Compositions and methods for immunotherapies

Abstract

Disclosed herein is a composition comprising one or more polynucleic acid molecules comprising a sequence encoding an anti-CD20 CAR, a sequence encoding an anti-CD22 CAR, and a sequence encoding an anti-CD19 CAR. Disclosed herein is a composition comprising one or more polynucleic acid molecules comprising a sequence encoding an anti-CD19 binding domain. Disclosed herein is a composition comprising one or more polynucleic acid molecules comprising a sequence encoding an anti-CD20 binding domain. Further provided herein are pharmaceutical compositions comprising the polynucleic acid molecules, and methods for cancer treatment.

Description

COMPOSITIONS AND METHODS FOR IMMUNOTHERAPIESCROSS-REFERENCE TO RELATED APPLICATION

[0001] The present application claims priority to U.S. Provisional Patent Application Serial No. 63 / 606,343, filed on December 5, 2023, and 63 / 536,301 filed on September 1, 2023. The contents of the above-referenced applications are herein expressly incorporated by reference in their entirety, including any drawings.BACKGROUND

[0002] Autologous CAR-T cell (chimeric antigen receptor T cell) therapies have been used for the treatment of hematologic malignancies such as B-cell lymphoma and for patients who have relapsed after receiving chemotherapy. Despite advances made by commercially available CAR-T cell therapies, there remain key resistance mechanisms that can limit the strength and quality of T cell response and lead to disease progression, including loss or down-regulation of target antigen expression, loss of co- stimulation and limited CAR-T-cell persistence. Therefore, there remains an urgent medical need for improved CAR-T immunotherapies.

[0003] Autologous cell therapy with T cells expressing synthetic chimeric antigen receptors (CARs) targeting one of the B cell antigens CD 19, CD20, and CD22 has revolutionized treatment of B cell malignancies since the FDA first approved the sale of Kymriah®, a CD19-specific CAR-T cell therapy, in August 2017. Although autologous CAR-T cells have induced beneficial therapeutic responses in many of the patients receiving such treatment, as many as 60% of those treated relapse, and 10-20% of those who relapse after CD 19 CAR-T cell treatment experience CD 19-negative relapse. In addition to down-regulation or loss of target antigen expression, relapse can be caused by mutation of the epitope recognized by the antigen-binding domain of the CAR, poor CAR-T cell persistence, transient B cell aplasia, or down-regulation or elimination of co-stimulatory signaling pathways necessary for T cell activation, among other things. Thus, there remains a substantial unmet clinical need for improved autologous CAR-T cell therapies for the treatment of B cell cancers.

[0004] One approach to improving the efficacy of such therapies has been to develop multi-specific CAR-T cells capable of targeting two or three of the B cell antigens referenced above. For CAR-T cells targeting two B cell antigens, common strategies include the use of (1) two mono-specific CARs (e.g., with T cells engineered to express a CD19-specific CAR and a CD20-specific CAR, a CD19-specific CAR and a CD22-specific CAR, or a CD20-specific CAR and a CD22-specific CAR); or (2) one CAR comprising a bi-specific binding domain (e.g., with T cells engineered toexpress a bispecific antibody or two single chain variable fragments (scFvs) in tandem to target CD 19 and CD20, CD 19 and CD22, or CD20 and CD22), referred to as a “tandem CAR”. For CAR T-cells targeting all three B cell antigens, strategies being explored include the use of: (1) one tandem CAR and a single mono-specific CAR; (2) a single CAR expressing a tri-specific binding domain; or (3) three mono-specific CARs, but all three of these approaches present technical challenges.

[0005] A variety of factors influence the ability of a CAR to recognize its cognate antigen(s) on tumor cells, induce CAR signaling, and activate the T cell including, for example: (1) the CAR’s binding affinity for its target antigen; (2) steric interactions, either between binding domains on tandem or other multi-specific binding domains or between binding domains and target epitopes; (3) target antigen density; (4) CAR receptor expression and density; (5) the combination of intracellular signaling domains used in the CAR; and (6) whether a binding domain is prone to ligandindependent tonic signaling.

[0006] Such bi- and tri-specific autologous CAR-T cells represent a promising approach to prevent relapse but prior embodiments leave room for improvement. For example, many of the multispecific CARs in development continue to use the same antigen-binding domains used in approved CAR-T cell therapies, some of which are not fully human (e.g., the mouse-human chimeric scFv FMC63 targeting CD 19), while others use humanized versions of new non-human antigen-binding domains e.g., single chain cam elid antibodies), despite the fact that such binding domains may provoke immune rejection and have not necessarily been optimized for antigen binding. In addition, with one exception, these multi-specific CARs generally rely on the same intracellular signaling domains to activate the CAR-T cells as used in the commercially available CAR-T cells, despite the fact that up to 60% of patients treated with those products are likely to relapse.

[0007] The complex nature of the interaction between a CAR-T cell and the target tumor cell - particularly with respect to the binding interaction between the antigen-specific domain(s) and its cognate antigen(s) - renders it difficult to predict what combination of physical parameters will result in an effective therapeutic. For example, optimal antigen affinity and avidity for a CAR binder must be sufficiently high to initiate the T cell activation signaling cascade but not high enough to result in activation-induced death of the CAR T cell. However, it has also been shown that even antigen-binding fragments (i.e., single chain variable fragments, or scFvs) with similar binding affinities for the same target antigen can differentially impact CAR T cell function. Thus, empirical testing of these parameters alone and in combination is essential to developing an effective mono-, bi- or multi-specific CAR-T cell.SUMMARY

[0008] Described herein are compositions comprising one or more recombinant polynucleic acid molecules comprising: (a) a sequence encoding an anti-CD19 chimeric antigen receptor (CAR), wherein the anti-CD19 CAR comprises: (i) an extracellular domain comprising an anti-CD19 binding domain, (ii) a transmembrane domain, and (iii) a cytoplasmic domain comprising an intracellular signaling domain comprising one or more Immunoreceptor Tyrosine-based Activation Motifs (IT AMs); and (b) a sequence encoding an anti-CD20 CAR, wherein the anti-CD20 CAR comprises: (i) an extracellular domain comprising an anti-CD20 binding domain, (ii) a transmembrane domain, and (iii) a cytoplasmic domain comprising an intracellular signaling domain comprising one or more ITAMs; and (c) a sequence encoding an anti-CD22 CAR, wherein the anti- CD22 CAR comprises: (i) an extracellular domain comprising an anti-CD22 binding domain, (ii) a transmembrane domain, and (iii) a cytoplasmic domain comprising an intracellular signaling domain comprising one or more ITAMs; wherein the cytoplasmic domain of the anti-CD19 CAR and / or the cytoplasmic domain of the anti-CD20 CAR further comprises a co-stimulatory signaling domain from CD2. In some embodiments, the cytoplasmic domain of the anti-CD20 CAR comprises a costimulatory signaling domain from CD2. In some embodiments, the cytoplasmic domain of the anti- CD19 CAR comprises a co-stimulatory signaling domain from CD2, CD28, or 4- IBB.

[0009] Described herein are compositions comprising one or more recombinant polynucleic acid molecules comprising: (a) a sequence encoding an anti-CD20 CAR, wherein the anti-CD20 CAR comprises: (i) an extracellular domain comprising an anti-CD20 binding domain and / or a hinge domain, (ii) a transmembrane domain, and (iii) a cytoplasmic domain comprising (A) a co- stimulatory signaling domain from CD2 and (B) an intracellular signaling domain comprising one or more ITAMs; and (b) a sequence encoding an anti-CD22 CAR, wherein the anti-CD22 CAR comprises: (i) an extracellular domain comprising an anti-CD22 binding domain and / or a hinge domain, (ii) a transmembrane domain, and (iii) a cytoplasmic domain comprising an intracellular signaling domain comprising one or more ITAMs. In some embodiments, the one or more recombinant polynucleic acid molecules further comprises a sequence encoding an anti-CD19 CAR. In some embodiments, the anti-CD19 CAR comprises (i) an extracellular domain comprising an anti- CD19 binding domain, (ii) a transmembrane domain, and (iii) a cytoplasmic domain comprising an intracellular signaling domain. In some embodiments, the cytoplasmic domain of the anti-CD19 CAR further comprises a co-stimulatory signaling domain from CD2, CD28, 4- IBB or CD3zeta.

[0010] In another aspect, provided herein is a composition comprising one or more recombinant polynucleic acid molecules comprising: (a) a sequence encoding an anti-CD19 CAR, wherein the anti-CD19 CAR comprises: (i) an extracellular domain comprising an anti-CD19 binding domain, (ii) a transmembrane domain, and (iii) a cytoplasmic domain comprising (A) a co-stimulatory signaling domain from CD2, CD28, or 4-1BB and (B) an intracellular signaling domain comprising one or more IT AMs; and (b) a sequence encoding an anti-CD22 CAR, wherein the anti-CD22 CAR comprises: (i) an extracellular domain comprising an anti-CD22 binding domain, (ii) a transmembrane domain, and (iii) a cytoplasmic domain comprising an intracellular signaling domain comprising one or more IT AMs.

[0011] In some embodiments, the one or more recombinant polynucleic acid molecules further comprises a sequence encoding an anti-CD20 CAR, wherein the anti-CD20 CAR comprises: (i) an extracellular domain comprising an anti-CD20 binding domain, (ii) a transmembrane domain, and (iii) a cytoplasmic domain comprising an intracellular signaling domain comprising one or more IT AMs. In some embodiments, the cytoplasmic domain of the anti-CD20 CAR further comprises a co-stimulatory signaling domain from CD2. In some embodiments, the cytoplasmic domain of the anti-CD19 CAR comprises a co-stimulatory signaling domain from CD28. In some embodiments, the intracellular signaling domain is an ITAM-containing domain.

[0012] In another aspect, provided herein is a composition comprising one or more recombinant polynucleic acid molecules comprising: (a) a sequence encoding an anti-CD20 CAR, wherein the anti-CD20 CAR comprises: (i) an extracellular domain comprising an anti-CD20 binding domain,(ii) a transmembrane domain, and (iii) a cytoplasmic domain comprising an intracellular signaling domain; and (b) a sequence encoding an anti-CD19 CAR, wherein the anti-CD19 CAR comprises: (i) an extracellular domain comprising an anti-CD19 binding domain, (ii) a transmembrane domain, and (iii) a cytoplasmic domain comprising an intracellular signaling domain wherein the cytoplasmic domain of the anti-CD19 CAR and / or the cytoplasmic domain of the anti-CD20 CAR further comprises a co-stimulatory signaling domain from CD2. In some embodiments, the intracellular signaling domain is an ITAM-containing domain.

[0013] In some embodiments, the one or more recombinant polynucleic acid molecules further comprises a sequence encoding an anti-CD22 CAR, wherein the anti-CD22 CAR comprises: (i) an extracellular domain comprising an anti-CD22 binding domain, (ii) a transmembrane domain, and(iii) a cytoplasmic domain comprising an intracellular signaling domain. In some embodiments, the intracellular signaling domain is an ITAM-containing domain.

