Compositions for modifying cells

Abstract

The present disclosure provides compositions and methods comprising recombinant particles suitable for specifically delivering one or more engineered receptors to immune effector cells in vivo.

Description

[0001] COMPOSITIONS FOR MODIFYING CELLS

[0002] CROSS REFERENCE TO RELATED APPLICATIONS

[0003] This application claims the benefit under 35 U.S. C. § 119(e) of U.S. Provisional Application No. 63 / 532,093, filed August 11, 2023, and U.S. Provisional Application No. 63 / 540,344, filed September 25, 2023, and each of which is incorporated by reference herein in its entirety.

[0004] STATEMENT REGARDING SEQUENCE LISTING

[0005] The Sequence Listing associated with this application is provided in XML format in lieu of a paper copy and is hereby incorporated by reference into the specification. The name of the XML file containing the Sequence Listing is KELO-010-WOl_ST26.xml. The XML file is 744 KB, was created on July 25, 2024, and is being submitted electronically via Patent Center, concurrent with the filing of the specification.

[0006] Technical Field

[0007] The present disclosure relates to recombinant particles engineered to deliver one or more engineered receptors to a cell. More particularly, the disclosure relates to recombinant particles engineered to deliver one or more engineered receptors to cells in vivo.

[0008] Description of the Related Art

[0009] Ex vivo gene therapies are potentially one-time therapeutic modalities that generally involve harvesting cells from a subject, modifying the cells by culturing them with a gene therapy vector, and delivering the modified cells back to the subject. Because ex vivo gene therapies are manufactured in a controlled environment, they do not generally require specialized targeting moieties and instead, targeting moieties with a broad tropism and that are highly efficient in delivering a gene therapy to most cell types are used.

[0010] In contrast, in vivo gene therapies are manufactured in the patient, in an uncontrolled environment. Accordingly, in vivo delivery of gene therapy vectors to specific cell types is orders of magnitude more complex than ex vivo delivery. In vivo gene therapy vectors encounter many non-target or off-target cells and may require a narrower or more specific tropism to deliver therapeutic payloads to a particular cell type. The potential of in vivo gene therapies has yet to be realized primarily due to inefficient delivery to desired cell types in combination with substantial off-target delivery. Use of specialized targeting moieties to deliver gene therapies in vivo has proven difficult in abrogating the off-target delivery to undesired cell types. In addition, on-target delivery of in vivo gene therapies using such specialized targeting moieties is often inefficient.

[0011] BRIEF SUMMARY

[0012] The present disclosure generally relates, in part, to a recombinant particle comprising a fusogen and one or more tropism polypeptides that are suitable for delivering a payload to one or more desired cell types in vivo. The present disclosure more particularly relates to recombinant viral, e.g., retroviral or lentiviral particles comprising a detargeted viral-based fusogen, one or more non- viral membrane bound tropism polypeptides and a vector, e.g., a viral, retroviral, or lentiviral vector encoding an expressible polynucleotide encoding an engineered receptor.

[0013] In various embodiments, a recombinant particle is contemplated, comprising: (a) a surface comprising (i) one or more mutated viral envelope glycoproteins that retain fusogenic activity and that lack cognate receptor binding activity and (ii) a non-viral membrane-bound tropism polypeptide comprising an antibody or antigen binding fragment thereof that binds an antigen expressed on an immune effector cell, a spacer polypeptide, and a GPI anchor domain or a transmembrane domain, and optionally, an intracellular domain comprising no more than 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 amino acids; and (b) a therapeutic payload.

[0014] In particular embodiments, the particle is a fusosome; an extracellular vesicle, including a microvesicle, an apoptotic body, and an exosome; a lipid nanoparticle; a vims- like particle (VLPs); or a recombinant viral particle.

[0015] In certain embodiments, the surface comprises a single layer of phospholipid, a phospholipid bilayer, a cell membrane, a capsid, or a viral envelope.

[0016] In some embodiments, the payload comprises one or more polynucleotides and / or polypeptides. In particular embodiments, the payload comprises a vector that encodes a therapeutic polynucleotide or polypeptide.

[0017] In particular embodiments, the payload comprises a vector that encodes one or more engineered receptors.

[0018] In further embodiments, the pay load comprises a vector that encodes a promoter operably linked to a polynucleotide encoding one or more engineered receptors.

[0019] In various embodiments, a recombinant lentiviral particle is contemplated, comprising: (a) a viral envelope comprising (i) one or more mutated viral envelope glycoproteins that retain fusogenic activity and that lack cognate receptor binding activity and (ii) a non- viral membrane-bound tropism polypeptide comprising an antibody or antigen binding fragment thereof that binds an antigen expressed on an immune effector cell, a spacer polypeptide, a GPI domain or transmembrane domain and optionally, an intracellular domain comprising no more than 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 amino acids; and (b) a recombinant lentiviral vector comprising a polynucleotide encoding a promoter operably linked to a polynucleotide encoding one or more engineered receptors.

[0020] In some embodiments, the one or more mutated viral envelope glycoproteins comprise a rhabdovirus envelope glycoprotein, or one or more paramyxovirus envelope glycoproteins.

[0021] In certain embodiments, the one or more mutated viral envelope glycoproteins comprise a vesiculovirus envelope glycoprotein, one or more morbillivirus envelope glycoproteins or one or more henipavirus envelope glycoproteins.

[0022] In particular embodiments, the vesiculovirus is selected from the group consisting of: vesicular stomatitis Alagoas virus (VSAV; Alagoas vesiculovirus), Carajas virus (CJSV; Carajas vesiculovirus), Chandipura vims (CHPV; Chandipura vesiculovirus), Cocal vims (COCV; Cocal vesiculovirus), vesicular stomatitis Indiana vims (VSIV; Indiana vesiculovims), Isfahan vims (ISFV ; Isfahan vesiculovirus), Maraba vims (MARAV ; Maraba vesiculovims), Morreton vims (MORV; Morreton vesiculovims), vesicular stomatitis New Jersey vims (VSNJV; New Jersey vesiculovims), and Piry vims (PIRYV; Piry vesiculovims).

[0023] In additional embodiments, the vesiculovims envelope glycoprotein is a vesiculovims G protein.

[0024] In additional embodiments, the vesiculovims G protein is a COCV G glycoprotein (COCV-G) or a VSIV G glycoprotein (VSIV-G). In certain embodiments, the VSIV-G envelope protein comprises one or more of: (a) one or more amino acid substitutions at H8, N9, Q10, K47, K50, A51, S183, S179, N180, 1182, M184, Y209, 1347, T350, T352, E353, and R354; (b) an insertion of TT between N9 and Q10, an insertion of GGS between H8 and N9, an insertion of GGS between N9 and Q10, an insertion of TT between N208 and Y209, an insertion of GGS between P46 and K47, and an insertion of GGS between N208 and Y209; or (c) amino acid substitutions at K47 and / or R354; or (d) a deletion of residues 1-8.

[0025] In particular embodiments, the VSIV-G envelope protein comprises one or more amino acid substitutions at H8, K47, Y209, and R354.

[0026] In particular embodiments, the VSIV-G polypeptide comprises one or more amino acid substitutions at K47, 1182, and / or R354 (substitution with any amino acid; a conservation substitution; a disruptive substitution; substitution with D, E, A, G, F, or Q; or substitution with A, G, F, or Q). In particular embodiments, the VSIV-G polypeptide comprises amino acid substitutions at K47, 1182, or R354; K47 and 1182; K47 and R354; 1182 and R354; or at K47, 1182, and R354 of an amino acid sequence at least 95% identical to, at least 96% identical to, at least 97% identical to, and least 98% identical to, at least 99% identical to, or identical to, SEQ ID NO: 1.

[0027] In particular embodiments, the VSIV-G polypeptide comprises one or more of the following amino acid substitutions: K47A, K47Q, I182E, I182D, R354A, and / or R354Q.

[0028] In particular embodiments, the VSIV-G polypeptide comprises the following amino acid substitutions: K47A and I182E; K47A and I182D; K47Q and I182E; K47Q and I182D; I182E and R354A; I182E and R354Q; I182D and R354A; I182D and R354Q; K47A and R354A; K47A and R354Q; K47Q and R354A; K47Q and R354Q; K47A, I182E, and R354A; K47A, I182D, and R354A; K47Q, I182E, and R354A; K47Q, I182D, and R354A; K47A, I182E, and R354Q; K47A, I182D, and R354Q; K47Q, I182E, and R354Q; or K47Q, I182D, and R354Q of an amino acid sequence at least 95% identical to, at least 96% identical to, at least 97% identical to, and least 98% identical to, at least 99% identical to, or identical to, SEQ ID NO: 1.

[0029] In particular embodiments, the VSIV-G envelope protein comprises amino acid substitutions selected from the group consisting of: K47A and R354A; K47A and R354G; K47A and R354F; K47A and R354Q; K47G and R354A; K47G and R354G; K47G and R354F; K47G and R354Q; K47F and R354A; K47F and R354G; K47F and R354F; K47F and R354Q; K47Q and R354A; K47Q and R354G; K47Q and R354F; and K47Q and R354Q.

[0030] In certain embodiments, the VSIV-G envelope protein comprises the amino acid sequence set forth in SEQ ID NO: 2 or an amino acid sequence at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical thereto, wherein Xi= I, X2= A, X3 = Q, and X4= A; Xi= I, X2= A, X3= Q, and X4= G; Xi= I, X2= A, X3= Q, and X4= F; Xi= I, X2= A, X3= Q, and X4= Q; Xi= L, X2= A, X3= Q, and X4= A; Xi= L, X2= A, X3= Q, and X4= G; Xi= L, X2= A, X3= Q, andX4= F; Xi= E, X2= A, X3= Q, andX4= Q; Xi= I, X2= A, X3= H, and X4= A; Xi= I, X2= A, X3= H, and X4= G; Xi= I, X2= A, X3= H, and X4= F; Xi= I, X2= A, X3= H, and X4= Q; Xi= E, X2= A, X3= H, and X4= A; Xi= E, X2= A, X3= H, and X4= G; Xi= L, X2= A, X3= H, and X4= F; Xi= L, X2= A, X3= H, and X4= Q; Xi= I, X2= G, X3= Q, and X4= A; Xi= I, X2= G, X3= Q, and X4= G; Xi= I, X2= G, X3= Q, and X4= F; Xi= I, X2= G, X3= Q, and X4= Q; Xi= E, X2= G, X3= Q, and X4= A; Xi= E, X2= G, X3= Q, and X4= G; Xi= L, X2= G, X3= Q, and X4= F; Xi= L, X2= G, X3= Q, and X4= Q; Xi= I, X2= G, X3= H, and X4= A; Xi= I, X2= G, X3= H, and X4= G; Xi= I, X2= G, X3= H, andX4= F; Xi= I, X2= G, X3= H, andX4= Q; Xi= E, X2= G, X3= H, and X4= A; Xi= E, X2= G, X3= H, and X4= G; Xi= L, X2= G, X3= H, and X4= F; Xi= L, X2= G, X3= H, and X4= Q; Xi= I, X2= F, X3= Q, and X4= A; Xi= I, X2= F, X3= Q, and X4= G; Xi= I, X2= F, X3= Q, and X4= F; Xi= I, X2= F, X3= Q, and X4= Q; Xi= E, X2= F, X3= Q, and X4= A; Xi= E, X2= F, X3= Q, and X4= G; Xi= E, X2= F, X3= Q, and X4= F; Xi= E, X2= F, X3= Q, and X4= Q; Xi= I, X2= F, X3= H, and X4= A; Xi= I, X2= F, X3= H, and X4= G; Xi= I, X2= F, X3= H, and X4= F; Xi= I, X2= F, X3= H, and X4= Q; Xi= E, X2= F, X3= H, and X4= A; Xi= E, X2= F, X3= H, and X4= G; Xi= E, X2= F, X3= H, and X4= F; Xi= E, X2= F, X3= H, and X4= Q; Xi= I, X2= Q, X3= Q, and X4= A; Xi= I, X2= Q, X3= Q, and X4= G; Xi= I, X2= Q, X3= Q, and X4= F; Xi= I, X2= Q, X3= Q, and X4= Q; Xi= E, X2= Q, X3= Q, and X4= A; Xi= E, X2= Q, X3= Q, and X4= G; Xi= E, X2= Q, X3= Q, and X4= F; Xi= E, X2= Q, X3= Q, andX4= Q; Xi= I, X2= Q, X3= H, andX4= A; Xi= I, X2= Q, X3= H, and X4= G; Xi= I, X2= Q, X3= H, and X4= F; Xi= I, X2= Q, X3= H, and X4= Q; Xi= E, X2= Q, X3= H, and X4= A; Xi= E, X2= Q, X3= H, and X4= G; Xi= E, X2= Q, X3= H, and X4= F; and Xi= E, X2= Q, X3= H, and X4= Q.

[0031] In some embodiments, the VSIV-G envelope protein comprises the amino acid sequence set forth in any one of SEQ ID NOs: 582-645 or an amino acid sequence at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical thereto. In particular embodiments, the VSIV-G envelope protein comprises the amino acid sequence set forth in any one of SEQ ID NOs: 630, 634, 638, and 642 or an amino acid sequence at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical thereto. In a particular preferred embodiment, the VSIV-G envelope protein comprises the amino acid sequence set forth in SEQ ID NO: 642 or an amino acid sequence at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical thereto.

[0032] In additional embodiments, the vesiculovirus G protein is COCV-G.

[0033] In some embodiments, the COCV-G envelope protein comprises one or more amino acid substitutions at K47 and / or R354.

[0034] In certain embodiments, the COCV-G envelope protein comprises amino acid substitutions selected from the group consisting of: K47A and R354A; K47A and R354G; K47A and R354F; K47A and R354Q; K47G and R354A; K47G and R354G; K47G and R354F; K47G and R354Q; K47F and R354A; K47F and R354G; K47F and R354F; K47F and R354Q; K47Q and R354A; K47Q and R354G; K47Q and R354F; and K47Q and R354Q.

[0035] In further embodiments, the COCV-G envelope protein comprises the amino acid sequence set forth in SEQ ID NO: 4 or an amino acid sequence at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical thereto, wherein Xi= A and X2= A; Xi= A and X2= G; Xi= A and X2= F; Xi= A and X2= Q; Xi= G and X2= A; Xi= G and X2= G; Xi= G and X2= F; Xi= G and X2= Q; Xi= F and X2= A; Xi= F and X2= G; Xi= F and X2= F; Xi= F and X2= Q; Xi= Q and X2= A; Xi= Q and X2= G; Xi= Q and X2= F; or Xi= A and X2= Q.

[0036] In particular embodiments, the COCV-G envelope protein comprises the amino acid sequence set forth in any one of SEQ ID NOs: 646-661 or an amino acid sequence at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical thereto.

[0037] In certain embodiments, the one or more morbillivirus envelope glycoproteins are measles virus F (MV-F) and measles virus H (MV-H).

[0038] In some embodiments, the MV-H protein comprises one or more amino acid substitutions at Y463, R515, S530, and F531.

[0039] In additional embodiments, the MV-H protein comprises one or more amino acid substitutions selected from the group consisting of: Y463A, R515A, S530, and F531.

[0040] In some embodiments, the MV-H protein comprises the amino acid sequence set forth in SEQ ID NO: 7. In certain embodiments, the one or more henipavirus envelope glycoproteins are nipah virus F (NiV-F) and nipah virus G (NiV-G).

[0041] In particular embodiments, the NiV-G protein comprises one or more amino acid substitutions at E468, W471, Q497, and E500.

[0042] In additional embodiments, the NiV-G protein comprises one or more amino acid substitutions selected from the group consisting of: E468A, W471A, Q497A, and E500A.

[0043] In particular embodiments, the NiV-G protein comprises the amino acid sequence set forth in SEQ ID NO: 10.

[0044] In further embodiments, the antibody or antigen binding fragment thereof is selected from the group consisting of: a Camel Ig, a Llama Ig, an Alpaca Ig, Ig NAR, a Fab' fragment, a F(ab')2 fragment, a bispecific Fab dimer (Fab2), a trispecific Fab trimer (Fab3), an Fv, a single chain Fv protein (scFv), a bis-scFv, (scFv)2, a minibody, a diabody, a triabody, a tetrabody, a disulfide stabilized Fv protein (“dsFv”), a Fibronectin type III (FN3) domain antibody, a single-domain antibody (sdAb, a camelid VHH, Nanobody), and a centyrin.

[0045] In certain embodiments, the antibody or antigen binding fragment thereof binds an antigen expressed on an immune effector cell selected from the group consisting of: the alpha, beta, gamma, or delta chain of the T cell receptor, CD2, CD35, CD3s CD3y, CD4, CD8a, and CD8p.

[0046] In particular embodiments, the antibody or antigen binding fragment thereof comprises (a) a heavy chain variable region (VH) comprising a CDRH1, a CDRH2, and a CDRH3 of an antibody or antigen binding fragment thereof set forth in Table 5; (b) a polypeptide linker; and (c) a light chain variable region (VL) comprising a CDRL1, a CDRL2, and a CDRL3 of an antibody or antigen binding fragment thereof set forth in Table 5.

[0047] In additional embodiments, the antibody or antigen binding fragment thereof comprises (a) a heavy chain variable region (VH) of an antibody or antigen binding fragment thereof set forth in Table 5; (b) a polypeptide linker; and (c) a light chain variable region (VL) of an antibody or antigen binding fragment thereof set forth in Table 5.

[0048] In some embodiments, the antibody or antigen binding fragment thereof comprises: (a) a heavy chain variable region (VH) of an antibody or antigen binding fragment thereof set forth in Table 5; (b) a polypeptide linker selected from the group consisting of: (GGGGS)n wherein n = 1, 2, 3, 4 or 5; S(GGGGS)n wherein n = 1, 2, 3, 4 or 5; GEGTSTGSGGSGGSGGAD, GSTSGSGKPGSGEGSTKG and variants thereof comprising an amino acid sequence 95% identical thereto; and (c) the corresponding light chain variable region (VL) of the antibody or antigen binding fragment thereof set forth in Table 5.

[0049] In particular embodiments, the antibody or antigen binding fragment thereof comprises the amino acid sequence set forth in any one of SEQ ID NOs: 19, 20, 29, 30, 39,

[0050] 40, 49, 50, 59, 60, 69, 70, 79, 80, 89, 90, 99, 100, 109, 110, 119, 120, 129, 130, 139, 140,

[0051] 149, 150, 159, 160, 169, 170, 179, 180, 189, 190, 199, 200, 209, 210, 219, 220, 229, 230,

[0052] 239, 240, 249, 250, 259, 260, 269, 270, 279, 280, 289, 290, 299, 300, 309, 310, 319, 320,

[0053] 329, 330, 339, 340, 349, 350, 359, 360, 369, 370, 379, 380, 389, 390, 399, 400, 409, and 410.

[0054] In particular embodiments, the antibody or antigen binding fragment thereof comprises the amino acid sequence set forth in any one of SEQ ID NOs: 99, 100, 109, 110, 119, 120, 129, 130, 139, 140, 149, 150, 159, 160, 169, and 170.

[0055] In further embodiments, the antibody or antigen binding fragment thereof comprises the amino acid sequence set forth in any one of SEQ ID NOs: 99 or 100, 109 or 110, 119 or 120, 129 or 130, 139 or 140, 149 or 150, 159 or 160, and 169 or 170. In a preferred embodiment, the antibody or antigen binding fragment thereof comprises the amino acid sequence set forth in SEQ ID NOs: 99.

[0056] In some embodiments, the spacer comprises an amino acid sequence at least 85% identical to, at least 90% identical to, at least 95% identical to, at least 96% identical to, at least 97% identical to, at least 98% identical to, at least 99% identical to, or identical to the amino acid sequence set forth in any one of SEQ ID NOs: 411-434.

[0057] In certain embodiments, the spacer comprises an amino acid sequence at least 85% identical to, at least 90% identical to, at least 95% identical to, at least 96% identical to, at least 97% identical to, at least 98% identical to, at least 99% identical to, or identical to the amino acid sequence set forth in any one of SEQ ID NOs: 411-413, SEQ ID NOs: 414-423, SEQ ID NOs: 424-427, or SEQ ID NOs: 428-434.

[0058] In particular embodiments, the transmembrane domain comprises an amino acid sequence at least 85% identical to, at least 90% identical to, at least 95% identical to, at least 96% identical to, at least 97% identical to, at least 98% identical to, at least 99% identical to, or identical to the amino acid sequence set forth in any one of SEQ ID NOs: 435, 436, 437, 438, 439, 440 or 441.

[0059] In some embodiments, the intracellular domain comprises an amino acid sequence at least 85% identical to, at least 90% identical to, at least 95% identical to, at least 96% identical to, at least 97% identical to, at least 98% identical to, at least 99% identical to, or identical to the amino acid sequence set forth in any one of SEQ ID NOs: 442, 443, 444, 445, 446, 447 or 448.

[0060] In particular embodiments, the non-viral membrane-bound tropism polypeptide comprises an amino acid sequence at least 85% identical to, at least 90% identical to, at least 95% identical to, at least 96% identical to, at least 97% identical to, at least 98% identical to, at least 99% identical to, or identical to the amino acid sequence set forth in any one of SEQ ID NOs: 455-576. In particular embodiments, the tropism polypeptide comprises an amino acid sequence set forth in any one of SEQ ID NOs: 461.

[0061] In additional embodiments, the non-viral membrane-bound tropism polypeptide comprises an amino acid sequence at least 85% identical to, at least 90% identical to, at least 95% identical to, at least 96% identical to, at least 97% identical to, at least 98% identical to, at least 99% identical to, or identical to the amino acid sequence set forth in any one of SEQ ID NOs: 471-486. In particular embodiments, the tropism polypeptide comprises an amino acid sequence set forth in any one of SEQ ID NOs: 471 or 483.

[0062] In certain embodiments, the non-viral membrane-bound tropism polypeptide comprises an amino acid sequence at least 85% identical to, at least 90% identical to, at least 95% identical to, at least 96% identical to, at least 97% identical to, at least 98% identical to, at least 99% identical to, or identical to the amino acid sequence set forth in any one of SEQ ID NOs: 535-554. In particular embodiments, the tropism polypeptide comprises an amino acid sequence set forth in any one of SEQ ID NOs: 537 or 553.

[0063] In particular embodiments, the non-viral membrane-bound tropism polypeptide comprises an amino acid sequence at least 85% identical to, at least 90% identical to, at least 95% identical to, at least 96% identical to, at least 97% identical to, at least 98% identical to, at least 99% identical to, or identical to the amino acid sequence set forth in any one of SEQ ID NOs: 555-562.

[0064] In additional embodiments, the non-viral membrane-bound tropism polypeptide comprises an amino acid sequence at least 85% identical to, at least 90% identical to, at least 95% identical to, at least 96% identical to, at least 97% identical to, at least 98% identical to, at least 99% identical to, or identical to the amino acid sequence set forth in any one of SEQ ID NOs: 563 or 564.

[0065] In additional embodiments, the non-viral membrane-bound tropism polypeptide comprises an amino acid sequence at least 85% identical to, at least 90% identical to, at least 95% identical to, at least 96% identical to, at least 97% identical to, at least 98% identical to, at least 99% identical to, or identical to the amino acid sequence set forth in any one of SEQ ID NOs: 565-576.

[0066] In additional embodiments, the non-viral membrane-bound tropism polypeptide comprises an amino acid sequence at least 85% identical to, at least 90% identical to, at least 95% identical to, at least 96% identical to, at least 97% identical to, at least 98% identical to, at least 99% identical to, or identical to the amino acid sequence set forth in SEQ ID NO: 471 or SEQ ID NO: 553.

[0067] In certain embodiments, the particle further comprises one or more secondary non- viral membrane bound tropism polypeptides.

[0068] In further embodiments, the one or more secondary non-viral membrane bound tropism polypeptides comprise one or more of CD80, CD86, CD137L, OX40L, and ICOSL or variants thereof.

[0069] In particular embodiments, the particle further comprises one or more secondary non- viral membrane-bound tropism polypeptides at least 85% identical to, at least 90% identical to, at least 95% identical to, at least 96% identical to, at least 97% identical to, at least 98% identical to, at least 99% identical to, or identical to the amino acid sequence set forth in any one of SEQ ID NOs: 577-581. In particular embodiments, the one or more secondary non- viral membrane-bound tropism polypeptide comprises an amino acid sequence set forth in any one of SEQ ID NOs: 577.

[0070] In some embodiments, the recombinant lend viral vector is engineered or derived from a lentiviral genome of a lentivirus selected from the group consisting of: human immunodeficiency virus 1 (HIV-1); human immunodeficiency vims 2 (HIV-2); visna-maedi vims (VMV); caprine arthritis-encephalitis vims (CAEV); equine infectious anemia vims (EIAV); feline immunodeficiency vims (FIV); bovine immune deficiency vims (BIV); and simian immunodeficiency vims (SIV).

[0071] In additional embodiments, the recombinant lentiviral vector further comprises one or more polynucleotides encoding one or more of the following: a signal peptide, a post- transcriptional regulatory element, an insulator element, a selectable marker, and a cell suicide gene.

[0072] In certain embodiments, the post-transcriptional regulatory element is a woodchuck hepatitis vims post-transcriptional regulatory element (WPRE) or a hepatitis B vims post- transcriptional regulatory element (HPRE). In particular embodiments, the promoter is selected from the group consisting of: a P-actin promoter, a cytomegalovirus (CMV) immediate early promoter, a simian vims 40 (SV40) (e.g. , early or late) promoter, a Moloney murine leukemia vims (MoMLV) promoter, a Rous sarcoma vims (RSV) promoter, a herpes simplex vims (HSV) (thymidine kinase) promoter, an SV40 / CD43 promoter, a spleen focus forming vims (SFFV) promoter, an elongation factor 1-alpha (EFla) short promoter (intronless), an EFla long promoter containing an intron, a Ubiquitin C (UBC) promoter, a phosphoglycerate kinase- 1 (PGK) promoter, a cytomegalovirus enhancer / chicken P-actin (CAG) promoter, and a myeloproliferative sarcoma vims enhancer, negative control region deleted, dl587rev primerbinding site substituted (MND) U3 promoter.

[0073] In further embodiments, the one or more engineered receptors are selected from the group consisting of: a chimeric antigen receptor (CAR), a chimeric costimulatory receptor (CCR), an alpha-beta T cell receptor (aP TCR), a gamma delta T cell receptor (y5 TCR), a dimerizing agent regulated immunoreceptor complex (DARIC), a chimeric TGF-P receptor (CTBR), and a zetakine receptor.

[0074] In certain embodiments, the one or more engineered receptors bind an antigen selected from the group consisting of: alpha folate receptor (FRa), avP6 integrin, BAFFR, 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), claudin 6, (CLDN6), claudin 18 isoform 2 (CLDN18.2), 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), delta like canonical Notch ligand 3 (DLL3), epidermal growth factor receptor (EGFR), epidermal growth factor receptor variant III (EGFRvIII), epithelial glycoprotein 2 (EGP2), epithelial glycoprotein 40 (EGP40), epithelial cell adhesion molecule (EPCAM), ephrin type-A receptor 2 (EPHA2), erb-b2 receptor tyrosine kinase 4 (ERBB4), fibroblast activation protein (FAP), Fc Receptor Like 5 (FCRL5), fetal acetylcholinesterase receptor (AchR), ganglioside G2 (GD2), ganglioside G3 (GD3), Glypican-3 (GPC3), G Protein-Coupled Receptor Class C Group 5 Member D (GPCR5D), EGFR family including ErbB2 (HER2), HER2 p95, IL-lORa, IL-13Ra2, Kappa, cancer / testis antigen 2 (LAGE- 1 A), Lambda, Lewis-Y (LeY), LI cell adhesion molecule (Ll-CAM), melanoma antigen gene (MAGE)-Al, MAGE-A3, MAGE-A4, MAGE-A6, MAGEA10, melanoma antigen recognized by T cells 1 (MelanA or MARTI), Mesothelin (MSLN), MUC1, MUC16, MHC class I chain related proteins A (MICA), MHC class I chain related proteins B (MICB), neural cell adhesion molecule (NCAM), cancer / testis antigen 1 (NY-ESO-1), placenta- specific 1 (PLAC1), preferentially expressed antigen in melanoma (PRAME), prostate stem cell antigen (PSCA), prostate-specific membrane antigen (PSMA), receptor tyrosine kinase-like orphan receptor 1 (R0R1), SLAMF7, synovial sarcoma, X breakpoint 2 (SSX2), Survivin, TACI, tumor associated glycoprotein 72 (TAG72), tumor endothelial marker 1 (TEM1 / CD248), tumor endothelial marker 7- related (TEM7R), transforming growth factor beta 1 (TFGpi), 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).

[0075] In some embodiments, the one or more engineered receptors bind an antigen selected from the group consisting of: BCMA, CD19, CD20, CD22, CD33, CD38, CD79A, CD79B, GPCR5D, MAGE-A4, MSLN, MUC16, NY-ESO-1, PSCA, PSMA and R0R1.

[0076] In particular embodiments, the one or more engineered receptors bind an antigen selected from the group consisting of: BCMA, CD19, CD20, CD22, CD38, CD79A, CD79B, and GPCR5D.

[0077] In additional embodiments, the one or more engineered receptors bind an antigen selected from the group consisting of: BCMA, CD38, and GPCR5D.

[0078] In particular embodiments, the one or more engineered receptors bind an antigen selected from the group consisting of: CD 19, CD20, CD22, CD79A, and CD79B.

[0079] In certain embodiments, the one or more engineered receptors comprises an extracellular antigen binding domain, a hinge domain, a transmembrane domain, and one or more intracellular signaling domains.

[0080] In some embodiments, the one or more engineered antigen receptors comprises an extracellular antigen binding domain selected from the group consisting of: a receptor ectodomain, a ligand, or an antibody or antigen binding fragment thereof selected from the group consisting of: a Camel Ig, a Llama Ig, an Alpaca Ig, Ig NAR, a Fab' fragment, a F(ab')2 fragment, a bispecific Fab dimer (Fab2), a trispecific Fab trimer (Fab3), an Fv, a single chain Fv protein (scFv), a bis-scFv, (scFv)2, a minibody, a diabody, a triabody, a tetrabody, a disulfide stabilized Fv protein (“dsFv”), a Fibronectin type III (FN3) domain antibody, a single-domain antibody (sdAb, a camelid VHH, Nanobody), and a centyrin. In further embodiments, the CAR comprises a hinge domain isolated or derived from a polypeptide selected from the group consisting of: CD4, CD8P, CD8a, CD28, CD134, CD137, CD152, CD278, IgGl, IgG2, IgG3, and IgG4.

[0081] In certain embodiments, the CAR comprises a transmembrane domain isolated or derived from a polypeptide selected from the group consisting of: alpha, beta, gamma, or delta chain of the T-cell receptor, CD35, CD3s, CD3y, CD3^, CD4, CD5, CD8a, CD9, CD16, CD22, CD27, CD28, CD33, CD37, CD45, CD64, CD80, CD86, CD134, CD137, CD152, CD154, CD278, amnionless (AMN), and programmed cell death 1 (PDCD1).

[0082] In particular embodiments, the CAR comprises a primary signaling domain isolated or derived from a polypeptide selected from the group consisting of: FcRy, FcRp, CD3y, CD35, CD3s, CD3 , CD22, CD79A, CD79B, and CD66d.

[0083] In some embodiments, the CAR comprises one or more costimulatory domains isolated or derived from a polypeptide 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 (CARD 11), CD2, CD7, CD27, CD28, CD30, CD40, CD54 (ICAM), CD83, CD94, CD134 (0X40), CD137 (4-1BB), CD278 (ICOS), DNAX-Activation Protein 10 (DAP10), Linker for activation of T-cells family member 1 (LAT), SH2 Domain-Containing Leukocyte Protein Of 76 kD (SLP76), T cell receptor associated transmembrane adaptor 1 (TRAT1), TNFR2, TNFRS14, TNFRS18, TNFRS25, and zeta chain of T cell receptor associated protein kinase 70 (ZAP70).

[0084] In particular embodiments, the CAR comprises an amino acid sequence set forth in any one of SEQ ID NOs: 662, 663, 664, 665, or 666.

[0085] In certain embodiments, the one or more engineered receptors further comprises a CCR comprising an extracellular antigen binding domain that binds an antigen different than the antigen bound by the CAR, a hinge domain, a transmembrane domain, and one or more intracellular signaling domains.

[0086] In particular embodiments, the one or more engineered receptors comprises a DARIC comprising a DARIC signaling component comprising a multimerization domain, a transmembrane domain and one or more intracellular signaling domains; and a DARIC binding component comprising an extracellular antigen binding domain, an extracellular antigen binding domain, a transmembrane domain, and optionally one or more intracellular signaling domains. In further embodiments, the one or more engineered receptors comprises a CTBR comprising (a) a first polypeptide comprising an extracellular TGFpi -binding domain of TGFPR2, a transmembrane domain, and a first interleukin receptor intracellular signaling domain; (b) a polypeptide cleavage signal; and (c) a second polypeptide comprising an extracellular TGFpi -binding domain of TGFpRl, a transmembrane domain, and a second interleukin receptor intracellular signaling domain.

[0087] In various embodiments, a recombinant particle is contemplated, comprising: (a) a surface comprising (i) one or more mutated viral envelope glycoproteins that retain fusogenic activity and that lack cognate receptor binding activity, wherein the one or more mutated viral envelope glycoproteins comprises an amino acid sequence at least 95%, at least 96%, at least 97%, at least 98%, at least 99% identical to or identical to any one of SEQ ID NOs: 630, 634, 638, and 642 and (ii) a non-viral membrane-bound tropism polypeptide comprising an antibody or antigen binding fragment thereof comprising an amino acid sequence set forth in any one of SEQ ID NOs: 91 and 95, and 99 or 100, that binds an antigen expressed on an immune effector cell, a spacer polypeptide, and a GPI anchor domain or a transmembrane domain, and optionally, an intracellular domain comprising no more than 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 amino acids; and (b) a therapeutic payload encoding an engineered antigen receptor.

[0088] In various embodiments, a recombinant particle is contemplated, comprising: (a) a surface comprising (i) a mutated VSIV-G envelope glycoprotein that comprises an amino acid sequence at least 95%, at least 96%, at least 97%, at least 98%, at least 99% identical to or identical to SEQ ID NO: 642 and (ii) a non-viral membrane-bound tropism polypeptide comprising an antibody or antigen binding fragment thereof comprising an amino acid sequence set forth in SEQ ID NO: 99 or 100, that binds an antigen expressed on an immune effector cell, a spacer polypeptide, and a GPI anchor domain or a transmembrane domain, and optionally, an intracellular domain comprising no more than 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 amino acids; and (b) a lentiviral vector encoding an engineered antigen receptor.

[0089] In various embodiments, a recombinant lentiviral particle is contemplated, comprising: (a) a viral envelope comprising (i) a mutated VSIV-G envelope glycoprotein that comprises an amino acid sequence at least 95%, at least 96%, at least 97%, at least 98%, at least 99% identical to or identical to SEQ ID NO: 642 and (ii) a non-viral membrane-bound tropism polypeptide comprising an amino acid sequence set forth in any one of SEQ ID NOs: 99 or 100, that binds an antigen expressed on an immune effector cell, a spacer polypeptide, and a GPI anchor domain or a transmembrane domain, and optionally, an intracellular domain comprising no more than 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 amino acids; and (b) a therapeutic payload encoding an engineered antigen receptor.

[0090] In various embodiments, a recombinant lentiviral particle is contemplated, comprising: (a) a viral envelope comprising (i) a mutated VSIV-G envelope glycoprotein that comprises an amino acid sequence at least 95%, at least 96%, at least 97%, at least 98%, at least 99% identical to or identical to SEQ ID NO: 642; (ii) a non-viral membranebound tropism polypeptide comprising an amino acid sequence set forth in any one of SEQ ID NOs: 461, 471, 483, 537, and 553; and optionally (iii) a secondary non-viral membranebound tropism polypeptide comprising an amino acid sequence set forth in SEQ ID NO: 577; and (b) a lentiviral vector encoding a chimeric antigen receptor.

[0091] In various embodiments, a recombinant lentiviral particle is contemplated, comprising: (a) a viral envelope comprising (i) a mutated VSIV-G envelope glycoprotein that comprises an amino acid sequence at least 95%, at least 96%, at least 97%, at least 98%, at least 99% identical to or identical to SEQ ID NO: 642; (ii) a non-viral membranebound tropism polypeptide that comprises an amino acid sequence at least 95%, at least 96%, at least 97%, at least 98%, at least 99% identical to or identical to SEQ ID NO: 471; and optionally (iii) a secondary non-viral membrane-bound tropism polypeptide that comprises an amino acid sequence at least 95%, at least 96%, at least 97%, at least 98%, at least 99% identical to or identical to SEQ ID NO: 577; and (b) a lentiviral vector encoding a chimeric antigen receptor comprising an amino acid sequence set forth in any one of SEQ ID NOs: 662-666.

[0092] In various embodiments, a recombinant lentiviral particle is contemplated, comprising: (a) a viral envelope comprising (i) a mutated VSIV-G envelope glycoprotein that comprises an amino acid sequence at least 95%, at least 96%, at least 97%, at least 98%, at least 99% identical to or identical to SEQ ID NO: 642; (ii) a non-viral membrane-bound tropism polypeptide that comprises an amino acid sequence at least 95%, at least 96%, at least 97%, at least 98%, at least 99% identical to or identical to SEQ ID NO: 471; and optionally (iii) a secondary non-viral membrane-bound tropism polypeptide that comprises an amino acid sequence at least 95%, at least 96%, at least 97%, at least 98%, at least 99% identical to or identical to SEQ ID NO: 577; and (b) a lentiviral vector encoding a chimeric antigen receptor that binds a polypeptide selected from any one of BCM A, CD 19, CD20, CD22, CD38, CD79A, CD79B, and GPCR5D. In various embodiments, a recombinant lentiviral particle is contemplated, comprising: (a) a viral envelope comprising (i) a mutated VSIV-G envelope glycoprotein that comprises an amino acid sequence at least 95%, at least 96%, at least 97%, at least 98%, at least 99% identical to or identical to SEQ ID NO: 642; (ii) a non-viral membrane-bound tropism polypeptide that comprises an amino acid sequence at least 95%, at least 96%, at least 97%, at least 98%, at least 99% identical to or identical to SEQ ID NO: 471; and optionally (iii) a secondary non-viral membrane-bound tropism polypeptide that comprises an amino acid sequence at least 95%, at least 96%, at least 97%, at least 98%, at least 99% identical to or identical to SEQ ID NO: 577; and (b) a lentiviral vector encoding a chimeric antigen receptor that binds BCMA.

[0093] In various embodiments, a recombinant lentiviral particle is contemplated, comprising: (a) a viral envelope comprising (i) a mutated VSIV-G envelope glycoprotein that comprises an amino acid sequence at least 95%, at least 96%, at least 97%, at least 98%, at least 99% identical to or identical to SEQ ID NO: 642; (ii) a non-viral membrane-bound tropism polypeptide that comprises an amino acid sequence at least 95%, at least 96%, at least 97%, at least 98%, at least 99% identical to or identical to SEQ ID NO: 471; and optionally (iii) a secondary non-viral membrane-bound tropism polypeptide that comprises an amino acid sequence at least 95%, at least 96%, at least 97%, at least 98%, at least 99% identical to or identical to SEQ ID NO: 577; and (b) a lentiviral vector encoding a chimeric antigen receptor that binds GPCR5D.

[0094] In various embodiments, a recombinant lentiviral particle is contemplated, comprising: (a) a viral envelope comprising (i) a mutated VSIV-G envelope glycoprotein that comprises an amino acid sequence at least 95%, at least 96%, at least 97%, at least 98%, at least 99% identical to or identical to SEQ ID NO: 642; (ii) a non-viral membrane-bound tropism polypeptide that comprises an amino acid sequence at least 95%, at least 96%, at least 97%, at least 98%, at least 99% identical to or identical to SEQ ID NO: 471; and optionally (iii) a secondary non-viral membrane-bound tropism polypeptide that comprises an amino acid sequence at least 95%, at least 96%, at least 97%, at least 98%, at least 99% identical to or identical to SEQ ID NO: 577; and (b) a lentiviral vector encoding a chimeric antigen receptor that binds CD 19.

[0095] In additional embodiments, a cell transduced with a particle contemplated herein.

[0096] In particular embodiments, the cell is an immune effector cell. In some embodiments, the cell is a T cell, a natural killer (NK) cell, or a natural killer T (NKT) cell.

[0097] In certain embodiments, a composition comprising a particle or a cell contemplated herein.

[0098] In particular embodiments, a pharmaceutical composition comprises a pharmaceutically acceptable carrier and a particle, a cell, or a composition contemplated herein.

[0099] In some embodiments, a method of treating, preventing, or ameliorating at least one symptom of a disease, disorder or condition associated therewith in a subject comprises administering to the subject an effective amount of a particle, a cell, a composition, or a pharmaceutical composition contemplated herein.

[0100] In further embodiments, the disease, disorder, or condition is a cancer.

[0101] In particular embodiments, the cancer is leukemia selected from the group consisting of: acute lymphocytic leukemia (ALL), an acute myeloid leukemia (AML), a myelodysplastic syndrome (MDS), a plasma cell leukemia (PCL), erythroleukemia, a hairy cell leukemia (HCL), chronic lymphocytic leukemia (CLL), chronic myeloid leukemia (CML), chronic myelomonocytic leukemia (CMML) and polycythemia vera.

[0102] In additional embodiments, the cancer is non-NHL or NHL selected from the group consisting of: diffuse large B cell lymphoma (DLBCL), follicular lymphoma (FL), mantlecell lymphoma (MCL), small lymphocytic lymphoma (SLL), primary mediastinal large B- cell lymphoma, a marginal zone B cell lymphoma (MZL), mucosa-associated lymphatic tissue lymphoma (MALT), Burkitt’s lymphoma (BL), immunoblastic large cell lymphoma, centroblastic large cell lymphoma, anaplastic B-cell lymphoma, mycosis fungoides, Sezary syndrome, T-lymphoblastic lymphoma, and anaplastic large-cell lymphoma (ALCL).

