In some embodiments, the IL7Rα intracellular signaling domain comprises an amino acid sequence at least 90%, 95%, 96%, 97%, 98%, or 99% identical to SEQ ID NO: 103. In one embodiment, the IL7Rα intracellular signaling domain comprises an amino acid sequence as shown in SEQ ID NO: 103. In some embodiments, the IL2Rβ intracellular signaling domain comprises an amino acid sequence at least 90%, 95%, 96%, 97%, 98%, or 99% identical to SEQ ID NO: 104 or SEQ ID NO: 105. In some embodiments, the IL2Rβ intracellular signaling domain comprises an amino acid sequence as shown in SEQ ID NO: 104 or SEQ ID NO: 105. In some embodiments, the common gamma chain intracellular signaling domain comprises an amino acid sequence at least 90%, 95%, 96%, 97%, 98%, or 99% identical to SEQ ID NO: 106. In one embodiment, the common gamma chain intracellular signaling domain comprises the amino acid sequence as shown in SEQ ID NO: 106. In some embodiments, the intracellular signaling domain or the costimulatory domain is a LAT domain. In some embodiments, the LAT domain comprises an amino acid sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity to the amino acid sequence as shown in SEQ ID NO: 102. In one embodiment, the LAT domain comprises an amino acid sequence as shown in SEQ ID NO: 102. In some embodiments, the intracellular signaling domain or the costimulatory domain is a CD4 co-receptor domain. In some embodiments, the CD4 co-receptor domain comprises an amino acid sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO: 100. In one embodiment, the CD4 co-receptor domain comprises an amino acid sequence as shown in SEQ ID NO: 100. In some embodiments, the intracellular signaling domain or the costimulatory domain is a CD8 co-receptor domain. In some embodiments, the CD8 co-receptor domain comprises an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO: 101. In one embodiment, the CD8 co-receptor domain comprises an amino acid sequence as shown in SEQ ID NO: 101.

[0062] In some embodiments, the targeting component does not comprise a functional intracellular signaling domain or a co-stimulatory domain. In some embodiments, the targeting component further comprises a truncated intracellular CD4 polypeptide. In some embodiments, the truncated intracellular CD4 polypeptide comprises the amino acid sequence as shown in SEQ ID NO: 48 or SEQ ID NO: 49. In some embodiments, the truncated intracellular CD4 polypeptide comprises the amino acid sequence as shown in SEQ ID NO: 48 or SEQ ID NO: 49.

[0063] In some embodiments, the extracellular domain comprises a first targeting domain. In some embodiments, the first targeting domain comprises a single-chain variable fragment (scFv) or a single domain antibody (sdAb). In some embodiments, the sdAb is a camel VHH, a nano antibody or a heavy chain antibody only (HcAb). In one embodiment, the sdAb is a camel VHH. In some embodiments, the scFv or the sdAb is human or humanized. In some embodiments, the first targeting domain comprises a sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity with any one of SEQ ID NO:50-83. In some embodiments, the first targeting domain comprises a sequence as shown in any one of SEQ ID NO:50-83. In some embodiments, the first targeting domain comprises a PD1 extracellular domain, a human A proliferation-inducing ligand (APRIL), a trimerized human APRIL or a NKG2D membrane protein. In some embodiments, the PD1 extracellular domain comprises a sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO: 84 or SEQ ID NO: 85. In one embodiment, the PD1 extracellular domain comprises a sequence as shown in SEQ ID NO: 85. In some embodiments, the APRIL comprises a sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO: 86, or the trimerized human APRIL comprises a sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO: 87. In some embodiments, the APRIL comprises a sequence as shown in SEQ ID NO: 86, or the trimerized human APRIL comprises a sequence as shown in SEQ ID NO: 87. In some embodiments, the NKG2D membrane protein comprises a sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO: 88. In some embodiments, the NKG2D membrane protein comprises the sequence shown in SEQ ID NO:88.

[0064] In some embodiments, the extracellular domain further comprises a second targeting domain. In some embodiments, the second targeting domain comprises a second single-chain variable fragment (scFv) or a second single domain antibody (sdAb). In some embodiments, the second sdAb is a camel VHH, a nano antibody or a heavy chain antibody only (HcAb). In one embodiment, the second sdAb is a camel VHH. In some embodiments, the second scFv or the second sdAb is human or humanized. In some embodiments, the second targeting domain comprises a sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity with any one of SEQ ID NO:50-83. In some embodiments, the second targeting domain comprises a sequence as shown in any one of SEQ ID NO:50-83. In some embodiments, the second targeting domain comprises a PD1 extracellular domain, a human A proliferation-inducing ligand (APRIL), a trimerized human APRIL or a NKG2D membrane protein. In some embodiments, the PD1 extracellular domain comprises a sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO: 84 or SEQ ID NO: 85. In one embodiment, the PD1 extracellular domain comprises a sequence as shown in SEQ ID NO: 85. In some embodiments, the APRIL comprises a sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO: 86, or the trimerized human APRIL comprises a sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO: 87. In some embodiments, the APRIL comprises a sequence as shown in SEQ ID NO: 86, or the trimerized human APRIL comprises a sequence as shown in SEQ ID NO: 87. In some embodiments, the NKG2D membrane protein comprises a sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO: 88. In one embodiment, the NKG2D membrane protein comprises a sequence as shown in SEQ ID NO: 88. In some embodiments, the first targeting domain and the second targeting domain bind to the same antigen or different antigens. In some embodiments, the targeting domain and the second targeting domain are separated by a third polypeptide linker having a length of 2 to 40 amino acids. In some embodiments, the third polypeptide linker is selected from the group consisting of: GG, GS, SG, SS, GSS, SSG, GSG, SGS, SGG, G4S, 2x G4S, 3xG4s, 4xG4S, 5xG4S, any one of SEQ ID NO: 16-31 and any combination thereof.

[0065] In one embodiment, the signaling component further comprises a signal sequence. In one embodiment, the signal sequence is a CD8 signal sequence. In one embodiment, the CD8 signal sequence comprises an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO: 96. In another embodiment, the CD8 signal sequence comprises an amino acid sequence as shown in SEQ ID NO: 96.

[0066] In another embodiment, the targeting component further comprises a signal sequence. In one embodiment, the signal sequence is an IgK signal sequence. In one embodiment, the IgK signal sequence comprises an amino acid sequence having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO: 95. In one embodiment, the IgK signal sequence comprises an amino acid sequence as shown in SEQ ID NO: 95.

[0067] In some embodiments, the signaling component comprises a sequence that is at least 90%, 95%, 96%, 97%, 98%, or 99% identical to SEQ ID NO: 107, SEQ ID NO: 108, SEQ ID NO: 109, SEQ ID NO: 110, SEQ ID NO: 111, SEQ ID NO: 112, SEQ ID NO: 113, SEQ NO: 114, or SEQ ID NO: 115. In some embodiments, the signaling component comprises a sequence as set forth in SEQ ID NO: 107, SEQ ID NO: 108, SEQ ID NO: 109, SEQ ID NO: 110, SEQ ID NO: 111, SEQ ID NO: 112, SEQ ID NO: 113, SEQ NO: 114, or SEQ ID NO: 115. In some embodiments, the targeting component comprises a sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to the sequence shown in SEQ ID NO:116, SEQ ID NO:117, SEQ ID NO:118, SEQ ID NO:119, SEQ ID NO:120, SEQ ID NO:121, SEQ ID NO:122, SEQ ID NO:123, SEQ ID NO:124, SEQ ID NO:125, SEQ ID NO:126, SEQ ID NO:127, SEQ ID NO:128, SEQ ID NO:129, SEQ ID NO:130, SEQ ID NO:131, SEQ ID NO:132, SEQ ID NO:133, SEQ ID NO:134, SEQ ID NO:135 or SEQ ID NO:136. In some embodiments, the targeting component comprises a sequence as shown in SEQ ID NO:116, SEQ ID NO:117, SEQ ID NO:118, SEQ ID NO:119, SEQ ID NO:120, SEQ ID NO:121, SEQ ID NO:122, SEQ ID NO:123, SEQ ID NO:124, SEQ ID NO:125, SEQ ID NO:126, SEQ ID NO:127, SEQ ID NO:128, SEQID NO:129, SEQ ID NO:130, SEQ ID NO:131, SEQ ID NO:132, SEQ ID NO:133, SEQ ID NO:134, SEQ ID NO:135 or SEQ ID NO:136.

[0068] In some embodiments, the polypeptide complex comprises a fusion polypeptide comprising the targeting component and the signaling component. In some embodiments, the polypeptide complex comprises a polypeptide complex comprising SEQ ID NO: 139, SEQ ID NO: 140, SEQ ID NO: 141, SEQ ID NO: 142, SEQ ID NO: 143, SEQ ID NO: 144, SEQ ID NO: 145, SEQ ID NO: 146, SEQ ID NO: 147, SEQ ID NO: 148, SEQ ID NO: 149, SEQ ID NO: 150, SEQ ID NO: 151, SEQ ID NO: 152, SEQ ID NO: 153, SEQ ID NO: 154, SEQ ID NO: 155, SEQ ID NO: 156, SEQ ID NO: 157, SEQ ID NO: 158, SEQ ID NO: 159, SEQ ID NO: 160, SEQ ID NO: 161, SEQ ID NO: 162, SEQ ID NO: 163, SEQ ID NO: 164, SEQ ID NO: 165, SEQ ID NO: 166, SEQ ID NO: 167, SEQ ID NO: 168, NO: 168, SEQ ID NO: 169 or SEQ ID NO: 170 having a sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical.In some embodiments, the polypeptide complex comprises SEQ ID NO: 139, SEQ ID NO: 140, SEQ ID NO: 141, SEQ ID NO: 142, SEQ ID NO: 143, SEQ ID NO: 144, SEQ ID NO: 145, SEQ ID NO: 146, SEQ ID NO: 147, SEQ ID NO: 148, SEQ ID NO: 149, SEQ ID NO: 150, SEQ ID NO: 151, SEQ ID NO: 152, SEQ ID NO: 153, SEQ ID NO: 154, SEQ ID NO: 155, SEQ ID NO: 156, SEQ ID NO: 157, SEQ ID NO: 158, SEQ ID NO: 159, SEQ ID NO: 160, SEQ ID NO: 161, SEQ ID NO: 162, SEQ ID NO: 163, SEQ ID NO: 164, SEQ ID NO: 165, SEQ ID NO: 166, SEQ ID NO: 167, SEQ ID NO: 168, A fusion polypeptide having the sequence shown in SEQ ID NO:168, SEQ ID NO:169 or SEQ ID NO:170.

[0069] In one aspect, the present disclosure provides a nucleic acid molecule encoding both the signaling component and the targeting component of any of the polypeptide complexes disclosed herein. In another aspect, the present disclosure provides a cell comprising any of the polypeptide complexes disclosed herein. In another aspect, the present disclosure provides a cell comprising any of the nucleic acid molecules disclosed herein.

[0070] In some embodiments, the cell further expresses exogenous co-stimulatory factors, immunomodulatory factors, agonists of co-stimulatory factors, antagonists of immunosuppressive factors, immune cell engagers, flip receptors or any combination thereof. In some embodiments, the cell further expresses exogenous lymphocyte receptors or auxiliary receptors. In some embodiments, the exogenous lymphocyte receptors or auxiliary receptors are selected from the group consisting of: TCRα (TCR alpha), TCRβ (TCR beta), TCRγ (TCR gamma), TCRδ (TCR delta), CD4, CD8, pre-T cell receptor α (pTα), Fc receptor α (FcRα), Fc receptor β (FcRβ), Fc receptor γ (FcRγ), natural killer group 2 member D (NKG2D), CD79A, CD79B and any combination thereof. In some aspects, the cell further expresses exogenous TCR.In some aspects, the exogenous TCR binds to a target antigen selected from the group consisting of alpha-fetoprotein (AFP), B melanoma antigen (BAGE) family members, brothers of regulatory factors at imprinted sites (BORIS), cancer-testis antigen, cancer-testis antigen 83 (CT-83), carbonic anhydrase IX (CA1X), carcinoembryonic antigen (CEA), cytomegalovirus (CMV) antigen, antigen recognized by cytotoxic T cells (CTL) on melanoma (CAMEL), Epstein-Barr virus (EBV) antigen, G antigen 1 (GAGE-1), GAGE-2, GAGE-3, GAGE-4, GAGE-5, GAGE-6, GAGE-7B, GAGE-8, glycoprotein 100 (GP100), hepatitis B virus (HBV) antigen, hepatitis C virus (HCV) nonstructural protein 3 (NS3), human epidermal growth factor receptor 2 (HER-2), human papillomavirus (HPV)-E6, HPV-E7, human telomerase reverse transcriptase (hTERT), IGF2BP3 / A3, K-Ras, K-Ras G12C, K-Ras G12D, K-Ras G12V, latent membrane protein 2 (LMP2), melanoma antigen family A, 1 (MAGE-A1), MAGE-A2, MAGE-A3, MAGE-A4, MAGE-A6, MAGE-A10, MAGE-A12, melanoma antigen recognized by T cells (MART-1), mesothelin (MSLN), mucin 1 (MUC1), mucin 16 (MUC16), New York esophageal squamous cell carcinoma-1 (NYESO-1), P53, P antigen (PAGE) family member, placenta-specific 1 (PLAC1), antigen preferentially expressed in melanoma (PRAME), survivin, synovial sarcoma X1 (SSX1), synovial sarcoma X2 (SSX2), synovial sarcoma X3 (SSX3), synovial sarcoma X4 (SSX4), synovial sarcoma X5 (SSX5), synovial sarcoma X 8 (SSX8), thyroglobulin, tyrosinase, tyrosinase-related protein (TRP) 1, TRP2, Wilms tumor protein (WT-1), X antigen family member 1 (XAGE1) and X antigen family member 2 (XAGE2). In some embodiments, the exogenous TCR is an αβ-TCR or a γδ-TCR.

[0071] In some embodiments, the cell further expresses a CAR, CCR, or flip receptor. In some embodiments, the cell further expresses a zetakine, an immune cell engager, or a BiTE. In one embodiment, the cell is a hematopoietic cell. In some embodiments, the cell is a T cell, an αβ-T cell, or a γδ-T cell. In some embodiments, the cell is a CD3 + cells, CD4+ cells and / or CD8 + Cell. In some embodiments, the cell is an immune effector cell. In some embodiments, the cell is a cytotoxic T lymphocyte (CTL), a tumor infiltrating lymphocyte (TIL) or a helper T cell. In some embodiments, the cell is a natural killer (NK) cell or a natural killer T (NKT) cell. In some embodiments, the source of the cell is a peripheral blood mononuclear cell, bone marrow, lymph node tissue, umbilical cord blood, thymus tissue, tissue from an infection site, ascites, pleural effusion, spleen tissue or a tumor. In one embodiment, the cell is an isolated cell. In one embodiment, the cell is obtained from a subject. In one embodiment, the cell is a human cell.

[0072] On the one hand, the present disclosure provides a polynucleotide encoding the signaling component and the targeting component of any one of the fusion polypeptides or polypeptide complexes disclosed herein. On the other hand, the present disclosure provides a cDNA encoding the signaling component and the targeting component of any one of the fusion polypeptides or polypeptide complexes disclosed herein. On the other hand, the present disclosure provides an RNA encoding the signaling component and the targeting component, or any one of the fusion polypeptides or polypeptide complexes disclosed herein. On the other hand, the present disclosure provides a vector comprising any one of the polynucleotides disclosed herein. In one embodiment, the vector is an expression vector. In one embodiment, the vector is a transposon. In one embodiment, the vector is a piggyBAC transposon or a Sleeping Beauty transposon. In one embodiment, the vector is a viral vector. In some embodiments, the vector is an adenoviral vector, an adeno-associated virus (AAV) vector, a herpes virus vector, a vaccinia virus vector, or a retroviral vector. In some embodiments, the retroviral vector is a lentiviral vector. In some embodiments, the lentiviral vector is selected from the group consisting of: human immunodeficiency virus 1 (HIV-1); human immunodeficiency virus 2 (HIV-2), visna-maedivirus (VMV); caprine arthritis-encephalitis virus (CAEV); equine infectious anemia virus (EIAV); feline immunodeficiency virus (FIV); bovine immunodeficiency virus (BIV); and simian immunodeficiency virus (SIV).

[0073] In one aspect, the present disclosure provides a cell comprising any of the fusion polypeptides, polynucleotides or vectors disclosed herein. In one embodiment, the cell is a hematopoietic cell. In one embodiment, the cell is an immune effector cell. In some embodiments, the cell is a T cell, an αβT cell or a γδT cell. In some embodiments, the cell expresses CD3+ 、CD4 + 、CD8 + Or a combination thereof. In some embodiments, the cell is a cytotoxic T lymphocyte (CTL), a tumor infiltrating lymphocyte (TIL) or a helper T cell. In some embodiments, the cell is a natural killer (NK) cell or a natural killer T (NKT) cell.

[0074] In one aspect, the present disclosure provides a composition comprising any of the cells or vectors disclosed herein. In one aspect, the present disclosure provides a composition comprising a physiologically acceptable carrier and any of the cells or vectors disclosed herein.

[0075] In one aspect, the present disclosure provides a method for treating a subject in need thereof, the method comprising administering to the subject an effective amount of any one of the compositions disclosed herein. In one aspect, the present disclosure provides a method for treating, preventing or ameliorating at least one symptom of cancer, infectious diseases, autoimmune diseases, inflammatory diseases and immunodeficiency or conditions associated therewith, the method comprising administering to the subject an effective amount of any one of the compositions disclosed herein.

[0076] On the other hand, the present disclosure provides a method for treating solid cancer, the method comprising administering to a subject an effective amount of any of the compositions disclosed herein. In some embodiments, the solid cancer is selected from the group consisting of lung cancer, squamous cell carcinoma, colorectal cancer, pancreatic cancer, breast cancer, thyroid cancer, bladder cancer, cervical cancer, esophageal cancer, ovarian cancer, gastric cancer, endometrial cancer, or brain cancer. In some embodiments, the solid cancer is non-small cell lung cancer, head and neck squamous cell carcinoma, colorectal cancer, pancreatic cancer, breast cancer, thyroid cancer, bladder cancer, cervical cancer, esophageal cancer, ovarian cancer, gastric cancer, endometrial cancer, glioma, glioblastoma, or oligodendroglioma.

[0077] On the one hand, the present disclosure provides a method for treating a hematological malignancy, the method comprising administering to a subject an effective amount of any of the compositions disclosed herein. In some embodiments, the hematological malignancy is leukemia, lymphoma or multiple myeloma. In one embodiment, the hematological malignancy is acute myeloid leukemia (AML). BRIEF DESCRIPTION OF THE DRAWINGS

[0078] Figures 1A-1H The architectures of representative engineered immune receptors and fusion constructs are shown. Specifically, Figures 1A-1C Expression constructs and components of several exemplary engineered immune receptors and fusion constructs of the invention are depicted. Figure 1D-1His a sketch depicting how a representative engineered immunoreceptor comprising a signaling component and a targeting component can form a complex with an immune receptor (e.g., TCR) to activate receptor signaling when a target cell expressing a target antigen is present, and in certain embodiments when a bridging factor is present. As a non-limiting example, in Figure 1D In the construct SR022 / SR001, a signaling component including an FRB multimerization domain, a linker, and a CD3ε actuator domain is included; and a targeting component including a CD4 transmembrane domain, a CD4 hinge, a FKBP multimerization domain, a linker, and a CD33 VHH targeting domain. When a cell expressing CD33 is present, the CD33 VHH targeting domain binds to the target cell, bringing it into close contact with an immune cell (e.g., a T cell) expressing an engineered immune receptor. When a bridging factor is present, the multimerization domains then interact, allowing the signaling component to activate the immune receptor (e.g., TCR).

[0079] Figure 2 Shown are the positive percentages and median fluorescence intensity of SR024 components on T cells as measured by flow cytometry.

[0080] Figure 3 Shown are IFNγ, IL2, IL4 and TNF secretion from non-transduced cells and SR024-T cells co-cultured with high CD33 expressing cancer cell lines in the presence of rapamycin.

[0081] Figure 4 Shown is IFNγ secretion from non-transduced cells and SR024-T cells cultured alone in the presence and absence of rapamycin.

[0082] Figure 5A and 5B The cytotoxicity and proliferation of control and SR024 T cells co-cultured with high CD33 expressing cancer cell lines are shown, respectively.

[0083] Figure 6 Shown are the percent positivity and median fluorescence intensity of SR020, SR022 and SR024 components on T cells as measured by flow cytometry.

[0084] Fig. 7A Shown are IFNγ, IL2, IL4 and TNF secretion from untransduced cells, SR020-T cells, SR022-T cells and SR024-T cells, each co-cultured with a high CD33 expressing cancer cell line in the presence of rapamycin.

[0085] Figure 7BShown is IFNγ secretion from untransduced cells, SR020-T cells, SR022-T cells, and SR024-T cells co-cultured with high, medium, and low CD33 expressing cancer cell lines, respectively, in the presence of rapamycin.

[0086] Figure 7C Shown is IL2 secretion from untransduced cells, SR020-T cells, SR022-T cells, and SR024-T cells co-cultured with high, medium, and low CD33 expressing cancer cell lines, respectively, in the presence of rapamycin.

[0087] Figure 8 Shown is IFNγ secretion from untransduced cells alone, SR020-T cells, SR022-T cells, and SR024-T cells in the presence and absence of rapamycin.

[0088] Fig. 9A and 9B The cytotoxicity and proliferation of control, SR020-T cells, SR022-T cells, and SR024-T cells, each co-cultured with a high CD33-expressing cancer cell line, are shown, respectively.

[0089] Fig.10 Shown are the positive percentage and median fluorescence intensity of SR021 components on T cells as measured by flow cytometry.

[0090] Fig.11 Shown are IFNγ, IL2, IL4 and TNF secretion from non-transduced cells and SR021-T cells co-cultured with high CD33 expressing cancer cell lines in the presence of rapamycin.

[0091] Fig.12 Shown is IFNγ secretion from untransduced cells and SR021-T cells co-cultured with individual high CD33 expressing cancer cell lines in the presence and absence of rapamycin.

[0092] Fig.13A and 13B Cytotoxicity and proliferation of control and SR021-T cells co-cultured with high CD33 expressing cancer cell lines are shown, respectively.

[0093] Fig.14 Shown are the percent positivity and median fluorescence intensity of SR022 and SR001 components on T cells as measured by flow cytometry.

[0094] Fig.15Shown are IFNγ, IL2, and TNF secretion from untransduced cells, SR022-T cells, and SR001-T cells, each co-cultured with a high CD33 expressing cancer cell line in the presence of rapamycin.

[0095] Fig.16 Shown is IFNγ secretion from untransduced cells, SR022-T cells, and SR001-T cells each co-cultured with a high CD33 expressing cancer cell line in the presence and absence of rapamycin.

[0096] Fig.17A and 17B The cytotoxicity and proliferation of control and SR001-T cells, each co-cultured with a high CD33-expressing cancer cell line, are shown, respectively.

[0097] Fig.18 Shown are the positive percentages and median fluorescence intensity of SR028 components on T cells as measured by flow cytometry.

[0098] Fig.19 Shown are IFNγ, IL2, IL4 and TNF secretion from non-transduced cells and SR028-T cells co-cultured with high CD33 expressing cancer cell lines in the presence of rapamycin.

[0099] Fig. 20 Shown is IFNγ secretion from non-transduced cells and SR028-T cells co-cultured with high CD33 expressing cancer cell lines in the presence and absence of rapamycin.

[0100] Fig.21A and 21B The cytotoxicity and proliferation of control and SR028-T cells, each co-cultured with a high CD33-expressing cancer cell line, are shown, respectively.

[0101] Fig. 22 Shown are the percent positivity and median fluorescence intensity of SR004 and SR006 components on T cells as measured by flow cytometry.

[0102] Fig.23 Shown are IFNγ, IL2, IL4 and TNF secretion from untransduced cells, SR004-T cells and SR006-T cells, each co-cultured with a high CD33 expressing cancer cell line in the presence of rapamycin.

[0103] Fig.24 Shown is IFNγ secretion from untransduced cells, SR004-T cells, and SR006-T cells each co-cultured with a high CD33 expressing cancer cell line in the presence and absence of rapamycin.

[0104] Fig.25A and 25B The cytotoxicity and proliferation of control, SR004-T cells, and SR006-T cells, each co-cultured with a high CD33-expressing cancer cell line, are shown, respectively.

[0105] Fig.26A and 26B Shown are the percent positivity and median fluorescence intensity of SR008 and SR030 components on T cells as measured by flow cytometry.

[0106] Fig.26C and 26D Shown are the phenotypes of CD4+ T cells and CD8+ T cells transduced with vectors encoding the indicated constructs.

[0107] Fig.27A and 27B Shown are IFNγ, IL2, IL4, and TNF secretion from untransduced cells, SR008-T cells, and SR030-T cells, each co-cultured with a high CD33-expressing cancer cell line or a high CLL1-expressing cancer cell line in the presence of rapamycin.

[0108] Fig.27C Shown is IFNγ secretion from non-transduced cells alone, SR008-T cells, and SR030-T cells in the presence or absence of rapamycin.

[0109] Fig.28A and 28B The cytotoxicity of non-transduced cells, SR008-T cells, and SR030-T cells each co-cultured with a high CD33-expressing cancer cell line or a high CLL1-expressing cancer cell line is shown, respectively.

[0110] Fig.29 Shown are the percent positivity and median fluorescence intensity of SR001 and SR001-28 components on T cells as measured by flow cytometry.

[0111] Fig.30 Shown are IFNγ, IL2, IL4 and TNF secretion from untransduced cells, SR001-T cells and SR001-28-T cells, each co-cultured with a high CD33 expressing cancer cell line in the presence of rapamycin.

[0112] Fig.31A and 31B The cytotoxicity and proliferation of control, SR001-T cells, and SR001-28-T cells, each co-cultured with a high CD33-expressing cancer cell line, are shown, respectively.

[0113] Fig.32A and 32B In vivo tumor growth in NSG mouse xenografts and controls following administration of 10E6 SR001±41BB or CD28 co-stimulatory domains is shown. All tumor controls were regulated by rapamycin.

[0114] Fig.32C and 32D Shown are in vivo tumor growth in NSG mouse xenografts and controls following administration of 3E6 SR001±41BB or CD28 co-stimulatory domains in the presence of rapamycin.

[0115] Fig.33 A and 33B show the percent positivity and median fluorescence intensity of SR022±CD4 or CD8 coreceptor signaling domain as measured by flow cytometry.

[0116] Fig.34 Shown are IFNγ and IL2 secretion from untransduced cells, SR022-T cells ± CD4 or CD8 coreceptor signaling domains, each co-cultured with a high CD33 expressing cancer cell line in the presence or absence of rapamycin.

[0117] Figures 35A-35C Shown are vector copy number, percent positive, and geometric mean fluorescence intensity of tetramer and TCR β chain staining for untransduced TCR, transgenic TCR, SR001, or a combination of transgenic TCR and SR001 as measured by flow cytometry.

[0118] Fig.35D and 35E Shown are the percent positive and geometric mean fluorescence intensity of anti-VHH staining of untransduced TCR, transgenic TCR, SR001, or the combination of transgenic TCR and SR001 as measured by flow cytometry.

[0119] Fig.36A and 36B Shown are IFNγ, IL2, and TNFα secretion from untransduced TCR, transgenic TCR, SR001, or a combination of transgenic TCR and SR001, each co-cultured with high HLA-A2+TCR epitope+ or CD33+ expressing cancer cell lines in the presence and absence of rapamycin.

[0120] Fig.36C The cytotoxicity of non-transduced TCR cells, transgenic TCR cells, SR001-T cells, or a combination of transgenic TCR and SR001 co-cultured with high HLA-A2+TCR epitope+ or CD33+ expressing cancer cell lines in the absence or presence of rapamycin is shown.

[0121] Fig.37A and 37B Shown is IL2 secretion from untransduced or SR001 co-cultured with very low CD33 or CLL1 expressing cancer cell lines, respectively, in the presence of rapamycin.

[0122] Fig.38 In vivo tumor growth in NSG mouse xenografts and controls following administration of 10E6 SR007 or comparator conditioned CAR T cells (targeting CD33 or CLL1) is shown. All tumor controls were conditioned by rapamycin.

[0123] Figures 39A-39C Shown are vector copy number, percentage of positive staining for FRB and VHH expression in the absence or presence of rapamycin.

[0124] Fig.40A and 40B Shown is IFNγ secretion by SR10167 alone or SR10168-T cells ± rapamycin compared to regulated CAR controls.

[0125] Fig.41A and 41B Shown is IFNγ secretion by SR10167 or SR10168-T cells ± rapamycin when cultured with CD19+ target cell lines compared to regulated CAR controls.

[0126] Figures 42A-42D Shown are IL2 and TNFα secretion by SR10167 or SR10168-T cells ± rapamycin when cultured with CD19+ target cell lines compared to conditioned CAR controls.

[0127] Fig.43A and 43B Cytotoxicity of control, SR10167-T cells and regulated CAR comparator-T cells co-cultured with CD19+Jeko-1 or Daudi cell lines, respectively, is shown. All tumor controls were regulated by rapamycin.

[0128] Fig.44 Sketches of representative constitutively active (non-regulatable) engineered immune receptors and fusion constructs are shown.

[0129] Fig.45A and 45B Shown are the positive percentages and median fluorescence intensity of VHH staining for expression of SR292, SR293, SR296, SR001, and relevant comparator molecules, including non-regulated CAR, regulated CAR, and non-regulated TCR-based architecture comparators.

[0130] Fig.46A and 46B Shown are IFNγ and IL2 secretion by SR292, SR293, SR296, SR001 and related comparator molecules (including non-regulated CAR, regulated CAR and non-regulated TCR-based architecture comparators) when cultured with CD33+ target cell lines.

[0131] Fig.47 IFNγ secretion by individual T cells expressing SR292, SR293, SR296, SR001 and related comparator molecules (including non-regulated CAR, regulated CAR and non-regulated TCR-based architecture comparators) is shown.

[0132] Fig.48A and 48B Shown are cytotoxicity and T cell proliferation of control, SR292, SR293, SR296, and a non-regulated CAR after co-culture with a CD33+ target line.

[0133] Fig.49 Shown are the positive percentages and median fluorescence intensity of FRB staining for expression of SR001 ± IL7-receptor α, common γ chain, IL2-receptor β, or both common γ chain and IL2-receptor β.

[0134] Figures 50A-50D Shown are IFNγ and IL2 secretion by SR001 ± IL7-receptor α, common γ chain, IL2-receptor β, or both common γ chain and IL2-receptor β when cultured with CD33+ target cell lines in the presence and absence of rapamycin.

[0135] Fig.51 Shown are T cell proliferation of control or SR001 ± IL7-receptor alpha, common gamma chain, IL2-receptor beta, or both common gamma chain and IL2-receptor beta after culture with CD33+ target lines.

[0136] Fig.52 Shown are the median fluorescence intensities of FRB or PD-1 staining of T cells expressing SR300 (affinity-enhanced PD1) or SR301 (wild-type PD1).

[0137] Figures 53A-53D Shown are the cytotoxicity of T cells expressing SR300 or SR301 when cultured with a PDL1+ target cell line or the same cell line with PDL1 and PDL2 knockout in the presence and absence of rapamycin.

[0138] Fig.54 Shown are the positive percentages and median fluorescence intensity of the expressed VHH staining of the SR354 constructs.

[0139] Fig.55A and 55B Shown is IFNγ secretion by NK cells expressing SR354 when cultured with a CD33+ target cell line or the same cell line with CD33 knockout.

[0140] Fig.55C and 55D Shown is the cytotoxicity of NK cells expressing SR354 when cultured with a CD33+ target cell line or the same cell line with CD33 knockout.

[0141] Fig.56 Shown are the positive percentages of SR303 components on T cells as measured by flow cytometry.

[0142] Fig.57A and 57B Shown is IFNγ secretion by SR303 cells when cultured with the ROR1+ target cell line in the presence and absence of rapamycin.

[0143] Fig.57C IFNγ secretion by individual T cells expressing the SR303 molecule is shown.

[0144] A brief description of sequence identifiers

[0145] SEQ ID NOs: 1-4 show the amino acid sequences of exemplary FRB and FKBP12 polypeptides.

[0146] SEQ ID NOs: 5-10 show the amino acid sequences of exemplary antibody-derived heterodimerization domains.

[0147] SEQ ID NOs: 11-31 show the amino acid sequences of exemplary linkers.

[0148] SEQ ID NOs:32-40 show the amino acid sequences of exemplary actuator domains.

[0149] SEQ ID NOs:41-44 show the amino acid sequences of exemplary hinge domains.

[0150] SEQ ID NOs:45-47 show the amino acid sequences of exemplary membrane-spanning polypeptides.

[0151] Amino acid sequences of exemplary truncated intracellular CD4 polypeptides are shown in SEQ ID NOs: 48 and 49. Amino acid sequences of exemplary targeting domains are shown in SEQ ID NOs: 50-94.

[0152] SEQ ID NOs:95-97 show the amino acid sequences of exemplary signal sequences.

[0153] SEQ ID NOs:98-106 set forth the amino acid sequences of exemplary intracellular signaling domains.

[0154] SEQ ID NOs: 107-115 show the amino acid sequences of exemplary signaling components.

[0155] SEQ ID NOs: 116-136 show the amino acid sequences of exemplary targeting components.

[0156] SEQ ID NOs: 137 and 138 show the amino acid sequences of exemplary co-signaling components.

[0157] SEQ ID NOs: 139-170 show the amino acid sequences of exemplary fusion polypeptides.

[0158] SEQ ID NOs: 171-192 show the amino acid sequences of the protease cleavage site and the self-cleaving polypeptide cleavage site.

[0159] SEQ ID NO: 193 shows the spacer amino acid sequence.

[0160] SEQ ID NO: 194 shows the furin recognition amino acid sequence.

[0161] SEQ ID NOs: 195-197 show the TEV (Tobacco Etch Virus) protease cleavage site.

[0162] SEQ ID NO: 198 shows the kozak amino acid sequence.

[0163] In the foregoing sequences, X, if present, refers to any amino acid or the absence of an amino acid. DETAILED DESCRIPTION

[0164] A. Overview

[0165] Cancer is one of the leading causes of death worldwide. Although adoptive cell therapy has been used to successfully treat some hematological malignancies, the treatment of solid tumors using chimeric antigen receptor (CAR) T cells and T cells expressing T cell receptors (TCRs) specific for tumor antigens remains largely ineffective.

[0166] In addition, although TCRs are known to be very sensitive to low levels of target antigens, they lack the ability to observe non-MHC-presented antigens. On the other hand, although CARs can be engineered to target almost any extracellular antigen through their antibody-like binding domains, they are generally less sensitive to low levels of target antigens and are prone to tonic and / or antigen-independent signaling. In addition, T cells engineered to express CARs or TCRs generally lack spatial and temporal control of T cell activity and / or exhibit insufficient activation of T cell signaling pathways.

[0167] Lack of control over the activity of engineered T cells can trigger a range of side effects, many of which start out small but rapidly worsen. A particularly serious complication is cytokine release syndrome (CRS) or "cytokine storm," in which CAR T cells induce massive and potentially lethal cytokine release. CRS can produce dangerously high fever, extreme fatigue, dyspnea, and a sharp drop in blood pressure. CRS can also produce a second wave of side effects involving the nervous system, including neurotoxicity, tremors, headaches, confusion, loss of balance, difficulty speaking, epilepsy, and hallucinations. Insufficient activation of T cell signaling pathways can result in the inability to eradicate cancer and / or lead to untreatable cancer. The compositions and methods contemplated herein provide solutions to these and other problems that plague adoptive cell therapy.

[0168] Thus, the present disclosure generally relates to improved compositions and methods for modulating the antigen recognition capacity, sensitivity and / or spatial and temporal control of adoptive cell therapy by using engineered immune receptor complexes that bind to selected target antigens and can recruit and activate native or transgenic immune receptor signaling complexes.

[0169] Without wishing to be bound by any particular theory, the engineered immune receptor compositions and methods contemplated herein provide many advantages over existing CAR T cells and TCR T cell therapies in the art, including but not limited to spatial and temporal control of immune effector cell signal transduction, binding and signal transduction activity, and activation of signal transduction pathways without the need for MHC complex recognition. In some embodiments, temporal control triggers the engineered immune receptor signal transduction mechanism by the association of the targeting component and the signal transduction component mediated by the bridging factor. In other embodiments, the mechanism is triggered by the association of the multimerization domain, without the need for a bridging factor. Spatial control participates in the signal transduction mechanism by the recognition of the target antigen by the extracellular domain or the targeting domain of the targeting component, and the signal transduction component comprises an actuator domain that forms a complex with a lymphocyte immune receptor. In this way, when there is a target cell expressing the target antigen, and in certain embodiments, when there is a bridging factor, the immune effector cell activates the receptor signal transduction.

[0170] In various embodiments, the present disclosure contemplates signaling components and targeting components that generate immune receptor-based responses to cells expressing target antigens without recognition of natural immune receptor target antigens. In various embodiments, the present disclosure contemplates signaling components and targeting components that generate immune receptor-based anti-cancer responses to cancers expressing target antigens without recognition of the target antigen and / or MHC complex of a natural or engineered target antigen by a lymphocyte immune receptor.

[0171] In certain embodiments, the engineered immune receptor complex includes a signaling component comprising a multimerization domain polypeptide or a variant thereof and an actuator domain or a variant thereof (e.g., CD3ε, CD3δ, CD3γ, FcεR1γ, Igα / CD79a, Igβ / CD79b, DAP10 or DAP12 polypeptide); and a targeting component comprising an extracellular domain, the extracellular domain comprising a targeting domain that binds to a target antigen expressed on a target cell, a multimerization domain polypeptide or a variant thereof, a transmembrane domain, and a hinge domain that is optionally disposed between the multimerization domain and the transmembrane domain. In some embodiments, in the presence of a bridging factor, the signaling and binding / targeting components associate with each other through the bridging factor to form a functionally active immune receptor. In some embodiments, a bridging factor is not required for component association.

[0172] In certain embodiments, the multimerization domains of the signaling component and the targeting component are located extracellularly. Compared to intracellular localization, extracellular localization of the multimerization domain provides many advantages, including but not limited to more effective localization of the targeting domain, higher temporal sensitivity to bridging factor regulation, and less toxicity due to the ability to use non-immunosuppressive doses of specific bridging factors.