[0014] In some embodiments, the cytoplasmic domain of the anti-CD19 CAR comprises a CD28 co- stimulatory signaling domain. In some embodiments, the cytoplasmic domain of the anti-CD22 CAR further comprises a co-stimulatory signaling domain from 4-1BB.

[0015] In some embodiments, the one or more recombinant polynucleic acid molecules comprises a first recombinant polynucleic acid molecule comprising a sequence encoding the anti-CD22 CAR and a second recombinant polynucleic acid molecule comprising a sequence encoding the anti-CD19 CAR.

[0016] In some embodiments, the one or more recombinant polynucleic acid molecules comprises a first recombinant polynucleic acid molecule comprising a sequence encoding the anti-CD22 CAR and a second recombinant polynucleic acid molecule comprising a sequence encoding the anti-CD20 CAR.

[0017] In some embodiments, the one or more recombinant polynucleic acid molecules comprises a first recombinant polynucleic acid molecule comprising a sequence encoding the anti-CD19 CAR and a second recombinant polynucleic acid molecule comprising a sequence encoding the anti-CD20 CAR.

[0018] In some embodiments, the one or more recombinant polynucleic acid molecules comprises a first recombinant polynucleic acid molecule comprising a sequence encoding the anti-CD22 CAR, a second recombinant polynucleic acid molecule comprising a sequence encoding the anti-CD20 CAR, and a third recombinant polynucleic acid molecule comprising a sequence encoding the anti-CD19 CAR.

[0019] In some embodiments, the first recombinant polynucleic acid molecule is a first viral vector, the second recombinant polynucleic acid molecule is a second viral vector, and a third recombinant polynucleic acid molecule is a third viral vector. In some embodiments, the first viral vector, the second viral vector and the third viral vector are each lentiviral vectors. In some embodiments, the intracellular signaling domain is an ITAM-containing domain.

[0020] In some embodiments, the one or more recombinant polynucleic acid molecules comprises a recombinant polynucleic acid molecule comprising a sequence encoding the anti-CD22 CAR and a sequence encoding the anti-CD19 CAR. In some embodiments, the one or more recombinant polynucleic acid molecules comprises a recombinant polynucleic acid molecule comprising a sequence encoding the anti-CD22 CAR and a sequence encoding the anti-CD20 CAR. In some embodiments, the one or more recombinant polynucleic acid molecules comprises a recombinant polynucleic acid molecule comprising a sequence encoding the anti-CD19 CAR and a sequence encoding the anti-CD20 CAR.

[0021] In some embodiments, the one or more recombinant polynucleic acid molecules comprises a recombinant polynucleic acid molecule comprising a sequence encoding the anti-CD22 CAR, a sequence encoding the anti-CD20 CAR, and a sequence encoding the anti -CD 19 CAR.

[0022] In some embodiments, the sequence encoding the anti-CD19 CAR, the sequence encoding the anti-CD20 CAR, and the sequence encoding the anti-CD22 CAR are separated by a sequenceencoding a P2A site, a T2A self-cleaving peptide, an E2A self-cleaving peptide or an F2A selfcleaving peptide.

[0023] In some embodiments, the recombinant polynucleic acid molecule comprises from 5’ to 3’ the sequence encoding the anti-CD22 CAR, the sequence encoding a P2A self-cleaving peptide, a T2A self-cleaving peptide, an E2A self-cleaving peptide or an F2A self-cleaving peptide, the sequence encoding the anti-CD20 CAR, the sequence encoding a P2A self-cleaving peptide, a T2A self-cleaving peptide, an E2A self-cleaving peptide or an F2A self-cleaving peptide, and the sequence encoding the anti-CD19 CAR.

[0024] In some embodiments, the recombinant polynucleic acid molecule comprises from 5’ to 3’ the sequence encoding the anti-CD22 CAR, the sequence encoding a P2A self-cleaving peptide, a T2A self-cleaving peptide, an E2A self-cleaving peptide or an F2A self-cleaving peptide, the sequence encoding the anti-CD19 CAR, the sequence encoding the P2A self-cleaving peptide, and the sequence encoding the anti-CD20 CAR.

[0025] In some embodiments, the recombinant polynucleic acid molecule comprises from 5’ to 3’ the sequence encoding the anti-CD19 CAR, the sequence encoding a P2A self-cleaving peptide, a T2A self-cleaving peptide, an E2A self-cleaving peptide or an F2A self-cleaving peptide, the sequence encoding the anti-CD20 CAR, the sequence encoding a P2A self-cleaving peptide, a T2A self-cleaving peptide, an E2A self-cleaving peptide or an F2A self-cleaving peptide, and the sequence encoding the anti-CD22 CAR.

[0026] In some embodiments, the recombinant polynucleic acid molecule comprises from 5’ to 3’ the sequence encoding the anti-CD19 CAR, the sequence encoding ta P2A self-cleaving peptide, a T2A self-cleaving peptide, an E2A self-cleaving peptide or an F2A self-cleaving peptide, the sequence encoding the anti-CD22 CAR, the sequence encoding a P2A self-cleaving peptide, a T2A self-cleaving peptide, an E2A self-cleaving peptide or an F2A self-cleaving peptide, and the sequence encoding the anti-CD20 CAR.

[0027] In some embodiments, the recombinant polynucleic acid molecule comprises from 5’ to 3’ the sequence encoding the anti-CD20 CAR, the sequence a P2A self-cleaving peptide, a T2A selfcleaving peptide, an E2A self-cleaving peptide or an F2A self-cleaving peptide, the sequence encoding the anti-CD19 CAR, the sequence encoding a P2A self-cleaving peptide, a T2A selfcleaving peptide, an E2A self-cleaving peptide or an F2A self-cleaving peptide, and the sequence encoding the anti-CD22 CAR.

[0028] In some embodiments, the recombinant polynucleic acid molecule comprises from 5’ to 3’ the sequence encoding the anti-CD20 CAR, the sequence encoding a P2A self-cleaving peptide, aT2A self-cleaving peptide, an E2A self-cleaving peptide or an F2A self-cleaving peptide, the sequence encoding the anti-CD19 CAR, the sequence encoding a P2A self-cleaving peptide, a T2A self-cleaving peptide, an E2A self-cleaving peptide or an F2A self-cleaving peptide, and the sequence encoding the anti-CD22 CAR.

[0029] In some embodiments, the recombinant polynucleic acid molecule is a viral vector. In some embodiments, the viral vector is a lentiviral vector. In some embodiments, the intracellular signaling domain is an ITAM-containing domain.

[0030] In some embodiments, the extracellular domain of the anti-CD22 CAR further comprises a hinge domain, wherein the hinge domain is from CD8a. In some embodiments, the transmembrane domain of the anti-CD22 CAR is from CD8a or CD28. In some embodiments, the intracellular signaling domain of the anti-CD22 CAR is from CD3zeta. In some embodiments, the anti-CD22 binding domain is an antibody or an antigen binding fragment thereof, or an scFv.

[0031] In some embodiments, the anti-CD22 binding domain comprises a heavy chain variable region (VH) that comprises a heavy chain CDR3 (HCDR3) sequence selected from the group consisting of the HCDR3 sequences in Table 19.

[0032] In some embodiments, the anti-CD22 binding domain comprises a VH that comprises a HCDR2 sequence selected from the group consisting of the HCDR2 sequences in Table 19.

[0033] In some embodiments, the anti-CD22 binding domain comprises a VH that comprises a HCDR1 sequence selected from the group consisting of the HCDR1 sequences in Table 19.

[0034] In some embodiments, the anti-CD22 binding domain comprises a light chain variable region (VL) that comprises a light chain CDR3 (LCDR3) sequence selected from the group consisting of the LCDR3 sequences in Table 19.

[0035] In some embodiments, the anti-CD22 binding domain comprises a VL that comprises a LCDR2 sequence selected from the group consisting of the LCDR2 sequences in Table 19.

[0036] In some embodiments, the anti-CD22 binding domain comprises a VL that comprises a LCDR1 sequence selected from the group consisting of the LCDR1 sequences in Table 19.

[0037] In some embodiments, the anti-CD22 binding domain comprises a heavy chain variable region (VH) with a sequence with at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to a sequence selected from the group consisting of the VH sequences in Table 19.

[0038] In some embodiments, the anti-CD22 binding domain comprises a heavy chain variable region (VH) with a sequence selected from the group consisting of the VH sequences in Table 19.

[0039] In some embodiments, the anti-CD22 binding domain comprises a VH with the sequence QVQLQQSGPGLVKPSQTLSLTCAISGDSVSSNSAAWNWIRQSPSRGLEWLGRTYYRSKWYNDYAVSVKSRITINPDTSKNQFSLQLNSVTPEDTAVYYCAREVTGDLEDAFDIWGQGTMVTV SS (SEQ ID NO: 665).

[0040] In some embodiments, the anti-CD22 binding domain comprises a light chain variable region (VL) with a sequence with at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to a sequence selected from the group consisting of the VL sequences in Table 19.

[0041] In some embodiments, the anti-CD22 binding domain comprises a light chain variable region (VL) a sequence selected from the group consisting of the VL sequences in Table 19.

[0042] In some embodiments, wherein the anti-CD22 binding domain comprises a VL with the sequenceDIQMTQSPSSLSASVGDRVTITCRASQTIWSYLNWYQQRPGKAPNLLIYAASSLQSGVPSRFS GRGSGTDFTLTISSLQAEDFATYYCQQSYSIPQTFGQGTKLEIK (SEQ ID NO: 666).

[0043] In some embodiments, the anti-CD22 binding domain comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to a sequence selected from the group consisting of the sequences in Table 19.

[0044] In some embodiments, the anti-CD22 binding domain comprises a sequence selected from the group consisting of the sequences in Table 19.

[0045] In some embodiments, the anti-CD22 binding domain is an scFv.

[0046] In some embodiments, the scFv of the anti-CD22 binding domain comprises a linker between the VH and the VL and comprises the sequence GGGGS.

[0047] In some embodiments, the anti-CD22 binding domain comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the sequence:QVQLQQSGPGLVKPSQTLSLTCAISGDSVSSNSAAWNWIRQSPSRGLEWLGRTYYRSKWYN DYAVSVKSRITINPDTSKNQFSLQLNSVTPEDTAVYYCAREVTGDLEDAFDIWGQGTMVTV SSGGGGSDIQMTQSPSSLSASVGDRVTITCRASQTIWSYLNWYQQRPGKAPNLLIYAASSLQ SGVPSRFSGRGSGTDFTLTISSLQAEDFATYYCQQSYSIPQTFGQGTKLEIK (SEQ ID NO: 664).

[0048] In some embodiments, the anti-CD22 binding domain comprises the sequence:QVQLQQSGPGLVKPSQTLSLTCAISGDSVSSNSAAWNWIRQSPSRGLEWLGRTYYRSKWYN DYAVSVKSRITINPDTSKNQFSLQLNSVTPEDTAVYYCAREVTGDLEDAFDIWGQGTMVTV SSGGGGSDIQMTQSPSSLSASVGDRVTITCRASQTIWSYLNWYQQRPGKAPNLLIYAASSLQ SGVPSRFSGRGSGTDFTLTISSLQAEDFATYYCQQSYSIPQTFGQGTKLEIK (SEQ ID NO: 664).