[0103] In certain embodiments, the cancer is MM selected from the group consisting of: active multiple myeloma, smoldering multiple myeloma, light chain myeloma, non- secretory myeloma, IgD myeloma, IgE myeloma, osteosclerotic myeloma, solitary plasmacytoma of bone, and extramedullary plasmacytoma.

[0104] In some embodiments, the cancer is relapsed and / or refractory.

[0105] In additional embodiments, the disease, disorder, or condition is an autoimmune disease. In certain embodiments, the autoimmune disease is systemic lupus erythematosus, rheumatoid arthritis, idiopathic thrombocytopenic purpura, myasthenia gravis, or autoimmune hemolytic anemia.

[0106] In some embodiments, a method of transducing an immune effector cell in vivo, comprises administering to a subject a pharmaceutical composition comprising a pharmaceutically acceptable carrier and an effective amount of a particle contemplated herein.

[0107] In particular embodiments, a method of making the recombinant lend virus comprises (a) transfecting a host cell with four polynucleotides: a first polynucleotide that encodes lentiviral gag-pol, a second polynucleotide that encodes lentiviral rev, a third polynucleotide that encodes one or more mutated viral envelope glycoproteins that retain fusogenic activity and lack cognate receptor binding activity and one or more non-viral membrane-bound tropism polypeptides, and a fourth polynucleotide that is a transfer plasmid encoding a recombinant lentiviral vector of any one of the preceding embodiments; and b) culturing the transduced cell for about 1 to 3 days to produce the recombinant lentivirus.

[0108] In certain embodiments, a kit comprises a particle contemplated herein, a pharmaceutically acceptable carrier, and instructions for use.

[0109] BRIEF DESCRIPTION OF SEVERAL VIEWS OF THE DRAWINGS

[0110] Figure 1A shows GFP expression (transduction) and CD69 expression (activation) in Jurkat cells targeted with lentiviral particles that have a viral envelope expressing tropism molecules that have T cell specific binding domains and a mutant fusogen.

[0111] Figure IB shows flow cytometry analysis of GFP expression (transduction) and CD69 expression (activation) in PBMCs cells targeted with lentiviral particles that have a viral envelope expressing tropism molecules that have T cell specific binding domains and a mutant fusogen.

[0112] Figure 2 shows flow cytometry analysis of GFP expression or CAR expression in HEK293T used to produce lentiviral particles that have a viral envelope expressing (i) tropism molecules that have T cell specific binding domains, a monomeric or dimeric CD8a stalk and a CD8a TM domain and an intracellular domain (ii) a mutant fusogen.

[0113] Figure 3A shows a diagram of various tropism molecule stalk architectures experimentally examined in the Examples. Figure 3B shows tropism molecule expression (measured by anti-CD3 scFv expression) in HEK293T cells used to produce lend viral particles that have a viral envelope expressing (i) anti-CD3-based tropism molecules and (ii) a mutant fusogen.

[0114] Figure 3C shows the transducing units (TU) per nanogram of P24 for lentiviral particles that have anti-CD3-based tropism molecules with various stalk architectures and a mutated fusogen used to transduce Jurkat cells.

[0115] Figure 3D shows the transduction specificity for lentiviral particles that express anti- CD3-based tropism molecules with various stalk architectures and a mutated fusogen. Left panel compares on-target transduction in PBMCs to off-target transduction in A549 cells. Right panel compares on-target transduction in Jurkat cells to off-target transduction in A549 cells.

[0116] Figure 4 shows the effects of anti-CD3-based tropism molecule stalk architecture on titer and specificity. Left panel shows the effect of estimated tropism molecule stalk length on titer (TU / ng P24). Right panel shows the effect of estimated tropism molecule stalk length on transduction specificity.

[0117] Figure 5A shows the GFP expression (transduction, top panel) and cell count (activation, bottom panel) in PBMCs transduced with lentiviral particles that express anti- CD3-based tropism molecules that have a variety of stalk architectures and a mutant fusogen.

[0118] Figure 5B shows the GFP expression (transduction, left panel) in PBMCs transduced with 0.5 ng P24 or 2 ng P24 of lentiviral particles that express anti-CD3-based tropism molecules that have a variety of stalk architectures and a mutant fusogen. Right panel shows the effect of tropism molecule stalk architecture on PBMC transduction compared to Jurkat cell transduction.

[0119] Figure 6 shows transduction (top panel) and activation (bottom panel) human T cells in an in vivo mouse model. Mice were administered lentiviral particles expressing T cell targeting tropism molecules, a mutant fusogen and a lentiviral vector encoding GFP.

[0120] Figure 7A shows the number (left panel) and percentage (right panel) of GFP positive T cells in the blood of mice administered 1 x 107TU, 2.5 x 107TU, 1 x 108TU, or 2.5 x 108TU of lentiviral particles that express (i) T cell targeting tropism molecules (LV 2) or lack T cell targeting tropism molecules (LV 26), (ii) a mutant fusogen and (iii) a lentiviral vector encoding GFP.

[0121] Figure 7B shows immunohistochemistry staining in the heart, brain, ovaries, and small intestines of mice administered 1 x 107TU, 2.5 x 107TU, 1 x 108TU, or 2.5 x 108TU of lentiviral particles that express (i) T cell targeting tropism molecules (LV 2) or lack T cell targeting tropism molecules (LV 26), (ii) a mutant fusogen and (iii) a lentiviral vector encoding GFP. The rightmost column shows flow cytometry data of GFP positive T cells in the spleen of these mice at each dose level.

[0122] Figure 8 shows that lentiviral particles expressing an anti-CD3-based tropism molecule, a mutant fusogen, and a lentiviral vector encoding an anti-BCMA CAR reduce tumor growth comparable to ex vivo manufactured anti-BCMA CAR T cells in an RPMI multiple myeloma mouse model.

[0123] Figure 9 shows that lentiviral particles expressing either of two different anti-CD3- based tropism molecules, and a mutant fusogen and a lentiviral vector encoding an anti- BCMA CAR reduce tumor growth comparable to ex vivo manufactured anti-BCMA CAR T cells in an Daudi multiple myeloma mouse model.

[0124] BRIEF DESCRIPTION OF THE SEQUENCE IDENTIFIERS

[0125] SEQ ID NOs: 1-10 set forth amino acid sequences of fusogens.

[0126] SEQ ID NOs: 11-410 set forth amino acid sequences of antibodies.

[0127] SEQ ID NOs: 411-434 set forth amino acid sequences of spacer domains.

[0128] SEQ ID NOs: 435-441 set forth amino acid sequences of transmembrane domains.

[0129] SEQ ID NOs: 442-448 set forth amino acid sequences of intracellular domains.

[0130] SEQ ID NOs: 449-454 set forth amino acid sequences of signal peptides.

[0131] SEQ ID NOs: 455-576 set forth amino acid sequences of non- viral membrane bound tropism polypeptides.

[0132] SEQ ID NOs: 577-581 set forth amino acid sequences of secondary tropism polypeptides.

[0133] SEQ ID NOs: 582-661 set forth amino acid sequences of fusogens.

[0134] SEQ ID NOs: 662-666 set forth amino acid sequences of engineered receptors.

[0135] SEQ ID NOs: 667-672 set forth nucleic acid sequences of promoters.

[0136] SEQ ID NOs: 673-693 set forth amino acid sequences of polypeptide linkers.

[0137] SEQ ID NOs: 694-713 set forth amino acid sequences of viral self-cleaving peptides.

[0138] SEQ ID NO: 714 sets forth an amino acid sequence of a lentivirus protease cleavage site. In the foregoing sequences, X, if present, refers to any amino acid, a specified group of amino acids or the absence of an amino acid.

[0139] Throughout the disclosure, the amino acid positions of a fusogen is with reference to the fusogen lacking a signal sequence (i.e., the amino acid sequence after the signal peptide has been cleaved).

[0140] DETAILED DESCRIPTION

[0141] A. OVERVIEW

[0142] The field of ex vivo gene therapy is not new and has been evolving for decades. Despite huge potential, ex vivo gene therapy has been met with limited success. Moreover, substantial obstacles still plague the field of ex vivo gene therapy, obstacles including limited precision and lack of commercial viability are likely among the reasons that it has yet to see widespread adoption in a clinical setting. In vivo gene therapy however, is new and offers the potential to deliver life-altering therapies on an unprecedented scale. In vivo gene therapy solves the commercial viability issues associated with the astronomical costs associated with manufacturing ex vivo gene therapies. But in vivo gene therapy comes with its own set of challenges, including potential off-target toxicity, low efficacy, and immunogenicity.

[0143] The present disclosure offers solutions to foregoing challenges and others that exist in the field of in vivo gene therapy.

[0144] The present disclosure generally relates to, in part, to an engineered cell-targeting particle (e.g., a fusosome; an extracellular vesicle, including a microvesicle, an apoptotic body, and an exosome; a lipid nanoparticle; a virus-like particle (VLPs); or a viral particle) that has improved specificity for a desired cell type, e.g., an immune effector cell. Without wishing to be bound by any particular theory, it is contemplated that an engineered celltargeting particle comprising an efficient fusogen and one or more cell-targeting polypeptides i.e., tropism polypeptides) increase the on-target delivery of a cargo or pay load to a desired cell type and decrease the off-target delivery to undesired cell types. It is further contemplated that particles comprising one or more non- viral tropism polypeptides and a viral envelope glycoprotein that is capable of cell fusion but not cell attachment or cell binding yields an engineered cell-targeting particle that has increased delivery specificity and that efficiently delivers one or more nucleic acids or proteins to a desired cell type. The disclosure contemplates, in part, recombinant particles with increased specificity for cellular vehicles to enable gene therapy or genome editing applications. Recombinant particles contemplated herein include but are not limited to, a fusosome, an extracellular vesicle, a microvesicle, an apoptotic body, an exosome, a lipid nanoparticle, a virus-like particle (VLPs), or a recombinant viral particle that has a single layer of phospholipid, a phospholipid bilayer, a cell membrane, a capsid, or a viral envelope comprising one or more membrane-bound viral glycoproteins modified to retain fusogenic activity and decrease, reduce, substantially ablate, ablate, abolish or eliminate cell binding or attachment activity; one or more non-viral membrane-bound tropism polypeptides; and a cargo or payload comprising one or more polynucleotides and / or polypeptides. The recombinant particles may be used for ex vivo gene therapy and genome editing but provide substantial advantages for use in in vivo gene therapy and genome editing.

[0145] The disclosure further contemplates, in part, gene therapy and genome editing compositions comprising a fusosome, an extracellular vesicle, a microvesicle, an apoptotic body, an exosome, a lipid nanoparticle, a virus-like particle (VLPs), or a recombinant viral particle that has a single layer of phospholipid, a phospholipid bilayer, a cell membrane, a capsid, or a viral envelope comprising one or more membrane-bound viral glycoproteins modified to retain fusogenic activity and that lack or substantially lack cell binding or attachment activity and / or cognate receptor binding activity; one or more non-viral membrane-bound tropism polypeptides; and a cargo or pay load comprising one or more polynucleotides and / or polypeptides.

[0146] The disclosure further contemplates, in part, methods of making the recombinant particles contemplated herein, along with methods of using the particles.

[0147] In preferred embodiments, the recombinant particle is a recombinant viral particle, preferably a recombinant retroviral particle, more preferably a recombinant lentiviral particle, and even more preferably, a recombinant HIV - 1 based lentiviral particle. In further preferred embodiments, the recombinant particle comprises a polynucleotide pay load, preferably a vector, more preferably a viral vector, and even more preferably a retroviral or lentiviral vector.

[0148] In particular embodiments, the disclosure contemplates, methods of using a recombinant particle contemplated herein to decrease off-target delivery and increase targeted delivery to an immune effector cell in vivo, to increase the specificity of an in vivo gene therapy or genome editing composition, to deliver a cargo or payload of one or more polynucleotides and / or polypeptides to an immune effector cell, to treat a wide variety of disorders, diseases and conditions or symptoms associated therewith, including but not limited to cancers, infectious diseases, autoimmune diseases, inflammatory diseases, immunodeficiencies, and monogeneic diseases.

[0149] Compositions, pharmaceutical compositions, and kits comprising one or more recombinant particles contemplated herein and methods of making and using the same are also provided in particular embodiments.

[0150] Techniques for recombinant (z.e., engineered) DNA, peptide and oligonucleotide synthesis, immunoassays, tissue culture, transformation (e.g., electroporation, lipofection), enzymatic reactions, purification and related techniques and procedures may be generally performed as described in various general and more specific references in microbiology, molecular biology, biochemistry, molecular genetics, cell biology, virology and immunology as cited and discussed throughout the present specification. See, e.g., Sambrook el al., Molecular Cloning: A Laboratory Manual, 4th ed., Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y.; Current Protocols in Molecular Biology (John 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 (2002); Glover, DNA Cloning: A Practical Approach, vol. I & II (IRL Press, Oxford Univ. Press USA, 1985); Current Protocols in Immunology (Edited by: John E. Coligan, Ada M. Kruisbeek, David H. Margulies, Ethan M. Shevach, Warren Strober 2001 John Wiley & Sons, NY, NY); Real-Time PCR: Current Technology and Applications, Edited by 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 the Hybridization (B. Hames & S. Higgins, Eds., 1985); Transcription and Translation (B. Hames & S. Higgins, Eds., 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 Edition, 2010 Humana Press); Immobilized Cells and Enzymes (IRL Press, 1986); the treatise, Methods in Enzymology (Academic Press, Inc., N.Y.); Gene Transfer Vectors for Mammalian Cells (J. H. Miller and M. P. Calos eds., 1987, Cold Spring Harbor Laboratory); Harlow and Lane, Antibodies, (Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y., 1998); Immunochemical Methods in Cell and Molecular Biology (Mayer and Walker, eds., Academic Press, London, 1987); Handbook of Experimental Immunology, Volumes I- IV (D. M. Weir and CC Blackwell, eds., 1986); Roitt, Essential Immunology, 6th Edition, (Blackwell Scientific Publications, Oxford, 1988); Current Protocols in Immunology (Q. E. Coligan, A. M. Kruisbeek, D. H. Margulies, E. M. Shevach and W. Strober, eds., 1991); Annual Review of Immunology, as well as monographs in journals such as Advances in Immunology.

[0151] B. DEFINITIONS

[0152] Prior to setting forth this disclosure in more detail, it may be helpful to an understanding thereof to provide definitions of certain terms to be used herein.

[0153] Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by those of ordinary skill in the art to which the invention belongs. Although any methods and materials similar or equivalent to those described herein can be used in the practice or testing of particular embodiments, preferred embodiments of compositions, methods and materials are described herein. For the purposes of the present disclosure, the following terms are defined below.

[0154] The articles “a,” “an,” and “the” are used herein to refer to one or to more than one (i.e., to at least one, or to one or more) of the grammatical object of the article. By way of example, “an element” means one element or one or more elements.

[0155] The use of the alternative (e.g., “or”) should be understood to mean either one, both, or any combination of the recited alternatives.

[0156] The term “and / or” should be understood to mean either one of, or both of, the alternatives.

[0157] 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 by as much as 15%, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2% or 1% to a reference quantity, level, value, number, frequency, percentage, dimension, size, amount, weight or length. In one embodiment, the term “about” or “approximately” refers a range of quantity, level, value, number, frequency, percentage, dimension, size, amount, weight or length ± 15%, ± 10%, ± 9%, ± 8%, ± 7%, ± 6%, ± 5%, ± 4%, ± 3%, ± 2%, or ± 1% of a reference quantity, level, value, number, frequency, percentage, dimension, size, amount, weight or length. In one embodiment, a range, e.g., 1 to 5, about 1 to 5, or about 1 to about 5, refers to each numerical value encompassed by the range. For example, in one non-limiting and merely illustrative embodiment, the range “1 to 5” is equivalent to the expression 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.

[0158] As used herein, the term “substantially” refers to a quantity, 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 higher compared to a reference quantity, level, value, number, frequency, percentage, dimension, size, amount, weight or length. In one embodiment, “substantially the same” refers to a quantity, level, value, number, frequency, percentage, dimension, size, amount, weight or length that produces an effect, e.g., a physiological effect, that is approximately the same as a reference quantity, level, value, number, frequency, percentage, dimension, size, amount, weight or length. In particular embodiments, “substantially lacks cell binding or cell attachment activity and / or cognate receptor binding activity” or “substantially abates cell binding or cell attachment activity and / or cognate receptor binding activity” refers to the negligible or undetectable or absent cell binding activity or cell attachment activity of a modified membrane-bound viral glycoprotein contemplated herein to bind or attach to its cognate receptor on the surface of a cell compared to the cell binding activity or cell attachment activity of the unmodified membrane-bound viral glycoprotein to bind or attach to its cognate receptor on the surface of the cell.

[0159] Throughout this specification, unless the context requires otherwise, the words “comprise”, “comprises” and “comprising” will be understood to imply the inclusion of a stated step or element or group of steps or elements but not the exclusion of any other step or element or group of steps or elements. By “consisting of’ is meant including, and limited to, whatever follows the phrase “consisting of.” Thus, the phrase “consisting of’ indicates that the listed elements are required or mandatory and that no other elements may be present. The phrase “consisting essentially of’ means including any elements listed after the phrase and other elements that do not interfere with or contribute to the activity or action specified in the disclosure for the listed elements. Thus, the phrase “consisting essentially of’ indicates that the listed elements are required or mandatory but that no other elements are present that materially affect the activity or action of the listed elements. Reference throughout this specification to “one embodiment,” “an embodiment,” “a particular embodiment,” “a related embodiment,” “a certain embodiment,” “an additional embodiment,” or “a further embodiment” or combinations thereof means 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 foregoing 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 is also understood that the positive recitation of a feature in one embodiment, serves as a basis for excluding the feature in a particular embodiment.

[0160] The terms, “binding domain,” “extracellular binding domain,” and “extracellular antigen binding domain” are used interchangeably and refers to a domain that enables an engineered receptor, e.g., a chimeric antigen receptor (CAR), a chimeric costimulatory receptor (CCR), an alpha-beta T cell receptor (aP TCR), a gamma delta T cell receptor (y5 TCR), a dimerizing agent regulated immunoreceptor complex (DARIC), a chimeric TGF-P receptor (CTBR), or a zetakine receptor to specifically bind to a target antigen. The binding domain may be derived either from a natural, synthetic, semi-synthetic, or recombinant source.

[0161] An “antibody” refers to a polypeptide that comprises at least a light chain immunoglobulin variable region and / or a heavy chain immunoglobulin variable region, which specifically recognizes and binds one or more epitopes of an antigen.

[0162] Antibodies include polyclonal and monoclonal antibodies and antigen binding fragments thereof; murine antibodies, camelid antibodies, and human antibodies, and antigen binding fragments thereof; and chimeric antibodies, an antibody that comprises variable regions from a non-human species and human constant regions, heteroconjugate antibodies, and humanized antibodies, an antibody that comprises complementarity determining regions (CDRs) from a non-human species and human framework and constant regions, and antigen binding fragments thereof.

[0163] Murine, chimeric, humanized, and human antibodies comprise two heavy chains and two light chains. Each heavy chain consists of a variable region (VH) and three constant regions (CHI, CH2, CH3), while each light chain consists of a variable region (VL) and a constant region (CL). Mammalian immunoglobulin heavy chains are classified as immunoglobulin (Ig)A, IgD, IgE, IgG, and IgM. Mammalian immunoglobulin light chains are classified as X or K.

[0164] Light and heavy chain variable regions contain a “framework” region interrupted by three hypervariable regions, also called “complementarity-determining regions” or “CDRs.” The sequences of the framework regions of different light or heavy chains are relatively conserved within a species, such as humans. The framework regions serve to position and align the CDRs in three-dimensional space to bind to an epitope. The CDRs of each chain are numbered sequentially starting from the N-terminus and are also typically identified by the chain in which the particular CDR is located. Heavy chain CDRs are referred to as CDRH1, CDRH2, and CDRH3, and light chain CDRs are referred to as CDRL1, CDRL2, and CDRL3. Although CDRs vary from antibody to antibody, the limited number of amino acid positions within the CDRs directly involved in antigen binding are called specificity determining residues (SDRs).

[0165] CDRs can be defined or identified by conventional methods, such as by sequence according to Wu and Kabat, J Exp Med. 132(2):211-50 (1970) and Kabat and Wu, Ami New York Acad Sci. 190:382-93 (1971), or by structure according to Chothia and Lesk, J Mol. Biol. 196(4): 901-917 (1987) and Chothia et al., Nature. 342:877-83(1989). Other boundaries defining CDRs overlapping with the Kabat CDRs have been described by Padlan et al., FASEB J. 9:133-9 (1995) and MacCallum et al., J Mol Biol. 262:732-745 (1996). Additional methods of determining CDRs include the Gelfand numbering system described in Gelfand and Kister, PNAS USA. 92:10884-8(1995), Gelfand etal., Protein Eng. 11:1015— 25 (1998), and Gelfand et al., PNAS USA. 93:3675-8 (1996); the Honneger number system described in Honegger and Pliickthun, J Mol Biol. 309:657-70 (2001); the AbM numbering system described by Abhinandan and Martin, Mol Immunol. 45:3832-9 (2008); and the IMGT numbering system described in Giudicelli et al., Nucleic Acids Res. 25:206-11 (1997), Lefranc, Immunol Today 18:509 (1997), and Lefranc et al., Dev Comp Immunol. 27:55-77 (2003). Proprietary and publicly programs that identify CDRs are available, e.g., abYsis (abysis.org / abysis / ) and IMGT / V-QUEST (imgt.org / IMGT_vquest).

[0166] “VL” or “VL” refers to the variable region of an immunoglobulin fight chain, including that of an antibody, Fv, scFv, dsFv, Fab, or other antigen binding fragment thereof. “VH” or “VH” refer to the variable region of an immunoglobulin heavy chain, including that of an antibody, Fv, scFv, dsFv, Fab, heavy chain only antibody (hcAb) or other antigen binding fragment thereof. An “antigen binding fragment” or “antigen binding portion” refers to one or more fragments of an antibody that retain the ability to specifically bind to an antigen. An “isolated antibody or antigen binding fragment thereof’ refers to an antibody or antigen binding fragment thereof that has been separated from its natural environment and / or that is derived from a natural, synthetic, semi-synthetic, or recombinant source. Illustrative examples of antigen binding fragments suitable for incorporation into engineered receptors contemplated in particular embodiments herein include, but are not limited to, a Camel Ig, a Llama Ig, an Alpaca Ig, Ig NAR, a Fab' fragment, a F(ab')2 fragment, a bispecific Fab dimer (Fab2), a trispecific Fab trimer (Fab3), an Fv, a single chain Fv protein (“scFv”), a bis-scFv, (scFv , a minibody, a diabody, a triabody, a tetrabody, a disulfide stabilized Fv protein (“dsFv”), a Fibronectin type III (FN3) domain antibody, a single-domain antibody (sdAb or nanobody, e.g., a camelid VHH or shark VNAR), a centyrin, and other portions of full length antibodies sufficient for antigen binding, and combinations thereof.

[0167] A “heavy chain antibody” or “he Ab” refers to an antibody that contains two heavy chain variable domains and no light chains. A “camelid antibody” or “camelid Ig” refers to an hcAb isolated from a Camel, Alpaca, or Llama that consists of a homodimer of a heavy chain variable domain (VHH) and CH2 and CH3 constant domains. An “IgNAR” or “immunoglobulin new antigen receptor” refers to an he Ab isolated from the shark immune repertoire that consists of a homodimer of a variable new antigen receptor (VNAR) domain and five constant new antigen receptor (CNAR) domains.

[0168] A “single domain antibody,” “sdAb,” or “nanobody” as used herein refers to an antibody fragment that contains the smallest known antigen binding unit of the variable region of a heavy chain antibody, e.g., a camelid VHH or shark VNAR. A “humanized VHH” refers to a single domain non-human VHH that has undergone humanization to reduce potential immunogenicity of the antibody in human recipients. A “humanized VNAR” refers to a single domain non-human VNAR that has undergone humanization to reduce potential immunogenicity of the antibody in human recipients.

[0169] A “single-chain Fv” or “scFv” antibody fragment comprises the VH and VL domains of an antibody, wherein these domains are present in a single polypeptide chain and in either orientation (e.g., VL-VH or VH-VL). Generally, the scFv polypeptide further comprises a polypeptide linker between the VH and VL domains which enables the scFv to form the desired structure for antigen binding. The terms “spacer,” “spacer domain,” or “spacer polypeptide” are used interchangeably and refer to a polypeptide domain or sequence of amino acids in a non- viral membrane-bound tropism polypeptide disposed between an extracellular antigen targeting domain and a transmembrane domain or an co amino acid, i.e., the amino acid serving as the attachment site of the GPI anchor. A spacer positions the extracellular antigen targeting domain away from the particle surface to enable proper particle / target cell contact, attachment, or binding. A spacer may be derived either from a natural, synthetic, semisynthetic, or recombinant source. Illustrative examples of spacer domains include but are not limited to hinge or stalk domains derived, obtained, or isolated from IgGl, IgG2, IgG4, CD2, CD3, CD4, CD8a, CD8p, CD28, CD45ABC, CD45RO, CD45RAB, CD45RAC, CD45RBC, CD45RA, CD45RB, CD45RC, CD45del-d3, CD45del-d2-d3 (e.g., SEQ ID NOs: 411-423), CADHERIN 1 and variants thereof (e.g., SEQ ID NOs: 424-427), CEACAM 5 and variants thereof (e.g., SEQ ID NOs: 428-433), and polypeptide linkers of similar amino acid composition, rigidity, flexibility and / or length.

[0170] A “hinge domain,” is a polypeptide domain or sequence of amino acids that plays a role in positioning the antigen binding domain away from the effector cell surface to enable proper cell / cell contact, antigen binding and activation. A hinge domain is placed between a binding domain and a transmembrane domain (TM). A hinge domain may be derived either from a natural, synthetic, semi-synthetic, or recombinant source. A hinge domain may be altered by substituting one or more cysteine and / or proline residues in a naturally occurring immunoglobulin hinge domain with one or more other amino acid residues (e.g., one or more serine residues).

[0171] A “transmembrane domain” or “TM domain” refers to a hydrophobic portion of polypeptide that anchors the polypeptide to the plasma membrane of the cell. The TM domain may be derived either from a natural, synthetic, semi-synthetic, or recombinant source.

[0172] A “glycosylphosphatidylinositol anchor” or “GPI anchor” refers to a moiety comprising a hydrophilic oligosaccharide and a lipophilic fatty acid that attaches a protein to the endoplasmatic reticulum (ER) membrane by a transamidase enzyme complex, the protein subsequently delivered to the outer leaflet of the plasma membrane.

[0173] An “intracellular signaling domain” refers to a portion of a polypeptide that participates in transducing the message of effective binding of a target antigen by a receptor expressed on an immune effector cell to the immune effector cell’s interior to elicit one or more effector functions (an “effector function” refers to a specialized function of an immune effector cell), e.g., activation, cytokine production, proliferation and cytotoxic activity, including the release of cytotoxic factors, or other cellular responses elicited with antigen binding to the receptor expressed on the immune effector cell. “Intracellular signaling domains” include a polypeptide domain or functional fragment thereof, which transduces an effector function signal and that directs a cell to perform a specialized function. The term intracellular signaling domain is meant to include any truncated portion of an intracellular signaling domain sufficient to transducing effector function signal.

[0174] T cell activation can be said to be mediated by two distinct classes of intracellular signaling domains: primary signaling domains that initiate antigen-dependent primary activation through the TCR (e.g. , a TCR / CD3 complex) and costimulatory signaling domains that act in an antigen-independent manner to provide a secondary or costimulatory signal.

[0175] A “primary signaling domain” refers to a signaling domain that regulates the primary activation of a TCR complex either in a stimulatory way, or in an inhibitory way. Primary signaling domains that act in a stimulatory manner may contain one or more signaling motifs which are known as immunoreceptor tyrosine-based activation motifs or IT AMs.

[0176] A “co stimulatory signaling domain” or “co stimulatory domain” refers to an 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 upon binding to antigen.

[0177] The terms “individual” and “subject” are often used interchangeably and refer to any animal that exhibits a symptom of a disease, disorder, or condition, e.g., cancer, that can be treated with the recombinant particles, e.g., recombinant retroviral particles or lentiviral particles, gene therapy vectors, compositions, and methods contemplated elsewhere herein. Suitable subjects (e.g., patients) include laboratory animals (e.g., mouse, rat, rabbit, or guinea pig), farm animals, and domestic animals or pets (e.g., cat or dog). Non-human primates and, preferably, human patients, are preferred subjects.

[0178] A “patient” refers to a subject that has been diagnosed with a particular disease, disorder, or condition that can be treated with the recombinant particles, e.g., recombinant retroviral particles or lentiviral particles, gene therapy vectors, compositions and methods disclosed elsewhere herein.

[0179] “Treatment” or “treating,” as used herein includes any beneficial or desirable effect on the symptoms or pathology of a disease or pathological condition and may include even minimal reductions in one or more measurable markers of the disease or condition being treated. Optionally, treatment can include reducing the disease burden or delaying disease progression. “Treatment” may, but does not necessarily, indicate complete eradication or cure of the disease or condition, or associated symptoms thereof.

[0180] “Prevent,” “prevention,” “preventing” and the like, as used herein, indicate an approach for preventing, inhibiting, or reducing the likelihood of occurrence or recurrence of a disease or condition. It also refers to delaying the onset or recurrence of a disease or condition or delaying the occurrence or recurrence of the symptoms of a disease or condition. Prevention includes reducing the intensity, effect, symptoms and / or burden of a disease or condition prior to onset or recurrence of the disease or condition.

[0181] “Ameliorating at least one symptom of’ as used herein, refers to decreasing one or more symptoms of the disease or condition for which a subject is being treated. In particular embodiments, the disease or condition being treated is a cancer, and the one or more symptoms ameliorated include, but are not limited to, weakness, fatigue, shortness of breath, easy bruising and bleeding, frequent infections, enlarged lymph nodes, distended or painful abdomen (due to enlarged abdominal organs), bone or joint pain, fractures, unplanned weight loss, poor appetite, night sweats, persistent mild fever, and decreased urination (due to impaired kidney function).

[0182] Additional definitions are set forth throughout this disclosure.

[0183] C. RECOMBINANT PARTICLES

[0184] Various particles have been proposed as gene delivery vehicles. The present disclosure contemplates, in part, a recombinant particle engineered to bind to a desired or target cell type and deliver a payload. Recombinant particles comprise a surface and a payload: the surface comprising one or more mutated viral envelope glycoproteins that retain fusogenic activity and substantially lack or lack cognate receptor binding activity and one or more non-viral membrane-bound tropism polypeptides engineered to bind a cell, e.g., an immune effector cell; and the payload comprising one or more polynucleotide and / or polypeptides. Suitable recombinant particles for use in particular embodiments contemplated herein include but are not limited to a fusosome; an extracellular vesicle, including a microvesicle, an apoptotic body, and an exosome; a lipid nanoparticle; a virus-like particle (VLPs); or a recombinant viral particle. In preferred embodiments, a recombinant particle is a recombinant viral particle or recombinant virus. The terms “recombinant viral particle” and “recombinant virus” are used interchangeably.

[0185] Recombinant viral particles have been used as a gene delivery platform for treatments of severe genetic diseases and cancer. Illustrative examples of recombinant viral particles suitable for use in particular embodiments contemplated herein are derived from an adenovirus (Ad), an adeno-associated virus (AAV), a rhabdovirus (e.g., lyssavirus, vesiculovirus), a paramyxovirus (e.g., henipavirus, morbillivirus, respirovirus, rubelavirus), a herpes simplex vims (e.g., HSV-1, HSV-2), vaccinia vims, or a retrovirus (e.g., gammaretrovims, lend vims).

[0186] Retrovimses are a commonly used tool for gene delivery (Miller, 2000, Nature. 357: 455-460). The term “retrovirus” refers to an enveloped RNA vims that reverse transcribes its genomic RNA into a linear double- stranded DNA copy and subsequently covalently integrates its genomic DNA into a host genome. A recombinant retroviral particle comprises a viral envelope, one or more retroviral proteins, e.g., matrix protein (MA), capsid protein (CA), nucleocapsid protein (NC), reverse transcriptase (RT), and integrase (IN), and a retroviral vector derived or engineered from a retrovirus genome.

[0187] Illustrative retrovimses suitable for deriving or engineering recombinant retroviral particles contemplated in particular embodiments herein include but are not limited to an alpharetrovims, a betaretrovims, a gammaretrovims, a deltaretrovims, an epsilonretrovims, a spumavims, and a lentivims.

[0188] Alpharetrovimses include but are not limited to avian leukosis vims, avian carcinoma Mill Hill vims 2, avian myeloblastosis vims, avian myelocytomatosis vims 29, avian sarcoma vims CT10, fujinami sarcoma vims, rous sarcoma vims, UR2 sarcoma vims and Y73 sarcoma vims.

[0189] Betaretrovimses include but are not limited to mouse mammary tumor vims, Jaagsiekte sheep retrovirus, langur vims, Mason-Pfizer monkey vims, and squirrel monkey retro vims (SMRV).

[0190] Deltaretrovimses include but are not limited to bovine leukemia vims, primate T- lymphotropic vims 1, primate T-lymphotropic vims 2, primate T-lymphotropic vims 3, and primate T-lymphotropic vims 4.

[0191] Epsilonretrovimses include but are not limited to walleye dermal sarcoma vims, walleye epidermal hyperplasia vims 1, and walleye epidermal hyperplasia vims 2. Gammaretroviruses include but are not limited to baboon endogenous virus (BaEV), chick syncytial virus, feline endogenous virus (e.g., RD 114), feline leukemia virus (FeLV), Finkel-Biskis-Jinkins murine sarcoma virus, Gardner-Amstein feline sarcoma virus, gibbon ape leukemia virus (GAEV), guinea pig type-C oncovirus, Hardy -Zuckerman feline sarcoma virus, Harvey murine sarcoma virus, Kirsten murine sarcoma vims, koala retrovirus, murine leukemia vims (MEV), Moloney murine leukemia vims (MoMLV), Moloney murine sarcoma vims, porcine endogenous vims (PERV), Porcine type-C oncovims, reticuloendotheliosis vims (REV), Snyder-Theilen feline sarcoma vims, Trager duck spleen necrosis vims, viper retrovirus, xenotropic murine leukemia vims- related vims (XMRV), and woolly monkey sarcoma vims.

[0192] Spumavimses include but are not limited to simian foamy vims, bovine foamy vims, equine foamy vims, feline foamy vims, human foamy vims (HFV), and brown greater galago prosimian foamy vims.

[0193] In particular embodiments, a recombinant retroviral particle is derived or engineered from a retro vims selected from the group consisting of: SMRV, BaEV, RD 114, FeLV, GALV, MLV, MoMLV, PERV, REV, XMRV, and HFV.

[0194] A “lentivims” refers to a complex retrovims. Among retroviruses, lentivimses are the most efficient at transducing resting or growth-arrested cells. In preferred embodiments, a recombinant retroviral particle is a recombinant lentiviral particle or recombinant lentivims.

[0195] Lentivimses suitable for deriving or engineering recombinant lentivimses contemplated in particular embodiments herein include but are not limited to human immunodeficiency vims (HIV), including HIV type 1 (HIV-1) and HIV type 2 (HIV-2); visna-maedi vims (VMV); caprine arthritis-encephalitis vims (CAEV); equine infectious anemia vims (EIAV); feline immunodeficiency vims (FIV); bovine immune deficiency vims (BIV); and simian immunodeficiency vims (SIV).

[0196] In particular embodiments, a recombinant lentiviral particle is derived or engineered from an HIV-1 or HIV-2 lentivims.

[0197] D. PARTICLE SURFACE

[0198] Recombinant particles engineered to bind to a particular cell type and deliver a payload comprise an outer surface that comprises, expresses, or displays (i) one or more polypeptides that direct the particle to, and allow it to bind or attach to, a desired cell type and (ii) one or more polypeptides that mediate fusion with the cell. In particular embodiments, a recombinant particle selected from the group consisting of a fusosome; a microvesicle, an apoptotic body, and an exosome; a lipid nanoparticle; a virus-like particle (VLPs); or a recombinant viral particle, comprises an outer surface selected from the group consisting of a single layer of phospholipid, a phospholipid bilayer, a cell membrane, a capsid, or a viral envelope.

[0199] In particular embodiments, the outer surface of the particle is engineered to express or display one or more mutated viral envelope glycoproteins that retain fusogenic activity and that substantially lack or lack cognate receptor binding activity and one or more non- viral membrane-bound tropism polypeptides engineered to bind a specific cell type, e.g. , an immune effector cell. Without wishing to be bound by any particular theory, it is contemplated that the viral envelope glycoproteins herein do not contribute to particle tropism. The viral envelope glycoproteins contemplated herein comprise one or more mutations e.g., amino acid substitution) that substantially reduce or eliminate their native antigen binding activities and / or blind the viral envelope glycoprotein from recognizing and binding or attaching to its cognate receptor expressed on a cell while still retaining the ability to mediate fusion with the cell. In this way, the native tropism activities of viral envelope glycoproteins contemplated herein are substantially extinguished or ablated while the fusion activity is retained or preserved.

[0200] Recombinant retroviral or lentiviral particles comprise an outer surface selected from the group consisting of a lipid bilayer, a cell membrane, or a viral envelope comprising polypeptides that mediate virus - cell attachment and fusion; and one or more polynucleotide and / or polypeptide pay loads. The outer surface of the retroviral or lentiviral particle is engineered to uncouple or separate particle fusion and tropism activities.

[0201] In particular embodiments, the outer surface of the retroviral or lentiviral particle is engineered to express or display one or more non- viral membrane-bound tropism polypeptides that enable selective and / or specific binding of the virus to a desired cell type. The outer surface is also engineered to express or display a viral envelope glycoprotein that mediates virus - cell fusion but that substantially lacks or lacks the ability to bind its cognate receptor. In other words, the viral envelope glycoprotein provides the fusion activity of the recombinant particle and does not provide or contribute to the tropism activity of the particle. Likewise, the non- viral membrane-bound tropism polypeptides provide the tropism activity of the recombinant particle and do not provide or contribute to the fusion activity of the particle. The foregoing engineering strategy enables highly efficient on-target delivery of therapeutic pay loads to the desired cell types while minimizing, reducing and / or eliminating delivery to undesired cell types.

[0202] In particular embodiments, a recombinant retroviral particle comprises a lipid bilayer, cell membrane, or viral envelope comprising one or more mutated viral envelope glycoproteins that mediate fusion of the viral particle with a cell, but that do not substantially bind or bind its cognate receptor, and one or more non- viral membrane-bound tropism polypeptides.

[0203] In particular embodiments, one or more mutated viral envelope glycoproteins comprise one or more rhabdovirus (e.g., vesiculovirus) envelope glycoproteins or one or more paramyxovirus (e.g., morbillivirus or henipavirus) envelope glycoproteins. In particular embodiments, a mutated viral envelope glycoprotein comprises a heterologous signal peptide.

[0204] In particular embodiments, one or more mutated viral envelope glycoproteins comprise one or more rhabdovirus (e.g., vesiculovirus) envelope glycoproteins or one or more paramyxovirus (e.g., morbillivirus or henipavirus) envelope glycoproteins.

[0205] In particular embodiments, one or more mutated viral envelope glycoproteins comprise one or more rhabdovirus (e.g., vesiculovirus) envelope glycoproteins and a native or heterologous signal peptide, which is subsequently cleaved.

[0206] In particular embodiments, one or more mutated viral envelope glycoproteins comprise one or more paramyxovirus (e.g., morbillivirus or henipavirus) envelope glycoproteins and a native or heterologous signal peptide, which is subsequently cleaved.

[0207] Illustrative examples of heterologous signal peptides are set forth in SEQ ID NOs: 449-454.

[0208] I. RHABDOVIRUS ENVELOPE GLYCOPROTEINS

[0209] The Rhabdoviridae family of viruses are negative- sense, single-stranded RNA viruses with a genome of about 10.8-16.1 kb. The prototypic mammalian rhabdoviruses include the rabies virus (RABV) and vesiculoviruses, which cause severe human and animal disease. In particular embodiments, a recombinant particle comprises one or more rhabdovirus envelope glycoproteins modified to substantially ablate or ablate the glycoprotein’s ability bind its cognate receptor expressed on a cell while retaining fusogenic activity. The Rhabdoviridae family includes cytorhabdo viruses, dichorhaviruses, ephemero viruses, lyssaviruses, no virhabdo viruses, nucleorhabdoviruses, perhabdo viruses, sigmaviruses, spriviviruses, tibroviruses, tupaviruses, varicosaviruses, and vesiculoviruses, among others. a. Vesiculovirus Envelope Glycoproteins

[0210] Vesiculoviruses use a single glycoprotein to enter a cell: glycoprotein G mediates both cell attachment (tropism) and cell fusion. Glycoprotein G is normally responsible for virus attachment to specific receptors (e.g., low-density lipoprotein receptor (LDL-R) and other members of this receptor family) and fusion between the viral and endosomal membranes. In particular embodiments, a modified vesiculovirus G envelope protein comprises one or more amino acid substitutions that that enable the polypeptide to mediate fusion of the viral particle and a cell but that substantially ablate or ablate the polypeptide’s ability to bind its cognate receptor expressed on a cell, e.g., LDL-R.

[0211] Illustrative examples of vesiculoviruses suitable for use in particular embodiments from which G glycoproteins can be isolated include, but are not limited to vesicular stomatitis Alagoas virus (VSAV; Alagoas vesiculovirus), Carajas virus (CJSV; Carajas vesiculovirus), Chandipura virus (CHPV; Chandipura vesiculovirus), Cocal virus (COCV; Cocal vesiculovirus), vesicular stomatitis Indiana vims (VSIV, f.k.a. VSV; Indiana vesiculovirus), Isfahan vims (ISFV; Isfahan vesiculovirus), Maraba vims (MARAV; Maraba vesiculovirus), Morreton vims (MORV; Morreton vesiculovirus), vesicular stomatitis New Jersey vims (VSNJV; New Jersey vesiculovirus), and Piry vims (PIRYV; Piry vesiculovirus).