[0173] Contemplated herein are polynucleotides encoding engineered immune receptors, targeting components, signaling components, protein / polypeptide complexes, and fusion proteins; polypeptides comprising engineered immune receptors, targeting components, signaling components, protein / polypeptide complexes, and fusion proteins; cells comprising polynucleotides encoding and / or expressing engineered immune receptors, targeting components, and signaling components; vectors encoding engineered immune receptors, targeting components, signaling components, protein / polypeptide complexes, and fusion proteins; and methods of using the same to treat a disease or disorder (e.g., cancer or an immune disorder).

[0174] Techniques for recombinant (i.e., engineered) DNA, peptide and oligonucleotide synthesis, immunoassays, tissue culture, transformation (e.g., electroporation, lipofection), enzymatic reactions, purification, and related techniques and procedures generally can be performed as described in various general and more specific references in microbiology, molecular biology, biochemistry, molecular genetics, cell biology, virology, and immunology that are cited and discussed throughout the present specification. See, e.g., Sambrook et al., Molecular Cloning: A Laboratory Manual, 3rd ed., Cold Spring Harbor Laboratory Press, Cold Spring Harbor, New York; Current Protocols in Molecular Biology (Wiley and Sons, updated July 2008); Short Protocols in Molecular Biology: A Compendium of Methods from Current Protocols in Molecular Biology, Greene Pub. Associates and Wiley-Interscience; Glover, DNA Cloning: A Practical Approach, Vols. I and II (IRL Press, Oxford Univ. Press USA, 1985); Current Protocols in Immunology: A Compendium of Methods from Current Protocols in Molecular Biology, Greene Pub. Associates and Wiley-Interscience; Glover, DNA Cloning: A Practical Approach, Vols. I and II (IRL Press, Oxford Univ. Press USA, 1985); Current Protocols in Immunology: A Compendium of Methods from Current Protocols in Molecular Biology, Greene Pub. Associates and Wiley-Interscience; Protocols in Immunology)》(Editors: John E.Coligan,Ada M.Kruisbeek,David H.Margulies,EthanM.Shevach, Warren Strober 2001 John Wiley & Sons, New York, NY); Real-Time PCR: Current Technology and Applications, ed. Julie Logan, Kirstin Edwards and Nick Saunders, 2009, Caister Academic Press, Norfolk, UK; Anand, Techniques for the Analysis of Complex Genomes, (Academic Press, New York, 1992); Guthrie and Fink, Guide to Yeast Genetics and Molecular Biology (Academic Press, New York, 1991); Oligonucleotide Synthesis (N. Gait, ed., 1984); Nucleic Acid Hybridization: A Review of the Invention. The Hybridization (B.Hames and S.Higgins, 1985); Transcription and Translation (B.Hames and S.Higgins, 1984); Animal Cell Culture (R.Freshney, ed., 1986); Perbal, A Practical Guide to Molecular Cloning (1984); Next-Generation Genome Sequencing (Janitz, 2008 Wiley-VCH); PCR Protocols (Methods in Molecular Biology) (Park, ed., 3rd ed., 2010, Humana Press); Immobilized Cells and Enzymes (IRL Press, 1986); Methods in Enzymology (Academic Press, Inc., New York); Gene Transfer Vectors for Mammalian Cells (2003). Cells (JH Miller and MP Calos, eds., 1987, Cold Spring Harbor Laboratory Press); Harlow and Lane, Antibodies (Cold Spring Harbor Laboratory Press, Cold Spring Harbor, New York, 1998); Immunochemical Methods in Cell and Molecular Biology (Mayer and Walker, eds., Academic Press, London, 1987); Handbook of Experimental Immunology, Vols. I-IV (DM Weir and CC Blackwell, eds., 1986); Roitt, Essential Immunology, 6th ed. (Blackwell Scientific Publications, Oxford, 1988); Current Protocols in Immunology (Current Protocols in Immunology, 1996). Immunology" (QE Coligan, A.M. Kruisbeek, D.H. Margulies, E.M. Shevach and W. Strober, eds., 1991); Annual Review of Immunology; and monographs in journals such as Advances in Immunology.

[0175] B. Definition

[0176] Before describing the present disclosure in greater detail, it may be helpful to understand the present disclosure by providing definitions of certain terms to be used herein.

[0177] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as those generally understood by those of ordinary skill in the art to which the invention belongs. Although any methods and materials similar to or equivalent to the methods and materials described herein may be used to practice or test specific embodiments, preferred embodiments of compositions, methods and materials are described herein. For purposes of this disclosure, the following terms are defined as follows.

[0178] The articles "a," "an," and "the" are used herein to refer to one or more than one (ie, at least one, or one or more) of the grammatical object of the article. By way of example, "an element" means one element or one or more elements.

[0179] The use of alternatives (eg, "or") should be understood to mean either one, both, or any combination of the alternatives.

[0180] The term "and / or" should be understood to mean one or both of the alternatives.

[0181] As used herein, the term "about" or "approximately" refers to a quantity, level, value, number, frequency, percentage, dimension, size, amount, weight, or length that varies up to 15%, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, or 1% relative to a reference quantity, level, value, number, frequency, percentage, dimension, size, amount, weight, or length. In one embodiment, the term "about" or "approximately" refers to a range of ±15%, ±10%, ±9%, ±8%, ±7%, ±6%, ±5%, ±4%, ±3%, ±2%, or ±1% relative to a reference quantity, level, value, number, frequency, percentage, dimension, size, amount, weight, or length.

[0182] In one embodiment, a range, for example, 1 to 5, about 1 to 5, or about 1 to about 5, refers to every value encompassed within the range. For example, in one non-limiting and exemplary embodiment, the range "1 to 5" is equivalent to stating 1, 2, 3, 4, 5; or 1.0, 1.5, 2.0, 2.5, 3.0, 3.5, 4.0, 4.5, or 5.0; or 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2.0, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9, 3.0, 3.1, 3.2, 3.3, 3.4, 3.5, 3.6, 3.7, 3.8, 3.9, 4.0, 4.1, 4.2, 4.3, 4.4, 4.5, 4.6, 4.7, 4.8, 4.9, or 5.0.

[0183] As used herein, the term "substantially" refers to an amount, level, value, number, frequency, percentage, dimension, size, amount, weight, or length that is 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more of a reference amount, level, value, number, frequency, percentage, dimension, size, amount, weight, or length. In one embodiment, "substantially the same" refers to an effect produced by an amount, level, value, number, frequency, percentage, dimension, size, amount, weight, or length, e.g., a physiological effect that is about the same as a reference amount, level, value, number, frequency, percentage, dimension, size, amount, weight, or length.

[0184] Throughout this specification, unless the context requires otherwise, the term "comprise / comprises / comprising" should be understood to imply the inclusion of the stated step or element or group of steps or elements, but not the exclusion of any other step or element or any other group of steps or elements. "Consisting of" means including and limited to what follows the phrase "consisting of." Thus, the phrase "consisting of" indicates that the listed elements are essential or required, and no other elements can be present. "Consisting essentially of" means including any element listed after the phrase, and is limited to other elements that do not interfere with or affect the activity or action described in the disclosure regarding the listed elements. Thus, the phrase "consisting essentially of" indicates that the listed elements are essential or required, and no other elements are present that substantially affect the activity or action of the listed elements.

[0185] References throughout this specification to "one embodiment," "an embodiment," "a specific embodiment," "a related embodiment," "an embodiment," "another embodiment," or "another embodiment," or combinations thereof, mean that a particular feature, structure, or characteristic described in connection with the embodiment is included in at least one embodiment. Thus, the appearances of the above phrases in various places throughout this specification are not necessarily all referring to the same embodiment. Furthermore, the particular features, structures, or characteristics may be combined in any suitable manner in one or more embodiments. It should also be understood that the affirmative recitation of a feature in one embodiment serves as a basis for excluding the feature in a specific embodiment.

[0186] As used herein, the term "TCR complex" refers to a complex formed by the association of CD3 with TCR. For example, the TCR complex may be composed of: a CD3γ chain, a CD3δ chain, two CD3ε chains, a homodimer of a CD3ζ chain, a TCRα chain, and a TCRβ chain. In some embodiments, the TCR complex may be composed of: a CD3γ chain, a CD3δ chain, two CD3ε chains, a homodimer of a CD3ζ chain, a TCRγ chain, and a TCRδ chain.

[0187] As used herein, "component of a TCR complex" refers to a TCR chain (i.e., TCRα, TCRβ, TCRγ or TCRδ), a CD3 chain (i.e., CD3γ, CD3δ, CD3ε or CD3ζ), or a complex formed by two or more TCR chains or CD3 chains (e.g., a complex of TCRα and TCRβ, a complex of TCRγ and TCRδ, a complex of CD3ε and CD3δ, a complex of CD3γ and CD3ε, or a sub-TCR complex of TCRα, TCRβ, CD3γ, CD3δ and two CD3ε chains).

[0188] As used herein, "actuator polypeptide", "actuator domain" or "actuator" refers to a polypeptide that directly or indirectly associates, integrates or complexes with a multimeric immune receptor complex to promote signal transduction and does not itself contain direct antigen binding properties. In certain embodiments, the actuator domain is part of a protein or protein complex that signals when bound to a target molecule. When the actuator domain contains a signaling domain or motif (such as an immunoreceptor tyrosine-based activation motif (ITAM)), it may directly contribute to the cellular response. In other embodiments, the actuator domain will indirectly promote the cellular response by associating with one or more other proteins that directly signal and thereby promote the cellular response. Exemplary actuator domains include, for example, a CD3 polypeptide, an FcεR1γ polypeptide, an Igα / CD79a polypeptide, an Igβ / CD79b polypeptide, a DAP10 polypeptide, or a DAP12 polypeptide, or any combination thereof.

[0189] As used herein, "multimerization domain" or "multimerization domain polypeptide" refers to a polypeptide that preferentially interacts or associates with another different polypeptide, directly or via a bridging molecule, e.g., a chemically inducible dimer, wherein the interaction of different multimerization domains substantially facilitates or efficiently promotes multimerization (i.e., the formation of a dimer, trimer, or multipart complex, which may be a homodimer, heterodimer, homotrimer, heterotrimer, homomultimer, heteromultimer). A multimerization domain may be derived from natural, synthetic, semisynthetic, or recombinant sources.

[0190] Illustrative examples of multimerization domains suitable for use in specific embodiments contemplated herein include FK506 binding protein (FKBP) polypeptides or variants thereof, FKBP-rapamycin binding (FRB) polypeptides or variants thereof, calcineurin polypeptides or variants thereof, cyclophilin polypeptides or variants thereof, bacterial dihydrofolate reductase (DHFR) polypeptides or variants thereof, PYR1-like 1 (PYL1) polypeptides or variants thereof, abscisic acid insensitive 1 (ABI1) polypeptides or variants thereof, GIB1 polypeptides or variants thereof, or GAI polypeptides or variants thereof.

[0191] As used herein, the term "FKBP-rapamycin binding polypeptide" refers to a FRB polypeptide. In specific embodiments, the FRB polypeptide is a FKBP12-rapamycin binding polypeptide. FRB polypeptides suitable for use in specific embodiments contemplated herein typically contain at least about 85 to about 100 amino acid residues. In certain embodiments, with reference to GenBank accession number L34075.1, the FRB polypeptide comprises a 93 amino acid sequence Ile-2021 to Lys-2113 and a mutation of T2098L (T82L is the equivalent position in the 93 amino acid FRB polypeptide). As used herein, the terms "FRB star", "FRBstar" or "FRB*" refer to such FRB T2098L (T82L) mutants. The FRB polypeptide contemplated herein binds to the FKBP polypeptide via a bridging factor, thereby forming a ternary complex.

[0192] As used herein, the term "FK506 binding protein" refers to a FKBP polypeptide. In a specific embodiment, the FKBP polypeptide is a FKBP12 polypeptide or a FKBP12 polypeptide comprising an F36V mutation. In certain embodiments, the FKBP domain may also be referred to as a "rapamycin binding domain". Information on nucleotide sequences, cloning and other aspects of various FKBP species is known in the art (see, Staendart et al., Nature 346:671, 1990 (human FKBP12); Kay, Biochem. J. 314:361, 1996). The FKBP polypeptide contemplated herein binds to the FRB polypeptide via a bridging factor, thereby forming a ternary complex.

[0193] "Bridging factor" refers to a molecule that associates with two or more multimerization domains and is disposed between the two or more multimerization domains. In specific embodiments, the multimerization domain substantially contributes to or efficiently promotes the formation of a polypeptide complex only in the presence of a bridging factor. In specific embodiments, the multimerization domain does not contribute to or efficiently promote the formation of a polypeptide complex in the absence of a bridging factor. Illustrative examples of bridging factors suitable for use in specific embodiments contemplated herein include, but are not limited to, AP21967, rapamycin (sirolimus) or rapamycin analogs, coumermycin or derivatives thereof, gibberellins or derivatives thereof, abscisic acid (ABA) or derivatives thereof, methotrexate or derivatives thereof, cyclosporin A or derivatives thereof, FKCsA or derivatives thereof, trimethoprim (Tmp) synthetic ligand (SLF) for FKBP or derivatives thereof, or any combination thereof.

[0194] Rapamycin analogs (rapalogs) include, but are not limited to, those disclosed in U.S. Pat. No. 6,649,595, the structure of which is incorporated herein by reference in its entirety. In certain embodiments, the bridging factor is a rapamycin analog that has a significantly lower immunosuppressive effect than rapamycin. In a preferred embodiment, the rapamycin analog is AP21967 (also known as C-16-(S)-7-methylindorapamycin, IC 50 = 10 nM, chemically modified non-immunosuppressive rapamycin analogs). Other exemplary rapamycin analogs suitable for use in the specific embodiments contemplated herein include, but are not limited to, AP1903, AP20187, everolimus, norvolimus, pimecrolimus, defostiolimus, sirolimus, tacrolimus, temsirolimus, usololimus, zotarolimus, or BPC015.

[0195] "Significantly reduced immunosuppressive effect" means at least less than 0.1 to 0.005 times the immunosuppressive effect observed or expected for the same dose measured clinically or in an appropriate in vitro (e.g., inhibition of T cell proliferation) or in vivo surrogate of human immunosuppressive activity.

[0196] A "transmembrane domain" or "TM domain" is a domain that anchors a polypeptide to the plasma membrane of a cell. A TM domain may be derived from natural, synthetic, semisynthetic or recombinant sources.

[0197] As used herein, the term "extracellular domain" refers to a domain or portion of a polypeptide located or present outside a cell. As disclosed herein, the extracellular domain of a targeting component comprises at least one targeting domain, which redirects activation of an immune receptor complex (e.g., TCR) to, for example, one or more selected target antigens on a target cell (e.g., a cancer cell) when the targeting component is associated with a signaling component (e.g., by connecting the multimerization domain of the targeting component to the multimerization domain of the signaling component via a bridging factor, or by association of the multimerization domain of the targeting component without a bridging factor to the multimerization domain of the signaling component).

[0198] The term "effector function" or "effector cell function" refers to a specialized function of an immune effector cell. Effector functions include, but are not limited to, activation, cytokine production, proliferation, and cytotoxic activity, including the release of cytotoxic factors, or other cellular responses elicited by antigen binding to receptors expressed on immune effector cells. An "intracellular signaling domain" or "intracellular domain" refers to a portion of a protein that transduces an effector function signal and directs the cell to perform a specialized function. Although the entire intracellular signaling domain can generally be employed, in many cases it is not necessary to use the entire domain. To the extent that a truncated portion of an intracellular signaling domain is used, such a truncated portion can be used in place of the entire domain, as long as it transduces an effector function signal. The term intracellular signaling domain means any truncated portion that is necessary or sufficient to transduce an effector function signal comprising an intracellular signaling domain.

[0199] It is well known that the signal generated by the TCR alone is insufficient to fully activate T cells and that secondary or co-stimulatory signals are also required. Therefore, T cell activation can be considered to be mediated by two different classes of intracellular signaling domains: primary signaling domains that initiate antigen-dependent primary activation by the TCR (e.g., the TCR / CD3 complex), and co-stimulatory signaling domains that act in an antigen-independent manner to provide secondary or co-stimulatory signals.

[0200] As used herein, the term "costimulatory signaling domain" or "costimulatory domain" refers to the intracellular signaling domain of a costimulatory molecule. Costimulatory molecules are cell surface molecules other than antigen receptors or Fc receptors that provide a second signal required for efficient activation and function of T lymphocytes after binding to an antigen. Exemplary examples of such costimulatory molecules from which costimulatory domains can be separated include, but are not limited to, Toll-like receptor 1 (TLR1), TLR2, TLR3, TLR4, TLR5, TLR6, TLR7, TLR8, TLR9, TLR10, caspase recruitment domain family member 11 (CARD11), CD2, CD3ε, CD3γ, CD3δ, CD7, CD27, CD28, CD30, CD40, CD54 (ICAM), CD83, CD94, CD134 (OX40), CD137 (4-1BB), CD27 8 (ICOS), DNAX-activating protein 10 (DAP10), FYN, adaptor for activating T-cell family member 1 (LAT), SH2 domain-containing leukocyte protein of 76 kD (SLP76), LCK, T-cell receptor-associated transmembrane adaptor 1 (TRAT1), TNFR2, TNF receptor superfamily member 14 (TNFRS14; HVEM), TNF receptor superfamily member 18 (TNFRS18; GITR), TNF receptor superfamily member 25 (TNFRS25; DR3), and T-cell receptor zeta chain-associated protein kinase 70 (ZAP70).

[0201] "Hinge domain" refers to a polypeptide that plays a role in spatially positioning a domain away from the effector cell surface to achieve appropriate cell / cell contact, antigen binding and activation. In particular embodiments, the polypeptide may include one or more hinge domains between the extracellular domain and the transmembrane domain (TM), between the multimerization domain and the transmembrane domain, or between the multimerization domain and the actuator domain. The hinge domain may be derived from a natural, synthetic, semisynthetic or recombinant source. The hinge domain may include the amino acid sequence of a naturally occurring immunoglobulin hinge region or an altered immunoglobulin hinge region.

[0202] "Linker" or "linker polypeptide" refers to a plurality of amino acid residues added between polypeptide domains for proper spacing and conformation of the molecule. In a specific embodiment, the linker is a variable region linker sequence. A "variable region linker sequence" is an amino acid sequence that connects the V H Domain and V LThe linker is a spacer that is compatible with the interaction of the two sub-binding domains, so that the resulting polypeptide retains specific binding affinity for the same target molecule as an antibody comprising the same light chain variable region and heavy chain variable region. In a specific embodiment, the linker separates one or more heavy chain variable domains or light chain variable domains, hinge domains, multimerization domains, transmembrane domains, costimulatory domains, and / or primary signaling domains.

[0203] Illustrative examples of linkers suitable for use in the specific embodiments contemplated herein include, but are not limited to, the following amino acid sequences: GGG; DGGGS (SEQ ID NO: 16); TGEKP (SEQ ID NO: 17) (see, e.g., Liu et al., Proceedings of the National Academy of Sciences of the United States of America (PNAS) 5525-5530 (1997)); GGRR (SEQ ID NO: 18) (Pomerantz et al., 1995, supra); (GGGGS) n , wherein n=1, 2, 3, 4 or 5 (SEQ ID NOs: 11-15) (Kim et al., Proc. Natl. Acad. Sci. USA 93, 1156-1160 (1996)); EGKSSGSGSESKVD (SEQ ID NO: 19) (Chaudhary et al., 1990, Proc. Natl. Acad. Sci. USA 87: 1066-1070); KESGSVSSEQLAQFRSLD (SEQ ID NO: 20) (Bird et al., 1988, Science 242: 423-426); GGRRGGGS (SEQ ID NO: 21); LRQRDGERP (SEQ ID NO: 22); LRQKDGGGSERP (SEQ ID NO: 23); LRQKD (GGGS) 2 ERP (SEQ ID NO: 24). Alternatively, flexible linkers can be rationally designed using computer programs capable of modeling the DNA binding site and the peptide itself (Desjarlais and Berg, Proc. Natl. Acad. Sci. USA 90: 2256-2260 (1993), Proc. Natl. Acad. Sci. USA 91: 11099-11103 (1994)) or by phage display methods. In one embodiment, the linker comprises the following amino acid sequence: GSTSGSGKPGSGEGSTKG (SEQ ID NO: 25) (Cooper et al., Blood, 101 (4): 1637-1644 (2003)).

[0204] "Spacer domain" refers to a polypeptide that separates two domains. In one embodiment, the spacer domain moves or targets the domain away from the effector cell surface to enable appropriate cell / cell contact, antigen binding and activation (Patel et al., Gene Therapy, 1999; 6: 412-419). In a specific embodiment, the spacer domain separates one or more heavy chain variable domains or light chain variable domains, multimerization domains, transmembrane domains, costimulatory domains and / or primary signaling domains. The spacer domain can be derived from a natural, synthetic, semisynthetic or recombinant source. In certain embodiments, the spacer domain is a part of an immunoglobulin, including but not limited to one or more heavy chain constant regions, for example, CH2 and CH3. The spacer domain may include a naturally occurring immunoglobulin hinge region or an altered immunoglobulin hinge region amino acid sequence.

[0205] "Antigen (Ag)" refers to a compound, composition or substance that can stimulate antibody production or T cell response in an animal, including compositions injected or absorbed into an animal (such as compositions including cancer-specific proteins). Exemplary antigens include, but are not limited to, lipids, carbohydrates, polysaccharides, glycoproteins, peptides or nucleic acids. Antigens react with products having specific humoral or cellular immunity, including those induced by heterologous antigens such as the disclosed antigens.

[0206] "Target antigen" or "target antigen of interest" refers to a molecule expressed on the cell surface of a target cell to which a binding domain or targeting domain contemplated herein is designed to bind. In particular embodiments, the target antigen is an epitope of a polypeptide expressed on the surface of a cancer cell.

[0207] As used herein, the term "targeting domain" refers to a domain that provides a polypeptide with the ability to specifically bind to a target antigen of interest, such as a domain of a targeting component. The targeting domain can be derived from a natural, synthetic, semisynthetic or recombinant source. The targeting domain can, for example, comprise an antibody or antigen-binding fragment thereof or, for example, an extracellular domain.

[0208] As used herein, the term "specific binding affinity" or "specifically binds or specifically bound or specific binding" or "specific targeting" describes that a targeting domain binds to a target antigen with a binding affinity greater than background binding. 5 M -1 Affinity or K aThe targeting domain "specifically binds" to the target antigen if the targeting domain binds or associates with the antigen at a specific equilibrium association constant (i.e., the equilibrium association constant for a specific binding interaction in units of 1 / M). In certain embodiments, the targeting domain (or fusion protein comprising the same) binds or associates with the antigen at a specific equilibrium association constant of greater than or equal to about 10 6 M -1 , 10 7 M -1 , 10 8 M -1 , 10 9 M -1 , 10 10 M -1 , 10 11 M -1 , 10 12 M -1 or 10 13 M -1 K a Binds to the target. A "high affinity" targeting domain (or a single-chain fusion protein thereof) refers to a K a For at least 10 7 M 1 , at least 10 8 M -1 , at least 10 9 M -1 , at least 10 10 M -1 , at least 10 11 M -1 , at least 10 12 M -1 , at least 10 13 M -1 or higher targeting domain.

[0209] The terms "selectively binds" or "selectively bound" or "selectively binding" or "selectively targets" describe the preferential binding of a molecule to a target molecule in the presence of multiple off-target molecules (on-target binding).

[0210] "Antibody" refers to a binding agent that is a polypeptide comprising at least a light or heavy chain immunoglobulin variable region that specifically recognizes and binds an epitope of an antigen, such as a lipid, carbohydrate, polysaccharide, glycoprotein, peptide, or nucleic acid containing antigenic determinants, such as those recognized by immune cells.

[0211] "Epitope" or "antigenic determinant" refers to the region of an antigen to which a binding agent binds.

[0212] Antibodies include antigen-binding fragments thereof, such as camel Ig, llama Ig, alpaca Ig, Ig NAR, Fab' fragments, F(ab') 2 fragment, bispecific Fab dimer (Fab2), trispecific Fab trimer (Fab3), Fv, single-chain Fv protein ("scFv"), bi-scFv, (scFv) 2 , miniantibodies, diabodies, triabodies, tetrabodies, disulfide-stabilized Fv proteins ("dsFv") and single domain antibodies (sdAbs, camelid VHHs, nanobodies) as well as the portion of a full-length antibody responsible for antigen binding. The term also includes genetically engineered forms such as chimeric antibodies (e.g., humanized murine antibodies), heteroconjugate antibodies (e.g., bispecific antibodies), and antigen-binding fragments thereof. See also Pierce Catalog and Handbook, 1994-1995 (Pierce Chemical Co., Rockford, IL); Kuby, J., Immunology, 3rd Edition, WH Freeman & Co., New York, 1997.

[0213] As used herein, the term "cancer" generally refers to a class of diseases or conditions in which abnormal cells divide without control and may invade nearby tissue.

[0214] As used herein, the term "malignant" refers to a group of cancer cells that show one or more of uncontrolled growth (i.e., division beyond normal limits), invasion (i.e., invasion and destruction of adjacent tissues), and metastasis (i.e., spread to other locations in the body via the lymph or blood). As used herein, the term "metastasis" refers to the spread of cancer from one part of the body to another. Tumors formed by cells that have spread are called "metastatic tumors" or "metastasis". Metastatic tumors contain cells similar to those in the original (primary) tumor.

[0215] As used herein, the term "benign" or "non-malignant" refers to a tumor that can grow larger but does not spread to other parts of the body. Benign tumors are self-limited and usually do not invade or metastasize.

[0216] "Cancer cell" refers to a single cell of a cancerous growth or tissue. Cancer cells include solid cancers and liquid cancers. "Tumor" or "tumor cell" generally refers to a swelling or lesion formed by an abnormal growth of cells, which can be benign, pre-malignant or malignant. Most cancers form tumors, but liquid cancers such as leukemias do not necessarily form tumors. For those cancers that form tumors, the terms cancer (cell) and tumor (cell) are used interchangeably. The amount of tumor in an individual is the "tumor burden" which can be measured as the number, volume or weight of tumors.

[0217] The term "relapse" refers to the return of cancer diagnosis or signs and symptoms after a period of improvement or remission.

[0218] "Remission" is also called "clinical remission" and includes both partial remission and complete remission. In a partial remission, some but not all signs and symptoms of cancer have disappeared. In a complete remission, all signs and symptoms of cancer have disappeared, although the cancer may still be in the body.

[0219] "Refractory" means that the cancer is resistant or unresponsive to therapy with a particular therapeutic agent. A cancer can be refractory at the start of treatment (i.e., unresponsive to initial exposure to the therapeutic agent), or because it develops resistance to the therapeutic agent during the first treatment period or during subsequent treatment periods.

[0220] "Antigen negative" refers to cells that do not express an antigen or express negligible amounts of an undetectable antigen. In one embodiment, an antigen negative cell does not bind to a receptor directed to the antigen. In one embodiment, an antigen negative cell does not substantially bind to a receptor directed to the antigen.

[0221] As used herein, the terms "individual" and "subject" are often used interchangeably and refer to any animal that exhibits symptoms of cancer or other immune disorders that can be treated with the compositions and methods contemplated elsewhere herein. Suitable subjects (e.g., patients) include experimental animals (such as mice, rats, rabbits, or guinea pigs), farm animals, and domestic animals or pets (such as cats or dogs). Non-human primates and preferably human patients are included. Typical subjects include human patients who have, have been diagnosed as having, or are at risk of having cancer or another immune disorder.

[0222] As used herein, the term "patient" refers to a subject who has been diagnosed as having cancer or another immune disorder that can be treated with the compositions and methods disclosed elsewhere herein.

[0223] As used herein, "treatment" or "treating" includes any beneficial or desired effect on the symptoms or pathology of a disease or pathological condition and may even include a minimal reduction in one or more measurable markers of the disease or condition being treated. Optionally, treatment may involve a reduction in the disease or condition or a delay in the progression of the disease or condition, such as a delay in tumor growth. "Treatment" does not necessarily indicate complete eradication or cure of a disease or condition or its associated symptoms.

[0224] As used herein, "prevent" and similar words such as "prevented / preventing" indicate the use of preventing, inhibiting, or reducing the likelihood of the occurrence or recurrence of a disease or condition. Prevention also refers to delaying the onset or recurrence of a disease or condition or delaying the onset or recurrence of symptoms of a disease or condition. As used herein, "prevention" and similar words also include reducing the intensity, effects, symptoms, and / or burden of a disease or condition before the onset or recurrence of the disease or condition.

[0225] As used herein, the phrase "improving at least one symptom of..." refers to reducing one or more symptoms of the disease or condition being treated in the subject. In certain embodiments, the disease or condition being treated is cancer, wherein the one or more symptoms reduced include, but are not limited to, weakness, fatigue, shortness of breath, easy bruising and bleeding, frequent infections, swollen lymph nodes, abdominal swelling or pain (due to enlarged abdominal organs), bone or joint pain, bone fractures, unintentional weight loss, loss of appetite, night sweats, persistent mild fever, and decreased urination (due to impaired kidney function).

[0226] "Enhancement" or "promotion" or "increase" or "amplification" generally refers to the ability of the composition contemplated herein to produce, induce or cause a greater physiological response (i.e., downstream effect) compared to the response caused by a vehicle or control molecule / composition. A measurable physiological response may include an increase in T cell expansion, activation, persistence, cytokine secretion, and / or an increase in cancer cell killing ability, as well as other aspects apparent from the understanding of the art and the description herein. An "increased" or "enhanced" amount is typically a "statistically significant" amount and may include an increase of 1.1 times, 1.2 times, 1.5 times, 2 times, 3 times, 4 times, 5 times, 6 times, 7 times, 8 times, 9 times, 10 times, 15 times, 20 times, 30 times or more (e.g., 500 times, 1000 times) (including all integers and decimal points therebetween and above 1, e.g., 1.5, 1.6, 1.7, 1.8, etc.) of the response produced by a vehicle or control composition.

[0227] "Reduced" or "diminished" or "lessened" or "reduced" or "mitigated" generally refers to the ability of the compositions contemplated herein to produce, elicit or cause less physiological responses (i.e., downstream effects) than responses elicited by vehicle or control molecules / compositions. A "reduced" or "reduced" amount is generally a "statistically significant" amount and may include a decrease of 1.1-fold, 1.2-fold, 1.5-fold, 2-fold, 3-fold, 4-fold, 5-fold, 6-fold, 7-fold, 8-fold, 9-fold, 10-fold, 15-fold, 20-fold, 30-fold or more (e.g., 500-fold, 1000-fold) (including all integers and decimal points between and above 1, such as 1.5, 1.6, 1.7, 1.8, etc.) of the response produced by vehicle, control compositions, or responses in a particular cell lineage (reference response).

[0228] "Maintain" or "maintain" or "no change" or "no substantial change" or "no substantial decrease" generally refers to the ability of the compositions contemplated herein to produce, elicit or cause a substantially similar or comparable physiological response (i.e., downstream effect) in a cell compared to the response elicited by a vehicle, a control molecule / composition, or a response in a particular cell lineage. A comparable response is one that is not significantly different or measurably different from a reference response.

[0229] Additional definitions are set forth throughout this disclosure.

[0230] C. Engineered immune receptor architecture

[0231] In a specific embodiment, it is envisioned that the cytotoxicity of immune effector cells is redirected toward cells expressing target antigens and recruiting one or more engineered immune receptors of immune receptor complexes. As used herein, the term "engineered immune receptor", "engineered immune receptor complex" or "engineered immune receptor system" refers to one or more non-naturally occurring polypeptides that promote the transduction of immunostimulatory signals in immune effector cells when exposed to a predefined or selected target antigen and associated / multimerized with an immune receptor complex. In certain embodiments, a multimerizing agent or bridging factor is required to promote the multimerization of non-naturally occurring polypeptides, and thereby stimulate the activity and function of immune effector cells by the activation of immune receptor complexes. In a preferred embodiment, an engineered immune receptor or system is a multi-chain chimeric receptor comprising a signaling component associated with a member of an immune receptor complex and a targeting component that redirects or supplements the antigen-specificity of a receptor / complex.

[0232] In one embodiment, the signaling component and the targeting component are expressed from the same cell. In another embodiment, the signaling component and the targeting component are expressed from different cells. In another embodiment, the signaling component is expressed from a cell and the targeting component is supplied exogenously as a polypeptide. In a specific embodiment, the targeting component pre-loaded with a bridging factor is supplied exogenously to a cell expressing the signaling component.

[0233] 1. Signal transduction components

[0234] "Signaling component" refers to a polypeptide comprising one or more multimerization domains or variants thereof and an actuator domain capable of recruiting an immune receptor complex (e.g., TCR, FcR, BCR or NKG2D complex). Such actuator domains include, but are not limited to, 1) a CD3 polypeptide, domain or fragment thereof associated or bound to a TCR complex, such as CD3ε, CD3γ or CD3δ, 2) an FcεR1γ polypeptide chain, domain or fragment thereof associated or bound to an Fc receptor complex, and 3) an Igα / CD79A polypeptide, domain or fragment thereof, 4) an Igβ / CD79B polypeptide, domain or fragment thereof, 5) a DAP10 polypeptide, domain or fragment thereof, and 6) a DAP12 polypeptide, domain or fragment thereof; or any combination thereof.

[0235] In one embodiment, the signaling component does not comprise a primary signaling domain (e.g., CD3z).

[0236] In various embodiments, the actuator domain comprises a CD3 polypeptide or a functional fragment or variant thereof. In some embodiments, the CD3 polypeptide is a CD3ε (CD3 epsilon) polypeptide, domain, or a functional fragment or variant thereof; a CD3γ (CD3 gamma) polypeptide, domain, or a functional fragment or variant thereof or a CD3δ (CD3 delta) polypeptide, domain, or a functional fragment or variant thereof. In some embodiments, the CD3 polypeptide comprises the extracellular and intracellular portions of a CD3ε, CD3γ, or CD3δ polypeptide.

[0237] In various embodiments, the CD3 polypeptide comprises a CD3 epsilon peptide or a functional fragment or variant thereof comprising an amino acid sequence having at least 90% identity with SEQ ID NO: 32. In some embodiments, the CD3 polypeptide comprises a CD3 epsilon peptide or a functional fragment or variant thereof comprising an amino acid sequence having at least 95% identity with SEQ ID NO: 32. In some embodiments, the CD3 polypeptide comprises a CD3 epsilon peptide or a functional fragment or variant thereof comprising an amino acid sequence having at least 96% identity with SEQ ID NO: 32. In some embodiments, the CD3 polypeptide comprises a CD3 epsilon peptide or a functional fragment or variant thereof comprising an amino acid sequence having at least 97% identity with SEQ ID NO: 32. In some embodiments, the CD3 polypeptide comprises a CD3 epsilon peptide or a functional fragment or variant thereof comprising an amino acid sequence having at least 98% identity with SEQ ID NO: 32. In some embodiments, the CD3 polypeptide comprises a CD3 epsilon peptide or a functional fragment or variant thereof comprising an amino acid sequence having at least 99% identity with SEQ ID NO: 32. In a specific embodiment, the CD3 polypeptide comprises a CD3 epsilon peptide comprising an amino acid sequence as shown in SEQ ID NO: 32.

[0238] In various embodiments, the CD3 polypeptide comprises a CD3γ peptide or a functional fragment or variant thereof comprising an amino acid sequence having at least 90% identity with SEQ ID NO: 33. In some embodiments, the CD3 polypeptide comprises a CD3γ peptide or a functional fragment or variant thereof comprising an amino acid sequence having at least 95% identity with SEQ ID NO: 33. In some embodiments, the CD3 polypeptide comprises a CD3γ peptide or a functional fragment or variant thereof comprising an amino acid sequence having at least 96% identity with SEQ ID NO: 33. In some embodiments, the CD3 polypeptide comprises a CD3γ peptide or a functional fragment or variant thereof comprising an amino acid sequence having at least 97% identity with SEQ ID NO: 33. In some embodiments, the CD3 polypeptide comprises a CD3γ peptide or a functional fragment or variant thereof comprising an amino acid sequence having at least 98% identity with SEQ ID NO: 33. In some embodiments, the CD3 polypeptide comprises a CD3γ peptide or a functional fragment or variant thereof comprising an amino acid sequence having at least 99% identity with SEQ ID NO: 33. In a specific embodiment, the CD3 polypeptide comprises a CD3γ peptide comprising an amino acid sequence as shown in SEQ ID NO: 33.

[0239] In various embodiments, the CD3 polypeptide comprises a CD3 δ peptide, or a functional fragment or variant thereof, comprising an amino acid sequence having at least 90% identity with SEQ ID NO: 34. In some embodiments, the CD3 polypeptide comprises a CD3 δ peptide, or a functional fragment or variant thereof, comprising an amino acid sequence having at least 95% identity with SEQ ID NO: 34. In some embodiments, the CD3 polypeptide comprises a CD3 δ peptide, or a functional fragment or variant thereof, comprising an amino acid sequence having at least 96% identity with SEQ ID NO: 34. In some embodiments, the CD3 polypeptide comprises a CD3 δ peptide, or a functional fragment or variant thereof, comprising an amino acid sequence having at least 97% identity with SEQ ID NO: 34. In some embodiments, the CD3 polypeptide comprises a CD3 δ peptide, or a functional fragment or variant thereof, comprising an amino acid sequence having at least 98% identity with SEQ ID NO: 34. In some embodiments, the CD3 polypeptide comprises a CD3 δ peptide, or a functional fragment or variant thereof, comprising an amino acid sequence having at least 99% identity with SEQ ID NO: 34. In a specific embodiment, the CD3 polypeptide comprises a CD3 δ peptide comprising an amino acid sequence as shown in SEQ ID NO: 34.

[0240] In various embodiments, the CD3 polypeptide is a human CD3 polypeptide.

[0241] In certain embodiments, the signaling component comprises a FRB or FRB T2098L multimerization domain, a linker polypeptide, and a CD3ε polypeptide. In certain embodiments, the signaling component comprises an antibody-derived heterodimerization domain, a linker polypeptide, and a CD3ε polypeptide. In certain embodiments, the signaling component comprises a lock and key multimerization domain, a linker polypeptide, and a CD3ε polypeptide. In certain embodiments, the signaling component comprises a DEKK multimerization domain, a linker polypeptide, and a CD3ε polypeptide. In certain embodiments, the signaling component comprises a SEEDbody multimerization domain, a linker polypeptide, and a CD3ε polypeptide.

[0242] In certain embodiments, the signaling component comprises a FRB or FRB T2098L multimerization domain, a linker polypeptide, and a CD3δ polypeptide. In certain embodiments, the signaling component comprises an antibody-derived heterodimerization domain, a linker polypeptide, and a CD3δ polypeptide. In certain embodiments, the signaling component comprises a lock and key multimerization domain, a linker polypeptide, and a CD3δ polypeptide. In certain embodiments, the signaling component comprises a DEKK multimerization domain, a linker polypeptide, and a CD3δ polypeptide. In certain embodiments, the signaling component comprises a SEEDbody multimerization domain, a linker polypeptide, and a CD3δ polypeptide.