[0049] In some embodiments, the anti-CD22 binding domain is a humanized binding domain.

[0050] In some embodiments, wherein the anti-CD22 CAR comprises a linker between the anti- CD22 binding domain and the hinge domain from CD8a or CD28.

[0051] In some embodiments, the linker between the anti-CD22 binding domain and the hinge domain from CD8a comprises the sequence AAA.

[0052] In some embodiments, the hinge domain from CD8a of the anti-CD22 CAR comprises the sequence TTTPAPRPPTPAPTIASQPLSLRPEACRPAAGGAVHTRGLDFACD (SEQ ID NO: 679).

[0053] In some embodiments, the transmembrane domain from CD8a of the anti-CD22 CAR comprises the sequence IYIWAPLAGTCGVLLLSLVIT (SEQ ID NO: 680).

[0054] In some embodiments, the anti-CD22 CAR comprises a portion of a CD8a cytoplasmic domain. In some embodiments, the portion of a CD8a cytoplasmic domain of the anti-CD22 CAR comprises the sequence LYC.

[0055] In some embodiments, the co-stimulatory signaling domain from 4- IBB comprises the sequence KRGRKKLLYIFKQPFMRPVQTTQEEDGCSCRFPEEEEGGCEL (SEQ ID NO: 998).

[0056] In some embodiments, the intracellular signaling domain from CD3zeta comprises the sequence RVKFSRSADAPAYKQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLY NELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 686).

[0057] In some embodiments, wherein the anti-CD22 CAR comprises a signal peptide sequence.

[0058] In some embodiments, the signal peptide sequence of the anti-CD22 CAR comprises the sequence MLLLVTSLLLCELPHPAFLLIP (SEQ ID NO: 1000).

[0059] In some embodiments, the anti-CD22 CAR comprises a sequence with at least 80%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the sequenceQVQLQQSGPGLVKPSQTLSLTCAISGDSVSSNSAAWNWIRQSPSRGLEWLGRTYYRSKWYN DYAVSVKSRITINPDTSKNQFSLQLNSVTPEDTAVYYCAREVTGDLEDAFDIWGQGTMVTV SSGGGGSDIQMTQSPSSLSASVGDRVTITCRASQTIWSYLNWYQQRPGKAPNLLIYAASSLQ SGVPSRFSGRGSGTDFTLTISSLQAEDFATYYCQQSYSIPQTFGQGTKLEIKAAATTTPAPRPP TPAPTIASQPLSLRPEACRPAAGGAVHTRGLDFACDIYIWAPLAGTCGVLLLSLVITLYCKRG RKKLLYIFKQPFMRP VQTTQEEDGC SCRFPEEEEGGCELRVKF SRS ADAP AYKQGQNQLYN ELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLYNELQKDKMAEAYSEIGMKGERR RGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 1009).

[0060] In some embodiments, the anti-CD22 CAR comprises the sequenceQVQLQQSGPGLVKPSQTLSLTCAISGDSVSSNSAAWNWIRQSPSRGLEWLGRTYYRSKWYN DYAVSVKSRITINPDTSKNQFSLQLNSVTPEDTAVYYCAREVTGDLEDAFDIWGQGTMVTV SSGGGGSDIQMTQSPSSLSASVGDRVTITCRASQTIWSYLNWYQQRPGKAPNLLIYAASSLQ SGVPSRFSGRGSGTDFTLTISSLQAEDFATYYCQQSYSIPQTFGQGTKLEIKAAATTTPAPRPP TPAPTIASQPLSLRPEACRPAAGGAVHTRGLDFACDIYIWAPLAGTCGVLLLSLVITLYCKRG RKKLLYIFKQPFMRP VQTTQEEDGC SCRFPEEEEGGCELRVKF SRS ADAP AYKQGQNQLYN ELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLYNELQKDKMAEAYSEIGMKGERR RGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 1009).

[0061] In some embodiments, the anti-CD22 CAR comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the sequenceMLLLVTSLLLCELPHPAFLLIPQVQLQQSGPGLVKPSQTLSLTCAISGDSVSSNSAAWNWIRQ SPSRGLEWLGRTYYRSKWYNDYAVSVKSRITINPDTSKNQFSLQLNSVTPEDTAVYYCARE VTGDLEDAFDIWGQGTMVTVSSGGGGSDIQMTQSPSSLSASVGDRVTITCRASQTIWSYLN WYQQRPGKAPNLLIYAASSLQSGVPSRFSGRGSGTDFTLTISSLQAEDFATYYCQQSYSIPQT FGQGTKLEIKAAATTTPAPRPPTPAPTIASQPLSLRPEACRPAAGGAVHTRGLDFACDIYIWA PLAGTCGVLLLSLVITLYCKRGRKKLLYIFKQPFMRPVQTTQEEDGCSCRFPEEEEGGCELR VKFSRSADAPAYKQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLYN ELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 1009).

[0062] In some embodiments, the anti-CD22 CAR comprises the sequenceMLLLVTSLLLCELPHPAFLLIPQVQLQQSGPGLVKPSQTLSLTCAISGDSVSSNSAAWNWIRQ SPSRGLEWLGRTYYRSKWYNDYAVSVKSRITINPDTSKNQFSLQLNSVTPEDTAVYYCARE VTGDLEDAFDIWGQGTMVTVSSGGGGSDIQMTQSPSSLSASVGDRVTITCRASQTIWSYLN WYQQRPGKAPNLLIYAASSLQSGVPSRFSGRGSGTDFTLTISSLQAEDFATYYCQQSYSIPQT FGQGTKLEIKAAATTTPAPRPPTPAPTIASQPLSLRPEACRPAAGGAVHTRGLDFACDIYIWA PLAGTCGVLLLSLVITLYCKRGRKKLLYIFKQPFMRPVQTTQEEDGCSCRFPEEEEGGCELR VKFSRSADAPAYKQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLYN ELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 1009).

[0063] In some embodiments, the extracellular domain of the anti-CD20 CAR further comprises a hinge domain, wherein the hinge domain is from CD8a or CD28.

[0064] In some embodiments, the transmembrane domain of the anti-CD20 CAR is from CD8a or CD28.

[0065] In some embodiments, the intracellular signaling domain of the anti-CD20 CAR is from CD3zeta.

[0066] In some embodiments, the anti-CD20 binding domain is an antibody or an antigen binding fragment thereof, or an scFv.

[0067] In some embodiments, the anti-CD20 binding domain comprises a heavy chain variable region (VH) that comprises a heavy chain CDR3 (HCDR1) sequence selected from the group consisting of the HCDR1 sequences in Tables 13-15.

[0068] In some embodiments, the anti-CD20 binding domain comprises a VH that comprises a HCDR2 sequence selected from the group consisting of the HCDR2 sequences in Tables 13-15.

[0069] In some embodiments, the anti-CD20 binding domain comprises a VH that comprises a HCDR3 sequence selected from the group consisting of the HCDR3 sequences in Tables 13-15.

[0070] In some embodiments, the anti-CD20 binding domain comprises a light chain variable region (VL) that comprises a light chain CDR3 (LCDR1) sequence selected from the group consisting of the LCDRl sequences in Tables 16-18.

[0071] In some embodiments, the anti-CD20 binding domain comprises a VL that comprises a LCDR2 sequence selected from the group consisting of the LCDR2 sequences in Tables 16-18.

[0072] In some embodiments, the anti-CD20 binding domain comprises a VL that comprises a LCDR3 sequence selected from the group consisting of the LCDR3 sequences in Tables 16-18.

[0073] In some embodiments, the anti-CD20 binding domain comprises a heavy chain variable region (VH) with a sequence with at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to a sequence selected from the group consisting of the VH sequences in Table 12.

[0074] In some embodiments, the anti-CD20 binding domain comprises a heavy chain variable region (VH) with a sequence selected from the group consisting of the VH sequences in Table 12.

[0075] In some embodiments, the anti-CD20 binding domain comprises a heavy chain variable region (VH) with a sequence selected from the group consisting of the VH sequences in Table 12.

[0076] In some embodiments, the anti-CD20 binding domain comprises a light chain variable region (VL) with a sequence selected from the group consisting of the VL sequences in Table 12.

[0077] In some embodiments, the anti-CD20 binding domain comprises a sequence with at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to a sequence selected from the group consisting of the sequences in Table 11.

[0078] In some embodiments, the anti-CD20 binding domain comprises a sequence selected from the group consisting of the sequences in Table 11.

[0079] In some embodiments, wherein the anti-CD20 binding domain is an scFv.

[0080] In some embodiments, the scFv of the anti-CD20 binding domain comprises a linker between the VH and the VL and comprises the sequence GGGGSGGGGSGGGGS (SEQ ID NO: 995).

[0081] In some embodiments, the CD28 hinge domain of the anti-CD20 CAR comprises the sequence lEVMYPPPYLDNEKSNGTIIHVKGKHLCPSPLFPGPSKP (SEQ ID NO: 682).

[0082] In some embodiments, the CD28 transmembrane domain of the anti-CD20 CAR comprises the sequence FWVLWVGGVLACYSLLVTVAFIIFWV (SEQ ID NO: 683).

[0083] In some embodiments, the cytoplasmic domain of the anti-CD20 CAR comprises a costimulatory signaling domain from CD2, and wherein the CD2 co-stimulatory signaling domain of the anti-CD20 CAR comprises the sequenceKRKKQRSRRNDEELETRAHRVATEERGRKPHQIPASTPQNPATSQHPPPPPGHRSQAPSHRP PPPGHRVQHQPQKRPPAPSGTQVHQQKGPPLPRPRVQPKPPHGAAENSLSPSSN (SEQ ID NO: 685).

[0084] In some embodiments, the intracellular signaling domain from CD3zeta of the anti-CD20 CAR comprises the sequenceRVKFSRSADAPAYKQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLY NELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 686).

[0085] In some embodiments, the anti-CD20 CAR comprises a signal peptide sequence. In some embodiments, the signal peptide sequence of the anti-CD20 CAR comprises the sequence MALPVTALLLPLALLLHAARP (SEQ ID NO: 1000) or MLLLVTSLLLCELPHPAFLLIP (SEQ ID NO: 1001).

[0086] In some embodiments, wherein the anti-CD20 CAR comprises the sequenceIEVMYPPPYLDNEKSNGTIIHVKGKHLCPSPLFPGPSKPFWVLVVVGGVLACYSLLVTVAFII FWVKRKKQRSRRNDEELETRAHRVATEERGRKPHQIPASTPQNPATSQHPPPPPGHRSQAPS HRPPPPGHRVQHQPQKRPPAPSGTQVHQQKGPPLPRPRVQPKPPHGAAENSLSPSSNRVKFS RSADAPAYKQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLYNELQK DKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPP (SEQ ID NO: 690).