[0212] In particular embodiments, a vesiculovirus G envelope protein comprises an amino acid sequence set forth in any one of SEQ ID NOs: 15-322 disclosed in U.S. Patent Application No. 20200216502, each said sequence incorporated by reference herein in its entirety, or an amino acid sequence at least 95%, 96%, 97%, 98%, or 99% identical thereto that enables the polypeptide to mediate fusion of the viral particle with a cell but that substantially ablates or ablates the polypeptide’s ability to bind its cognate receptor expressed on a cell, e.g., LDL-R.

[0213] In particular embodiments, a vesiculovirus is vesicular stomatitis Indiana vims (VSIV). In particular embodiments, a mutated viral envelope glycoprotein is derived from a VSIV envelope glycoprotein (VSIV-G; e.g., SEQ ID NO: 1: KFTIVFPHNQKGNWKNVPSNYHYCPSSSDLNWHNDLIGTALQVKMPKSHKAIQA DGWMCHASKWVTTCDFRWYGPKYITHSIRSFTPSVEQCKESIEQTKQGTWLNPG FPPQSCGYATVTDAEAVIVQVTPHHVLVDEYTGEWVDSQFINGKCSNYICPTVHN STTWHSDYKVKGLCDSNLISMDITFFSEDGELSSLGKEGTGFRSNYFAYETGGKA CKMQYCKHWGVRLPSGVWFEMADKDLFAAARFPECPEGSSISAPSQTSVDVSLI QDVERILDYSLCQETWSKIRAGLPISPVDLSYLAPKNPGTGPAFTnNGTLKYFETR YIRVDIAAPILSRMVGMISGTTTERELWDDWAPYEDVEIGPNGVLRTSSGYKFPLY MIGHGMLDSDLHLSSKAQVFEHPHIQDAASQLPDDESLFFGDTGLSKNPIELVEG WFSSWKSSIASFFFnGLnGLFLVLRVGIHLCIKLKHTKKRQIYTDIEMNRLGK) or an amino acid sequence at least 95%, 96%, 97%, 98%, or 99% identical thereto comprising one or more modifications that enable the polypeptide to mediate fusion of the viral particle and a cell but that substantially ablate or ablate the polypeptide’s ability to bind its cognate receptor expressed on a cell, e.g., LDL-R. In particular embodiments, a mutated viral envelope glycoprotein is derived from a VSIV-G polypeptide set forth in SEQ ID NO: 1 comprising L47I and / or H80Q amino acid substitutions, such substitutions being present in naturally occurring variants of VSIV.

[0214] In particular embodiments, a mutated VSIV-G envelope protein comprises one or more of: one or more amino acid substitutions at H8, N9, Q10, K47, K50, A51, S183, S179, N180, 1182, M184, Y209, 1347, T350, T352, E353, and R354 (substitution with any amino acid; a conservation substitution; a disruptive substitution; substitution with D, E, A, G, F, or Q; or substitution with A, G, F, or Q); an insertion of TT between N9 and Q10, an insertion of GGS between H8 and N9, an insertion of GGS between N9 and Q10, an insertion of TT between N208 and Y209, an insertion of GGS between P46 and K47, and an insertion of GGS between N208 and Y209; or a deletion of residues 1-8. In particular embodiments, a VSIV-G envelope protein comprises one or more amino acid substitutions at H8, K47, Y209, and R354. In particular embodiments, a VSIV-G envelope protein comprises one or more amino acid substitutions at H8, K47, Y209, and R354, wherein each substitution can be with A, G, F, or Q. Amino acid positions are with reference to a VSIV-G envelope protein that lacks a signal peptide, e.g., SEQ ID NO: 1. In particular embodiments, a mutated viral envelope glycoprotein is derived from a VSIV-G polypeptide set forth in SEQ ID NO: 1 comprising L47I and / or H80Q amino acid substitutions, such substitutions present in naturally occurring variants of VSIV.

[0215] In particular embodiments, a mutated VSIV-G polypeptide comprises one or more amino acid substitutions at K47, 1182, and / or R354 (substitution with any amino acid; a conservation substitution; a disruptive substitution; substitution with D, E, A, G, F, or Q; or substitution with A, G, F, or Q). In particular embodiments, a mutated VSIV-G polypeptide comprises amino acid substitutions at K47, 1182, or R354; K47 and 1182; K47 and R354; 1182 and R354; or at K47, 1182, and R354 of SEQ ID NO: 1.

[0216] In particular embodiments, a mutated VSIV-G polypeptide comprises one or more of the following amino acid substitutions: K47A, K47Q, I182E, I182D, R354A, and / or R354Q. In particular embodiments, a mutated VSIV-G polypeptide comprises the following amino acid substitutions: K47A, K47Q, I182E, I182D, R354A, orR354Q; K47A andI182E; K47A and I182D; K47Q and I182E; K47Q and I182D; I182E and R354A; I182E and R354Q; I182D and R354A; I182D and R354Q; K47A and R354A; K47A and R354Q; K47Q and R354A; K47Q and R354Q; K47A, I182E, and R354A; K47A, I182D, and R354A; K47Q, I182E, and R354A; K47Q, I182D, and R354A; K47A, I182E, and R354Q; K47A, I182D, and R354Q; K47Q, I182E, and R354Q; or K47Q, I182D, and R354Q of SEQ ID NO: 1.

[0217] In particular embodiments, a VSIV-G envelope protein comprises one or more amino acid substitutions at K47 and R354 (with reference to a VSIV-G envelope protein that lacks a signal peptide, e.g., SEQ ID NO: 1). In particular embodiments, a VSIV-G envelope protein comprises one or more amino acid substitutions selected from the group consisting of: K47A and R354A; K47A and R354G; K47A and R354F; K47A and R354Q; K47G and R354A; K47G and R354G; K47G and R354F; K47G and R354Q;K47F and R354A; K47F and R354G; K47F and R354F; K47F and R354Q; K47Q and R354A; K47Q and R354G; K47Q and R354F; and K47Q and R354Q. In a preferred embodiment, a VSIV-G envelope comprises the amino acid substitutions K47Q and R354A.

[0218] In particular embodiments, a mutated VSIV-G envelope protein comprises an amino acid sequence set forth in SEQ ID NO: 2 (wherein Xi= I, X2= A, X3 = Q, and X4= A; Xi= I, X2= A, X3= Q, and X4= G; Xi= I, X2= A, X3= Q, and X4= F; Xi= I, X2= A, X3= Q, and X4= Q; Xi= L, X2= A, X3= Q, and X4= A; Xi= L, X2= A, X3= Q, and X4= G; Xi= L, X2= A, X3= Q, and X4= F; Xi= L, X2= A, X3= Q, and X4= Q; Xi= I, X2= A, X3= H, and X4= A; Xi= I, X2= A, X3= H, and X4= G; Xi= I, X2= A, X3= H, and X4= F; Xi= I, X2= A, X3= H, and X4= Q; Xi= L, X2= A, X3= H, and X4= A; Xi= L, X2= A, X3= H, and X4= G; Xi= L, X2= A, X3= H, and X4= F; Xi= L, X2= A, X3= H, and X4= Q; Xi= I, X2= G, X3= Q, and X4= A; Xi= I, X2= G, X3= Q, and X4= G; Xi= I, X2= G, X3= Q, and X4= F; Xi= I, X2= G, X3= Q, and X4= Q; Xi= L, X2= G, X3= Q, and X4= A; Xi= L, X2= G, X3= Q, and X4= G; Xi= L, X2= G, X3= Q, and X4= F; Xi= L, X2= G, X3= Q, and X4= Q; X1= I, X2= G, X3= H, and X4= A; Xi= I, X2= G, X3= H, and X4= G; Xi= I, X2= G, X3= H, and X4= F; Xi= I, X2= G, X3= H, and X4= Q; Xi= L, X2= G, X3= H, and X4= A; Xi= L, X2= G, X3= H, and X4= G; Xi= L, X2= G, X3= H, and X4= F; Xi= L, X2=

[0219] G, X3= H, and X4= Q; Xi= I, X2= F, X3= Q, and X4= A; Xi= I, X2= F, X3= Q, and X4= G; Xi= I, X2= F, X3= Q, and X4= F; Xi= I, X2= F, X3= Q, and X4= Q; Xi= L, X2= F, X3= Q, and X4= A; Xi= L, X2= F, X3= Q, and X4= G; Xi= L, X2= F, X3= Q, and X4= F; Xi= L, X2= F, X3= Q, and X4= Q; Xi= I, X2= F, X3= H, and X4= A; Xi= I, X2= F, X3=

[0220] H, and X4= G; Xi= I, X2= F, X3= H, and X4= F; Xi= I, X2= F, X3= H, and X4= Q; Xi= L, X2= F, X3= H, and X4= A; Xi= L, X2= F, X3= H, and X4= G; Xi= L, X2= F, X3= H, and X4= F; Xi= L, X2= F, X3= H, and X4= Q; Xi= I, X2= Q, X3= Q, and X4= A; Xi= I, X2= Q, X3= Q, and X4= G; Xi= I, X2= Q, X3= Q, and X4= F; Xi= I, X2= Q, X3= Q, and X4= Q; Xi= L, X2= Q, X3= Q, and X4= A; Xi= L, X2= Q, X3= Q, and X4= G; Xi= L, X2= Q, X3= Q, and X4= F; Xi= L, X2= Q, X3= Q, andX4= Q; Xi= I, X2= Q, X3= H, andX4= A; Xi= I, X2= Q, X3= H, and X4= G; Xi= I, X2= Q, X3= H, and X4= F; Xi= I, X2= Q, X3= H, and X4= Q; Xi= L, X2= Q, X3= H, and X4= A; Xi= L, X2= Q, X3= H, and X4= G; Xi= L, X2= Q, X3= H, andX4= F; andXi= L, X2= Q, X3= H, andX4= Q) or an amino acid sequence at least 95%, 96%, 97%, 98%, or 99% identical thereto that mediates fusion of the viral particle and a cell but that substantially ablates or ablates the polypeptide’s ability to bind its cognate receptor expressed on a cell, e.g., LDL-R.

[0221] Table 1

[0222] In particular embodiments, a mutated VSIV-G envelope protein comprises an amino acid sequence set forth in any one of SEQ ID NOs: 582-645 or an amino acid sequence at least 95%, 96%, 97%, 98%, or 99% identical thereto that mediates fusion of the viral particle and a cell but that substantially ablates or ablates the polypeptide’s ability to bind its cognate receptor expressed on a cell, e.g., LDL-R. In particular embodiments, a mutated VSIV-G envelope protein comprises an amino acid sequence set forth in any one of SEQ ID NOs: 630-645 or an amino acid sequence at least 95%, 96%, 97%, 98%, or 99% identical thereto that mediates fusion of the viral particle and a cell but that substantially ablates or ablates the polypeptide’s ability to bind its cognate receptor expressed on a cell, e.g., LDL-R. In particular embodiments, a mutated VSIV-G envelope protein comprises an amino acid sequence set forth in any one of SEQ ID NOs: 630, 634, 638, and 642 or an amino acid sequence at least 95%, 96%, 97%, 98%, or 99% identical thereto that mediates fusion of the viral particle and a cell but that substantially ablates or ablates the polypeptide’s ability to bind its cognate receptor expressed on a cell, e.g., LDL-R.

[0223] Table 2 In particular embodiments, a vesiculovirus is cocal virus (COCV). In particular embodiments, a mutated viral envelope glycoprotein is derived from a COCV envelope glycoprotein (COCV-G; e.g., SEQ ID NO: 3:

[0224] KFSIVFPQSQKGNWKNVPSSYHYCPSSSDQNWHNDEEGITMKVKMPKTHKAIQA DGWMCHAAKWITTCDFRWYGPKYITHSIHSIQPTSEQCKESIKQTKQGTWMSPGF PPQNCGYATVTDSVAVVVQATPHHVLVDEYTGEWIDSQFPNGKCETEECETVHN STVWYSDYKVTGLCDATLVDTEITFFSEDGKKESIGKPNTGYRSNYFAYEKGDKV CKMNYCKHAGVRLPSGVWFEFVDQDVYAAAKLPECPVGATISAPTQTSVDVSLI LDVERILDYSLCQETWSKIRSKQPVSPVDLSYLAPKNPGTGPAFTIINGTLKYFETR YIRIDIDNPIISKMVGKISGSQTERELWTEWFPYEGVEIGPNGILKTPTGYKFPLFMI GHGMLDSDLHKTSQAEVFEHPHLAEAPKQLPEEETLFFGDTGISKNPVELIEGWFS SWKSTVVTFFFAIGVFILLYVVARIVIAVRYRYQGSNNKRIYNDIEMSRFRK) or an amino acid sequence at least 95%, 96%, 97%, 98%, or 99% identical thereto comprising one or more modifications that enable the polypeptide to mediate fusion of the viral particle and a cell but that substantially ablate or ablate the polypeptide’s ability to bind its cognate receptor expressed on a cell, e.g., LDL-R. In particular embodiments, a COCV-G envelope protein comprises one or more amino acid substitutions at K47 and / or R354. In particular embodiments, a COCV-G envelope protein comprises one or more amino acid substitutions at K47 and / or R354, wherein each amino acid can be substituted with A, G, F, or Q. In particular embodiments, a COCV-G envelope protein comprises one or more amino acid substitutions at K47 and / or R354 (with reference to a COCV-G envelope protein that lacks a signal peptide, e.g., SEQ ID NO: 3). In particular embodiments, a COCV-G envelope protein comprises one or more amino acid substitutions selected from the group consisting of: K47A and R354A; K47A and R354G; K47A and R354F; K47A and R354Q; K47G and R354A; K47G and R354G; K47G and R354F; K47G and R354Q;K47F and R354A; K47F and R354G; K47F and R354F; K47F and R354Q; K47Q and R354A; K47Q and R354G; K47Q and R354F; and K47Q and R354Q. In particular embodiments, a COCV-G envelope protein comprises an amino acid sequence set forth in SEQ ID NO: 4 (wherein Xi= A and X2= A; Xi= A andX2= G; Xi= A andX2= F; Xi= A andX2= Q; Xi= G andX2= A; Xi= G and X2= G; Xi= G andX2= F; Xi= G andX2= Q; Xi= F andX2= A; Xi= F andX2= G; Xi= F and X2= F; Xi= F and X2= Q; Xi= Q and X2= A; Xi= Q and X2= G; Xi= Q and X2= F; or Xi= A and X2= Q) or an amino acid sequence at least 95%, 96%, 97%, 98%, or 99% identical thereto that mediates fusion of the viral particle and a cell but that substantially ablates or ablates the polypeptide’s ability to bind its cognate receptor expressed on a cell, e.g., LDL-

[0225] R.

[0226] Table 3

[0227] In particular embodiments, a mutated COCV-G envelope protein comprises an amino acid sequence set forth in any one of SEQ ID NOs: 646-661 or an amino acid sequence at least 95%, 96%, 97%, 98%, or 99% identical thereto that mediates fusion of the viral particle and a cell but that substantially ablates or ablates the polypeptide’s ability to bind its cognate receptor expressed on a cell, e.g., LDL-R. In particular embodiments, a mutated COCV-G envelope protein comprises an amino acid sequence set forth in any one of SEQ ID NOs: 658-661 or an amino acid sequence at least 95%, 96%, 97%, 98%, or 99% identical thereto that mediates fusion of the viral particle and a cell but that substantially ablates or ablates the polypeptide’s ability to bind its cognate receptor expressed on a cell, e.g., LDL- R. In particular embodiments, a mutated VSIV-G envelope protein comprises an amino acid sequence set forth in SEQ ID NOs: 658 or 661 or an amino acid sequence at least 95%, 96%, 97%, 98%, or 99% identical thereto that mediates fusion of the viral particle and a cell but that substantially ablates or ablates the polypeptide’s ability to bind its cognate receptor expressed on a cell, e.g., LDL-R.

[0228] Table 4 2. PARAMYXOVIRUS ENVELOPE GL COPROTEINS

[0229] The Paramyxoviridae family of negative- sense single- stranded RNA viruses is known to cause different types of infections in vertebrates. Examples of these infections in humans include the measles vims, encephalitis, mumps vims, parainfluenza vims, and respiratory syncytial vims. In particular embodiments, a recombinant particle comprises one or more paramyxovirus envelope glycoproteins modified to substantially ablate or ablate the glycoprotein’s ability bind its cognate receptor expressed on a cell while retaining fusogenic activity. The Paramyxoviridae family includes morbillivimses and henipavimses, among others. a. Morbillivirus Envelope Glycoproteins

[0230] Morbillivimses use two types of glycoproteins to enter a cell: glycoprotein F (a fusion polypeptide) and glycoprotein H (an attachment protein). Although glycoprotein F mediates fusion and glycoprotein H mediates binding, both H and F glycoproteins are required for viral particle fusion with a cell, glycoprotein H supports glycoprotein F in its membrane fusion function.

[0231] Efficient pseudotyping with morbillivirus envelope proteins may be accomplished using glycoprotein F comprising a tmncated cytoplasmic portion comprising at least 1 positively charged amino acid residue and no more than 9 consecutive amino acid residues as counted from the N-terminal end of the cytoplasmic portion and glycoprotein H comprising a tmncated cytoplasmic portion comprising at least 9 and no more than 19 consecutive amino acid residues as counted from the C-terminal end of the cytoplasmic portion plus an additional methionine at the N-terminus. Moreover, glycoprotein H can be modified to ablate its antigen binding activity.

[0232] In particular embodiments, the vector comprises modified morbillivirus F and H viral envelope glycoproteins, wherein glycoprotein H is modified to ablate cell binding activity and both H and F glycoproteins retain the ability to cooperatively mediate cell fusion.

[0233] Illustrative examples of morbillivimses suitable for use in particular embodiments from which F and H glycoproteins can independently be isolated from include but are not limited to canine distemper vims (CDV), cetacean morbillivirus (CeMV), feline morbillivirus (FeMV), measles vims (MV), peste-des-petits-mminants vims (PPRV), phocine distemper vims (PDV), rinderpest vims (RPV). In particular embodiments, one or more mutated morbillivirus envelope glycoproteins are derived from measles vims F (MV-F) and measles vims H (MV-H). In particular embodiments, a recombinant particle comprises one or more measles vims viral envelope glycoproteins modified to lack cell binding activity and retain fusogenic activity. In some embodiments, a recombinant particle comprises a modified MV-F glycoprotein and an MV-H glycoprotein modified to lack cell binding activity and retain fusogenic activity.

[0234] In particular embodiments, one or more mutated morbillivirus envelope glycoproteins are derived from measles vims F (MV-F) polypeptide (e.g., SEQ ID NO: 5: QIHWGNLSKIGVVGIGSASYKVMTRSSHQSLVIKLMPNITLLNNCTRVEIAEYRRL LRTVLEPIRDALNAMTQNIRPVQSVASSRRHKRFAGVVLAGAALGVATAAQITA GIALHQSMLNSQAIDNLRASLETTNQAIEAIRQAGQEMILAVQGVQDYINNELIPS MNQLSCDLIGQKLGLKLLRYYTEILSLFGPSLRDPISAEISIQALSYALGGDINKVLE KLGYSGGDLLGILESRGIKARITHVDTESYFIVLSIAYPTLSEIKGVIVHRLEGVSYN IGSQEWYTTVPKYVATQGYLISNFDESSCTFMPEGTVCSQNALYPMSPLLQECLR GSTKSCARTLVSGSFGNRFILSQGNLIANCASILCKCYTTGTIINQDPDKILTYIAAD HCPVVEVNGVTIQVGSRRYPDAVYLHRIDLGPPISLERLDVGTNLGNAIAKLEDA KELLESSDQILRSMKGLSSTSIVYILIAVCLGGLIGIPALICCCRGR) or an amino acid sequence at least 95%, 96%, 97%, 98%, or 99% identical thereto and a measles vims H (MV- H) polypeptide (e.g., SEQ ID NO: 6:

[0235] MGSRIVINREHLMIDRPYVLLAVLFVMFLSLIGLLAIAGIRLHRAAIYTAEIHKSLST NLDVTNSIEHQVKDVLTPLFKIIGDEVGLRTPQRFTDLVKFISDKIKFLNPDREYDF RDLTWCINPPERIKLDYDQYCADVAAEELMNALVNSTLLETRTTNQFLAVSKGN CSGPTTIRGQFSNMSLSLLDLYLGRGYNVSSIVTMTSQGMYGGTYLVEKPNLSSK RSELSQLSMYRVFEVGVIRNPGLGAPVFHMTNYLEQPVSNDLSNCMVALGELKL AALCHGEDSmPYQGSGKGVSFQLVKLGVWKSPTDMQSWVPLSTDDPVIDRLYL SSHRGVIADNQAKWAVPTTRTDDKLRMETCFQQACKGKIQALCENPEWAPLKD NRIPSYGVLSVDLSLTVELKIKIASGFGPLITHGSGMDLYKSNHNNVYWLTIPPMK NLALGVINTLEWIPRFKVSPYLFTVPIKEAGGDCHAPTYLPAEVDGDVKLSSNLVI LPGQDLQYVLATYDTSRVEHAVVYYVYSPSRSFSYFYPFRLPIKGVPIELQVECFT WDQKLWCRHFCVLADSESGGHITHSGMVGMGVSCTVTREDGTN) or an amino acid sequence at least 95%, 96%, 97%, 98%, or 99% identical thereto, wherein the MV-H protein lacks cell binding activity and retains fusogenic activity. In particular embodiments, the MV-H polypeptide comprises one or more amino acid substitutions at positions Y463, R515, S530, and F531 of a MV-H polypeptide (e.g., SEQ ID NO 6). In particular embodiments, the MV-H polypeptide comprises one or more of the amino acid substitutions Y463A, R515A, S530L, and F531S in an MV-H polypeptide (e.g., SEQ ID NO: 7: MGSRIVINREHLMIDRPYVLLAVLFVMFLSLIGLLAIAGIRLHRAAIYTAEIHKSLST NEDVTNSIEHQVKDVETPEFKIIGDEVGERTPQRFrDEVKFISDKIKFENPDREYDF RDETWCINPPERIKEDYDQYCADVAAEEEMNAEVNSTEEETRTTNQFEAVSKGN CSGPTTIRGQFSNMSESEEDEYEGRGYNVSSIVTMTSQGMYGGTYEVEKPNESSK RSEESQESMYRVFEVGVIRNPGEGAPVFHMTNYEEQPVSNDESNCMVAEGEEKE AAECHGEDSmPYQGSGKGVSFQEVKEGVWKSPTDMQSWVPESTDDPVIDREYE SSHRGVIADNQAKWAVPTTRTDDKERMETCFQQACKGKIQAECENPEWAPEKD NRIPSYGVESVDESETVEEKIKIASGFGPEITHGSGMDEYKSNHNNVYWETIPPMK NEAEGVINTEEWIPRFKVSPAEFNVPIKEAGGDCHAPTYEPAEVDGDVKESSNEVI EPGQDEQYVEATYDTSAVEHAVVYYVYSPSRESSYFYPFREPIKGVPIEEQVECFr WDQKEWCRHFCVEADSESGGHITHSGMVGMGVSCTVTREDGTN) or an amino acid sequence at least 95%, 96%, 97%, 98%, or 99% identical thereto, wherein the MV-H protein lacks cell binding activity and retains fusogenic activity. a. Henipavirus Envelope Glycoproteins

[0236] Henipaviruses use two types of glycoproteins to enter a cell: glycoprotein F (a fusion polypeptide) and glycoprotein G (an attachment protein). Although glycoprotein F mediates fusion and glycoprotein G mediates binding, both F and G glycoproteins are required for viral particle fusion with a target cell, glycoprotein G supports glycoprotein F in its membrane fusion function.

[0237] Efficient pseudotyping with henipavirus envelope proteins may be accomplished using glycoprotein F comprising a truncated cytoplasmic portion comprising at least 6 consecutive amino acid residues as counted from the N-terminal end of the cytoplasmic portion and glycoprotein G comprising a truncated cytoplasmic portion comprising at least 11 to at least 15 consecutive amino acid residues as counted from the C-terminal end of the cytoplasmic portion plus an additional methionine at the N-terminus. Moreover, glycoprotein G can be modified to ablate its antigen binding activity. In particular embodiments, a recombinant particle comprises henipaviruses F and G viral envelope glycoproteins, wherein glycoprotein G is modified to ablate cell binding activity and both G and F glycoproteins retain the ability to cooperatively mediate cell fusion.

[0238] Illustrative examples of henipaviruses suitable for use in particular embodiments from which F and H glycoproteins can independently be isolated from include but are not limited to Cedar vims (CedV; Cedar henipavirus), Kumasi vims (KV; Ghanaian bat henipavims), Hendra vims (HeV), (MojV; Mojiang henipavims), and Nipah vims (NiV).

[0239] In particular embodiments, one or more mutated henipaviruses envelope glycoproteins are derived from nipah vims F (NiV-F) and nipah vims G (NiV-G). In particular embodiments, a recombinant particle comprises one or more nipah vims viral envelope glycoproteins modified to lack cell binding activity and retain fusogenic activity. In some embodiments, a recombinant particle comprises a modified NiV-F glycoprotein and a NiV-G glycoprotein modified to lack cell binding activity and retain fusogenic activity.

[0240] In particular embodiments, one or more mutated henipavims envelope glycoproteins are derived from a nipah vims F (NiV-F) polypeptide (e.g., SEQ ID NO: 8: LHYEKLSKIGLVKGVTRKYKIKSNPLTKDIVIKMIPNVSNMSQCTGSVMENYKTR LNGILTPIKGALEIYKNNTHDLVGDVRLAGVIMAGVAIGIATAAQITAGVALYEA MKNADNINKLKSSIESTNEAVVKLQETAEKTVYVLTALQDYINTNLVPTIDKISCK QTELSLDLALSKYLSDLLFVFGPNLQDPVSNSMTIQAISQAFGGNYETLLRTLGYA TEDFDDLLESDSITGQIIYVDLSSYYUVRVYFPILTEIQQAYIQELLPVSFNNDNSEW ISIVPNFILVRNTLISNIEIGFCLITKRSVICNQDYATPMTNNMRECLTGSTEKCPREL VVSSHVPRFALSNGVLFANCISVTCQCQTTGRAISQSGEQTLLMIDNTTCPTAVLG NVnSLGKYLGSVNYNSEGIAIGPPVFTDKVDISSQISSMNQSLQQSKDYIKEAQRL LDTVNPSLISMLSMIILYVLSIASLCIGLITFISFIIVEKKRNT) or an amino acid sequence at least 95%, 96%, 97%, 98%, or 99% identical thereto and a nipah vims G (NiV- G) polypeptide (e.g., SEQ ID NO: 9:

[0241] MKKINEGLLDSKILSAFNTVIALLGSIVUVMNIMIIQNYTRSTDNQAVIKDALQGIQ QQIKGLADKIGTEIGPKVSLIDTSSTITIPANIGLLGSKISQSTASINENVNEKCKFTL PPLKIHECNISCPNPLPFREYRPQTEGVSNLVGLPNNICLQKTSNQILKPKLISYTLP VVGQSGTCITDPLLAMDEGYFAYSHLERIGSCSRGVSKQRIIGVGEVLDRGDEVPS LFMTNVWTPPNPNTVYHCSAVYNNEFYYVLCAVSTVGDPILNSTYWSGSLMMT RLAVKPKSNGGGYNQHQLALRSIEKGRYDKVMPYGPSGIKQGDTLYFPAVGFLV RTEFKYNDSNCPITKCQYSKPENCRLSMGIRPNSHYILRSGLLKYNLSDGENPKVV FIEISDQRLSIGSPSKIYDSLGQPVFYQASFSWDTMIKFGDVLTVNPLVVNWRNNT VISRPGQSQCPRFNTCPEICWEGVYNDAFLIDRINWISAGVFLDSNQTAENPVFTV FKDNEILYRAQLASEDTNAQKTITNCFLLKNKIWCISLVEIYDTGDNVIRPKLFAV KIPEQCT) or an amino acid sequence at least 95%, 96%, 97%, 98%, or 99% identical thereto, wherein the NiV-G polypeptide lacks cell binding activity and retains fusogenic activity. In particular embodiments, the NiV-G polypeptide comprises one or more amino acid substitutions at positions E468, W471, Q497, and E500 of an NiV-G polypeptide (e.g., SEQ ID NO: 9). In particular embodiments, the NiV-G polypeptide comprises one or more of the amino acid substitutions E468A, W471A, Q497A, and E500A in an NiV-G polypeptide SEQ ID NO: 10:

[0242] MKKINEGLLDSKILSAFNTVIALLGSIVUVMNIMIIQNYTRSTDNQAVIKDALQGIQ QQIKGLADKIGTEIGPKVSLIDTSSTITIPANIGLLGSKISQSTASINENVNEKCKFTL PPLKIHECNISCPNPLPFREYRPQTEGVSNLVGLPNNICLQKTSNQILKPKLISYTLP VVGQSGTCITDPLLAMDEGYFAYSHLERIGSCSRGVSKQRIIGVGEVLDRGDEVPS LFMTNVWTPPNPNTVYHCSAVYNNEFYYVLCAVSTVGDPILNSTYWSGSLMMT RLAVKPKSNGGGYNQHQLALRSIEKGRYDKVMPYGPSGIKQGDTLYFPAVGFLV RTEFKYNDSNCPITKCQYSKPENCRLSMGIRPNSHYILRSGLLKYNLSDGENPKVV FIEISDQRLSIGSPSKIYDSLGQPVFYQASFSWDTMIKFGDVLTVNPLVVNWRNNT VISRPGQSQCPRFNTCPAICAEGVYNDAFLIDRINWISAGVFLDSNATAANPVFTV FKDNEILYRAQLASEDTNAQKTITNCFLLKNKIWCISLVEIYDTGDNVIRPKLFAV KIPEQCT) or an amino acid sequence at least 95%, 96%, 97%, 98%, or 99% identical thereto, wherein the NiV-G polypeptide lacks cell binding activity and retains fusogenic activity.

[0243] 3. TROPISM POLYPEPTIDES

[0244] Recombinant particles contemplated herein comprise one or more tropism polypeptides that govern particle targeting. Without wishing to be bound by any particular theory, it is contemplated that decoupling particle fusion and tropism solves specificity and delivery issues associated with in vivo gene therapy and gene editing. In particular embodiments, recombinant particles comprise one or more viral envelope glycoproteins modified to retain cell fusion activity and substantially ablate or ablate native tropism activity and further comprise one or more non-viral membrane-bound tropism polypeptides to support particle targeting. Decoupling particle fusion and tropism activities enables particle on-target delivery to the intended cells and decreases off-target delivery to other cells thereby increasing the safety and efficacy of the therapy.

[0245] In particular embodiments, a recombinant particle contemplated herein comprises an outer surface comprising, expressing, or displaying one or more non- viral membrane bound tropism polypeptides. A “tropism polypeptide” is a polypeptide that binds one or more antigens on a desired cell type. A “non-viral membrane bound tropism polypeptide” is a polypeptide that binds one or more antigens on a desired cell type; that is not native to, or derived from, either in whole or in part, a virus.

[0246] In particular embodiments, a recombinant particle comprises an outer surface comprising one or more mutated viral envelope glycoproteins that mediate fusion of the particle with a cell but that do not bind its cognate receptor expressed on a cell, and one or more non-viral membrane-bound tropism polypeptides that direct the particle to bind a specific antigen expressed on a cell, e.g., an immune effector cell. In particular embodiments, a recombinant particle contemplated herein comprises an outer surface comprising, expressing or displaying a primary non-viral membrane bound tropism polypeptide and optionally, a secondary non-viral membrane bound tropism polypeptide.

[0247] In particular embodiments, a recombinant particle, e.g., retroviral or lentiviral particle, comprises a lipid bilayer, cell membrane, or viral envelope comprising one or more mutated viral envelope glycoproteins that mediate fusion of the viral particle with a cell but that do not bind its cognate receptor expressed on a cell, and one or more non-viral membrane-bound tropism polypeptides, e.g., a primary and optionally a secondary non-viral membrane bound tropism polypeptide, that direct the particle to bind one or more specific antigens expressed on a cell, e.g., an immune effector cell.

[0248] In particular embodiments, a non-viral membrane bound tropism polypeptide comprises an extracellular antigen targeting domain, a spacer polypeptide, a transmembrane domain or GPI anchor, and optionally an intracellular domain. In particular embodiments, a non-viral membrane bound tropism polypeptide comprises an intracellular domain of no more than one, two, three, four, five, six, seven, eight, nine, or ten amino acids.

[0249] In particular embodiments, a non-viral membrane bound tropism polypeptide comprises an amino acid sequence set forth in any one of SEQ ID NOs: 455-576.

[0250] In particular embodiments, a non-viral membrane bound tropism polypeptide comprises an amino acid sequence set forth in any one of SEQ ID NOs: 471, 472, 473, 474, 475, 476, 477, 478, 479, 480, 481, 482, 483, 484, 485, and 486. In particular embodiments, a non-viral membrane bound tropism polypeptide comprises an amino acid sequence set forth in any one of SEQ ID NOs: 535, 536, 537, 538, 539, 540, 541, 542, 543, 544, 545, 546, 547, 548, 549, 550, 551, 552, 553, and 554.

[0251] In particular embodiments, a non-viral membrane bound tropism polypeptide comprises an amino acid sequence set forth in any one of SEQ ID NOs: 555, 556, 557, 558, 559, 560, 561, and 562.

[0252] In particular embodiments, a non-viral membrane bound tropism polypeptide comprises an amino acid sequence set forth in any one of SEQ ID NOs: 563, 564, 564, 566, 567, 568, 569, 570, 571, 572, 573, 574, 575, and 576.

[0253] In particular embodiments, a non-viral membrane bound tropism polypeptide comprises an amino acid sequence set forth in SEQ ID NO: 471 and SEQ ID NO: 472.

[0254] In particular embodiments, a non-viral membrane bound tropism polypeptide comprises an amino acid sequence set forth in SEQ ID NO: 553 and SEQ ID NO: 554.

[0255] In particular embodiments, a non-viral membrane bound tropism polypeptide comprises an amino acid sequence set forth in SEQ ID NO: 563 and SEQ ID NO: 564. a. Extracellular Antigen Targeting Domain

[0256] Recombinant particles contemplated in particular embodiments comprise a detargeted fusogen and one or more non-viral membrane bound tropism polypeptides that comprise an extracellular antigen targeting domain that binds one or more antigens specifically or selectively expressed on a particular cell type, e.g., an immune effector cell. As used herein, an “extracellular antigen targeting domain” (also referred to as an “extracellular targeting domain,” “antigen targeting domain,” or “targeting domain”) refers to any naturally occurring, synthetic, semi-synthetic, or recombinantly produced binding partner that binds a biological molecule or target antigen expressed or displayed on the surface of a cell. In particular embodiments, an extracellular antigen targeting domain binds an antigen expressed on a desired or specified cell type. In some embodiments, expression of the antigen is specific to the cell. In preferred embodiments, an extracellular antigen targeting domain binds an antigen expressed on an immune effector cell.

[0257] In various embodiments, a non-viral membrane bound tropism polypeptide comprises one or more extracellular targeting domains selected from the group consisting of: an antibody, a ligand (e.g., cytokines, chemokines, or cell surface associated ligands), a receptor ectodomain (e.g., c-Met, EGFR), or an aptamer.

[0258] In particular embodiments, an extracellular antigen targeting domain comprises an antibody or antigen binding fragment thereof that binds an antigen expressed on the surface of a cell, e.g., an immune effector cell. In certain embodiments, an extracellular antigen targeting domain comprises an antibody or antigen binding fragment thereof selected from the group consisting of: a Camel Ig, a Llama Ig, an Alpaca Ig, Ig NAR, a Fab' fragment, a F(ab')2 fragment, a bispecific Fab dimer (Fab2), a trispecific Fab trimer (Fab3), an Fv, a single chain Fv protein (scFv), a bis-scFv, (scFv , a minibody, a diabody, a triabody, a tetrabody, a disulfide stabilized Fv protein (dsFv), a Fibronectin type III (FN3) domain antibody, a single-domain antibody (sdAb or nanobody, e.g., a camelid VHH or shark VNAR), and a centyrin; a spacer polypeptide; a transmembrane domain or GPI anchor; and optionally an intracellular domain.

[0259] In particular embodiments, a non-viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain that binds an antigen expressed on an immune effector cell selected from the group consisting of: alpha, beta, gamma, or delta chain of the T-cell receptor, CDla, CDlb, CDlc, CD2, CD35, CD3s CD3y, CD4, CD5, CD6, CD7, CD8a, CD8p, CDlla, CDllb, CDllc, CD25, CD26, CD27, CD28, CD30, CD35, CD37, CD38, CD39, CD43, CD45RO, CD45RA, CD45RB, CD45RAB, CD45RBC, CD45ABC, CD49a, CD49f, CD49d, CD56, CD57, CD62L, CD69, CD70, CD73, CD74, CDw75, CDw76, CD80 (B7-1), CD83, CDw84, CD86 (B7-2), CD89, CD94, CD95, CD96, CD98, CD99, CD100, CDwlOl, CD102, CD103, CD104, CDwlO8, CDwl21a (IL-1RI), CD122 (IL-2RP), CDwl24( IL-4R), CDwl27 (IL-7R), CDwl28 (IL-8R), CD134, CD137, CD152 (CTLA-4), CD154 (CD40L), CD161, CD183, CD184, CD185, CD193, CD194, CD195, CD196, CD197, CD223 (LAG-3), CD244 (SlamF4), CD270 (TNFRSF14; HVEM), CD272 (BTLA-4), CD273 (PD-L2), CD274 (PD-L1), CD278 (ICOS), CD279, CD300, CD357 (TNFRSF18; GUR), CMRF35, CCR10, IL-2RY, IL-10R, IL-12R, IL-13Ral, P2RX7, Sca-1, SlamF6, TIGIT, TIM-3, TNRF2, and TNFRS25; a spacer polypeptide; a transmembrane domain or GPI anchor; and optionally an intracellular domain.

[0260] In particular embodiments, a non-viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain that binds an antigen expressed on an immune effector cell selected from the group consisting of: the alpha, beta, gamma, or delta chain of the T cell receptor, CD2, CD35, CD3s CD3y, CD4, CD7, CD8a, and CD8P; a spacer polypeptide; a transmembrane domain or GPI anchor; and optionally an intracellular domain.

[0261] In particular embodiments, a non-viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain comprising an scFv, a murine scFv, a humanized scFv, a human scFv, or one or more VHH or VNAR single domain antibodies that binds an antigen expressed on an immune effector cell selected from the group consisting of: the alpha, beta, gamma, or delta chain of the T cell receptor, CD2, CD35, CD3s CD3y, CD4, CD7, CD8a, and CD8P; a spacer polypeptide; a transmembrane domain or GPI anchor; and optionally an intracellular domain.

[0262] In particular embodiments, a non-viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain comprising an anti-CD3s antibody or antigen binding fragment thereof. Illustrative examples of anti-CD3s antibodies or antigen binding fragments thereof suitable for use in particular embodiments contemplated herein include but are not limited to scFvs or other antigen binding fragments isolated from 0KT3, UCHT1, YTH12.5, and TR66, and variants thereof, having at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, or at least 99% identity thereto.

[0263] In particular embodiments, a non-viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain comprising an anti-CD3s antibody or antigen binding fragment thereof comprising a CDRH1, a CDRH2, and a CDRH3 and a CDRL1, a CDRL2, and a CDRL3 of an antibody or antigen binding fragment thereof set forth in Table 5; a spacer polypeptide; a transmembrane domain or GPI anchor; and optionally an intracellular domain.