[0243] In certain embodiments, the signaling component comprises a FRB or FRB T2098L multimerization domain, a linker polypeptide, and a CD3γ polypeptide. In certain embodiments, the signaling component comprises an antibody-derived heterodimerization domain, a linker polypeptide, and a CD3γ polypeptide. In certain embodiments, the signaling component comprises a lock and key multimerization domain, a linker polypeptide, and a CD3γ polypeptide. In certain embodiments, the signaling component comprises a DEKK multimerization domain, a linker polypeptide, and a CD3γ polypeptide. In certain embodiments, the signaling component comprises a SEEDbody multimerization domain, a linker polypeptide, and a CD3γ polypeptide.

[0244] In various embodiments, the signaling component comprises an amino acid sequence that is at least 90% identical to SEQ ID NO: 107. In one embodiment, the signaling component comprises an amino acid sequence that is at least 95% identical to SEQ ID NO: 107. In one embodiment, the signaling component comprises an amino acid sequence that is at least 96% identical to SEQ ID NO: 107. In one embodiment, the signaling component comprises an amino acid sequence that is at least 97% identical to SEQ ID NO: 107. In one embodiment, the signaling component comprises an amino acid sequence that is at least 98% identical to SEQ ID NO: 107. In one embodiment, the signaling component comprises an amino acid sequence that is at least 99% identical to SEQ ID NO: 107. In a specific embodiment, the signaling component comprises an amino acid sequence as shown in SEQ ID NO: 107.

[0245] In various embodiments, the signaling component comprises an amino acid sequence that is at least 90% identical to SEQ ID NO: 108. In various embodiments, the signaling component comprises an amino acid sequence that is at least 95% identical to SEQ ID NO: 108. In various embodiments, the signaling component comprises an amino acid sequence that is at least 96% identical to SEQ ID NO: 108. In various embodiments, the signaling component comprises an amino acid sequence that is at least 97% identical to SEQ ID NO: 108. In various embodiments, the signaling component comprises an amino acid sequence that is at least 98% identical to SEQ ID NO: 108. In various embodiments, the signaling component comprises an amino acid sequence that is at least 99% identical to SEQ ID NO: 108. In specific embodiments, the signaling component comprises an amino acid sequence as shown in SEQ ID NO: 108.

[0246] In various embodiments, the signaling component comprises an amino acid sequence that is at least 90% identical to SEQ ID NO: 109. In various embodiments, the signaling component comprises an amino acid sequence that is at least 95% identical to SEQ ID NO: 109. In various embodiments, the signaling component comprises an amino acid sequence that is at least 96% identical to SEQ ID NO: 109. In various embodiments, the signaling component comprises an amino acid sequence that is at least 97% identical to SEQ ID NO: 109. In various embodiments, the signaling component comprises an amino acid sequence that is at least 98% identical to SEQ ID NO: 109. In various embodiments, the signaling component comprises an amino acid sequence that is at least 99% identical to SEQ ID NO: 109. In specific embodiments, the signaling component comprises an amino acid sequence as shown in SEQ ID NO: 109.

[0247] In various embodiments, the signaling component comprises an amino acid sequence that is at least 90% identical to SEQ ID NO: 110. In various embodiments, the signaling component comprises an amino acid sequence that is at least 95% identical to SEQ ID NO: 110. In various embodiments, the signaling component comprises an amino acid sequence that is at least 96% identical to SEQ ID NO: 110. In various embodiments, the signaling component comprises an amino acid sequence that is at least 97% identical to SEQ ID NO: 110. In various embodiments, the signaling component comprises an amino acid sequence that is at least 98% identical to SEQ ID NO: 110. In various embodiments, the signaling component comprises an amino acid sequence that is at least 99% identical to SEQ ID NO: 110. In specific embodiments, the signaling component comprises an amino acid sequence as shown in SEQ ID NO: 110.

[0248] In various embodiments, the signaling component comprises an amino acid sequence that is at least 90% identical to SEQ ID NO: 111. In various embodiments, the signaling component comprises an amino acid sequence that is at least 95% identical to SEQ ID NO: 111. In various embodiments, the signaling component comprises an amino acid sequence that is at least 96% identical to SEQ ID NO: 111. In various embodiments, the signaling component comprises an amino acid sequence that is at least 97% identical to SEQ ID NO: 111. In various embodiments, the signaling component comprises an amino acid sequence that is at least 98% identical to SEQ ID NO: 111. In various embodiments, the signaling component comprises an amino acid sequence that is at least 99% identical to SEQ ID NO: 111. In specific embodiments, the signaling component comprises an amino acid sequence as shown in SEQ ID NO: 111.

[0249] In various embodiments, the signaling component comprises an amino acid sequence that is at least 90% identical to SEQ ID NO: 112. In various embodiments, the signaling component comprises an amino acid sequence that is at least 95% identical to SEQ ID NO: 112. In various embodiments, the signaling component comprises an amino acid sequence that is at least 96% identical to SEQ ID NO: 112. In various embodiments, the signaling component comprises an amino acid sequence that is at least 97% identical to SEQ ID NO: 112. In various embodiments, the signaling component comprises an amino acid sequence that is at least 98% identical to SEQ ID NO: 112. In various embodiments, the signaling component comprises an amino acid sequence that is at least 99% identical to SEQ ID NO: 112. In specific embodiments, the signaling component comprises an amino acid sequence as shown in SEQ ID NO: 112.

[0250] In various embodiments, the signaling component comprises an amino acid sequence that is at least 90% identical to SEQ ID NO: 113. In various embodiments, the signaling component comprises an amino acid sequence that is at least 95% identical to SEQ ID NO: 113. In various embodiments, the signaling component comprises an amino acid sequence that is at least 96% identical to SEQ ID NO: 113. In various embodiments, the signaling component comprises an amino acid sequence that is at least 97% identical to SEQ ID NO: 113. In various embodiments, the signaling component comprises an amino acid sequence that is at least 98% identical to SEQ ID NO: 113. In various embodiments, the signaling component comprises an amino acid sequence that is at least 99% identical to SEQ ID NO: 113. In specific embodiments, the signaling component comprises an amino acid sequence as shown in SEQ ID NO: 113.

[0251] In various embodiments, the signaling component comprises an amino acid sequence that is at least 90% identical to SEQ ID NO: 114. In various embodiments, the signaling component comprises an amino acid sequence that is at least 95% identical to SEQ ID NO: 114. In various embodiments, the signaling component comprises an amino acid sequence that is at least 96% identical to SEQ ID NO: 114. In various embodiments, the signaling component comprises an amino acid sequence that is at least 97% identical to SEQ ID NO: 114. In various embodiments, the signaling component comprises an amino acid sequence that is at least 98% identical to SEQ ID NO: 114. In various embodiments, the signaling component comprises an amino acid sequence that is at least 99% identical to SEQ ID NO: 114. In specific embodiments, the signaling component comprises an amino acid sequence as shown in SEQ ID NO: 114.

[0252] In various embodiments, the actuator domain comprises an FcεR1γ polypeptide or a functional fragment or variant thereof. In some embodiments, the FcεR1γ polypeptide comprises an extracellular and intracellular portion of an FcεR1γ polypeptide. In various embodiments, the FcεR1γ polypeptide comprises an amino acid sequence having at least 90% identity to SEQ ID NO: 35. In some embodiments, the FcεR1γ chain polypeptide comprises an amino acid sequence having at least 95% identity to SEQ ID NO: 35. In some embodiments, the FcεR1γ polypeptide comprises an amino acid sequence having at least 96% identity to SEQ ID NO: 35. In some embodiments, the FcεR1γ polypeptide comprises an amino acid sequence having at least 97% identity to SEQ ID NO: 35. In some embodiments, the FcεR1γ polypeptide comprises an amino acid sequence having at least 98% identity to SEQ ID NO: 35. In some embodiments, the FcεR1γ polypeptide comprises an amino acid sequence having at least 99% identity to SEQ ID NO: 35. In a specific embodiment, the FcεR1γ polypeptide comprises an amino acid sequence as shown in SEQ ID NO: 35.

[0253] In various embodiments, the FcεR1γ polypeptide comprises an amino acid sequence that is at least 90% identical to SEQ ID NO: 36. In some embodiments, the FcεR1γ chain polypeptide comprises an amino acid sequence that is at least 95% identical to SEQ ID NO: 36. In some embodiments, the FcεR1γ polypeptide comprises an amino acid sequence that is at least 96% identical to SEQ ID NO: 36. In some embodiments, the FcεR1γ polypeptide comprises an amino acid sequence that is at least 97% identical to SEQ ID NO: 36. In some embodiments, the FcεR1γ polypeptide comprises an amino acid sequence that is at least 98% identical to SEQ ID NO: 36. In some embodiments, the FcεR1γ chain polypeptide comprises an amino acid sequence that is at least 99% identical to SEQ ID NO: 36. In specific embodiments, the FcεR1γ chain polypeptide comprises an amino acid sequence as shown in SEQ ID NO: 36.

[0254] In one embodiment, the signaling component comprises a sequence having at least 90% identity to SEQ ID NO: 115. In one embodiment, the signaling component comprises a sequence having at least 95% identity to SEQ ID NO: 115. In one embodiment, the signaling component comprises a sequence having at least 96% identity to SEQ ID NO: 115. In one embodiment, the signaling component comprises a sequence having at least 97% identity to SEQ ID NO: 115. In one embodiment, the signaling component comprises a sequence having at least 98% identity to SEQ ID NO: 115. In one embodiment, the signaling component comprises a sequence having at least 99% identity to SEQ ID NO: 115. In one embodiment, the signaling component comprises a sequence as set forth in SEQ ID NO: 115.

[0255] In certain embodiments, the signaling component comprises a FRB or FRB T2098L multimerization domain, a linker polypeptide, and an FcεR1γ polypeptide. In certain embodiments, the signaling component comprises an antibody-derived heterologous dimerization domain, a linker polypeptide, and an FcεR1γ polypeptide. In certain embodiments, the signaling component comprises a lock and key multimerization domain, a linker polypeptide, and an FcεR1γ polypeptide. In certain embodiments, the signaling component comprises a DEKK multimerization domain, a linker polypeptide, and an aFcεR1γ chain polypeptide. In certain embodiments, the signaling component comprises a SEEDbody multimerization domain, a linker polypeptide, and an FcεR1γ polypeptide.

[0256] In various embodiments, the signaling component comprises an amino acid sequence that is at least 90% identical to SEQ ID NO: 115. In various embodiments, the signaling component comprises an amino acid sequence that is at least 95% identical to SEQ ID NO: 115. In various embodiments, the signaling component comprises an amino acid sequence that is at least 96% identical to SEQ ID NO: 115. In various embodiments, the signaling component comprises an amino acid sequence that is at least 97% identical to SEQ ID NO: 115. In various embodiments, the signaling component comprises an amino acid sequence that is at least 98% identical to SEQ ID NO: 115. In various embodiments, the signaling component comprises an amino acid sequence that is at least 99% identical to SEQ ID NO: 115. In specific embodiments, the signaling component comprises an amino acid sequence as shown in SEQ ID NO: 115.

[0257] In various embodiments, the actuator domain comprises an Igα / CD79A polypeptide or a functional fragment or variant thereof. In one embodiment, the Igα / CD79A polypeptide comprises a sequence having at least 90% identity to SEQ ID NO:37. In one embodiment, the Igα / CD79A polypeptide comprises a sequence having at least 95% identity to SEQ ID NO:37. In one embodiment, the Igα / CD79A polypeptide comprises a sequence having at least 96% identity to SEQ ID NO:37. In one embodiment, the Igα / CD79A polypeptide comprises a sequence having at least 97% identity to SEQ ID NO:37. In one embodiment, the Igα / CD79A polypeptide comprises a sequence having at least 98% identity to SEQ ID NO:37. In one embodiment, the Igα / CD79A polypeptide comprises a sequence having at least 99% identity to SEQ ID NO:37. In one embodiment, the Igα / CD79A polypeptide comprises a sequence as shown in SEQ ID NO:37.

[0258] In various embodiments, the actuator domain comprises an Igβ / CD79A polypeptide or a functional fragment or variant thereof. In one embodiment, the Igβ / CD79A polypeptide comprises a sequence having at least 90% identity to SEQ ID NO:38. In one embodiment, the Igβ / CD79A polypeptide comprises a sequence having at least 95% identity to SEQ ID NO:38. In one embodiment, the Igβ / CD79A polypeptide comprises a sequence having at least 96% identity to SEQ ID NO:38. In one embodiment, the Igβ / CD79A polypeptide comprises a sequence having at least 97% identity to SEQ ID NO:38. In one embodiment, the Igβ / CD79A polypeptide comprises a sequence having at least 98% identity to SEQ ID NO:38. In one embodiment, the Igβ / CD79A polypeptide comprises a sequence having at least 99% identity to SEQ ID NO:38. In one embodiment, the Igβ / CD79A polypeptide comprises a sequence as shown in SEQ ID NO:38.

[0259] In various embodiments, the actuator domain comprises a DAP10 polypeptide or a functional fragment or variant thereof. In one embodiment, the DAP10 polypeptide comprises a sequence having at least 90% identity to SEQ ID NO:39. In one embodiment, the DAP10 polypeptide comprises a sequence having at least 95% identity to SEQ ID NO:39. In one embodiment, the DAP10 polypeptide comprises a sequence having at least 96% identity to SEQ ID NO:39. In one embodiment, the DAP10 polypeptide comprises a sequence having at least 97% identity to SEQ ID NO:39. In one embodiment, the DAP10 polypeptide comprises a sequence having at least 98% identity to SEQ ID NO:39. In one embodiment, the DAP10 polypeptide comprises a sequence having at least 99% identity to SEQ ID NO:39. In one embodiment, the DAP10 polypeptide comprises a sequence as set forth in SEQ ID NO:39.

[0260] In various embodiments, the actuator domain comprises a DAP12 polypeptide or a functional fragment or variant thereof. In one embodiment, the DAP12 polypeptide comprises a sequence having at least 90% identity to SEQ ID NO:40. In one embodiment, the DAP12 polypeptide comprises a sequence having at least 95% identity to SEQ ID NO:40. In one embodiment, the DAP12 polypeptide comprises a sequence having at least 96% identity to SEQ ID NO:40. In one embodiment, the DAP12 polypeptide comprises a sequence having at least 97% identity to SEQ ID NO:40. In one embodiment, the DAP12 polypeptide comprises a sequence having at least 98% identity to SEQ ID NO:40. In one embodiment, the DAP12 polypeptide comprises a sequence having at least 99% identity to SEQ ID NO:40. In one embodiment, the DAP12 polypeptide comprises a sequence as set forth in SEQ ID NO:40.

[0261] In one embodiment, the signaling component does not comprise the amino acid sequence of SEQ ID NO:111.

[0262] In one embodiment, the signaling component comprises a multimerization domain. In various embodiments, when the signaling component is expressed, the multimerization domain is located extracellularly.

[0263] Illustrative examples of multimerization domains suitable for use in the specific signaling components contemplated herein include, but are not limited to, FKBP polypeptides or variants thereof, FRB polypeptides or variants thereof, calcineurin polypeptides or variants thereof, cyclophilin polypeptides or variants thereof, DHFR polypeptides or variants thereof, PYL1 polypeptides or variants thereof, and ABI1 polypeptides or variants thereof.

[0264] In certain embodiments, the signaling component comprises a FRB polypeptide or a variant thereof. In various embodiments, the signaling component comprises a FRB polypeptide or a variant thereof comprising a T2098L (T82L) mutation. In certain embodiments, the FRB polypeptide comprises an amino acid sequence having at least 90% identity to SEQ ID NO: 1. In certain embodiments, the FRB polypeptide comprises an amino acid sequence having at least 95% identity to SEQ ID NO: 1. In certain embodiments, the FRB polypeptide comprises an amino acid sequence having at least 96% identity to SEQ ID NO: 1. In certain embodiments, the FRB polypeptide comprises an amino acid sequence having at least 97% identity to SEQ ID NO: 1. In certain embodiments, the FRB polypeptide comprises an amino acid sequence having at least 98% identity to SEQ ID NO: 1. In certain embodiments, the FRB polypeptide comprises an amino acid sequence having at least 99% identity to SEQ ID NO: 1. In certain embodiments, the FRB polypeptide comprises an amino acid sequence as shown in SEQ ID NO: 1. In certain embodiments, the FRB polypeptide comprises an amino acid sequence as shown in SEQ ID NO: 2. In specific embodiments, the FRB polypeptide comprises an amino acid sequence that is at least 90% identical to SEQ ID NO:2. In specific embodiments, the FRB polypeptide comprises an amino acid sequence that is at least 95% identical to SEQ ID NO:2. In specific embodiments, the FRB polypeptide comprises an amino acid sequence that is at least 96% identical to SEQ ID NO:2. In specific embodiments, the FRB polypeptide comprises an amino acid sequence that is at least 97% identical to SEQ ID NO:2. In specific embodiments, the FRB polypeptide comprises an amino acid sequence that is at least 98% identical to SEQ ID NO:2. In specific embodiments, the FRB polypeptide comprises an amino acid sequence that is at least 99% identical to SEQ ID NO:2. In specific embodiments, the FRB polypeptide comprises an amino acid sequence as shown in SEQ ID NO:2.

[0265] In certain embodiments, the signaling component comprises a FKBP12 polypeptide or a variant thereof. In various embodiments, the signaling component comprises a FKBP12 polypeptide comprising a F36V mutation. In one embodiment, the FKBP12 polypeptide comprises an amino acid sequence having at least 90% identity to SEQ ID NO: 3. In one embodiment, the FKBP12 polypeptide comprises an amino acid sequence having at least 95% identity to SEQ ID NO: 3. In one embodiment, the FKBP12 polypeptide comprises an amino acid sequence having at least 96% identity to SEQ ID NO: 3. In one embodiment, the FKBP12 polypeptide comprises an amino acid sequence having at least 97% identity to SEQ ID NO: 3. In one embodiment, the FKBP12 polypeptide comprises an amino acid sequence having at least 98% identity to SEQ ID NO: 3. In one embodiment, the FKBP12 polypeptide comprises an amino acid sequence having at least 99% identity to SEQ ID NO: 3. In certain embodiments, the FKBP12 polypeptide comprises an amino acid sequence as shown in SEQ ID NO: 3. In certain embodiments, the FKBP12 polypeptide comprises an amino acid sequence as shown in SEQ ID NO: 4. In one embodiment, the FKBP12 polypeptide comprises an amino acid sequence that is at least 90% identical to SEQ ID NO: 4. In one embodiment, the FKBP12 polypeptide comprises an amino acid sequence that is at least 95% identical to SEQ ID NO: 4. In one embodiment, the FKBP12 polypeptide comprises an amino acid sequence that is at least 96% identical to SEQ ID NO: 4. In one embodiment, the FKBP12 polypeptide comprises an amino acid sequence that is at least 97% identical to SEQ ID NO: 4. In one embodiment, the FKBP12 polypeptide comprises an amino acid sequence that is at least 98% identical to SEQ ID NO: 4. In one embodiment, the FKBP12 polypeptide comprises an amino acid sequence that is at least 99% identical to SEQ ID NO: 4. In a specific embodiment, the FKBP12 polypeptide comprises an amino acid sequence as shown in SEQ ID NO: 4.

[0266] Other illustrative examples of multimerization domains suitable for use in the specific signaling components contemplated herein include antibody-derived heterodimerization domains / pairs. Such antibody heterodimerization domains include, but are not limited to, lock and key, DEKK, SEEDbody, DuoBody, dual variable domain immunoglobulin (DVD-Ig), and tandem Fab immunoglobulin (FIT-Ig) multimerization domains (see, e.g., Ma et al., Front Immunol. 2021 May 5; 12: 626616).

[0267] In various embodiments, the signaling component comprises a lock and key multimerization domain / polypeptide or a variant thereof. In some embodiments, the lock and key multimerization domain / polypeptide or a variant thereof comprises the amino acid sequence as shown in SEQ ID NO: 5. In some embodiments, the lock and key multimerization domain / polypeptide or a variant thereof comprises the amino acid sequence as shown in SEQ ID NO: 6.

[0268] In various embodiments, the signaling component comprises a DEKK multimerization domain / polypeptide or a variant thereof. In some embodiments, the DEKK multimerization domain / polypeptide or a variant thereof comprises the amino acid sequence as shown in SEQ ID NO: 7. In some embodiments, the DEKK multimerization domain / polypeptide or a variant thereof comprises the amino acid sequence as shown in SEQ ID NO: 8.

[0269] In various embodiments, the signaling component comprises a SEEDbody multimerization domain / polypeptide or variant thereof. In some embodiments, the SEEDbody multimerization domain / polypeptide or variant thereof comprises the amino acid sequence as shown in SEQ ID NO: 9. In some embodiments, the SEEDbody multimerization domain / polypeptide or variant thereof comprises the amino acid sequence as shown in SEQ ID NO: 10.

[0270] Other illustrative examples of multimerization domains suitable for use in the specific signaling components contemplated herein include, but are not limited to, leucine zipper multimerization domains, mFos:mJun helical multimerization domains, p53 and MDM2 multimerization domains, VPS36 and VPS28 multimerization domains. Other suitable multimerization or dimerization domains are known to those skilled in the art.

[0271] In various embodiments, the first multimerization domain and the actuator domain are separated by a first polypeptide linker of 2 to 40 amino acids in length. In various embodiments, preferably a short oligopeptide linker or polypeptide linker of 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19 or 20 amino acids in length connects one or more domains in the signaling component. Glycine-serine based linkers provide particularly suitable linkers. In some embodiments, the first polypeptide linker is selected from the group consisting of: GG, GS, SG, SS, GSS, SSG, GSG, SGS, SGG, G4S, 2xG4S, 3xG4S, 4xG4S and any combination thereof. In some embodiments, the first polypeptide linker is a G4S (GGGGS; SEQ ID NO: 11) linker. In some embodiments, the first polypeptide linker is a 2xG4S (GGGGSGGGGS; SEQ ID NO: 12) linker. In some embodiments, the first polypeptide linker is a 3xG4S (GGGGSGGGGSGGGGS; SEQ ID NO: 13) linker. In some embodiments, the first polypeptide linker is a 4xG4S (GGGGSGGGGSGGGGSGGGGS; SEQ ID NO: 14) linker. In some embodiments, the first polypeptide linker is a 5xG4S (GGGGSGGGGSGGGGSGGGGSGGGGS; SEQ ID NO: 15) linker. In one embodiment, the linker has a sequence as set forth in SEQ ID NO: 16. In one embodiment, the linker has a sequence as set forth in SEQ ID NO: 17. In one embodiment, the linker has a sequence as set forth in SEQ ID NO: 18. In one embodiment, the linker has a sequence as set forth in SEQ ID NO: 19. In one embodiment, the linker has a sequence as set forth in SEQ ID NO: 20. In one embodiment, the linker has a sequence as set forth in SEQ ID NO: 21. In one embodiment, the linker has a sequence as set forth in SEQ ID NO: 22. In one embodiment, the linker has a sequence as shown in SEQ ID NO:23. In one embodiment, the linker has a sequence as shown in SEQ ID NO:24. In one embodiment, the linker has a sequence as shown in SEQ ID NO:25. In one embodiment, the linker has a sequence as shown in SEQ ID NO:26. In one embodiment, the linker has a sequence as shown in SEQ ID NO:27. In one embodiment, the linker has a sequence as shown in SEQ ID NO:28. In one embodiment, the linker has a sequence as shown in SEQ ID NO:29. In one embodiment, the linker has a sequence as shown in SEQ ID NO:30.In one embodiment, the linker has the sequence shown in SEQ ID NO:31.

[0272] In various embodiments, the signaling components contemplated herein comprise a signal peptide, such as a secretory signal polypeptide or a signal sequence. Illustrative examples of signal peptides / sequences suitable for use in specific signaling components include, but are not limited to, IgG1 heavy chain signal polypeptides, Igκ light chain signal polypeptides, CD8α signal polypeptides, or human GM-CSF receptor α signal polypeptides. In various embodiments, the signaling component comprises an Igκ light chain signal polypeptide. In some embodiments, the Igκ light chain signal polypeptide comprises an amino acid sequence as shown in SEQ ID NO: 95. In various embodiments, the signaling component comprises a CD8α signal polypeptide. In some embodiments, the CD8α light chain signal polypeptide comprises an amino acid sequence as shown in SEQ ID NO: 96. In various embodiments, the signaling component comprises a PD1 signal polypeptide. In one embodiment, the PD1 signal polypeptide comprises an amino acid sequence as shown in SEQ ID NO: 97.

[0273] 2. Targeting components

[0274] "Targeting component" refers to a polypeptide comprising an extracellular domain, one or more multimerization domains and a transmembrane domain, the conjugation of which with a signaling component redirects activation of an immune receptor complex to one or more selected target antigens. In some embodiments, the extracellular domain comprises one or more targeting domains that associate and / or bind to one or more target antigens. In specific embodiments, the targeting component comprises a first targeting domain, a second multimerization domain, a hinge domain and a transmembrane domain.

[0275] In certain embodiments, the targeting component comprises an extracellular domain comprising at least one targeting domain, wherein the targeting domain is an antibody or antigen-binding fragment thereof to one or more target antigens. Antigen-binding fragments suitable for use in certain embodiments contemplated herein to one or more target antigens include antigen-binding fragments selected from the group consisting of: camel Ig, llama Ig, alpaca Ig, Ig NAR, Fab' fragment, F(ab')2 fragment, bispecific Fab dimer (Fab2), trispecific Fab trimer (Fab3), Fv, single-chain Fv protein ("scFv"), double scFv, (scFv)2, mini-antibodies, bifunctional antibodies, trifunctional antibodies, tetrafunctional antibodies, disulfide-stabilized Fv proteins ("dsFv"), and single domain antibodies (e.g., sdAb, camel VHH or nanobodies).

[0276] In various embodiments, the targeting domain comprises a single chain variable fragment (scFv) or a single domain antibody (sdAb). In various embodiments, the targeting domain comprises a single chain variable fragment (scFv). In certain embodiments, the sdAb is a camel VHH, a nano antibody or a heavy chain antibody only (HcAb). In certain embodiments, the sdAb is a camel VHH. In certain embodiments, the scFv or the sdAb is human or humanized. In certain embodiments, the VHH is humanized.

[0277] The present disclosure provides a platform technology that can be applied to many types and fragments of targeting domains and should not be limited to, for example, one specific targeting domain, such as an antibody, antigen-binding fragment, extracellular domain, etc., for targeting a specific target antigen.

[0278] For example, in some embodiments, the target antigen is an antigen expressed on a target cell (including, for example, a cancer cell). In some embodiments, the targeting domain binds to a target antigen selected from the group consisting of: alpha folate receptor (FRα), alpha v β 6Integrin, B cell maturation antigen (BCMA), B7-H3 (CD276), B7-H6, carbonic anhydrase IX (CAIX), CD16, CD19, CD20, CD22, CD30, CD33, CD37, CD38, CD44, CD44v6, CD44v7 / 8, CD70, CD79a, CD79b, CD123, CD133, CD138, CD171, carcinoembryonic antigen (CEA), C-type lectin-like molecule 1 (CLL-1), CD2 subset 1 (CS-1), chondroitin sulfate proteoglycan 4 (CSPG4), cutaneous T-cell lymphoma-associated antigen 1 (CTAGE1), epidermal growth factor receptor (EGFR), epidermal growth factor receptor 2 (EGFR), EGFR variant III (EGFRvIII), epithelial glycoprotein 2 (EGP2), epithelial glycoprotein 40 (EGP40), epithelial cell adhesion molecule (EPCAM), ephrin type A receptor 2 (EPHA2), fibroblast activation protein (FAP), Fc receptor-like 5 (FCRL5), fetal acetylcholinesterase receptor (AchR), ganglioside G2 (GD2), ganglioside G3 (GD3), glypican-3 (GPC3), EGFR family including ErbB2 (HER2), IGF2BP3 / A3, IL-10Rα, IL-13Rα2, κ, cancer / testis antigen 2 (LAGE-1A), K-Ras, K-Ras G12C, K-Ras G12D, K-Ras G12V, λ, Lewis-Y (Lewis-Y,LeY), L1 cell adhesion molecule (L1-CAM), melanoma antigen gene (MAGE)-A1, MAGE-A3, MAGE-A4, MAGE-A6, MAGEA10, melanoma antigen recognized by T cells 1 (MelanA or MART1), mesothelin (MSLN), MUC1, MUC16, MHC class I chain-associated protein A (MICA), MHC Class I chain-associated protein B (MICB), neural cell adhesion molecule (NCAM), cancer / testis antigen 1 (NY-ESO-1), polysialic acid, placenta-specific 1 (PLAC1), antigen preferentially expressed in melanoma (PRAME), prostate stem cell antigen (PSCA), prostate-specific membrane antigen (PSMA), receptor tyrosine kinase-like orphan receptor 1 (ROR1), synovial sarcoma, breakpoint X 2 (SSX2), survivin, tumor-associated glycoprotein 72 (TAG72), transmembrane activator and CAML interactor (TACI), tumor endothelial marker 1 (TEM1 / CD248), tumor endothelial marker 7-related (TEM7R), trophoblast glycoprotein (TPBG), UL16 binding protein (ULBP) 1, ULBP2, ULBP3, ULBP4, ULBP5, ULBP6, vascular endothelial growth factor receptor 2 (VEGFR2), and Wilms' tumor 1 (WT-1).

[0279] In particular embodiments, the targeting domain binds to a target antigen selected from the group consisting of BCMA, B7-H3 (CD276), CD19, CD20, CD22, CD33, CD79a, CD79b, CD123, CLL-1, EGFR, EGFRvIII, MUC16, and PRAME.

[0280] In particular embodiments, the targeting domain binds to CD33 or CLL-1.

[0281] In various embodiments, the targeting component comprises an extracellular domain comprising a second targeting domain.

[0282] In various embodiments, the second targeting domain comprises a single chain variable fragment (scFv) or a single domain antibody (sdAb). In various embodiments, the second targeting domain comprises a single chain variable fragment (scFv). In certain embodiments, the sdAb is a camel VHH, a nano antibody or a heavy chain antibody only (HcAb). In certain embodiments, the sdAb is a camel VHH. In certain embodiments, the scFv or the sdAb is human or humanized. In certain embodiments, the VHH is humanized.

[0283] In certain embodiments, the extracellular domain contemplated comprises a second targeting domain that binds to a target antigen selected from the group consisting of: alpha folate receptor (FRα), alpha v β 6Integrin, B cell maturation antigen (BCMA), B7-H3 (CD276), B7-H6, carbonic anhydrase IX (CAIX), CD16, CD19, CD20, CD22, CD30, CD33, CD37, CD38, CD44, CD44v6, CD44v7 / 8, CD70, CD79a, CD79b, CD123, CD133, CD138, CD171, carcinoembryonic antigen (CEA), C-type lectin-like molecule 1 (CLL-1), CD2 subset 1 (CS-1), chondroitin sulfate proteoglycan 4 (CSPG4), cutaneous T-cell lymphoma-associated antigen 1 (CTAGE1), epidermal growth factor receptor (EGFR), epidermal growth factor receptor 2 (EGFR), EGF receptor variant III (EGFRvIII), epithelial glycoprotein 2 (EGP2), epithelial glycoprotein 40 (EGP40), epithelial cell adhesion molecule (EPCAM), ephrin type A receptor 2 (EPHA2), fibroblast activation protein (FAP), Fc receptor-like 5 (FCRL5), fetal acetylcholinesterase receptor (AchR), ganglioside G2 (GD2), ganglioside G3 (GD3), glypican-3 (GPC3), EGFR family including ErbB2 (HER2), IGF2BP3 / A3, IL-10Rα, IL-13Rα2, κ, cancer / testis antigen 2 (LAGE-1A), K-Ras, K-Ras G12C, K-RasG12D, K-Ras G12V, λ, Lewis-Y (LeY), L1 cell adhesion molecule (L1-CAM), melanoma antigen gene (MAGE)-A1, MAGE-A3, MAGE-A4, MAGE-A6, MAGEA10, melanoma antigen recognized by T cells 1 (MelanA or MART1), mesothelin (MSLN), MUC1, MUC16, MHC class I chain-associated protein A (MICA), MHCClass I chain-associated protein B (MICB), neural cell adhesion molecule (NCAM), cancer / testis antigen 1 (NY-ESO-1), polysialic acid, placenta-specific 1 (PLAC1), antigen preferentially expressed in melanoma (PRAME), prostate stem cell antigen (PSCA), prostate-specific membrane antigen (PSMA), receptor tyrosine kinase-like orphan receptor 1 (ROR1), synovial sarcoma, breakpoint X 2 (SSX2), survivin, tumor-associated glycoprotein 72 (TAG72), transmembrane activator and CAML interactor (TACI), tumor endothelial marker 1 (TEM1 / CD248), tumor endothelial marker 7-related (TEM7R), trophoblast glycoprotein (TPBG), UL16 binding protein (ULBP) 1, ULBP2, ULBP3, ULBP4, ULBP5, ULBP6, vascular endothelial growth factor receptor 2 (VEGFR2), and Wilms' tumor 1 (WT-1).

[0284] In particular embodiments, the second targeting domain binds to a target antigen selected from the group consisting of BCMA, B7-H3 (CD276), CD19, CD20, CD22, CD33, CD79a, CD79b, CD123, CLL-1, EGFR, EGFRvIII, MUC16, and PRAME.

[0285] In particular embodiments, the second targeting domain binds to CD33 or CLL-1.

[0286] In one embodiment, the targeting domain is an anti-BCMA targeting domain.

[0287] In one embodiment, the targeting domain peptide comprises a sequence that is at least 90% identical to SEQ ID NO:50. In one embodiment, the targeting domain peptide comprises a sequence that is at least 95% identical to SEQ ID NO:50. In one embodiment, the targeting domain peptide comprises a sequence that is at least 96% identical to SEQ ID NO:50. In one embodiment, the targeting domain peptide comprises a sequence that is at least 97% identical to SEQ ID NO:50. In one embodiment, the targeting domain peptide comprises a sequence that is at least 98% identical to SEQ ID NO:50. In one embodiment, the targeting domain peptide comprises a sequence that is at least 99% identical to SEQ ID NO:50. In one embodiment, the targeting domain peptide comprises a sequence as set forth in SEQ ID NO:50.

[0288] In one embodiment, the targeting domain peptide comprises a sequence that is at least 90% identical to SEQ ID NO:51. In one embodiment, the targeting domain peptide comprises a sequence that is at least 95% identical to SEQ ID NO:51. In one embodiment, the targeting domain peptide comprises a sequence that is at least 96% identical to SEQ ID NO:51. In one embodiment, the targeting domain peptide comprises a sequence that is at least 97% identical to SEQ ID NO:51. In one embodiment, the targeting domain peptide comprises a sequence that is at least 98% identical to SEQ ID NO:51. In one embodiment, the targeting domain peptide comprises a sequence that is at least 99% identical to SEQ ID NO:51. In one embodiment, the targeting domain peptide comprises a sequence as set forth in SEQ ID NO:51.

[0289] In one embodiment, the targeting domain peptide comprises a sequence that is at least 90% identical to SEQ ID NO:52. In one embodiment, the targeting domain peptide comprises a sequence that is at least 95% identical to SEQ ID NO:52. In one embodiment, the targeting domain peptide comprises a sequence that is at least 96% identical to SEQ ID NO:52. In one embodiment, the targeting domain peptide comprises a sequence that is at least 97% identical to SEQ ID NO:52. In one embodiment, the targeting domain peptide comprises a sequence that is at least 98% identical to SEQ ID NO:52. In one embodiment, the targeting domain peptide comprises a sequence that is at least 99% identical to SEQ ID NO:52. In one embodiment, the targeting domain peptide comprises a sequence as set forth in SEQ ID NO:52.

[0290] In one embodiment, the targeting domain is a CD19 targeting domain. In one embodiment, the targeting domain peptide comprises a sequence having at least 90% identity to SEQ ID NO:53. In one embodiment, the targeting domain peptide comprises a sequence having at least 95% identity to SEQ ID NO:53. In one embodiment, the targeting domain peptide comprises a sequence having at least 96% identity to SEQ ID NO:53. In one embodiment, the targeting domain peptide comprises a sequence having at least 97% identity to SEQ ID NO:53. In one embodiment, the targeting domain peptide comprises a sequence having at least 98% identity to SEQ ID NO:53. In one embodiment, the targeting domain peptide comprises a sequence having at least 99% identity to SEQ ID NO:53. In one embodiment, the targeting domain peptide comprises a sequence as shown in SEQ ID NO:53.

[0291] In one embodiment, the targeting domain is an nCD20 targeting domain.

[0292] In one embodiment, the targeting domain peptide comprises a sequence that is at least 90% identical to SEQ ID NO:54. In one embodiment, the targeting domain peptide comprises a sequence that is at least 95% identical to SEQ ID NO:54. In one embodiment, the targeting domain peptide comprises a sequence that is at least 96% identical to SEQ ID NO:54. In one embodiment, the targeting domain peptide comprises a sequence that is at least 97% identical to SEQ ID NO:54. In one embodiment, the targeting domain peptide comprises a sequence that is at least 98% identical to SEQ ID NO:54. In one embodiment, the targeting domain peptide comprises a sequence that is at least 99% identical to SEQ ID NO:54. In one embodiment, the targeting domain peptide comprises a sequence as set forth in SEQ ID NO:54.

[0293] In one embodiment, the targeting domain peptide comprises a sequence that is at least 90% identical to SEQ ID NO:55. In one embodiment, the targeting domain peptide comprises a sequence that is at least 95% identical to SEQ ID NO:55. In one embodiment, the targeting domain peptide comprises a sequence that is at least 96% identical to SEQ ID NO:55. In one embodiment, the targeting domain peptide comprises a sequence that is at least 97% identical to SEQ ID NO:55. In one embodiment, the targeting domain peptide comprises a sequence that is at least 98% identical to SEQ ID NO:55. In one embodiment, the targeting domain peptide comprises a sequence that is at least 99% identical to SEQ ID NO:55. In one embodiment, the targeting domain peptide comprises a sequence as set forth in SEQ ID NO:55.

[0294] In one embodiment, the targeting domain peptide comprises a sequence that is at least 90% identical to SEQ ID NO:56. In one embodiment, the targeting domain peptide comprises a sequence that is at least 95% identical to SEQ ID NO:56. In one embodiment, the targeting domain peptide comprises a sequence that is at least 96% identical to SEQ ID NO:56. In one embodiment, the targeting domain peptide comprises a sequence that is at least 97% identical to SEQ ID NO:56. In one embodiment, the targeting domain peptide comprises a sequence that is at least 98% identical to SEQ ID NO:56. In one embodiment, the targeting domain peptide comprises a sequence that is at least 99% identical to SEQ ID NO:56. In one embodiment, the targeting domain peptide comprises a sequence as set forth in SEQ ID NO:56.