[0087] In some embodiments, wherein the anti-CD20 CAR comprises a CD28 hinge domain, a CD28 transmembrane domain, a CD2 intracellular costimulatory signaling domain, and a CD3^ intracellular signaling domain having the sequenceIEVMYPPPYLDNEKSNGTIIHVKGKHLCPSPLFPGPSKPFWVLVVVGGVLACYSLLVTVAFII FWVKRKKQRSRRNDEELETRAHRVATEERGRKPHQIPASTPQNPATSQHPPPPPGHRSQAPS HRPPPPGHRVQHQPQKRPPAPSGTQVHQQKGPPLPRPRVQPKPPHGAAENSLSPSSNRVKFS RSADAPAYKQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLYNELQK DKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 1005).

[0088] In some embodiments, the anti-CD20 CAR comprises a sequence with at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to a sequence selected from the group of the CAR sequences in Table 21.

[0089] In some embodiments, the anti-CD20 CAR comprises a sequence selected from the group of the CAR sequences in Table 21.

[0090] In some embodiments, the extracellular domain of the anti-CD19 CAR further comprises a hinge domain, wherein the hinge domain is from CD8a or CD28.

[0091] In some embodiments, the transmembrane domain of the anti-CD19 CAR is from CD8a or CD28.

[0092] In some embodiments, the intracellular signaling domain of the anti-CD19 CAR is from CD3zeta.

[0093] In some embodiments, the anti-CD19 binding domain is an antibody or an antigen binding fragment thereof, or an scFv.

[0094] In some embodiments, the anti-CD19 binding domain comprises a heavy chain variable region (VH) that comprises a heavy chain CDR1 (HCDR1) sequence selected from the group consisting of the HCDR1 sequences in Tables 5-7.

[0095] In some embodiments, wherein the anti-CD19 binding domain comprises a VH that comprises a HCDR2 sequence selected from the group consisting of the HCDR2 sequences in Tables 5-7.

[0096] In some embodiments, the anti-CD19 binding domain comprises a VH that comprises a HCDR3 sequence selected from the group consisting of the HCDR3 sequences in Tables 5-7.

[0097] In some embodiments, the anti-CD19 binding domain comprises a light chain variable region (VL) that comprises a light chain CDR3 (LCDR1) sequence selected from the group consisting of the LCDR1 sequences in Tables 8-10.

[0098] In some embodiments, the anti-CD19 binding domain comprises a VL that comprises a LCDR2 sequence selected from the group consisting of the LCDR2 sequences in Tables 8-10.

[0099] In some embodiments, the anti-CD19 binding domain comprises a VL that comprises a LCDR3 sequence selected from the group consisting of the LCDR3 sequences in Tables 8-10.

[0100] In some embodiments, the anti-CD19 binding domain comprises a VH with a sequence with at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to a sequence selected from the group consisting of the VH sequences in Table 4.

[0101] In some embodiments, wherein the anti-CD19 binding domain comprises a VH with a sequence selected from the group consisting of the VH sequences in Table 4.

[0102] In some embodiments, the anti-CD19 binding domain comprises a light chain variable region (VL) a sequence with at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to a sequence selected from the group consisting of the VL sequences in Table 4.

[0103] In some embodiments, the anti-CD19 binding domain comprises a light chain variable region (VL) a sequence selected from the group consisting of the VL sequences in Table 4.

[0104] In some embodiments, the anti-CD19 binding domain comprises a sequence with at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to a sequence selected from the group consisting of the sequences in Table 3.

[0105] In some embodiments, the anti-CD19 binding domain comprises a sequence selected from the group consisting of the sequences in Table 3.

[0106] In some embodiments, the anti-CD19 binding domain is an scFv.

[0107] In some embodiments, the scFv of the anti-CD19 binding domain comprises a linker between the VH and the VL. In some embodiments, the linker comprises the sequence AAASGGGGSGGGGSGGGGSAL (SEQ ID NO: 994).

[0108] In some embodiments, the hinge domain from CD28 of the anti-CD19 CAR comprises the sequence IEVMYPPPYLDNEKSNGTIIHVKGKHLCPSPLFPGPSKP (SEQ ID NO: 682).

[0109] In some embodiments, the transmembrane domain from CD28 of the anti-CD19 CAR comprises the sequence FWVLVVVGGVLACYSLLVTVAFIIFWV (SEQ ID NO: 683).

[0110] In some embodiments, the cytoplasmic domain of the anti-CD19 CAR comprises a costimulatory signaling domain from CD28, and wherein the co-stimulatory signaling domain from CD28 of the anti-CD19 CAR comprises the sequenceRSKRSRLLHSDYMNMTPRRPGPTRKHYQPYAPPRDFAAYRS (SEQ ID NO: 684).[OHl] In some embodiments, the intracellular signaling domain from CD3zeta of the anti-CD19 CAR comprises the sequenceRVKFSRSADAPAYKQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLYNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 686).

[0112] In some embodiments, the anti-CD19 CAR comprises a signal peptide sequence.

[0113] In some embodiments, the signal peptide sequence of the anti-CD19 CAR comprises the sequence MALPVTALLLPLALLLHAARP (SEQ ID NO: 1000) or MLLLVTSLLLCELPHPAFLLIP (SEQ ID NO: 1001).

[0114] In some embodiments, the anti-CD19 CAR comprises a CD28 hinge domain, a CD28 transmembrane domain, a CD28 intracellular costimulatory signaling domain and a CD3^ intracellular signaling domain having the sequence IEVMYPPPYLDNEKSNGTIIHVKGKHLCPSPLFPGPSKPFWVLVVVGGVLACYSLLVTVAFII FWVRSKRSRLLHSDYMNMTPRRPGPTRKHYQPYAPPRDFAAYRSRVKFSRSADAPAYKQG QNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLYNELQKDKMAEAYSEIG MKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 1004).

[0115] In some embodiments, the anti-CD19 CAR comprises a sequence with at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to a sequence selected from the group of the CAR sequences in Table 21.

[0116] In some embodiments, the anti-CD19 CAR comprises a sequence selected from the group of the CAR sequences in Table 21.

[0117] In some embodiments, (a)the anti-CD19 CAR comprises: (i) an extracellular domain comprising an anti-CD19 binding domain and a hinge domain from CD28, (ii) a transmembrane domain from CD28, and (iii) a cytoplasmic domain comprising a co-stimulatory signaling domain from CD28 and an intracellular signaling domain from CD3zeta; and (b) the anti-CD20 CAR comprises: (i) an extracellular domain comprising an anti-CD20 binding domain and a hinge domain from CD28, (ii) a transmembrane domain from CD28, and (iii) a cytoplasmic domain comprising a co-stimulatory signaling domain from CD2 and an intracellular signaling domain from CD3zeta; and (c) the anti-CD22 CAR comprises: (i) an extracellular domain comprising an anti-CD22 binding domain and a hinge domain from CD8a, (ii) a transmembrane domain from CD8a, and (iii) a cytoplasmic domain comprising a co-stimulatory signaling domain from 4-1BB and an intracellular signaling domain from CD3zeta.

[0118] In another aspect, provided herein is a composition comprising a recombinant polynucleic acid molecule comprising a sequence encoding polypeptide with an anti-CD19 binding domain, wherein the anti-CD19 binding domain comprises a heavy chain variable region (VH) that comprises a heavy chain CDR3 (HCDR3) sequence selected from the group consisting of the HCDR3sequences in Tables 5-7. In some embodiments, the anti-CD19 binding domain comprises a VH that comprises a HCDR2 sequence selected from the group consisting of the HCDR2 sequences in Tables 5-7. In some embodiments, the anti-CD19 binding domain comprises a VH that comprises a HCDR1 sequence selected from the group consisting of the HCDR1 sequences in Tables 5-7. In some embodiments, the anti-CD19 binding domain comprises a light chain variable region (VL) that comprises a light chain CDR3 (HCDR3) sequence selected from the group consisting of the LCDR3 sequences in Tables 8-10. In some embodiments, the anti-CD19 binding domain comprises a VL that comprises a LCDR2 sequence selected from the group consisting of the LCDR2 sequences in Tables 8-10. In some embodiments, the anti-CD19 binding domain comprises a VL that comprises a LCDR1 sequence selected from the group consisting of the LCDR1 sequences in Tables 8-10. In some embodiments, the anti-CD19 binding domain comprises a VH with a sequence with at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to a sequence selected from the group consisting of the VH sequences in Table 4. In some embodiments, the anti-CD19 binding domain comprises a VH with a sequence selected from the group consisting of the VH sequences in Table 4. In some embodiments, the anti-CD19 binding domain comprises a light chain variable region (VL) a sequence with at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to a sequence selected from the group consisting of the VL sequences in Table 4. In some embodiments, the antiCD 19 binding domain comprises a light chain variable region (VL) a sequence selected from the group consisting of the VL sequences in Table 4. In some embodiments, the polypeptide comprises an scFv. In some embodiments, the polypeptide is an antibody. In some embodiments, the polypeptide is an anti-CD19 CAR comprising (i) an extracellular domain comprising the anti-CD19 binding domain, (ii) a transmembrane domain, and (iii) a cytoplasmic domain comprising an intracellular signaling domain. In some embodiments, the anti-CD19 CAR comprises: (i) an extracellular domain comprising the anti-CD19 binding domain and a hinge domain from CD8a or CD28, (ii) a transmembrane domain from CD8a or CD28, and (iii) a cytoplasmic domain comprising a co- stimulatory signaling domain from CD28 and an intracellular signaling domain from CD3zeta. In some embodiments, the hinge domain of the anti-CD19 CAR comprises the sequence lEVMYPPPYLDNEKSNGTIIHVKGKHLCPSPLFPGPSKP (SEQ ID NO: 682). In some embodiments, the transmembrane domain of the anti-CD20 CAR comprises the sequence FWVLVVVGGVLACYSLLVTVAFIIFWV (SEQ ID NO: 683). the co-stimulatory signaling domain of the anti-CD19 CAR comprises the sequenceRSKRSRLLHSDYMNMTPRRPGPTRKHYQPYAPPRDFAAYRS (SEQ ID NO: 684). In some embodiments, the intracellular signaling domain of the anti-CD19 CAR comprises the sequenceRVKFSRSADAPAYKQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLY NELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 686). In some embodiments, the anti-CD19 CAR comprises a signal peptide sequence. In some embodiments, the signal peptide sequence comprises the sequence MALPVTALLLPL ALLLEIA ARP (SEQ ID NO: 1000) or MLLLVTSLLLCELPHPAFLLIP (SEQ ID NO: 1001).