[0264] In particular embodiments, a non-viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain comprising an anti-CD3s antibody or antigen binding fragment thereof comprising: CDRH1, CDRH2, and CDRH3 amino acid sequences set forth in SEQ ID NOs: 12-14, and CDRL1, CDRL2, and CDRL3 amino acid sequences set forth in SEQ ID NOs: 16-18; CDRH1, CDRH2, and CDRH3 amino acid sequences set forth in SEQ ID NOs: 22-24, and CDRL1, CDRL2, and CDRL3 amino acid sequences set forth in SEQ ID NOs: 26-28; CDRH1, CDRH2, and CDRH3 amino acid sequences set forth in SEQ ID NOs: 32-34, and CDRL1, CDRL2, and CDRL3 amino acid sequences set forth in SEQ ID NOs: 36-38; CDRH1, CDRH2, and CDRH3 amino acid sequences set forth in SEQ ID NOs: 42-44, and CDRL1, CDRL2, and CDRL3 amino acid sequences set forth in SEQ ID NOs: 46-48; CDRH1, CDRH2, and CDRH3 amino acid sequences set forth in SEQ ID NOs: 52-54, and CDRL1, CDRL2, and CDRL3 amino acid sequences set forth in SEQ ID NOs: 56-58; CDRH1, CDRH2, and CDRH3 amino acid sequences set forth in SEQ ID NOs: 62-64, and CDRL1, CDRL2, and CDRL3 amino acid sequences set forth in SEQ ID NOs: 66-68; CDRH1, CDRH2, and CDRH3 amino acid sequences set forth in SEQ ID NOs: 72-74, and CDRL1, CDRL2, and CDRL3 amino acid sequences set forth in SEQ ID NOs: 76-78; CDRH1, CDRH2, and CDRH3 amino acid sequences set forth in SEQ ID NOs: 82-84, and CDRL1, CDRL2, and CDRL3 amino acid sequences set forth in SEQ ID NOs: 86-88; CDRH1, CDRH2, and CDRH3 amino acid sequences set forth in SEQ ID NOs: 92-94, and CDRL1, CDRL2, and CDRL3 amino acid sequences set forth in SEQ ID NOs: 96-98; CDRH1, CDRH2, and CDRH3 amino acid sequences set forth in SEQ ID NOs: 102-104, and CDRL1, CDRL2, and CDRL3 amino acid sequences set forth in SEQ ID NOs: 106-108; CDRH1, CDRH2, and CDRH3 amino acid sequences set forth in SEQ ID NOs: 112-114, and CDRL1, CDRL2, and CDRL3 amino acid sequences set forth in SEQ ID NOs: 116-118; CDRH1, CDRH2, and CDRH3 amino acid sequences set forth in SEQ ID NOs: 122-124, and CDRL1, CDRL2, and CDRL3 amino acid sequences set forth in SEQ ID NOs: 126-128; CDRH1, CDRH2, and CDRH3 amino acid sequences set forth in SEQ ID NOs: 132-134, and CDRL1, CDRL2, and CDRL3 amino acid sequences set forth in SEQ ID NOs: 136-138; CDRH1, CDRH2, and CDRH3 amino acid sequences set forth in SEQ ID NOs: 142-144, and CDRL1, CDRL2, and CDRL3 amino acid sequences set forth in SEQ ID NOs: 146-148; CDRH1, CDRH2, and CDRH3 amino acid sequences set forth in SEQ ID NOs: 152-154, and CDRL1, CDRL2, and CDRL3 amino acid sequences set forth in SEQ ID NOs: 156-158; CDRH1, CDRH2, and CDRH3 amino acid sequences set forth in SEQ ID NOs: 162-164, and CDRL1, CDRL2, and CDRL3 amino acid sequences set forth in SEQ ID NOs: 166-168; CDRH1, CDRH2, and CDRH3 amino acid sequences set forth in SEQ ID NOs: 172-174, and CDRL1, CDRL2, and CDRL3 amino acid sequences set forth in SEQ ID NOs: 176-178; CDRH1, CDRH2, and CDRH3 amino acid sequences set forth in SEQ ID NOs: 182-184, and CDRL1, CDRL2, and CDRL3 amino acid sequences set forth in SEQ ID NOs: 186-188; CDRH1, CDRH2, and CDRH3 amino acid sequences set forth in SEQ ID NOs: 192-194, and CDRL1, CDRL2, and CDRL3 amino acid sequences set forth in SEQ ID NOs: 196-198; CDRH1, CDRH2, and CDRH3 amino acid sequences set forth in SEQ ID NOs: 202-204, and CDRL1, CDRL2, and CDRL3 amino acid sequences set forth in SEQ ID NOs: 206-208; CDRH1, CDRH2, and CDRH3 amino acid sequences set forth in SEQ ID NOs: 212-214, and CDRL1, CDRL2, and CDRL3 amino acid sequences set forth in SEQ ID NOs: 216-218; CDRH1, CDRH2, and CDRH3 amino acid sequences set forth in SEQ ID NOs: 222-224, and CDRL1, CDRL2, and CDRL3 amino acid sequences set forth in SEQ ID NOs: 226-228; CDRH1, CDRH2, and CDRH3 amino acid sequences set forth in SEQ ID NOs: 232-234, and CDRL1, CDRL2, and CDRL3 amino acid sequences set forth in SEQ ID NOs: 236-238; CDRH1, CDRH2, and CDRH3 amino acid sequences set forth in SEQ ID NOs: 242-244, and CDRL1, CDRL2, and CDRL3 amino acid sequences set forth in SEQ ID NOs: 246-248; CDRH1, CDRH2, and CDRH3 amino acid sequences set forth in SEQ ID NOs: 252-254, and CDRL1, CDRL2, and CDRL3 amino acid sequences set forth in SEQ ID NOs: 256-258; CDRH1, CDRH2, and CDRH3 amino acid sequences set forth in SEQ ID NOs: 262-264, and CDRL1, CDRL2, and CDRL3 amino acid sequences set forth in SEQ ID NOs: 266-268; CDRH1, CDRH2, and CDRH3 amino acid sequences set forth in SEQ ID NOs: 272-274, and CDRL1, CDRL2, and CDRL3 amino acid sequences set forth in SEQ ID NOs: 276-278; CDRH1, CDRH2, and CDRH3 amino acid sequences set forth in SEQ ID NOs: 282-284, and CDRL1, CDRL2, and CDRL3 amino acid sequences set forth in SEQ ID NOs: 286-288; CDRH1, CDRH2, and CDRH3 amino acid sequences set forth in SEQ ID NOs: 292-294, and CDRL1, CDRL2, and CDRL3 amino acid sequences set forth in SEQ ID NOs: 296-298; CDRH1, CDRH2, and CDRH3 amino acid sequences set forth in SEQ ID NOs: 302-304, and CDRL1, CDRL2, and CDRL3 amino acid sequences set forth in SEQ ID NOs: 306-308; CDRH1, CDRH2, and CDRH3 amino acid sequences set forth in SEQ ID NOs: 312-314, and CDRLl, CDRL2, andCDRL3 amino acid sequences set forth in SEQ ID NOs: 316-318; CDRH1, CDRH2, and CDRH3 amino acid sequences set forth in SEQ ID NOs: 322-324, and CDRL1, CDRL2, and CDRL3 amino acid sequences set forth in SEQ ID NOs: 326-328; CDRH1, CDRH2, and CDRH3 amino acid sequences set forth in SEQ ID NOs: 332-334, and CDRL1, CDRL2, and CDRL3 amino acid sequences set forth in SEQ ID NOs: 336-338; CDRH1, CDRH2, and CDRH3 amino acid sequences set forth in SEQ ID NOs: 342-344, and CDRL1, CDRL2, and CDRL3 amino acid sequences set forth in SEQ ID NOs: 346-348; CDRH1, CDRH2, and CDRH3 amino acid sequences set forth in SEQ ID NOs: 352-354, and CDRL1, CDRL2, and CDRL3 amino acid sequences set forth in SEQ ID NOs: 356-358; CDRH1, CDRH2, and CDRH3 amino acid sequences set forth in SEQ ID NOs: 362-364, and CDRL1, CDRL2, and CDRL3 amino acid sequences set forth in SEQ ID NOs: 366-368; CDRH1, CDRH2, and CDRH3 amino acid sequences set forth in SEQ ID NOs: 372-374, and CDRL1, CDRL2, and CDRL3 amino acid sequences set forth in SEQ ID NOs: 376-378; CDRH1, CDRH2, and CDRH3 amino acid sequences set forth in SEQ ID NOs: 382-384, and CDRL1, CDRL2, and CDRL3 amino acid sequences set forth in SEQ ID NOs: 386-388; CDRH1, CDRH2, and CDRH3 amino acid sequences set forth in SEQ ID NOs: 392-394, and CDRL1, CDRL2, and CDRL3 amino acid sequences set forth in SEQ ID NOs: 396-398; or CDRH1, CDRH2, and CDRH3 amino acid sequences set forth in SEQ ID NOs: 402-404, and CDRL1, CDRL2, and CDRL3 amino acid sequences set forth in SEQ ID NOs: 406-408; a spacer polypeptide; a transmembrane domain or GPI anchor; and optionally an intracellular domain.

[0265] In particular embodiments, a non-viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain comprising an anti-CD3s antibody or antigen binding fragment thereof comprising CDRH1, CDRH2, and CDRH3 amino acid sequences set forth in SEQ ID NOs: 92-94, and CDRL1, CDRL2, and CDRL3 amino acid sequences set forth in SEQ ID NOs: 96-98; a spacer polypeptide; a transmembrane domain or GPI anchor; and optionally an intracellular domain.

[0266] In particular embodiments, a non-viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain comprising an anti-CD3s antibody or antigen binding fragment thereof comprising CDRH1, CDRH2, and CDRH3 amino acid sequences set forth in SEQ ID NOs: 102-104, and CDRL1, CDRL2, and CDRL3 amino acid sequences setforth in SEQ ID NOs: 106-108; a spacer polypeptide; a transmembrane domain or GPI anchor; and optionally an intracellular domain.

[0267] In particular embodiments, a non-viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain comprising an anti-CD3s antibody or antigen binding fragment thereof comprising CDRH1, CDRH2, and CDRH3 amino acid sequences set forth in SEQ ID NOs: 112-114, and CDRL1, CDRL2, and CDRL3 amino acid sequences set forth in SEQ ID NOs: 116- 118 ; a spacer polypeptide; a transmembrane domain or GPI anchor; and optionally an intracellular domain.

[0268] In particular embodiments, a non-viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain comprising an anti-CD3s antibody or antigen binding fragment thereof comprising CDRH1, CDRH2, and CDRH3 amino acid sequences set forth in SEQ ID NOs: 122-124, and CDRL1, CDRL2, and CDRL3 amino acid sequences setforth in SEQ ID NOs: 126-128; a spacer polypeptide; a transmembrane domain or GPI anchor; and optionally an intracellular domain.

[0269] In particular embodiments, a non-viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain comprising an anti-CD3s antibody or antigen binding fragment thereof comprising CDRH1, CDRH2, and CDRH3 amino acid sequences set forth in SEQ ID NOs: 132-134, and CDRL1, CDRL2, and CDRL3 amino acid sequences setforth in SEQ ID NOs: 136-138; a spacer polypeptide; a transmembrane domain or GPI anchor; and optionally an intracellular domain.

[0270] In particular embodiments, a non-viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain comprising an anti-CD3s antibody or antigen binding fragment thereof comprising CDRH1, CDRH2, and CDRH3 amino acid sequences set forth in SEQ ID NOs: 142-144, and CDRL1, CDRL2, and CDRL3 amino acid sequences setforth in SEQ ID NOs: 146-148; a spacer polypeptide; a transmembrane domain or GPI anchor; and optionally an intracellular domain.

[0271] In particular embodiments, a non-viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain comprising an anti-CD3s antibody or antigen binding fragment thereof comprising CDRH1, CDRH2, and CDRH3 amino acid sequences set forth in SEQ ID NOs: 152-154, and CDRL1, CDRL2, and CDRL3 amino acid sequences setforth in SEQ ID NOs: 156-158; a spacer polypeptide; a transmembrane domain or GPI anchor; and optionally an intracellular domain.

[0272] In particular embodiments, a non-viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain comprising an anti-CD3s antibody or antigen binding fragment thereof comprising CDRH1, CDRH2, and CDRH3 amino acid sequences set forth in SEQ ID NOs: 162-164, and CDRL1, CDRL2, and CDRL3 amino acid sequences setforth in SEQ ID NOs: 166-168; a spacer polypeptide; a transmembrane domain or GPI anchor; and optionally an intracellular domain.

[0273] In particular embodiments, a non-viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain comprising an anti-CD3s antibody or antigen binding fragment thereof comprising a heavy chain variable (VH) domain and a light chain variable (VL) domain of an antibody or antigen binding fragment thereof set forth in Table 5; a polypeptide linker disposed between the VH domain and the VL domain; a spacer polypeptide; a transmembrane domain or GPI anchor; and optionally an intracellular domain. In particular embodiments the polypeptide linker is selected from the group consisting of: (GGGGS)n wherein n = 1, 2, 3, 4 or 5; S(GGGGS)nwherein n = 1, 2, 3, 4 or 5; GEGTSTGSGGSGGSGGAD, GSTSGSGKPGSGEGSTKG and variants thereof comprising an amino acid sequence at least 90% identical thereto. In particular embodiments, a non-viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain comprising an anti-CD3s antibody or antigen binding fragment thereof comprising: a VH domain comprising an amino acid sequence set forth in SEQ ID NO: 11 and a VL domain comprising an amino acid sequence set forth in SEQ ID NO: 15; a VH domain comprising an amino acid sequence set forth in SEQ ID NO: 21 and a VL domain comprising an amino acid sequence set forth in SEQ ID NO: 25; a VH domain comprising an amino acid sequence set forth in SEQ ID NO: 31 and a VL domain comprising an amino acid sequence set forth in SEQ ID NO: 35; a VH domain comprising an amino acid sequence set forth in SEQ ID NO: 41 and a VL domain comprising an amino acid sequence set forth in SEQ ID NO: 45; a VH domain comprising an amino acid sequence set forth in SEQ ID NO: 51 and a VL domain comprising an amino acid sequence set forth in SEQ ID NO: 55; a VH domain comprising an amino acid sequence set forth in SEQ ID NO: 61 and a VL domain comprising an amino acid sequence set forth in SEQ ID NO: 65; a VH domain comprising an amino acid sequence set forth in SEQ ID NO: 71 and a VL domain comprising an amino acid sequence set forth in SEQ ID NO: 75; a VH domain comprising an amino acid sequence set forth in SEQ ID NO: 81 and a VL domain comprising an amino acid sequence set forth in SEQ ID NO: 85; a VH domain comprising an amino acid sequence set forth in SEQ ID NO: 91 and a VL domain comprising an amino acid sequence set forth in SEQ ID NO: 95; a VH domain comprising an amino acid sequence set forth in SEQ ID NO: 101 and a VL domain comprising an amino acid sequence set forth in SEQ ID NO: 105; a VH domain comprising an amino acid sequence set forth in SEQ ID NO: 111 and a VL domain comprising an amino acid sequence set forth in SEQ ID NO: 115; a VH domain comprising an amino acid sequence set forth in SEQ ID NO: 121 and a VL domain comprising an amino acid sequence set forth in SEQ ID NO: 125; a VH domain comprising an amino acid sequence set forth in SEQ ID NO: 131 and a VL domain comprising an amino acid sequence set forth in SEQ ID NO: 135; a VH domain comprising an amino acid sequence set forth in SEQ ID NO: 141 and a VL domain comprising an amino acid sequence set forth in SEQ ID NO: 145; a VH domain comprising an amino acid sequence set forth in SEQ ID NO: 151 and a VL domain comprising an amino acid sequence set forth in SEQ ID NO: 155; a VH domain comprising an amino acid sequence set forth in SEQ ID NO: 161 and a VL domain comprising an amino acid sequence set forth in SEQ ID NO: 165; a VH domain comprising an amino acid sequence set forth in SEQ ID NO: 171 and a VL domain comprising an amino acid sequence set forth in SEQ ID NO: 175; a VH domain comprising an amino acid sequence set forth in SEQ ID NO: 181 and a VL domain comprising an amino acid sequence set forth in SEQ ID NO: 185; a VH domain comprising an amino acid sequence set forth in SEQ ID NO: 191 and a VL domain comprising an amino acid sequence set forth in SEQ ID NO: 195; a VH domain comprising an amino acid sequence set forth in SEQ ID NO: 201 and a VL domain comprising an amino acid sequence set forth in SEQ ID NO: 205; a VH domain comprising an amino acid sequence set forth in SEQ ID NO: 211 and a VL domain comprising an amino acid sequence set forth in SEQ ID NO: 215; a VH domain comprising an amino acid sequence set forth in SEQ ID NO: 221 and a VL domain comprising an amino acid sequence set forth in SEQ ID NO: 225; a VH domain comprising an amino acid sequence set forth in SEQ ID NO: 231 and a VL domain comprising an amino acid sequence set forth in SEQ ID NO: 235; a VH domain comprising an amino acid sequence set forth in SEQ ID NO: 241 and a VL domain comprising an amino acid sequence set forth in SEQ ID NO: 245; a VH domain comprising an amino acid sequence set forth in SEQ ID NO: 251 and a VL domain comprising an amino acid sequence set forth in SEQ ID NO: 255; a VH domain comprising an amino acid sequence set forth in SEQ ID NO: 261 and a VL domain comprising an amino acid sequence set forth in SEQ ID NO: 265; a VH domain comprising an amino acid sequence set forth in SEQ ID NO: 271 and a VL domain comprising an amino acid sequence set forth in SEQ ID NO: 275; a VH domain comprising an amino acid sequence set forth in SEQ ID NO: 281 and a VL domain comprising an amino acid sequence set forth in SEQ ID NO: 285; a VH domain comprising an amino acid sequence set forth in SEQ ID NO: 291 and a VL domain comprising an amino acid sequence set forth in SEQ ID NO: 295; a VH domain comprising an amino acid sequence set forth in SEQ ID NO: 301 and a VL domain comprising an amino acid sequence set forth in SEQ ID NO:305; a VH domain comprising an amino acid sequence set forth in SEQ ID NO: 311 and a VL domain comprising an amino acid sequence set forth in SEQ ID NO: 315; a VH domain comprising an amino acid sequence set forth in SEQ ID NO: 321 and a VL domain comprising an amino acid sequence set forth in SEQ ID NO: 325; a VH domain comprising an amino acid sequence set forth in SEQ ID NO: 331 and a VL domain comprising an amino acid sequence set forth in SEQ ID NO: 335; a VH domain comprising an amino acid sequence set forth in SEQ ID NO: 341 and a VL domain comprising an amino acid sequence set forth in SEQ ID NO: 345; a VH domain comprising an amino acid sequence set forth in SEQ ID NO: 351 and a VL domain comprising an amino acid sequence set forth in SEQ ID NO: 355; a VH domain comprising an amino acid sequence set forth in SEQ ID NO: 361 and a VL domain comprising an amino acid sequence set forth in SEQ ID NO: 365; a VH domain comprising an amino acid sequence set forth in SEQ ID NO: 371 and a VL domain comprising an amino acid sequence set forth in SEQ ID NO: 375; a VH domain comprising an amino acid sequence set forth in SEQ ID NO: 381 and a VL domain comprising an amino acid sequence set forth in SEQ ID NO: 385; a VH domain comprising an amino acid sequence set forth in SEQ ID NO: 391 and a VL domain comprising an amino acid sequence set forth in SEQ ID NO: 395; or a VH domain comprising an amino acid sequence set forth in SEQ ID NO: 401 and a VL domain comprising an amino acid sequence set forth in SEQ ID NO:405; a polypeptide linker disposed between the VH domain and the VH domain; a spacer polypeptide; a transmembrane domain or GPI anchor; and optionally an intracellular domain. In particular embodiments the polypeptide linker is selected from the group consisting of: (GGGGS)nwherein n = 1, 2, 3, 4 or 5; S(GGGGS)nwherein n = 1, 2, 3, 4 or 5; GEGTSTGSGGSGGSGGAD, GSTSGSGKPGSGEGSTKG and variants thereof comprising an amino acid sequence at least 90% identical thereto.

[0274] In particular embodiments, a non-viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain comprising an anti-CD3s antibody or antigen binding fragment thereof comprising: a VH domain comprising an amino acid sequence set forth in SEQ ID NO: 91 and a VL domain comprising an amino acid sequence set forth in SEQ ID NO: 95; a polypeptide linker disposed between the VH domain and the VL domain; a spacer polypeptide; a transmembrane domain or GPI anchor; and optionally an intracellular domain.

[0275] In particular embodiments, a non-viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain comprising an anti-CD3s antibody or antigen binding fragment thereof comprising: a VH domain comprising an amino acid sequence set forth in SEQ ID NO: 101 and a VL domain comprising an amino acid sequence set forth in SEQ ID NO: 105; a polypeptide linker disposed between the VH domain and the VL domain; a spacer polypeptide; a transmembrane domain or GPI anchor; and optionally an intracellular domain.

[0276] In particular embodiments, a non-viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain comprising an anti-CD3s antibody or antigen binding fragment thereof comprising: a VH domain comprising an amino acid sequence set forth in SEQ ID NO: 111 and a VL domain comprising an amino acid sequence set forth in SEQ ID NO: 115; a polypeptide linker disposed between the VH domain and the VL domain; a spacer polypeptide; a transmembrane domain or GPI anchor; and optionally an intracellular domain.

[0277] In particular embodiments, a non-viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain comprising an anti-CD3s antibody or antigen binding fragment thereof comprising: a VH domain comprising an amino acid sequence set forth in SEQ ID NO: 121 and a VL domain comprising an amino acid sequence set forth in SEQ ID NO: 125; a polypeptide linker disposed between the VH domain and the VL domain; a spacer polypeptide; a transmembrane domain or GPI anchor; and optionally an intracellular domain.

[0278] In particular embodiments, a non-viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain comprising an anti-CD3s antibody or antigen binding fragment thereof comprising: a VH domain comprising an amino acid sequence set forth in SEQ ID NO: 131 and a VL domain comprising an amino acid sequence set forth in SEQ ID NO: 135; a polypeptide linker disposed between the VH domain and the VL domain; a spacer polypeptide; a transmembrane domain or GPI anchor; and optionally an intracellular domain.

[0279] In particular embodiments, a non-viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain comprising an anti-CD3s antibody or antigen binding fragment thereof comprising: a VH domain comprising an amino acid sequence set forth in SEQ ID NO: 141 and a VL domain comprising an amino acid sequence set forth in SEQ ID NO: 145; a polypeptide linker disposed between the VH domain and the VL domain; a spacer polypeptide; a transmembrane domain or GPI anchor; and optionally an intracellular domain.

[0280] In particular embodiments, a non-viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain comprising an anti-CD3s antibody or antigen binding fragment thereof comprising: a VH domain comprising an amino acid sequence set forth in SEQ ID NO: 151 and a VL domain comprising an amino acid sequence set forth in SEQ ID NO: 155; a polypeptide linker disposed between the VH domain and the VL domain; a spacer polypeptide; a transmembrane domain or GPI anchor; and optionally an intracellular domain.

[0281] In particular embodiments, a non-viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain comprising an anti-CD3s antibody or antigen binding fragment thereof comprising: or a VH domain comprising an amino acid sequence set forth in SEQ ID NO: 161 and a VL domain comprising an amino acid sequence set forth in SEQ ID NO: 165; a polypeptide linker disposed between the VH domain and the VL domain; a spacer polypeptide; a transmembrane domain or GPI anchor; and optionally an intracellular domain.

[0282] In particular embodiments the polypeptide linker is selected from the group consisting of: (GGGGS)nwherein n = 1, 2, 3, 4 or 5; S(GGGGS)nwherein n = 1, 2, 3, 4 or 5; GEGTSTGSGGSGGSGGAD, GSTSGSGKPGSGEGSTKG and variants thereof comprising an amino acid sequence at least 90% identical thereto.

[0283] In particular embodiments, a non-viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain comprising an anti-CD3s antibody or antigen binding fragment thereof set forth in Table 5; a spacer polypeptide; a transmembrane domain or GPI anchor; and optionally an intracellular domain.

[0284] In particular embodiments, a non-viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain comprising an anti-CD3s antibody or antigen binding fragment thereof comprising the amino acid sequence set forth in any one of SEQ ID NOs: 19, 20, 29, 30, 39, 40, 49, 50, 59, 60, 69, 70, 79, 80, 89, 90, 99, 100, 109, 110, 119, 120, 129, 130, 139, 140, 149, 150, 159, 160, 169, 170, 179, 180, 189, 190, 199, 200,

[0285] 209, 210, 219, 220, 229, 230, 239, 240, 249, 250, 259, 260, 269, 270, 279, 280, 289, 290,

[0286] 299, 300, 309, 310, 319, 320, 329, 330, 339, 340, 349, 350, 359, 360, 369, 370, 379, 380,

[0287] 389, 390, 399, 400, 409, and 410; a spacer polypeptide; a transmembrane domain or GPI anchor; and optionally an intracellular domain.

[0288] In particular embodiments, a non-viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain comprising the amino acid sequence set forth in SEQ ID NO: 79. In particular embodiments, a non-viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain comprising the amino acid sequence set forth in SEQ ID NO: 80. In particular embodiments, a non-viral membranebound tropism polypeptide comprises an extracellular antigen targeting domain comprising the amino acid sequence set forth in SEQ ID NO: 89. In particular embodiments, a non-viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain comprising the amino acid sequence set forth in SEQ ID NO: 90. In particular embodiments, a non-viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain comprising the amino acid sequence set forth in SEQ ID NO: 99. In particular embodiments, a non-viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain comprising the amino acid sequence set forth in SEQ ID NO: 100. In particular embodiments, a non- viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain comprising the amino acid sequence set forth in SEQ ID NO: 109. In particular embodiments, a non- viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain comprising the amino acid sequence set forth in SEQ ID NO: 110. In particular embodiments, a non- viral membranebound tropism polypeptide comprises an extracellular antigen targeting domain comprising the amino acid sequence set forth in SEQ ID NO: 119. In particular embodiments, a non- viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain comprising the amino acid sequence set forth in SEQ ID NO: 120. In particular embodiments, a non- viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain comprising the amino acid sequence set forth in SEQ ID NO: 129. In particular embodiments, a non- viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain comprising the amino acid sequence set forth in SEQ ID NO: 130. In particular embodiments, a non-viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain comprising the amino acid sequence set forth in SEQ ID NO: 139. In particular embodiments, a non-viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain comprising the amino acid sequence set forth in SEQ ID NO: 140. In particular embodiments, a non-viral membranebound tropism polypeptide comprises an extracellular antigen targeting domain comprising the amino acid sequence set forth in SEQ ID NO: 149. In particular embodiments, a non- viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain comprising the amino acid sequence set forth in SEQ ID NO: 150. In particular embodiments, a non-viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain comprising the amino acid sequence set forth in SEQ ID NO: 159. In particular embodiments, a non-viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain comprising the amino acid sequence set forth in SEQ ID NO: 160. In particular embodiments, a non-viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain comprising the amino acid sequence set forth in SEQ ID NO: 169. In particular embodiments, a non-viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain comprising the amino acid sequence set forth in SEQ ID NO: 170.

[0289] Table 5 b. Spacer

[0290] Non-viral membrane-bound tropism polypeptides contemplated in particular embodiments comprise a spacer domain. In particular embodiments, a spacer domain impacts the manufacturability of the particle, the expression of the tropism polypeptide and the on-target / off-target cell binding profile of the tropism polypeptide. Without wishing to be bound to any particular theory, it is contemplated that in particular embodiments, disposing the extracellular targeting domains of the one or more non-viral membrane bound tropism polypeptides a particular distance from a particle surface (e.g., lipid bilayer, cell membrane, or envelope) out-competes the host cell derived membrane proteins and leads to increased targeting specificity of the vector. Increased vector specificity should correspond to efficacy at lower vector doses, which corresponds to increased safety. Lower doses also lead to manufacturing cost advantages and efficiencies, which has been an insurmountable obstacle for current ex vivo gene therapies. In particular embodiments, a particle comprises a non-viral membrane bound tropism polypeptide that comprises a spacer domain of about 12 nm in length, about 13 nm in length, about 14 nm in length, about 15 nm in length, about 16 nm in length, about 17 nm in length, about 18 nm in length, about 19 nm in length, about 20 nm in length, about 21 nm in length, about 22 nm in length, about 23 nm in length, about 24 nm in length, about 25 nm in length, about 26 nm in length, about 27 nm in length, about 28 nm in length, about 29 nm in length, about 30 nm in length, about 31 nm in length, about 32 nm in length, about 33 nm in length, about 34 nm in length, about 35 nm in length, about 36 nm in length, about 37 nm in length, about 38 nm in length, about 39 nm in length, about 40 nm in length, about 41 nm in length, about 42 nm in length, about 43 nm in length, about 44 nm in length, about 45 nm in length, about 46 nm in length, about 47 nm in length, about 48 nm in length, about 49 nm in length, about 50 nm in length, about 51 nm in length, about 52 nm in length, about 53 nm in length, about 54 nm in length, or about 55 nm in length, wherein the targeting domain is disposed about 12 nm to about 25 nm, about 13 nm to about 20 nm, or about 12 nm, about 13, nm, about 14 nm, about 15 nm, about 16 nm, about 17 nm, about 18 nm, about 19 nm, or about 20 nm from the vector surface.

[0291] In particular embodiments, a particle comprises a non-viral membrane bound tropism polypeptide that comprises a spacer domain about 12 nm in length to about 20 nm in length, wherein the extracellular antigen targeting domain is disposed at least about 12 nm, about 13, nm, about 14 nm, about 15 nm, about 16 nm, about 17 nm, about 18 nm, about 19 nm, or about 20 nm from the particle surface.

[0292] In particular embodiments, a particle comprises a tropism polypeptide that comprises a spacer comprising a hinge or stalk polypeptide about 12 nm in length, about 13 nm in length, about 14 nm in length, about 15 nm in length, about 16 nm in length, about 17 nm in length, about 18 nm in length, about 19 nm in length, or about 20 nm in length, wherein the targeting domain is disposed about 12 nm to about 20 nm from the particle surface.

[0293] In particular embodiments, a spacer comprises one, two, three, four, five, six, seven, eight, nine, ten or more domains are linked together to achieve the desired length and distance to dispose the extracellular targeting domain from the particle surface. The following table lists exemplary spacers domains based on hinge or stalk polypeptides and their predicted lengths.

[0294] Table 6

[0295] | Spacer | Es . Length | SEQUENCE |

[0296] In particular embodiments, a non-viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain; a spacer comprising a hinge or stalk domain derived, obtained, or isolated from PDGFR, IgGl, IgG2, IgG4, CD2, CD3, CD4, CD8a, CD8P, CD28, CD45ABC, CD45RO, CD45RAB, CD45RAC, CD45RBC, CD45RA, CD45RB, CD45RC, CD45del-d3, CD45del-d2-d3, CADHERIN 1 (E-CADHERIN; ECAD), CEACAM 5, variants thereof, an amino acid sequence at least 85% identical to, at least 90% identical to, at least 95 % identical to, at least 96% identical to, at least 97 % identical to, at least 98% identical to, or at least 99% identical thereto, and polypeptide linkers of similar amino acid composition and lengths; a transmembrane domain or GPI anchor, and optionally an intracellular domain.

[0297] In particular embodiments, a non-viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain comprising an anti-CD3s antibody or antigen binding fragment thereof comprising a CDRH1, CDRH2, CDRH3, CDRL1, CDRL2, and CDRL3; a VH and VL; and / or an scFv set forth in Table 5; a spacer comprising a hinge or stalk domain derived, obtained, or isolated from PDGFR, IgGl, IgG2, IgG4, CD2, CD3, CD4, CD8a, CD8p, CD28, CD45ABC, CD45RO, CD45RAB, CD45RAC, CD45RBC, CD45RA, CD45RB, CD45RC, CD45del-d3, CD45del-d2-d3, CADHERIN 1, CEACAM 5, or variants thereof; a transmembrane domain or GPI anchor, and optionally an intracellular domain.

[0298] In particular embodiments, a non-viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain comprising an anti-CD3s antibody or antigen binding fragment thereof comprising the amino acid sequence set forth in any one of SEQ ID NOs: 19, 20, 29, 30, 39, 40, 49, 50, 59, 60, 69, 70, 79, 80, 89, 90, 99, 100, 109, 110, 119, 120, 129, 130, 139, 140, 149, 150, 159, 160, 169, 170, 179, 180, 189, 190, 199, 200,

[0299] 209, 210, 219, 220, 229, 230, 239, 240, 249, 250, 259, 260, 269, 270, 279, 280, 289, 290,

[0300] 299, 300, 309, 310, 319, 320, 329, 330, 339, 340, 349, 350, 359, 360, 369, 370, 379, 380,

[0301] 389, 390, 399, 400, 409, and 410; a spacer comprising a hinge or stalk domain derived, obtained, or isolated from PDGFR, IgGl, IgG2, IgG4, CD2, CD3, CD4, CD8a, CD8P, CD28, CD45ABC, CD45RO, CD45RAB, CD45RAC, CD45RBC, CD45RA, CD45RB, CD45RC, CD45del-d3, CD45del-d2-d3, CADHERIN 1, CEACAM 5, or variants thereof; a transmembrane domain or GPI anchor, and optionally an intracellular domain.

[0302] In particular embodiments, a non-viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain comprising an anti-CD3s antibody or antigen binding fragment thereof comprising the amino acid sequence set forth in any one of SEQ ID NOs: 19, 20, 29, 30, 39, 40, 49, 50, 59, 60, 69, 70, 79, 80, 89, 90, 99, 100, 109, 110, 119, 120, 129, 130, 139, 140, 149, 150, 159, 160, 169, 170, 179, 180, 189, 190, 199, 200,

[0303] 209, 210, 219, 220, 229, 230, 239, 240, 249, 250, 259, 260, 269, 270, 279, 280, 289, 290,

[0304] 299, 300, 309, 310, 319, 320, 329, 330, 339, 340, 349, 350, 359, 360, 369, 370, 379, 380,

[0305] 389, 390, 399, 400, 409, and 410; a spacer comprising a hinge or stalk domain derived, obtained, or isolated from PDGFR, IgGl, IgG2, IgG4, CD2, CD3, CD4, CD8a, CD8P, CD28, CD45ABC, CD45RO, CD45RAB, CD45RAC, CD45RBC, CD45RA, CD45RB, CD45RC, CD45del-d3, CD45del-d2-d3, CADHERIN 1, CEACAM 5, or variants thereof; a transmembrane domain or GPI anchor, and optionally an intracellular domain.

[0306] In particular embodiments, a non-viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain comprising an anti-CD3s antibody or antigen binding fragment thereof comprising the amino acid sequence set forth in any one of SEQ ID NOs: 99, 100, 109, 110, 119, 120, 129, 130, 139, 140, 149, 150, 159, 160, 169, and 170; a spacer comprising a hinge or stalk domain derived, obtained, or isolated from PDGFR, IgGl, IgG2, IgG4, CD2, CD3, CD4, CD8a, CD8p, CD28, CD45ABC, CD45RO, CD45RAB, CD45RAC, CD45RBC, CD45RA, CD45RB, CD45RC, CD45del-d3, CD45del- d2-d3, CADHERIN 1, CEACAM 5, or variants thereof; a transmembrane domain or GPI anchor, and optionally an intracellular domain.

[0307] In particular embodiments, a non-viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain; a spacer comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 411-413, SEQ ID NOs: 414- 423, SEQ ID NOs: 424-427, and SEQ ID NOs: 428-434; a transmembrane domain or GPI anchor, and optionally an intracellular domain.

[0308] In particular embodiments, a non-viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain comprising an anti-CD3s antibody or antigen binding fragment thereof comprising a CDRH1, CDRH2, CDRH3, CDRL1, CDRL2, and CDRL3, a VH and VL, and / or an scFv set forth in Table 5; a spacer comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 411, 412, and 413, SEQ ID NOs: 414, 415, 416, 417, 418, 419, 420, 421, 422, and 423, SEQ ID NOs: 424, 425, 426, and 427, and SEQ ID NOs: 428, 429, 430, 431, 432, 433, and 434; a transmembrane domain or GPI anchor, and optionally an intracellular domain.

[0309] In particular embodiments, a non-viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain comprising an anti-CD3s antibody or antigen binding fragment thereof comprising a CDRH1, CDRH2, CDRH3, CDRL1, CDRL2, and CDRL3; a VH and VL; and / or an scFv set forth in Table 5; a spacer comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 411, 423, and 428; a transmembrane domain or GPI anchor, and optionally an intracellular domain

[0310] In particular embodiments, a non-viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain comprising an anti-CD3s antibody or antigen binding fragment thereof comprising the amino acid sequence set forth in any one of SEQ ID NOs: 19, 20, 29, 30, 39, 40, 49, 50, 59, 60, 69, 70, 79, 80, 89, 90, 99, 100, 109, 110,

[0311] 119, 120, 129, 130, 139, 140, 149, 150, 159, 160, 169, 170, 179, 180, 189, 190, 199, 200,

[0312] 209, 210, 219, 220, 229, 230, 239, 240, 249, 250, 259, 260, 269, 270, 279, 280, 289, 290,

[0313] 299, 300, 309, 310, 319, 320, 329, 330, 339, 340, 349, 350, 359, 360, 369, 370, 379, 380,

[0314] 389, 390, 399, 400, 409, and 410; a spacer comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 411, 412, and 413, SEQ ID NOs: 414, 415, 416, 417, 418, 419, 420, 421, 422, and 423, SEQ ID NOs: 424, 425, 426, and 427, and SEQ ID NOs: 428, 429, 430, 431, 432, 433, and 434; a transmembrane domain or GPI anchor, and optionally an intracellular domain.

[0315] In particular embodiments, a non-viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain comprising an anti-CD3s antibody or antigen binding fragment thereof comprising the amino acid sequence set forth in any one of SEQ ID NOs: 19, 20, 29, 30, 39, 40, 49, 50, 59, 60, 69, 70, 79, 80, 89, 90, 99, 100, 109, 110, 119, 120, 129, 130, 139, 140, 149, 150, 159, 160, 169, 170, 179, 180, 189, 190, 199, 200, 209, 210, 219, 220, 229, 230, 239, 240, 249, 250, 259, 260, 269, 270, 279, 280, 289, 290, 299, 300, 309, 310, 319, 320, 329, 330, 339, 340, 349, 350, 359, 360, 369, 370, 379, 380, 389, 390, 399, 400, 409, and 410; a spacer comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 411, 412, and 413, SEQ ID NOs: 414, 415, 416, 417, 418, 419, 420, 421, 422, and 423, SEQ ID NOs: 424, 425, 426, and 427, SEQ ID NOs: 428, 429, 430, 431, 432, 433, and 434, and amino acid sequences at least 85% identical to, at least 90% identical to, at least 95% identical to, at least 96% identical to, at least 97% identical to, at least 98% identical to, or at least 99% identical thereto; a transmembrane domain or GPI anchor, and optionally an intracellular domain.

[0316] In particular embodiments, a non-viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain comprising an anti-CD3s antibody or antigen binding fragment thereof comprising the amino acid sequence set forth in any one of SEQ ID NOs: 99, 100, 109, 110, 119, 120, 129, 130, 139, 140, 149, 150, 159, 160, 169, and 170; a spacer comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 411, 412, and 413, SEQ ID NOs: 414, 415, 416, 417, 418, 419, 420, 421, 422, and 423, SEQ ID NOs: 424, 425, 426, and 427, SEQ ID NOs: 428, 429, 430, 431, 432, 433, and 434, and amino acid sequences at least 85% identical to, at least 90% identical to, at least 95% identical to, at least 96% identical to, at least 97% identical to, at least 98% identical to, or at least 99% identical thereto; a transmembrane domain or GPI anchor, and optionally an intracellular domain.

[0317] In particular embodiments, a non-viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain comprising an anti-CD3s antibody or antigen binding fragment thereof comprising the amino acid sequence set forth in any one of SEQ ID NOs: 99, 100, 109, 110, 119, 120, 129, 130, 139, 140, 149, 150, 159, 160, 169, and 170; a spacer comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 411, 423, 428, and amino acid sequences at least 85% identical to, at least 90% identical to, at least 95% identical to, at least 96% identical to, at least 97% identical to, at least 98% identical to, or at least 99% identical thereto; a transmembrane domain or GPI anchor, and optionally an intracellular domain.

[0318] Table 7 c. Transmembrane Domain

[0319] Non-viral membrane-bound tropism polypeptides contemplated in particular embodiments comprise an extracellular antigen targeting domain that binds an antigen expressed on an immune effector cell; a spacer polypeptide; a transmembrane domain and optionally, an intracellular domain.

[0320] In particular embodiments, a non-viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain that binds an antigen expressed on an immune effector cell; a spacer polypeptide; a transmembrane domain derived, obtained, or isolated from a polypeptide selected from the group consisting of: alpha, beta, gamma, or delta chain of the T-cell receptor, CD2, CD35, CD3s CD3y, CD4, CD5, CD7, CD8a, CD9, CD16, CD22, CD27, CD28, CD33, CD37, CD40, CD45, CD64, CD80, CD86, CD95, CD 134, CD137, CD152, CD154, CD200R, CD223, CD235a, CD244, CD270, CD272, CD273, CD274, CD278, CD279, CD300, CD357, TNRF2, TNFRS14, TNFRS25, TLR2, TLR4, TLR5, TIM1, SlamFl, SlamF5, SlamF6, and an amino acid sequence at least 85% identical to, at least 90% identical to, at least 95% identical to, at least 96% identical to, at least 97% identical to, at least 98% identical to, or at least 99% identical thereto; and optionally, an intracellular domain of no more than 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acids.

[0321] In particular embodiments, a non-viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain; a spacer comprising a hinge or stalk domain derived, obtained, or isolated from PDGFR, IgGl, IgG2, IgG4, CD2, CD3, CD4, CD8a, CD8P, CD28, CD45ABC, CD45RO, CD45RAB, CD45RAC, CD45RBC, CD45RA, CD45RB, CD45RC, CD45del-d3, CD45del-d2-d3, CADHERIN 1, CEACAM 5, variants thereof, and polypeptide linkers of similar amino acid composition and lengths; a transmembrane domain derived, obtained, or isolated from a polypeptide selected from the group consisting of: alpha, beta, gamma, or delta chain of the T-cell receptor, CD2, CD35, CD3s CD3y, CD4, CD5, CD7, CD8a, CD9, CD 16, CD22, CD27, CD28, CD33, CD37, CD40, CD45, CD64, CD80, CD86, CD95, CD 134, CD137, CD152, CD154, CD200R, CD223, CD235a, CD244, CD270, CD272, CD273, CD274, CD278, CD279, CD300, CD357, PDGFR, TNRF2, TNFRS14, TNFRS25, TLR2, TLR4, TLR5, TIM1, SlamFl, SlamF5, SlamF6, and an amino acid sequence at least 85% identical to, at least 90% identical to, at least 95% identical to, at least 96% identical to, at least 97% identical to, at least 98% identical to, or at least 99% identical thereto; and optionally, an intracellular domain of no more than 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acids.