[0295] In one embodiment, the targeting domain is a CD22 targeting domain. In one embodiment, the targeting domain peptide comprises a sequence having at least 90% identity to SEQ ID NO:57. In one embodiment, the targeting domain peptide comprises a sequence having at least 95% identity to SEQ ID NO:57. In one embodiment, the targeting domain peptide comprises a sequence having at least 96% identity to SEQ ID NO:57. In one embodiment, the targeting domain peptide comprises a sequence having at least 97% identity to SEQ ID NO:57. In one embodiment, the targeting domain peptide comprises a sequence having at least 98% identity to SEQ ID NO:57. In one embodiment, the targeting domain peptide comprises a sequence having at least 99% identity to SEQ ID NO:57. In one embodiment, the targeting domain peptide comprises a sequence as shown in SEQ ID NO:57.

[0296] In one embodiment, the targeting domain is a CD33 targeting domain. In one embodiment, the targeting domain peptide comprises a sequence having at least 90% identity to SEQ ID NO:58. In one embodiment, the targeting domain peptide comprises a sequence having at least 95% identity to SEQ ID NO:58. In one embodiment, the targeting domain peptide comprises a sequence having at least 96% identity to SEQ ID NO:58. In one embodiment, the targeting domain peptide comprises a sequence having at least 97% identity to SEQ ID NO:58. In one embodiment, the targeting domain peptide comprises a sequence having at least 98% identity to SEQ ID NO:58. In one embodiment, the targeting domain peptide comprises a sequence having at least 99% identity to SEQ ID NO:58. In one embodiment, the targeting domain peptide comprises a sequence as shown in SEQ ID NO:58.

[0297] In one embodiment, the targeting domain is a CD79A targeting domain. In one embodiment, the targeting domain peptide comprises a sequence having at least 90% identity to SEQ ID NO:59. In one embodiment, the targeting domain peptide comprises a sequence having at least 95% identity to SEQ ID NO:59. In one embodiment, the targeting domain peptide comprises a sequence having at least 96% identity to SEQ ID NO:59. In one embodiment, the targeting domain peptide comprises a sequence having at least 97% identity to SEQ ID NO:59. In one embodiment, the targeting domain peptide comprises a sequence having at least 98% identity to SEQ ID NO:59. In one embodiment, the targeting domain peptide comprises a sequence having at least 99% identity to SEQ ID NO:59. In one embodiment, the targeting domain peptide comprises a sequence as shown in SEQ ID NO:59.

[0298] In one embodiment, the targeting domain is an anti-CD79B targeting domain. In one embodiment, the targeting domain peptide comprises a sequence having at least 90% identity to SEQ ID NO:60. In one embodiment, the targeting domain peptide comprises a sequence having at least 95% identity to SEQ ID NO:60. In one embodiment, the targeting domain peptide comprises a sequence having at least 96% identity to SEQ ID NO:60. In one embodiment, the targeting domain peptide comprises a sequence having at least 97% identity to SEQ ID NO:60. In one embodiment, the targeting domain peptide comprises a sequence having at least 98% identity to SEQ ID NO:60. In one embodiment, the targeting domain peptide comprises a sequence having at least 99% identity to SEQ ID NO:60. In one embodiment, the targeting domain peptide comprises a sequence as shown in SEQ ID NO:60.

[0299] In one embodiment, the targeting domain peptide comprises a sequence that is at least 90% identical to SEQ ID NO:61. In one embodiment, the targeting domain peptide comprises a sequence that is at least 95% identical to SEQ ID NO:61. In one embodiment, the targeting domain peptide comprises a sequence that is at least 96% identical to SEQ ID NO:61. In one embodiment, the targeting domain peptide comprises a sequence that is at least 97% identical to SEQ ID NO:61. In one embodiment, the targeting domain peptide comprises a sequence that is at least 98% identical to SEQ ID NO:61. In one embodiment, the targeting domain peptide comprises a sequence that is at least 99% identical to SEQ ID NO:61. In one embodiment, the targeting domain peptide comprises a sequence as set forth in SEQ ID NO:61.

[0300] In one embodiment, the targeting domain is a B7H3 targeting domain. In one embodiment, the targeting domain peptide comprises a sequence having at least 90% identity to SEQ ID NO:62. In one embodiment, the targeting domain peptide comprises a sequence having at least 95% identity to SEQ ID NO:62. In one embodiment, the targeting domain peptide comprises a sequence having at least 96% identity to SEQ ID NO:62. In one embodiment, the targeting domain peptide comprises a sequence having at least 97% identity to SEQ ID NO:62. In one embodiment, the targeting domain peptide comprises a sequence having at least 98% identity to SEQ ID NO:62. In one embodiment, the targeting domain peptide comprises a sequence having at least 99% identity to SEQ ID NO:62. In one embodiment, the targeting domain peptide comprises a sequence as shown in SEQ ID NO:62.

[0301] In one embodiment, the targeting domain is a MUC16 targeting domain. In one embodiment, the targeting domain peptide comprises a sequence having at least 90% identity to SEQ ID NO:63. In one embodiment, the targeting domain peptide comprises a sequence having at least 95% identity to SEQ ID NO:63. In one embodiment, the targeting domain peptide comprises a sequence having at least 96% identity to SEQ ID NO:63. In one embodiment, the targeting domain peptide comprises a sequence having at least 97% identity to SEQ ID NO:63. In one embodiment, the targeting domain peptide comprises a sequence having at least 98% identity to SEQ ID NO:63. In one embodiment, the targeting domain peptide comprises a sequence having at least 99% identity to SEQ ID NO:63. In one embodiment, the targeting domain peptide comprises a sequence as shown in SEQ ID NO:63.

[0302] In one embodiment, the targeting domain is a HER2 targeting domain. In one embodiment, the targeting domain peptide comprises a sequence having at least 90% identity to SEQ ID NO:64. In one embodiment, the targeting domain peptide comprises a sequence having at least 95% identity to SEQ ID NO:64. In one embodiment, the targeting domain peptide comprises a sequence having at least 96% identity to SEQ ID NO:64. In one embodiment, the targeting domain peptide comprises a sequence having at least 97% identity to SEQ ID NO:64. In one embodiment, the targeting domain peptide comprises a sequence having at least 98% identity to SEQ ID NO:64. In one embodiment, the targeting domain peptide comprises a sequence having at least 99% identity to SEQ ID NO:64. In one embodiment, the targeting domain peptide comprises a sequence as shown in SEQ ID NO:64.

[0303] In one embodiment, the targeting domain is an EGFR targeting domain. In one embodiment, the targeting domain peptide comprises a sequence having at least 90% identity to SEQ ID NO:65. In one embodiment, the targeting domain peptide comprises a sequence having at least 95% identity to SEQ ID NO:65. In one embodiment, the targeting domain peptide comprises a sequence having at least 96% identity to SEQ ID NO:65. In one embodiment, the targeting domain peptide comprises a sequence having at least 97% identity to SEQ ID NO:65. In one embodiment, the targeting domain peptide comprises a sequence having at least 98% identity to SEQ ID NO:65. In one embodiment, the targeting domain peptide comprises a sequence having at least 99% identity to SEQ ID NO:65. In one embodiment, the targeting domain peptide comprises a sequence as shown in SEQ ID NO:65.

[0304] In one embodiment, the targeting domain is a FN-EDB targeting domain. In one embodiment, the targeting domain peptide comprises a sequence having at least 90% identity to SEQ ID NO: 66. In one embodiment, the targeting domain peptide comprises a sequence having at least 95% identity to SEQ ID NO: 66. In one embodiment, the targeting domain peptide comprises a sequence having at least 96% identity to SEQ ID NO: 66. In one embodiment, the targeting domain peptide comprises a sequence having at least 97% identity to SEQ ID NO: 66. In one embodiment, the targeting domain peptide comprises a sequence having at least 98% identity to SEQ ID NO: 66. In one embodiment, the targeting domain peptide comprises a sequence having at least 99% identity to SEQ ID NO: 66. In one embodiment, the targeting domain peptide comprises a sequence as set forth in SEQ ID NO: 66.

[0305] In one embodiment, the targeting domain is a CLDN18.2 targeting domain. In one embodiment, the targeting domain peptide comprises a sequence having at least 90% identity to SEQ ID NO: 67. In one embodiment, the targeting domain peptide comprises a sequence having at least 95% identity to SEQ ID NO: 67. In one embodiment, the targeting domain peptide comprises a sequence having at least 96% identity to SEQ ID NO: 67. In one embodiment, the targeting domain peptide comprises a sequence having at least 97% identity to SEQ ID NO: 67. In one embodiment, the targeting domain peptide comprises a sequence having at least 98% identity to SEQ ID NO: 67. In one embodiment, the targeting domain peptide comprises a sequence having at least 99% identity to SEQ ID NO: 67. In one embodiment, the targeting domain peptide comprises a sequence as shown in SEQ ID NO: 67.

[0306] In one embodiment, the targeting domain is a DLL3 targeting domain. In one embodiment, the targeting domain peptide comprises a sequence having at least 90% identity to SEQ ID NO:68. In one embodiment, the targeting domain peptide comprises a sequence having at least 95% identity to SEQ ID NO:68. In one embodiment, the targeting domain peptide comprises a sequence having at least 96% identity to SEQ ID NO:68. In one embodiment, the targeting domain peptide comprises a sequence having at least 97% identity to SEQ ID NO:68. In one embodiment, the targeting domain peptide comprises a sequence having at least 98% identity to SEQ ID NO:68. In one embodiment, the targeting domain peptide comprises a sequence having at least 99% identity to SEQ ID NO:68. In one embodiment, the targeting domain peptide comprises a sequence as set forth in SEQ ID NO:68.

[0307] In one embodiment, the targeting domain is a FLT3 targeting domain. In one embodiment, the targeting domain peptide comprises a sequence having at least 90% identity to SEQ ID NO:69. In one embodiment, the targeting domain peptide comprises a sequence having at least 95% identity to SEQ ID NO:69. In one embodiment, the targeting domain peptide comprises a sequence having at least 96% identity to SEQ ID NO:69. In one embodiment, the targeting domain peptide comprises a sequence having at least 97% identity to SEQ ID NO:69. In one embodiment, the targeting domain peptide comprises a sequence having at least 98% identity to SEQ ID NO:69. In one embodiment, the targeting domain peptide comprises a sequence having at least 99% identity to SEQ ID NO:69. In one embodiment, the targeting domain peptide comprises a sequence as shown in SEQ ID NO:69.

[0308] In one embodiment, the targeting domain peptide comprises a sequence that is at least 90% identical to SEQ ID NO:70. In one embodiment, the targeting domain peptide comprises a sequence that is at least 95% identical to SEQ ID NO:70. In one embodiment, the targeting domain peptide comprises a sequence that is at least 96% identical to SEQ ID NO:70. In one embodiment, the targeting domain peptide comprises a sequence that is at least 97% identical to SEQ ID NO:70. In one embodiment, the targeting domain peptide comprises a sequence that is at least 98% identical to SEQ ID NO:70. In one embodiment, the targeting domain peptide comprises a sequence that is at least 99% identical to SEQ ID NO:70. In one embodiment, the targeting domain peptide comprises a sequence as set forth in SEQ ID NO:70.

[0309] In one embodiment, the targeting domain is a ROR1 targeting domain. In one embodiment, the targeting domain peptide comprises a sequence having at least 90% identity to SEQ ID NO: 71. In one embodiment, the targeting domain peptide comprises a sequence having at least 95% identity to SEQ ID NO: 71. In one embodiment, the targeting domain peptide comprises a sequence having at least 96% identity to SEQ ID NO: 71. In one embodiment, the targeting domain peptide comprises a sequence having at least 97% identity to SEQ ID NO: 71. In one embodiment, the targeting domain peptide comprises a sequence having at least 98% identity to SEQ ID NO: 71. In one embodiment, the targeting domain peptide comprises a sequence having at least 99% identity to SEQ ID NO: 71. In one embodiment, the targeting domain peptide comprises a sequence as shown in SEQ ID NO: 71.

[0310] In one embodiment, the targeting domain is a ROR1 targeting domain. In one embodiment, the targeting domain peptide comprises a sequence having at least 90% identity to SEQ ID NO: 71. In one embodiment, the targeting domain peptide comprises a sequence having at least 95% identity to SEQ ID NO: 71. In one embodiment, the targeting domain peptide comprises a sequence having at least 96% identity to SEQ ID NO: 71. In one embodiment, the targeting domain peptide comprises a sequence having at least 97% identity to SEQ ID NO: 71. In one embodiment, the targeting domain peptide comprises a sequence having at least 98% identity to SEQ ID NO: 71. In one embodiment, the targeting domain peptide comprises a sequence having at least 99% identity to SEQ ID NO: 71. In one embodiment, the targeting domain peptide comprises a sequence as shown in SEQ ID NO: 71.

[0311] In one embodiment, the targeting domain is a CD33 targeting domain. In one embodiment, the targeting domain peptide comprises a sequence having at least 90% identity to SEQ ID NO: 72. In one embodiment, the targeting domain peptide comprises a sequence having at least 95% identity to SEQ ID NO: 72. In one embodiment, the targeting domain peptide comprises a sequence having at least 96% identity to SEQ ID NO: 72. In one embodiment, the targeting domain peptide comprises a sequence having at least 97% identity to SEQ ID NO: 72. In one embodiment, the targeting domain peptide comprises a sequence having at least 98% identity to SEQ ID NO: 72. In one embodiment, the targeting domain peptide comprises a sequence having at least 99% identity to SEQ ID NO: 72. In one embodiment, the targeting domain peptide comprises a sequence as shown in SEQ ID NO: 72.

[0312] In one embodiment, the targeting domain peptide comprises a sequence that is at least 90% identical to SEQ ID NO: 73. In one embodiment, the targeting domain peptide comprises a sequence that is at least 95% identical to SEQ ID NO: 73. In one embodiment, the targeting domain peptide comprises a sequence that is at least 96% identical to SEQ ID NO: 73. In one embodiment, the targeting domain peptide comprises a sequence that is at least 97% identical to SEQ ID NO: 73. In one embodiment, the targeting domain peptide comprises a sequence that is at least 98% identical to SEQ ID NO: 73. In one embodiment, the targeting domain peptide comprises a sequence that is at least 99% identical to SEQ ID NO: 73. In one embodiment, the targeting domain peptide comprises a sequence as set forth in SEQ ID NO: 73.

[0313] In one embodiment, the targeting domain peptide comprises a sequence that is at least 90% identical to SEQ ID NO: 74. In one embodiment, the targeting domain peptide comprises a sequence that is at least 95% identical to SEQ ID NO: 74. In one embodiment, the targeting domain peptide comprises a sequence that is at least 96% identical to SEQ ID NO: 74. In one embodiment, the targeting domain peptide comprises a sequence that is at least 97% identical to SEQ ID NO: 74. In one embodiment, the targeting domain peptide comprises a sequence that is at least 98% identical to SEQ ID NO: 74. In one embodiment, the targeting domain peptide comprises a sequence that is at least 99% identical to SEQ ID NO: 74. In one embodiment, the targeting domain peptide comprises a sequence as set forth in SEQ ID NO: 74.

[0314] In one embodiment, the targeting domain is a CLL1 targeting domain. In one embodiment, the targeting domain peptide comprises a sequence having at least 90% identity to SEQ ID NO: 75. In one embodiment, the targeting domain peptide comprises a sequence having at least 95% identity to SEQ ID NO: 75. In one embodiment, the targeting domain peptide comprises a sequence having at least 96% identity to SEQ ID NO: 75. In one embodiment, the targeting domain peptide comprises a sequence having at least 97% identity to SEQ ID NO: 75. In one embodiment, the targeting domain peptide comprises a sequence having at least 98% identity to SEQ ID NO: 75. In one embodiment, the targeting domain peptide comprises a sequence having at least 99% identity to SEQ ID NO: 75. In one embodiment, the targeting domain peptide comprises a sequence as shown in SEQ ID NO: 75.

[0315] In one embodiment, the targeting domain peptide comprises a sequence that is at least 90% identical to SEQ ID NO: 76. In one embodiment, the targeting domain peptide comprises a sequence that is at least 95% identical to SEQ ID NO: 76. In one embodiment, the targeting domain peptide comprises a sequence that is at least 96% identical to SEQ ID NO: 76. In one embodiment, the targeting domain peptide comprises a sequence that is at least 97% identical to SEQ ID NO: 76. In one embodiment, the targeting domain peptide comprises a sequence that is at least 98% identical to SEQ ID NO: 76. In one embodiment, the targeting domain peptide comprises a sequence that is at least 99% identical to SEQ ID NO: 76. In one embodiment, the targeting domain peptide comprises a sequence as set forth in SEQ ID NO: 76.

[0316] In one embodiment, the targeting domain is a CD123 targeting domain. In one embodiment, the targeting domain peptide comprises a sequence with at least 90% identity to SEQ ID NO:77. In one embodiment, the targeting domain peptide comprises a sequence with at least 95% identity to SEQ ID NO:77. In one embodiment, the targeting domain peptide comprises a sequence with at least 96% identity to SEQ ID NO:77. In one embodiment, the targeting domain peptide comprises a sequence with at least 97% identity to SEQ ID NO:77. In one embodiment, the targeting domain peptide comprises a sequence with at least 98% identity to SEQ ID NO:77. In one embodiment, the targeting domain peptide comprises a sequence with at least 99% identity to SEQ ID NO:77. In one embodiment, the targeting domain peptide comprises a sequence as shown in SEQ ID NO:77.

[0317] In one embodiment, the targeting domain is a CD123 targeting domain. In one embodiment, the targeting domain peptide comprises a sequence with at least 90% identity to SEQ ID NO:78. In one embodiment, the targeting domain peptide comprises a sequence with at least 95% identity to SEQ ID NO:78. In one embodiment, the targeting domain peptide comprises a sequence with at least 96% identity to SEQ ID NO:78. In one embodiment, the targeting domain peptide comprises a sequence with at least 97% identity to SEQ ID NO:78. In one embodiment, the targeting domain peptide comprises a sequence with at least 98% identity to SEQ ID NO:78. In one embodiment, the targeting domain peptide comprises a sequence with at least 99% identity to SEQ ID NO:78. In one embodiment, the targeting domain peptide comprises a sequence as shown in SEQ ID NO:78.

[0318] In one embodiment, the targeting domain is a CD20 targeting domain. In one embodiment, the targeting domain peptide comprises a sequence having at least 90% identity to SEQ ID NO: 79. In one embodiment, the targeting domain peptide comprises a sequence having at least 95% identity to SEQ ID NO: 79. In one embodiment, the targeting domain peptide comprises a sequence having at least 96% identity to SEQ ID NO: 79. In one embodiment, the targeting domain peptide comprises a sequence having at least 97% identity to SEQ ID NO: 79. In one embodiment, the targeting domain peptide comprises a sequence having at least 98% identity to SEQ ID NO: 79. In one embodiment, the targeting domain peptide comprises a sequence having at least 99% identity to SEQ ID NO: 79. In one embodiment, the targeting domain peptide comprises a sequence as shown in SEQ ID NO: 79.

[0319] In one embodiment, the targeting domain is an EGFR targeting domain. In one embodiment, the targeting domain peptide comprises a sequence having at least 90% identity to SEQ ID NO:80. In one embodiment, the targeting domain peptide comprises a sequence having at least 95% identity to SEQ ID NO:80. In one embodiment, the targeting domain peptide comprises a sequence having at least 96% identity to SEQ ID NO:80. In one embodiment, the targeting domain peptide comprises a sequence having at least 97% identity to SEQ ID NO:80. In one embodiment, the targeting domain peptide comprises a sequence having at least 98% identity to SEQ ID NO:80. In one embodiment, the targeting domain peptide comprises a sequence having at least 99% identity to SEQ ID NO:80. In one embodiment, the targeting domain peptide comprises a sequence as shown in SEQ ID NO:80.

[0320] In one embodiment, the targeting domain is a BCMA targeting domain. In one embodiment, the targeting domain peptide comprises a sequence having at least 90% identity to SEQ ID NO: 81. In one embodiment, the targeting domain peptide comprises a sequence having at least 95% identity to SEQ ID NO: 81. In one embodiment, the targeting domain peptide comprises a sequence having at least 96% identity to SEQ ID NO: 81. In one embodiment, the targeting domain peptide comprises a sequence having at least 97% identity to SEQ ID NO: 81. In one embodiment, the targeting domain peptide comprises a sequence having at least 98% identity to SEQ ID NO: 81. In one embodiment, the targeting domain peptide comprises a sequence having at least 99% identity to SEQ ID NO: 81. In one embodiment, the targeting domain peptide comprises a sequence as shown in SEQ ID NO: 81.

[0321] In one embodiment, the targeting domain is a BCMA targeting domain. In one embodiment, the targeting domain peptide comprises a sequence having at least 90% identity to SEQ ID NO: 82. In one embodiment, the targeting domain peptide comprises a sequence having at least 95% identity to SEQ ID NO: 82. In one embodiment, the targeting domain peptide comprises a sequence having at least 96% identity to SEQ ID NO: 82. In one embodiment, the targeting domain peptide comprises a sequence having at least 97% identity to SEQ ID NO: 82. In one embodiment, the targeting domain peptide comprises a sequence having at least 98% identity to SEQ ID NO: 82. In one embodiment, the targeting domain peptide comprises a sequence having at least 99% identity to SEQ ID NO: 82. In one embodiment, the targeting domain peptide comprises a sequence as shown in SEQ ID NO: 82.

[0322] In one embodiment, the targeting domain is a CD19 targeting domain. In one embodiment, the targeting domain peptide comprises a sequence having at least 90% identity to SEQ ID NO:83. In one embodiment, the targeting domain peptide comprises a sequence having at least 95% identity to SEQ ID NO:83. In one embodiment, the targeting domain peptide comprises a sequence having at least 96% identity to SEQ ID NO:83. In one embodiment, the targeting domain peptide comprises a sequence having at least 97% identity to SEQ ID NO:83. In one embodiment, the targeting domain peptide comprises a sequence having at least 98% identity to SEQ ID NO:83. In one embodiment, the targeting domain peptide comprises a sequence having at least 99% identity to SEQ ID NO:83. In one embodiment, the targeting domain peptide comprises a sequence as shown in SEQ ID NO:83.

[0323] In one embodiment, the targeting domain comprises a CD33 targeting domain. In one embodiment, the CD33 targeting domain is a VHH. In one embodiment, the CD33 VHH comprises a CDR1 comprising SEQ ID NO: 89, a CDR2 comprising SEQ ID NO: 90, and a CDR3 comprising SEQ ID NO: 91.

[0324] In one embodiment, the targeting domain comprises a CLL1 targeting domain. In one embodiment, the CLL1 targeting domain is a VHH. In one embodiment, the CD33 VHH comprises a CDR1 comprising SEQ ID NO: 92, a CDR2 comprising SEQ ID NO: 93, and a CDR3 comprising SEQ ID NO: 94.

[0325] In one embodiment, the targeting component comprises a first targeting domain and a second targeting domain. In one embodiment, the first targeting domain is a CD33 targeting domain and the second targeting domain is a CLL1 targeting domain. In another embodiment, the first targeting domain is a CLL1 targeting domain and the second targeting domain is a CD33 targeting domain. In one embodiment, the first targeting domain is a CD33 VHH, wherein the CD33 VHH comprises a CDR1 comprising SEQ ID NO: 89, a CDR2 comprising SEQ ID NO: 90, and a CDR3 comprising SEQ ID NO: 91; and the second targeting domain is a CLL1 VHH, wherein the CLL1 VHH comprises a CDR1 comprising SEQ ID NO: 92, a CDR2 comprising SEQ ID NO: 93, and a CDR3 comprising SEQ ID NO: 94.

[0326] In various embodiments, the first targeting domain and / or the second targeting domain comprises a receptor extracellular domain. In some embodiments, the extracellular domain is derived from a receptor selected from the group consisting of PD1 or high affinity PD1.

[0327] In various embodiments, the extracellular domain is a PD1 extracellular domain. In some embodiments, the PD1 extracellular domain comprises an amino acid sequence having at least 90% identity to SEQ ID NO: 84. In some embodiments, the PD1 extracellular domain comprises an amino acid sequence having at least 95% identity to SEQ ID NO: 84. In some embodiments, the PD1 extracellular domain comprises an amino acid sequence having at least 96% identity to SEQ ID NO: 84. In some embodiments, the PD1 extracellular domain comprises an amino acid sequence having at least 97% identity to SEQ ID NO: 84. In some embodiments, the PD1 extracellular domain comprises an amino acid sequence having at least 98% identity to SEQ ID NO: 84. In some embodiments, the PD1 extracellular domain comprises an amino acid sequence having at least 99% identity to SEQ ID NO: 84. In a specific embodiment, the PD1 extracellular domain comprises an amino acid sequence as shown in SEQ ID NO: 84.

[0328] In various embodiments, the extracellular domain is a high-affinity PD1 extracellular domain. See, e.g., US20220378873A1. In some embodiments, the high-affinity PD1 extracellular domain comprises an amino acid sequence having at least 90% identity to SEQ ID NO: 85. In some embodiments, the high-affinity PD1 extracellular domain comprises an amino acid sequence having at least 95% identity to SEQ ID NO: 85. In some embodiments, the high-affinity PD1 extracellular domain comprises an amino acid sequence having at least 96% identity to SEQ ID NO: 85. In some embodiments, the high-affinity PD1 extracellular domain comprises an amino acid sequence having at least 97% identity to SEQ ID NO: 85. In some embodiments, the high-affinity PD1 extracellular domain comprises an amino acid sequence having at least 98% identity to SEQ ID NO: 85. In some embodiments, the high-affinity PD1 extracellular domain comprises an amino acid sequence having at least 99% identity to SEQ ID NO: 85. In a specific embodiment, the high-affinity PD1 extracellular domain comprises an amino acid sequence as shown in SEQ ID NO: 85.

[0329] In various embodiments, the first targeting domain and / or the second targeting domain comprises human A proliferation inducing ligand (APRIL), trimerized human APRIL, or NKG2D membrane protein.

[0330] In various embodiments, the first targeting domain and / or the second targeting domain comprises an APRIL. In some embodiments, the APRIL comprises an amino acid sequence having at least 90% identity to SEQ ID NO: 86. In some embodiments, the APRIL comprises an amino acid sequence having at least 95% identity to SEQ ID NO: 86. In some embodiments, the APRIL comprises an amino acid sequence having at least 96% identity to SEQ ID NO: 86. In some embodiments, the APRIL comprises an amino acid sequence having at least 97% identity to SEQ ID NO: 86. In some embodiments, the APRIL comprises an amino acid sequence having at least 98% identity to SEQ ID NO: 86. In some embodiments, the APRIL comprises an amino acid sequence having at least 99% identity to SEQ ID NO: 86. In some embodiments, the APRIL comprises an amino acid sequence as set forth in SEQ ID NO: 86.

[0331] In various embodiments, the first targeting domain and / or the second targeting domain comprises a trimerized human APRIL. In some embodiments, the APRIL comprises an amino acid sequence having at least 90% identity to SEQ ID NO: 87. In some embodiments, the APRIL comprises an amino acid sequence having at least 95% identity to SEQ ID NO: 87. In some embodiments, the APRIL comprises an amino acid sequence having at least 96% identity to SEQ ID NO: 87. In some embodiments, the APRIL comprises an amino acid sequence having at least 97% identity to SEQ ID NO: 87. In some embodiments, the APRIL comprises an amino acid sequence having at least 98% identity to SEQ ID NO: 87. In some embodiments, the APRIL comprises an amino acid sequence having at least 99% identity to SEQ ID NO: 87. In some embodiments, the APRIL comprises an amino acid sequence as set forth in SEQ ID NO: 87.

[0332] In various embodiments, the first targeting domain and / or the second targeting domain comprises a NKG2D membrane protein. In some embodiments, the NKG2D membrane protein comprises an amino acid sequence having at least 90% identity to SEQ ID NO: 88. In some embodiments, the NKG2D membrane protein comprises an amino acid sequence having at least 95% identity to SEQ ID NO: 88. In some embodiments, the NKG2D membrane protein comprises an amino acid sequence having at least 96% identity to SEQ ID NO: 88. In some embodiments, the NKG2D membrane protein comprises an amino acid sequence having at least 97% identity to SEQ ID NO: 88. In some embodiments, the NKG2D membrane protein comprises an amino acid sequence having at least 98% identity to SEQ ID NO: 88. In some embodiments, the NKG2D membrane protein comprises an amino acid sequence having at least 99% identity to SEQ ID NO: 88. In some embodiments, NKG2D comprises an amino acid sequence as shown in SEQ ID NO: 88.

[0333] In various embodiments, the first targeting domain and the second targeting domain associate or bind to the same or different target antigens.

[0334] In various embodiments, the first targeting domain and the second targeting domain are separated by a third polypeptide linker having a length of 2 to 40 amino acids. In some embodiments, the third polypeptide linker is selected from the group consisting of GG, GS, SG, SS, GSS, SSG, GSG, SGS, SGG, G4S, 2xG4S, 3xG4s, 4xG4S, and any combination thereof.

[0335] In certain embodiments, the targeting component comprises one or more multimerization domains.

[0336] Illustrative examples of multimerization domains suitable for use in the specific targeting components contemplated herein include, but are not limited to, FK506 binding protein (FKBP) polypeptides or variants thereof, FKBP-rapamycin binding (FRB) polypeptides or variants thereof, calcineurin polypeptides or variants thereof, cyclophilin polypeptides or variants thereof, bacterial dihydrofolate reductase (DHFR) polypeptides or variants thereof, PYR1-like 1 (PYL1) polypeptides or variants thereof, and abscisic acid insensitive 1 (ABI1) polypeptides or variants thereof.

[0337] In various embodiments, the targeting component comprises a FRB polypeptide. In various embodiments, the targeting component comprises a FRB polypeptide or a variant thereof comprising a T2098L (T82L) mutation. In specific embodiments, the FRB polypeptide comprises the amino acid sequence as shown in SEQ ID NO: 1. In specific embodiments, the FRB polypeptide comprises the amino acid sequence as shown in SEQ ID NO: 2.

[0338] In various embodiments, the targeting component comprises a FKBP12 polypeptide or a variant thereof. In various embodiments, the targeting component comprises a FKBP12 polypeptide comprising a F36V mutation. In specific embodiments, the FKBP12 polypeptide comprises the amino acid sequence as shown in SEQ ID NO:3. In specific embodiments, the FKBP12 polypeptide comprises the amino acid sequence as shown in SEQ ID NO:4.

[0339] Other illustrative examples of multimerization domains suitable for use in the specific targeting components contemplated herein include antibody-derived heterodimerization domains. Such antibody heterodimerization domains include, but are not limited to, lock and key, DEKK, SEEDbody, DuoBody, dual variable domain immunoglobulin (DVD-Ig) and tandem Fab immunoglobulin (FIT-Ig) multimerization domains (see, e.g., Ma et al., Frontiers in Immunology (Front Immunol) . May 5, 2021; 12: 626616).

[0340] In various embodiments, the targeting component comprises a lock and key multimerization domain / polypeptide or a variant thereof. In some embodiments, the lock and key multimerization domain / polypeptide or a variant thereof comprises an amino acid sequence having at least 90% identity to SEQ ID NO:5. In one embodiment, the lock and key multimerization domain / polypeptide or a variant thereof comprises an amino acid sequence having at least 95% identity to SEQ ID NO:5. In one embodiment, the lock and key multimerization domain / polypeptide or a variant thereof comprises an amino acid sequence having at least 96% identity to SEQ ID NO:5. In one embodiment, the lock and key multimerization domain / polypeptide or a variant thereof comprises an amino acid sequence having at least 97% identity to SEQ ID NO:5. In one embodiment, the lock and key multimerization domain / polypeptide or a variant thereof comprises an amino acid sequence having at least 98% identity to SEQ ID NO:5. In one embodiment, the lock and key multimerization domain / polypeptide or a variant thereof comprises an amino acid sequence having at least 99% identity to SEQ ID NO:5. In one embodiment, the lock and key multimerization domain / polypeptide or a variant thereof comprises an amino acid sequence as shown in SEQ ID NO:5. In some embodiments, the lock and key multimerization domain / polypeptide or variant thereof comprises an amino acid sequence that is at least 90% identical to SEQ ID NO:6. In one embodiment, the lock and key multimerization domain / polypeptide or variant thereof comprises an amino acid sequence that is at least 95% identical to SEQ ID NO:6. In one embodiment, the lock and key multimerization domain / polypeptide or variant thereof comprises an amino acid sequence that is at least 96% identical to SEQ ID NO:6. In one embodiment, the lock and key multimerization domain / polypeptide or variant thereof comprises an amino acid sequence that is at least 97% identical to SEQ ID NO:6. In one embodiment, the lock and key multimerization domain / polypeptide or variant thereof comprises an amino acid sequence that is at least 98% identical to SEQ ID NO:6. In one embodiment, the lock and key multimerization domain / polypeptide or variant thereof comprises an amino acid sequence that is at least 99% identical to SEQ ID NO:6. In some embodiments, the lock and key multimerization domain / polypeptide or variant thereof comprises an amino acid sequence as shown in SEQ ID NO:6.

[0341] In various embodiments, the targeting component comprises a DEKK multimerization domain / polypeptide or a variant thereof. In some embodiments, the DEKK multimerization domain / polypeptide or a variant thereof comprises an amino acid sequence having at least 90% identity to SEQ ID NO: 7. In some embodiments, the DEKK multimerization domain / polypeptide or a variant thereof comprises an amino acid sequence having at least 95% identity to SEQ ID NO: 7. In some embodiments, the DEKK multimerization domain / polypeptide or a variant thereof comprises an amino acid sequence having at least 96% identity to SEQ ID NO: 7. In some embodiments, the DEKK multimerization domain / polypeptide or a variant thereof comprises an amino acid sequence having at least 97% identity to SEQ ID NO: 7. In some embodiments, the DEKK multimerization domain / polypeptide or a variant thereof comprises an amino acid sequence having at least 98% identity to SEQ ID NO: 7. In some embodiments, the DEKK multimerization domain / polypeptide or a variant thereof comprises an amino acid sequence having at least 99% identity to SEQ ID NO: 7. In some embodiments, the DEKK multimerization domain / polypeptide or a variant thereof comprises an amino acid sequence as shown in SEQ ID NO: 7. In some embodiments, the DEKK multimerization domain / polypeptide or variant thereof comprises an amino acid sequence having at least 90% identity to SEQ ID NO: 8. In some embodiments, the DEKK multimerization domain / polypeptide or variant thereof comprises an amino acid sequence having at least 95% identity to SEQ ID NO: 8. In some embodiments, the DEKK multimerization domain / polypeptide or variant thereof comprises an amino acid sequence having at least 96% identity to SEQ ID NO: 8. In some embodiments, the DEKK multimerization domain / polypeptide or variant thereof comprises an amino acid sequence having at least 97% identity to SEQ ID NO: 8. In some embodiments, the DEKK multimerization domain / polypeptide or variant thereof comprises an amino acid sequence having at least 98% identity to SEQ ID NO: 8. In some embodiments, the DEKK multimerization domain / polypeptide or variant thereof comprises an amino acid sequence having at least 99% identity to SEQ ID NO: 8. In some embodiments, the DEKK multimerization domain / polypeptide or variant thereof comprises an amino acid sequence as shown in SEQ ID NO: 8.

[0342] In various embodiments, the targeting component comprises a SEEDbody multimerization domain / polypeptide or a variant thereof. In some embodiments, the SEEDbody multimerization domain / polypeptide or a variant thereof comprises an amino acid sequence having at least 90% identity to SEQ ID NO: 9. In some embodiments, the SEEDbody multimerization domain / polypeptide or a variant thereof comprises an amino acid sequence having at least 95% identity to SEQ ID NO: 9. In some embodiments, the SEEDbody multimerization domain / polypeptide or a variant thereof comprises an amino acid sequence having at least 96% identity to SEQ ID NO: 9. In some embodiments, the SEEDbody multimerization domain / polypeptide or a variant thereof comprises an amino acid sequence having at least 97% identity to SEQ ID NO: 9. In some embodiments, the SEEDbody multimerization domain / polypeptide or a variant thereof comprises an amino acid sequence having at least 98% identity to SEQ ID NO: 9. In some embodiments, the SEEDbody multimerization domain / polypeptide or a variant thereof comprises an amino acid sequence having at least 99% identity to SEQ ID NO: 9. In some embodiments, the SEEDbody multimerization domain / polypeptide or a variant thereof comprises an amino acid sequence as shown in SEQ ID NO: 9. In some embodiments, the SEEDbody multimerization domain / polypeptide or variant thereof comprises an amino acid sequence that is at least 90% identical to SEQ ID NO: 10. In some embodiments, the SEEDbody multimerization domain / polypeptide or variant thereof comprises an amino acid sequence that is at least 95% identical to SEQ ID NO: 10. In some embodiments, the SEEDbody multimerization domain / polypeptide or variant thereof comprises an amino acid sequence that is at least 96% identical to SEQ ID NO: 10. In some embodiments, the SEEDbody multimerization domain / polypeptide or variant thereof comprises an amino acid sequence that is at least 97% identical to SEQ ID NO: 10. In some embodiments, the SEEDbody multimerization domain / polypeptide or variant thereof comprises an amino acid sequence that is at least 98% identical to SEQ ID NO: 10. In some embodiments, the SEEDbody multimerization domain / polypeptide or variant thereof comprises an amino acid sequence that is at least 99% identical to SEQ ID NO: 10. In some embodiments, the SEEDbody multimerization domain / polypeptide or variant thereof comprises an amino acid sequence as shown in SEQ ID NO: 10.

[0343] Other illustrative examples of multimerization domains suitable for use in the specific targeting components contemplated herein include, but are not limited to, leucine zipper multimerization domains, mFos:mJun helical multimerization domains, p53 and MDM2 multimerization domains, VPS36 and VPS28 multimerization domains. Other suitable multimerization or dimerization domains are known to those skilled in the art.