[0119] In another aspect, provided herein is a composition comprising a recombinant polynucleic acid molecule comprising a sequence encoding polypeptide with an anti-CD20 binding domain, wherein the anti-CD20 binding domain comprises a heavy chain variable region (VH) that comprises a heavy chain CDR3 (HCDR3) sequence selected from the group consisting of the HCDR3 sequences in Table 13-15. In some embodiments, the anti-CD20 binding domain comprises a VH that comprises a HCDR2 sequence selected from the group consisting of the HCDR2 sequences in Table 13-15. In some embodiments, the anti-CD20 binding domain comprises a VH that comprises a HCDR1 sequence selected from the group consisting of the HCDR1 sequences in Table 13-15. In some embodiments, the anti-CD20 binding domain comprises a light chain variable region (VL) that comprises a light chain CDR3 (HCDR3) sequence selected from the group consisting of the LCDR3 sequences in Table 15-17. In some embodiments, the anti-CD20 binding domain comprises a VL that comprises a LCDR2 sequence selected from the group consisting of the LCDR2 sequences in Table 15-17. In some embodiments, the anti-CD20 binding domain comprises a VL that comprises a LCDR1 sequence selected from the group consisting of the LCDR1 sequences in Table 15-17. In some embodiments, the anti-CD20 binding domain comprises a heavy chain variable region (VH) with a sequence having at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to a sequence selected from the group consisting of the VH sequences in Table 12. In some embodiments, the anti-CD20 binding domain comprises a heavy chain variable region (VH) with a sequence selected from the group consisting of the VH sequences in Table 12. In some embodiments, the anti-CD20 binding domain comprises a light chain variable region (VL) a sequence with at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to a sequence selected from the group consisting of the VL sequences in Table 12. In some embodiments, the anti- CD20 binding domain comprises a light chain variable region (VL) a sequence selected from the group consisting of the VL sequences in Table 12. In some embodiments, the polypeptide comprises an scFv. In some embodiments, the polypeptide is an antibody. In some embodiments, the polypeptide is an anti-CD20 CAR comprising (i) an extracellular domain comprising the anti-CD20 binding domain, (ii) a transmembrane domain, and (iii) a cytoplasmic domain comprising an intracellular signaling domain. In some embodiments, the anti-CD20 CAR comprises (i) anextracellular domain comprising an anti-CD20 binding domain and a hinge domain from CD28, (ii) a transmembrane domain from CD28, and (iii) a cytoplasmic domain comprising a co-stimulatory signaling domain from CD2 and an intracellular signaling domain from CD3zeta. In some embodiments, the hinge domain of the anti-CD20 CAR comprises the sequence IEVMYPPPYLDNEKSNGTIIHVKGKHLCPSPLFPGPSKP (SEQ ID NO: 682). In some embodiments, the transmembrane domain of the anti-CD20 CAR comprises the sequence FWVLVVVGGVLACYSLLVTVAFIIFWV (SEQ ID NO: 683). In some embodiments, the costimulatory signaling domain of the anti-CD20 CAR comprises the sequence KRKKQRSRRNDEELETRAHRVATEERGRKPHQIPASTPQNPATSQHPPPPPGHRSQAPSHRP PPPGHRVQHQPQKRPPAPSGTQVHQQKGPPLPRPRVQPKPPHGAAENSLSPSSN (SEQ ID NO: 685). In some embodiments, the intracellular signaling domain of the anti-CD20 CAR comprises the sequence RVKFSRSADAPAYKQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLY NELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 686). In some embodiments, the anti-CD20 CAR comprises a signal peptide sequence. In some embodiments, the signal peptide sequence comprises the sequence MALPVTALLLPLALLLHAARP (SEQ ID NO: 1000) or MLLLVTSLLLCELPHPAFLLIP (SEQ ID NO: 1001).

[0120] In another aspect, provided herein is a composition comprising one or more recombinant polynucleic acid molecules comprising: (a) a sequence encoding a bispecific chimeric antigen receptor (CAR), wherein the bispecific CAR comprises: (i) an extracellular domain comprising an anti-CD19 binding domain and an anti-CD20 binding domain, (ii) a transmembrane domain, and (iii) a cytoplasmic domain comprising an intracellular signaling domain; and wherein the cytoplasmic domain further comprises a co-stimulatory signaling domain from CD2. In some embodiments, the one or more recombinant polynucleic acid molecules further comprises a sequence encoding an anti- CD22 CAR, wherein the anti-CD22 CAR comprises: (i) an extracellular domain comprising an anti- CD22 binding domain, (ii) a transmembrane domain, and (iii) a cytoplasmic domain comprising an intracellular signaling domain.

[0121] In another aspect, provided herein is a composition comprising one or more recombinant polynucleic acid molecules comprising: (a) a sequence encoding a bispecific CAR, wherein the bispecific CAR comprises: (i) an extracellular domain comprising an anti-CD19 binding domain and an anti-CD22 binding domain, (ii) a transmembrane domain; and (iii) a cytoplasmic domain comprising (A) a co-stimulatory signaling domain from CD2, CD28, or 4- IBB and (B) an intracellular signaling domain.

[0122] In some embodiments, the one or more recombinant polynucleic acid molecules further comprises a sequence encoding an anti-CD20 CAR, wherein the anti-CD20 CAR comprises: (i) an extracellular domain comprising an anti-CD20 binding domain, (ii) a transmembrane domain, and (iii) a cytoplasmic domain comprising an intracellular signaling domain. In some embodiments, the cytoplasmic domain of the anti-CD20 CAR further comprises a co-stimulatory signaling domain from CD2.

[0123] In another aspect, provided herein is a composition comprising one or more recombinant polynucleic acid molecules comprising: (a) a sequence encoding a bispecific CAR, wherein the bispecific CAR comprises: (i) an extracellular domain comprising an anti-CD20 binding domain and anti-CD22 binding domain, (ii) a transmembrane domain, and (iii) a cytoplasmic domain comprising (A) a co-stimulatory signaling domain from CD2 and (B) an intracellular signaling domain.

[0124] In some embodiments, the one or more recombinant polynucleic acid molecules further comprises a sequence encoding an anti-CD19 CAR, wherein the anti-CD19 CAR comprises: (i) an extracellular domain comprising an anti-CD19 binding domain, (ii) a transmembrane domain, and (iii) a cytoplasmic domain comprising an intracellular signaling domain. In some embodiments, the cytoplasmic domain of the anti-CD19 CAR further comprises a co-stimulatory signaling domain from CD2, CD28, 4- IBB or CD3zeta. In some embodiments, the cytoplasmic domain of the anti- CD19 CAR comprises a co-stimulatory signaling domain from CD28.

[0125] In another aspect, provided herein is a composition comprising one or more recombinant polynucleic acid molecules comprising: (a) a sequence encoding a bispecific chimeric antigen receptor (CAR), wherein the bispecific CAR comprises: (i) an extracellular domain comprising an anti-CD19 binding domain and an anti-CD20 binding domain, (ii) a transmembrane domain, and (iii) a cytoplasmic domain comprising an intracellular signaling domain; and (b) a sequence encoding an anti-CD22 CAR, wherein the anti-CD22 CAR comprises: (i) an extracellular domain comprising an anti-CD22 binding domain, (ii) a transmembrane domain, and (iii) a cytoplasmic domain comprising an intracellular signaling domain; wherein the cytoplasmic domain of the bispecific CAR further comprises a co-stimulatory signaling domain from CD2. In some embodiments, the extracellular domain of the bispecific CAR further comprises a hinge domain, wherein the hinge domain is from CD8a or CD28. In some embodiments, the transmembrane domain of the bispecific CAR is from CD8a or CD28. In some embodiments, the intracellular signaling domain of the bispecific CAR is from CD3zeta. In some embodiments, the hinge domain from CD8a or CD28 comprises the sequence IEVMYPPPYLDNEKSNGTIIHVKGKHLCPSPLFPGPSKP (SEQ ID NO: 682). In some embodiments, the transmembrane domain from CD28 comprises the sequenceFWVLVVVGGVLACYSLLVTVAFIIFWV (SEQ ID NO: 683). In some embodiments, the wherein the cytoplasmic domain of the bispecific CAR or the anti-CD20 CAR comprises a co-stimulatory signaling domain from CD2, and wherein the co-stimulatory signaling domain from CD2 comprises the sequence KRKKQRSRRNDEELETRAHRVATEERGRKPHQIPASTPQNPATSQHPPPPPGHRSQAPSHRP PPPGHRVQHQPQKRPPAPSGTQVHQQKGPPLPRPRVQPKPPHGAAENSLSPSSN (SEQ ID NO: 685).

[0126] In some embodiments, the intracellular signaling domain from CD3zeta of the bispecific CAR comprises the sequence RVKFSRSADAPAYKQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLY NELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 686).

[0127] In some embodiments, the anti-CD20 binding domain is an antibody or an antigen binding fragment thereof, or an scFv.

[0128] In some embodiments, the anti-CD20 binding domain comprises a heavy chain variable region (VH) that comprises a heavy chain CDR1 (HCDR1) sequence selected from the group consisting of the HCDR3 sequences in Tables 13-15.

[0129] In some embodiments, the anti-CD20 binding domain comprises a VH that comprises a HCDR2 sequence selected from the group consisting of the HCDR2 sequences in Tables 13-15.

[0130] In some embodiments, the anti-CD20 binding domain comprises a VH that comprises a HCDR3 sequence selected from the group consisting of the HCDR3 sequences in Tables 13-15.

[0131] In some embodiments, the anti-CD20 binding domain comprises a light chain variable region (VL) that comprises a light chain CDR1 (LCDR1) sequence selected from the group consisting of the LCDR3 sequences in Tables 16-18.

[0132] In some embodiments, the anti-CD20 binding domain comprises a VL that comprises a LCDR2 sequence selected from the group consisting of the LCDR2 sequences in Tables 16-18.

[0133] In some embodiments, the anti-CD20 binding domain comprises a VL that comprises a LCDR3 sequence selected from the group consisting of the LCDR3 sequences in Tables 16-18.

[0134] In some embodiments, the anti-CD20 binding domain comprises a heavy chain variable region (VH) with a sequence with at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to a sequence selected from the group consisting of the VH sequences in Table 12.

[0135] In some embodiments, the anti-CD20 binding domain comprises a heavy chain variable region (VH) with a sequence selected from the group consisting of the VH sequences in Table 12.

[0136] In some embodiments, the anti-CD20 binding domain comprises a light chain variable region (VL) with a sequence with at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to selected from the group consisting of the VH sequences in Table 12.

[0137] In some embodiments, the anti-CD20 binding domain comprises a light chain variable region (VL) with a sequence selected from the group consisting of the VL sequences in Table 12.

[0138] In some embodiments, the anti-CD20 binding domain comprises a sequence with at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to a sequence selected from the group consisting of the sequences in Table 11.

[0139] In some embodiments, the anti-CD20 binding domain comprises a sequence selected from the group consisting of the sequences in Table 11.

[0140] In some embodiments, the anti-CD20 binding domain is an scFv.

[0141] In some embodiments, the scFv of the anti-CD20 binding domain comprises a linker between the VH and the VL and comprises the sequence GGGGSGGGGSGGGGS (SEQ ID NO: 995).

[0142] In some embodiments, the anti-CD19 binding domain is an antibody or an antigen binding fragment thereof, or an scFv.

[0143] In some embodiments, the anti-CD19 binding domain comprises a heavy chain variable region (VH) that comprises a heavy chain CDR1 (HCDR1) sequence selected from the group consisting of the HCDR3 sequences in Tables 5-7.

[0144] In some embodiments, the anti-CD19 binding domain comprises a VH that comprises a HCDR2 sequence selected from the group consisting of the HCDR2 sequences in Tables 5-7.

[0145] In some embodiments, the anti-CD19 binding domain comprises a VH that comprises a HCDR3 sequence selected from the group consisting of the HCDR3 sequences in Tables 5-7.