[0322] In particular embodiments, a non-viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain comprising an anti-CD3s antibody or antigen binding fragment thereof comprising a CDRH1, CDRH2, CDRH3, CDRL1, CDRL2, and CDRL3, a VH and VL, and / or an scFv set forth in Table 5; a spacer comprising a hinge or stalk domain derived, obtained, or isolated from PDGFR, IgGl, IgG2, IgG4, CD2, CD3, CD4, CD8a, CD8p, CD28, CD45ABC, CD45RO, CD45RAB, CD45RAC, CD45RBC, CD45RA, CD45RB, CD45RC, CD45del-d3, CD45del-d2-d3, CADHERIN 1, CEACAM 5, or variants thereof; a transmembrane domain derived, obtained, or isolated from a polypeptide selected from the group consisting of: alpha, beta, gamma, or delta chain of the T-cell receptor, CD2, CD35, CD3s CD3y, CD4, CD5, CD7, CD8a, CD9, CD16, CD22, CD27, CD28, CD33, CD37, CD40, CD45, CD64, CD80, CD86, CD95, CD 134, CD137, CD152, CD154, CD200R, CD223, CD235a, CD244, CD270, CD272, CD273, CD274, CD278, CD279, CD300, CD357, PDGFR, TNRF2, TNFRS14, TNFRS25, TLR2, TLR4, TLR5, TIM1, SlamFl, SlamF5, SlamF6, and an amino acid sequence at least 85% identical to, at least 90% identical to, at least 95% identical to, at least 96% identical to, at least 97% identical to, at least 98% identical to, or at least 99% identical thereto; and optionally, an intracellular domain of no more than 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acids.

[0323] In particular embodiments, a non-viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain comprising an anti-CD3s antibody or antigen binding fragment thereof comprising the amino acid sequence set forth in any one of SEQ ID NOs: 19, 20, 29, 30, 39, 40, 49, 50, 59, 60, 69, 70, 79, 80, 89, 90, 99, 100, 109, 110, 119, 120, 129, 130, 139, 140, 149, 150, 159, 160, 169, 170, 179, 180, 189, 190, 199, 200, 209, 210, 219, 220, 229, 230, 239, 240, 249, 250, 259, 260, 269, 270, 279, 280, 289, 290, 299, 300, 309, 310, 319, 320, 329, 330, 339, 340, 349, 350, 359, 360, 369, 370, 379, 380, 389, 390, 399, 400, 409, and 410; a spacer comprising a hinge or stalk domain derived, obtained, or isolated from PDGFR, IgGl, IgG2, IgG4, CD2, CD3, CD4, CD8a, CD8P, CD28, CD45ABC, CD45RO, CD45RAB, CD45RAC, CD45RBC, CD45RA, CD45RB, CD45RC, CD45del-d3, CD45del-d2-d3, CADHERIN 1, CEACAM 5, or variants thereof; a transmembrane domain derived, obtained, or isolated from a polypeptide selected from the group consisting of: alpha, beta, gamma, or delta chain of the T-cell receptor, CD2, CD35, CD3s CD3y, CD4, CD5, CD7, CD8a, CD9, CD 16, CD22, CD27, CD28, CD33, CD37, CD40, CD45, CD64, CD80, CD86, CD95, CD 134, CD137, CD152, CD154, CD200R, CD223, CD235a, CD244, CD270, CD272, CD273, CD274, CD278, CD279, CD300, CD357, PDGFR, TNRF2, TNFRS14, TNFRS25, TLR2, TLR4, TLR5, TIM1, SlamFl, SlamF5, SlamF6, and an amino acid sequence at least 85% identical to, at least 90% identical to, at least 95% identical to, at least 96% identical to, at least 97% identical to, at least 98% identical to, or at least 99% identical thereto; and optionally, an intracellular domain of no more than 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acids.

[0324] In particular embodiments, a non-viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain comprising an anti-CD3s antibody or antigen binding fragment thereof comprising the amino acid sequence set forth in any one of SEQ ID NOs: 19, 20, 29, 30, 39, 40, 49, 50, 59, 60, 69, 70, 79, 80, 89, 90, 99, 100, 109, 110, 119, 120, 129, 130, 139, 140, 149, 150, 159, 160, 169, 170, 179, 180, 189, 190, 199, 200,

[0325] 209, 210, 219, 220, 229, 230, 239, 240, 249, 250, 259, 260, 269, 270, 279, 280, 289, 290,

[0326] 299, 300, 309, 310, 319, 320, 329, 330, 339, 340, 349, 350, 359, 360, 369, 370, 379, 380,

[0327] 389, 390, 399, 400, 409, and 410; a spacer comprising a hinge or stalk domain derived, obtained, or isolated from PDGFR, IgGl, IgG2, IgG4, CD2, CD3, CD4, CD8a, CD8P, CD28, CD45ABC, CD45RO, CD45RAB, CD45RAC, CD45RBC, CD45RA, CD45RB, CD45RC, CD45del-d3, CD45del-d2-d3, CADHERIN 1, CEACAM 5, or variants thereof; a transmembrane domain comprising an amino acid sequence set forth in any one of SEQ ID NOs: 435, 436, 437, 438, 439, 440, and 441 and an amino acid sequence at least 85% identical to, at least 90% identical to, at least 95% identical to, at least 96% identical to, at least 97% identical to, at least 98% identical to, or at least 99% identical thereto; and optionally, an intracellular domain of no more than 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acids.

[0328] In particular embodiments, a non-viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain comprising an anti-CD3s antibody or antigen binding fragment thereof comprising the amino acid sequence set forth in any one of SEQ ID NOs: 99, 100, 109, 110, 119, 120, 129, 130, 139, 140, 149, 150, 159, 160, 169, and 170; a spacer comprising a hinge or stalk domain derived, obtained, or isolated from PDGFR, IgGl, IgG2, IgG4, CD2, CD3, CD4, CD8a, CD8p, CD28, CD45ABC, CD45RO, CD45RAB, CD45RAC, CD45RBC, CD45RA, CD45RB, CD45RC, CD45del-d3, CD45del- d2-d3, CADHERIN 1, CEACAM 5, or variants thereof; a transmembrane domain comprising an amino acid sequence set forth in any one of SEQ ID NOs: 435, 436, 437, 438, 439, 440, and 441, and an amino acid sequence at least 85% identical to, at least 90% identical to, at least 95% identical to, at least 96% identical to, at least 97% identical to, at least 98% identical to, or at least 99% identical thereto; and optionally, an intracellular domain of no more than 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acids.

[0329] In particular embodiments, a non-viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain; a spacer comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 411-413, SEQ ID NOs: 414- 423, SEQ ID NOs: 424-427, and SEQ ID NOs: 428-434; a transmembrane domain comprising an amino acid sequence set forth in any one of SEQ ID NOs: 435, 436, 437, 438, 439, 440, and 441; and optionally, an intracellular domain of no more than 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acids.

[0330] In particular embodiments, a non-viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain comprising an anti-CD3s antibody or antigen binding fragment thereof comprising a CDRH1, CDRH2, CDRH3, CDRL1, CDRL2, and CDRL3, a VH and VL, and / or an scFv set forth in Table 5; a spacer comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 411, 412, and 413, SEQ ID NOs: 414, 415, 416, 417, 418, 419, 420, 421, 422, and 423, SEQ ID NOs: 424, 425, 426, and 427, and SEQ ID NOs: 428, 429, 430, 431, 432, 433, and 434; a transmembrane domain comprising an amino acid sequence set forth in any one of SEQ ID NOs: 435, 436, 437, 438, 439, 440, and 441; and optionally, an intracellular domain of no more than 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acids.

[0331] In particular embodiments, a non-viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain comprising an anti-CD3s antibody or antigen binding fragment thereof comprising the amino acid sequence set forth in any one of SEQ ID NOs: 19, 20, 29, 30, 39, 40, 49, 50, 59, 60, 69, 70, 79, 80, 89, 90, 99, 100, 109, 110, 119, 120, 129, 130, 139, 140, 149, 150, 159, 160, 169, 170, 179, 180, 189, 190, 199, 200, 209, 210, 219, 220, 229, 230, 239, 240, 249, 250, 259, 260, 269, 270, 279, 280, 289, 290, 299, 300, 309, 310, 319, 320, 329, 330, 339, 340, 349, 350, 359, 360, 369, 370, 379, 380, 389, 390, 399, 400, 409, and 410; a spacer comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 411, 412, and 413, SEQ ID NOs: 414, 415, 416, 417, 418, 419, 420, 421, 422, and 423, SEQ ID NOs: 424, 425, 426, and 427, and SEQ ID NOs: 428, 429, 430, 431, 432, 433, and 434; a transmembrane domain comprising an amino acid sequence set forth in any one of SEQ ID NOs: 435, 436, 437, 438, 439, 440, and 441; and optionally, an intracellular domain of no more than 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acids.

[0332] In particular embodiments, a non-viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain comprising an anti-CD3s antibody or antigen binding fragment thereof comprising the amino acid sequence set forth in any one of SEQ ID NOs: 19, 20, 29, 30, 39, 40, 49, 50, 59, 60, 69, 70, 79, 80, 89, 90, 99, 100, 109, 110,

[0333] 119, 120, 129, 130, 139, 140, 149, 150, 159, 160, 169, 170, 179, 180, 189, 190, 199, 200,

[0334] 209, 210, 219, 220, 229, 230, 239, 240, 249, 250, 259, 260, 269, 270, 279, 280, 289, 290,

[0335] 299, 300, 309, 310, 319, 320, 329, 330, 339, 340, 349, 350, 359, 360, 369, 370, 379, 380,

[0336] 389, 390, 399, 400, 409, and 410; a spacer comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 411, 412, and 413, SEQ ID NOs: 414, 415, 416, 417, 418, 419, 420, 421, 422, and 423, SEQ ID NOs: 424, 425, 426, and 427, and SEQ ID NOs: 428, 429, 430, 431, 432, 433, and 434; a transmembrane domain comprising an amino acid sequence set forth in any one of SEQ ID NOs: 435, 436, 437, 438, 439, 440, and 441, and an amino acid sequence at least 85% identical to, at least 90% identical to, at least 95% identical to, at least 96% identical to, at least 97 % identical to, at least 98% identical to, or at least 99% identical thereto; and optionally, an intracellular domain of no more than 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acids.

[0337] In particular embodiments, a non-viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain comprising an anti-CD3s antibody or antigen binding fragment thereof comprising the amino acid sequence set forth in any one of SEQ ID NOs: 99, 100, 109, 110, 119, 120, 129, 130, 139, 140, 149, 150, 159, 160, 169, and 170; a spacer comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 411, 423, and 428; a transmembrane domain comprising an amino acid sequence set forth in any one of SEQ ID NOs: 437 and 439; and optionally, an intracellular domain of no more than 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acids.

[0338] Table 8

[0339] In particular embodiments, a non-viral membrane-bound tropism polypeptide comprises a transmembrane domain and optionally, an intracellular domain of no more than 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acids. In preferred embodiments, an intracellular domain comprises, consists essentially of, or consists of one, two, three, four, five, or six of the amino acids set forth in any one of SEQ ID NOs: 442, 443, 444, 445, 446, 447, and 448.

[0340] Table 9

[0341] In particular embodiments, a non-viral membrane bound tropism polypeptide comprises an extracellular antigen targeting domain, a spacer polypeptide, and a GPI anchor. A “GPI anchor” or “glycosylphosphatidylinositol anchor” refers to a posttranslational modification that anchors a non-viral membrane bound tropism polypeptide in the outer leaflet of a lipid bilayer, a cell membrane, or a viral envelope of a recombinant particle contemplated herein. A GPI anchor is a complex structure comprising a phosphoethanolamine linker, glycan core, and phospholipid tail. In particular embodiments, a nascently translated non-viral membrane-bound tropism polypeptide comprises a GPI attachment signal peptide at its C-terminus. The signal peptide is recognized by GPI transamidase, which cleaves and replaces it with a preassembled GPI by transamidation at an amino acid termed the co site amino acid thereby generating a GPI anchored non-viral membrane-bound tropism polypeptide.

[0342] In particular embodiments, a non-viral membrane bound tropism polypeptide comprises an amino acid sequence set forth in any one of SEQ ID NOs: 455-576. In particular preferred embodiments, a non- viral membrane bound tropism polypeptide comprises an amino acid sequence set forth in any one of SEQ ID NOs: 461, 471, 483, 537, and 553. Table 10 d. Additional Tropism Polypeptides

[0343] Recombinant particles contemplated herein comprise at least one non-viral membrane-bound tropism polypeptide. In particular embodiments, it may be preferred, and in some cases necessary or advantageous, to engineer a recombinant particle that comprises at least one additional non-viral membrane-bound tropism polypeptide. In some embodiments, a recombinant particle comprises two non-viral membrane-bound tropism polypeptides. In some embodiments, a recombinant particle comprises three non-viral membrane-bound tropism polypeptides.

[0344] In particular embodiments, a recombinant particle comprises a lipid bilayer, cell membrane, or viral envelope comprising one or more mutated viral envelope glycoproteins that mediate fusion of the viral particle with a cell but that do not bind its cognate receptor expressed on a cell, and at least two non-viral membrane-bound tropism polypeptides, e.g., a primary and / or secondary non-viral membrane bound tropism polypeptide, that direct the particle to bind one or more specific antigens expressed on a desired cell type, e.g., an immune effector cell. In preferred embodiments, the recombinant particle is a recombinant viral particle, e.g., a retroviral or lentiviral particle. In particular embodiments, wherein a recombinant particle comprises at least two non- viral membrane-bound tropism polypeptides, each non- viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain that binds an antigen expressed on an immune effector cell; a spacer polypeptide, a transmembrane domain or GPI anchor; and optionally an intracellular domain.

[0345] In particular embodiments, a recombinant particle comprises at least two non-viral membrane-bound tropism polypeptides, one non-viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain that binds an antigen expressed on an immune effector cell including but not limited to the alpha, beta, gamma, or delta chain of the T cell receptor, CD2, CD35, CD3s CD3y, CD4, CD8a, and CD8P; a spacer polypeptide; a transmembrane domain or GPI anchor; and optionally an intracellular domain; and another non-viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain that an antigen expressed on an immune effector cell including but not limited to the alpha or beta chain of a TCR, CD28, CD134 (0X40), CD137 (4-1BB), and CD278 (ICOS); a spacer polypeptide, a transmembrane domain or GPI anchor; and optionally an intracellular domain.

[0346] In particular embodiments, a recombinant particle comprises at least two non-viral membrane-bound tropism polypeptides, one non-viral membrane-bound tropism polypeptide comprises an extracellular antigen targeting domain that binds an antigen expressed on an immune effector cell including but not limited to the alpha, beta, gamma, or delta chain of the T cell receptor, CD2, CD35, CD3s CD3y, CD4, CD8a, and CD8P; a spacer polypeptide; a transmembrane domain or GPI anchor; and optionally an intracellular domain; and another non-viral membrane bound tropism polypeptide comprises all or part of a costimulatory molecule including but not limited to CD80, CD86, OX40L, 4-1BBL, and ICOSL or a functional fragment thereof. In particular embodiments, the functional fragment is a CD80, CD86, OX40L, 4-1BBL, and ICOSL comprising a C-terminal truncation that results in a cytoplasmic domain of about 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acids or more). In particular embodiments, the functional fragment comprises, consists essentially of, or consists of the amino acid sequence set forth in any one of SEQ ID NOs: 577, 578, 579, 580, and 581.

[0347] In particular embodiments, a recombinant particle comprises a first non-viral membrane-bound tropism polypeptide comprising an extracellular antigen targeting domain comprising an anti-CD3s antibody or antigen binding fragment thereof comprising a CDRH1, CDRH2, CDRH3, CDRL1, CDRL2, and CDRL3, a VH and VL, and / or an scFv set forth in Table 5; a spacer comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 411, 412, and 413, SEQ ID NOs: 414, 415, 416, 417, 418, 419, 420, 421, 422, and 423, SEQ ID NOs: 424, 425, 426, and 427, and SEQ ID NOs: 428, 429, 430, 431, 432, 433, and 434; a transmembrane domain comprising an amino acid sequence set forth in any one of SEQ ID NOs: 435, 436, 437, 438, 439, 440, and 441; and optionally, an intracellular domain of no more than 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acids; and a second non-viral membrane bound tropism polypeptide comprising, consisting essentially of, or consisting of the amino acid sequence set forth in any one of SEQ ID NOs: 577, 578, 579, 580, and 581.

[0348] In particular embodiments, a recombinant particle comprising a first non-viral membrane-bound tropism polypeptide comprising an extracellular antigen targeting domain comprising an anti-CD3s antibody or antigen binding fragment thereof comprising the amino acid sequence set forth in any one of SEQ ID NOs: 19, 20, 29, 30, 39, 40, 49, 50, 59, 60, 69, 70, 79, 80, 89, 90, 99, 100, 109, 110, 119, 120, 129, 130, 139, 140, 149, 150, 159, 160, 169, 170, 179, 180, 189, 190, 199, 200, 209, 210, 219, 220, 229, 230, 239, 240, 249, 250, 259, 260, 269, 270, 279, 280, 289, 290, 299, 300, 309, 310, 319, 320, 329, 330, 339, 340, 349, 350, 359, 360, 369, 370, 379, 380, 389, 390, 399, 400, 409, and 410; a spacer comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 411, 412, and 413, SEQ ID NOs: 414, 415, 416, 417, 418, 419, 420, 421, 422, and 423, SEQ ID NOs: 424, 425, 426, and 427, and SEQ ID NOs: 428, 429, 430, 431, 432, 433, and 434; a transmembrane domain comprising an amino acid sequence set forth in any one of SEQ ID NOs: 435, 436, 437, 438, 439, 440, and 441; and optionally, an intracellular domain of no more than 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acids; and a second non-viral membrane bound tropism polypeptide comprising, consisting essentially of, or consisting of the amino acid sequence set forth in any one of SEQ ID NOs: 577, 578, 579, 580, and 581.

[0349] In particular embodiments, a recombinant particle comprising a non-viral membranebound tropism polypeptide comprising an extracellular antigen targeting domain comprising an anti-CD3s antibody or antigen binding fragment thereof comprising the amino acid sequence set forth in any one of SEQ ID NOs: 19, 20, 29, 30, 39, 40, 49, 50, 59, 60, 69, 70, 79, 80, 89, 90, 99, 100, 109, 110, 119, 120, 129, 130, 139, 140, 149, 150, 159, 160, 169, 170, 179, 180, 189, 190, 199, 200, 209, 210, 219, 220, 229, 230, 239, 240, 249, 250, 259, 260, 269, 270, 279, 280, 289, 290, 299, 300, 309, 310, 319, 320, 329, 330, 339, 340, 349, 350, 359, 360, 369, 370, 379, 380, 389, 390, 399, 400, 409, and 410; a spacer polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 411, 412, and 413, SEQ ID NOs: 414, 415, 416, 417, 418, 419, 420, 421, 422, and 423, SEQ ID NOs: 424, 425, 426, and 427, and SEQ ID NOs: 428, 429, 430, 431, 432, 433, and 434; a transmembrane domain comprising an amino acid sequence set forth in any one of SEQ ID NOs: 435, 436, 437, 438, 439, 440, and 441; and optionally, an intracellular domain of no more than 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acids; and a second non-viral membrane bound tropism polypeptide comprising, consisting essentially of, or consisting of the amino acid sequence set forth in any one of SEQ ID NOs: 577, 578, 579, 580, and 581.

[0350] In particular embodiments, a recombinant particle comprising a non-viral membranebound tropism polypeptide comprising an extracellular antigen targeting domain comprising an anti-CD3s antibody or antigen binding fragment thereof comprising the amino acid sequence set forth in any one of SEQ ID NOs: 99, 100, 109, 110, 119, 120, 129, 130, 139, 140, 149, 150, 159, 160, 169, and 170; a spacer comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 411, 412, and 413, SEQ ID NOs: 414, 415, 416, 417, 418, 419, 420, 421, 422, and 423, SEQ ID NOs: 424, 425, 426, and 427, and SEQ ID NOs: 428, 429, 430, 431, 432, 433, and 434; a transmembrane domain comprising an amino acid sequence set forth in any one of SEQ ID NOs: 435, 436, 437, 438, 439, 440, and 441; and optionally, an intracellular domain comprising, consisting essentially of, or consisting of one, two, three, four, five, or six of the amino acids set forth in any one of SEQ ID NOs: 442, 443, 444, 445, 446, 447 and 448; and a second non-viral membrane bound tropism polypeptide comprising, consisting essentially of, or consisting of the amino acid sequence set forth in any one of SEQ ID NOs: 577, 578, 579, 580, and 581.

[0351] In particular embodiments, a recombinant particle comprising a non-viral membranebound tropism polypeptide comprising an extracellular antigen targeting domain comprising an anti-CD3s antibody or antigen binding fragment thereof comprising the amino acid sequence set forth in any one of SEQ ID NOs: 99, 100, 109, 110, 119, 120, 129, 130, 139, 140, 149, 150, 159, 160, 169, and 170; a spacer comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 411, 423, and 428; a transmembrane domain comprising an amino acid sequence set forth in any one of SEQ ID NOs: 437 and 439; and optionally, an intracellular domain comprising, consisting essentially of, or consisting of one, two, three, four, five, or six of the amino acids set forth in any one of SEQ ID NOs 444 and 446; and a second non-viral membrane bound tropism polypeptide comprising, consisting essentially of, or consisting of the amino acid sequence set forth in any one of SEQ ID NOs: 577, 578, 579, 580, and 581.

[0352] In particular embodiments, a recombinant particle comprising a non- viral membranebound tropism polypeptide comprising an amino acid sequence set forth in any one of SEQ ID NOs: 461, 471, 483, 537, and 553; and a second non-viral membrane bound tropism polypeptide comprising, consisting essentially of, or consisting of the amino acid sequence set forth in any one of SEQ ID NOs: 577, 578, 579, 580, and 581. In particular embodiments, a recombinant particle comprising a non-viral membrane-bound tropism polypeptide comprising an amino acid sequence set forth in SEQ ID NO: 461; and a second non-viral membrane bound tropism polypeptide comprising, consisting essentially of, or consisting of the amino acid sequence set forth in any one of SEQ ID NOs: 577, 578, 579, 580, and 581. In particular embodiments, a recombinant particle comprising a non-viral membrane-bound tropism polypeptide comprising an amino acid sequence set forth in SEQ ID NO: 471; and a second non-viral membrane bound tropism polypeptide comprising, consisting essentially of, or consisting of the amino acid sequence set forth in any one of SEQ ID NOs: 577, 578, 579, 580, and 581. In particular embodiments, a recombinant particle comprising a non-viral membrane-bound tropism polypeptide comprising an amino acid sequence set forth in SEQ ID NO: 483; and a second non-viral membrane bound tropism polypeptide comprising, consisting essentially of, or consisting of the amino acid sequence set forth in any one of SEQ ID NOs: 577, 578, 579, 580, and 581. In particular embodiments, a recombinant particle comprising a non-viral membrane-bound tropism polypeptide comprising an amino acid sequence set forth in SEQ ID NO: 537; and a second non-viral membrane bound tropism polypeptide comprising, consisting essentially of, or consisting of the amino acid sequence set forth in any one of SEQ ID NOs: 577, 578, 579, 580, and 581. In particular embodiments, a recombinant particle comprising a non-viral membrane-bound tropism polypeptide comprising an amino acid sequence set forth in SEQ ID NO: 553; and a second non-viral membrane bound tropism polypeptide comprising, consisting essentially of, or consisting of the amino acid sequence set forth in any one of SEQ ID NOs: 577, 578, 579, 580, and 581.

[0353] Table 11

[0354] E. PAYLOAD

[0355] Recombinant particles contemplated herein are engineered to efficiently deliver a payload comprising one or more polynucleotides, polypeptides, and / or small molecules to a specific cell type recognized by one or more tropism molecules expressed or displayed on the surface of the particle. A payload may comprise one or more polynucleotides, polypeptides, and / or small molecules sufficient to enable gene therapy, gene editing, or cell therapy. In particular embodiments, the pay load is a therapeutic pay load. Without wishing to be bound to any particular theory it is further contemplated in particular embodiments that once the particle binds to and fuses with a specific cell, the payload is introduced into the cell and either genetically modifies the cell or is maintained episomally in the cell. In particular embodiments, a recombinant particle delivers a pay load comprising one or more gene therapy vectors, therapeutic proteins, and / or genome editing compositions to a specified celltype in a subject in vivo.

[0356] Recombinant particles contemplated in particular embodiments comprise a surface and a payload: the surface comprising one or more mutated viral envelope glycoproteins that retain fusogenic activity and substantially lack or lack cognate receptor binding activity and one or more non- viral membrane-bound tropism polypeptides engineered to bind a desired cell type, e.g., an immune effector cell; and the payload comprising one or more polynucleotide and / or polypeptides. Suitable recombinant particles include but are not limited to a fusosome; an extracellular vesicle, including a microvesicle, an apoptotic body, and an exosome; a lipid nanoparticle; a virus-like particle (VLPs); or a recombinant viral particle. In particular embodiments, a recombinant particle comprises (i) a surface comprising one or more mutated viral envelope glycoproteins that retain fusogenic activity and lack cognate receptor binding activity and one or more non-viral membrane-bound tropism polypeptides that bind an antigen expressed on a cell and (ii) a payload comprising one or more vectors.

[0357] In particular embodiments, a recombinant particle comprises (i) a surface comprising one or more mutated viral envelope glycoproteins that retain fusogenic activity and lack cognate receptor binding activity and one or more non-viral membrane-bound tropism polypeptides that bind an antigen expressed on a cell and (ii) a pay load comprising a genome editing composition.

[0358] / . VECTOR PAYLOADS

[0359] Recombinant particles contemplated in particular embodiments comprise (i) a surface comprising one or more mutated viral envelope glycoproteins that retain fusogenic activity and lack cognate receptor binding activity and one or more non-viral membranebound tropism polypeptides that bind an antigen expressed on a cell and (ii) a payload comprising one or more polynucleotides.

[0360] In particular embodiments, a payload comprises one or more polynucleotides. Illustrative examples of polynucleotide payloads include but are not limited to: RNAs including but not limited to circRNAs, miRNAs, siRNAs, sgRNAs, shRNAs, or ribozymes; RNAs include therapeutic polypeptides; RNAs including genome editing enzymes and / or donor templates; non-viral vectors and viral vectors.

[0361] Illustrative examples non-viral vectors include but are not limited to: plasmids, phagemids, cosmids, artificial chromosomes such as yeast artificial chromosomes (YAC), bacterial artificial chromosomes (BAC), or Pl-derived artificial chromosomes (PAC), and bacteriophages such as lambda phage or M13 phage.

[0362] Illustrative examples viral vectors include but are not limited to: Adenoviral (Ad) vectors, adeno-associated virus (AAV) vectors, rhabdovirus (e.g., lyssavirus, vesiculovirus) vectors, paramyxovirus (e.g., henipavirus, morbillivirus, respirovirus, rubelavirus) vectors, herpes simplex virus (e.g., HSV-1, HSV-2) vectors, vaccinia virus vectors, and retroviral vectors, preferably lentiviral vectors (LVV). A “viral vector” is a nucleic acid molecule derived from a viral genome that is used to transfer or deliver another nucleic acid to a cell. A viral vector is based on, and derived from, a virus genome that has been engineered to remove viral accessory proteins but leave elements intact for packaging, reverse transcription and integration. In preferred embodiments, viral vectors contemplated herein comprise a polynucleotide encoding a promoter operably linked to a polynucleotide encoding an engineered receptor, e.g., a chimeric antigen receptor (CAR), a chimeric costimulatory receptor (CCR), an alpha-beta T cell receptor (aP TCR), a gamma delta T cell receptor (y5 TCR), a dimerizing agent regulated immunoreceptor complex (DARIC), a chimeric TGF-P receptor (CTBR), or a zetakine receptor.

[0363] In particular embodiments, a recombinant particle comprises (i) a surface comprising one or more mutated viral envelope glycoproteins that retain fusogenic activity and lack cognate receptor binding activity and one or more non-viral membrane-bound tropism polypeptides that bind an antigen expressed on a cell and (ii) a payload comprising an adenoviral vector. High-capacity adenoviral vectors (HC-Ads) (third generation) only retain short non-coding regions from the Ad genome (ITRs and y signal), which enables the vector tp carry large polynucleotide payloads (~37kb).

[0364] In particular embodiments, a recombinant particle comprises (i) a surface comprising one or more mutated viral envelope glycoproteins that retain fusogenic activity and lack cognate receptor binding activity and one or more non-viral membrane-bound tropism polypeptides that bind an antigen expressed on a cell and (ii) a payload comprising an AAV vector. Recombinant AAV (rAAV) vectors are primarily episomally maintained and have a polynucleotide payload capacity of about 4.7kb. rAAV vectors are typically composed of, at a minimum, a transgene and its regulatory sequences, and 5' and 3' AAV inverted terminal repeats (ITRs). rAAV vectors may comprise ITRs from any one of AAV1, AAV2, AAV3, AAV4, AAV5, AAV6, AAV7, AAV8, AAV9, or AAV10. Construction of rAAV vectors, and production, and purificationof AAV have been disclosed, e.g., in U.S. Patent Nos. 9,169,494; 9,169,492; 9,012,224; 8,889,641; 8,809,058; and 8,784,799, each of which is incorporated by reference herein, in its entirety.

[0365] In particular embodiments, a recombinant particle comprises (i) a surface comprising one or more mutated viral envelope glycoproteins that retain fusogenic activity and lack cognate receptor binding activity and one or more non-viral membrane-bound tropism polypeptides that bind an antigen expressed on a cell and (ii) a payload comprising an HSV vector. HSV vectors are relatively large, e.g., up to 152 kb. Typically, HSV vectors are rendered replication deficient; moreover, one or more essential or non-essential HSV genes are removed from the vector backbone to make room for polynucleotide payloads. Most replication deficient HSV vectors contain a deletion to remove one or more intermediate- early, early, or late HSV genes to prevent replication. Advantages of the HSV vector are its ability to enter a latent stage that can result in long-term DNA expression and its large viral DNA genome that can accommodate exogenous DNA inserts of up to 25 kb. HSV-based vectors are described in, for example, U.S. Pat. Nos. 5,837,532, 5,846,782, and 5,804,413, and International Patent Applications WO 91 / 02788, WO 96 / 04394, WO 98 / 15637, and WO 99 / 06583, each of which are incorporated by reference herein in its entirety.

[0366] In particular embodiments, a recombinant particle comprises (i) a surface comprising one or more mutated viral envelope glycoproteins that retain fusogenic activity and lack cognate receptor binding activity and one or more non-viral membrane-bound tropism polypeptides that bind an antigen expressed on a cell and (ii) a payload comprising a retroviral vector or a lentiviral vector. In particular embodiments, a recombinant particle comprises two copies of a vector, a genomic RNA comprising backbone sequences derived from a retrovirus genome, e.g., a lentivirus genome.

[0367] In various embodiments, a retroviral vector is engineered or derived from a retrovirus genome selected from the group consisting of: an alpharetrovirus genome, a betaretrovirus genome, a gammaretrovirus genome, a deltaretrovirus genome, or a spumavirus genome e.g., an epsilonretrovirus genome, a simiispumavirus genome, a bovispumavirus genome, an equispumavirus genome, a felispumavirus genome, and a pro simiispumavirus genome).

[0368] In particular embodiments, a retroviral vector comprises a 5' LTR and a 3' LTR each isolated, obtained, or derived from a retrovirus genome selected from the group consisting of: an alpharetrovirus genome, a betaretrovirus genome, a gammaretrovirus genome, a deltaretrovirus genome, an epsilonretrovirus genome, and a spumavirus genome.

[0369] Illustrative examples of alpharetroviruses from which a retroviral vector may be isolated, obtained, or derived from include but are not limited to avian leukosis vims, avian carcinoma Mill Hill vims 2, avian myeloblastosis vims, avian myelocytomatosis vims 29, avian sarcoma vims CT10, fujinami sarcoma vims, rous sarcoma vims, UR2 sarcoma vims and Y73 sarcoma vims.

[0370] Illustrative examples of betaretrovimses from which a retroviral vector may be isolated, obtained, or derived from include but are not limited to mouse mammary tumor vims, Jaagsiekte sheep retrovirus, langur vims, Mason-Pfizer monkey vims, and squirrel monkey retrovims (SMRV). Illustrative examples of deltaretroviruses from which a retroviral vector may be isolated, obtained, or derived from include but are not limited to bovine leukemia virus, primate T-lymphotropic virus 1, primate T-lymphotropic vims 2, primate T-lympho tropic vims 3, and primate T-lymphotropic vims 4.

[0371] Illustrative examples of epsilonretrovimses from which a retroviral vector may be isolated, obtained, or derived from include but are not limited to walleye dermal sarcoma vims, walleye epidermal hyperplasia vims 1, and walleye epidermal hyperplasia vims 2.

[0372] Illustrative examples of gammaretrovims from which a retroviral vector may be isolated, obtained, or derived from include but are not limited to baboon endogenous vims (BaEV), chick syncytial vims, feline endogenous vims (e.g., RD 114), feline leukemia vims (FeLV), Finkel-Biskis-Jinkins murine sarcoma vims, Gardner- Amstein feline sarcoma vims, gibbon ape leukemia vims (GALV), guinea pig type-C oncovirus, Hardy-Zuckerman feline sarcoma vims, Harvey murine sarcoma vims, Kirsten murine sarcoma vims, koala retrovirus, murine leukemia vims (MLV), Moloney murine leukemia vims (MoMLV), Moloney murine sarcoma vims, porcine endogenous vims (PERV), Porcine type-C oncovims, reticuloendotheliosis vims (REV), Snyder-Theilen feline sarcoma vims, Trager duck spleen necrosis vims, viper retrovirus, xenotropic murine leukemia vims- related vims (XMRV), and woolly monkey sarcoma vims.

[0373] Illustrative examples of spumavimses from which a retroviral vector may be isolated, obtained, or derived from include but are not limited to simian foamy vims, bovine foamy vims, equine foamy vims, feline foamy vims, human foamy vims (HFV), and brown greater galago prosimian foamy vims.

[0374] In particular embodiments, a recombinant particle comprises a recombinant retroviral vector engineered or derived from a retrovirus genome selected from the group consisting of: SMRV, BaEV, RD 114, FeLV, GALV, MLV, MoMLV, PERV, REV, XMRV, and HFV; gag and pol from an alpharetrovims, a betaretrovims, a gammaretrovims, a deltaretrovims, an epsilonretrovims, or a spumavims; and a 5' LTR and a 3' LTR each isolated, obtained, or derived from a retrovirus genome selected from the group consisting of: an alpharetrovims genome, a betaretrovims genome, a gammaretrovims genome, a deltaretrovims genome, an epsilonretrovims genome, and a spumavims genome.

[0375] In particular embodiments, a recombinant particle comprises a recombinant lentiviral vector engineered or derived from a lentivims genome. Illustrative lentivimses include, but are not limited to, HIV (human immunodeficiency vims; including HIV type 1 , and HIV type 2); visna-maedi virus (VMV); caprine arthritis-encephalitis virus (CAEV); equine infectious anemia vims (EIAV); feline immunodeficiency vims (FIV); bovine immune deficiency vims (BIV); and simian immunodeficiency vims (SIV). In particular embodiments, lentiviral vectors are derived from HIV-1 viral genomes, preferably HIV-1 or HIV-2 viral genomes and more preferably, HIV-1 viral genomes (z.e., HIV-1 cis-acting sequence elements are preferred).

[0376] In various embodiments, a lentiviral vector payload contemplated herein comprises two copies of a lentiviral vector-based RNA genome comprising a 5' long terminal repeat (LTR) comprising R and U5 regions, a Psi ( ) packaging signal, a cPPT / FLAP, an export element, a polynucleotide comprising a promoter operably linked to a therapeutic polynucleotide or a polynucleotide encoding a therapeutic polypeptide, a 3' LTR comprising U3 and R regions, a polyadenylation signal, a poly(A) tail; and optionally one or more of a WPRE or HPRE, one or more microRNA (miRNA) target sequences, an insulator element, a selectable marker, and a cell suicide gene.

[0377] The term “long terminal repeat (LTR),” as used herein, refers to elements located at the ends of retroviral polynucleotides which, in their natural sequence context, are direct repeats and contain U3, R and U5 regions. LTRs generally provide functions fundamental to the expression of retroviral genes (e.g., promotion, initiation and polyadenylation of gene transcripts) and to viral replication. The LTR contains numerous regulatory signals including transcriptional control elements, poly adenylation signals and sequences needed for replication and integration of the viral genome. The viral LTR is divided into three regions called U3, R and U5. The U3 region contains the enhancer and promoter elements. The U5 region is the sequence between the primer binding site and the R region and contains the polyadenylation signal. The R (repeat) region is flanked by the U3 and U5 regions. A transfer plasmid, which is used as the vector genome template comprises a 5' LTR comprising U3, R and / or U5 regions and a 3' LTR comprising U3, R and / or U5 regions. Adjacent to the 5' LTR are sequences necessary for reverse transcription of the genome (the tRNA primer binding site) and for efficient packaging of viral RNA into particles (the Psi “T” site). A retroviral vector-based genome packaged in a particle comprises a 5' LTR comprising R and U5 regions and a 3' LTR comprising U3 and R regions. The retroviral vector-based genome is reverse transcribed and integrated into the host cell genome as a provector. Through reverse transcription and second strand synthesis of the retroviral vector genome, pro vectors comprise two copies of the 3' LTR, one copy that replaces the 5' LTR and the 3' LTR.

[0378] A “TAR” element as used herein, refers to the “trans-activation response” genetic element located in the R region of lentiviral vector LTRs. This element interacts with the lentiviral trans-activator (tat) genetic element to enhance lentiviral vector genome replication. In third generation lentiviral vectors, this element is not usually present because lentiviral vector transfer vectors comprise a 5' LTR U3 region replaced by a heterologous promoter.

[0379] An “R region,” as used herein, refers to the region within LTRs beginning at the start of the capping group (z.e. , the start of transcription) and ending immediately prior to the start of the polyA tract. The R region is also defined as being flanked by the U3 and U5 regions. The R region plays a role during reverse transcription in permitting the transfer of nascent DNA from one end of the genome to the other.

[0380] As used herein, a “packaging signal” or “packaging sequence” refers to sequences located within the retroviral genome which are required for insertion of the viral RNA into the viral capsid or particle, see e.g., Clever et al., 1995. J. of Virology, Vol. 69, No. 4; pp. 2101-2109. Several retroviral vectors use the minimal packaging signal (also referred to as the psi | | or [T / +] sequence) needed for encapsidation of the viral genome. Thus, as used herein, the terms “packaging sequence,” “packaging signal,” “psi” and the symbol “VF,” are used in reference to the non-coding sequence required for encapsidation of retroviral RNA strands during viral particle formation.

[0381] A “FLAP element” or “cPPT / FLAP,” as used herein refers to a nucleic acid whose sequence includes the central polypurine tract and central termination sequences (cPPT and CTS) of a lentivirus, e.g., HIV-1 or HIV-2. “FLAP element” and “cPPT / FLAP” may used interchangeably to refer to the foregoing FLAP element. Suitable FLAP elements are described in U.S. Pat. No. 6,682,907 and in Zennou, et al. , 2000, Cell, 101:173. During HIV- 1 reverse transcription, central initiation of the plus-strand DNA at the central polypurine tract (cPPT) and central termination at the central termination sequence (CTS) lead to the formation of a three- stranded DNA structure: the HIV-1 central DNA flap. While not wishing to be bound by any particular theory, the DNA flap may act as a cis-active determinant of lentiviral genome nuclear import and / or may increase virus titer.

[0382] As used herein, an “export element” refers to a cis-acting post-transcriptional regulatory element which regulates the transport of an RNA transcript from the nucleus to the cytoplasm of a cell. Examples of RNA export elements include, but are not limited to, the human immunodeficiency vims (HIV) rev response element (RRE) (see e.g., Cullen et al., 1991. J. Virol. 65: 1053; and Cullen et al., 1991. Cell 58: 423), the woodchuck hepatitis vims posttranscriptional regulatory element (WPRE), and the hepatitis B vims post- transcriptional regulatory element (HPRE).

[0383] Expression of heterologous sequences in viral vectors may be increased by incorporating posttranscriptional regulatory elements, efficient polyadenylation signals, and optionally, transcription termination signals into the vectors. A variety of posttranscriptional regulatory elements can increase expression of a heterologous nucleic acid at the protein, e.g. , WPRE, HPRE.

[0384] Lentiviral vectors may contain one or more safety enhancements to reduce the risk of replication, insertional mutagenesis, and off-target transduction and / or expression. In particular embodiments, a lentiviral vector comprises one or more or the following safety enhancements: one or more modifications of the 5' and 3' LTRs, and cell or tissue specific expression control sequences, e.g., promoters, enhancers, miRNA target sequences. A “modified LTR,” as used herein, refers to one or more nucleotide additions, deletions or substitutions in the native HIV-1 5' LTR and / or 3' LTR. The skilled artisan would be able to determine whether an LTR is modified by comparison to a reference LTR.

[0385] “Self-inactivating” (SIN) vectors, as used herein, refer to replication-defective vectors, e.g., retroviral or lentiviral vectors, in which the right (3 ) LTR enhancer-promoter region, known as the U3 region, has been modified (e.g., by deletion or substitution) to prevent viral transcription beyond the first round of viral replication. Self-inactivation is achieved through a deletion in the U3 region of the 3' LTR of the lentiviral vector transfer plasmid that removes the LTR TATA box (e.g., deletions from -418 to -18), without significant reductions in titers.

[0386] An additional safety enhancement is provided by replacing the U3 region of the 5' LTR with a heterologous promoter to drive transcription of the viral genome during production of recombinant viral particles. Examples of heterologous promoters which can be used include, for example, viral simian vims 40 (SV40) (e.g., early or late), cytomegalovirus (CMV) (e.g., immediate early), Moloney murine leukemia vims (MoMLV), Rous sarcoma vims (RSV), and herpes simplex vims (HSV) (thymidine kinase) promoters.