[0344] In various embodiments, the extracellular domain or the first targeting domain and the second multimerization domain are separated by a second polypeptide linker of 2 to 40 amino acids in length. In various embodiments, preferably a short oligopeptide linker or polypeptide linker of 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19 or 20 amino acids in length connects one or more domains in the targeting component. A linker based on glycine-serine provides a particularly suitable linker. In certain embodiments, the second polypeptide linker is selected from the group consisting of: GG, GS, SG, SS, GSS, SSG, GSG, SGS, SGG, G4S, 2xG4S, 3xG4S, 4xG4S and any combination thereof. In certain embodiments, the second polypeptide linker is a G4S (GGGGS; SEQ ID NO: 11) linker. In some embodiments, the second polypeptide linker is a 2xG4S (GGGGSGGGGS; SEQ ID NO: 12) linker. In some embodiments, the second polypeptide linker is a 3xG4S (GGGGSGGGGSGGGGS; SEQ ID NO: 13) linker. In some embodiments, the second polypeptide linker is a 4xG4S

[0345] (GGGGSGGGGSGGGGSGGGGS; SEQ ID NO: 14) linker. In some embodiments, the second polypeptide linker is 5xG4S (GGGGSGGGGSGGGGSGGGGSGGGGS; SEQ ID NO: 15) linker. In some embodiments, the second polypeptide linker comprises an amino acid sequence as shown in any one or more of SEQ ID NOs: 16-31.

[0346] In various embodiments, the second multimerization domain and the transmembrane domain are separated by a hinge domain.

[0347] In some embodiments, the hinge domain is selected from the group consisting of: a CD4 hinge, a CD8 hinge, a CD28 hinge, an IgG4 hinge, and any fragment or variant or combination thereof. In some embodiments, the hinge domain is a CD4 hinge. In some embodiments, the CD4 hinge comprises an amino acid sequence having at least 90% identity to SEQ ID NO: 41. In some embodiments, the CD4 hinge comprises an amino acid sequence having at least 95% identity to SEQ ID NO: 41. In some embodiments, the CD4 hinge comprises an amino acid sequence having at least 96% identity to SEQ ID NO: 41. In some embodiments, the CD4 hinge comprises an amino acid sequence having at least 97% identity to SEQ ID NO: 41. In some embodiments, the CD4 hinge comprises an amino acid sequence having at least 98% identity to SEQ ID NO: 41. In some embodiments, the CD4 hinge comprises an amino acid sequence having at least 99% identity to SEQ ID NO: 41. In some embodiments, the CD4 hinge comprises an amino acid sequence as shown in SEQ ID NO: 41.

[0348] In some embodiments, the hinge domain is a CD28 hinge. In some embodiments, the CD28 hinge comprises an amino acid sequence having at least 90% identity to SEQ ID NO: 42. In some embodiments, the CD28 hinge comprises an amino acid sequence having at least 95% identity to SEQ ID NO: 42. In some embodiments, the CD28 hinge comprises an amino acid sequence having at least 96% identity to SEQ ID NO: 42. In some embodiments, the CD28 hinge comprises an amino acid sequence having at least 97% identity to SEQ ID NO: 1042. In some embodiments, the CD28 hinge comprises an amino acid sequence having at least 98% identity to SEQ ID NO: 42. In some embodiments, the CD28 hinge comprises an amino acid sequence having at least 99% identity to SEQ ID NO: 42. In some embodiments, the CD28 hinge comprises an amino acid sequence as shown in SEQ ID NO: 42.

[0349] In some embodiments, the hinge domain is an IgG4 hinge. In one embodiment, the IgG4 hinge comprises an amino acid sequence that is at least 90% identical to SEQ ID NO: 43. In one embodiment, the IgG4 hinge comprises an amino acid sequence that is at least 95% identical to SEQ ID NO: 43. In one embodiment, the IgG4 hinge comprises an amino acid sequence that is at least 96% identical to SEQ ID NO: 43. In one embodiment, the IgG4 hinge comprises an amino acid sequence that is at least 97% identical to SEQ ID NO: 43. In one embodiment, the IgG4 hinge comprises an amino acid sequence that is at least 98% identical to SEQ ID NO: 43. In one embodiment, the IgG4 hinge comprises an amino acid sequence that is at least 99% identical to SEQ ID NO: 43. In one embodiment, the IgG4 hinge comprises an amino acid sequence as shown in SEQ ID NO: 43.

[0350] In one embodiment, the hinge domain is a CD8 hinge. In one embodiment, the CD8 hinge comprises an amino acid sequence that is at least 90% identical to SEQ ID NO: 44. In one embodiment, the CD8 hinge comprises an amino acid sequence that is at least 95% identical to SEQ ID NO: 44. In one embodiment, the CD8 hinge comprises an amino acid sequence that is at least 96% identical to SEQ ID NO: 44. In one embodiment, the CD8 hinge comprises an amino acid sequence that is at least 97% identical to SEQ ID NO: 44. In one embodiment, the CD8 hinge comprises an amino acid sequence that is at least 98% identical to SEQ ID NO: 44. In one embodiment, the CD8 hinge comprises an amino acid sequence that is at least 99% identical to SEQ ID NO: 44. In one embodiment, the CD8 hinge comprises an amino acid sequence as set forth in SEQ ID NO: 44.

[0351] In some embodiments, the targeting component does not comprise a hinge domain. In some embodiments, the targeting component comprises a linker comprising a GGR amino acid sequence. In some embodiments, the targeting component comprises a linker comprising an amino acid sequence as shown in SEQ ID NO: 13 or 14.

[0352] In certain embodiments, the targeting component comprises a transmembrane domain.

[0353] Illustrative examples of transmembrane domains suitable for use in the specific targeting components contemplated herein include, but are not limited to, the transmembrane regions of the α chain, β chain, γ chain, or δ chain of the T cell receptor, CD3ε, CD3ζ, CD4, CD5, CD8α, CD9, CD 16, CD22, CD27, CD28, CD33, CD37, CD45, CD64, CD71, CD80, CD86, CD 134, CD137, CD152, CD 154, amniotic membrane protein (AMN), and programmed cell death 1 (PDCD1). In various embodiments, the targeting component comprises a CD4 transmembrane domain, a CD28 transmembrane domain, or a CD8α transmembrane domain.

[0354] In a specific embodiment, the targeting component comprises a CD4 transmembrane domain. In some embodiments, the CD4 transmembrane domain comprises an amino acid sequence that is at least 90% identical to SEQ ID NO: 45. In some embodiments, the CD4 transmembrane domain comprises an amino acid sequence that is at least 95% identical to SEQ ID NO: 45. In some embodiments, the CD4 transmembrane domain comprises an amino acid sequence that is at least 96% identical to SEQ ID NO: 45. In some embodiments, the CD4 transmembrane domain comprises an amino acid sequence that is at least 97% identical to SEQ ID NO: 45. In some embodiments, the CD4 transmembrane domain comprises an amino acid sequence that is at least 98% identical to SEQ ID NO: 45. In some embodiments, the CD4 transmembrane domain comprises an amino acid sequence that is at least 99% identical to SEQ ID NO: 45. In some embodiments, the CD4 transmembrane domain comprises an amino acid sequence as shown in SEQ ID NO: 45.

[0355] In a specific embodiment, the targeting component comprises a CD28 transmembrane domain. In some embodiments, the CD28 transmembrane domain comprises an amino acid sequence having at least 90% identity to SEQ ID NO: 46. In some embodiments, the CD28 transmembrane domain comprises an amino acid sequence having at least 95% identity to SEQ ID NO: 46. In some embodiments, the CD28 transmembrane domain comprises an amino acid sequence having at least 96% identity to SEQ ID NO: 46. In some embodiments, the CD28 transmembrane domain comprises an amino acid sequence having at least 97% identity to SEQ ID NO: 46. In some embodiments, the CD28 transmembrane domain comprises an amino acid sequence having at least 98% identity to SEQ ID NO: 46. In some embodiments, the CD28 transmembrane domain comprises an amino acid sequence having at least 99% identity to SEQ ID NO: 46. In some embodiments, the CD28 transmembrane domain comprises an amino acid sequence as shown in SEQ ID NO: 46.

[0356] In a specific embodiment, the targeting component comprises a CD8 transmembrane domain. In some embodiments, the CD28 transmembrane domain comprises an amino acid sequence having at least 90% identity to SEQ ID NO: 47. In some embodiments, the CD8 transmembrane domain comprises an amino acid sequence having at least 95% identity to SEQ ID NO: 47. In some embodiments, the CD8 transmembrane domain comprises an amino acid sequence having at least 96% identity to SEQ ID NO: 47. In some embodiments, the CD8 transmembrane domain comprises an amino acid sequence having at least 97% identity to SEQ ID NO: 47. In some embodiments, the CD8 transmembrane domain comprises an amino acid sequence having at least 98% identity to SEQ ID NO: 47. In some embodiments, the CD8 transmembrane domain comprises an amino acid sequence having at least 99% identity to SEQ ID NO: 47. In some embodiments, the CD8 transmembrane domain comprises an amino acid sequence as shown in SEQ ID NO: 47.

[0357] In certain embodiments, the targeting component comprises a truncated CD4 intracellular (CD4ic) domain. In one embodiment, the truncated CD4ic domain comprises an amino acid sequence having at least 90% identity to SEQ ID NO: 48. In one embodiment, the truncated CD4ic domain comprises an amino acid sequence having at least 95% identity to SEQ ID NO: 48. In one embodiment, the truncated CD4ic domain comprises an amino acid sequence having at least 96% identity to SEQ ID NO: 48. In one embodiment, the truncated CD4ic domain comprises an amino acid sequence having at least 97% identity to SEQ ID NO: 48. In one embodiment, the truncated CD4ic domain comprises an amino acid sequence having at least 98% identity to SEQ ID NO: 48. In one embodiment, the truncated CD4ic domain comprises an amino acid sequence having at least 99% identity to SEQ ID NO: 48. In various embodiments, the truncated CD4ic domain comprises an amino acid sequence as shown in SEQ ID NO: 48. In one embodiment, the truncated CD4ic domain comprises an amino acid sequence having at least 90% identity to SEQ ID NO: 49. In one embodiment, the truncated CD4ic domain comprises an amino acid sequence that is at least 95% identical to SEQ ID NO: 49. In one embodiment, the truncated CD4ic domain comprises an amino acid sequence that is at least 96% identical to SEQ ID NO: 49. In one embodiment, the truncated CD4ic domain comprises an amino acid sequence that is at least 97% identical to SEQ ID NO: 49. In one embodiment, the truncated CD4ic domain comprises an amino acid sequence that is at least 98% identical to SEQ ID NO: 49. In one embodiment, the truncated CD4ic domain comprises an amino acid sequence that is at least 99% identical to SEQ ID NO: 49. In various embodiments, the truncated CD4ic domain comprises an amino acid sequence as shown in SEQ ID NO: 49.

[0358] The targeting components contemplated in specific embodiments herein comprise one or more functional intracellular signaling domains or co-stimulatory domains (i.e., having intracellular or co-stimulatory signaling capability). In various embodiments, the intracellular signaling domain or co-stimulatory domain is derived from a protein selected from the group consisting of an antigen receptor, a co-receptor, a co-stimulatory receptor, a growth receptor or a cytokine receptor, an adaptor signaling protein, an intracellular signaling protein, or any fragment or variant thereof.

[0359] In various embodiments, the targeting component comprises one or more intracellular signaling domains or co-stimulatory domains, and optionally a CD3zeta primary signaling domain. A primary signaling domain; and one or more intracellular signaling domains or co-stimulatory domains; may be connected in series with the carboxyl terminus of the transmembrane domain in any order. In specific embodiments, the targeting component does not comprise a primary signaling domain (e.g., CD3zeta).

[0360] In various embodiments, the one or more intracellular signaling domains or co-stimulatory domains are selected from CD3ε, CD3γ, CD3δ, CD4 or CD8.

[0361] In various embodiments, the auxiliary receptor domain is a CD4 auxiliary receptor domain. In some embodiments, the CD4 auxiliary receptor domain comprises an amino acid sequence that is at least 90% identical to SEQ ID NO: 100. In some embodiments, the CD4 auxiliary receptor domain comprises an amino acid sequence that is at least 95% identical to SEQ ID NO: 100. In some embodiments, the CD4 auxiliary receptor domain comprises an amino acid sequence that is at least 96% identical to SEQ ID NO: 100. In some embodiments, the CD4 auxiliary receptor domain comprises an amino acid sequence that is at least 97% identical to SEQ ID NO: 100. In some embodiments, the CD4 auxiliary receptor domain comprises an amino acid sequence that is at least 98% identical to SEQ ID NO: 100. In some embodiments, the CD4 auxiliary receptor domain comprises an amino acid sequence that is at least 99% identical to SEQ ID NO: 100. In some embodiments, the CD4 auxiliary receptor domain comprises an amino acid sequence as shown in SEQ ID NO: 100.

[0362] In various embodiments, the auxiliary receptor domain is a CD8 auxiliary receptor domain. In some embodiments, the CD8 auxiliary receptor domain comprises an amino acid sequence that is at least 90% identical to SEQ ID NO: 101. In some embodiments, the CD8 auxiliary receptor domain comprises an amino acid sequence that is at least 95% identical to SEQ ID NO: 101. In some embodiments, the CD8 auxiliary receptor domain comprises an amino acid sequence that is at least 96% identical to SEQ ID NO: 101. In some embodiments, the CD8 auxiliary receptor domain comprises an amino acid sequence that is at least 97% identical to SEQ ID NO: 101. In some embodiments, the CD8 auxiliary receptor domain comprises an amino acid sequence that is at least 98% identical to SEQ ID NO: 101. In some embodiments, the CD8 auxiliary receptor domain comprises an amino acid sequence that is at least 99% identical to SEQ ID NO: 101. In some embodiments, the CD8 auxiliary receptor domain comprises an amino acid sequence as shown in SEQ ID NO: 101.

[0363] In various embodiments, one or more intracellular signaling domains or co-stimulatory domains comprise an immunoreceptor tyrosine-based activation motif (ITAM). Illustrative examples of primary signaling domains containing ITAMs suitable for use in the specific signaling components contemplated herein include, but are not limited to, those derived from FcRγ, FcRβ, CD3γ, CD3δ, CD3ε, CD3ζ, CD22, CD79a, CD79b, and CD66d.

[0364] Illustrative examples of intracellular signaling domains or co-stimulatory domains suitable for use in the specific targeting components contemplated herein include, but are not limited to, those isolated from the following co-stimulatory molecules: Toll-like receptor 1 (TLR1), TLR2, TLR3, TLR4, TLR5, TLR6, TLR7, TLR8, TLR9, TLR10, caspase recruitment domain family member 11 (CARD11), CD2, CD3ε, CD3γ, CD3δ, CD7, CD27, CD28, CD30, CD40, CD54 (ICAM), CD83, CD94, CD134 (OX40), CD137 (4- 1BB), CD278 (ICOS), common gamma chain cytokine DNAX-activating protein 10 (DAP10), FYN, adaptor for activating T cells family member 1 (LAT), LCK, interleukin 2 receptor (IL-2R), IL-4R, IL-7R, IL-9R, IL-12R, IL-13R, IL-15R, IL-21R, SH2 domain-containing leukocyte protein of 76 kD (SLP76), T cell receptor-associated transmembrane adaptor 1 (TRAT1), TNFR2, TNFRS14, TNFRS18, TNRFS25, and T cell receptor zeta chain-associated protein kinase 70 (ZAP70).

[0365] In various embodiments, the costimulatory domain is selected from the group consisting of: 4-1BB, CD28, TNFR2, OX40, ICOS, and DAP10 costimulatory domains. In some embodiments, the costimulatory domain is a 4-1BB costimulatory domain.

[0366] In some embodiments, the 4-1BB costimulatory domain comprises an amino acid sequence having at least 90% identity to SEQ ID NO: 98. In some embodiments, the 4-1BB costimulatory domain comprises an amino acid sequence having at least 95% identity to SEQ ID NO: 98. In some embodiments, the 4-1BB costimulatory domain comprises an amino acid sequence having at least 96% identity to SEQ ID NO: 98. In some embodiments, the 4-1BB costimulatory domain comprises an amino acid sequence having at least 97% identity to SEQ ID NO: 98. In some embodiments, the 4-1BB costimulatory domain comprises an amino acid sequence having at least 98% identity to SEQ ID NO: 98. In some embodiments, the 4-1BB costimulatory domain comprises an amino acid sequence having at least 99% identity to SEQ ID NO: 98. In some embodiments, the 4-1BB costimulatory domain comprises an amino acid sequence as shown in SEQ ID NO: 98. In some embodiments, the costimulatory domain is a CD28 costimulatory domain. In some embodiments, the CD28 costimulatory domain comprises an amino acid sequence as shown in SEQ ID NO: 99. In some embodiments, the CD28 costimulatory domain comprises an amino acid sequence that is at least 90% identical to SEQ ID NO: 99. In some embodiments, the CD28 costimulatory domain comprises an amino acid sequence that is at least 95% identical to SEQ ID NO: 99. In some embodiments, the CD28 costimulatory domain comprises an amino acid sequence that is at least 96% identical to SEQ ID NO: 99. In some embodiments, the CD28 costimulatory domain comprises an amino acid sequence that is at least 97% identical to SEQ ID NO: 99. In some embodiments, the CD28 costimulatory domain comprises an amino acid sequence that is at least 98% identical to SEQ ID NO: 99. In some embodiments, the CD28 costimulatory domain comprises an amino acid sequence that is at least 99% identical to SEQ ID NO: 99.

[0367] In various embodiments, the intracellular signaling domain or co-stimulatory domain is a LAT domain. In one embodiment, the LAT domain comprises an amino acid sequence having at least 90% identity to SEQ ID NO: 102. In one embodiment, the LAT domain comprises an amino acid sequence having at least 95% identity to SEQ ID NO: 102. In one embodiment, the LAT domain comprises an amino acid sequence having at least 96% identity to SEQ ID NO: 102. In one embodiment, the LAT domain comprises an amino acid sequence having at least 97% identity to SEQ ID NO: 102. In one embodiment, the LAT domain comprises an amino acid sequence having at least 98% identity to SEQ ID NO: 102. In one embodiment, the LAT domain comprises an amino acid sequence having at least 99% identity to SEQ ID NO: 102. In one embodiment, the LAT domain comprises an amino acid sequence as shown in SEQ ID NO: 102.

[0368] In various embodiments, the intracellular signaling domain or co-stimulatory domain is one or more cytokine receptor intracellular signaling domains. In some embodiments, the one or more cytokine receptor intracellular signaling domains are selected from the group consisting of: an IL7Rα intracellular signaling domain, an IL2Rβ intracellular signaling domain, a common γ chain intracellular signaling domain, and both IL2Rβ and common γ chain intracellular signaling domains.

[0369] In various embodiments, the IL7Rα intracellular signaling domain comprises an amino acid sequence that is at least 90% identical to SEQ ID NO: 103. In some embodiments, the IL7Rα intracellular signaling domain comprises an amino acid sequence that is at least 95% identical to SEQ ID NO: 103. In some embodiments, the IL7Rα intracellular signaling domain comprises an amino acid sequence that is at least 96% identical to SEQ ID NO: 103. In some embodiments, the IL7Rα intracellular signaling domain comprises an amino acid sequence that is at least 97% identical to SEQ ID NO: 103. In some embodiments, the IL7Rα intracellular signaling domain comprises an amino acid sequence that is at least 98% identical to SEQ ID NO: 103. In some embodiments, the IL7Rα intracellular signaling domain comprises an amino acid sequence that is at least 99% identical to SEQ ID NO: 103. In some embodiments, the IL7Rα intracellular signaling domain comprises an amino acid sequence as shown in SEQ ID NO: 103.

[0370] In various embodiments, the IL2Rβ intracellular signaling domain comprises an amino acid sequence that is at least 90% identical to SEQ ID NO: 104. In some embodiments, the IL2Rβ intracellular signaling domain comprises an amino acid sequence that is at least 95% identical to SEQ ID NO: 104. In some embodiments, the IL2Rβ intracellular signaling domain comprises an amino acid sequence that is at least 96% identical to SEQ ID NO: 104. In some embodiments, the IL2Rβ intracellular signaling domain comprises an amino acid sequence that is at least 97% identical to SEQ ID NO: 104. In some embodiments, the IL2Rβ intracellular signaling domain comprises an amino acid sequence that is at least 98% identical to SEQ ID NO: 104. In some embodiments, the IL2Rβ intracellular signaling domain comprises an amino acid sequence that is at least 99% identical to SEQ ID NO: 104. In some embodiments, the IL2Rβ intracellular signaling domain comprises an amino acid sequence as set forth in SEQ ID NO: 104.

[0371] In various embodiments, the IL2Rβ intracellular signaling domain comprises an amino acid sequence that is at least 90% identical to SEQ ID NO: 105. In some embodiments, the IL2Rβ intracellular signaling domain comprises an amino acid sequence that is at least 95% identical to SEQ ID NO: 105. In some embodiments, the IL2Rβ intracellular signaling domain comprises an amino acid sequence that is at least 96% identical to SEQ ID NO: 105. In some embodiments, the IL2Rβ intracellular signaling domain comprises an amino acid sequence that is at least 97% identical to SEQ ID NO: 105. In some embodiments, the IL2Rβ intracellular signaling domain comprises an amino acid sequence that is at least 98% identical to SEQ ID NO: 105. In some embodiments, the IL2Rβ intracellular signaling domain comprises an amino acid sequence that is at least 99% identical to SEQ ID NO: 105. In some embodiments, the IL2Rβ intracellular signaling domain comprises an amino acid sequence as set forth in SEQ ID NO: 105.

[0372] In various embodiments, the common gamma chain intracellular signaling domain comprises an amino acid sequence that is at least 90% identical to SEQ ID NO: 106. In some embodiments, the common gamma chain intracellular signaling domain comprises an amino acid sequence that is at least 95% identical to SEQ ID NO: 106. In some embodiments, the common gamma chain intracellular signaling domain comprises an amino acid sequence that is at least 96% identical to SEQ ID NO: 106. In some embodiments, the common gamma chain intracellular signaling domain comprises an amino acid sequence that is at least 97% identical to SEQ ID NO: 106. In some embodiments, the common gamma chain intracellular signaling domain comprises an amino acid sequence that is at least 98% identical to SEQ ID NO: 106. In some embodiments, the common gamma chain intracellular signaling domain comprises an amino acid sequence that is at least 99% identical to SEQ ID NO: 106. In some embodiments, the common gamma chain intracellular signaling domain comprises an amino acid sequence as shown in SEQ ID NO: 106.

[0373] In various embodiments, the targeting component does not comprise an intracellular signaling domain or a co-stimulatory domain.

[0374] In various embodiments, the targeting component further comprises a truncated intracellular CD4 polypeptide. In some embodiments, the truncated intracellular CD4 polypeptide comprises the amino acid sequence as shown in SEQ ID NO: 48. In some embodiments, the truncated intracellular CD4 polypeptide comprises the amino acid sequence as shown in SEQ ID NO: 49.

[0375] In a preferred embodiment, the targeting component comprises an extracellular domain (comprising a first targeting domain of a VHH or scFv that binds to a target antigen), a linker polypeptide, a FRB or FKBP12 multimerization domain, a CD4 hinge, and a CD4 transmembrane domain. In various embodiments, the targeting component comprises an extracellular domain (comprising a first targeting domain of a VHH or scFv that binds to a target antigen), a linker polypeptide, a FRB or FKBP12 multimerization domain, a CD4 hinge, a CD4 transmembrane domain, and a truncated CD4ic domain. In various embodiments, the targeting component comprises an extracellular domain (comprising a first targeting domain of a VHH or scFv that binds to a target antigen), a linker polypeptide, a FRB or FKBP12 multimerization domain, a CD4 hinge, a CD4 transmembrane domain, a truncated CD4ic domain, and a 4-1BB costimulatory domain. In various embodiments, the targeting component comprises an extracellular domain (comprising a first targeting domain comprising a VHH or scFv that binds to a target antigen), a linker polypeptide, a FRB or FKBP12 multimerization domain, a CD4 hinge, a CD4 transmembrane domain, a truncated CD4ic domain, and does not comprise an intracellular signaling domain or a co-stimulatory domain.

[0376] In a preferred embodiment, the targeting component comprises an extracellular domain (comprising a first targeting domain of a VHH or scFv that binds to a target antigen), a linker polypeptide, an antibody-derived heterodimerization domain, a CD4 hinge, and a CD4 transmembrane domain. In various embodiments, the targeting component comprises an extracellular domain (comprising a first targeting domain of a VHH or scFv that binds to a target antigen), a linker polypeptide, an antibody-derived heterodimerization domain, a CD4 hinge, a CD4 transmembrane domain, and a truncated CD4ic domain. In various embodiments, the targeting component comprises an extracellular domain (comprising a first targeting domain of a VHH or scFv that binds to a target antigen), a linker polypeptide, an antibody-derived heterodimerization domain, a CD4 hinge, a CD4 transmembrane domain, a truncated CD4ic domain, and a 4-1BB costimulatory domain. In various embodiments, the targeting component comprises an extracellular domain (comprising a first targeting domain comprising a VHH or scFv that binds to a target antigen), a linker polypeptide, an antibody-derived heterodimerization domain, a CD4 hinge, a CD4 transmembrane domain, a truncated CD4ic domain, and does not comprise an intracellular signaling domain or a co-stimulatory domain.

[0377] In a preferred embodiment, the targeting component comprises an extracellular domain (comprising a first targeting domain of a VHH or scFv that binds to a target antigen), a linker polypeptide, a lock and key multimerization domain, a CD4 hinge, and a CD4 transmembrane domain. In various embodiments, the targeting component comprises an extracellular domain (comprising a first targeting domain of a VHH or scFv that binds to a target antigen), a linker polypeptide, a lock and key multimerization domain, a CD4 hinge, a CD4 transmembrane domain, and a truncated CD4ic domain. In various embodiments, the targeting component comprises an extracellular domain (comprising a first targeting domain of a VHH or scFv that binds to a target antigen), a linker polypeptide, a lock and key multimerization domain, a CD4 hinge, a CD4 transmembrane domain, a truncated CD4ic domain, and a 4-1BB costimulatory domain. In various embodiments, the targeting component comprises an extracellular domain (comprising a first targeting domain comprising a VHH or scFv that binds to a target antigen), a linker polypeptide, a lock and key multimerization domain, a CD4 hinge, a CD4 transmembrane domain, a truncated CD4ic domain, and does not comprise an intracellular signaling domain or a co-stimulatory domain.

[0378] In a preferred embodiment, the targeting component comprises an extracellular domain (comprising a first targeting domain of a VHH or scFv that binds to a target antigen), a linker polypeptide, a DEKK multimerization domain, a CD4 hinge, and a CD4 transmembrane domain. In various embodiments, the targeting component comprises an extracellular domain (comprising a first targeting domain of a VHH or scFv that binds to a target antigen), a linker polypeptide, a DEKK multimerization domain, a CD4 hinge, a CD4 transmembrane domain, and a truncated CD4ic domain. In various embodiments, the targeting component comprises an extracellular domain (comprising a first targeting domain of a VHH or scFv that binds to a target antigen), a linker polypeptide, a DEKK multimerization domain, a CD4 hinge, a CD4 transmembrane domain, a truncated CD4ic domain, and a 4-1BB costimulatory domain. In various embodiments, the targeting component comprises an extracellular domain (comprising a first targeting domain comprising a VHH or scFv that binds to a target antigen), a linker polypeptide, a DEKK multimerization domain, a CD4 hinge, a CD4 transmembrane domain, a truncated CD4ic domain, and does not comprise an intracellular signaling domain or a co-stimulatory domain.

[0379] In a preferred embodiment, the targeting component comprises an extracellular domain (comprising a first targeting domain of a VHH or scFv that binds to a target antigen), a linker polypeptide, a SEEDbody multimerization domain, a CD4 hinge, and a CD4 transmembrane domain. In various embodiments, the targeting component comprises an extracellular domain (comprising a first targeting domain of a VHH or scFv that binds to a target antigen), a linker polypeptide, a SEEDbody multimerization domain, a CD4 hinge, a CD4 transmembrane domain, and a truncated CD4ic domain. In various embodiments, the targeting component comprises an extracellular domain (comprising a first targeting domain of a VHH or scFv that binds to a target antigen), a linker polypeptide, a SEEDbody multimerization domain, a CD4 hinge, a CD4 transmembrane domain, a truncated CD4ic domain, and a 4-1BB costimulatory domain. In various embodiments, the targeting component comprises an extracellular domain (comprising a first targeting domain comprising a VHH or scFv that binds to a target antigen), a linker polypeptide, a SEEDbody multimerization domain, a CD4 hinge, a CD4 transmembrane domain, a truncated CD4ic domain, and does not comprise an intracellular signaling domain or a co-stimulatory domain.

[0380] In one embodiment, the targeting component comprises a sequence having at least 90% identity to SEQ ID NO: 116. In one embodiment, the targeting component comprises a sequence having at least 95% identity to SEQ ID NO: 116. In one embodiment, the targeting component comprises a sequence having at least 96% identity to SEQ ID NO: 116. In one embodiment, the targeting component comprises a sequence having at least 97% identity to SEQ ID NO: 116. In one embodiment, the targeting component comprises a sequence having at least 98% identity to SEQ ID NO: 116. In one embodiment, the targeting component comprises a sequence having at least 99% identity to SEQ ID NO: 116. In one embodiment, the targeting component comprises a sequence as set forth in SEQ ID NO: 116.

[0381] In one embodiment, the targeting component comprises a sequence having at least 90% identity to SEQ ID NO: 117. In one embodiment, the targeting component comprises a sequence having at least 95% identity to SEQ ID NO: 117. In one embodiment, the targeting component comprises a sequence having at least 96% identity to SEQ ID NO: 117. In one embodiment, the targeting component comprises a sequence having at least 97% identity to SEQ ID NO: 117. In one embodiment, the targeting component comprises a sequence having at least 98% identity to SEQ ID NO: 117. In one embodiment, the targeting component comprises a sequence having at least 99% identity to SEQ ID NO: 117. In one embodiment, the targeting component comprises a sequence as set forth in SEQ ID NO: 117.

[0382] In one embodiment, the targeting component comprises a sequence having at least 90% identity to SEQ ID NO: 118. In one embodiment, the targeting component comprises a sequence having at least 95% identity to SEQ ID NO: 118. In one embodiment, the targeting component comprises a sequence having at least 96% identity to SEQ ID NO: 118. In one embodiment, the targeting component comprises a sequence having at least 97% identity to SEQ ID NO: 118. In one embodiment, the targeting component comprises a sequence having at least 98% identity to SEQ ID NO: 118. In one embodiment, the targeting component comprises a sequence having at least 99% identity to SEQ ID NO: 118. In one embodiment, the targeting component comprises a sequence as set forth in SEQ ID NO: 118.

[0383] In one embodiment, the targeting component comprises a sequence that is at least 90% identical to SEQ ID NO: 119. In one embodiment, the targeting component comprises a sequence that is at least 95% identical to SEQ ID NO: 119. In one embodiment, the targeting component comprises a sequence that is at least 96% identical to SEQ ID NO: 119. In one embodiment, the targeting component comprises a sequence that is at least 97% identical to SEQ ID NO: 119. In one embodiment, the targeting component comprises a sequence that is at least 98% identical to SEQ ID NO: 119. In one embodiment, the targeting component comprises a sequence that is at least 99% identical to SEQ ID NO: 119. In one embodiment, the targeting component comprises a sequence as set forth in SEQ ID NO: 119.

[0384] In one embodiment, the targeting component comprises a sequence having at least 90% identity to SEQ ID NO: 120. In one embodiment, the targeting component comprises a sequence having at least 95% identity to SEQ ID NO: 120. In one embodiment, the targeting component comprises a sequence having at least 96% identity to SEQ ID NO: 120. In one embodiment, the targeting component comprises a sequence having at least 97% identity to SEQ ID NO: 120. In one embodiment, the targeting component comprises a sequence having at least 98% identity to SEQ ID NO: 120. In one embodiment, the targeting component comprises a sequence having at least 99% identity to SEQ ID NO: 120. In one embodiment, the targeting component comprises a sequence as set forth in SEQ ID NO: 120.

[0385] In one embodiment, the targeting component comprises a sequence having at least 90% identity to SEQ ID NO: 121. In one embodiment, the targeting component comprises a sequence having at least 95% identity to SEQ ID NO: 121. In one embodiment, the targeting component comprises a sequence having at least 96% identity to SEQ ID NO: 121. In one embodiment, the targeting component comprises a sequence having at least 97% identity to SEQ ID NO: 121. In one embodiment, the targeting component comprises a sequence having at least 98% identity to SEQ ID NO: 121. In one embodiment, the targeting component comprises a sequence having at least 99% identity to SEQ ID NO: 121. In one embodiment, the targeting component comprises a sequence as set forth in SEQ ID NO: 121.

[0386] In one embodiment, the targeting component comprises a sequence that is at least 90% identical to SEQ ID NO: 122. In one embodiment, the targeting component comprises a sequence that is at least 95% identical to SEQ ID NO: 122. In one embodiment, the targeting component comprises a sequence that is at least 96% identical to SEQ ID NO: 122. In one embodiment, the targeting component comprises a sequence that is at least 97% identical to SEQ ID NO: 122. In one embodiment, the targeting component comprises a sequence that is at least 98% identical to SEQ ID NO: 122. In one embodiment, the targeting component comprises a sequence that is at least 99% identical to SEQ ID NO: 122. In one embodiment, the targeting component comprises a sequence as set forth in SEQ ID NO: 122.

[0387] In one embodiment, the targeting component comprises a sequence having at least 90% identity to SEQ ID NO: 123. In one embodiment, the targeting component comprises a sequence having at least 95% identity to SEQ ID NO: 123. In one embodiment, the targeting component comprises a sequence having at least 96% identity to SEQ ID NO: 123. In one embodiment, the targeting component comprises a sequence having at least 97% identity to SEQ ID NO: 123. In one embodiment, the targeting component comprises a sequence having at least 98% identity to SEQ ID NO: 123. In one embodiment, the targeting component comprises a sequence having at least 99% identity to SEQ ID NO: 123. In one embodiment, the targeting component comprises a sequence as set forth in SEQ ID NO: 123.

[0388] In one embodiment, the targeting component comprises a sequence that is at least 90% identical to SEQ ID NO: 124. In one embodiment, the targeting component comprises a sequence that is at least 95% identical to SEQ ID NO: 124. In one embodiment, the targeting component comprises a sequence that is at least 96% identical to SEQ ID NO: 124. In one embodiment, the targeting component comprises a sequence that is at least 97% identical to SEQ ID NO: 124. In one embodiment, the targeting component comprises a sequence that is at least 98% identical to SEQ ID NO: 124. In one embodiment, the targeting component comprises a sequence that is at least 99% identical to SEQ ID NO: 124. In one embodiment, the targeting component comprises a sequence as set forth in SEQ ID NO: 124.

[0389] In one embodiment, the targeting component comprises a sequence that is at least 90% identical to SEQ ID NO: 125. In one embodiment, the targeting component comprises a sequence that is at least 95% identical to SEQ ID NO: 125. In one embodiment, the targeting component comprises a sequence that is at least 96% identical to SEQ ID NO: 125. In one embodiment, the targeting component comprises a sequence that is at least 97% identical to SEQ ID NO: 125. In one embodiment, the targeting component comprises a sequence that is at least 98% identical to SEQ ID NO: 125. In one embodiment, the targeting component comprises a sequence that is at least 99% identical to SEQ ID NO: 125. In one embodiment, the targeting component comprises a sequence as set forth in SEQ ID NO: 125.

[0390] In one embodiment, the targeting component comprises a sequence having at least 90% identity to SEQ ID NO: 126. In one embodiment, the targeting component comprises a sequence having at least 95% identity to SEQ ID NO: 126. In one embodiment, the targeting component comprises a sequence having at least 96% identity to SEQ ID NO: 126. In one embodiment, the targeting component comprises a sequence having at least 97% identity to SEQ ID NO: 126. In one embodiment, the targeting component comprises a sequence having at least 98% identity to SEQ ID NO: 126. In one embodiment, the targeting component comprises a sequence having at least 99% identity to SEQ ID NO: 126. In one embodiment, the targeting component comprises a sequence as set forth in SEQ ID NO: 126.

[0391] In one embodiment, the targeting component comprises a sequence having at least 90% identity to SEQ ID NO: 127. In one embodiment, the targeting component comprises a sequence having at least 95% identity to SEQ ID NO: 127. In one embodiment, the targeting component comprises a sequence having at least 96% identity to SEQ ID NO: 127. In one embodiment, the targeting component comprises a sequence having at least 97% identity to SEQ ID NO: 127. In one embodiment, the targeting component comprises a sequence having at least 98% identity to SEQ ID NO: 127. In one embodiment, the targeting component comprises a sequence having at least 99% identity to SEQ ID NO: 127. In one embodiment, the targeting component comprises a sequence as set forth in SEQ ID NO: 127.

[0392] In one embodiment, the targeting component comprises a sequence that is at least 90% identical to SEQ ID NO: 128. In one embodiment, the targeting component comprises a sequence that is at least 95% identical to SEQ ID NO: 128. In one embodiment, the targeting component comprises a sequence that is at least 96% identical to SEQ ID NO: 128. In one embodiment, the targeting component comprises a sequence that is at least 97% identical to SEQ ID NO: 128. In one embodiment, the targeting component comprises a sequence that is at least 98% identical to SEQ ID NO: 128. In one embodiment, the targeting component comprises a sequence that is at least 99% identical to SEQ ID NO: 128. In one embodiment, the targeting component comprises a sequence as set forth in SEQ ID NO: 128.

[0393] In one embodiment, the targeting component comprises a sequence having at least 90% identity to SEQ ID NO: 129. In one embodiment, the targeting component comprises a sequence having at least 95% identity to SEQ ID NO: 129. In one embodiment, the targeting component comprises a sequence having at least 96% identity to SEQ ID NO: 129. In one embodiment, the targeting component comprises a sequence having at least 97% identity to SEQ ID NO: 129. In one embodiment, the targeting component comprises a sequence having at least 98% identity to SEQ ID NO: 129. In one embodiment, the targeting component comprises a sequence having at least 99% identity to SEQ ID NO: 129. In one embodiment, the targeting component comprises a sequence as set forth in SEQ ID NO: 129.

[0394] In one embodiment, the targeting component comprises a sequence having at least 90% identity to SEQ ID NO: 130. In one embodiment, the targeting component comprises a sequence having at least 95% identity to SEQ ID NO: 130. In one embodiment, the targeting component comprises a sequence having at least 96% identity to SEQ ID NO: 130. In one embodiment, the targeting component comprises a sequence having at least 97% identity to SEQ ID NO: 130. In one embodiment, the targeting component comprises a sequence having at least 98% identity to SEQ ID NO: 130. In one embodiment, the targeting component comprises a sequence having at least 99% identity to SEQ ID NO: 130. In one embodiment, the targeting component comprises a sequence as set forth in SEQ ID NO: 130.