[0146] In some embodiments, the anti-CD19 binding domain comprises a light chain variable region (VL) that comprises a light chain CDR1 (LCDR1) sequence selected from the group consisting of the LCDR1 sequences in Tables 8-10.

[0147] In some embodiments, the anti-CD19 binding domain comprises a VL that comprises a LCDR2 sequence selected from the group consisting of the LCDR2 sequences in Tables 8-10.

[0148] In some embodiments, the anti-CD19 binding domain comprises a VL that comprises a LCDR3 sequence selected from the group consisting of the LCDR3 sequences in Tables 8-10.

[0149] In some embodiments, the anti-CD19 binding domain comprises a VH with a sequence with at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to a sequence selected from the group consisting of the VH sequences in Table 4.

[0150] In some embodiments, the anti-CD19 binding domain comprises a VH with a sequence selected from the group consisting of the VH sequences in Table 4.

[0151] In some embodiments, the anti-CD19 binding domain comprises a light chain variable region (VL) a sequence with at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to a sequence selected from the group consisting of the VL sequences in Table 4.

[0152] In some embodiments, the anti-CD19 binding domain comprises a light chain variable region (VL) a sequence selected from the group consisting of the VL sequences in Table 4.

[0153] In some embodiments, the anti-CD19 binding domain comprises a sequence with at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to a sequence selected from the group consisting of the sequences in Table 3.

[0154] In some embodiments, the anti-CD19 binding domain comprises a sequence selected from the group consisting of the sequences in Table 3.

[0155] In some embodiments, the anti-CD19 binding domain is an scFv.

[0156] In some embodiments, the scFv of the anti-CD19 binding domain comprises a linker between the VH and the VL and comprises the sequence AAASGGGGSGGGGSGGGGSAL (SEQ ID NO: 994).

[0157] In some embodiments, the anti-CD22 binding domain is an antibody or an antigen binding fragment thereof, or an scFv.

[0158] In some embodiments, the anti-CD22 binding domain comprises a heavy chain variable region (VH) that comprises a heavy chain CDR1 (HCDR1) sequence selected from the group consisting of the HCDR1 sequences in Table 19.

[0159] In some embodiments, the anti-CD22 binding domain comprises a VH that comprises a HCDR2 sequence selected from the group consisting of the HCDR2 sequences in Table 19.

[0160] In some embodiments, the anti-CD22 binding domain comprises a VH that comprises a HCDR3 sequence selected from the group consisting of the HCDR3 sequences in Table 19.

[0161] In some embodiments, the anti-CD22 binding domain comprises a light chain variable region (VL) that comprises a light chain CDR1 (LCDR1) sequence selected from the group consisting of the LCDR3 sequences in Table 19.

[0162] In some embodiments, the anti-CD22 binding domain comprises a VL that comprises a LCDR2 sequence selected from the group consisting of the LCDR2 sequences in Table 19.

[0163] In some embodiments, the anti-CD22 binding domain comprises a VL that comprises a LCDR3 sequence selected from the group consisting of the LCDR3 sequences in Table 19.

[0164] In some embodiments, the anti-CD22 binding domain comprises a heavy chain variable region (VH) with a sequence with at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to a sequence selected from the group consisting of the VH sequences in Table 19.

[0165] In some embodiments, wherein the anti-CD22 binding domain comprises a heavy chain variable region (VH) with a sequence selected from the group consisting of the VH sequences in Table 19.

[0166] In some embodiments, wherein the anti-CD22 binding domain comprises a light chain variable region (VL) with a sequence with at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to a sequence selected from the group consisting of the VL sequences in Table 19.

[0167] In some embodiments, the anti-CD22 binding domain comprises a light chain variable region (VL) a sequence selected from the group consisting of the VL sequences in Table 19.

[0168] In some embodiments, the anti-CD22 binding domain comprises a VL with the sequence DIQMTQSPSSLSASVGDRVTITCRASQTIWSYLNWYQQRPGKAPNLLIYAASSLQSGVPSRFS GRGSGTDFTLTISSLQAEDFATYYCQQSYSIPQTFGQGTKLEIK (SEQ ID NO: 666).

[0169] In some embodiments, the anti-CD22 binding domain comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to a sequence selected from the group consisting of the sequences in Table 19.

[0170] In some embodiments, the anti-CD22 binding domain comprises a sequence selected from the group consisting of the sequences in Table 19.

[0171] In some embodiments, the anti-CD22 binding domain is an scFv.

[0172] In some embodiments, the scFv of the anti-CD22 binding domain comprises a linker between the VH and the VL and comprises the sequence GGGGS (SEQ ID NO: 1007).

[0173] In some embodiments, the bispecific CAR comprises a sequence with at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to a sequence selected from the group of sequences in Table 23.

[0174] In some embodiments, the bispecific CAR comprises a sequence selected from the group of sequences in Table 23.

[0175] In some embodiments, the anti-CD20 CAR comprises a sequence with at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to a sequence selected from the group of the CAR sequences in Table 21.

[0176] In some embodiments, the anti-CD20 CAR comprises a sequence selected from the group of the CAR sequences in Table 21.

[0177] In some embodiments, the anti-CD19 CAR comprises a sequence with at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to a sequence selected from the group of the CAR sequences in Table 21.

[0178] In some embodiments, the anti-CD19 CAR comprises a sequence selected from the group of the CAR sequences in Table 21.

[0179] In some embodiments, the anti-CD22 CAR comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the sequenceMLLLVTSLLLCELPHPAFLLIPQVQLQQSGPGLVKPSQTLSLTCAISGDSVSSNSAAWNWIRQ SPSRGLEWLGRTYYRSKWYNDYAVSVKSRITINPDTSKNQFSLQLNSVTPEDTAVYYCARE VTGDLEDAFDIWGQGTMVTVSSGGGGSDIQMTQSPSSLSASVGDRVTITCRASQTIWSYLN WYQQRPGKAPNLLIYAASSLQSGVPSRFSGRGSGTDFTLTISSLQAEDFATYYCQQSYSIPQT FGQGTKLEIKAAATTTPAPRPPTPAPTIASQPLSLRPEACRPAAGGAVHTRGLDFACDIYIWA PLAGTCGVLLLSLVITLYCKRGRKKLLYIFKQPFMRPVQTTQEEDGCSCRFPEEEEGGCELR VKFSRSADAPAYKQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLYN ELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 1009).

[0180] In some embodiments, the anti-CD22 CAR comprises the sequenceMLLLVTSLLLCELPHPAFLLIPQVQLQQSGPGLVKPSQTLSLTCAISGDSVSSNSAAWNWIRQ SPSRGLEWLGRTYYRSKWYNDYAVSVKSRITINPDTSKNQFSLQLNSVTPEDTAVYYCARE VTGDLEDAFDIWGQGTMVTVSSGGGGSDIQMTQSPSSLSASVGDRVTITCRASQTIWSYLN WYQQRPGKAPNLLIYAASSLQSGVPSRFSGRGSGTDFTLTISSLQAEDFATYYCQQSYSIPQT FGQGTKLEIKAAATTTPAPRPPTPAPTIASQPLSLRPEACRPAAGGAVHTRGLDFACDIYIWA PLAGTCGVLLLSLVITLYCKRGRKKLLYIFKQPFMRPVQTTQEEDGCSCRFPEEEEGGCELR VKFSRSADAPAYKQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLYN ELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 1009).

[0181] In another aspect, the one or more recombinant polynucleic acid molecules of the composition encode one or more polypeptides.

[0182] In another aspect, provided herein is a cell comprising one or more recombinant polynucleic acid molecules of the composition disclosed herein. In some embodiments, the cell is an immune cell. In some embodiments, the cell is a T cell. In some embodiments, the cell is an NK cell. In some embodiments, the cell is a population of cells comprising at least 1 >< 1OA3 cells. In some embodiments, at least 60%, 70%, 80%, 90%, 95% or more of the cells in the population of cellsexpress the anti-CD19 CAR and the anti-CD22 CAR. In some embodiments, at least 60%, 70%, 80%, 90%, 95% or more of the cells in the population of cells express the anti-CD20 CAR and the anti-CD22 CAR. In some embodiments, at least 60%, 70%, 80%, 90%, 95% or more of the cells in the population of cells express the anti-CD19 CAR and the anti-CD20 CAR. In some embodiments, at least 60%, 70%, 80%, 90%, 95% or more of the cells in the population of cells express the anti- CD19 CAR, the anti-CD20 CAR, and the anti-CD22 CAR. In some embodiments, at least 70% or more of the cells in the population of cells express the anti-CD19 CAR, the anti-CD20 CAR, and the anti-CD22 CAR. In some embodiments, at least 80% or more of the cells in the population of cells express the anti-CD19 CAR, the anti-CD20 CAR, and the anti-CD22 CAR. In some embodiments, at least 90% or more of the cells in the population of cells express the anti-CD19 CAR, the anti-CD20 CAR, and the anti-CD22 CAR.

[0183] In another aspect, provided herein is a pharmaceutical composition comprising one or more of the recombinant polynucleic acid molecules of the compositions disclosed herein. In some embodiments, the pharmaceutical the composition comprises one or more polypeptides disclosed herein. In some embodiments, the pharmaceutical composition further comprises a cell disclosed herein.

[0184] In another aspect, provided herein is a method of making a cell comprising introducing one or more recombinant polynucleic acid molecules of the compositions disclosed herein or any one or more of the polypeptides disclosed herein into a cell.

[0185] In another aspect, provided herein is a method of treating a disease or condition in a subject in need thereof comprising administering a therapeutically effective amount of a pharmaceutical composition disclosed herein. In some embodiments, the disease or condition is cancer or a relapsed / refractory cancer. In some embodiments, the disease or condition is a lymphoma or a leukemia. In some embodiments, the disease or condition is relapsed B-cell acute lymphoblastic leukemia, chronic lymphocytic leukemia, adult acute lymphoblastic leukemia, large B-cell lymphoma, follicular lymphoma or mantle cell lymphoma.

[0186] In another aspect, provided herein is a viral vector comprising a polynucleic acid sequence that is at least 3000 nucleotides in length, wherein the polynucleic acid sequence encodes a polypeptide that is at least 1000 amino acids in length, and wherein the polypeptide is expressed in at least 50% of the cells in a cell population when transduced into the cell population.

[0187] In another aspect, provided herein is a viral vector comprising a polynucleic acid sequence that is at least 3000 nucleotides in length, wherein the polynucleic acid sequence encodes at least two polypeptides, wherein the sum of the lengths of the at least two polypeptides is at least 1000 aminoacids in length, and wherein each of the at least two polypeptides is expressed in at least 50% of the cells in a cell population when transduced into the cell population.

[0188] In some embodiments, the polynucleic acid sequence is at least 4500 nucleotides in length and encodes one or more polypeptides that are at least 1500 amino acids in length in total. In some embodiments, the polynucleic acid sequence is at least 4200 nucleotides in length and encodes one or more polypeptides that are at least 1400 amino acids in length in total. In some embodiments, the polynucleic acid sequence is at least 3900 nucleotides in length and encodes one or more polypeptides that are at least 1300 amino acids in length in total. In some embodiments, the polynucleic acid sequence is at least 3600 nucleotides in length and encodes one or more polypeptides that are at least 1200 amino acids in length in total. In some embodiments, the polypeptide is a CAR. In some embodiments, each of the at least two polypeptides is a CAR. In some embodiments, the cell is a T cell.