[0387] In particular embodiments, a lentiviral vector integrates into the host cell genome. In certain embodiments, a lentiviral vector is integration defective, episomal, and does not integrate in the cell genome. As used herein, the term “integration defective lentivirus” or “IDLV” refers to a lentivirus having an integrase that lacks the capacity to integrate the viral vector into the host cell genome. Integration-incompetent viral vectors have been described in patent application WO 2006 / 010834, which is herein incorporated by reference in its entirety. Illustrative mutations in HIV-1 integrase suitable to reduce integrase activity include, but are not limited to: H12N, H12C, H16C, H16V, S81R, D41A, K42A, H51A, Q53C, D55V, D64E, D64V, E69A, K71A, E85A, E87A, D116N, D116I, D116A, N120G, N120I, N120E, E152G, E152A, K156E, K156A, E157A, K159E, K159A, K160A, R166A, D167A, E170A, H171A, K173A, K186Q, K186T, K188T, E198A, R199C, R199T, R199A, D202A, K211A, Q214L, Q216L, Q221L, W235F, W235E, K236S, K236A, K246A, G247W, D253A, R262A, R263A and K264H. In particular embodiments, an HIV- 1 integration deficient integrase comprises a D64V, D161I, D116A, E152G, or E152A mutation; D64V, DI 16A, and E152G mutations; D64V, DI 16A, and E152A mutations; or a D64V mutation.

[0388] In preferred embodiments, a recombinant particle, e.g., a lentiviral particle, comprises a lentiviral vector comprising a polynucleotide encoding a promoter operably linked to a polynucleotide encoding an engineered receptor, e.g., a chimeric antigen receptor (CAR), a chimeric costimulatory receptor (CCR), an alpha-beta T cell receptor (aP TCR), a gamma delta T cell receptor (y5 TCR), a dimerizing agent regulated immunoreceptor complex (DARIC), a chimeric TGF-P receptor (CTBR), or a zetakine receptor.

[0389] In particular embodiments, a recombinant particle, e.g., a lentiviral particle, comprises a lentiviral vector comprising a polynucleotide encoding a promoter operably linked to a polynucleotide encoding an engineered receptor that binds an antigen selected from the group consisting of alpha folate receptor (FRa), avP6 integrin, BAFFR, 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), claudin 6, (CEDN6), claudin 18 isoform 2 (CEDN18.2), C-type lectin-like molecule-1 (CEE-1), CD2 subset 1 (CS-1), chondroitin sulfate proteoglycan 4 (CSPG4), cutaneous T cell lymphoma- associated antigen 1 (CTAGE1), delta like canonical Notch ligand 3 (DLL3), epidermal growth factor receptor (EGFR), epidermal growth factor receptor variant III (EGFRvIII), epithelial glycoprotein 2 (EGP2), epithelial glycoprotein 40 (EGP40), epithelial cell adhesion molecule (EPCAM), ephrin type-A receptor 2 (EPHA2), erb-b2 receptor tyrosine kinase 4 (ERBB4), fibroblast activation protein (FAP), Fc Receptor Like 5 (FCRL5), fetal acetylcholinesterase receptor (AchR), ganglioside G2 (GD2), ganglioside G3 (GD3), Glypican-3 (GPC3), G Protein-Coupled Receptor Class C Group 5 Member D (GPCR5D), EGFR family including ErbB2 (HER2), HER2 p95, IL-lORa, IL-13Ra2, Kappa, cancer / testis antigen 2 (LAGE- 1 A), Lambda, Lewis-Y (LeY), LI cell adhesion molecule (LI -CAM), melanoma antigen gene (MAGE)-Al, MAGE- A3, MAGE-A4, MAGE-A6, MAGEA10, melanoma antigen recognized by T cells 1 (MelanA or MARTI), Mesothelin (MSLN), MUC1, MUC16, MHC class I chain related proteins A (MICA), MHC class I chain related proteins B (MICB), neural cell adhesion molecule (NCAM), cancer / testis antigen 1 (NY-ESO-1); placenta- specific 1 (PLAC1), preferentially expressed antigen in melanoma (PRAME), prostate stem cell antigen (PSCA), prostate-specific membrane antigen (PSMA), receptor tyrosine kinase-like orphan receptor 1 (R0R1), SLAMF7, synovial sarcoma, X breakpoint 2 (SSX2), Survivin, TACI, tumor associated glycoprotein 72 (TAG72), 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) a. Chimeric antigen receptor (CAR)

[0390] A CAR is a molecule that combines antibody-based specificity for a target antigen with one or more intracellular T cell signaling domains to generate a chimeric protein that exhibits a specific anti- tumor cellular immune activity. In particular embodiments, a recombinant particle, e.g., a retroviral or lentiviral particles, comprises (a) a phospholipid bilayer, a cellular membrane, or a viral envelope comprising (i) one or more mutated viral envelope glycoproteins that retain fusogenic activity and lack cognate receptor binding activity and (ii) a non- viral membrane-bound tropism polypeptide comprising an antibody or antigen binding fragment thereof that binds an antigen expressed on an immune effector cell, a spacer polypeptide, a transmembrane domain, and optionally, an intracellular domain comprising no more than 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 amino acids; and (b) a recombinant retroviral or lentiviral vector comprising a polynucleotide encoding a promoter operably linked to a polynucleotide encoding a CAR. In particular embodiments, a retroviral or lentiviral vector comprises a polynucleotide comprising or encoding a promoter operably linked to a polynucleotide encoding a CAR comprising a signal peptide, an extracellular antigen binding domain that specifically binds to a target polypeptide expressed on the surface of a cell, a hinge domain, a transmembrane domain, and one or more intracellular signaling domains, and optionally a polynucleotide encoding a posttranscriptional regulatory element, e.g., a WPRE or HPRE.

[0391] In particular embodiments, a CAR comprises an extracellular antigen binding domain that comprises an antibody or antigen binding fragment thereof.

[0392] In particular embodiments, a CAR comprises an extracellular antigen binding domain that comprises an antibody or antigen binding fragment thereof selected from the group consisting of: a Camel Ig, a Llama Ig, an Alpaca Ig, Ig NAR, a Fab' fragment, a F(ab')2 fragment, a bispecific Fab dimer (Fab2), a trispecific Fab trimer (Fab3), an Fv, a single chain Fv protein (“scFv”), a bis-scFv, (scFv , a minibody, a diabody, a triabody, a tetrabody, a disulfide stabilized Fv protein (“dsFv”), a Fibronectin type III (FN3) domain antibody, a single-domain antibody (sdAb or nanobody, e.g., a camelid VHH or shark VNAR) and a centyrin that binds an antigen selected from the group consisting of FRa, avP6 integrin, BAFFR, BCMA, CD276, B7-H6, CAIX, CD16, CD19, CD20, CD22, CD30, CD33, CD37, CD38, CD44, CD44v6, CD44v7 / 8, CD70, CD79a, CD79b, CD123, CD133, CD138, CD171, CEA, CLDN6, CLDN18.2, CLL-1, CS-1, CSPG4, CTAGE1, DLL3, EGFR, EGFRvIII, EGP2, EGP40, EPCAM, EPHA2, ERBB4, FAP, FCRL5, AchR, GD2, GD3, GPC3, GPCR5D, HER2, HER2p95, IL-lORa, IL-13Ra2, Kappa, LAGE- 1 A, Lambda, LeY, Ll-CAM, MAGE-A1, MAGE-A3, MAGE-A4, MAGE-A6, MAGEA10, MARTI, MSLN, MUC1, MUC16, MICA, MICB, NCAM, NY-ESO-1, PLAC1, PRAME, PSCA, PSMA, R0R1, SLAMF7, SSX2, Survivin, TACI, TAG72, TEM1 / CD248, TEM7R, TPBG, ULBP1, ULBP2, ULBP3, ULBP4, ULBP5, ULBP6, VEGFR2, and WT-1; a hinge domain selected from the group consisting of a CD4 hinge, a CD8P hinge, a CD8a hinge, a CD28 hinge, a CD 134 hinge, a CD 137 hinge, a CD 152 hinge, a CD278 hinge, an IgGl hinge, an IgG2 hinge, an IgG3 hinge, an IgG4 hinge; a transmembrane domain isolated or derived from a polypeptide selected from the group consisting of an alpha, beta, gamma, or delta chain of the T-cell receptor, CD35, CD3s, CD3y, CD3^, CD4, CD5, CD8a, CD9, CD16, CD22, CD27, CD28, CD33, CD37, CD45, CD64, CD80, CD86, CD134, CD137, CD152, CD154, CD278, amnionless (AMN), and programmed cell death 1 (PDCD1); a primary signaling domain isolated or derived from a polypeptide selected from the group consisting of FcRy, FcRp, CD3y, CD35, CD3s, CD3^ CD22, CD79a, CD79b, and CD66d; and optionally, one or more costimulatory signaling domains isolated or derived from a polypeptide selected from the group consisting of TLR1, TLR2, TLR3, TLR4, TLR5, TLR6, TLR7, TLR8, TLR9, TLR10, CARD11, CD2, CD7, CD27, CD28, CD30, CD40, ICAM, CD83, CD94, CD134 (0X40), CD137 (4-1BB), CD278 (ICOS), DAP10, LAT, SLP76, TRAT1, TNFR2, TNFRS14, TNFRS18, TNFRS25, andZAP70.

[0393] In particular embodiments, a retroviral or lentiviral vector comprises a polynucleotide comprising or encoding a promoter operably linked to a polynucleotide encoding a CAR having an amino acid sequence set forth in any one of SEQ ID NOs: 662-666. Table 12 | | DTYDALHMQALPPR b. Chimeric costimulatory receptor (CCR)

[0394] A chimeric costimulatory receptor (CCR) is a molecule that combines antibodybased specificity to a desired antigen with a costimulatory domain but that lacks a primary signaling domain. CCRs redirect immune effector cell specificity in an MHC independent manner and enhance the immune effector cell response in the presence of a CAR. In particular embodiments, a CAR comprises a first costimulatory domain and a primary signaling domain and a CCR comprises a second co stimulatory domain, wherein the first and second co stimulatory domains are different from each other and synergize to immune effector cell proliferation, persistence, cytokine production, and / or phagocytosis or production of molecules that can mediate cell death of the target antigen expressing cell.

[0395] In particular embodiments, a retroviral or lentiviral vector comprises a polynucleotide comprising or encoding a promoter operably linked to a polynucleotide encoding a CAR comprising a signal peptide, an extracellular antigen binding domain that specifically binds to a target polypeptide expressed on the surface of a cell, a hinge domain, a transmembrane domain, and one or more intracellular signaling domains; a polypeptide cleavage signal; a CCR that comprises a signal peptide, an extracellular antigen binding domain, a hinge domain, a transmembrane domain, and a costimulatory signaling domain; and optionally a polynucleotide encoding a posttranscriptional regulatory element, e.g., a WPRE or HPRE.

[0396] In particular embodiments, the signal peptide of the CAR is the same as, or different from, the signal peptide of the CCR; the extracellular antigen binding domain of the CAR is different than the extracellular antigen binding domain of the CCR; the hinge domain of the CAR is the same as, or different from, the hinge domain of the CCR; the transmembrane domain of the CAR is the same as, or different from, the transmembrane domain of the CCR; the costimulatory domain of the CAR is the same as, or different from, the costimulatory domain of the CCR; and the CAR comprises a primary signaling domain.

[0397] In particular embodiments, a retroviral or lentiviral vector comprises a polynucleotide comprising or encoding a promoter operably linked to (a) a polynucleotide encoding a CAR comprising an extracellular antigen binding domain that comprises an antibody or antigen binding fragment thereof selected from the group consisting of: a Camel Ig, a Llama Ig, an Alpaca Ig, Ig NAR, a Fab' fragment, a F(ab')2 fragment, a bispecific Fab dimer (Fab2), a trispecific Fab trimer (Fab3), an Fv, a single chain Fv protein (“scFv”), a bis-scFv, (scFv)2, a minibody, a diabody, a triabody, a tetrabody, a disulfide stabilized Fv protein (“dsFv”), a Fibronectin type III (FN3) domain antibody, a single-domain antibody (sdAb or nanobody, e.g., a camelid VHH or shark VNAR) and a centyrin that binds an antigen selected from the group consisting of FRa, avP6 integrin, BAFFR, BCMA, CD276, B7-H6, CAIX, CD 16, CD19, CD20, CD22, CD30, CD33, CD37, CD38, CD44, CD44v6, CD44v7 / 8, CD70, CD79a, CD79b, CD123, CD133, CD138, CD171, CEA, CLDN6, CLDN18.2, CLL-1, CS- 1, CSPG4, CTAGE1, DLL3, EGFR, EGFRvin, EGP2, EGP40, EPCAM, EPHA2, ERBB4, FAP, FCRL5, AchR, GD2, GD3, GPC3, GPCR5D, HER2, HER2 p95, IL-lORa, IL-13Ra2, Kappa, LAGE-1A, Lambda, LeY, Ll-CAM, MAGE-A1, MAGE-A3, MAGE-A4, MAGE- A6, MAGEA10, MARTI, MSLN, MUC1, MUC16, MICA, MICB, NCAM, NY-ESO-1, PLAC1, PRAME, PSCA, PSMA, R0R1, SLAMF7, SSX2, Survivin, TACI, TAG72, TEM1 / CD248, TEM7R, TPBG, ULBP1, ULBP2, ULBP3, ULBP4, ULBP5, ULBP6, VEGFR2, and WT-1; a hinge domain selected from the group consisting of a CD4 hinge, a CD8P hinge, a CD8a hinge, a CD28 hinge, a CD134 hinge, a CD137 hinge, a CD152 hinge, a CD278 hinge, an IgGl hinge, an IgG2 hinge, an IgG3 hinge, an IgG4 hinge; a transmembrane domain isolated or derived from a polypeptide selected from the group consisting of an alpha, beta, gamma, or delta chain of the T-cell receptor, CD35, CD3s, CD3y, CD3 , CD4, CD5, CD8a, CD9, CD16, CD22, CD27, CD28, CD33, CD37, CD45, CD64, CD80, CD86, CD134, CD137, CD152, CD154, CD278, amnionless (AMN), and programmed cell death 1 (PDCD1); a primary signaling domain isolated or derived from a polypeptide selected from the group consisting of FcRy, FcRp, CD3y, CD35, CD3s, CD3^, CD22, CD79a, CD79b, and CD66d; and optionally, one or more costimulatory signaling domains isolated or derived from a polypeptide selected from the group consisting of TLR1, TLR2, TLR3, TLR4, TLR5, TLR6, TLR7, TLR8, TLR9, TLR10, CARD11, CD2, CD7, CD27, CD28, CD30, CD40, ICAM, CD83, CD94, CD134 (0X40), CD137 (4-1BB), CD278 (ICOS), DAP10, LAT, SLP76, TRAT1, TNFR2, TNFRS14, TNFRS18, TNFRS25, and ZAP70; (b) a self-cleaving viral polypeptide or ribosomal skipping polypeptide; (c) a polynucleotide encoding a CCR that comprises an antibody or antigen binding fragment thereof selected from the group consisting of: a Camel Ig, a Llama Ig, an Alpaca Ig, Ig NAR, a Fab' fragment, a F(ab')2 fragment, a bispecific Fab dimer (Fab2), a trispecific Fab trimer (Fab3), an Fv, a single chain Fv protein (“scFv”), a bis-scFv, (scFv)2, a minibody, a diabody, a triabody, a tetrabody, a disulfide stabilized Fv protein (“dsFv”), a Fibronectin type III (FN3) domain antibody, a single-domain antibody (sdAb or nanobody, e.g., a camelid VHH or shark VNAR) and a centyrin that binds an antigen selected from the group consisting of FRa, avP6 integrin, BAFFR, BCMA, CD276, B7-H6, CAIX, CD16, CD19, CD20, CD22, CD30, CD33, CD37, CD38, CD44, CD44v6, CD44v7 / 8, CD70, CD79a, CD79b, CD123, CD133, CD138, CD171, CEA, CLDN6, CLDN18.2, CLL-1, CS-1, CSPG4, CTAGE1, DLL3, EGFR, EGFRvIII, EGP2, EGP40, EPCAM, EPHA2, ERBB4, FAP, FCRL5, AchR, GD2, GD3, GPC3, GPCR5D, HER2, HER2 p95, IL-lORa, IL-13Ra2, Kappa, LAGE-1A, Lambda, LeY, Ll-CAM, MAGE-A1, MAGE-A3, MAGE-A4, MAGE-A6, MAGEA10, MARTI, MSLN, MUC1, MUC16, MICA, MICB, NCAM, NY-ESO-1, PLAC1, PRAME, PSCA, PSMA, ROR1, SLAMF7, SSX2, Survivin, TACI, TAG72, TEM1 / CD248, TEM7R, TPBG, ULBP1, ULBP2, ULBP3, ULBP4, ULBP5, ULBP6, VEGFR2, and WT-1 that is different than the antigen bound by the CAR; a hinge domain selected from the group consisting of a CD4 hinge, a CD8P hinge, a CD8a hinge, a CD28 hinge, a CD134 hinge, a CD 137 hinge, a CD 152 hinge, a CD278 hinge, an IgGl hinge, an IgG2 hinge, an IgG3 hinge, an IgG4 hinge, optionally that comprises one or more amino acid substitutions that reduces antigen independent signaling of the CCR compared to the unmodified hinge domain; a transmembrane domain isolated or derived from a polypeptide selected from the group consisting of an alpha, beta, gamma, or delta chain of the T-cell receptor, CD35, CD3s, CD3y, CD3 , CD4, CD5, CD8a, CD9, CD16, CD22, CD27, CD28, CD33, CD37, CD45, CD64, CD80, CD86, CD134, CD137, CD152, CD154, CD278, amnionless (AMN), and programmed cell death 1 (PDCD1); and one or more co stimulatory signaling domains isolated or derived from a polypeptide selected from the group consisting of TLR1, TLR2, TLR3, TLR4, TLR5, TLR6, TLR7, TLR8, TLR9, TLR10, CARD11, CD2, CD7, CD27, CD28, CD30, CD40, ICAM, CD83, CD94, CD134 (0X40), CD137 (4-1BB), CD278 (ICOS), DAP10, LAT, SLP76, TRAT1, TNFR2, TNFRS14, TNFRS18, TNFRS25, and ZAP70 that are different than the costimulatory domain of the CAR. c. Engineered T Cell Receptors (TCR)

[0398] Naturally occurring T cell receptors have two chains: an aPTCR comprises an alpha chain and a beta chain and a ySTCR comprises a gamma chain and a delta chain. Each TCR is a unique protein produced by a recombination event in each T cell’s genome. Libraries of TCRs may be screened for their selectivity to particular target antigens and natural TCRs with high-avidity and reactivity toward target antigens may be selected, cloned, and incorporated into a payload to deliver to an immune effector cell.

[0399] In particular embodiments, a recombinant particle, e.g., a retroviral or lentiviral particles, comprises (a) a phospholipid bilayer, a cellular membrane, or a viral envelope comprising (i) one or more mutated viral envelope glycoproteins that retain fusogenic activity and lack cognate receptor binding activity and (ii) a non- viral membrane-bound tropism polypeptide comprising an antibody or antigen binding fragment thereof that binds an antigen expressed on an immune effector cell, a spacer polypeptide, a transmembrane domain, and optionally, an intracellular domain comprising no more than 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 amino acids; and (b) a recombinant retroviral or lentiviral vector comprising a polynucleotide encoding a promoter operably linked to a polynucleotide encoding an alpha chain and / or a beta chain of an aPTCR or a gamma chain and / or a delta chain of a ySTCR.

[0400] In particular embodiments, a retroviral or lentiviral vector comprises a polynucleotide comprising or encoding a promoter operably linked to a polynucleotide encoding an alpha chain and / or a beta chain of an aPTCR or a gamma chain and / or a delta chain of a ySTCR, and optionally a polynucleotide encoding a posttranscriptional regulatory element, e.g., a WPRE or HPRE. In particular embodiments, the vector comprises a polynucleotide encoding a self-cleaving viral polypeptide or ribosomal skipping polypeptide disposed between the chains of the aPTCR or ySTCR.

[0401] In particular embodiments, an engineered TCR comprises an extracellular antigen binding domain that binds an antigen selected from the group consisting of FRa, avP6 integrin, BAFFR, BCMA, CD276, B7-H6, CAIX, CD16, CD19, CD20, CD22, CD30, CD33, CD37, CD38, CD44, CD44v6, CD44v7 / 8, CD70, CD79a, CD79b, CD123, CD133, CD138, CD171, CEA, CLDN6, CLDN18.2, CLL-1, CS-1, CSPG4, CTAGE1, DLL3, EGFR, EGFRvIII, EGP2, EGP40, EPCAM, EPHA2, ERBB4, FAP, FCRL5, AchR, GD2, GD3, GPC3, GPCR5D, HER2, HER2 p95, IL-lORa, IL-13Ra2, Kappa, LAGE-1A, Lambda, LeY, Ll-CAM, MAGE-A1, MAGE-A3, MAGE-A4, MAGE-A6, MAGEA10, MARTI, MSLN, MUC1, MUC16, MICA, MICB, NCAM, NY-ESO-1, PLAC1, PRAME, PSCA, PSMA, ROR1, SLAMF7, SSX2, Survivin, TACI, TAG72, TEM1 / CD248, TEM7R, TPBG, ULBP1, ULBP2, ULBP3, ULBP4, ULBP5, ULBP6, VEGFR2, and WT-1. d. Dimerizing agent regulated immunoreceptor complex

[0402] (DARIC)

[0403] A DARIC is a multi-component signaling receptor comprising one or more non- naturally occurring polypeptides that transduces an immunostimulatory signal in an immune effector cell upon exposure to a multimerizing agent or bridging factor, e.g., stimulating immune effector cell activity and function, increasing production and / or secretion of proinflammatory cytokines.

[0404] In particular embodiments, a recombinant particle, e.g., a retroviral or lentiviral particles, comprises (a) a phospholipid bilayer, a cellular membrane, or a viral envelope comprising (i) one or more mutated viral envelope glycoproteins that retain fusogenic activity and lack cognate receptor binding activity and (ii) a non- viral membrane-bound tropism polypeptide comprising an antibody or antigen binding fragment thereof that binds an antigen expressed on an immune effector cell, a spacer polypeptide, a transmembrane domain, and optionally, an intracellular domain comprising no more than 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 amino acids; and (b) a recombinant retroviral or lentiviral vector comprising a polynucleotide encoding a promoter operably linked to a polynucleotide encoding a DARIC signaling component and a DARIC binding component.

[0405] In particular embodiments, a retroviral or lentiviral vector comprises a polynucleotide comprising or encoding a promoter operably linked to a polynucleotide encoding a DARIC signaling component comprising a signal peptide, a multimerization domain, a transmembrane domain, a costimulatory domain and / or a primary signaling domain; a selfcleaving viral polypeptide or ribosomal skipping polypeptide; and a DARIC binding component comprising a signal peptide, an extracellular antigen binding domain, a multimerization domain, a transmembrane domain, and optionally, a costimulatory domain, and optionally a polynucleotide encoding a posttranscriptional regulatory element, e.g., a WPRE or HPRE; wherein a bridging factor regulates association of the DARIC binding component and DARIC signaling component by interacting with the multimerization domains.

[0406] In particular embodiments, a DARIC signaling component comprises a multimerization domain selected from the group consisting of an FK506 binding protein (FKBP) polypeptide or variants thereof (e.g., FKBP12), or an FKBP-rapamycin binding (FRB) polypeptide or variants thereof (e.g., T2098L); a transmembrane domain isolated or derived from a polypeptide selected from the group consisting of an alpha, beta, gamma, or delta chain of the T-cell receptor, CD35, CD3s, CD3y, CD3^, CD4, CD5, CD8a, CD9, CD16, CD22, CD27, CD28, CD33, CD37, CD45, CD64, CD80, CD86, CD134, CD137, CD152, CD154, CD278, amnionless (AMN), and programmed cell death 1 (PDCD1); one or more costimulatory signaling domains isolated or derived from a polypeptide selected from the group consisting of TLR1, TLR2, TLR3, TLR4, TLR5, TLR6, TLR7, TLR8, TLR9, TLR10, CARD11, CD2, CD7, CD27, CD28, CD30, CD40, ICAM, CD83, CD94, CD134 (0X40), CD137 (4-1BB), CD278 (ICOS), DAP10, LAT, SLP76, TRAT1, TNFR2, TNFRS14, TNFRS18, TNFRS25, and ZAP70; and a primary signaling domain isolated or derived from a polypeptide selected from the group consisting of FcRy, FcRp, CD3y, CD35, CD3s, CD3 , CD22, CD79a, CD79b, and CD66d.

[0407] In particular embodiments, a DARIC binding component comprises an extracellular antigen binding domain comprising an antibody or antigen binding fragment thereof selected from the group consisting of a Camel Ig, a Llama Ig, an Alpaca Ig, Ig NAR, a Fab' fragment, a F(ab')2 fragment, a bispecific Fab dimer (Fab2), a trispecific Fab trimer (Fab3), an Fv, a single chain Fv protein (“scFv”), a bis-scFv, (scFv , a minibody, a diabody, a triabody, a tetrabody, a disulfide stabilized Fv protein (“dsFv”), a Fibronectin type III (FN3) domain antibody, and a single-domain antibody (sdAb or nanobody, e.g., a camelid VHH or shark VNAR) that binds an antigen selected from the group consisting of FRa, avP6 integrin, BAFFR, BCMA, CD276, B7-H6, CAIX, CD16, CD19, CD20, CD22, CD30, CD33, CD37, CD38, CD44, CD44v6, CD44v7 / 8, CD70, CD79a, CD79b, CD123, CD133, CD138, CD171, CEA, CLDN6, CLDN18.2, CLL-1, CS-1, CSPG4, CTAGE1, DLL3, EGFR, EGFRvIII, EGP2, EGP40, EPCAM, EPHA2, ERBB4, FAP, FCRL5, AchR, GD2, GD3, GPC3, GPCR5D, HER2, HER2p95, IL-lORa, IL-13Ra2, Kappa, LAGE- 1 A, Lambda, LeY, Ll-CAM, MAGE-A1, MAGE-A3, MAGE-A4, MAGE-A6, MAGEA10, MARTI, MSLN, MUC1, MUC16, MICA, MICB, NCAM, NY-ESO-1, PLAC1, PRAME, PSCA, PSMA, R0R1, SLAMF7, SSX2, Survivin, TACI, TAG72, TEM1 / CD248, TEM7R, TPBG, ULBP1, ULBP2, ULBP3, ULBP4, ULBP5, ULBP6, VEGFR2, and WT-1; a multimerization domain selected from the group consisting of an FK506 binding protein (FKBP) polypeptide or variants thereof (e.g., FKBP12), or an FKBP-rapamycin binding (FRB) polypeptide or variants thereof (e.g., T2098L); a transmembrane domain isolated or derived from a polypeptide selected from the group consisting of an alpha, beta, gamma, or delta chain of the T-cell receptor, CD35, CD3s, CD3y, CD3^, CD4, CD5, CD8a, CD9, CD16, CD22, CD27, CD28, CD33, CD37, CD45, CD64, CD80, CD86, CD134, CD137, CD152, CD154, CD278, amnionless (AMN), and programmed cell death 1 (PDCD1); and optionally one or more costimulatory signaling domains isolated or derived from a polypeptide selected from the group consisting of TLR1, TLR2, TLR3, TLR4, TLR5, TLR6, TLR7, TLR8, TLR9, TLR10, CARD11, CD2, CD7, CD27, CD28, CD30, CD40, ICAM, CD83, CD94, CD134 (0X40), CD137 (4-1BB), CD278 (ICOS), DAP10, LAT, SLP76, TRAT1, TNFR2, TNFRS14, TNFRS18, TNFRS25, andZAP70.

[0408] Illustrative examples of bridging factors suitable for use in particular embodiments comprising a DARIC receptor contemplated herein include, but are not limited to, AP1903, AP20187, AP21967 (also known as C-16-(S)-7-methylindolerapamycin), everolimus, novolimus, pimecrolimus, ridaforolimus, tacrolimus, temsirolimus, umirolimus, and zotarolimus. e. Chimeric TGF-p receptor (CTBR)

[0409] A chimeric TGF-P receptor (CTBR) is a fusion polypeptide that when expressed on an immune effector cell, transduces an immunostimulatory signal to the immune effector cell in the presence of immunosuppressive cytokines (e.g., TGFP) produced by tumor cells and tumor-infiltrating lymphocytes in the tumor microenvironment (TME).

[0410] In particular embodiments, a retroviral or lentiviral vector comprises a polynucleotide comprising or encoding a promoter operably linked to a polynucleotide encoding a CAR, aP TCR, y5 TCR, DARIC, or zetakine; a self-cleaving viral polypeptide or ribosomal skipping polypeptide; a CTBR; and optionally a polynucleotide encoding a posttranscriptional regulatory element, e.g., a WPRE or HPRE.

[0411] In particular embodiments, a CTBR comprises (a) a first polypeptide comprising an extracellular TGFpi -binding domain of TGFPR2, a transmembrane domain, and a first interleukin receptor intracellular signaling domain; (b) a polypeptide cleavage signal; and (c) a second polypeptide comprising an extracellular TGFpi -binding domain of TGFpRl, a transmembrane domain, and a second interleukin receptor intracellular signaling domain.

[0412] Illustrative examples of first and second interleukin receptor intracellular signaling domains suitable for use in a CTBR include but are not limited to IL-12RP1 and IL-12RP2 (IL- 12 receptor); IL-7Ra and IL-2Ry (IL-7 receptor); IL-2Ry and IL-2RY (IL-2 receptor); IL- 21R and IL-2Ry (IL-21 receptor); IL-18R1 and IL-18RAP (IL- 18 receptor); and IL-1R1 and IL- 1 RAP (IL-1 receptor).

[0413] In particular embodiments, a retroviral or lentiviral vector comprises a polynucleotide comprising or encoding a promoter operably linked to (a) a CAR, aP TCR, y5 TCR, DARIC, or zetakine; (b) a self-cleaving viral polypeptide or ribosomal skipping polypeptide; and (c) a polynucleotide encoding a CTBR comprising (i) a first polypeptide comprising an extracellular TGFpi -binding domain of TGFPR2, a TGFPR2 or an IL-12RP2 transmembrane domain and an IL-12RP2 intracellular signaling domain; (ii) a self-cleaving viral peptide or ribosomal skipping polypeptide and (iii) a second polypeptide comprising an extracellular TGFpi -binding domain of TGFpRl; a TGFpRl or an IL-12RP1 transmembrane domain, and an IL-12RP1 intracellular signaling domain.

[0414] In particular embodiments, a retroviral or lentiviral vector comprises a polynucleotide comprising or encoding a promoter operably linked to (a) a CAR, aP TCR, y5 TCR, DARIC, or zetakine; (b) a self-cleaving viral polypeptide or ribosomal skipping polypeptide; and (c) a polynucleotide encoding a CTBR comprising (i) a first polypeptide comprising an extracellular TGFpi -binding domain of TGFPR2, a TGFPR2 or an IL-7Ra transmembrane domain and an IL-7Ra intracellular signaling domain; (ii) a self-cleaving viral peptide or ribosomal skipping polypeptide; and (iii) a second polypeptide comprising an extracellular TGFpi -binding domain of TGFpRl; a TGFpRl or an IL-2Ry transmembrane domain, and an IL-2Ry intracellular signaling domain.

[0415] In particular embodiments, a retroviral or lentiviral vector comprises a polynucleotide comprising or encoding a promoter operably linked to (a) a CAR, aP TCR, y5 TCR, DARIC, or zetakine; (b) a self-cleaving viral polypeptide or ribosomal skipping polypeptide; and (c) a polynucleotide encoding a CTBR comprising (i) a first polypeptide comprising an extracellular TGFpi -binding domain of TGFPR2, a TGFPR2 or an IL-2RP transmembrane domain and an IL-2RP intracellular signaling domain; (ii) a self-cleaving viral peptide or ribosomal skipping polypeptide; and (iii) a second polypeptide comprising an extracellular TGFpi -binding domain of TGFpRl; a TGFpRl or an IL-2Ry transmembrane domain, and an IL-2Ry intracellular signaling domain.

[0416] In particular embodiments, a retroviral or lentiviral vector comprises a polynucleotide comprising or encoding a promoter operably linked to (a) a CAR, aP TCR, y5 TCR, DARIC, or zetakine; (b) a self-cleaving viral polypeptide or ribosomal skipping polypeptide; and (c) a polynucleotide encoding a CTBR comprising (i) a first polypeptide comprising an extracellular TGFpi -binding domain of TGFPR2, a TGFPR2 or an IL-21R transmembrane domain and an IL-21R intracellular signaling domain; (ii) a self-cleaving viral peptide or ribosomal skipping polypeptide; and (iii) a second polypeptide comprising an extracellular TGFpi -binding domain of TGFpRl; a TGFpRl or an fL-2Ry transmembrane domain, and an fL-2Ry intracellular signaling domain.

[0417] 2. GENE EDITI G PA YLOADS

[0418] Recombinant particles contemplated in particular embodiments comprise (i) a surface comprising one or more mutated viral envelope glycoproteins that retain fusogenic activity and lack cognate receptor binding activity and one or more non- viral membranebound tropism polypeptides that bind an antigen expressed on a cell and (ii) a payload comprising a genome editing composition comprising one or more polynucleotides and / or polypeptides. Without wishing to be bound to any particular theory it is believed that genome editing compositions can be delivered using the recombinant particles contemplated herein to modify the target cell genome and introduce one or more exogenous genes or modulate expression of one or more endogenous genes in the cell.

[0419] In a particular embodiment, a genome editing payload comprises a template and a nuclease both formatted as circRNAs, mRNAs or DNAs.

[0420] In a particular embodiment, a genome editing payload comprises a template formatted as RNA or DNA and a nuclease formatted as a polypeptide.

[0421] In a particular embodiment, a genome editing payload comprises a nucleic acid- polypeptide complex comprising a template formatted as RNA or DNA and a nuclease formatted as a polypeptide.

[0422] In a particular embodiment, a genome editing payload comprises a nuclease formatted as circRNA, mRNA, DNA, or polypeptide.

[0423] In particular embodiments, a genome editing payload comprises a fusion protein comprising a viral accessory protein and a nuclease and optionally, a DNA repair template.

[0424] In particular embodiments, a genome editing payload comprises a fusion protein comprising a lentiviral accessory protein, a lentivirus protease cleavage site (e.g., SQNYPIVQ (SEQ ID NO: 714)) and a nuclease and optionally, a DNA repair template.

[0425] In particular embodiments, the viral accessory protein is a gag or vpr polypeptide, and in preferred embodiments, a lentiviral (e.g., HIV-1) gag or vpr polypeptide. In particular embodiments, the lentivirus protease cleavage site is an HIV-1 protease cleavage site. Illustrative examples of genome editing compositions suitable for use in particular embodiments contemplated herein include, but are not limited to base editors (e.g., cytosine base editors (Cas9-cytosine deaminase fusion proteins; CBEs), adenosine base editors (Cas9- deoxyadenosine deaminase fusion proteins; ABEs), sgRNA), prime editors (Cas9-reverse trancriptase fusion proteins; pegRNAs), clustered regularly -interspaced short palindromic repeats (CRISPR) / Cas nuclease systems (Cas9, sgRNA), retrotransposons, homing endonucleases (meganucleases), megaTALs, transcription activator-like effector nucleases (TALENs), zinc finger nucleases (ZFNs), and ARCUS nucleases.

[0426] F. POLYNUCLEOTIDES

[0427] Polynucleotides comprising vectors, vector-based genomes, transfer vectors, packaging plasmids, expression control sequences, therapeutic polynucleotides, nucleases, DNA repair templates and polynucleotides encoding polypeptides, fusion polypeptides, viral envelope glycoproteins, non-viral membrane-bound tropism polypeptides, CARs, CCRs, TCRs, DARICs, CTBRs, nucleases, and therapeutic polypeptides and variants thereof are contemplated herein.

[0428] As used herein, the terms “polynucleotide” or “nucleic acid” refer to deoxyribonucleic acid (DNA), ribonucleic acid (RNA) and DNA / RNA hybrids. Polynucleotides may be single-stranded or double-stranded and either recombinant, synthetic, or isolated. Polynucleotides include but are not limited to: pre-messenger RNA (pre-mRNA), messenger RNA (mRNA), RNA, circular RNA (circRNA), synthetic RNA, short interfering RNA (siRNA), short hairpin RNA (shRNA), microRNA (miRNA), ribozymes, genomic RNA (gRNA), viral genomic RNA, plus strand RNA (RNA(+)), minus strand RNA (RNA(-)), tracrRNA, crRNA, single guide RNA (sgRNA), PCR amplified DNA, complementary DNA (cDNA), synthetic DNA, or recombinant DNA. Polynucleotides refer to a polymeric form of nucleotides of at least 5, at least 10, at least 15, at least 20, at least 25, at least 30, at least 40, at least 50, at least 100, at least 200, at least 300, at least 400, at least 500, at least 1000, at least 5000, at least 10000, or at least 15000 or more nucleotides in length, either ribonucleotides or deoxyribonucleotides or a modified form of either type of nucleotide, as well as all intermediate lengths. It will be readily understood that “intermediate lengths,” in this context, means any length between the quoted values, such as 6, 7, 8, 9, etc., 101, 102, 103, etc., 151, 152, 153, etc., 201, 202, 203, etc. In particular embodiments, polynucleotides contemplated herein are polynucleotide variants. As used herein, the terms “polynucleotide variant” and “variant” and the like refer to polynucleotides displaying substantial sequence identity with a reference polynucleotide sequence or polynucleotides that hybridize with a reference sequence under stringent conditions that are defined hereinafter. These terms also encompass polynucleotides that are distinguished from a reference polynucleotide by the addition, deletion, substitution, or modification of one or more nucleotides. Accordingly, the terms “polynucleotide variant” and “variant” include polynucleotides in which one or more nucleotides have been added or deleted, or modified, or replaced with different nucleotides. In this regard, it is well understood in the art that certain alterations inclusive of mutations, additions, deletions and substitutions can be made to a reference polynucleotide whereby the altered polynucleotide retains the biological function or activity of the reference polynucleotide or wherein the function or activity of the altered polynucleotide is modulated.

[0429] In particular embodiments, polynucleotides or polynucleotide variants have at least or about 50%, 55%, 60%, 65%, 70%, 71%, 72%, 73%, 74%, 75%, 76%, 77%, 78%, 79%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to a reference sequence.

[0430] As used herein, “isolated polynucleotide” refers to a polynucleotide that has been isolated from or purified from the sequences which flank it in a naturally-occurring state. In particular embodiments, an isolated polynucleotide is a synthetic polynucleotide, a semisynthetic polynucleotide, or a polynucleotide obtained or derived from a recombinant source, or other polynucleotide that does not exist in nature and that has been made by the hand of man.

[0431] In various embodiments, a polynucleotide comprises a vector, optionally a retroviral vector, optionally a lentiviral vector.

[0432] In various embodiments, a polynucleotide comprises a retroviral vector-based RNA genome, optionally a lentiviral vector-based RNA genome.

[0433] Illustrative examples of polynucleotides include, but are not limited to, polynucleotide sequences set forth in any one of SEQ ID NOs: 667-672 and polynucleotides encoding polypeptides set forth in SEQ ID NOs: 1-666 and 673-714.

[0434] In particular embodiments, polynucleotides may be codon-optimized. As used herein, the term “codon-optimized” refers to substituting codons in a polynucleotide encoding a polypeptide in order to modulate polypeptide expression, stability and / or activity. Factors that influence codon optimization include, but are not limited to one or more of: (i) variation of codon biases between two or more organisms or genes or synthetically constructed bias tables, (ii) variation in the degree of codon bias within an organism, gene, or set of genes, (iii) systematic variation of codons including context, (iv) variation of codons according to their decoding tRNAs, (v) variation of codons according to GC %, either overall or in one position of the triplet, (vi) variation in degree of similarity to a reference sequence for example a naturally occurring sequence, (vii) variation in the codon frequency cutoff, (viii) structural properties of mRNAs transcribed from the DNA sequence, (ix) prior knowledge about the function of the DNA sequences upon which design of the codon substitution set is to be based, (x) systematic variation of codon sets for each amino acid, and / or (xi) isolated removal of spurious translation initiation sites.

[0435] Polynucleotides contemplated in particular embodiments, regardless of the length of the coding sequence itself, may be combined with other polynucleotide sequences, such as expression control sequences, promoters and / or enhancers, untranslated regions (UTRs), polynucleotides encoding signal peptides, Kozak sequences, polyadenylation signals, restriction enzyme sites, multiple cloning sites, internal ribosomal entry sites (IRES), recombinase recognition sites (e.g., LoxP, FRT, and Att sites), termination codons, transcriptional termination signals, and polynucleotides encoding self-cleaving polypeptides or epitope tags, as disclosed elsewhere herein or as known in the art.

[0436] In particular embodiments, a polynucleotide encodes a fusion polypeptide, viral envelope glycoprotein, non-viral membrane-bound tropism polypeptide, CAR, CCR, aP TCR, y5 TCR, DARIC, CTBR, nuclease, or therapeutic polypeptide comprising signal peptide, e.g., a polynucleotide encoding any one of SEQ ID NOs: 449-454.

[0437] Signal peptides are short 16 to 30 amino acid N-terminal sequences of nascently synthesized polypeptide chains that mediate protein targeting to the membrane of the endoplasmic reticulum (ER). Typically, signal peptides are cleaved cotranslationally by signal peptidase, a heterooligomeric polypeptide complex. In particular embodiments, a polypeptide comprises a signal peptide. In preferred embodiments, a polynucleotide encoding a polypeptide comprises a polynucleotide encoding a signal polypeptide; and the translated polypeptide does not comprise a signal peptide.

[0438] In particular embodiments, a vector comprises a polynucleotide comprising or encoding one or more exogenous, endogenous, or heterologous expression control sequences operably linked to a polynucleotide encoding one or more polynucleotides and / or polypeptides contemplated herein.

[0439] “Expression control sequences,” “control elements,” or “regulatory sequences” contemplated in particular embodiments include but not limited to promoters, enhancers, translation initiation signals (Shine Dalgamo sequence or Kozak sequence), introns, polyadenylation signals, 5' and 3' untranslated regions, all of which may interact with host cell proteins to carry out transcription and translation.