[0395] In one embodiment, the targeting component comprises a sequence having at least 90% identity to SEQ ID NO: 131. In one embodiment, the targeting component comprises a sequence having at least 95% identity to SEQ ID NO: 131. In one embodiment, the targeting component comprises a sequence having at least 96% identity to SEQ ID NO: 131. In one embodiment, the targeting component comprises a sequence having at least 97% identity to SEQ ID NO: 131. In one embodiment, the targeting component comprises a sequence having at least 98% identity to SEQ ID NO: 131. In one embodiment, the targeting component comprises a sequence having at least 99% identity to SEQ ID NO: 131. In one embodiment, the targeting component comprises a sequence as set forth in SEQ ID NO: 131.

[0396] In one embodiment, the targeting component comprises a sequence having at least 90% identity to SEQ ID NO: 132. In one embodiment, the targeting component comprises a sequence having at least 95% identity to SEQ ID NO: 132. In one embodiment, the targeting component comprises a sequence having at least 96% identity to SEQ ID NO: 132. In one embodiment, the targeting component comprises a sequence having at least 97% identity to SEQ ID NO: 132. In one embodiment, the targeting component comprises a sequence having at least 98% identity to SEQ ID NO: 132. In one embodiment, the targeting component comprises a sequence having at least 99% identity to SEQ ID NO: 132. In one embodiment, the targeting component comprises a sequence as set forth in SEQ ID NO: 132.

[0397] In one embodiment, the targeting component comprises a sequence having at least 90% identity to SEQ ID NO: 133. In one embodiment, the targeting component comprises a sequence having at least 95% identity to SEQ ID NO: 133. In one embodiment, the targeting component comprises a sequence having at least 96% identity to SEQ ID NO: 133. In one embodiment, the targeting component comprises a sequence having at least 97% identity to SEQ ID NO: 133. In one embodiment, the targeting component comprises a sequence having at least 98% identity to SEQ ID NO: 133. In one embodiment, the targeting component comprises a sequence having at least 99% identity to SEQ ID NO: 133. In one embodiment, the targeting component comprises a sequence as set forth in SEQ ID NO: 133.

[0398] In one embodiment, the targeting component comprises a sequence having at least 90% identity to SEQ ID NO: 134. In one embodiment, the targeting component comprises a sequence having at least 95% identity to SEQ ID NO: 134. In one embodiment, the targeting component comprises a sequence having at least 96% identity to SEQ ID NO: 134. In one embodiment, the targeting component comprises a sequence having at least 97% identity to SEQ ID NO: 134. In one embodiment, the targeting component comprises a sequence having at least 98% identity to SEQ ID NO: 134. In one embodiment, the targeting component comprises a sequence having at least 99% identity to SEQ ID NO: 134. In one embodiment, the targeting component comprises a sequence as set forth in SEQ ID NO: 134.

[0399] In one embodiment, the targeting component comprises a sequence having at least 90% identity to SEQ ID NO: 135. In one embodiment, the targeting component comprises a sequence having at least 95% identity to SEQ ID NO: 135. In one embodiment, the targeting component comprises a sequence having at least 96% identity to SEQ ID NO: 135. In one embodiment, the targeting component comprises a sequence having at least 97% identity to SEQ ID NO: 135. In one embodiment, the targeting component comprises a sequence having at least 98% identity to SEQ ID NO: 135. In one embodiment, the targeting component comprises a sequence having at least 99% identity to SEQ ID NO: 135. In one embodiment, the targeting component comprises a sequence as set forth in SEQ ID NO: 135.

[0400] In one embodiment, the targeting component comprises a sequence having at least 90% identity to SEQ ID NO: 136. In one embodiment, the targeting component comprises a sequence having at least 95% identity to SEQ ID NO: 136. In one embodiment, the targeting component comprises a sequence having at least 96% identity to SEQ ID NO: 136. In one embodiment, the targeting component comprises a sequence having at least 97% identity to SEQ ID NO: 136. In one embodiment, the targeting component comprises a sequence having at least 98% identity to SEQ ID NO: 136. In one embodiment, the targeting component comprises a sequence having at least 99% identity to SEQ ID NO: 136. In one embodiment, the targeting component comprises a sequence as set forth in SEQ ID NO: 136.

[0401] In one embodiment, the targeting component does not comprise the sequence shown in SEQ ID NO:122.

[0402] In various embodiments, the targeting component contemplated herein comprises a signal peptide, such as a secretory signal polypeptide or sequence. Illustrative examples of signal polypeptides / sequences suitable for use in specific targeting components include, but are not limited to, IgG1 heavy chain signal polypeptides, Igκ light chain signal polypeptides, CD8α signal polypeptides, or human GM-CSF receptor α signal polypeptides. In various preferred embodiments, the targeting component comprises an Igκ light chain signal polypeptide. In some embodiments, the Igκ light chain signal polypeptide comprises an amino acid sequence having at least 90% identity to SEQ ID NO: 95. In some embodiments, the Igκ light chain signal polypeptide comprises an amino acid sequence having at least 95% identity to SEQ ID NO: 95. In some embodiments, the Igκ light chain signal polypeptide comprises an amino acid sequence having at least 96% identity to SEQ ID NO: 95. In some embodiments, the Igκ light chain signal polypeptide comprises an amino acid sequence having at least 97% identity to SEQ ID NO: 95. In some embodiments, the Igκ light chain signal polypeptide comprises an amino acid sequence having at least 98% identity to SEQ ID NO: 95. In some embodiments, the Ig kappa light chain signal polypeptide comprises an amino acid sequence that is at least 99% identical to SEQ ID NO: 95. In some embodiments, the Ig kappa light chain signal polypeptide comprises an amino acid sequence as shown in SEQ ID NO:95.

[0403] In various embodiments, the signaling component comprises a CD8α signaling polypeptide. In some embodiments, the CD8α light chain signaling polypeptide comprises an amino acid sequence having at least 90% identity with SEQ ID NO: 96. In some embodiments, the CD8α light chain signaling polypeptide comprises an amino acid sequence having at least 95% identity with SEQ ID NO: 96. In some embodiments, the CD8α light chain signaling polypeptide comprises an amino acid sequence having at least 96% identity with SEQ ID NO: 96. In some embodiments, the CD8α light chain signaling polypeptide comprises an amino acid sequence having at least 97% identity with SEQ ID NO: 96. In some embodiments, the CD8α light chain signaling polypeptide comprises an amino acid sequence having at least 98% identity with SEQ ID NO: 96. In some embodiments, the CD8α light chain signaling polypeptide comprises an amino acid sequence having at least 99% identity with SEQ ID NO: 96. In some embodiments, the CD8α light chain signaling polypeptide comprises an amino acid sequence as shown in SEQ ID NO: 96.

[0404] In some embodiments, the signal polypeptide is a PD1 signal polypeptide. In one embodiment, the PD1 signal polypeptide comprises an amino acid sequence having at least 90% identity with SEQ ID NO: 97. In one embodiment, the PD1 signal polypeptide comprises an amino acid sequence having at least 95% identity with SEQ ID NO: 97. In one embodiment, the PD1 signal polypeptide comprises an amino acid sequence having at least 96% identity with SEQ ID NO: 97. In one embodiment, the PD1 signal polypeptide comprises an amino acid sequence having at least 97% identity with SEQ ID NO: 97. In one embodiment, the PD1 signal polypeptide comprises an amino acid sequence having at least 98% identity with SEQ ID NO: 97. In one embodiment, the PD1 signal polypeptide comprises an amino acid sequence having at least 99% identity with SEQ ID NO: 97. In one embodiment, the PD1 signal polypeptide comprises an amino acid sequence as shown in SEQ ID NO: 97.

[0405] 3. Bridging Factor

[0406] The bridging factors envisioned in the specific embodiments herein mediate or promote the association of one or more signaling components with one or more targeting components through the multimerization domains in the corresponding components. The bridging factor associates with the multimerization domain and is placed between the multimerization domains to promote the association of the signaling component with the targeting component. In the presence of the bridging factor, when the targeting component binds to the target antigen on the target cell, the targeting component and the signaling component bind to the immune receptor complex and initiate the immune effector cell activity against the target cell. In the absence of the bridging factor, the targeting component does not associate with the signaling component, does not recruit the immune receptor complex, and the engineered receptor has no activity against the target antigen of the targeting component.

[0407] In particular embodiments, the signaling component and the targeting component comprise a homologous multimerization domain pair selected from the group consisting of: FKBP and FKBP12-rapamycin binding (FRB), FKBP and calcineurin, FKBP and cyclophilin, FKBP and bacterial dihydrofolate reductase (DHFR), calcineurin and cyclophilin, and PYR1-like 1 (PYL1) and abscisic acid insensitive 1 (ABI1).

[0408] In certain embodiments, the multimerization domains of the signaling component and the targeting component are associated with a bridging factor selected from the group consisting of rapamycin or a rapamycin analog thereof, kumarins or a derivative thereof, gibberellins or a derivative thereof, abscisic acid (ABA) or a derivative thereof, methotrexate or a derivative thereof, cyclosporine A or a derivative thereof, FK506 / cyclosporine A (FKCsA) or a derivative thereof, and a trimethoprim (Tmp) synthetic ligand (SLF) for FK506 binding protein (FKBP) or a derivative thereof.

[0409] In certain embodiments, the signaling component and the targeting component comprise one or more FRB and / or FKBP multimerization domains or variants thereof. In certain embodiments, the signaling component comprises a FKBP12 multimerization domain or a variant thereof, and the targeting component comprises a FRB multimerization domain or a variant thereof. In certain embodiments, the signaling component comprises a FRB multimerization domain or a variant thereof, and the targeting component comprises a FKBP12 multimerization domain or a variant thereof. In certain preferred embodiments, the signaling component comprises a FKBP12 or FKBP12 F36V multimerization domain or a variant thereof, and the targeting component comprises a FRBT2098L multimerization domain or a variant thereof. In certain preferred embodiments, the CD3 signaling component comprises a FRB T2098L (T82L) multimerization domain or a variant thereof, and the targeting component comprises a FKBP12 or FKBP12 F36V multimerization domain or a variant thereof.

[0410] Illustrative examples of bridging factors suitable for use in the specific embodiments contemplated herein include, but are not limited to, AP1903, AP20187, AP21967, (also known as C-16-(S)-7-methylindorapamycin), everolimus, norvolimus, pimecrolimus, dafoslimus, sirolimus, tacrolimus, temsirolimus, usololimus, zotarolimus, or BPC015. In certain preferred embodiments, the bridging factor is AP21967. In certain preferred embodiments, the bridging factor is a non-immunosuppressive dose of sirolimus (rapamycin).

[0411] D. Antigen receptors and binding molecules

[0412] In certain embodiments, the immune effector cells contemplated herein further express exogenous lymphocyte receptors or engineered antigen receptors and one or more components of engineered...

Claims

1. A non-natural cell comprising: (a) a signaling component comprising a first multimerization domain and an actuator domain; and (b) a targeting component, which comprises an extracellular domain, a second multimerization domain and a transmembrane domain.

2. The non-natural cell of claim 1, wherein the second multimerization domain and the transmembrane domain are separated by a hinge domain.

3. The non-natural cell of claim 2, wherein the hinge domain is selected from the group consisting of a CD4 hinge, a CD8 hinge, a CD28 hinge, an IgG4 hinge, and any fragment or variant or combination thereof.

4. The non-natural cell of claim 2 or claim 3, wherein the hinge domain is a CD4 hinge.

5. The non-natural cell of claim 4, wherein the CD4 hinge comprises an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

41.

6. The non-natural cell of claim 5, wherein the CD4 hinge comprises the amino acid sequence as shown in SEQ ID NO:

41.

7. The non-natural cell of claim 2 or claim 3, wherein the hinge domain is a CD28 hinge.

8. The non-natural cell of claim 7, wherein the CD28 hinge comprises an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

42.

9. The non-natural cell of claim 8, wherein the CD28 hinge comprises the amino acid sequence as shown in SEQ ID NO:

42.

10. The non-natural cell of claim 1 or claim 2, wherein the hinge domain is a CD8 hinge.

11. The non-natural cell of claim 10, wherein the CD8 hinge comprises an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

44.

12. The non-natural cell of claim 11, wherein the CD8 hinge comprises the amino acid sequence as shown in SEQ ID NO:

44.

13. The non-natural cell of claim 1 or claim 2, wherein the hinge domain is an IgG4 hinge.

14. The non-natural cell of claim 13, wherein the IgG4 hinge comprises an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

43.

15. The non-natural cell of claim 14, wherein the IgG4 hinge comprises the amino acid sequence as shown in SEQ ID NO:

43.

16. The non-natural cell of any one of the preceding claims, wherein the actuator domain is a CD3 polypeptide, an FcεR1γ polypeptide, an Igα / CD79a polypeptide, an Igβ / CD79b polypeptide, a DAP10 polypeptide, or a DAP12 polypeptide.

17. The non-natural cell of any one of the preceding claims, wherein the CD3 polypeptide is a CD3 epsilon polypeptide or a variant thereof, CD3 gamma or a variant thereof, or CD3 delta or a variant thereof.

18. The non-natural cell of any one of the preceding claims, wherein the actuator domain is a CD3ε polypeptide or a variant thereof.

19. The non-natural cell of any of the preceding claims, wherein the actuator domain is a CD3ε polypeptide comprising an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

32.

20. The non-natural cell of any one of the preceding claims, wherein the actuator domain is a CD3ε polypeptide comprising the amino acid sequence as shown in SEQ ID NO:

32.

21. The non-natural cell of any one of claims 1 to 17, wherein the actuator domain is a CD3γ polypeptide or a variant thereof.

22. The non-natural cell of any one of claims 1 to 17 or 21, wherein the actuator domain is a CD3γ polypeptide comprising an amino acid sequence at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

33.

23. The non-natural cell of any one of claims 1 to 17, 21 or 22, wherein the actuator domain is a CD3γ polypeptide comprising the amino acid sequence as shown in SEQ ID NO:

33.

24. The non-natural cell of any one of claims 1 to 17, wherein the actuator domain is a CD3 delta polypeptide or a variant thereof.

25. The non-natural cell of any one of claims 1 to 17 or 24, wherein the actuator domain is a CD3 delta polypeptide comprising an amino acid sequence at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

34.

26. The non-natural cell of any one of claims 1 to 7, 24 or 25, wherein the actuator domain is a CD3 delta polypeptide comprising the amino acid sequence as shown in SEQ ID NO:

34.

27. The non-natural cell of any one of claims 1 to 16, wherein the actuator domain is an FcεR1γ polypeptide or a variant thereof.

28. The non-natural cell of any one of claims 1 to 16 or claim 27, wherein the actuator domain is an FcεR1γ polypeptide comprising an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

35.

29. The non-natural cell of any one of claims 1 to 16, 27 or 28, wherein the actuator domain is an FcεR1γ polypeptide comprising the amino acid sequence as shown in SEQ ID NO:

35.

30. The non-natural cell of any one of claims 1 to 16 or claim 27, wherein the actuator domain is an FcεR1γ polypeptide comprising an amino acid sequence at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

36.

31. The non-natural cell of any one of claims 1 to 16, 27 or 30, wherein the actuator domain is an FcεR1γ polypeptide comprising the amino acid sequence as shown in SEQ ID NO:

36.

32. The non-natural cell of any one of claims 1 to 16, wherein the actuator domain is an Igα / CD79a polypeptide or a variant thereof.

33. The non-natural cell of any one of claims 1 to 16 or 32, wherein the actuator domain is an Igα / CD79a polypeptide comprising an amino acid sequence at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

37.

34. The non-natural cell of any one of claims 1 to 16, 32 or 33, wherein the actuator domain is an Igα / CD79a polypeptide comprising the amino acid sequence shown in SEQ ID NO:

37.

35. The non-natural cell of any one of claims 1 to 16, wherein the actuator domain is an Igβ / CD79b polypeptide or a variant thereof.

36. The non-natural cell of any one of claims 1 to 16 or 35, wherein the actuator domain is an Igβ / CD79b polypeptide comprising an amino acid sequence at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

38.

37. The non-natural cell of any one of claims 1 to 16, 35 or 36, wherein the actuator domain is an Igβ / CD79b polypeptide comprising the amino acid sequence shown in SEQ ID NO:

38.

38. The non-natural cell of any one of claims 1 to 16, wherein the actuator domain is a DAP10 polypeptide or a variant thereof.

39. The non-natural cell of any one of claims 1 to 16 or 38, wherein the actuator domain is a DAP10 polypeptide comprising an amino acid sequence at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

39.

40. The non-natural cell of any one of claims 1 to 16, 38 or 39, wherein the actuator domain is a DAP10 polypeptide comprising the amino acid sequence shown in SEQ ID NO:

39.

41. The non-natural cell of any one of claims 1 to 16, wherein the actuator domain is a DAP12 polypeptide or a variant thereof.

42. The non-natural cell of any one of claims 1 to 16 or 41, wherein the actuator domain is a DAP12 polypeptide comprising an amino acid sequence at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

40.

43. The non-natural cell of any one of claims 1 to 16, 41 or 42, wherein the actuator domain is a DAP12 polypeptide comprising the amino acid sequence shown in SEQ ID NO:

40.

44. The non-natural cell of any of the preceding claims, wherein the actuator domain comprises both an extracellular portion and an intracellular portion.

45. The non-natural cell of any of the preceding claims, wherein when the signaling component and the targeting component are expressed, the first multimerization domain and the second multimerization domain are located extracellularly.

46. ​​The non-natural cell of any of the preceding claims, wherein the first multimerization domain and the second multimerization domain are different.

47. The non-natural cell of any of the preceding claims, wherein the multimerization domains of the signaling component and the targeting component are associated with a bridging factor selected from the group consisting of rapamycin or a rapamycin analog thereof, gibberellin or a derivative thereof, abscisic acid (ABA) or a derivative thereof, methotrexate or a derivative thereof, cyclosporine A or a derivative thereof, FK506 / cyclosporine A or a derivative thereof, trimethoprim (Tmp) synthetic ligand (SLF) for FK506 binding protein (FKBP) or a derivative thereof, wherein the bridging factor promotes the formation of a polypeptide complex, wherein the bridging factor is associated with the multimerization domains of the signaling component and the targeting component and is positioned between the multimerization domains.

48. The non-natural cell of any of the preceding claims, wherein the first and second multimerization domains are a pair selected from the group consisting of: FK506 binding protein 1A (FKBP12) and FKBP12-rapamycin binding (FRB), FKBP12 and calcineurin, FKBP and cyclophilin A or any other member of the peptidyl-prolyl cis-trans isomerase (PPIase) family, FKBP and dihydrofolate reductase (DHFR), calcineurin and cyclophilin A or any other member of the peptidyl-prolyl cis-trans isomerase (PPIase) family, and PYR1-like 1 (PYL1) and abscisic acid insensitive 1 (ABI1).

49. The non-natural cell of any of the preceding claims, wherein the first multimerization domain comprises a first FRB polypeptide or a variant thereof, and the second multimerization domain comprises a first FKBP12 polypeptide or a variant thereof.

50. The non-natural cell of any one of claims 1 to 48, wherein the first multimerization domain comprises a first FKBP12 polypeptide or a variant thereof, and the second multimerization domain comprises a first FRB polypeptide or a variant thereof.

51. The non-natural cell of any one of claims 48 to 50, wherein the FRB polypeptide is the FRB T2098L variant.

52. The non-natural cell of any one of claims 48 to 51, wherein the FRB polypeptide comprises an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO: 1 or SEQ ID NO: 2, or comprises the sequence as shown in SEQ ID NO: 1 or SEQ ID NO:

2.

53. The non-natural cell of any one of claims 48 to 52, wherein the FKBP12 polypeptide comprises an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to, or comprises the sequence as shown in, SEQ ID NO: 3 or SEQ ID NO:

4.

54. The non-natural cell of any of the preceding claims, wherein the bridging factor is AP1903, AP20187, AP21967 (also known as C16-(S)-7-methylindorapamycin), everolimus, novolimus, pimecrolimus, ridaforolimus, sirolimus, tacrolimus, temsirolimus, umirolimus, zotarolimus, or BPC015.

55. The non-natural cell of any one of claims 1 to 46, wherein the first multimerization domain and the second multimerization domain are a pair of antibody-derived heterodimerization domains.

56. A non-natural cell according to any one of claims 1 to 46 or 55, wherein the first multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98% or 99% identity with SEQ ID NO:5 or comprises the amino acid sequence as shown in SEQ ID NO:5; and wherein the second multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98% or 99% identity with SEQ ID NO:6 or comprises the amino acid sequence as shown in SEQ ID NO:

6.

57. A non-natural cell according to any one of claims 1 to 46 or 55, wherein the first multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98% or 99% identity with SEQ ID NO:6 or comprises the amino acid sequence as shown in SEQ ID NO:6; and wherein the second multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98% or 99% identity with SEQ ID NO:5 or comprises the amino acid sequence as shown in SEQ ID NO:

5.

58. A non-natural cell according to any one of claims 1 to 46 or 55, wherein the first multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98% or 99% identity with SEQ ID NO:7 or comprises the amino acid sequence as shown in SEQ ID NO:7; and wherein the second multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98% or 99% identity with SEQ ID NO:8 or comprises the amino acid sequence as shown in SEQ ID NO:

8.

59. A non-natural cell according to any one of claims 1 to 46 or 55, wherein the first multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98% or 99% identity with SEQ ID NO:8 or comprises the amino acid sequence as shown in SEQ ID NO:8; and wherein the second multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98% or 99% identity with SEQ ID NO:7 or comprises the amino acid sequence as shown in SEQ ID NO:

7.

60. The non-natural cell of any one of claims 1 to 46 or 55, wherein the first multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO:9 or comprises the amino acid sequence as shown in SEQ ID NO:9; and wherein the second multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98% or 99% identity to SEQ ID NO:10 or comprises the amino acid sequence as shown in SEQ ID NO:

10.

61. A non-natural cell according to any one of claims 1 to 46 or 55, wherein the first multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98% or 99% identity with SEQ ID NO:10 or comprises the amino acid sequence as shown in SEQ ID NO:10; and wherein the second multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98% or 99% identity with SEQ ID NO:9 or comprises the amino acid sequence as shown in SEQ ID NO:

9.

62. The non-natural cell of any of the preceding claims, wherein the first multimerization domain and the actuator domain are separated by a first polypeptide linker having a length of 2 to 40 amino acids.

63. The non-natural cell of claim 62, wherein the first polypeptide linker is selected from the group consisting of GG, GS, SG, SS, GSS, SSG, GSG, SGS, SGG, G4S, 2xG4S, 3xG4S, 4xG4S, 5xG4S, and any combination thereof.

64. The non-natural cell of claim 63, wherein the first polypeptide linker is a 3xG4S linker.

65. The non-natural cell of claim 62, wherein the first polypeptide linker comprises a polypeptide comprising an amino acid sequence as shown in any one of SEQ ID NOs: 16-31.

66. The non-natural cell of any of the preceding claims, wherein the extracellular domain and the second multimerization domain are separated by a second polypeptide linker having a length of 2 to 40 amino acids.

67. The non-natural cell of claim 66, wherein the second polypeptide linker is selected from the group consisting of GG, GS, SG, SS, GSS, SSG, GSG, SGS, SGG, G4S, 2xG4S, 3xG4s, 4xG4S, and any combination thereof.

68. The non-natural cell of claim 67, wherein the second polypeptide linker is a G4S linker.

69. The non-natural cell of claim 66, wherein the second polypeptide linker comprises a polypeptide comprising an amino acid sequence as shown in any one of SEQ ID NOs: 16-31.

70. The non-natural cell of any one of claims 1 to 69, wherein the transmembrane domain is a CD4 transmembrane domain.

71. The non-natural cell of claim 70, wherein the CD4 transmembrane domain comprises an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

45.

72. The non-natural cell of claim 71, wherein the CD4 transmembrane domain comprises the amino acid sequence shown in SEQ ID NO:

45.

73. The non-natural cell of any one of claims 1 to 69, wherein the transmembrane domain is a CD28 transmembrane domain.

74. The non-natural cell of claim 73, wherein the CD28 transmembrane domain comprises an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

46.

75. The non-natural cell of claim 74, wherein the CD28 transmembrane domain comprises the amino acid sequence shown in SEQ ID NO:

46.

76. The non-natural cell of any one of claims 1 to 69, wherein the transmembrane domain is a CD8 transmembrane domain.

77. The non-natural cell of claim 76, wherein the CD8 transmembrane domain comprises an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

47.

78. The non-natural cell of claim 77, wherein the CD8 transmembrane domain comprises the amino acid sequence shown in SEQ ID NO:

47.

79. The non-natural cell of any of the preceding claims, wherein the targeting component further comprises an intracellular signaling domain or a co-stimulatory domain derived from a protein selected from the group consisting of an antigen receptor, a co-stimulatory receptor, a growth receptor, a cytokine receptor, an adaptor signaling protein, an intracellular signaling protein, or any fragment or variant thereof.

80. The non-natural cell of claim 79, wherein the intracellular signaling domain or the co-stimulatory domain on the targeting component is selected from the group consisting of: Toll-like receptor 1 (TLR1), TLR2, TLR3, TLR4, TLR5, TLR6, TLR7, TLR8, TLR9, TLR10, caspase recruitment domain family member 11 (CARD11), CD2, CD3ε, CD3γ, CD3δ, CD4, CD7, CD8, CD27, CD28, CD30, CD40, CD54 (ICAM), CD83, CD94, CD134 (OX40), CD1 37 (4-1BB), CD278 (ICOS), common gamma chain cytokine, DNAX-activating protein 10 (DAP10), adaptor for activating T cells family member 1 (LAT), interleukin 2 receptor (IL-2R), IL-4R, IL-7R, IL-9R, IL-12R, IL-13R, IL-15R, IL-21R, SH2 domain-containing leukocyte protein of 76 kD (SLP76), T cell receptor-associated transmembrane adaptor 1 (TRAT1), TNFR2, TNFRS14, TNFRS18, TNRFS25, and T cell receptor zeta chain-associated protein kinase 70 (ZAP70).

81. The non-natural cell of claim 79 or claim 80, wherein the intracellular signaling domain or the co-stimulatory domain on the targeting component is selected from the group consisting of: 4-1BB, CD28, TNFR2, OX40, ICOS, and DAP10 co-stimulatory domains.

82. The non-natural cell of any one of claims 79 to 81, wherein the co-stimulatory domain on the targeting component is a 4-1BB co-stimulatory domain, optionally wherein the 4-1BB co-stimulatory domain comprises the amino acid sequence shown in SEQ ID NO:

98.

83. The non-natural cell of any one of claims 79 to 81, wherein the co-stimulatory domain on the targeting component is a CD28 co-stimulatory domain, optionally wherein the CD28 co-stimulatory domain comprises the amino acid sequence as shown in SEQ ID NO:

99.

84. The non-natural cell of claim 79, wherein the intracellular signaling domain or the co-stimulatory domain is one or more cytokine receptor intracellular signaling domains.

85. The non-natural cell of claim 84, wherein the one or more cytokine receptor intracellular signaling domains are selected from the group consisting of an IL7Rα intracellular signaling domain, an IL2Rβ intracellular signaling domain, a common gamma chain intracellular signaling domain, and both IL2Rβ and common gamma chain intracellular signaling domains.

86. The non-natural cell of claim 85, wherein the IL7Rα intracellular signaling domain comprises an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

103.

87. The non-natural cell of claim 86, wherein the IL7Rα intracellular signaling domain comprises the amino acid sequence as shown in SEQ ID NO:

103.

88. The non-natural cell of claim 85, wherein the IL2Rβ intracellular signaling domain comprises an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO: 104 or SEQ ID NO:

105.

89. The non-natural cell of claim 88, wherein the IL2Rβ intracellular signaling domain comprises the amino acid sequence as shown in SEQ ID NO: 104 or SEQ ID NO:

105.

90. The non-natural cell of claim 85, wherein the common gamma chain intracellular signaling domain comprises an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

106.

91. The non-natural cell of claim 90, wherein the common gamma chain intracellular signaling domain comprises the amino acid sequence shown in SEQ ID NO:

106.

92. The non-natural cell of claim 79, wherein the intracellular signaling domain or the co-stimulatory domain is a LAT domain.

93. The non-natural cell of claim 92, wherein the LAT domain comprises an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to the amino acid sequence shown in SEQ ID NO:

102.

94. The non-natural cell of claim 93, wherein the LAT domain comprises the amino acid sequence shown in SEQ ID NO:

102.

95. The non-natural cell of claim 79, wherein the intracellular signaling domain or the co-stimulatory domain is a CD4 coreceptor domain.

96. The non-natural cell of claim 95, wherein the CD4 coreceptor domain comprises an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

100.

97. The non-natural cell of claim 96, wherein the CD4 coreceptor domain comprises the amino acid sequence shown in SEQ ID NO:

100.

98. The non-natural cell of claim 79, wherein the intracellular signaling domain or the co-stimulatory domain is a CD8 coreceptor domain.

99. The non-natural cell of claim 98, wherein the CD8 coreceptor domain comprises an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

101.

100. The non-natural cell of claim 99, wherein the CD8 co-receptor domain comprises the amino acid sequence shown in SEQ ID NO:

101.

101. The non-natural cell of any one of claims 1 to 78, wherein the targeting component does not comprise a functional intracellular signaling domain or a co-stimulatory domain.

102. The non-natural cell of any one of claims 1 to 101, wherein the targeting component further comprises a truncated intracellular CD4 polypeptide.

103. The non-natural cell of claim 102, wherein the truncated intracellular CD4 polypeptide comprises the amino acid sequence shown in SEQ ID NO:48 or SEQ ID NO:

49.

104. The non-natural cell of claim 103, wherein the truncated intracellular CD4 polypeptide comprises the amino acid sequence shown in SEQ ID NO:48 or SEQ ID NO:

49.

105. The non-natural cell of any one of claims 1 to 104, wherein the extracellular domain comprises a first targeting domain.

106. The non-natural cell of any of the preceding claims, wherein the first targeting domain comprises a single-chain variable fragment (scFv) or a single domain antibody (sdAb).

107. The non-natural cell of claim 106, wherein the sdAb is a camelid VHH, a nanobody, or a heavy chain only antibody (HcAb).

108. The non-natural cell of claim 107, wherein the sdAb is a camelid VHH.

109. The non-natural cell of any one of claims 106 to 108, wherein the scFv or the sdAb is human or humanized.

110. The non-natural cell of any one of claims 105 to 109, wherein the first targeting domain comprises a sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to any one of SEQ ID NOs: 50-83.

111. The non-natural cell of claim 110, wherein the first targeting domain comprises a sequence as shown in any one of SEQ ID NOs: 50-83.

112. The non-natural cell of claim 105, wherein the first targeting domain comprises the extracellular domain of PD1, human A proliferation-inducing ligand (APRIL), trimerized human APRIL, or NKG2D membrane protein.

113. The non-natural cell of claim 112, wherein the PD1 extracellular domain comprises a sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:84 or SEQ ID NO:

85.

114. The non-natural cell of claim 113, wherein the PD1 extracellular domain comprises the sequence shown in SEQ ID NO:

85.

115. The non-natural cell of claim 112, wherein the APRIL comprises a sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:86, or wherein the trimerized human APRIL comprises a sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

87.

116. The non-natural cell of claim 115, wherein the APRIL comprises the sequence as shown in SEQ ID NO:86, or wherein the trimerized human APRIL comprises the sequence as shown in SEQ ID NO:

87.

117. The non-natural cell of claim 112, wherein the NKG2D membrane protein comprises a sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

88.

118. The non-natural cell of claim 117, wherein the NKG2D membrane protein comprises the sequence shown in SEQ ID NO:

88.

119. The non-natural cell of any one of claims 105 to 118, wherein the extracellular domain further comprises a second targeting domain.

120. The non-natural cell of claim 119, wherein the second targeting domain comprises a second single chain variable fragment (scFv) or a second single domain antibody (sdAb).

121. The non-natural cell of claim 120, wherein the second sdAb is a camelid VHH, a nanobody, or a heavy chain only antibody (HcAb).

122. The non-natural cell of claim 121, wherein the second sdAb is a camelid VHH.

123. The non-natural cell of any one of claims 119 to 122, wherein the second scFv or the second sdAb is human or humanized.

124. The non-natural cell of any one of claims 119 to 123, wherein the second targeting domain comprises a sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to any one of SEQ ID NOs: 50-83.

125. The non-natural cell of claim 124, wherein the second targeting domain comprises a sequence as shown in any one of SEQ ID NOs: 50-83.

126. The non-natural cell of claim 119, wherein the second targeting domain comprises the PD1 extracellular domain, human A proliferation-inducing ligand (APRIL), trimerized human APRIL, or NKG2D membrane protein.

127. The non-natural cell of claim 126, wherein the PD1 extracellular domain comprises a sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:84 or SEQ ID NO:

85.

128. The non-natural cell of claim 127, wherein the PD1 extracellular domain comprises the sequence shown in SEQ ID NO:

85.

129. The non-natural cell of claim 126, wherein the APRIL comprises a sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:86, or wherein the trimerized human APRIL comprises a sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

87.

130. The non-natural cell of claim 129, wherein the APRIL comprises the sequence as shown in SEQ ID NO:86, or wherein the trimerized human APRIL comprises the sequence as shown in SEQ ID NO:

87.

131. The non-natural cell of claim 126, wherein the NKG2D membrane protein comprises a sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

88.

132. The non-natural cell of claim 131, wherein the NKG2D membrane protein comprises the sequence shown in SEQ ID NO:

88.

133. The non-natural cell of any one of claims 119 to 132, wherein the targeting domain and the second targeting domain bind to the same antigen or different antigens.

134. The non-natural cell of any one of claims 119 to 132, wherein the targeting domain and the second targeting domain are separated by a third polypeptide linker having a length of 2 to 40 amino acids.

135. The non-natural cell of claim 134, wherein the third polypeptide linker is selected from the group consisting of GG, GS, SG, SS, GSS, SSG, GSG, SGS, SGG, G4S, 2xG4S, 3xG4s, 4xG4S, 5xG4S, any one of SEQ ID NOs: 16-31, and any combination thereof.

136. A non-natural cell according to any of the preceding claims, wherein the signaling component comprises a sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO: 107, SEQ ID NO: 108, SEQ ID NO: 109, SEQ ID NO: 110, SEQ ID NO: 111, SEQ ID NO: 112, SEQ ID NO: 113, SEQ NO: 114 or SEQ ID NO:

115.

137. The non-natural cell of claim 136, wherein the signaling component comprises a sequence as shown in SEQ ID NO: 107, SEQ ID NO: 108, SEQ ID NO: 109, SEQ ID NO: 110, SEQ ID NO: 111, SEQ ID NO: 112, SEQ ID NO: 113, SEQ NO: 114, or SEQ ID NO:

115.

138. A non-natural cell according to any of the preceding claims, wherein the targeting component comprises a sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to the sequence shown in SEQ ID NO: 116, SEQ ID NO: 117, SEQ ID NO: 118, SEQ ID NO: 119, SEQ ID NO: 120, SEQ ID NO: 121, SEQ ID NO: 122, SEQ ID NO: 123, SEQ ID NO: 124, SEQ ID NO: 125, SEQ ID NO: 126, SEQ ID NO: 127, SEQ ID NO: 128, SEQ ID NO: 129, SEQ ID NO: 130, SEQ ID NO: 131, SEQ ID NO: 132, SEQ ID NO: 133, SEQ ID NO: 134, SEQ ID NO: 135 or SEQ ID NO:

136.

139. The non-natural cell of claim 138, wherein the targeting component comprises a sequence as shown in SEQ ID NO: 116, SEQ ID NO: 117, SEQ ID NO: 118, SEQ ID NO: 119, SEQ ID NO: 120, SEQ ID NO: 121, SEQ ID NO: 122, SEQ ID NO: 123, SEQ ID NO: 124, SEQ ID NO: 125, SEQ ID NO: 126, SEQ ID NO: 127, SEQ ID NO: 128, SEQ ID NO: 129, SEQ ID NO: 130, SEQ ID NO: 131, SEQ ID NO: 132, SEQ ID NO: 133, SEQ ID NO: 134, SEQ ID NO: 135, or SEQ ID NO:

136.

140. The non-natural cell of any of the preceding claims, comprising a fusion polypeptide comprising the targeting component and the signaling component.

141. The non-natural cell of claim 140, wherein the fusion polypeptide comprises a polypeptide having a residue of SEQ ID NO: 139, SEQ ID NO: 140, SEQ ID NO: 141, SEQ ID NO: 142, SEQ ID NO: 143, SEQ ID NO: 144, SEQ ID NO: 145, SEQ ID NO: 146, SEQ ID NO: 147, SEQ ID NO: 148, SEQ ID NO: 149, SEQ ID NO: 150, SEQ ID NO: 151, SEQ ID NO: 152, SEQ ID NO: 153, SEQ ID NO: 154, SEQ ID NO: 155, SEQ ID NO: 156, SEQ ID NO: 157, SEQ ID NO: 158, SEQ ID NO: 159, SEQ ID NO: 160, SEQ ID NO: 161, SEQ ID NO: 162, SEQ ID NO: 163, SEQ ID NO: 164, SEQ ID NO: 165, SEQ ID NO: 166, SEQ ID NO: 167, SEQ ID NO: 167, SEQ ID NO: 168, SEQ ID NO: 169 or SEQ ID NO: 170 having a sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical.

142. The non-natural cell of claim 141, wherein the fusion polypeptide comprises SEQ ID NO: 139, SEQ ID NO: 140, SEQ ID NO: 141, SEQ ID NO: 142, SEQ ID NO: 143, SEQ ID NO: 144, SEQ ID NO: 145, SEQ ID NO: 146, SEQ ID NO: 147, SEQ ID NO: 148, SEQ ID NO: 149, SEQ ID NO: 150, SEQ ID NO: 151, SEQ ID NO: 152, SEQ ID NO: 153, SEQ ID NO: 154, SEQ ID NO: 155, SEQ ID NO: 156, SEQ ID NO: 157, SEQ ID NO: 158, SEQ ID NO: 159, SEQ ID NO: 160, SEQ ID NO: 161, SEQ ID NO: 162, SEQ ID NO: 163, SEQ ID NO: 164, SEQ ID NO: 165, SEQ ID NO: 166, SEQ ID NO: 167, SEQ ID NO: 168, The sequence shown in SEQ ID NO:167, SEQ ID NO:168, SEQ ID NO:169 or SEQ ID NO:

170.

143. The non-natural cell of any of the preceding claims, wherein the cell comprises a first nucleic acid molecule encoding the signaling component.

144. The non-natural cell of any of the preceding claims, wherein the cell comprises a second nucleic acid molecule encoding the targeting component.

145. The non-natural cell of any of the preceding claims, wherein the cell comprises a nucleic acid molecule encoding both the signaling component and the targeting component.