[0189] In another aspect, provided herein is a population of at least 1 x 10A3 cells expressing a polypeptide encoded by a polynucleic acid sequence of a single viral vector that is at least 3000 nucleotides in length, wherein the polypeptide is at least 1000 amino acids in length. In some embodiments, the polynucleic acid sequence is at least 4500 nucleotides in length and encodes one or more polypeptides that are at least 1500 amino acids in length in total. In some embodiments, the polynucleic acid sequence is at least 4200 nucleotides in length and encodes one or more polypeptides that are at least 1400 amino acids in length in total. In some embodiments, the polynucleic acid sequence is at least 3900 nucleotides in length and encodes one or more polypeptides that are at least 1300 amino acids in length in total. In some embodiments, the polypeptide is a CAR. In some embodiments, the cell is a T cell.

[0190] Also provided herein is a composition comprising one or more recombinant polynucleic acid molecules comprising a sequence encoding an anti-CD19 chimeric antigen receptor (CAR), the antiCD 19 CAR comprising: an extracellular domain comprising an anti-CD19 binding domain; a transmembrane domain; and a cytoplasmic domain comprising an intracellular signaling domain comprising one or more immunoreceptor tyrosine-based activation motifs (IT AMs) and optionally an intracellular costimulatory signaling domain; and a sequence encoding an anti-CD20 CAR, the anti- CD20 CAR comprising: an extracellular domain comprising an anti-CD20 binding domain; a transmembrane domain; and a cytoplasmic domain comprising an intracellular signaling domain comprising one or more ITAMs and optionally an intracellular costimulatory signaling domain derived from CD2; and a sequence encoding an anti-CD22 CAR, the anti-CD22 CAR comprising: an extracellular domain comprising an anti-CD22 binding domain; a transmembrane domain; and

[0191] a cytoplasmic domain comprising an intracellular signaling domain comprising one or more IT AMs and optionally an intracellular costimulatory signaling domain.

[0192] In some embodiments, the sequence encoding an anti-CD19 CAR, the sequence encoding an anti-CD20 CAR, and the sequence encoding an anti-CD22 CAR are encoded on the same polynucleic acid molecule.

[0193] In some embodiments, the sequence encoding the anti-CD19 CAR, the sequence encoding the anti-CD20 CAR, and the sequence encoding the anti-CD22 CAR are each separated from the sequences encoding the other CARs by a sequence encoding a viral ribosome skipping peptide selected from the group consisting of a P2A peptide, a T2A peptide, an E2A peptide, and an F2A peptide.

[0194] In some embodiments, the single recombinant polynucleic acid comprises, from 5’ to 3’, a sequence encoding the anti-CD22 CAR; a sequence encoding a viral ribosome skipping peptide selected from the group consisting of a P2A peptide, a T2A peptide, an E2A peptide, and an F2A peptide; a sequence encoding the anti-CD19 CAR; a sequence encoding a viral ribosome skipping peptide selected from the group consisting of a P2A peptide, a T2A peptide, an E2A peptide, and an F2A peptide; and a sequence encoding the anti-CD20 CAR.

[0195] In some embodiments, the single recombinant polynucleic acid comprises, from 5’ to 3’, a sequence encoding the anti-CD22 CAR; a sequence encoding a viral P2A ribosome skipping peptide; a sequence encoding the anti-CD19 CAR; a sequence encoding a viral T2A ribosome skipping peptide; and a sequence encoding the anti-CD20 CAR.

[0196] In some embodiments, the anti-CD22 CAR binding domain comprises an anti-CD22 antibody or an antigen-binding fragment thereof.

[0197] In some embodiments, the anti-CD22 CAR binding domain is an antigen-binding fragment of an anti-CD22 antibody.

[0198] In some embodiments, the antigen-binding fragment of an anti-CD22 antibody is a single chain variable fragment (scFv).

[0199] In some embodiments, the anti-CD22 binding domain comprises a heavy chain variable region (VH) that comprises a first heavy chain complementarity determining region (HCDR1) having the sequence GDSVSSNSA (SEQ ID NO: 667), a second heavy chain complementarity determining region (HCDR2) having the sequence TYYRSKWYN (SEQ ID NO: 675), and a third heavy chain complementarity determining region (HCDR3) having the sequence AREVTGDLEDAFDI (SEQ ID NO: 676).

[0200] In some embodiments, the anti-CD22 binding domain comprises a light chain variable region (VL) that comprises a first light chain complementarity determining region (LCDR1) having the sequence QTIWSY (SEQ ID NO: 677), a second light chain complementarity determining region (LCDR2) having the sequence AASSLQS (SEQ ID NO: 671), and a third light chain complementarity determining region (LCDR3) having the sequence AAS (SEQ ID NO: 678).

[0201] In some embodiments, the anti-CD22 binding domain comprises a VH comprising a sequence having at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the sequence QVQLQQSGPGLVKPSQTLSLTCAISGDSVSSNSAAWNWIRQSPSRGLEWLGRTYYRSKWYN DYAVSVKSRITINPDTSKNQFSLQLNSVTPEDTAVYYCAREVTGDLEDAFDIWGQGTMVTV SS (SEQ ID NO: 665) and a VL comprising a sequence having at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the sequenceDIQMTQSPSSLSASVGDRVTITCRASQTIWSYLNWYQQRPGKAPNLLIYAASSLQSGVPSRFS GRGSGTDFTLTISSLQAEDFATYYCQQSYSIPQTFGQGTKLEIK (SEQ ID NO: 666).

[0202] In some embodiments, the anti-CD22 binding domain comprises a VH having the sequence QVQLQQSGPGLVKPSQTLSLTCAISGDSVSSNSAAWNWIRQSPSRGLEWLGRTYYRSKWYN DYAVSVKSRITINPDTSKNQFSLQLNSVTPEDTAVYYCAREVTGDLEDAFDIWGQGTMVTV SS (SEQ ID NO: 665) and a VL having the sequence DIQMTQSPSSLSASVGDRVTITCRASQTIWSYLNWYQQRPGKAPNLLIYAASSLQSGVPSRFS GRGSGTDFTLTISSLQAEDFATYYCQQSYSIPQTFGQGTKLEIK (SEQ ID NO: 666).

[0203] In some embodiments, the VH and VL are linked by a linker comprising the sequence GGGGS (SEQ ID NO: 1007).

[0204] In some embodiments, the anti-CD22 binding domain comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the sequence QVQLQQSGPGLVKPSQTLSLTCAISGDSVSSNSAAWNWIRQSPSRGLEWLGRTYYRSKWYN DYAVSVKSRITINPDTSKNQFSLQLNSVTPEDTAVYYCAREVTGDLEDAFDIWGQGTMVTV SSGGGGSDIQMTQSPSSLSASVGDRVTITCRASQTIWSYLNWYQQRPGKAPNLLIYAASSLQ SGVPSRFSGRGSGTDFTLTISSLQAEDFATYYCQQSYSIPQTFGQGTKLEIK (SEQ ID NO: 664).

[0205] In some embodiments, the anti-CD22 binding domain comprises an scFv having the sequence QVQLQQSGPGLVKPSQTLSLTCAISGDSVSSNSAAWNWIRQSPSRGLEWLGRTYYRSKWYN DYAVSVKSRITINPDTSKNQFSLQLNSVTPEDTAVYYCAREVTGDLEDAFDIWGQGTMVTV SSGGGGSDIQMTQSPSSLSASVGDRVTITCRASQTIWSYLNWYQQRPGKAPNLLIYAASSLQSGVPSRFSGRGSGTDFTLTISSLQAEDFATYYCQQSYSIPQTFGQGTKLEIK (SEQ ID NO: 664).

[0206] In some embodiments, the anti-CD22 CAR further comprises a hinge domain.

[0207] In some embodiments, the hinge domain of the anti-CD22 CAR is derived from CD8a or from CD28 and the transmembrane domain of the anti-CD22 CAR is derived from CD8 a or from CD28.

[0208] In some embodiments, the hinge domain of the anti-CD22 CAR is derived from CD8a and the transmembrane domain is derived from CD8a.

[0209] In some embodiments, the CD8a hinge domain has the sequence TTTPAPRPPTPAPTIASQPLSLRPEACRPAAGGAVHTRGLDFACD (SEQ ID NO: 679).

[0210] In some embodiments, the CD8a transmembrane domain has the sequence IYIWAPLAGTCGVLLLSLVIT (SEQ ID NO: 680).

[0211] In some embodiments, the CD8a transmembrane domain further comprises a portion of the CD8a cytoplasmic domain having the sequence LYC (SEQ ID NO: 681).

[0212] In some embodiments, the cytoplasmic domain of the anti-CD22 CAR comprises an intracellular co-stimulatory domain derived from a molecule selected from the group consisting of CD27, CD28, 4-1BB / CD137, 0X40, ICOS, and CD2.

[0213] In some embodiments, the intracellular co-stimulatory domain is derived from 4- 1BB / CD137.

[0214] In some embodiments, the 4-1BB / CD137 intracellular co-stimulatory domain comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the sequence KRGRKKLLYIFKQPFMRPVQTTQEEDGCSCRFPEEEEGGCEL (SEQ ID NO: 998).

[0215] In some embodiments, the 4-1BB / CD137 intracellular co-stimulatory domain has the sequence KRGRKKLLYIFKQPFMRPVQTTQEEDGCSCRFPEEEEGGCEL (SEQ ID NO: 998).

[0216] In some embodiments, the cytoplasmic domain of the anti-CD22 CAR comprises an intracellular signaling domain comprising one or more ITAMs derived from CD3 □.

[0217] In some embodiments, the CD3(^ intracellular signaling domain comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the sequence RVKFSRSADAPAYKQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLY NELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 686).

[0218] In some embodiments, the CD3(^ intracellular signaling domain has the sequence RVKFSRSADAPAYKQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLYNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 686).

[0219] In some embodiments, the CD3(^ intracellular signaling domain comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the sequence RVKFSRSADAPAYQQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLY NELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 687).

[0220] In some embodiments, the CD3(^ intracellular signaling domain has the sequence RVKFSRSADAPAYQQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLY NELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 687).

[0221] In some embodiments, the CD3(^ intracellular signaling domain comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the sequence RVKFSRSADAPAYQQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPQRRKNPQEGL YNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 688).

[0222] In some embodiments, the CD3(^ intracellular signaling domain has the sequence RVKFSRSADAPAYQQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPQRRKNPQEGL YNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 688).

[0223] In some embodiments, the CD3(^ intracellular signaling domain comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the sequence RVKFSRSADAPAYKQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPQRRKNPQEGL YNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 689).

[0224] In some embodiments, the CD3(^ intracellular signaling domain has the sequence RVKFSRSADAPAYKQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPQRRKNPQEGL YNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 689).