[0440] The term “promoter” as used herein refers to a recognition site of a polynucleotide (DNA or RNA) to which an RNA polymerase binds. An RNA polymerase initiates and transcribes polynucleotides operably linked to the promoter. In particular embodiments, promoters operative in mammalian cells comprise an AT-rich region located approximately 25 to 30 bases upstream from the site where transcription is initiated and / or another sequence found 70 to 80 bases upstream from the start of transcription, a CNCAAT region where N may be any nucleotide. The term “enhancer” refers to a segment of DNA which contains sequences capable of providing enhanced transcription and in some instances can function independent of their orientation relative to another control sequence. An enhancer can function cooperatively or additively with promoters and / or other enhancer elements. The term “promoter / enhancer” refers to a segment of DNA which contains sequences capable of providing both promoter and enhancer functions.

[0441] The term “operably linked”, refers to a juxtaposition wherein the components described are in a relationship permitting them to function in their intended manner. In one embodiment, the term refers to a functional linkage between an expression control sequence (such as a promoter, and / or enhancer) and a second polynucleotide sequence encoding a polypeptide, wherein the expression control sequence directs transcription of the nucleic acid corresponding to the second sequence.

[0442] Illustrative expression control sequences suitable for use in particular embodiments include, but are not limited to, a P-actin promoter, a cytomegalovirus (CMV) immediate early promoter, a simian virus 40 (SV40) (e.g., early or late) promoter, a Moloney murine leukemia virus (MoMLV) promoter, a Rous sarcoma virus (RSV) promoter, a herpes simplex virus (HSV) (thymidine kinase) promoter, an SV40 / CD43 promoter, a spleen focus forming vims (SFFV) promoter, an elongation factor 1-alpha (EFla) short promoter (intronless), an EFla long promoter containing an intron, a Ubiquitin C (UBC) promoter, a phosphoglycerate kinase- 1 (PGK) promoter, a cytomegalovirus enhancer / chicken P-actin (CAG) promoter, and a myeloproliferative sarcoma virus enhancer, negative control region deleted, dl587rev primer-binding site substituted (MND) U3 promoter (Haas et al., Journal of Virology. 2003;77(17): 9439-9450).

[0443] In particular embodiments, a polynucleotide comprises one or more cell type- or tissue-specific expression control sequences. In particular embodiments a cell type-specific expression control sequence is specific for immune effector cells. In particular embodiments a cell type- specific expression control sequence is a T cell specific promoter, an NK cell specific promoter, an NKT cell specific promoter, or a mucosal-associated invariant T (MAIT) cell promoter.

[0444] In particular embodiments, a cell type-specific expression control sequence is selected from the group consisting of a distal lymphocyte protein tyrosine kinase (LCK) promoter (Brenner et al., Proc. Natl. Acad. Sci. USA 99:2936-2941 (2002)), a CD35 promoter (Ji et al., J Biol Chem. 277(49):47898-906 (2002)), a CD4 gene promoter (Salmon et al., Proc. Natl. Acad. Sci. USA 90:7739 (1993), a CD2 promoter (Greaves et al., Cell 56:979-86 (1989)), and a TCF7 promoter (van de Wetering et al. J. of Bio. Chem. 267: 8530- 8536 (1992)).

[0445] Efficient expression of polynucleotides can be increased in some embodiments, by using sequences that increase translational efficiency, e.g., through an increase in mRNA ribosomal binding or an increase in mRNA stability. In certain embodiments, polynucleotides encoding a polypeptide of the invention comprise a short recognition sequence, i.e., a Kozak sequence, that greatly facilitates the initial binding of mRNA to the small subunit of the ribosome and increases translation. The consensus Kozak sequence is (GCC)RCCATGG, where R is a purine (A or G) (Kozak, Cell. 44:283-92 (1986), and Kozak, Nucleic Acids Res. 15:8125-48 (1987)).

[0446] Elements directing the efficient termination and polyadenylation of heterologous nucleic acid transcripts increases heterologous gene expression. Transcription termination signals are generally found downstream of the polyadenylation signal. In particular embodiments, vectors comprise a poly adenylation sequence 3' to a sequence to be transcribed and / or expressed. The term “polyadenylation (or poly(A)) signal” refers to a DNA sequence which directs both the termination and polyadenylation of the nascent RNA transcript by RNA polymerase II. Polyadenylation signals can promote mRNA stability by addition of a poly(A) tail to the 3' end of the coding sequence and thus, contribute to increased translational efficiency. Cleavage and poly adenylation are directed by a poly (A) signal in the RNA. The core poly(A) signal for mammalian pre-mRNAs has two recognition elements flanking a cleavage-polyadenylation site. Typically, an almost invariant AAUAAA hexamer lies 20-50 nucleotides upstream of a more variable element rich in U or GU residues. Cleavage of the nascent transcript occurs between these two elements and is coupled to the addition of up to 250 adenosines to the 5' cleavage product. In particular embodiments, the core poly(A) signal is an ideal poly(A) signal (e.g., AATAAA, ATTAAA, AGTAAA). In particular embodiments, the poly (A) signal is an SV40 poly (A) signal, a bovine growth hormone poly(A) signal (BGHpA), a rabbit P-globin poly(A) signal (rPgpA), variants thereof, or another suitable heterologous or endogenous poly(A) signal known in the art. In particular embodiments, the poly (A) signal is synthetic.

[0447] In particular embodiments, a polynucleotide, e.g., a vector, a viral vector, a retroviral vector, or lentiviral vector, comprises one or more off-target (e.g., brain, heart, liver, lung) miR target sequences inserted into a 5' UTR, intron, and / or 3' UTR.

[0448] In some embodiments, a polynucleotide comprises an inducible suicide gene to reduce the risk of direct toxicity and / or uncontrolled proliferation. In some embodiment, the suicide gene is caspase-8 or caspase-9. Caspase-9 can be activated using a specific chemical inducer of dimerization (CID).

[0449] In some embodiments, a polynucleotide comprises a gene or gene segment that when introduced into a cell, renders the cell susceptible to negative selection. Negative selection suitable for use in particular embodiments include but are not limited to the HSV-TK gene which confers ganciclovir sensitivity; the cellular hypoxanthine phosphribosyltransferase (HPRT) gene, the cellular adenine phosphoribosyltransferase (APRT) gene, and bacterial cytosine deaminase.

[0450] In some embodiments, a polynucleotide comprises a gene or gene segment that when introduced into a cell, renders the cell susceptible to positive selection. Positive selection genes suitable for use in particular embodiments contemplated herein include but are not limited to hygromycin-B phosphotransferase gene (hph) which confers resistance to hygromycin B, the amino glycoside phosphotransferase gene (neo or aph) from Tn5 which codes for resistance to the antibiotic G418, the dihydrofolate reductase (DHFR) gene, the adenosine deaminase gene (ADA), and the multi-drug resistance (MDR) gene.

[0451] In particular embodiments, a polynucleotide is a vector comprising or encoding a promoter operably linked to a polynucleotide encoding one or more engineered receptors, e.g., CAR, CCR, aP TCR, y5 TCR, DARIC, and / or CTBR, or components thereof. In particular embodiments, a polynucleotide is a retroviral or lentiviral vector comprising or encoding a promoter active in immune effector cells operably linked to a polynucleotide encoding one or more engineered receptors, e.g., CAR, CCR, aP TCR, y5 TCR, DARIC, and / or CTBR, or components thereof.

[0452] In particular embodiments, a polynucleotide is a retroviral or lentiviral vector comprising or encoding a promoter active in immune effector cells operably linked to a polynucleotide encoding an engineered receptor comprising an extracellular antigen binding domain that binds BCMA, CD19, CD20, CD22, CD38, CD79A, CD79B, GPCR5D, MAGE- A4, MSLN, MUC16, NY-ESO-1, PSCA, PSMA and R0R1.

[0453] In particular embodiments, a polynucleotide is a retroviral or lentiviral vector comprising or encoding a promoter selected from the group consisting of a CMV promoter, an SFFV promoter, an EFla short promoter, a UBC promoter, a PGK promoter, a CAG promoter, and an MNDU3 promoter operably linked to a polynucleotide encoding an engineered receptor comprising an extracellular antigen binding domain that binds BCMA, CD19, CD20, CD22, CD38, CD79A, CD79B, GPCR5D, MAGE-A4, MSLN, MUC16, NY- ESO-1, PSCA, PSMA and R0R1.

[0454] In particular embodiments, a polynucleotide is a retroviral or lentiviral vector comprising or encoding an SFFV promoter, operably linked to a polynucleotide encoding an engineered receptor comprising an extracellular antigen binding domain that binds BCMA, CD19, CD20, CD22, CD38, CD79A, CD79B, GPCR5D, MAGE-A4, MSLN, MUC16, NY- ESO-1, PSCA, PSMA and R0R1.

[0455] In particular embodiments, a polynucleotide is a retroviral or lentiviral vector comprising or encoding an EFla short promoter operably linked to a polynucleotide encoding an engineered receptor comprising an extracellular antigen binding domain that binds BCMA, CD19, CD20, CD22, CD38, CD79A, CD79B, GPCR5D, MAGE-A4, MSLN, MUC16, NY-ESO-1, PSCA, PSMA and R0R1.

[0456] In particular embodiments, a polynucleotide is a retroviral or lentiviral vector comprising or encoding a UBC promoter operably linked to a polynucleotide encoding an engineered receptor comprising an extracellular antigen binding domain that binds BCMA, CD19, CD20, CD22, CD38, CD79A, CD79B, GPCR5D, MAGE-A4, MSLN, MUC16, NY- ESO-1, PSCA, PSMA and R0R1.

[0457] In particular embodiments, a polynucleotide is a retroviral or lentiviral vector comprising or encoding a CAG promoter operably linked to a polynucleotide encoding an engineered receptor comprising an extracellular antigen binding domain that binds BCMA, CD19, CD20, CD22, CD38, CD79A, CD79B, GPCR5D, MAGE-A4, MSLN, MUC16, NY- ESO-1, PSCA, PSMA and ROR1.

[0458] In particular embodiments, a polynucleotide is a retroviral or lentiviral vector comprising or encoding an MND promoter operably linked to a polynucleotide encoding an engineered receptor comprising an extracellular antigen binding domain that binds BCMA, CD19, CD20, CD22, CD38, CD79A, CD79B, GPCR5D, MAGE-A4, MSLN, MUC16, NY- ESO-1, PSCA, PSMA and R0R1.

[0459] In particular embodiments, a polynucleotide is a retroviral or lentiviral vector comprising or encoding a promoter comprising a polynucleotide sequence set forth in any one of SEQ ID NOs: 667-672 operably linked to a polynucleotide encoding an engineered receptor comprising an amino acid sequence set forth in any one of SEQ ID NOs: 662-666.

[0460] Table 13: Exemplary Promoters

[0461] G. POLYPEPTIDES

[0462] Polypeptides, fusion polypeptides, and polypeptide variants are contemplated herein. Exemplary polypeptides contemplated herein include but not limited to, fusion polypeptides, viral envelope glycoproteins, non-viral membrane-bound tropism polypeptides, CARs, CCRs, TCRs, DARICs, CTBRs, nucleases, and therapeutic polypeptides, and variants and / or fragments thereof, e.g., SEQ ID NOs: 1-666 and 673-714.

[0463] Polypeptide,” “polypeptide,” “peptide,” and “protein” are used interchangeably, unless specified to the contrary, and according to conventional meaning, i.e., as a sequence of amino acids. In particular embodiments, a “polypeptide” is a fusion polypeptide or polypeptide variant. Polypeptides can be prepared using any of a variety of well-known recombinant and / or synthetic techniques. Polypeptides are not limited to a specific length, e.g., they may comprise a full-length protein sequence, a fragment of a full-length protein, or a fusion protein, and may include post-translational modifications, e.g., glycosylations, acetylations, phosphorylations and the like, as well as other modifications known in the art, both naturally occurring and non-naturally occurring.

[0464] An “isolated peptide,” “isolated protein” or an “isolated polypeptide” as used herein, refers to isolation, separation, and / or purification of a polypeptide molecule from a cellular environment, and from association with other components of the cell, i.e., it is not significantly associated with in vivo substances.

[0465] Polypeptides include “polypeptide variants.” In particular embodiments, a polypeptide variant is referred to as a “modified polypeptide.” Polypeptide variants may differ from a naturally occurring polypeptide in one or more amino acid substitutions, deletions, additions and / or insertions. For example, in particular embodiments, it may be desirable to modulate one or more biological activities of a viral envelope glycoprotein, non- viral membrane-bound tropism polypeptide, or engineered receptor by introducing one or more amino acid substitutions, deletions, additions and / or insertions into the polypeptide. Such variants may be naturally occurring or may be synthetically generated. In particular embodiments, polypeptides include polypeptide variants having at least about 65%, 66%, 67%, 68%, 69%, 70%, 71%, 72%, 73%, 74%, 75%, 76%, 77%, 78%, 79%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 86%, 97%, 98%, or 99% amino acid identity to any reference sequence contemplated herein, typically where the variant maintains at least one biological activity of the reference sequence.

[0466] In particular embodiments, a polypeptide variant is a viral envelope glycoprotein that has been modified to preserve its fusogenic activity and remove, disable, decrease, eliminate, or abate its binding activity to its cognate receptor on a cell. In particular embodiments, a polypeptide variant is a viral envelope glycoprotein that has at least about 65%, 66%, 67%, 68%, 69%, 70%, 71%, 72%, 73%, 74%, 75%, 76%, 77%, 78%, 79%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 86%, 97%, 98%, or 99% amino acid identity to any one of SEQ ID NOs: 582-645 or 646-661, wherein the variant comprises fusogenic activity and but is unable to bind or substantially bind to its cognate receptor on a cell.

[0467] Polypeptides variants include biologically active “polypeptide fragments.”

[0468] Illustrative examples of biologically active polypeptide fragments include but are not limited to binding domains, hinges, transmembrane domains, intracellular domains, and the like. As used herein, the term “biologically active fragment” or “minimal biologically active fragment” refers to a polypeptide fragment that retains at least 100%, at least 95%, at least 90%, at least 85%, at least 80%, at least 75%, at least 70%, at least 65%, at least 60%, at least 55%, or at least 50% of the naturally occurring polypeptide activity. In particular embodiments, a biologically active fragment is a polypeptide comprising an N-terminal and / or C-terminal truncation of 1, 2, 3, 4, 5, 6, 7, 8, 9, 10 or more amino acids. In particular embodiments, a mutated viral envelope glycoprotein or non-viral membrane-bound tropism polypeptide comprises a truncation of the cytoplasmic domain that results in a cytoplasmic tail or stub comprising, consisting essentially of, or consisting of 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acids.

[0469] In particular embodiments, polypeptides contemplated herein may comprise one or more amino acids denoted as “X” or “Xn” wherein n is an integer that denotes the particular X amino acid. “X” if present in an amino acid SEQ ID NO, refers to any one or more amino acids or particular amino acids if disclosed.

[0470] As noted above, polypeptides may be altered in various ways including amino acid substitutions, deletions, truncations, and insertions. Methods for such manipulations are generally known in the art. For example, amino acid sequence variants of a reference polypeptide can be prepared by mutations in the DNA. Methods for mutagenesis and nucleotide sequence alterations are well known in the art. See, for example, Kunkel (Proc. Natl. Acad. Sci. USA. 82: 488-492. (1985)), Kunkel et al., (Methods in Enzymol, 154: 367- 382. (1987)), U.S. Pat. No. 4,873,192, Watson, J. D. et al., (Molecular Biology of the Gene , Fourth Edition, Benjamin / Cummings, Menlo Park, Calif. (1987)) and the references cited therein. Guidance as to appropriate amino acid substitutions that do not affect biological activity of the protein of interest may be found in the model of Dayhoff et al. , Atlas of Protein Sequence and Structure (Natl. Biomed. Res. Found., Washington, D.C. (1978)).

[0471] In certain embodiments, a polypeptide variant comprises one or more conservative substitutions or disruptive substitutions. A “conservative substitution” is one in which an amino acid is substituted for another amino acid that has similar properties. In particular embodiments, polypeptide variants contemplated herein comprise one or more conservative amino acid changes compared to a reference polypeptide. In particular embodiments, a conservative amino acid substitution involves substituting an amino acid with an amino acid having a related side chain. A “disruptive substitution” is one in which an amino acid is substituted for another amino acid that has different properties, e.g., polar vs. non-polar, bulky vs. non-bulky, charged vs. uncharged, acidic vs. basic. In particular embodiments, polypeptide variants contemplated herein comprise one or more disruptive amino acid changes compared to a reference polypeptide. In particular embodiments, a disruptive amino acid substitution involves substituting an amino acid with an amino acid having an unrelated side chain or side change with a different chemical property. Guidance in determining which amino acid residues can be substituted, inserted, or deleted can be found using computer programs well known in the art, such as DNASTAR, DNA Strider, Geneious, Mac Vector, or Vector NTI software.

[0472] Naturally occurring amino acids are generally divided into four groups or families: acidic (aspartate, glutamate), basic (lysine, arginine, histidine), non-polar (alanine, valine, leucine, isoleucine, proline, phenylalanine, methionine, tryptophan), and uncharged polar (glycine, asparagine, glutamine, cysteine, serine, threonine, tyrosine) amino acids. Phenylalanine, tryptophan, and tyrosine are sometimes classified jointly as aromatic amino acids. In particular embodiments, a conservative amino acid substitution refers to substituting amino acids within the same group or family.

[0473] Those of skill in this art recognize that, in general, conservative single amino acid substitutions in non-essential regions of a polypeptide do not substantially alter biological activity (see, e.g., Watson el al. Molecular Biology of the Gene, 4th Edition, 1987, The Benjamin / Cummings Pub. Co., p.224), whereas disruptive single amino acid substitutions may.

[0474] In particular embodiments, a conservative amino acid substitution refers to substituting amino acids having a similar hydropathic index or score. The importance of the hydropathic amino acid index in conferring interactive biologic function on a protein is generally understood in the art (Kyte and Doolittle, 1982, incorporated herein by reference). Each amino acid has been assigned a hydropathic index on the basis of its hydrophobicity and charge characteristics (Kyte and Doolittle, 1982). These values are: isoleucine (+4.5); valine (+4.2); leucine (+3.8); phenylalanine (+2.8); cysteine / cysteine (+2.5); methionine (+1.9); alanine (+1.8); glycine (-0.4); threonine (-0.7); serine (-0.8); tryptophan (-0.9); tyrosine (-1.3); proline (-1.6); histidine (-3.2); glutamate (-3.5); glutamine (-3.5); aspartate (- 3.5); asparagine (-3.5); lysine (-3.9); and arginine (-4.5). In particular embodiments, a conservative amino acid substitution refers to substituting amino acids having a similar hydropathic index or score. In particular embodiments, substitution of amino acids whose hydropathic indices are within +2 is preferred, those within +1 are particularly preferred, and those within ±0.5 are even more particularly preferred. It is also understood in the art that the substitution of like amino acids can be made effectively on the basis of hydrophilicity.

[0475] In particular embodiments, a conservative amino acid substitution refers to substituting amino acids having a similar hydrophilic index or score. As detailed in U.S. Patent No. 4,554,101, the following hydrophilicity values have been assigned to amino acid residues: arginine (+3.0); lysine (+3.0); aspartate (+3.0 + 1); glutamate (+3.0 + 1); serine (+0.3); asparagine (+0.2); glutamine (+0.2); glycine (0); threonine (-0.4); proline (-0.5 + 1); alanine (-0.5); histidine (-0.5); cysteine (-1.0); methionine (-1.3); valine (-1.5); leucine (-1.8); isoleucine (-1.8); tyrosine (-2.3); phenylalanine (-2.5); tryptophan (-3.4). In particular embodiments, a conservative amino acid substitution refers to substituting amino acids having a similar hydrophilic index or score. In particular embodiments, substitution of amino acids whose hydrophilic indices are substitution of amino acids whose hydrophilicity values are within +2 is preferred, those within +1 are particularly preferred, and those within +0.5 are even more particularly preferred.

[0476] In particular embodiments, a conservative amino acid substitution may be based on the relative similarity of the amino acid side-chain substituents, for example, their hydrophobicity, hydrophilicity, charge, size, and the like. In particular embodiments, a disruptive amino acid substitution may be based on the relative dissimilarity of the amino acid side-chain substituents, for example, their hydrophobicity, hydrophilicity, charge, size, and the like.

[0477] In particular embodiments, a vector comprises one or more polynucleotides encoding two or more polypeptides. In particular embodiments, a polynucleotide encoding a polypeptide cleavage signal is disposed between the polynucleotide sequences encoding the two or more polypeptides to enable efficient translation and subsequent cleavage of a fusion polypeptide.

[0478] Polypeptides contemplated in particular embodiments include fusion polypeptides. In particular embodiments, fusion polypeptides and polynucleotides encoding fusion polypeptides are provided. Fusion polypeptides can include one or more polypeptide domains or segments including signal peptides, cell permeable peptide domains (CPP), binding domains, signaling domains, etc., epitope tags (e.g., maltose binding protein (“MBP”), glutathione S transferase (GST), HIS6, MYC, FLAG, V5, VSV-G, and HA), polypeptide linkers, and polypeptide cleavage signals. Fusion proteins and polypeptides are typically linked C-terminus to N-terminus, although they can also be linked C-terminus to C- terminus, N-terminus to N-terminus, or N-terminus to C-terminus. Fusion polypeptides and fusion proteins refer to a polypeptide having at least two, three, four, five, six, seven, eight, nine, or ten polypeptide segments.

[0479] In particular embodiments, a fusion polypeptide, viral envelope glycoprotein, non- viral membrane-bound tropism polypeptide, CAR, CCR, aP TCR, y5 TCR, DARIC, CTBR, nuclease, or therapeutic polypeptide comprises signal peptide set forth in any one of SEQ ID NOs: 449-454 that is subsequently cleaved from the fusion polypeptide, viral envelope glycoprotein, non-viral membrane-bound tropism polypeptide, CAR, CCR, aP TCR, y5 TCR, DARIC, CTBR, nuclease, or therapeutic polypeptide.

[0480] Table 14: Exemplary Signal Peptides

[0481] Fusion polypeptides may optionally comprise a polypeptide linker contemplated elsewhere herein that can be used to link one or more polypeptides or domains within a polypeptide. “Linker,” “peptide linker,” and “polypeptide linker” are used interchangeably and refer to a plurality of amino acid residues between various polypeptide domains added for appropriate spacing, conformation, and function. A polypeptide linker sequence may be employed to separate any two or more polypeptide components by a distance sufficient to ensure that each polypeptide folds into its appropriate secondary and tertiary structures so as to allow the polypeptide domains to exert their desired functions. Linkers include a “variable domain linking sequence,” an amino acid sequence that connects two or more domains of an antibody or antigen binding fragments thereof and provides a spacer function compatible with interaction of the two sub-binding domains so that the resulting polypeptide retains a specific binding affinity to the same target molecule as an antibody that comprises the same light and / or heavy chain variable domains. A linker may be 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, or 50 or more amino acids long. Illustrative examples of linkers include, but are not limited to the following amino acid sequences: AS, ASP, GGG; DGGGS (SEQ ID NO: 673); TGEKP (SEQ ID NO: 674); GGRR (SEQ ID NO: 675); (GGGGS)nwherein n = 1, 2, 3, 4 or 5 (SEQ ID NO: 676-680); S(GGGGS)nwherein n = 1, 2, 3, 4 or 5 (SEQ ID NO: 681-685); EGKSSGSGSESKVD (SEQ ID NO: 686); KESGSVSSEQLAQFRSLD (SEQ ID NO: 687); GGRRGGGS (SEQ ID NO: 688); LRQRDGERP (SEQ ID NO: 689); LRQKDGGGSERP (SEQ ID NO: 690); LRQKDGGGSGGGSERP (SEQ ID NO: 691), GEGTSTGSGGSGGSGGAD (SEQ ID NO: 692), and GSTSGSGKPGSGEGSTKG (SEQ ID NO: 693).

[0482] In particular embodiments, two or more polypeptides can be expressed as a fusion polypeptides that comprises one or more polypeptide cleavage signals disposed between the two or more polypeptides.

[0483] In particular embodiments, a fusion polypeptide comprises a CAR, a polypeptide cleavage signal and a CCR. In particular embodiments, a fusion polypeptide comprises a DARIC binding component, a polypeptide cleavage signal and a DARIC signaling component. In particular embodiments, a fusion polypeptide comprises a CAR, aP TCR, y5 TCR, or DARIC, a polypeptide cleavage signal, and a CTBR.

[0484] Exemplary polypeptide cleavage signals include, but are not limited to, protease cleavage sites, nuclease cleavage sites and ribosomal skipping polypeptide or self-cleaving viral polypeptides (see, e.g., in Ryan et al., 1997. J. Gener. Virol. 78, 699-722; deFelipe and Ryan, 2004. Traffic, 5(8); 616-26; and Scymczak et al. (2004) Nature Biotech. 5, 589-594).

[0485] Exemplary protease cleavage sites include, but are not limited to the cleavage sites of potyvirus NIa proteases (e.g, tobacco etch virus protease), poty virus HC proteases, potyvirus PI (P35) proteases, byovirus NIa proteases, byovirus RNA-2-encoded proteases, aphtho virus L proteases, enterovirus 2A proteases, rhino virus 2 A proteases, picoma 3C proteases, como virus 24K proteases, nepo virus 24K proteases, RTSV (rice tungro spherical virus) 3C-like protease, PYVF (parsnip yellow fleck virus) 3C-like protease, heparin, thrombin, factor Xa and enterokinase.

[0486] Illustrative examples of ribosomal skipping polypeptides include but are not limited to: a viral 2A peptide or sequence (Donnelly et al., 2001. J. Gen. Virol. 82: 1027-1041). In a particular embodiment, the viral 2A peptide is an aphthovirus 2A peptide, a potyvirus 2A peptide, or a cardiovirus 2A peptide.

[0487] In one embodiment, the viral 2A peptide is selected from the group consisting of: a foot-and-mouth disease vims (FMDV) 2A peptide, an equine rhinitis A vims (ERAV) 2A peptide, a Thosea asigna virus (TaV) 2A peptide, a porcine teschovirus-1 (PTV-1) 2A peptide, a Theilo virus 2A peptide, and an encephalomyocarditis virus 2A peptide.

[0488] Illustrative examples of viral 2A sequences include, but are not limited to: GSGATNFSLLKQAGDVEENPGP (SEQ ID NO: 694); ATNFSLLKQAGDVEENPGP (SEQ ID NO: 695); LLKQAGDVEENPGP (SEQ ID NO: 696); GSGEGRGSLLTCGDVEENPGP (SEQ ID NO: 697); EGRGSLLTCGDVEENPGP (SEQ ID NO: 698); LLTCGDVEENPGP (SEQ ID NO: 699); GSGQCTNYALLKLAGDVESNPGP (SEQ ID NO: 700);

[0489] QCTNYALLKLAGDVESNPGP (SEQ ID NO: 701); LLKLAGDVESNPGP (SEQ ID NO: 702); GSGVKQTLNFDLLKLAGDVESNPGP (SEQ ID NO: 703); VKQTLNFDLLKLAGDVESNPGP (SEQ ID NO: 704); LLNFDLLKLAGDVESNPGP (SEQ ID NO: 705); TLNFDLLKLAGDVESNPGP (SEQ ID NO: 706); NFDLLKLAGDVESNPGP (SEQ ID NO: 707); QLLNFDLLKLAGDVESNPGP (SEQ ID NO: 708); APVKQTLNFDLLKLAGDVESNPGP (SEQ ID NO: 709); VTELLYRMKRAETYCPRPLLAIHPTEARHKQKIVAPVKQT (SEQ ID NO: 710); LNFDLLKLAGDVESNPGP (SEQ ID NO: 711);

[0490] LLAIHPTEARHKQKIV APVKQTLNFDLLKLAGDVESNPGP (SEQ ID NO: 712); and EARHKQKIV APVKQTLNFDLLKLAGDVESNPGP (SEQ ID NO: 713).

[0491] H. CELLS

[0492] Recombinant particles contemplated herein are engineered to bind and transduce a cell. In particular embodiments, a recombinant particle contemplated herein is engineered to bind and transduce an immune effector cell. In particular embodiments, a cell transduced with a recombinant particle comprising a payload is modified to modulate the expression of one or more endogenous genes, and / or express one or more therapeutic polynucleotides and / or therapeutic polypeptides (e.g., an engineered receptor) contemplated herein.

[0493] In particular embodiments, a recombinant particle, e.g., a recombinant retroviral or lentiviral particle comprises a viral envelope comprising one or more mutated viral envelope glycoproteins that mediate virus-target cell fusion and one or more non-viral membranebound tropism polypeptides that bind an antigen expressed on a cell, e.g., an immune effector cell.

[0494] An “immune effector cell” is any cell of the immune system that has one or more effector functions (e.g., cytotoxic cell killing activity, secretion of cytokines, induction of ADCC and / or CDC). Illustrative types of immune effector cells contemplated in particular embodiments include, without limitation, T lymphocytes, dendritic cells (DC), Treg cells, natural killer (NK) cells, natural killer T (NKT) cells, and macrophages. The terms “T cell” or “T lymphocyte” are art-recognized and are intended, in particular embodiments, to include thymocytes, immature T lymphocytes, mature T lymphocytes, resting T lymphocytes, and / or activated T lymphocytes. Illustrative examples of T lymphocytes suitable for use in particular embodiments, include but not limited to cytotoxic T cells (CTLs; CD8+T cells), TILs, helper T cells (HTLs; CD4+T cells), CD4+CD8+T cells, CD4' CD8' T cells, or any other subset of T cells that has an effector function. In a particular embodiment, the cells comprise «P T cells. In a particular embodiment, the cells comprise y5 T cells.

[0495] In particular embodiments, a recombinant particle, e.g., a recombinant retroviral or lentiviral particle comprises a viral envelope comprising one or more mutated viral envelope glycoproteins that mediate virus-target cell fusion and one or more non-viral membranebound tropism polypeptides that bind an antigen on an immune effector cell, wherein the antigen is selected from the group consisting of: alpha, beta, gamma, or delta chain of the T cell receptor, CD2, CD35, CD3s CD3y, CD4, CD8a, CD8p, CD28, CD134 (0X40), CD137 (4-1BB), and CD278 (ICOS).

[0496] In particular embodiments, immune effector cells include natural killer (NK) cells. NK cells do not express T cell antigen receptors (TCR), CD3 or surface immunoglobulins (Ig) B cell receptor, but usually express the surface markers CD16 (FcyRIII) and CD56 in humans. In particular embodiments, a recombinant particle, e.g., a recombinant retroviral or lentiviral particle comprises a viral envelope comprising one or more mutated viral envelope glycoproteins that mediate virus-target cell fusion and one or more non-viral membranebound tropism polypeptides that bind an antigen on an immune effector cell, wherein the antigen is CD 16 (FcyRIII) and / or CD56.

[0497] In particular embodiments, immune effector cells include natural killer T (NKT) cells.

[0498] In particular embodiments, a progenitor of an immune effector cell is transduced with a recombinant particle contemplated herein and is subsequently induced to differentiate, or differentiates, into one or more immune effector cells. In particular embodiments, progenitors of immune effectors cells include hematopoietic stem cells (HSCs) contained within the CD34+population of cells derived from cord blood, bone marrow or mobilized peripheral blood which naturally differentiate into mature immune effector cells, or which can be induced to differentiate into mature immune effector cells. In particular embodiments, a recombinant particle, e.g., a recombinant retroviral or lentiviral particle comprises a viral envelope comprising one or more mutated viral envelope glycoproteins that mediate virustarget cell fusion and one or more non-viral membrane-bound tropism polypeptides that bind an antigen on a hematopoietic stem cell, wherein the antigen is selected from the group consisting of: CD7, CD33, CD34, CD45, CD49f, CD90, CD98, CD110, CD117, CD123, CD133, CD184, CD201, FMS-like tyrosine kinase 3 (FLT3), and thrombopoietin receptor.

[0499] I. COMPOSITIONS AND FORMULATIONS

[0500] Formulations and compositions contemplated herein comprise one or more polypeptides, polynucleotides, vectors, genome editing components, recombinant particles and / or immune effector cells modified ex vivo formulated in pharmaceutically acceptable or physiologically-acceptable compositions for administration to a cell, tissue, organ, or an animal, either alone, or in combination with one or more other modalities of therapy.

[0501] In particular embodiments, a composition comprises a recombinant particle, e.g., a recombinant retroviral or lentiviral particle comprises a viral envelope comprising one or more mutated viral envelope glycoproteins that mediate virus-target cell fusion and one or more non-viral membrane-bound tropism polypeptides, and a pay load comprising one or more polynucleotides and / or polypeptides.

[0502] In particular embodiments, a composition comprises a recombinant particle, e.g., a recombinant retroviral or lentiviral particle comprises a viral envelope comprising one or more mutated viral envelope glycoproteins that mediate virus-target cell fusion and one or more non-viral membrane-bound tropism polypeptides, and a pay load comprising a vector comprising a polynucleotide comprising or encoding a promoter operably linked to one or more polynucleotide encoding one or more engineered receptors.

[0503] In particular embodiments, a composition comprises a recombinant particle, e.g., a recombinant retroviral or lentiviral particle comprises a viral envelope comprising one or more mutated viral envelope glycoproteins that mediate virus-target cell fusion and one or more non-viral membrane-bound tropism polypeptides, and a pay load comprising a genome editing composition comprising one or more polynucleotides encoding a nuclease, a nuclease, or one or more polynucleotides encoding a DNA repair template and a nuclease. In particular embodiments, the composition is a pharmaceutical composition. A “pharmaceutical composition” refers to a composition formulated in a pharmaceutically - acceptable or physiologically-acceptable solution for administration to a cell or a subject, either alone, or in combination with one or more other modalities of therapy.

[0504] “Pharmaceutically acceptable” refers to molecular entities and compositions that do not produce excessive toxicity, irritation, allergic response, or other problems or complications, commensurate with a reasonable benefit / risk ratio when administered to a human.

[0505] In particular embodiments, a composition comprises a pharmaceutically acceptable carrier and a recombinant particle contemplated herein. The term “pharmaceutically acceptable carrier” refers to a diluent, adjuvant, excipient, vehicle and the like with which a recombinant particle, e.g., a recombinant retroviral or lentiviral particle, is physiologically compatible with administration to a human, including but not limited to pharmaceutically acceptable cell culture media, Dulbecco's phosphate buffered saline (PBS), Ringer's solution, 5% dextrose in water (D5W), and normal / physiologic saline (0.9% NaCl).

[0506] In particular embodiments, a composition comprises a recombinant particle and a pharmaceutically acceptable carrier suitable for enteral or parenteral, e.g., intravascular (intravenous or intraarterial), intraosseous, intraperitoneal, intraventricular, intracerebral, intracranial, intraspinal, intrathecal, intramuscular, and intramedullary, administration and formulation.

[0507] In particular embodiments, a composition is substantially free of mycoplasma, endotoxin, and microbial contamination. By “substantially free” with respect to endotoxin is meant that there is less endotoxin per dose of cells than is allowed by the FDA for a biologic, which is a total endotoxin of 5 EU / kg body weight per day, which for an average 70 kg person is 350 EU per total dose of cells. In particular embodiments, compositions contemplated herein contain about 0.5 EU / mL to about 5.0 EU / mL, or about 0.5 EU / mL, 1.0 EU / mL, 1.5 EU / mL, 2.0 EU / mL, 2.5 EU / mL, 3.0 EU / mL, 3.5 EU / mL, 4.0 EU / mL, 4.5 EU / mL, or 5.0 EU / mL.

[0508] In particular embodiments, compositions contemplated herein are used in the treatment of a cancer, GVHD, an infectious disease, an autoimmune disease, an inflammatory disease, or an immunodeficiency. In particular embodiments, a composition comprises a recombinant particle contemplated herein and one or more cytokines, growth factors, steroids, NSAIDs, DMARDs, anti-inflammatories, chemotherapeutics, radiotherapeutics, therapeutic antibodies, or other active and ancillary agents, either alone or in combination.

[0509] It would be understood by the skilled artisan that particular embodiments contemplated herein may comprise other formulations, such as those that are well known in the pharmaceutical art, and are described, for example, in Remington: The Science and Practice of Pharmacy, Volume I and Volume II. 23rdEdition. Edited by Adeboye Adejare. Academic Press, 2020, which is incorporated by reference herein, in its entirety.

[0510] J. METHODS OF MAKING

[0511] The manufacturing processes contemplated herein comprise an upstream process that produces a recombinant particle and a downstream process that purifies the recombinant particle. In preferred embodiments, a method of manufacturing a recombinant retroviral particle or recombinant lentiviral particle comprises an upstream process that produces the recombinant viral particle and a downstream process that purifies the recombinant viral particle. Methods of manufacturing lentiviral particles are described in W02023 / 003844, which is hereby incorporated by reference in its entirety. See, also Kutner et al., BMC Biotechnol. 2009;9:10. doi: 10.1186 / 1472-6750-9-10 and Kutner et al. Nat. Protoc. 2009;4(4):495-505. doi: 10.1038 / nprot.2009.22.

[0512] In particular embodiments, a method of manufacturing recombinant retroviral particles or recombinant lentiviral particles comprises transfecting a host cell culture with packaging plasmids and a transfer plasmid, culturing transfected host cells to produce viral particles; and collecting and processing the culture supernatant that contains the crude viral particles to remove impurities and concentrate and formulate the particles for clinical use.

[0513] In particular embodiments, retroviral particles are manufactured by transfecting host cells with a multi-plasmid system comprising (i) an envelope plasmid encoding one or more mutated viral envelope glycoproteins that retain fusogenic activity and substantially lack or lack cognate receptor binding activity and one or more non- viral membrane-bound tropism polypeptides, (ii) a packaging plasmid encoding gag-pol and (iii) a transfer plasmid or vector.

[0514] In particular embodiments, lentiviral particles are manufactured by transfecting host cells with a third-generation lentiviral vector plasmid system comprising (i) an envelope plasmid encoding one or more mutated viral envelope glycoproteins that retain fusogenic activity and substantially lack or lack cognate receptor binding activity and one or more non- viral membrane-bound tropism polypeptides, (ii) a packaging plasmid encoding gag-pol, (iii) a packaging plasmid encoding rev, and (iv) a transfer plasmid or vector.

[0515] A “transfer plasmid” or “transfer vector” encodes a viral genomic RNA modified to package a viral vector that comprises a polynucleotide sequence delivered by recombinant viral particle to a cell. In particular embodiments, a transfer plasmid comprises one or more polynucleotide sequences of interest flanked by long terminal repeat (LTR) sequences, which facilitate packaging, reverse transcription and integration of the viral vector and associated polynucleotide sequences into the host genome. Viral vectors contemplated herein are replication incompetent, i.e., lack the genetic elements necessary for generation of infective particles in the host cell. For example, the viral vector may be designed with a deletion of the 3' LTR, rendering the virus “self-inactivating” (SIN).

[0516] Illustrative examples of host cells suitable for transfection with the aforementioned plasmid systems include but are not limited to HEK293 cells, HEK293S cells, HEK293T cells, HEK293T cells adapted for suspension culture (HEK293Ts), HEK293F cells, HEK293FT cells, HEK293FTM cells, and HEK293E cells.

[0517] Once host cells are transfected and produce viral particles, the cell culture is subjected to a downstream process that yields particles sufficient for clinical use, In particular embodiments, a downstream process comprises treating the contents of the bioreactor with a DNA endonuclease, .e.g., benzonase; harvesting and clarifying the suspension culture supernatant by filtration; capturing and concentrating the viral particles in the resultant filtrate using affinity chromatography or cation exchange chromatography; filtering the eluate comprising the viral particles; ultrafiltering and diafiltering the viral particles using tangential flow filtration (TFF); and formulating the viral particles in a culture medium to produce a formulated bulk viral particles. In one embodiment, the formulated bulk lend viral particles are sterile filtered, filled, and frozen; and subsequently thawed, sterile filtered, subjected to a final fill finish, and frozen. In another embodiment, the bulk lentiviral particles are sterile filtered, subjected to a final fill finish, and frozen.

[0518] K. METHODS OF USE

[0519] Recombinant particles contemplated herein are engineered to modify a cell in vivo to express one or more engineered receptors that bind one or more target antigens and redirect the cell to a target cell expressing the target antigen, thereby preventing, treating, or ameliorating at least one symptom associated with, a cancer, GVHD, an infectious disease, an autoimmune disease, an inflammatory disease, or an immunodeficiency in a subject.

[0520] In particular embodiments, a method of preventing, treating, or ameliorating at least one symptom of a cancer, GVHD, an infectious disease, an autoimmune disease, an inflammatory disease, or an immunodeficiency comprises administering the subject an amount of recombinant particle contemplated herein. The term “amount” as used herein, refers to “an amount effective” or “an effective amount” of a recombinant particle contemplated herein comprising a pay load contemplated herein, etc., to achieve a beneficial or desired prophylactic or therapeutic result, including clinical results. A “prophylactically effective amount” refers to an amount of recombinant particle contemplated herein comprising payload contemplated herein, effective to achieve the desired prophylactic result. A “therapeutically effective amount” refers to an amount of recombinant particle contemplated herein comprising a paylaod contemplated herein, that is effective to “treat” a subject (e.g., a patient). When a therapeutic amount is indicated, the precise amount of the compositions to be administered can be determined by a physician with consideration of individual differences in age, weight, tumor size, extent of infection or metastasis, and condition of the patient (subject).

[0521] In particular embodiments, a recombinant retroviral or lentiviral particle is engineered to modify an immune effector cell in vivo to express one or more engineered receptors that bind one or more target antigens to redirect the immune effector cell to a target cell expressing the target antigens, thereby preventing, treating, or ameliorating at least one symptom associated with, a cancer, GVHD, an infectious disease, an autoimmune disease, an inflammatory disease, or an immunodeficiency in a subject.