146. The non-natural cell of any of the preceding claims, wherein the cell further expresses an exogenous co-stimulatory factor, an immunomodulatory factor, an agonist of a co-stimulatory factor, an antagonist of an immunosuppressive factor, an immune cell engager, a flip receptor, or any combination thereof.

147. The non-natural cell of any of the preceding claims, wherein the cell further expresses an exogenous lymphocyte receptor or co-receptor.

148. The non-natural cell of claim 147, wherein the exogenous lymphocyte receptor or co-receptor is selected from the group consisting of TCRα (TCR alpha), TCRβ (TCR beta), TCRγ (TCR gamma), TCRδ (TCRdelta), CD4, CD8, pre-T cell receptor alpha (pTα), Fc receptor alpha (FcRα), Fc receptor beta (FcRβ), Fc receptor gamma (FcRγ), natural killer group 2 member D (NKG2D), CD79A, CD79B, and any combination thereof.

149. The non-natural cell of any of the preceding claims, wherein the cell further expresses an exogenous TCR.

150. The non-natural cell of claim 149, wherein the exogenous TCR binds to a target antigen selected from the group consisting of alpha-fetoprotein (AFP), members of the B melanoma antigen (BAGE) family, brothers of regulatory factors at imprinted sites (BORIS), cancer-testis antigen, cancer-testis antigen 83 (CT-83), carbonic anhydrase IX (CA1X), carcinoembryonic antigen (CEA), cytomegalovirus (CMV) antigen, antigen recognized by cytotoxic T cells (CTL) on melanoma (CAMEL), Epstein-Barr virus (Epstein-Barr virus, EBV) antigen, G antigen 1 (GAGE-1), GAGE-2, GAGE-3, GAGE-4, GAGE-5, GAGE-6, GAGE-7B, GAGE-8, glycoprotein 100 (GP100), hepatitis B virus (HBV) antigen, hepatitis C virus (HCV) nonstructural protein 3 (NS3), human epidermal growth factor receptor 2 (HER-2), human papillomavirus (HPV)-E6, HPV-E7, human telomerase reverse transcriptase (hTERT), IGF2BP3 / A3, K-Ras, K-Ras G12C, K-Ras G12D, K-Ras G12V, latent membrane protein 2 (LMP2), melanoma antigen family A, 1 (MAGE-A1), MAGE-A2, MAGE-A3, MAGE-A4, MAGE-A6, MAGE-A10, MAGE-A12, melanoma antigen recognized by T cells (MART-1), mesothelin (MSLN), mucin 1 (MUC1), mucin 16 (MUC16), New York esophageal squamous cell carcinoma-1 (NYESO-1), P53, P antigen (PAGE) family member, placenta-specific 1 (PLAC1), antigen preferentially expressed in melanoma (PRAME), survivin, synovial sarcoma X1 (SSX1), synovial sarcoma X2 (SSX2), synovial sarcoma X3 (SSX3), synovial sarcoma X4 (SSX4), synovial sarcoma X5 (SSX5), synovial sarcoma X 8 (SSX8), thyroglobulin, tyrosinase, tyrosinase-related protein (TRP) 1, TRP2, Wilms tumor protein (WT-1), X antigen family member 1 (XAGE1), and X antigen family member 2 (XAGE2).

151. The non-natural cell of claim 149 or claim 150, wherein the exogenous TCR is an αβ-TCR or a γδ-TCR.

152. A non-natural cell according to any of the preceding claims, wherein the cell further expresses a CAR, a CCR or a flip receptor.

153. The non-natural cell of any of the preceding claims, wherein the cell further expresses a zetakine, an immune cell engager, or a BiTE.

154. The non-natural cell of any of the preceding claims, wherein the cell is a hematopoietic cell.

155. The non-natural cell of any of the preceding claims, wherein the cell is a T cell, an αβ-T cell, or a γδ-T cell.

156. The non-natural cell of any of the preceding claims, wherein the cell is a CD3 + cells, CD4 + cells and / or CD8 + cell.

157. The non-natural cell of any of the preceding claims, wherein the cell is an immune effector cell.

158. The non-natural cell of any of the preceding claims, wherein the cell is a cytotoxic T lymphocyte (CTL), a tumor infiltrating lymphocyte (TIL), or a helper T cell.

159. The non-natural cell of any of the preceding claims, wherein the cell is a Natural Killer (NK) cell or a Natural Killer T (NKT) cell.

160. The non-natural cell of any of the preceding claims, wherein the source of the cell is peripheral blood mononuclear cells, bone marrow, lymph node tissue, umbilical cord blood, thymus tissue, tissue from a site of infection, ascites, pleural effusion, spleen tissue, or a tumor.

161. The non-naturally occurring cell of any of the preceding claims, wherein the non-naturally occurring cell is an isolated non-naturally occurring cell.

162. The non-natural cell of any of the preceding claims, wherein the non-natural cell is obtained from a subject.

163. The non-natural cell of any of the preceding claims, wherein the non-natural cell is a human cell.

164. A fusion polypeptide comprising: (a) a signaling component comprising a first multimerization domain and an actuator domain; (b) a polypeptide cleavage signal; and (c) a targeting component, which comprises an extracellular domain, a second multimerization domain and a transmembrane domain.

165. The fusion polypeptide of claim 164, wherein the first multimerization domain and the second multimerization domain are located extracellularly when the signaling component and the targeting component are expressed.

166. A fusion polypeptide according to claim 164 or claim 165, wherein the second multimerization domain and the transmembrane domain are separated by a hinge domain.

167. The fusion polypeptide of claim 166, wherein the hinge domain is selected from the group consisting of a CD4 hinge, a CD8 hinge, a CD28 hinge, an IgG4 hinge, and any fragment or variant or combination thereof.

168. The fusion polypeptide of claim 166 or claim 167, wherein the hinge domain is a CD4 hinge.

169. The fusion polypeptide cell of claim 168, wherein the CD4 hinge comprises an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

41.

170. The fusion polypeptide of claim 169, wherein the CD4 hinge comprises the amino acid sequence shown in SEQ ID NO:

41.

171. The fusion polypeptide of claim 166 or claim 167, wherein the hinge domain is a CD28 hinge.

172. The fusion polypeptide of claim 171, wherein the CD28 hinge comprises an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

42.

173. The fusion polypeptide of claim 172, wherein the CD28 hinge comprises the amino acid sequence shown in SEQ ID NO:

42.

174. The fusion polypeptide of claim 166 or claim 167, wherein the hinge domain is a CD8 hinge.

175. The fusion polypeptide of claim 174, wherein the CD8 hinge comprises an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

44.

176. The fusion polypeptide of claim 175, wherein the CD8 hinge comprises the amino acid sequence shown in SEQ ID NO:

44.

177. The fusion polypeptide of claim 166 or claim 167, wherein the hinge domain is an IgG4 hinge.

178. The fusion polypeptide of claim 177, wherein the IgG4 hinge comprises an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

43.

179. The fusion polypeptide of claim 178, wherein the IgG4 hinge comprises the amino acid sequence shown in SEQ ID NO:

43.

180. The fusion polypeptide of any one of claims 164 to 179, wherein the actuator domain is a CD3 polypeptide, an FcεR1γ polypeptide, an Igα / CD79a polypeptide, an Igβ / CD79b polypeptide, a DAP10 polypeptide, or a DAP12 polypeptide.

181. A fusion polypeptide according to any one of claims 164 to 180, wherein the CD3 polypeptide is CD3ε (CD3 epsilon) or a fragment or variant thereof, CD3γ (CD3 gamma) or a fragment or variant thereof, or CD3δ (CD3 delta) or a fragment or variant thereof.

182. The fusion polypeptide of any one of claims 164 to 181, wherein the actuator domain is a CD3ε polypeptide comprising an amino acid sequence at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

32.

183. The fusion polypeptide of any one of claims 164 to 182, wherein the actuator domain is a CD3ε polypeptide comprising the amino acid sequence as shown in SEQ ID NO:

32.

184. The fusion polypeptide of any one of claims 164 to 181, wherein the actuator domain is a CD3γ polypeptide or a variant thereof.

185. The fusion polypeptide of any one of claims 164 to 181 or 184, wherein the actuator domain is a CD3γ polypeptide comprising an amino acid sequence at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

33.

186. The fusion polypeptide of any one of claims 164 to 181, 184 or 185, wherein the actuator domain is a CD3γ polypeptide comprising the amino acid sequence as shown in SEQ ID NO:

33.

187. The fusion polypeptide of any one of claims 164 to 181, wherein the actuator domain is a CD3 delta polypeptide or a variant thereof.

188. The fusion polypeptide of any one of claims 164 to 181 or 187, wherein the actuator domain is a CD3 delta polypeptide comprising an amino acid sequence at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

34.

189. The fusion polypeptide of any one of claims 164 to 181, 187 or 188, wherein the actuator domain is a CD3 delta polypeptide comprising the amino acid sequence as shown in SEQ ID NO:

34.

190. The fusion polypeptide of any one of claims 164 to 181 or 1 89, wherein the actuator domain is an FcεR1γ polypeptide or a variant thereof.

191. The fusion polypeptide of any one of claims 164 to 181 or claim 190, wherein the actuator domain is an FcεR1γ polypeptide comprising an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

35.

192. The fusion polypeptide of any one of claims 164 to 181, 190 or 191, wherein the actuator domain is an FcεR1γ polypeptide comprising the amino acid sequence as shown in SEQ ID NO:

35.

193. The fusion polypeptide of any one of claims 164 to 181 or 190, wherein the actuator domain is an FcεR1γ polypeptide comprising an amino acid sequence at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

36.

194. The fusion polypeptide of any one of claims 164 to 181, 190 or 193, wherein the actuator domain is an FcεR1γ polypeptide comprising the amino acid sequence as shown in SEQ ID NO:

36.

195. The fusion polypeptide of any one of claims 164 to 180, wherein the actuator domain is an Igα / CD79a polypeptide or a variant thereof.

196. The fusion polypeptide of any one of claims 164 to 180 or 195, wherein the actuator domain is an Igα / CD79a polypeptide comprising an amino acid sequence at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

37.

197. The fusion polypeptide of any one of claims 164 to 180, 195 or 196, wherein the actuator domain is an Igα / CD79a polypeptide comprising the amino acid sequence as shown in SEQ ID NO:

37.

198. The fusion polypeptide of any one of claims 164 to 180, wherein the actuator domain is an Igβ / CD79b polypeptide or a variant thereof.

199. The fusion polypeptide of any one of claims 164 to 180 or 198, wherein the actuator domain is an Igβ / CD79b polypeptide comprising an amino acid sequence at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

38.

200. The fusion polypeptide of any one of claims 164 to 180, 198 or 199, wherein the actuator domain is an Igβ / CD79b polypeptide comprising the amino acid sequence as shown in SEQ ID NO:

38.

201. The fusion polypeptide of any one of claims 164 to 180, wherein the actuator domain is a DAP10 polypeptide or a variant thereof.

202. The fusion polypeptide of any one of claims 164 to 180 or 201, wherein the actuator domain is a DAP10 polypeptide comprising an amino acid sequence at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

39.

203. The fusion polypeptide of any one of claims 164 to 180, 201 or 202, wherein the actuator domain is a DAP10 polypeptide comprising the amino acid sequence as shown in SEQ ID NO:

39.

204. The fusion polypeptide of any one of claims 164 to 180, wherein the actuator domain is a DAP12 polypeptide or a variant thereof.

205. The fusion polypeptide of any one of claims 164 to 180 or 204, wherein the actuator domain is a DAP12 polypeptide comprising an amino acid sequence at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

40.

206. The fusion polypeptide of any one of claims 164 to 180, 204 or 205, wherein the actuator domain is a DAP12 polypeptide comprising the amino acid sequence shown in SEQ ID NO:

40.

207. The fusion polypeptide of any one of claims 164 to 206, wherein the actuator domain comprises both an extracellular portion and an intracellular portion.

208. A fusion polypeptide according to any one of claims 164 to 207, further comprising a signal sequence, optionally wherein the signal sequence has at least 90%, 95%, 96%, 97%, 98% or 99% identity with SEQ ID NO:95, SEQ ID NO:96 or SEQ ID NO:97 or comprises SEQ ID NO:95, SEQ ID NO:96 or SEQ ID NO:

97.

209. The fusion polypeptide of any one of claims 164 to 208, wherein the first multimerization domain and the second multimerization domain are different.

210. The fusion polypeptide of any one of claims 164 to 209, wherein the signaling component and the multimerization domain of the targeting component are associated with a bridging factor selected from the group consisting of: rapamycin or a rapamycin analog thereof, gibberellins or derivatives thereof, abscisic acid (ABA) or derivatives thereof, methotrexate or derivatives thereof, cyclosporine A or derivatives thereof, FK506 / cyclosporine A or derivatives thereof, trimethoprim (Tmp) synthetic ligand (SLF) for FK506 binding protein (FKBP) or derivatives thereof, wherein the bridging factor promotes the formation of a polypeptide complex, wherein the bridging factor is associated with the multimerization domains of the signaling component and the targeting component and is positioned between the multimerization domains.

211. A fusion polypeptide according to any one of claims 164 to 210, wherein the first multimerization domain and the second multimerization domain are a pair selected from the group consisting of: FK506 binding protein 1A (FKBP12) and FKBP12-rapamycin binding (FRB), FKBP12 and calcineurin, FKBP and cyclophilin A or any other member of the peptidyl-prolyl cis-trans isomerase (PPIase) family, FKBP and dihydrofolate reductase (DHFR), calcineurin and cyclophilin A or any other member of the peptidyl-prolyl cis-trans isomerase (PPIase) family, and PYR1-like 1 (PYL1) and abscisic acid insensitive 1 (ABI1).

212. The fusion polypeptide of any one of claims 164 to 211, wherein the first multimerization domain comprises a first FRB polypeptide or a variant thereof, and the second multimerization domain comprises a first FKBP12 polypeptide or a variant thereof.

213. The fusion polypeptide of any one of claims 164 to 211, wherein the first multimerization domain comprises a first FKBP12 polypeptide or a variant thereof, and the second multimerization domain comprises a first FRB polypeptide or a variant thereof.

214. The fusion polypeptide of any one of claims 211 to 213, wherein the FRB polypeptide is the FRBT2098L variant.

215. The fusion polypeptide of any one of claims 211 to 214, wherein the FRB polypeptide comprises the amino acid sequence shown in SEQ ID NO: 1 or SEQ ID NO:

2.

216. The fusion polypeptide of any one of claims 211 to 215, wherein the FKBP12 polypeptide comprises the amino acid sequence shown in SEQ ID NO:3 or SEQ ID NO:

4.

217. A fusion polypeptide according to any one of claims 164 to 216, wherein the bridging factor is AP1903, AP20187, AP21967 (also known as C16-(S)-7-methylindorapamycin), everolimus, norvolimus, pimecrolimus, dafolimus, sirolimus, tacrolimus, temsirolimus, urolimus, zotarolimus or BPC015.

218. The fusion polypeptide of any one of claims 164 to 209, wherein the first multimerization domain and the second multimerization domain are a pair of antibody-derived heterologous dimerization domains.

219. A fusion polypeptide according to any one of claims 164 to 209 or 218, wherein the first multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98% or 99% identity with SEQ ID NO:5 or comprises the amino acid sequence as shown in SEQ ID NO:5; and wherein the second multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98% or 99% identity with SEQ ID NO:6 or comprises the amino acid sequence as shown in SEQ ID NO:

6.

220. A fusion polypeptide according to any one of claims 164 to 209 or 218, wherein the first multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98% or 99% identity with SEQ ID NO:6 or comprises the amino acid sequence as shown in SEQ ID NO:6; and wherein the second multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98% or 99% identity with SEQ ID NO:5 or comprises the amino acid sequence as shown in SEQ ID NO:

5.

221. A fusion polypeptide according to any one of claims 164 to 209 or 218, wherein the first multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98% or 99% identity with SEQ ID NO:7 or comprises the amino acid sequence as shown in SEQ ID NO:7; and wherein the second multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98% or 99% identity with SEQ ID NO:8 or comprises the amino acid sequence as shown in SEQ ID NO:

8.

222. A fusion polypeptide according to any one of claims 164 to 209 or 218, wherein the first multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98% or 99% identity with SEQ ID NO:8 or comprises the amino acid sequence as shown in SEQ ID NO:8; and wherein the second multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98% or 99% identity with SEQ ID NO:7 or comprises the amino acid sequence as shown in SEQ ID NO:

7.

223. A fusion polypeptide according to any one of claims 164 to 209 or 218, wherein the first multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98% or 99% identity with SEQ ID NO:9 or comprises the amino acid sequence as shown in SEQ ID NO:9; and wherein the second multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98% or 99% identity with SEQ ID NO:10 or comprises the amino acid sequence as shown in SEQ ID NO:

10.

224. A fusion polypeptide according to any one of claims 164 to 209 or 218, wherein the first multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98% or 99% identity with SEQ ID NO:10 or comprises the amino acid sequence as shown in SEQ ID NO:10; and wherein the second multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98% or 99% identity with SEQ ID NO:9 or comprises the amino acid sequence as shown in SEQ ID NO:

9.

225. The fusion polypeptide of any one of claims 164 to 224, wherein the first multimerization domain and the actuator domain are separated by a first polypeptide linker having a length of 2 to 40 amino acids.

226. The fusion polypeptide of claim 225, wherein the first polypeptide linker is selected from the group consisting of GG, GS, SG, SS, GSS, SSG, GSG, SGS, SGG, G4S, 2xG4S, 3xG4S, 4xG4S, 5xG4S, and any combination thereof.

227. The fusion polypeptide of claim 226, wherein the first polypeptide linker is a 3xG4S linker.

228. The fusion polypeptide of claim 226, wherein the first polypeptide linker comprises a polypeptide comprising an amino acid sequence as shown in any one of SEQ ID NOs: 16-31.

229. A fusion polypeptide according to any one of claims 164 to 228, wherein the extracellular domain and the second multimerization domain are separated by a second polypeptide linker having a length of 2 to 40 amino acids.

230. The fusion polypeptide of claim 229, wherein the second polypeptide linker is selected from the group consisting of GG, GS, SG, SS, GSS, SSG, GSG, SGS, SGG, G4S, 2x G4S, 3xG4s, 4xG4S, and any combination thereof.

231. The fusion polypeptide of claim 230, wherein the second polypeptide linker is a G4S linker.

232. The fusion polypeptide of claim 230, wherein the second polypeptide linker comprises a polypeptide comprising an amino acid sequence as shown in any one of SEQ ID NOs: 16-31.

233. The fusion polypeptide of any one of claims 164 to 232, wherein the transmembrane domain is a CD4 transmembrane domain.

234. The fusion polypeptide of claim 233, wherein the CD4 transmembrane domain comprises an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

45.

235. The fusion polypeptide of claim 234, wherein the CD4 transmembrane domain comprises the amino acid sequence shown in SEQ ID NO:

45.

236. The fusion polypeptide of any one of claims 164 to 232, wherein the transmembrane domain is a CD28 transmembrane domain.

237. The fusion polypeptide of claim 236, wherein the CD28 transmembrane domain comprises an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

46.

238. The fusion polypeptide of claim 237, wherein the CD28 transmembrane domain comprises the amino acid sequence shown in SEQ ID NO:

46.

239. The fusion polypeptide of any one of claims 164 to 232, wherein the transmembrane domain is a CD8 transmembrane domain.

240. The fusion polypeptide of claim 239, wherein the CD8 transmembrane domain comprises an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

47.

241. The fusion polypeptide of claim 240, wherein the CD8 transmembrane domain comprises the amino acid sequence shown in SEQ ID NO:

47.

242. The fusion polypeptide of any one of claims 164 to 241, wherein the targeting component further comprises an intracellular signaling domain or a co-stimulatory domain derived from a protein selected from the group consisting of an antigen receptor, a co-stimulatory receptor, a growth receptor, a cytokine receptor, an adaptor signaling protein, an intracellular signaling protein, or any fragment or variant thereof.

243. The fusion polypeptide of claim 242, wherein the intracellular signaling domain or the co-stimulatory domain on the targeting component is selected from the group consisting of: Toll-like receptor 1 (TLR1), TLR2, TLR3, TLR4, TLR5, TLR6, TLR7, TLR8, TLR9, TLR10, caspase recruitment domain family member 11 (CARD11), CD2, CD3ε, CD3γ, CD3δ, CD4, CD7, CD8, CD27, CD28, CD30, CD40, CD54 (ICAM), CD83, CD94, CD134 (OX40), CD1 37 (4-1BB), CD278 (ICOS), common gamma chain cytokine, DNAX-activating protein 10 (DAP10), adaptor for activating T cells family member 1 (LAT), interleukin 2 receptor (IL-2R), IL-4R, IL-7R, IL-9R, IL-12R, IL-13R, IL-15R, IL-21R, SH2 domain-containing leukocyte protein of 76 kD (SLP76), T cell receptor-associated transmembrane adaptor 1 (TRAT1), TNFR2, TNFRS14, TNFRS18, TNRFS25, and T cell receptor zeta chain-associated protein kinase 70 (ZAP70).

244. The fusion polypeptide of claim 242 or claim 243, wherein the intracellular signaling domain or the co-stimulatory domain on the targeting component is selected from the group consisting of: 4-1BB, CD28, TNFR2, OX40, ICOS, and DAP10 co-stimulatory domains.

245. The fusion polypeptide of claim 242, wherein the co-stimulatory domain on the targeting component is a 4-1BB co-stimulatory domain, optionally wherein the 4-1BB co-stimulatory domain comprises the amino acid sequence shown in SEQ ID NO:

98.

246. The fusion polypeptide of claim 242, wherein the co-stimulatory domain on the targeting component is a CD28 co-stimulatory domain, optionally wherein the CD28 co-stimulatory domain comprises the amino acid sequence shown in SEQ ID NO:

99.

247. The fusion polypeptide of claim 242, wherein the intracellular signaling domain or the co-stimulatory domain is one or more cytokine receptor intracellular signaling domains.

248. The fusion polypeptide of claim 247, wherein the one or more cytokine receptor intracellular signaling domains are selected from the group consisting of an IL7Rα intracellular signaling domain, an IL2Rβ intracellular signaling domain, a common γ chain intracellular signaling domain, and both IL2Rβ and common γ chain intracellular signaling domains.

249. The fusion polypeptide of claim 248, wherein the IL7Rα intracellular signaling domain comprises an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

103.

250. The fusion polypeptide of claim 249, wherein the IL7Rα intracellular signaling domain comprises the amino acid sequence shown in SEQ ID NO:

103.

251. The fusion polypeptide of claim 248, wherein the IL2Rβ intracellular signaling domain comprises an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO: 104 or SEQ ID NO:

105.

252. The fusion polypeptide of claim 251, wherein the IL2Rβ intracellular signaling domain comprises the amino acid sequence shown in SEQ ID NO: 104 or SEQ ID NO:

105.

253. The fusion polypeptide of claim 248, wherein the common gamma chain intracellular signaling domain comprises an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

106.

254. The fusion polypeptide of claim 253, wherein the common γ chain intracellular signaling domain comprises the amino acid sequence shown in SEQ ID NO:

106.

255. The fusion polypeptide of claim 242, wherein the intracellular signaling domain or the co-stimulatory domain is a LAT domain.

256. The fusion polypeptide of claim 255, wherein the LAT domain comprises an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to the amino acid sequence shown in SEQ ID NO:

102.

257. The fusion polypeptide of claim 256, wherein the LAT domain comprises the amino acid sequence shown in SEQ ID NO:

102.

258. The fusion polypeptide of claim 242, wherein the intracellular signaling domain or the co-stimulatory domain is a CD4 coreceptor domain.

259. The fusion polypeptide of claim 258, wherein the CD4 co-receptor domain comprises an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

100.

260. The fusion polypeptide of claim 259, wherein the CD4 co-receptor domain comprises the amino acid sequence shown in SEQ ID NO:

100.

261. The fusion polypeptide of claim 242, wherein the intracellular signaling domain or the co-stimulatory domain is a CD8 coreceptor domain.

262. The fusion polypeptide of claim 261, wherein the CD8 co-receptor domain comprises an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

101.

263. The fusion polypeptide of claim 262, wherein the CD8 co-receptor domain comprises the amino acid sequence shown in SEQ ID NO:

101.

264. The fusion polypeptide of any one of claims 164 to 241, wherein the targeting component does not comprise a functional intracellular signaling domain or a co-stimulatory domain.

265. The fusion polypeptide of any one of claims 164 to 264, wherein the targeting component further comprises a truncated intracellular CD4 polypeptide.

266. The fusion polypeptide of claim 265, wherein the truncated intracellular CD4 polypeptide comprises the amino acid sequence shown in SEQ ID NO:48 or SEQ ID NO:

49.

267. The fusion polypeptide of claim 266, wherein the truncated intracellular CD4 polypeptide comprises the amino acid sequence shown in SEQ ID NO:48 or SEQ ID NO:

49.

268. The fusion polypeptide of any one of claims 164 to 267, wherein the extracellular domain comprises a first targeting domain.

269. The fusion polypeptide of any one of claims 164 to 268, wherein the first targeting domain comprises a single-chain variable fragment (scFv) or a single domain antibody (sdAb).

270. The fusion polypeptide of claim 269, wherein the sdAb is a camelid VHH, a nanobody, or a heavy chain only antibody (HcAb).

271. The fusion polypeptide of claim 270, wherein the sdAb is a camelid VHH.

272. The fusion polypeptide of any one of claims 270 or 271, wherein the scFv or the sdAb is human or humanized.

273. The fusion polypeptide of any one of claims 268 to 272, wherein the first targeting domain comprises a sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to any one of SEQ ID NOs: 50-83.

274. The fusion polypeptide of claim 273, wherein the first targeting domain comprises a sequence as shown in any one of SEQ ID NOs: 50-83.

275. The fusion polypeptide of claim 274, wherein the first targeting domain comprises the extracellular domain of PD1, human A proliferation-inducing ligand (APRIL), trimerized human APRIL, or NKG2D membrane protein.

276. The fusion polypeptide of claim 275, wherein the PD1 extracellular domain comprises a sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:84 or SEQ ID NO:

85.

277. The fusion polypeptide of claim 276, wherein the PD1 extracellular domain comprises the sequence shown in SEQ ID NO:

85.

278. A fusion polypeptide according to claim 275, wherein the APRIL comprises a sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:86, or wherein the trimerized human APRIL comprises a sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

87.

279. The fusion polypeptide of claim 276, wherein the APRIL comprises the sequence shown in SEQ ID NO:86, or wherein the trimerized human APRIL comprises the sequence shown in SEQ ID NO:

87.

280. The fusion polypeptide of claim 275, wherein the NKG2D membrane protein comprises a sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

88.

281. The fusion polypeptide of claim 280, wherein the NKG2D membrane protein comprises the sequence shown in SEQ ID NO:

88.

282. The fusion polypeptide of any one of claims 268 to 281, wherein the extracellular domain further comprises a second targeting domain.

283. The fusion polypeptide of claim 282, wherein the second targeting domain comprises a second single-chain variable fragment (scFv) or a second single domain antibody (sdAb).

284. The fusion polypeptide of claim 283, wherein the second sdAb is a camelid VHH, a nanobody, or a heavy chain only antibody (HcAb).

285. The fusion polypeptide of claim 284, wherein the second sdAb is a camelid VHH.

286. The fusion polypeptide of any one of claims 282 to 285, wherein the second scFv or the second sdAb is human or humanized.

287. The fusion polypeptide of any one of claims 282 to 286, wherein the second targeting domain comprises a sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to any one of SEQ ID NOs: 50-83.

288. The fusion polypeptide of claim 287, wherein the second targeting domain comprises a sequence as shown in any one of SEQ ID NOs: 50-83.

289. The fusion polypeptide of claim 282, wherein the second targeting domain comprises the extracellular domain of PD1, human A proliferation-inducing ligand (APRIL), trimerized human APRIL, or NKG2D membrane protein.

290. The fusion polypeptide of claim 289, wherein the PD1 extracellular domain comprises a sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:84 or SEQ ID NO:

85.

291. The fusion polypeptide of claim 290, wherein the PD1 extracellular domain comprises the sequence shown in SEQ ID NO:

85.

292. A fusion polypeptide according to claim 289, wherein the APRIL comprises a sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:86, or wherein the trimerized human APRIL comprises a sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

87.

293. The fusion polypeptide of claim 292, wherein the APRIL comprises the sequence as shown in SEQ ID NO:86, or wherein the trimerized human APRIL comprises the sequence as shown in SEQ ID NO:

87.

294. The fusion polypeptide of claim 289, wherein the NKG2D membrane protein comprises a sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

88.

295. The fusion polypeptide of claim 294, wherein the NKG2D membrane protein comprises the sequence shown in SEQ ID NO:

88.

296. The fusion polypeptide of any one of claims 268 to 295, wherein the first targeting domain and the second targeting domain bind to the same antigen or different antigens.

297. The fusion polypeptide of any one of claims 268 to 296, wherein the first targeting domain and the second targeting domain are separated by a third polypeptide linker having a length of 2 to 40 amino acids.

298. A fusion polypeptide according to claim 297, wherein the third polypeptide linker is selected from the group consisting of: GG, GS, SG, SS, GSS, SSG, GSG, SGS, SGG, G4S, 2x G4S, 3xG4s, 4xG4S, 5xG4S, any one of SEQ ID NOs: 16-31, and any combination thereof.

299. The fusion polypeptide of any one of claims 164 to 298, wherein the polypeptide cleavage signal is a viral self-cleavage polypeptide.

300. The fusion polypeptide of any one of claims 164 to 299, wherein the polypeptide cleavage signal is a viral self-cleavage 2A polypeptide.

301. A fusion polypeptide according to any one of claims 164 to 299, wherein the polypeptide cleavage signal is a viral self-cleaving polypeptide, and the viral self-cleaving polypeptide is selected from the group consisting of: foot-and-mouth disease virus (FMDV) (F2A) peptide, equine rhinitis virus type A (ERAV) (E2A) peptide, Thosea asigna virus (TaV) (T2A) peptide, porcine teschovirus-1 (PTV-1) (P2A) peptide, Theilovirus 2A peptide and encephalomyocarditis virus 2A peptide.

302. A fusion polypeptide according to any one of claims 164 to 301, wherein the signaling component comprises a sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:107, SEQ ID NO:108, SEQ ID NO:109, SEQ ID NO:110, SEQ ID NO:111, SEQ ID NO:112, SEQ ID NO:113, SEQ NO:114 or SEQ ID NO:

115.

303. The fusion polypeptide of claim 302, wherein the signaling component comprises a sequence as shown in SEQ ID NO:107, SEQ ID NO:108, SEQ ID NO:109, SEQ ID NO:110, SEQ ID NO:111, SEQ ID NO:112, SEQ ID NO:113, SEQ NO:114, or SEQ ID NO:

115.

304. A fusion polypeptide according to any one of claims 164 to 303, wherein the targeting component comprises a sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to the sequence shown in SEQ ID NO:116, SEQ ID NO:117, SEQ ID NO:118, SEQ ID NO:119, SEQ ID NO:120, SEQ ID NO:121, SEQ ID NO:122, SEQ ID NO:123, SEQ ID NO:124, SEQ ID NO:125, SEQ ID NO:126, SEQ ID NO:127, SEQ ID NO:128, SEQ ID NO:129, SEQ ID NO:130, SEQ ID NO:131, SEQ ID NO:132, SEQ ID NO:133, SEQ ID NO:134, SEQ ID NO:135 or SEQ ID NO:

136.

305. The fusion polypeptide of claim 304, wherein the targeting component comprises a sequence as shown in SEQ ID NO: 116, SEQ ID NO: 117, SEQ ID NO: 118, SEQ ID NO: 119, SEQ ID NO: 120, SEQ ID NO: 121, SEQ ID NO: 122, SEQ ID NO: 123, SEQ ID NO: 124, SEQ ID NO: 125, SEQ ID NO: 126, SEQ ID NO: 127, SEQ ID NO: 128, SEQ ID NO: 129, SEQ ID NO: 130, SEQ ID NO: 131, SEQ ID NO: 132, SEQ ID NO: 133, SEQ ID NO: 134, SEQ ID NO: 135, or SEQ ID NO:

136.

306. The fusion polypeptide of any one of claims 164 to 305, comprising a fusion polypeptide comprising the targeting component and the signaling component.

307. The fusion polypeptide of claim 306, wherein the fusion polypeptide comprises the residues of SEQ ID NO: 139, SEQ ID NO: 140, SEQ ID NO: 141, SEQ ID NO: 142, SEQ ID NO: 143, SEQ ID NO: 144, SEQ ID NO: 145, SEQ ID NO: 146, SEQ ID NO: 147, SEQ ID NO: 148, SEQ ID NO: 149, SEQ ID NO: 150, SEQ ID NO: 151, SEQ ID NO: 152, SEQ ID NO: 153, SEQ ID NO: 154, SEQ ID NO: 155, SEQ ID NO: 156, SEQ ID NO: 157, SEQ ID NO: 158, SEQ ID NO: 159, SEQ ID NO: 160, SEQ ID NO: 161, SEQ ID NO: 162, SEQ ID NO: 163, SEQ ID NO: 164, SEQ ID NO: 165, SEQ ID NO: 166, SEQ ID NO: 167, NO: 167, SEQ ID NO: 168, SEQ ID NO: 169 or SEQ ID NO: 170 having a sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical.

308. The fusion polypeptide of claim 307, wherein the fusion polypeptide comprises SEQ ID NO: 139, SEQ ID NO: 140, SEQ ID NO: 141, SEQ ID NO: 142, SEQ ID NO: 143, SEQ ID NO: 144, SEQ ID NO: 145, SEQ ID NO: 146, SEQ ID NO: 147, SEQ ID NO: 148, SEQ ID NO: 149, SEQ ID NO: 150, SEQ ID NO: 151, SEQ ID NO: 152, SEQ ID NO: 153, SEQ ID NO: 154, SEQ ID NO: 155, SEQ ID NO: 156, SEQ ID NO: 157, SEQ ID NO: 158, SEQ ID NO: 159, SEQ ID NO: 160, SEQ ID NO: 161, SEQ ID NO: 162, SEQ ID NO: 163, SEQ ID NO: 164, SEQ ID NO: 165, SEQ ID NO: 166, SEQ ID NO: 167, The sequence shown in SEQ ID NO:167, SEQ ID NO:168, SEQ ID NO:169 or SEQ ID NO:

170.

309. A nucleic acid molecule encoding a fusion polypeptide according to any one of claims 164 to 308.

310. A cell comprising the fusion polypeptide of any one of claims 164 to 308.

311. A cell comprising the nucleic acid molecule of claim 309.

312. The cell of claim 310 or claim 311, further expressing an exogenous co-stimulatory factor, an immunomodulatory factor, an agonist of a co-stimulatory factor, an antagonist of an immunosuppressive factor, an immune cell engager, a flip receptor, or any combination thereof.

313. The cell of any one of claims 310 to 311, wherein the cell further expresses an exogenous lymphocyte receptor or co-receptor.

314. The cell of claim 313, wherein the exogenous lymphocyte receptor or co-receptor is selected from the group consisting of TCRα (TCR alpha), TCRβ (TCR beta), TCRγ (TCR gamma), TCRδ (TCR delta), CD4, CD8, pre-T cell receptor alpha (pTα), Fc receptor alpha (FcRα), Fc receptor beta (FcRβ), Fc receptor gamma (FcRγ), natural killer group 2 member D (NKG2D), CD79A, CD79B, and any combination thereof.

315. The cell of any one of claims 310 to 314, wherein the cell further expresses an exogenous TCR.

316. The cell of claim 315, wherein the exogenous TCR binds to a target antigen selected from the group consisting of: alpha-fetoprotein (AFP), members of the B melanoma antigen (BAGE) family, brothers of regulatory factors at imprinted sites (BORIS), cancer-testis antigen, cancer-testis antigen 83 (CT-83), carbonic anhydrase IX (CA1X), carcinoembryonic antigen (CEA), cytomegalovirus (CMV) antigen, antigen recognized by cytotoxic T cells (CTL) on melanoma (CAMEL), Epstein-Barr virus (EBV ) antigen, G antigen 1 (GAGE-1), GAGE-2, GAGE-3, GAGE-4, GAGE-5, GAGE-6, GAGE-7B, GAGE-8, glycoprotein 100 (GP100), hepatitis B virus (HBV) antigen, hepatitis C virus (HCV) nonstructural protein 3 (NS3), human epidermal growth factor receptor 2 (HER-2), human papillomavirus (HPV)-E6, HPV-E7, human telomerase reverse transcriptase (hTERT), IGF2BP3 / A3, K-Ras, K-Ras G12C, K-Ras G12D, K-Ras G12V, latent membrane protein 2 (LMP2), melanoma antigen family A, 1 (MAGE-A1), MAGE-A2, MAGE-A3, MAGE-A4, MAGE-A6, MAGE-A10, MAGE-A12, melanoma antigen recognized by T cells (MART-1), mesothelin (MSLN), mucin 1 (MUC1), mucin 16 (MUC16), New York esophageal squamous cell carcinoma-1 (NYESO-1), P53, P antigen (PAGE) family member, placenta-specific 1 (PLAC1), antigen preferentially expressed in melanoma (PRAME), survivin, synovial sarcoma X1 (SSX1), synovial sarcoma X2 (SSX2), synovial sarcoma X3 (SSX3), synovial sarcoma X4 (SSX4), synovial sarcoma X5 (SSX5), synovial sarcoma X 8 (SSX8), thyroglobulin, tyrosinase, tyrosinase-related protein (TRP) 1, TRP2, Wilms' tumor protein (WT-1), X antigen family member 1 (XAGE1), and X antigen family member 2 (XAGE2).

317. The cell of claim 315 or claim 316, wherein the exogenous TCR is an αβ-TCR or a γδ-TCR.

318. A cell according to any one of claims 310 to 317, wherein the cell further expresses CAR, CCR or flip receptor.

319. The cell of any one of claims 310 to 318, wherein the cell further expresses a zetakine, an immune cell engager, or a BiTE.

320. The cell of any one of claims 310 to 319, wherein the cell is a hematopoietic cell.

321. The cell of any one of claims 310 to 320, wherein the cell is a T cell, an αβ-T cell, or a γδ-T cell.

322. The cell according to any one of claims 310 to 321, wherein the cell is a CD3 + cells, CD4 + cells and / or CD8 + cell.

323. The cell of any one of claims 310 to 322, wherein the cell is an immune effector cell.

324. The cell of any one of claims 310 to 323, wherein the cell is a cytotoxic T lymphocyte (CTL), a tumor infiltrating lymphocyte (TIL), or a helper T cell.

325. The cell of any one of claims 164 to 324, wherein the cell is a Natural Killer (NK) cell or a Natural Killer T (NKT) cell.

326. The cell of any one of claims 310 to 325, wherein the source of the cell is peripheral blood mononuclear cells, bone marrow, lymph node tissue, umbilical cord blood, thymus tissue, tissue from an infection site, ascites, pleural effusion, spleen tissue, or a tumor.

327. The cell of any one of claims 310 to 326, wherein the cell is an isolated cell.

328. The cell of any one of claims 310 to 327, wherein the cell is obtained from a subject.

329. The cell of any one of claims 310 to 328, wherein the cell is a human cell.

330. A polypeptide complex comprising: a signaling component comprising a first multimerization domain and an actuator domain; and The targeting component comprises an extracellular domain, a second multimerization domain and a transmembrane domain.

331. The polypeptide complex of claim 330, wherein the first multimerization domain and the second multimerization domain are located extracellularly when the signaling component and the targeting component are expressed.

332. A polypeptide complex according to claim 330 or claim 331, wherein the second multimerization domain and the transmembrane domain are separated by a hinge domain.

333. The polypeptide complex of claim 332, wherein the hinge domain is selected from the group consisting of a CD4 hinge, a CD8 hinge, a CD28 hinge, an IgG4 hinge, and any fragment or variant or combination thereof.

334. A polypeptide complex according to claim 332 or claim 333, wherein the hinge domain is a CD4 hinge.

335. The polypeptide complex cell of claim 334, wherein the CD4 hinge comprises an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

41.

336. The polypeptide complex of claim 335, wherein the CD4 hinge comprises the amino acid sequence shown in SEQ ID NO:

41.

337. A polypeptide complex according to claim 332 or claim 333, wherein the hinge domain is a CD28 hinge.

338. The polypeptide complex of claim 337, wherein the CD28 hinge comprises an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

42.

339. The polypeptide complex of claim 338, wherein the CD28 hinge comprises the amino acid sequence shown in SEQ ID NO:

42.

340. The polypeptide complex of claim 332 or claim 333, wherein the hinge domain is a CD8 hinge.

341. The polypeptide complex of claim 340, wherein the CD8 hinge comprises an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

44.

342. The polypeptide complex of claim 341, wherein the CD8 hinge comprises the amino acid sequence shown in SEQ ID NO:

44.

343. A polypeptide complex according to claim 332 or claim 333, wherein the hinge domain is an IgG4 hinge.

344. The polypeptide complex of claim 343, wherein the IgG4 hinge comprises an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

43.

345. The polypeptide complex of claim 355, wherein the IgG4 hinge comprises the amino acid sequence shown in SEQ ID NO:

43.

346. The polypeptide complex of any one of claims 330 to 345, wherein the actuator domain is a CD3 polypeptide, an FcεR1γ polypeptide, an Igα / CD79a polypeptide, an Igβ / CD79b polypeptide, a DAP10 polypeptide or a DAP12 polypeptide.

347. The polypeptide complex according to any one of claims 330 to 346, wherein the CD3 polypeptide is CD3ε (CD3 epsilon) or a fragment or variant thereof, CD3γ (CD3 gamma) or a fragment or variant thereof, or CD3δ (CD3 delta) or a fragment or variant thereof.

348. The polypeptide complex of any one of claims 330 to 347, wherein the actuator domain is a CD3ε polypeptide comprising an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:-32.

349. The polypeptide complex of any one of claims 330 to 348, wherein the actuator domain is a CD3ε polypeptide comprising the amino acid sequence as shown in SEQ ID NO:

32.

350. The polypeptide complex of any one of claims 330 to 347, wherein the actuator domain is a CD3γ polypeptide or a variant thereof.

351. The polypeptide complex of any one of claims 330 to 347 or 350, wherein the actuator domain is a CD3γ polypeptide comprising an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

33.

352. The polypeptide complex of any one of claims 330 to 347, 350 or 351, wherein the actuator domain is a CD3γ polypeptide comprising the amino acid sequence as shown in SEQ ID NO:

33.

353. The polypeptide complex of any one of claims 330 to 347, wherein the actuator domain is a CD3δ polypeptide or a variant thereof.

354. The polypeptide complex of any one of claims 330 to 347 or 353, wherein the actuator domain is a CD3δ polypeptide comprising an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

34.

355. The polypeptide complex of any one of claims 330 to 347, 353 or 354, wherein the actuator domain is a CD3 delta polypeptide comprising the amino acid sequence as shown in SEQ ID NO:

34.

356. The polypeptide complex of any one of claims 330 to 346, wherein the actuator domain is an FcεR1γ polypeptide or a variant thereof.

357. The polypeptide complex of any one of claims 330 to 346 or claim 356, wherein the actuator domain is an FcεR1γ polypeptide comprising an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

35.

358. The polypeptide complex of any one of claims 330 to 346, 356 or 357, wherein the actuator domain is an FcεR1γ polypeptide comprising the amino acid sequence as shown in SEQ ID NO:

35.

359. The polypeptide complex of any one of claims 330 to 346 or 356, wherein the actuator domain is an FcεR1γ polypeptide comprising an amino acid sequence at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

36.

360. The polypeptide complex of any one of claims 330 to 346, 356 or 359, wherein the actuator domain is an FcεR1γ polypeptide comprising the amino acid sequence as shown in SEQ ID NO:

36.

361. The polypeptide complex of any one of claims 330 to 346, wherein the actuator domain is an Igα / CD79a polypeptide or a variant thereof.

362. The polypeptide complex of any one of claims 330 to 346 or 361, wherein the actuator domain is an Igα / CD79a polypeptide comprising an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

37.

363. The polypeptide complex of any one of claims 330 to 346, 361 or 362, wherein the actuator domain is an Igα / CD79a polypeptide comprising the amino acid sequence shown in SEQ ID NO:

37.

364. The polypeptide complex of any one of claims 330 to 346, wherein the actuator domain is an Igβ / CD79b polypeptide or a variant thereof.

365. The polypeptide complex of any one of claims 330 to 346 or 364, wherein the actuator domain is an Igβ / CD79b polypeptide comprising an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

38.

366. The polypeptide complex of any one of claims 330 to 346, 364 or 365, wherein the actuator domain is an Igβ / CD79b polypeptide comprising the amino acid sequence shown in SEQ ID NO:

38.

367. The polypeptide complex of any one of claims 330 to 346, wherein the actuator domain is a DAP10 polypeptide or a variant thereof.

368. The polypeptide complex of any one of claims 330 to 346 or 367, wherein the actuator domain is a DAP10 polypeptide comprising an amino acid sequence at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

39.

369. The polypeptide complex of any one of claims 330 to 346, 367 or 368, wherein the actuator domain is a DAP10 polypeptide comprising the amino acid sequence shown in SEQ ID NO:

39.

370. The polypeptide complex of any one of claims 330 to 346, wherein the actuator domain is a DAP12 polypeptide or a variant thereof.

371. The polypeptide complex of any one of claims 330 to 346 or 370, wherein the actuator domain is a DAP12 polypeptide comprising an amino acid sequence at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

40.

372. The polypeptide complex of any one of claims 330 to 346, 370 or 371, wherein the actuator domain is a DAP12 polypeptide comprising the amino acid sequence shown in SEQ ID NO:

40.

373. The polypeptide complex of any one of claims 330 to 372, wherein the actuator domain comprises both an extracellular portion and an intracellular portion.

374. The polypeptide complex of any one of claims 330 to 373, further comprising a signal sequence, optionally wherein the signal sequence is at least 90%, 95%, 96%, 97%, 98% or 99% identical to or comprises SEQ ID NO:95, SEQ ID NO:96 or SEQ ID NO:

97.

375. The polypeptide complex of any one of claims 330 to 374, wherein the first multimerization domain and the second multimerization domain are different.

376. The polypeptide complex of any one of claims 330 to 375, wherein the multimerization domains of the signaling component and the targeting component are associated with a bridging factor selected from the group consisting of: rapamycin or a rapamycin analog thereof, a gibberellin or a derivative thereof, abscisic acid (ABA) or a derivative thereof, methotrexate or a derivative thereof, cyclosporine A or a derivative thereof, FK506 / cyclosporine A or a derivative thereof, trimethoprim (Tmp) synthetic ligand (SLF) for FK506 binding protein (FKBP) or a derivative thereof, wherein the bridging factor promotes the formation of the polypeptide complex, wherein the bridging factor is associated with the multimerization domains of the signaling component and the targeting component and is positioned between the multimerization domains.

377. The polypeptide complex of any one of claims 330 to 376, wherein the first multimerization domain and the second multimerization domain are a pair selected from the group consisting of: FK506 binding protein 1A (FKBP12) and FKBP12-rapamycin binding (FRB), FKBP12 and calcineurin, FKBP and cyclophilin A or any other member of the peptidyl-prolyl cis-trans isomerase (PPIase) family, FKBP and dihydrofolate reductase (DHFR), calcineurin and cyclophilin A or any other member of the peptidyl-prolyl cis-trans isomerase (PPIase) family, and PYR1-like 1 (PYL1) and abscisic acid insensitive 1 (ABI1).

378. The polypeptide complex of any one of claims 330 to 377, wherein the first multimerization domain comprises a first FRB polypeptide or a variant thereof, and the second multimerization domain comprises a first FKBP12 polypeptide or a variant thereof.

379. The polypeptide complex of any one of claims 330 to 377, wherein the first multimerization domain comprises a first FKBP12 polypeptide or a variant thereof, and the second multimerization domain comprises a first FRB polypeptide or a variant thereof.

380. The polypeptide complex of any one of claims 377 to 379, wherein the FRB polypeptide is a FRBT2098L variant.

381. The polypeptide complex of any one of claims 377 to 380, wherein the FRB polypeptide comprises the amino acid sequence shown in SEQ ID NO: 1 or SEQ ID NO:

2.

382. The polypeptide complex of any one of claims 377 to 381, wherein the FKBP12 polypeptide comprises the amino acid sequence shown in SEQ ID NO:3 or SEQ ID NO:

4.

383. The polypeptide complex of any one of claims 376 to 382, ​​wherein the bridging factor is AP1903, AP20187, AP21967 (also known as C16-(S)-7-methylindorapamycin), everolimus, norvolimus, pimecrolimus, dafoslimus, sirolimus, tacrolimus, temsirolimus, urolimus, zotarolimus or BPC015.

384. The polypeptide complex of any one of claims 330 to 375, wherein the first multimerization domain and the second multimerization domain are a pair of antibody-derived heterologous dimerization domains.

385. A polypeptide complex according to any one of claims 330 to 375 or 384, wherein the first multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98% or 99% identity with SEQ ID NO:5 or comprises the amino acid sequence as shown in SEQ ID NO:5; and wherein the second multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98% or 99% identity with SEQ ID NO:6 or comprises the amino acid sequence as shown in SEQ ID NO:

6.

386. A polypeptide complex according to any one of claims 330 to 375 or 384, wherein the first multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98% or 99% identity with SEQ ID NO:6 or comprises the amino acid sequence as shown in SEQ ID NO:6; and wherein the second multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98% or 99% identity with SEQ ID NO:5 or comprises the amino acid sequence as shown in SEQ ID NO:

5.

387. A polypeptide complex according to any one of claims 330 to 375 or 384, wherein the first multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98% or 99% identity with SEQ ID NO:7 or comprises the amino acid sequence as shown in SEQ ID NO:7; and wherein the second multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98% or 99% identity with SEQ ID NO:8 or comprises the amino acid sequence as shown in SEQ ID NO:

8.

388. A polypeptide complex according to any one of claims 330 to 375 or 384, wherein the first multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98% or 99% identity with SEQ ID NO:8 or comprises the amino acid sequence as shown in SEQ ID NO:8; and wherein the second multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98% or 99% identity with SEQ ID NO:7 or comprises the amino acid sequence as shown in SEQ ID NO:

7.

389. A polypeptide complex according to any one of claims 330 to 375 or 384, wherein the first multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98% or 99% identity with SEQ ID NO:9 or comprises the amino acid sequence as shown in SEQ ID NO:9; and wherein the second multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98% or 99% identity with SEQ ID NO:10 or comprises the amino acid sequence as shown in SEQ ID NO:

10.

390. A polypeptide complex according to any one of claims 330 to 375 or 384, wherein the first multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98% or 99% identity with SEQ ID NO:10 or comprises the amino acid sequence as shown in SEQ ID NO:10; and wherein the second multimerization domain comprises a polypeptide having at least 90%, 95%, 96%, 97%, 98% or 99% identity with SEQ ID NO:9 or comprises the amino acid sequence as shown in SEQ ID NO:

9.

391. The polypeptide complex of any one of claims 330 to 390, wherein the first multimerization domain and the actuator domain are separated by a first polypeptide linker having a length of 2 to 40 amino acids.

392. The polypeptide complex of claim 391, wherein the first polypeptide linker is selected from the group consisting of GG, GS, SG, SS, GSS, SSG, GSG, SGS, SGG, G4S, 2xG4S, 3xG4S, 4xG4S, 5xG4S, and any combination thereof.

393. The polypeptide complex of claim 392, wherein the first polypeptide linker is a 3xG4S linker.

394. The polypeptide complex of claim 391, wherein the first polypeptide linker comprises a polypeptide comprising an amino acid sequence as shown in any one of SEQ ID NOs: 16-31.

395. The polypeptide complex of any one of claims 330 to 394, wherein the extracellular domain and the second multimerization domain are separated by a second polypeptide linker having a length of 2 to 40 amino acids.

396. The polypeptide complex of claim 395, wherein the second polypeptide linker is selected from the group consisting of GG, GS, SG, SS, GSS, SSG, GSG, SGS, SGG, G4S, 2x G4S, 3xG4s, 4xG4S, and any combination thereof.

397. The polypeptide complex of claim 396, wherein the second polypeptide linker is a G4S linker.

398. The polypeptide complex of claim 395, wherein the second polypeptide linker comprises a polypeptide comprising an amino acid sequence as shown in any one of SEQ ID NOs: 16-31.

399. The polypeptide complex of any one of claims 330 to 398, wherein the transmembrane domain is a CD4 transmembrane domain.

400. The polypeptide complex of claim 399, wherein the CD4 transmembrane domain comprises an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

45.

401. The polypeptide complex of claim 400, wherein the CD4 transmembrane domain comprises the amino acid sequence shown in SEQ ID NO:

45.

402. The polypeptide complex of any one of claims 330 to 398, wherein the transmembrane domain is a CD28 transmembrane domain.

403. The polypeptide complex of claim 402, wherein the CD28 transmembrane domain comprises an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

46.

404. The polypeptide complex of claim 403, wherein the CD28 transmembrane domain comprises the amino acid sequence shown in SEQ ID NO:

46.

405. The polypeptide complex of any one of claims 330 to 398, wherein the transmembrane domain is a CD8 transmembrane domain.

406. The polypeptide complex of claim 405, wherein the CD8 transmembrane domain comprises an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

47.

407. The polypeptide complex of claim 406, wherein the CD8 transmembrane domain comprises the amino acid sequence shown in SEQ ID NO:

47.

408. The polypeptide complex of any one of claims 330 to 407, wherein the targeting component further comprises an intracellular signaling domain or a co-stimulatory domain derived from a protein selected from the group consisting of an antigen receptor, a co-stimulatory receptor, a growth receptor, a cytokine receptor, an adaptor signaling protein, an intracellular signaling protein, or any fragment or variant thereof.

409. The polypeptide complex of claim 408, wherein the intracellular signaling domain or the co-stimulatory domain on the targeting component is selected from the group consisting of: Toll-like receptor 1 (TLR1), TLR2, TLR3, TLR4, TLR5, TLR6, TLR7, TLR8, TLR9, TLR10, caspase recruitment domain family member 11 (CARD11), CD2, CD3ε, CD3γ, CD3δ, CD4, CD7, CD8, CD27, CD28, CD30, CD40, CD54 (ICAM), CD83, CD94, CD134 (OX40), CD 137 (4-1BB), CD278 (ICOS), common gamma chain cytokine, DNAX-activating protein 10 (DAP10), adaptor for activating T-cell family member 1 (LAT), interleukin 2 receptor (IL-2R), IL-4R, IL-7R, IL-9R, IL-12R, IL-13R, IL-15R, IL-21R, SH2 domain-containing leukocyte protein of 76 kD (SLP76), T-cell receptor-associated transmembrane adaptor 1 (TRAT1), TNFR2, TNFRS14, TNFRS18, TNRFS25, and T-cell receptor zeta chain-associated protein kinase 70 (ZAP70).

410. The polypeptide complex of claim 408 or claim 409, wherein the intracellular signaling domain or the co-stimulatory domain on the targeting component is selected from the group consisting of: 4-1BB, CD28, TNFR2, OX40, ICOS, and DAP10 co-stimulatory domains.

411. The polypeptide complex of claim 410, wherein the co-stimulatory domain on the targeting component is a 4-1BB co-stimulatory domain, optionally wherein the 4-1BB co-stimulatory domain comprises an amino acid sequence as shown in SEQ ID NO:

98.

412. The polypeptide complex of claim 410, wherein the co-stimulatory domain on the targeting component is a CD28 co-stimulatory domain, optionally wherein the CD28 co-stimulatory domain comprises the amino acid sequence as shown in SEQ ID NO:

99.

413. The polypeptide complex of claim 408, wherein the intracellular signaling domain or the co-stimulatory domain is one or more cytokine receptor intracellular signaling domains.

414. The polypeptide complex of claim 413, wherein the one or more cytokine receptor intracellular signaling domains are selected from the group consisting of an IL7Rα intracellular signaling domain, an IL2Rβ intracellular signaling domain, a common γ chain intracellular signaling domain, and both IL2Rβ and common γ chain intracellular signaling domains.

415. The polypeptide complex of claim 414, wherein the IL7Rα intracellular signaling domain comprises an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

103.

416. The polypeptide complex according to claim 415, wherein the IL7Rα intracellular signaling domain comprises the amino acid sequence as shown in SEQ ID NO:

103.

417. The polypeptide complex of claim 414, wherein the IL2Rβ intracellular signaling domain comprises an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO: 104 or SEQ ID NO:

105.

418. The polypeptide complex of claim 417, wherein the IL2Rβ intracellular signaling domain comprises the amino acid sequence shown in SEQ ID NO: 104 or SEQ ID NO:

105.

419. The polypeptide complex of claim 414, wherein the common γ chain intracellular signaling domain comprises an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

106.

420. The polypeptide complex of claim 419, wherein the common γ chain intracellular signaling domain comprises the amino acid sequence shown in SEQ ID NO:

106.

421. The polypeptide complex of claim 408, wherein the intracellular signaling domain or the co-stimulatory domain is a LAT domain.

422. The polypeptide complex of claim 421, wherein the LAT domain comprises an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to the amino acid sequence shown in SEQ ID NO:

102.

423. The polypeptide complex of claim 422, wherein the LAT domain comprises the amino acid sequence shown in SEQ ID NO:

102.

424. The polypeptide complex of claim 408, wherein the intracellular signaling domain or the co-stimulatory domain is a CD4 co-receptor domain.

425. The polypeptide complex of claim 424, wherein the CD4 co-receptor domain comprises an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

100.

426. The polypeptide complex of claim 425, wherein the CD4 co-receptor domain comprises the amino acid sequence shown in SEQ ID NO:

100.

427. The polypeptide complex of claim 408, wherein the intracellular signaling domain or the co-stimulatory domain is a CD8 co-receptor domain.

428. The polypeptide complex of claim 427, wherein the CD8 co-receptor domain comprises an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

101.

429. The polypeptide complex of claim 428, wherein the CD8 co-receptor domain comprises the amino acid sequence shown in SEQ ID NO:

101.

430. The polypeptide complex of any one of claims 330 to 407, wherein the targeting component does not comprise a functional intracellular signaling domain or a co-stimulatory domain.

431. The polypeptide complex of any one of claims 330 to 430, wherein the targeting component further comprises a truncated intracellular CD4 polypeptide.

432. The polypeptide complex of claim 431, wherein the truncated intracellular CD4 polypeptide comprises the amino acid sequence shown in SEQ ID NO:48 or SEQ ID NO:

49.

433. The polypeptide complex of claim 432, wherein the truncated intracellular CD4 polypeptide comprises the amino acid sequence shown in SEQ ID NO:48 or SEQ ID NO:

49.

434. The polypeptide complex of any one of claims 330 to 433, wherein the extracellular domain comprises a first targeting domain.

435. The polypeptide complex of any one of claims 330 to 434, wherein the first targeting domain comprises a single-chain variable fragment (scFv) or a single domain antibody (sdAb).

436. The polypeptide complex of claim 435, wherein the sdAb is a camelid VHH, a nanobody or a heavy chain only antibody (HcAb).

437. The polypeptide complex of claim 436, wherein the sdAb is a camelid VHH.

438. The polypeptide complex of claim 436 or claim 437, wherein the scFv or the sdAb is human or humanized.

439. The polypeptide complex of any one of claims 434 to 438, wherein the first targeting domain comprises a sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to any one of SEQ ID NOs: 50-83.

440. The polypeptide complex of claim 439, wherein the first targeting domain comprises a sequence as shown in any one of SEQ ID NOs: 50-83.

441. The polypeptide complex of claims 434 to 438, wherein the first targeting domain comprises the PD1 extracellular domain, human A proliferation-inducing ligand (APRIL), trimerized human APRIL, or NKG2D membrane protein.

442. The polypeptide complex of claim 441, wherein the PD1 extracellular domain comprises a sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:84 or SEQ ID NO:

85.

443. The polypeptide complex according to claim 442, wherein the PD1 extracellular domain comprises the sequence shown in SEQ ID NO:

85.

444. A polypeptide complex according to claim 441, wherein the APRIL comprises a sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:86, or wherein the trimeric human APRIL comprises a sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

87.

445. The polypeptide complex of claim 444, wherein the APRIL comprises the sequence as shown in SEQ ID NO:86, or wherein the trimerized human APRIL comprises the sequence as shown in SEQ ID NO:

87.

446. The polypeptide complex of claim 441, wherein the NKG2D membrane protein comprises a sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

88.

447. The polypeptide complex of claim 446, wherein the NKG2D membrane protein comprises the sequence shown in SEQ ID NO:

88.

448. The polypeptide complex of any one of claims 434 to 447, wherein the extracellular domain further comprises a second targeting domain.

449. The polypeptide complex of claim 448, wherein the second targeting domain comprises a second single-chain variable fragment (scFv) or a second single domain antibody (sdAb).

450. The polypeptide complex of claim 449, wherein the second sdAb is a camelid VHH, a nanobody, or a heavy chain only antibody (HcAb).

451. The polypeptide complex of claim 450, wherein the second sdAb is a camelid VHH.

452. The polypeptide complex of any one of claims 449 to 451, wherein the second scFv or the second sdAb is human or humanized.

453. The polypeptide complex of any one of claims 448 to 453, wherein the second targeting domain comprises a sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to any one of SEQ ID NOs: 50-83.

454. The polypeptide complex of claim 454, wherein the second targeting domain comprises a sequence as shown in any one of SEQ ID NOs: 50-83.

455. The polypeptide complex of claim 448, wherein the second targeting domain comprises the PD1 extracellular domain, human A proliferation-inducing ligand (APRIL), trimerized human APRIL, or NKG2D membrane protein.

456. The polypeptide complex of claim 455, wherein the PD1 extracellular domain comprises a sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:84 or SEQ ID NO:

85.

457. The polypeptide complex according to claim 456, wherein the PD1 extracellular domain comprises the sequence shown in SEQ ID NO:

85.

458. A polypeptide complex according to claim 455, wherein the APRIL comprises a sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:86, or wherein the trimeric human APRIL comprises a sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

87.

459. The polypeptide complex of claim 458, wherein the APRIL comprises the sequence as shown in SEQ ID NO:86, or wherein the trimerized human APRIL comprises the sequence as shown in SEQ ID NO:

87.

460. The polypeptide complex of claim 455, wherein the NKG2D membrane protein comprises a sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

88.

461. The polypeptide complex of claim 460, wherein the NKG2D membrane protein comprises the sequence shown in SEQ ID NO:

88.

462. The polypeptide complex of any one of claims 448 to 461, wherein the first targeting domain and the second targeting domain bind to the same antigen or different antigens.

463. The polypeptide complex of any one of claims 448 to 462, wherein the targeting domain and the second targeting domain are separated by a third polypeptide linker having a length of 2 to 40 amino acids.

464. The polypeptide complex of claim 463, wherein the third polypeptide linker is selected from the group consisting of GG, GS, SG, SS, GSS, SSG, GSG, SGS, SGG, G4S, 2x G4S, 3xG4s, 4xG4S, 5xG4S, any one of SEQ ID NOs: 16-31, and any combination thereof.

465. The polypeptide complex of any one of claims 330 to 464, wherein the signaling component comprises a sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO: 107, SEQ ID NO: 108, SEQ ID NO: 109, SEQ ID NO: 110, SEQ ID NO: 111, SEQ ID NO: 112, SEQ ID NO: 113, SEQ NO: 114 or SEQ ID NO:

115.

466. The polypeptide complex of claim 165, wherein the signaling component comprises a sequence as shown in SEQ ID NO:107, SEQ ID NO:108, SEQ ID NO:109, SEQ ID NO:110, SEQ ID NO:111, SEQ ID NO:112, SEQ ID NO:113, SEQ NO:114, or SEQ ID NO:

115.

467. The polypeptide complex of any one of claims 330 to 466, wherein the targeting component comprises a sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to a sequence as shown in SEQ ID NO: 116, SEQ ID NO: 117, SEQ ID NO: 118, SEQ ID NO: 119, SEQ ID NO: 120, SEQ ID NO: 121, SEQ ID NO: 122, SEQ ID NO: 123, SEQ ID NO: 124, SEQ ID NO: 125, SEQ ID NO: 126, SEQ ID NO: 127, SEQ ID NO: 128, SEQ ID NO: 129, SEQ ID NO: 130, SEQ ID NO: 131, SEQ ID NO: 132, SEQ ID NO: 133, SEQ ID NO: 134, SEQ ID NO: 135 or SEQ ID NO:

136.

468. The polypeptide complex of claim 467, wherein the targeting component comprises a sequence as shown in SEQ ID NO: 116, SEQ ID NO: 117, SEQ ID NO: 118, SEQ ID NO: 119, SEQ ID NO: 120, SEQ ID NO: 121, SEQ ID NO: 122, SEQ ID NO: 123, SEQ ID NO: 124, SEQ ID NO: 125, SEQ ID NO: 126, SEQ ID NO: 127, SEQ ID NO: 128, SEQ ID NO: 129, SEQ ID NO: 130, SEQ ID NO: 131, SEQ ID NO: 132, SEQ ID NO: 133, SEQ ID NO: 134, SEQ ID NO: 135, or SEQ ID NO:

136.

469. The polypeptide complex of any one of claims 330 to 468, comprising a fusion polypeptide comprising the targeting component and the signaling component.

470. The polypeptide complex of claim 469, wherein the fusion polypeptide comprises SEQ ID NO: 139, SEQ ID NO: 140, SEQ ID NO: 141, SEQ ID NO: 142, SEQ ID NO: 143, SEQ ID NO: 144, SEQ ID NO: 145, SEQ ID NO: 146, SEQ ID NO: 147, SEQ ID NO: 148, SEQ ID NO: 149, SEQ ID NO: 150, SEQ ID NO: 151, SEQ ID NO: 152, SEQ ID NO: 153, SEQ ID NO: 154, SEQ ID NO: 155, SEQ ID NO: 156, SEQ ID NO: 157, SEQ ID NO: 158, SEQ ID NO: 159, SEQ ID NO: 160, SEQ ID NO: 161, SEQ ID NO: 162, SEQ ID NO: 163, SEQ ID NO: 164, SEQ ID NO: 165, SEQ ID NO: 166, SEQ ID NO: 167, SEQ ID NO: 167, SEQ ID NO: 168, SEQ ID NO: 169 or SEQ ID NO: 170 having a sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical.

471. The polypeptide complex of claim 470, wherein the fusion polypeptide comprises SEQ ID NO: 139, SEQ ID NO: 140, SEQ ID NO: 141, SEQ ID NO: 142, SEQ ID NO: 143, SEQ ID NO: 144, SEQ ID NO: 145, SEQ ID NO: 146, SEQ ID NO: 147, SEQ ID NO: 148, SEQ ID NO: 149, SEQ ID NO: 150, SEQ ID NO: 151, SEQ ID NO: 152, SEQ ID NO: 153, SEQ ID NO: 154, SEQ ID NO: 155, SEQ ID NO: 156, SEQ ID NO: 157, SEQ ID NO: 158, SEQ ID NO: 159, SEQ ID NO: 160, SEQ ID NO: 161, SEQ ID NO: 162, SEQ ID NO: 163, SEQ ID NO: 164, SEQ ID NO: 165, SEQ ID NO: 166, SEQ ID NO: 167, The sequence shown in SEQ ID NO:167, SEQ ID NO:168, SEQ ID NO:169 or SEQ ID NO:

170.

472. A nucleic acid molecule encoding both the signaling component and the targeting component of the polypeptide complex according to any one of claims 330 to 471.

473. A cell comprising the polypeptide complex of any one of claims 330 to 471.

474. A cell comprising a nucleic acid molecule according to claim 472.

475. The cell of any one of claims 473 or 474, wherein the cell further expresses an exogenous co-stimulatory factor, an immunomodulatory factor, an agonist of a co-stimulatory factor, an antagonist of an immunosuppressive factor, an immune cell engager, a flip receptor, or any combination thereof.

476. The cell of any one of claims 473 to 475, wherein the cell further expresses an exogenous lymphocyte receptor or co-receptor.

477. The cell of claim 476, wherein the exogenous lymphocyte receptor or co-receptor is selected from the group consisting of TCRα (TCR alpha), TCRβ (TCR beta), TCRγ (TCR gamma), TCRδ (TCR delta), CD4, CD8, pre-T cell receptor alpha (pTα), Fc receptor alpha (FcRα), Fc receptor beta (FcRβ), Fc receptor gamma (FcRγ), natural killer group 2 member D (NKG2D), CD79A, CD79B, and any combination thereof.

478. The cell of any one of claims 473 to 477, wherein the cell further expresses an exogenous TCR.

479. The cell of claim 478, wherein the exogenous TCR binds to a target antigen selected from the group consisting of alpha-fetoprotein (AFP), members of the B melanoma antigen (BAGE) family, brothers of regulatory factors at imprinted sites (BORIS), cancer-testis antigen, cancer-testis antigen 83 (CT-83), carbonic anhydrase IX (CA1X), carcinoembryonic antigen (CEA), cytomegalovirus (CMV) antigen, antigen recognized by cytotoxic T cells (CTL) on melanoma (CAMEL), Epstein-Barr virus (EBV ) antigen, G antigen 1 (GAGE-1), GAGE-2, GAGE-3, GAGE-4, GAGE-5, GAGE-6, GAGE-7B, GAGE-8, glycoprotein 100 (GP100), hepatitis B virus (HBV) antigen, hepatitis C virus (HCV) nonstructural protein 3 (NS3), human epidermal growth factor receptor 2 (HER-2), human papillomavirus (HPV)-E6, HPV-E7, human telomerase reverse transcriptase (hTERT), IGF2BP3 / A3, K-Ras, K-Ras G12C, K-Ras G12D, K-Ras G12V, latent membrane protein 2 (LMP2), melanoma antigen family A, 1 (MAGE-A1), MAGE-A2, MAGE-A3, MAGE-A4, MAGE-A6, MAGE-A10, MAGE-A12, melanoma antigen recognized by T cells (MART-1), mesothelin (MSLN), mucin 1 (MUC1), mucin 16 (MUC16), New York esophageal squamous cell carcinoma-1 (NYESO-1), P53, P antigen (PAGE) family member, placenta-specific 1 (PLAC1), antigen preferentially expressed in melanoma (PRAME), survivin, synovial sarcoma X1 (SSX1), synovial sarcoma X2 (SSX2), synovial sarcoma X3 (SSX3), synovial sarcoma X4 (SSX4), synovial sarcoma X5 (SSX5), synovial sarcoma X 8 (SSX8), thyroglobulin, tyrosinase, tyrosinase-related protein (TRP) 1, TRP2, Wilms' tumor protein (WT-1), X antigen family member 1 (XAGE1), and X antigen family member 2 (XAGE2).

480. The cell of claim 478 or claim 479, wherein the exogenous TCR is an αβ-TCR or a γδ-TCR.

481. The cell of any one of claims 473 to 480, wherein the cell further expresses a CAR, a CCR or a flip receptor.

482. The cell of any one of claims 473 to 481, wherein the cell further expresses a zetakine, an immune cell engager, or a BiTE.

483. The cell of any one of claims 473 to 482, wherein the cell is a hematopoietic cell.

484. The cell of any one of claims 473 to 483, wherein the cell is a T cell, an αβ-T cell, or a γδ-T cell.

485. The cell of any one of claims 473 to 484, wherein the cell is a CD3 + cells, CD4 + cells and / or CD8 + cell.

486. The cell of any one of claims 473 to 485, wherein the cell is an immune effector cell.

487. The cell of any one of claims 473 to 486, wherein the cell is a cytotoxic T lymphocyte (CTL), a tumor infiltrating lymphocyte (TIL), or a helper T cell.

488. The cell of any one of claims 473 to 487, wherein the cell is a Natural Killer (NK) cell or a Natural Killer T (NKT) cell.

489. The cell of any one of claims 473 to 488, wherein the source of the cell is peripheral blood mononuclear cells, bone marrow, lymph node tissue, umbilical cord blood, thymus tissue, tissue from a site of infection, ascites, pleural effusion, spleen tissue, or a tumor.

490. The cell of any one of claims 473 to 489, wherein the cell is an isolated cell.

491. The cell of any one of claims 473 to 490, wherein the cell is obtained from a subject.

492. The cell of any one of claims 473 to 491, wherein the cell is a human cell.

493. A polynucleotide encoding the signaling component and the targeting component of the fusion polypeptide of any one of claims 164 to 308 or the polypeptide complex of any one of claims 330 to 471.

494. A cDNA encoding the signaling component and the targeting component of the fusion polypeptide of any one of claims 164 to 308 or the polypeptide complex of any one of claims 330 to 471.

495. An RNA encoding the signaling component and the targeting component of any one of claims 1 to 81 and 144 to 203, or a fusion polypeptide according to any one of claims 82 to 143, a fusion polypeptide according to any one of claims 164 to 308, or a polypeptide complex according to any one of claims 330 to 471.

496. A vector comprising the polynucleotide of claim 493.

497. The vector of claim 496, wherein the vector is an expression vector.

498. The vector of claim 496, wherein the vector is a transposon.

499. The vector of claim 493, wherein the vector is a piggyBAC transposon or a Sleeping Beauty transposon.

500. The vector of claim 496, wherein the vector is a viral vector.

501. The vector of claim 500, wherein the vector is an adenoviral vector, an adeno-associated virus (AAV) vector, a herpes virus vector, a vaccinia virus vector, or a retroviral vector.

502. The vector of claim 501, wherein the retroviral vector is a lentiviral vector.

503. The vector of claim 502, wherein the lentiviral vector is selected from the group consisting of: human immunodeficiency virus 1 (HIV-1); human immunodeficiency virus 2 (HIV-2), visna-maedi virus (VMV) virus; caprine arthritis-encephalitis virus (CAEV); equine infectious anemia virus (EIAV); feline immunodeficiency virus (FIV); bovine immunodeficiency virus (BIV); and simian immunodeficiency virus (SIV).

504. A cell comprising the fusion polypeptide of any one of claims 164 to 308, the polynucleotide of claim 493, or the vector of any one of claims 496 to 503.

505. The cell of claim 504, wherein the cell is a hematopoietic cell.

506. The cell of claim 504 or claim 505, wherein the cell is an immune effector cell.

507. The cell of any one of claims 504 to 506, wherein the cell is a T cell, an αβ-T cell, or a γδ-T cell.

508. The cell of any one of claims 504 to 507, wherein the cell expresses CD3 + 、CD4 + 、CD8 + or a combination thereof.

509. The cell of any one of claims 504 to 508, wherein the cell is a cytotoxic T lymphocyte (CTL), a tumor infiltrating lymphocyte (TIL), or a helper T cell.

510. The cell of any one of claims 504 to 508, wherein the cell is a natural killer (NK) cell or a natural killer T (NKT) cell.

511. A composition comprising a cell according to any one of claims 1 to 163, 310 to 329, 473 to 492 or 504 to 510, or a vector according to any one of claims 496 to 503.

512. A composition comprising a physiologically acceptable carrier and a cell according to any one of claims 1 to 163, 310 to 329, 473 to 492 or 504 to 510, or a vector according to any one of claims 496 to 503.

513. A method of treating a subject in need thereof, the method comprising administering to the subject an effective amount of a composition according to claim 511 or claim 512.

514. A method for treating, preventing or ameliorating at least one symptom of cancer, infectious diseases, autoimmune diseases, inflammatory diseases and immunodeficiency or conditions associated therewith, the method comprising administering to a subject an effective amount of a composition according to claim 511 or claim 512.

515. A method of treating a solid cancer, the method comprising administering to a subject an effective amount of a composition according to claim 513 or claim 514.

516. The method of claim 515, wherein the solid cancer is selected from the group consisting of lung cancer, squamous cell carcinoma, colorectal cancer, pancreatic cancer, breast cancer, thyroid cancer, bladder cancer, cervical cancer, esophageal cancer, ovarian cancer, gastric cancer, endometrial cancer, or brain cancer.

517. The method of claim 515 or claim 516, wherein the solid cancer is non-small cell lung cancer, squamous cell carcinoma of the head and neck, colorectal cancer, pancreatic cancer, breast cancer, thyroid cancer, bladder cancer, cervical cancer, esophageal cancer, ovarian cancer, gastric cancer, endometrial cancer, glioma, glioblastoma, or oligodendroglioma.

518. A method of treating a hematological malignancy, the method comprising administering to a subject an effective amount of a composition according to claim 515 or claim 516.

519. The method of claim 518, wherein the hematological malignancy is a leukemia, a lymphoma, or multiple myeloma.

520. The method of claim 518, wherein the hematological malignancy is acute myeloid leukemia (AML).

521. The non-natural cell of any one of claims 1 to 163, wherein the cell further comprises a polypeptide that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

137.

522. The non-natural cell of any one of claims 1 to 163, wherein the cell further comprises the amino acid sequence shown in SEQ ID NO:

137.

523. The non-natural cell of any one of claims 1 to 163, wherein the cell further comprises a polypeptide that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO:

138.

524. The non-natural cell of any one of claims 1 to 163, wherein the cell further comprises the amino acid sequence shown in SEQ ID NO: 138.

Images