[0522] In particular embodiments, a recombinant particle contemplated herein is administered to a subject to treat, prevent, or ameliorate at least one symptom of a solid tumor in the subject.

[0523] In particular embodiments, a recombinant particle contemplated herein is administered to a subject to treat, prevent, or ameliorate at least one symptom of a solid tumor selected from the group consisting of prostate cancer, lung cancer, non-small cell lung cancer (NSCLC), liver cancer, cervical cancer, colon cancer, breast cancer, ovarian cancer, endometrial cancer, pancreatic cancer, melanoma, esophageal cancer, gastric cancer, stomach cancer, renal carcinoma, bladder cancer, hepatocellular carcinoma, renal cell carcinoma, urothelial carcinoma, head and neck cancer, glioma, glioblastoma, colorectal cancer, thyroid cancer, epithelial cancers, or adenocarcinomas.

[0524] In particular embodiments, a recombinant particle contemplated herein is administered to a subject to treat, prevent, or ameliorate at least one symptom of a liquid or hematological cancer in the subject.

[0525] In particular embodiments, a recombinant particle contemplated herein is administered to a subject to treat, prevent, or ameliorate at least one symptom of a liquid or hematological cancer selected from the group consisting of leukemias, lymphomas, and multiple myeloma. In particular embodiments, a recombinant particle contemplated herein is administered to a subject to treat, prevent, or ameliorate at least one symptom of a relapsed and / or refractory leukemia, a relapsed and / or refractory lymphoma, or a relapsed and / or refractory multiple myeloma.

[0526] In particular embodiments, a recombinant particle contemplated herein is administered to a subject to treat, prevent, or ameliorate at least one symptom of a leukemia or precursor thereof selected from the group consisting of: an acute lymphocytic leukemia (ALL), an acute myeloid leukemia (AML), a myelodysplastic syndrome (MDS), a plasma cell leukemia (PCL), erythroleukemia, a hairy cell leukemia (HCL), chronic lymphocytic leukemia (CLL), chronic myeloid leukemia (CML), chronic myelomonocytic leukemia (CMML) and polycythemia vera.

[0527] In particular embodiments, a recombinant particle contemplated herein is administered to a subject to treat, prevent, or ameliorate at least one symptom of a lymphoma or precursor thereof selected from the group consisting of: a Hodgkin’ s lymphoma or a nonHodgkin’ s lymphoma, e.g., diffuse large B cell lymphoma (DLBCL), follicular lymphoma (FL), mantle-cell lymphoma (MCL), small lymphocytic lymphoma (SLL), primary mediastinal large B-cell lymphoma, a marginal zone B cell lymphoma (MZL), mucosa- associated lymphatic tissue lymphoma (MALT), Waldenstrom’s macroglobulinema, Burkitt’s lymphoma (BL), immunoblastic large cell lymphoma, centroblastic large cell lymphoma, anaplastic B-cell lymphoma, mycosis fungoides, Sezary syndrome, T- lymphoblastic lymphom...

Claims

CLAIMSWhat is claimed is:

1. A recombinant particle comprising:(a) a surface comprising (i) one or more mutated viral envelope glycoproteins that retain fusogenic activity and that lack cognate receptor binding activity and (ii) a non-viral membrane-bound tropism polypeptide comprising an antibody or antigen binding fragment thereof that binds an antigen expressed on an immune effector cell, a spacer polypeptide, a transmembrane domain, and optionally, an intracellular domain comprising no more than 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 amino acids; and(b) a therapeutic payload.

2. The particle of claim 1, wherein the particle is a fusosome; an extracellular vesicle, a microvesicle, an apoptotic body, or an exosome; a lipid nanoparticle; a virus-like particle (VLP); or a recombinant viral particle.

3. The particle of claim 1 or claim 2, wherein the surface comprises a single layer of phospholipid, a phospholipid bilayer, a cell membrane, a capsid, or a viral envelope.

4. The particle of any one of claims 1 to 3, wherein the payload comprises one or more polynucleotides and / or polypeptides.

5. The particle of any one of claims 1 to 4, wherein the pay load comprises a vector that encodes a therapeutic polynucleotide or polypeptide.

6. The particle of any one of claims 1 to 5, wherein the pay load comprises a vector that encodes one or more engineered receptors.

7. The particle of any one of claims 1 to 6, wherein the pay load comprises a vector that encodes a promoter operably linked to a polynucleotide encoding one or more engineered receptors.

8. A recombinant lentiviral particle comprising:(a) a viral envelope comprising (i) one or more mutated viral envelope glycoproteins that retain fusogenic activity and that lack cognate receptor binding activity and (ii) a non-viral membrane-bound tropism polypeptide comprising an antibody or antigen binding fragment thereof that binds an antigen expressed on an immune effector cell, a spacer polypeptide, a transmembrane domain, and optionally, an intracellular domain comprising no more than 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 amino acids; and(b) a recombinant lentiviral vector comprising a polynucleotide encoding a promoter operably linked to a polynucleotide encoding one or more engineered receptors.

9. The particle of any one of claims 1 to 8, wherein the one or more mutated viral envelope glycoproteins comprise a rhabdovirus envelope glycoprotein, or one or more paramyxovirus envelope glycoproteins.

10. The particle of any one of claims 1 to 9, wherein the one or more mutated viral envelope glycoproteins comprise a vesiculovirus envelope glycoprotein, one or more morbillivirus envelope glycoproteins or one or more henipavirus envelope glycoproteins.

11. The particle of claim 10, wherein the vesiculovirus is selected from the group consisting of: vesicular stomatitis Alagoas virus (VSAV; Alagoas vesiculovirus), Carajas virus (CJSV; Carajas vesiculovirus), Chandipura virus (CHPV; Chandipura vesiculovirus), Cocal virus (COCV; Cocal vesiculovirus), vesicular stomatitis Indiana virus (VSIV; Indiana vesiculovirus), Isfahan virus (ISFV; Isfahan vesiculovirus), Maraba virus (MARAV; Maraba vesiculovirus), Morreton virus (MORV; Morreton vesiculovirus), vesicular stomatitis New Jersey virus (VSNJV; New Jersey vesiculovirus), and Piry virus (PIRYV; Piry vesiculovirus).

12. The particle of claim 10 or claim 11, wherein the vesiculovirus envelope glycoprotein is a vesiculovirus G protein.

13. The particle of any one of claims 10 to 12, wherein the vesiculovirus G protein is a COCV G glycoprotein (COCV-G) or a VSIV G glycoprotein (VSIV-G).

14. The particle of any one of claims 10 to 13, wherein the VSIV-G envelope protein comprises one or more of:(a) one or more amino acid substitutions at H8, N9, Q10, K47, K50, A51, S183, S179, N180, 1182, M184, Y209, 1347, T350, T352, E353, and R354;(b) an insertion of TT between N9 and Q10, an insertion of GGS between H8 and N9, an insertion of GGS between N9 and Q10, an insertion of TT between N208 and Y209, an insertion of GGS between P46 and K47, and an insertion of GGS between N208 and Y209; or(c) amino acid substitutions at K47 and / or R354; or(d) a deletion of residues 1-8.

15. The particle of any one of claims 10 to 14, wherein the VSIV-G envelope protein comprises one or more amino acid substitutions at H8, K47, Y209, and R354.

16. The particle of any one of claims 10 to 15, wherein the VSIV-G envelope protein comprises amino acid substitutions selected from the group consisting of: K47A and R354A; K47A and R354G; K47A and R354F; K47A and R354Q; K47G and R354A; K47G and R354G; K47G and R354F; K47G and R354Q;K47F and R354A; K47F and R354G; K47F and R354F; K47F and R354Q; K47Q and R354A; K47Q and R354G; K47Q and R354F; and K47Q and R354Q.

17. The particle of any one of claims 10 to 16, wherein the VSIV-G envelope protein comprises the amino acid sequence set forth in SEQ ID NO: 2 or an amino acid sequence at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical thereto, wherein Xi= I, X2= A, X3= Q, and X4= A; Xi= I, X2= A, X3= Q, and X4= G; Xi= I, X2= A, X3= Q, and X4= F; Xi= I, X2= A, X3= Q, and X4= Q; Xi= L, X2= A, X3= Q, and X4= A; Xi= L, X2= A, X3= Q, and X4= G; Xi= L, X2= A, X3= Q, and X4= F; Xi= L, X2= A, X3= Q, and X4= Q; Xi= I, X2= A, X3= H, and X4= A; Xi= I, X2= A, X3= H, and X4= G; Xi= I, X2= A, X3= H, and X4= F; Xi= I, X2= A, X3= H, and X4= Q; Xi= L, X2= A, X3= H, and X4= A; Xi= L, X2= A, X3= H, and X4= G; Xi= L, X2= A, X3= H, and X4= F; Xi= L, X2= A, X3= H, and X4= Q; Xi= I, X2= G, X3= Q, and X4= A; Xi= I, X2= G, X3= Q, and X4= G; Xi= I, X2= G, X3= Q, and X4= F; Xi= I, X2= G, X3= Q, and X4= Q; Xi= L, X2= G, X3= Q, and X4= A; Xi= L, X2= G, X3= Q, and X4= G; Xi= L, X2= G, X3= Q, and X4= F; Xi= L, X2= G, X3= Q, and X4= Q; Xi= I, X2= G, X3= H, and X4= A; Xi= I, X2= G, X3= H, and X4= G; Xi= I, X2= G, X3= H, and X4= F; Xi= I, X2= G, X3= H, and X4= Q; Xi= L, X2= G, X3= H, and X4= A; Xi= L, X2= G, X3= H, and X4= G; Xi= L, X2= G, X3= H, and X4= F; Xi= L, X2= G, X3= H, and X4= Q; Xi= I, X2= F, X3= Q, and X4= A; Xi= I, X2= F, X3= Q, and X4= G; Xi= I, X2= F, X3= Q, and X4= F; Xi= I, X2= F, X3= Q, and X4= Q; Xi= L,X2= F, X3= Q, and X4= A; Xi= L, X2= F, X3= Q, and X4= G; Xi= L, X2= F, X3= Q, and X4= F; Xi= L, X2= F, X3= Q, and X4= Q; Xi= I, X2= F, X3= H, and X4= A; Xi= I, X2= F, X3= H, and X4= G; Xi= I, X2= F, X3= H, and X4= F; Xi= I, X2= F, X3= H, and X4= Q; Xi= L, X2= F, X3= H, and X4= A; Xi= L, X2= F, X3= H, and X4= G; Xi= L, X2= F, X3= H, and X4= F; Xi= L, X2= F, X3= H, and X4= Q; Xi= I, X2= Q, X3= Q, and X4= A; Xi= I, X2= Q, X3= Q, and X4= G; Xi= I, X2= Q, X3= Q, and X4= F; Xi= I, X2= Q, X3= Q, and X4= Q; Xi= L, X2= Q, X3= Q, and X4= A; Xi= L, X2= Q, X3= Q, and X4= G; Xi= L, X2= Q, X3= Q, and X4= F; Xi= L, X2= Q, X3= Q, and X4= Q; Xi= I, X2= Q, X3= H, and X4= A; Xi= I, X2= Q, X3= H, and X4= G; Xi= I, X2= Q, X3= H, and X4= F; Xi= I, X2= Q, X3= H, and X4= Q; Xi= L, X2= Q, X3= H, and X4= A; Xi= L, X2= Q, X3= H, and X4= G; Xi= L, X2= Q, X3= H, and X4= F; and Xi= L, X2= Q, X3= H, and X4= Q.

18. The particle of any one of claims 10 to 17, wherein the VSIV-G envelope protein comprises the amino acid sequence set forth in any one of SEQ ID NOs: 582-645 or an amino acid sequence at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical thereto.

19. The particle of any one of claims 10 to 18, wherein the VSIV-G envelope protein comprises the amino acid sequence set forth in any one of SEQ ID NOs: 630, 634, 638, and 642 or an amino acid sequence at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical thereto.

20. The particle of any one of claims 10 to 13, wherein the vesiculovirus G protein is COCV-G.

21. The recombinant lentivirus of claim 20, wherein the COCV-G envelope protein comprises one or more amino acid substitutions at K47 and / or R354.

22. The recombinant lentivirus of claim 20 or claim 21, wherein the COCV-G envelope protein comprises amino acid substitutions selected from the group consisting of: K47A and R354A; K47A and R354G; K47A and R354F; K47A and R354Q; K47G and R354A; K47G and R354G; K47G and R354F; K47G and R354Q;K47F and R354A; K47F and R354G; K47F and R354F; K47F and R354Q; K47Q and R354A; K47Q and R354G; K47Q and R354F; and K47Q and R354Q.

23. The recombinant lentivirus of any one of claims 20 to 22, wherein the COCV- G envelope protein comprises the amino acid sequence set forth in SEQ ID NO: 4 or an amino acid sequence at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical thereto, wherein Xi = A and X2 = A; Xi = A and X2 = G; Xi = A and X2 = F; Xi = A and X2 = Q; Xi = G and X2= A; Xi = G and X2= G; Xi = G and X2= F; Xi = G and X2= Q; Xi = F and X2= A; Xi = F and X2= G; Xi = F and X2= F; Xi = F and X2= Q; Xi = Q and X2= A; Xi = Q and X2= G; Xi = Q and X2= F; or Xi = Q and X2= Q.

24. The recombinant lentivirus of any one of claims 20 to 23, wherein the COCV- G envelope protein comprises the amino acid sequence set forth in any one of SEQ ID NOs: 646-661 or an amino acid sequence at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical thereto.

25. The particle of claim 10, wherein the one or more morbillivirus envelope glycoproteins are measles virus F (MV-F) and measles virus H (MV-H).

26. The particle of claim 25, wherein the MV-H protein comprises one or more amino acid substitutions at Y463, R515, S530, and F531.

27. The particle of claim 25 or claim 26, wherein the MV-H protein comprises one or more amino acid substitutions selected from the group consisting of: Y463A, R515A, S530, and F531.

28. The particle of any one of claims 25 to 27, wherein the MV-H protein comprises the amino acid sequence set forth in SEQ ID NO: 7.

29. The particle of claim 10, wherein the one or more henipavirus envelope glycoproteins are nipah virus F (NiV-F) and nipah virus G (NiV-G).

30. The particle of claim 29, wherein the NiV-G protein comprises one or more amino acid substitutions at E468, W471, Q497, and E500.

31. The particle of claim 29 or claim 30, wherein the NiV-G protein comprises one or more amino acid substitutions selected from the group consisting of: E468A, W471A, Q497A, and E500A.

32. The particle of any one of claims 29 to 31 , wherein the NiV-G protein comprises the amino acid sequence set forth in SEQ ID NO: 10.

33. The particle of any one of claims 1 to 32, wherein the antibody or antigen binding fragment thereof is selected from the group consisting of: a Camel Ig, a Llama Ig, an Alpaca Ig, Ig NAR, a Fab' fragment, a F(ab')2 fragment, a bispecific Fab dimer (Fab2), a trispecific Fab trimer (Fab3), an Fv, a single chain Fv protein (scFv), a bis-scFv, (scFv)2, a minibody, a diabody, a triabody, a tetrabody, a disulfide stabilized Fv protein (“dsFv”), a Fibronectin type III (FN3) domain antibody, a single-domain antibody (sdAb, a camelid VHH, Nanobody), and a centyrin.

34. The particle of any one of claims 1 to 33, wherein the antibody or antigen binding fragment thereof binds an antigen expressed on an immune effector cell selected from the group consisting of: the alpha, beta, gamma, or delta chain of the T cell receptor, CD2, CD35, CD3s CD3y, CD4, CD8a, and CD8p.

35. The particle of any one of claims 1 to 34, wherein the antibody or antigen binding fragment thereof comprises (a) a heavy chain variable region (VH) comprising a CDRH1, a CDRH2, and a CDRH3 of an antibody or antigen binding fragment thereof set forth in Table 5; (b) a polypeptide linker; and (c) a light chain variable region (VL) comprising a CDRL1, a CDRL2, and a CDRL3 of an antibody or antigen binding fragment thereof set forth in Table 5.

36. The particle of any one of claims 1 to 35, wherein the antibody or antigen binding fragment thereof comprises (a) a heavy chain variable region (VH) of an antibody or antigen binding fragment thereof set forth in Table 5; (b) a polypeptide linker; and (c) a light chain variable region (VL) of an antibody or antigen binding fragment thereof set forth in Table 5.

37. The particle of any one of claims 1 to 36, wherein the antibody or antigen binding fragment thereof comprises:(a) a heavy chain variable region (VH) of an antibody or antigen binding fragment thereof set forth in Table 5;(b) a polypeptide linker selected from the group consisting of: (GGGGS)nwherein n = 1, 2, 3, 4 or 5; S(GGGGS)nwherein n = 1, 2, 3, 4 or 5; GEGTSTGSGGSGGSGGAD, GSTSGSGKPGSGEGSTKG and variants thereof comprising an amino acid sequence 95% identical thereto; and(c) the corresponding light chain variable region (VL) of the antibody or antigen binding fragment thereof set forth in Table 5.

38. The particle of any one of claims 1 to 37, wherein the antibody or antigen binding fragment thereof comprises the amino acid sequence set forth in any one of SEQ ID NOs: 19, 20, 29, 30, 39, 40, 49, 50, 59, 60, 69, 70, 79, 80, 89, 90, 99, 100, 109, 110, 119, 120,129, 130, 139, 140, 149, 150, 159, 160, 169, 170, 179, 180, 189, 190, 199, 200, 209, 210, 219,220, 229, 230, 239, 240, 249, 250, 259, 260, 269, 270, 279, 280, 289, 290, 299, 300, 309, 310,319, 320, 329, 330, 339, 340, 349, 350, 359, 360, 369, 370, 379, 380, 389, 390, 399, 400, 409, and 410.

39. The particle of any one of claims 1 to 38, wherein the antibody or antigen binding fragment thereof comprises the amino acid sequence set forth in any one of SEQ ID NOs: 99, 100, 109, 110, 119, 120, 129, 130, 139, 140, 149, 150, 159, 160, 169, and 170.

40. The particle of any one of claims 1 to 39, wherein the antibody or antigen binding fragment thereof comprises the amino acid sequence set forth in any one of SEQ ID NOs: 99 or 100, 109 or 110, 119 or 120, 129 or 130, 139 or 140, 149 or 150, 159 or 160, and 169 or 170.

41. The particle of any one of claims 1 to 40, wherein the spacer comprises an amino acid sequence at least 85% identical to, at least 90% identical to, at least 95% identical to, at least 96% identical to, at least 97% identical to, at least 98% identical to, at least 99% identical to, or identical to the amino acid sequence set forth in any one of SEQ ID NOs: 411-434.

42. The particle of any one of claims 1 to 41, wherein the spacer comprises an amino acid sequence at least 85% identical to, at least 90% identical to, at least 95% identical to, at least 96% identical to, at least 97% identical to, at least 98% identical to, at least 99% identicalto, or identical to the amino acid sequence set forth in any one of SEQ ID NOs: 411-413, SEQ ID NOs: 414-423, SEQ ID NOs: 424-427, or SEQ ID NOs: 428-434.

43. The particle of any one of claims 1 to 42, wherein the transmembrane domain comprises an amino acid sequence at least 85% identical to, at least 90% identical to, at least 95% identical to, at least 96% identical to, at least 97% identical to, at least 98% identical to, at least 99% identical to, or identical to the amino acid sequence set forth in any one of SEQ ID NOs: 435, 436, 437, 438, 439, 440 or 441.

44. The particle of any one of claims 1 to 43, wherein the intracellular domain comprises an amino acid sequence at least 85% identical to, at least 90% identical to, at least 95% identical to, at least 96% identical to, at least 97% identical to, at least 98% identical to, at least 99% identical to, or identical to the amino acid sequence set forth in any one of SEQ ID NOs: 442, 443, 444, 445, 446, 447 or 448.

45. The particle of any one of claims 1 to 44, wherein the tropism polypeptide comprises an amino acid sequence at least 85% identical to, at least 90% identical to, at least 95% identical to, at least 96% identical to, at least 97% identical to, at least 98% identical to, at least 99% identical to, or identical to the amino acid sequence set forth in any one of SEQ ID NOs: 455-576.

46. The particle of any one of claims 1 to 45, wherein the tropism polypeptide comprises an amino acid sequence at least 85% identical to, at least 90% identical to, at least 95% identical to, at least 96% identical to, at least 97% identical to, at least 98% identical to, at least 99% identical to, or identical to the amino acid sequence set forth in any one of SEQ ID NOs: 471-486.

47. The particle of any one of claims 1 to 45, wherein the tropism polypeptide comprises an amino acid sequence at least 85% identical to, at least 90% identical to, at least 95% identical to, at least 96% identical to, at least 97% identical to, at least 98% identical to, at least 99% identical to, or identical to the amino acid sequence set forth in any one of SEQ ID NOs: 535-554.

48. The particle of any one of claims 1 to 45, wherein the tropism polypeptide comprises an amino acid sequence at least 85% identical to, at least 90% identical to, at least 95% identical to, at least 96% identical to, at least 97% identical to, at least 98% identical to, at least 99% identical to, or identical to the amino acid sequence set forth in any one of SEQ ID NOs: 555-562.

49. The particle of any one of claims 1 to 45, wherein the tropism polypeptide comprises an amino acid sequence at least 85% identical to, at least 90% identical to, at least 95% identical to, at least 96% identical to, at least 97% identical to, at least 98% identical to, at least 99% identical to, or identical to the amino acid sequence set forth in any one of SEQ ID NOs: 563 or 564.

50. The particle of any one of claims 1 to 49, further comprising one or more secondary non-viral membrane bound tropism polypeptides.

51. The particle of claim 50, wherein the one or more secondary non-viral membrane bound tropism polypeptides comprise one or more of CD80, CD86, CD137L, OX40L, and ICOSL or variants thereof.

52. The particle of any one of claims 1 to 51, further comprising one or more secondary non-viral membrane-bound tropism polypeptides at least 85% identical to, at least 90% identical to, at least 95% identical to, at least 96% identical to, at least 97% identical to, at least 98% identical to, at least 99% identical to, or identical to the amino acid sequence set forth in any one of SEQ ID NOs: 577-581.

53. The particle of any one of claims 8 to 52, wherein the recombinant lentiviral vector is engineered or derived from a lentiviral genome of a lentivirus selected from the group consisting of: human immunodeficiency virus 1 (HIV-1); human immunodeficiency virus 2 (HIV-2); visna-maedi virus (VMV); caprine arthritis-encephalitis virus (CAEV); equine infectious anemia virus (EIAV); feline immunodeficiency virus (FIV); bovine immune deficiency virus (BIV); and simian immunodeficiency virus (SIV).

54. The particle of any one of claims 8 to 53, wherein the recombinant lentiviral vector further comprises one or more polynucleotides encoding one or more of the following:a signal peptide, a post-transcriptional regulatory element, an insulator element, a selectable marker, and a cell suicide gene.

55. The particle of claim 54, wherein the post-transcriptional regulatory element is a woodchuck hepatitis virus post-transcriptional regulatory element (WPRE) or a hepatitis B virus post-transcriptional regulatory element (HPRE).

56. The particle of any one of claims 7 to 55, wherein the promoter is selected from the group consisting of: a P-actin promoter, a cytomegalovirus (CMV) immediate early promoter, a simian virus 40 (SV40) (e.g., early or late) promoter, a Moloney murine leukemia virus (MoMLV) promoter, a Rous sarcoma virus (RSV) promoter, a herpes simplex virus (HSV) (thymidine kinase) promoter, an SV40 / CD43 promoter, a spleen focus forming virus (SFFV) promoter, an elongation factor 1 -alpha (EFla) short promoter (intronless), an EFla long promoter containing an intron, a Ubiquitin C (UBC) promoter, a phosphoglycerate kinase- 1 (PGK) promoter, a cytomegalovirus enhancer / chicken P-actin (CAG) promoter, and a myeloproliferative sarcoma virus enhancer, negative control region deleted, dl587rev primerbinding site substituted (MND) U3 promoter.

57. The particle of any one of claims 6 to 56, wherein the one or more engineered receptors are selected from the group consisting of: a chimeric antigen receptor (CAR), a chimeric costimulatory receptor (CCR), an alpha-beta T cell receptor (aP TCR), a gamma delta T cell receptor (y5 TCR), a dimerizing agent regulated immunoreceptor complex (DARIC), a chimeric TGF-P receptor (CTBR), and a zetakine receptor.

58. The particle of any one of claims 6 to 57, wherein the one or more engineered receptors bind an antigen selected from the group consisting of: alpha folate receptor (FRa), avP6 integrin, BAFFR, 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), claudin 6, (CEDN6), claudin 18 isoform 2 (CEDN18.2), C- type lectin-like molecule- 1 (CEE-1), CD2 subset 1 (CS-1), chondroitin sulfate proteoglycan 4 (CSPG4), cutaneous T cell lymphoma-associated antigen 1 (CTAGE1), delta like canonical Notch ligand 3 (DLL3), epidermal growth factor receptor (EGFR), epidermal growth factor receptor variant III (EGFRvIII), epithelial glycoprotein 2 (EGP2), epithelial glycoprotein 40(EGP40), epithelial cell adhesion molecule (EPCAM), ephrin type-A receptor 2 (EPHA2), erb- b2 receptor tyrosine kinase 4 (ERBB4), fibroblast activation protein (FAP), Fc Receptor Like 5 (FCRL5), fetal acetylcholinesterase receptor (AchR), ganglioside G2 (GD2), ganglioside G3 (GD3), Glypican-3 (GPC3), G Protein-Coupled Receptor Class C Group 5 Member D (GPCR5D), EGFR family including ErbB2 (HER2), HER2 p95, IL-lORa, IL-13Ra2, Kappa, cancer / testis antigen 2 (LAGE- 1 A), Lambda, Lewis-Y (LeY), LI cell adhesion molecule (LI- CAM), melanoma antigen gene (MAGE)-Al, MAGE- A3, MAGE-A4, MAGE-A6, MAGEA10, melanoma antigen recognized by T cells 1 (MelanA or MARTI), Mesothelin (MSLN), MUC1, MUC16, MHC class I chain related proteins A (MICA), MHC class I chain related proteins B (MICB), neural cell adhesion molecule (NCAM), cancer / testis antigen 1 (NY-ESO-1), placenta- specific 1 (PLAC1), preferentially expressed antigen in melanoma (PRAME), prostate stem cell antigen (PSCA), pro state- specific membrane antigen (PSMA), receptor tyrosine kinase-like orphan receptor 1 (ROR1), SLAMF7, synovial sarcoma, X breakpoint 2 (SSX2), Survivin, TACI, tumor associated glycoprotein 72 (TAG72), tumor endothelial marker 1 (TEM1 / CD248), tumor endothelial marker 7-related (TEM7R), transforming growth factor beta 1 (TFGpi), 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).

59. The particle of any one of claims 6 to 58, wherein the one or more engineered receptors bind an antigen selected from the group consisting of: BCMA, CD19, CD20, CD22, CD33, CD38, CD79A, CD79B, GPCR5D, MAGE-A4, MSLN, MUC16, NY-ESO-1, PSCA, PSMA and ROR1.

60. The particle of any one of claims 6 to 58, wherein the one or more engineered receptors bind an antigen selected from the group consisting of: BCMA, CD19, CD20, CD22, CD38, CD79A, CD79B, and GPCR5D.

61. The particle of any one of claims 6 to 60, wherein the one or more engineered receptors bind an antigen selected from the group consisting of: BCMA, CD38, and GPCR5D.

62. The particle of any one of claims 6 to 60, wherein the one or more engineered receptors bind an antigen selected from the group consisting of: CD19, CD20, CD22, CD79A, and CD79B.

63. The particle of any one of claims 6 to 62, wherein the one or more engineered receptors comprises an extracellular antigen binding domain, a hinge domain, a transmembrane domain, and one or more intracellular signaling domains.

64. The particle of any one of claims 6 to 63, wherein the one or more engineered antigen receptors comprises an extracellular antigen binding domain selected from the group consisting of: a receptor ectodomain, a ligand, or an antibody or antigen binding fragment thereof selected from the group consisting of: a Camel Ig, a Llama Ig, an Alpaca Ig, Ig NAR, a Fab' fragment, a F(ab')2 fragment, a bispecific Fab dimer (Fab2), a trispecific Fab trimer (Fab3), an Fv, a single chain Fv protein (scFv), a bis-scFv, (scFv)2, a minibody, a diabody, a triabody, a tetrabody, a disulfide stabilized Fv protein (“dsFv”), a Fibronectin type III (FN3) domain antibody, and a single-domain antibody (sdAb, a camelid VHH, Nanobody).

65. The particle of any one of claims 57 to 64, wherein the CAR comprises a hinge domain isolated or derived from a polypeptide selected from the group consisting of: CD4, CD8P, CD8a, CD28, CD134, CD137, CD152, CD278, IgGl, IgG2, IgG3, and IgG4.

66. The particle of any one of claims 57 to 65, wherein the CAR comprises a transmembrane domain isolated or derived from a polypeptide selected from the group consisting of: alpha, beta, gamma, or delta chain of the T-cell receptor, CD35, CD3s, CD3y, CD3 , CD4, CD5, CD8a, CD9, CD16, CD22, CD27, CD28, CD33, CD37, CD45, CD64, CD80, CD86, CD134, CD137, CD152, CD154, CD278, amnionless (AMN), and programmed cell death 1 (PDCD1).

67. The particle of any one of claims 57 to 66, wherein the CAR comprises a primary signaling domain isolated or derived from a polypeptide selected from the group consisting of: FcRy, FcRp, CD3y, CD35, CD3s, CD3^, CD22, CD79A, CD79B, and CD66d.

68. The particle of any one of claims 57 to 67, wherein the CAR comprises one or more costimulatory domains isolated or derived from a polypeptide 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, CD7, CD27, CD28, CD30, CD40, CD54 (ICAM), CD83, CD94, CD134 (0X40), CD137 (4-1BB), CD278 (ICOS), DNAX-Activation Protein 10 (DAP10), Linker for activation of T-cells family member 1 (LAT), SH2 Domain-Containing Leukocyte Protein Of 76 kD (SLP76), T cellreceptor associated transmembrane adaptor 1 (TRAT1), TNFR2, TNFRS14, TNFRS18, TNFRS25, and zeta chain of T cell receptor associated protein kinase 70 (ZAP70).

69. The particle of any one of claims 6 to 68, wherein the one or more engineered receptors further comprises a CCR comprising an extracellular antigen binding domain that binds an antigen different than the antigen bound by the CAR, a hinge domain, a transmembrane domain, and one or more intracellular signaling domains.

70. The particle of any one of claims 6 to 57, wherein the one or more engineered receptors comprises a DARIC comprising a DARIC signaling component comprising a multimerization domain, a transmembrane domain and one or more intracellular signaling domains; and a DARIC binding component comprising an extracellular antigen binding domain, an extracellular antigen binding domain, a transmembrane domain, and optionally one or more intracellular signaling domains.

71. The particle of any one of claims 6 to 70, wherein the one or more engineered receptors comprises a CTBR comprising (a) a first polypeptide comprising an extracellular TGFpi -binding domain of TGFPR2, a transmembrane domain, and a first interleukin receptor intracellular signaling domain; (b) a polypeptide cleavage signal; and (c) a second polypeptide comprising an extracellular TGFpi -binding domain of TGFpRl, a transmembrane domain, and a second interleukin receptor intracellular signaling domain.

72. A recombinant lentiviral particle comprising: (a) a surface comprising (i) one or more mutated viral envelope glycoproteins that retain fusogenic activity and that lack cognate receptor binding activity, wherein the one or more mutated viral envelope glycoproteins comprises an amino acid sequence at least 95%, at least 96%, at least 97%, at least 98%, at least 99% identical to or identical to any one of SEQ ID NOs: 630, 634, 638, and 642 and (ii) a non-viral membrane-bound tropism polypeptide comprising an antibody or antigen binding fragment thereof comprising an amino acid sequence set forth in any one of SEQ ID NOs: 91 and 95, and 99 or 100, that binds an antigen expressed on an immune effector cell, a spacer polypeptide, and a GPI anchor domain or a transmembrane domain, and optionally, an intracellular domain comprising no more than 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 amino acids; and (b) a therapeutic pay load encoding an engineered antigen receptor.

73. A recombinant lentiviral particle comprising: (a) a surface comprising (i) a mutated VSIV-G envelope glycoprotein that comprises an amino acid sequence at least 95%, at least 96%, at least 97%, at least 98%, at least 99% identical to or identical to SEQ ID NO: 642 and (ii) a non-viral membrane-bound tropism polypeptide comprising an antibody or antigen binding fragment thereof comprising an amino acid sequence set forth in SEQ ID NO: 99 or 100, that binds an antigen expressed on an immune effector cell, a spacer polypeptide, and a GPI anchor domain or a transmembrane domain, and optionally, an intracellular domain comprising no more than 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 amino acids; and (b) a lentiviral vector encoding an engineered antigen receptor.

74. A recombinant lentiviral particle comprising: (a) a viral envelope comprising (i) a mutated VSIV-G envelope glycoprotein that comprises an amino acid sequence at least 95%, at least 96%, at least 97%, at least 98%, at least 99% identical to or identical to SEQ ID NO: 642 and (ii) a non-viral membrane-bound tropism polypeptide comprising an amino acid sequence set forth in any one of SEQ ID NOs: 99 or 100, that binds an antigen expressed on an immune effector cell, a spacer polypeptide, and a GPI anchor domain or a transmembrane domain, and optionally, an intracellular domain comprising no more than 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 amino acids; and (b) a therapeutic pay load encoding an engineered antigen receptor.

75. A recombinant lentiviral particle comprising: (a) a viral envelope comprising (i) a mutated VSIV-G envelope glycoprotein that comprises an amino acid sequence at least 95%, at least 96%, at least 97%, at least 98%, at least 99% identical to or identical to SEQ ID NO: 642; (ii) a non-viral membrane -bound tropism polypeptide comprising an amino acid sequence set forth in any one of SEQ ID NOs: 461, 471, 483, 537, and 553; and optionally (iii) a secondary non-viral membrane-bound tropism polypeptide comprising an amino acid sequence set forth in SEQ ID NO: 577; and (b) a lentiviral vector encoding a chimeric antigen receptor.

76. A recombinant lentiviral particle comprising: (a) a viral envelope comprising (i) a mutated VSIV-G envelope glycoprotein that comprises an amino acid sequence at least 95%, at least 96%, at least 97%, at least 98%, at least 99% identical to or identical to SEQ ID NO: 642; (ii) a non-viral membrane -bound tropism polypeptide that comprises an amino acid sequence at least 95%, at least 96%, at least 97%, at least 98%, at least 99% identical to or identical to SEQ ID NO: 471; and optionally (iii) a secondary non-viral membrane-boundtropism polypeptide that comprises an amino acid sequence at least 95%, at least 96%, at least 97 %, at least 98%, at least 99% identical to or identical to SEQ ID NO: 577 ; and (b) a lentiviral vector encoding a chimeric antigen receptor comprising an amino acid sequence set forth in any one of SEQ ID NOs: 662-666.

77. A recombinant lentiviral particle comprising: (a) a viral envelope comprising (i) a mutated VSIV-G envelope glycoprotein that comprises an amino acid sequence at least 95%, at least 96%, at least 97%, at least 98%, at least 99% identical to or identical to SEQ ID NO: 642; (ii) a non-viral membrane -bound tropism polypeptide that comprises an amino acid sequence at least 95%, at least 96%, at least 97%, at least 98%, at least 99% identical to or identical to SEQ ID NO: 471; and optionally (iii) a secondary non-viral membrane-bound tropism polypeptide that comprises an amino acid sequence at least 95%, at least 96%, at least 97 %, at least 98%, at least 99% identical to or identical to SEQ ID NO: 577 ; and (b) a lentiviral vector encoding a chimeric antigen receptor that binds a polypeptide selected from any one of BCMA, CD19, CD20, CD22, CD38, CD79A, CD79B, and GPCR5D.

78. A recombinant lentiviral particle comprising: (a) a viral envelope comprising (i) a mutated VSIV-G envelope glycoprotein that comprises an amino acid sequence at least 95%, at least 96%, at least 97%, at least 98%, at least 99% identical to or identical to SEQ ID NO: 642; (ii) a non-viral membrane -bound tropism polypeptide that comprises an amino acid sequence at least 95%, at least 96%, at least 97%, at least 98%, at least 99% identical to or identical to SEQ ID NO: 471; and optionally (iii) a secondary non-viral membrane-bound tropism polypeptide that comprises an amino acid sequence at least 95%, at least 96%, at least 97 %, at least 98%, at least 99% identical to or identical to SEQ ID NO: 577 ; and (b) a lentiviral vector encoding a chimeric antigen receptor that binds BCMA.

79. A recombinant lentiviral particle comprising: (a) a viral envelope comprising (i) a mutated VSIV-G envelope glycoprotein that comprises an amino acid sequence at least 95%, at least 96%, at least 97%, at least 98%, at least 99% identical to or identical to SEQ ID NO: 642; (ii) a non-viral membrane -bound tropism polypeptide that comprises an amino acid sequence at least 95%, at least 96%, at least 97%, at least 98%, at least 99% identical to or identical to SEQ ID NO: 471; and optionally (iii) a secondary non-viral membrane-bound tropism polypeptide that comprises an amino acid sequence at least 95%, at least 96%, at least97 %, at least 98%, at least 99% identical to or identical to SEQ ID NO: 577 ; and (b) a lentiviral vector encoding a chimeric antigen receptor that binds GPCR5D.

80. A recombinant lentiviral particle comprising: (a) a viral envelope comprising (i) a mutated VSIV-G envelope glycoprotein that comprises an amino acid sequence at least 95%, at least 96%, at least 97%, at least 98%, at least 99% identical to or identical to SEQ ID NO: 642; (ii) a non-viral membrane -bound tropism polypeptide that comprises an amino acid sequence at least 95%, at least 96%, at least 97%, at least 98%, at least 99% identical to or identical to SEQ ID NO: 471; and optionally (iii) a secondary non-viral membrane-bound tropism polypeptide that comprises an amino acid sequence at least 95%, at least 96%, at least 97 %, at least 98%, at least 99% identical to or identical to SEQ ID NO: 577 ; and (b) a lentiviral vector encoding a chimeric antigen receptor that binds CD 19.

81. A cell transduced with the particle of any one of claims 1 to 80.

82. The cell of claim 81, wherein the cell is an immune effector cell.

83. The cell of claim 81 or claim 82, wherein the cell is a T cell, a natural killer (NK) cell, or a natural killer T (NKT) cell.

84. A composition comprising the particle of any one of claims 1 to 80 or the cell of any one of claims 81 to 83.

85. A pharmaceutical composition comprising a pharmaceutically acceptable carrier and the particle of any one of claims 1 to 80, the cell of any one of claims 81 to 83, or the composition of claim 84.

86. A method of treating, preventing, or ameliorating at least one symptom of a disease, disorder or condition associated therewith in a subject, comprising administering to the subject an effective amount of the particle of any one of claims 1 to 80, the cell of any one of claims 81 to 83, the composition of claim 84, or the pharmaceutical composition of claim 85.

87. The method of claim 86, wherein the disease, disorder, or condition is a cancer.

88. The method of claim 86 or claim 87, wherein the cancer is leukemia selected from the group consisting of: acute lymphocytic leukemia (ALL), an acute myeloid leukemia (AML), a myelodysplastic syndrome (MDS), a plasma cell leukemia (PCL), erythroleukemia, a hairy cell leukemia (HCL), chronic lymphocytic leukemia (CLL), chronic myeloid leukemia (CML), chronic myelomonocytic leukemia (CMML) and polycythemia vera.

89. The method of claim 86 or claim 87, wherein the cancer is non-NHL or NHL selected from the group consisting of: diffuse large B cell lymphoma (DLBCL), follicular lymphoma (FL), mantle-cell lymphoma (MCL), small lymphocytic lymphoma (SLL), primary mediastinal large B-cell lymphoma, a marginal zone B cell lymphoma (MZL), mucosa- associated lymphatic tissue lymphoma (MALT), Burkitt’s lymphoma (BL), immunoblastic large cell lymphoma, centroblastic large cell lymphoma, anaplastic B-cell lymphoma, mycosis fungoides, Sezary syndrome, T-lymphoblastic lymphoma, and anaplastic large-cell lymphoma (ALCL).

90. The method of claim 86 or claim 87, wherein the cancer is MM selected from the group consisting of: active multiple myeloma, smoldering multiple myeloma, light chain myeloma, non-secretory myeloma, IgD myeloma, IgE myeloma, osteosclerotic myeloma, solitary plasmacytoma of bone, and extramedullary plasmacytoma.

91. The method of any one of claims 86 to 90, wherein the cancer is relapsed and / or refractory.

92. The method of claim 86, wherein the disease, disorder, or condition is an autoimmune disease.

93. The method of claim 92, wherein the autoimmune disease is systemic lupus erythematosus, rheumatoid arthritis, idiopathic thrombocytopenic purpura, myasthenia gravis, or autoimmune hemolytic anemia.

94. A method of transducing an immune effector cell in vivo, comprising administering to a subject a pharmaceutical composition comprising a pharmaceutically acceptable carrier and an effective amount of the particle of any one of claims 1 to 80.

95. A method of making the particle of any one of claims 1 to 80 comprising (a) transfecting a host cell with four polynucleotides: a first polynucleotide that encodes lentiviral gag-pol, a second polynucleotide that encodes lentiviral rev, a third polynucleotide that encodes the one or more mutated viral envelope glycoproteins that retain fusogenic activity and lack cognate receptor binding activity and the one or more non-viral membrane-bound tropism polypeptides, and a fourth polynucleotide that is a transfer plasmid encoding a recombinant lentiviral vector encoding an engineered antigen receptor; and b) culturing the transduced cell for about 1 to 3 days to produce the particle.

96. A kit comprising the particle of any one of claims 1 to 80, a pharmaceutically acceptable carrier, and instructions for use.

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