[0225] In some embodiments, the anti-CD22 CAR comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the sequence QVQLQQSGPGLVKPSQTLSLTCAISGDSVSSNSAAWNWIRQSPSRGLEWLGRTYYRSKWYN DYAVSVKSRITINPDTSKNQFSLQLNSVTPEDTAVYYCAREVTGDLEDAFDIWGQGTMVTVSSGGGGSDIQMTQSPSSLSASVGDRVTITCRASQTIWSYLNWYQQRPGKAPNLLIYAASSLQ SGVPSRFSGRGSGTDFTLTISSLQAEDFATYYCQQSYSIPQTFGQGTKLEIKAAATTTPAPRPP TPAPTIASQPLSLRPEACRPAAGGAVHTRGLDFACDIYIWAPLAGTCGVLLLSLVITLYCKRG RKKLLYIFKQPFMRP VQTTQEEDGC SCRFPEEEEGGCELRVKF SRS ADAP AYKQGQNQLYN ELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLYNELQKDKMAEAYSEIGMKGERR RGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 1008).

[0226] In some embodiments, the anti-CD22 CAR has the sequenceQVQLQQSGPGLVKPSQTLSLTCAISGDSVSSNSAAWNWIRQSPSRGLEWLGRTYYRSKWYN DYAVSVKSRITINPDTSKNQFSLQLNSVTPEDTAVYYCAREVTGDLEDAFDIWGQGTMVTV SSGGGGSDIQMTQSPSSLSASVGDRVTITCRASQTIWSYLNWYQQRPGKAPNLLIYAASSLQ SGVPSRFSGRGSGTDFTLTISSLQAEDFATYYCQQSYSIPQTFGQGTKLEIKAAATTTPAPRPP TPAPTIASQPLSLRPEACRPAAGGAVHTRGLDFACDIYIWAPLAGTCGVLLLSLVITLYCKRG RKKLLYIFKQPFMRP VQTTQEEDGC SCRFPEEEEGGCELRVKF SRS ADAP AYKQGQNQLYN ELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLYNELQKDKMAEAYSEIGMKGERR RGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 1008).

[0227] In some embodiments, the anti-CD22 CAR further comprises a signal peptide.

[0228] In some embodiments, the signal peptide sequence is derived from CD8a or granulocyte macrophage colony stimulating factor receptor alpha (GMCSFRa).

[0229] In some embodiments, the signal peptide sequence is derived from GMCSFRa and has the sequence of MLLLVTSLLLCELPHPAFLLIP (SEQ ID NO: 691).

[0230] In some embodiments, the anti-CD22 CAR comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the sequenceMLLLVTSLLLCELPHPAFLLIPQVQLQQSGPGLVKPSQTLSLTCAISGDSVSSNSAAWNWIRQ SPSRGLEWLGRTYYRSKWYNDYAVSVKSRITINPDTSKNQFSLQLNSVTPEDTAVYYCARE VTGDLEDAFDIWGQGTMVTVSSGGGGSDIQMTQSPSSLSASVGDRVTITCRASQTIWSYLN WYQQRPGKAPNLLIYAASSLQSGVPSRFSGRGSGTDFTLTISSLQAEDFATYYCQQSYSIPQT FGQGTKLEIKAAATTTPAPRPPTPAPTIASQPLSLRPEACRPAAGGAVHTRGLDFACDIYIWA PLAGTCGVLLLSLVITLYCKRGRKKLLYIFKQPFMRPVQTTQEEDGCSCRFPEEEEGGCELR VKFSRSADAPAYKQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLYN ELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 996).

[0231] In some embodiments, the anti-CD19 CAR binding domain comprises an anti-CD19 antibody or an antigen-binding fragment thereof.

[0232] In some embodiments, the anti-CD19 CAR binding domain is an antigen-binding fragment of an anti-CD19 antibody.

[0233] In some embodiments, the antigen-binding fragment of an anti-CD19 antibody is a single chain variable fragment (scFv).

[0234] In some embodiments, the anti-CD19 binding domain comprises a heavy chain variable region (VH) that comprises a first heavy chain complementarity determining region (HCDR1) having the sequence GYRFTNYW (SEQ ID NO: 173), a second heavy chain complementarity determining region (HCDR2) having the sequence IDPSDSYT (SEQ ID NO: 174), and a third heavy chain complementarity determining region (HCDR3) having the sequence ARPGDILTGWAMDV (SEQ ID NO: 175).

[0235] In some embodiments, the anti-CD19 binding domain comprises a light chain variable region (VL) that comprises a first light chain complementarity determining region (LCDR1) having the sequence SSNIGNNY (SEQ ID NO: 297), a second light chain complementarity determining region (LCDR2) having the sequence DNN (SEQ ID NO: 298), and a third light chain complementarity determining region (LCDR3) having the sequence QSYDSSLSGNYV (SEQ ID NO: 299).

[0236] In some embodiments, the anti-CD19 binding domain comprises a VH comprising a sequence having at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the sequence EVQLVQSGAEVKKPGESLKISCKASGYRFTNYWIAWVRQRPGKGLEWMGRIDPSDSYTHY SPSFQGHVTMSTDKSISTAYLQWSSLKASDTAMYYCARPGDILTGWAMDVWGQGTLVTVS S (SEQ ID NO: 47) and a VL comprising a sequence having at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the sequence QSVLTQPPSVSAAPGQKVTISCSGSSSNIGNNYVSWYQQLPGTAPKLLIYDNNKRPSGIPDRF SGSKSGTSATLGITGLQAEDEADYYCQSYDSSLSGNYVFGTGTKVTVLK (SEQ ID NO: 48).

[0237] In some embodiments, the anti-CD19 binding domain comprises a VH having the sequence EVQLVQSGAEVKKPGESLKISCKASGYRFTNYWIAWVRQRPGKGLEWMGRIDPSDSYTHY SPSFQGHVTMSTDKSISTAYLQWSSLKASDTAMYYCARPGDILTGWAMDVWGQGTLVTVS S (SEQ ID NO: 47) and a VL having the sequenceQSVLTQPPSVSAAPGQKVTISCSGSSSNIGNNYVSWYQQLPGTAPKLLIYDNNKRPSGIPDRF SGSKSGTSATLGITGLQAEDEADYYCQSYDSSLSGNYVFGTGTKVTVLK (SEQ ID NO: 48).

[0238] In some embodiments, the VH and VL are linked by a linker comprising the sequence GGGGSGGGGSGGGGS (SEQ ID NO: 995).

[0239] In some embodiments, the anti-CD19 binding domain comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the sequence EVQLVQSGAEVKKPGESLKISCKASGYRFTNYWIAWVRQRPGKGLEWMGRIDPSDSYTHY SPSFQGHVTMSTDKSISTAYLQWSSLKASDTAMYYCARPGDILTGWAMDVWGQGTLVTVS SAAASGGGGSGGGGSGGGGSALQSVLTQPPSVSAAPGQKVTISCSGSSSNIGNNYVSWYQQ LPGTAPKLLIYDNNKRPSGIPDRFSGSKSGTSATLGITGLQAEDEADYYCQSYDSSLSGNYVF GTGTKVTVL (SEQ ID NO: 11).

[0240] In some embodiments, the anti-CD19 binding domain comprises an scFv having the sequence EVQLVQSGAEVKKPGESLKISCKASGYRFTNYWIAWVRQRPGKGLEWMGRIDPSDSYTHY SPSFQGHVTMSTDKSISTAYLQWSSLKASDTAMYYCARPGDILTGWAMDVWGQGTLVTVS SAAASGGGGSGGGGSGGGGSALQSVLTQPPSVSAAPGQKVTISCSGSSSNIGNNYVSWYQQ LPGTAPKLLIYDNNKRPSGIPDRFSGSKSGTSATLGITGLQAEDEADYYCQSYDSSLSGNYVF GTGTKVTVL (SEQ ID NO: 11).

[0241] In some embodiments, the anti-CD19 CAR further comprises a hinge domain.

[0242] In some embodiments, the hinge domain of the anti-CD19 CAR is derived from CD8a or from CD28 and the transmembrane domain of the anti-CD19 CAR is derived from CD8a or from CD28.

[0243] In some embodiments, the hinge domain of the anti-CD19 CAR is derived from CD28 and the transmembrane domain is derived from CD28.

[0244] In some embodiments, the CD28 hinge domain has the sequenceIEVMYPPPYLDNEKSNGTIIHVKGKHLCPSPLFPGPSKP (SEQ ID NO: 682).

[0245] In some embodiments, the CD28 transmembrane domain has the sequence FWVLVVVGGVLACYSLLVTVAFIIFWV (SEQ ID NO: 683).

[0246] In some embodiments, the cytoplasmic domain of the anti-CD19 CAR comprises an intracellular co-stimulatory domain derived from a molecule selected from the group consisting of CD27, CD28, 4-1BB / CD137, 0X40, ICOS, and CD2.

[0247] In some embodiments, the intracellular co-stimulatory domain is derived from CD28.

[0248] In some embodiments, the CD28 intracellular co-stimulatory domain comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the sequence RSKRSRLLHSDYMNMTPRRPGPTRKHYQPYAPPRDFAAYRS (SEQ ID NO: 684).

[0249] In some embodiments, the CD28 intracellular co-stimulatory domain has the sequence RSKRSRLLHSDYMNMTPRRPGPTRKHYQPYAPPRDFAAYRS (SEQ ID NO: 684).

[0250] In some embodiments, the cytoplasmic domain of the anti-CD19 CAR comprises an intracellular signaling domain comprising one or more ITAMs derived from CD3(^.

[0251] In some embodiments, the CD3(^ intracellular signaling domain comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the sequence RVKFSRSADAPAYKQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLY NELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 686).

[0252] In some embodiments, the CD3(^ intracellular signaling domain has the sequence RVKFSRSADAPAYKQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLY NELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 686).

[0253] In some embodiments, the CD3(^ intracellular signaling domain comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the sequence RVKFSRSADAPAYQQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLY NELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 687).

[0254] In some embodiments, the CD3(^ intracellular signaling domain has the sequence RVKFSRSADAPAYQQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLY NELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 687).

[0255] In some embodiments, the CD3(^ intracellular signaling domain comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the sequence RVKFSRSADAPAYQQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPQRRKNPQEGL YNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 688).

[0256] In some embodiments, the CD3(^ intracellular signaling domain has the sequence RVKFSRSADAPAYQQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPQRRKNPQEGL YNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 688).

[0257] In some embodiments, the CD3(^ intracellular signaling domain comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the sequence RVKFSRSADAPAYKQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPQRRKNPQEGLYNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 689).

[0258] In some embodiments, the CD3(^ intracellular signaling domain has the sequence RVKFSRSADAPAYKQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPQRRKNPQEGL YNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 689).

[0259] In some embodiments, the anti-CD19 CAR comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the sequence EVQLVQSGAEVKKPGESLKISCKASGYRFTNYWIAWVRQRPGKGLEWMGRIDPSDSYTHY SPSFQGHVTMSTDKSISTAYLQWSSLKASDTAMYYCARPGDILTGWAMDVWGQGTLVTVS SAAASGGGGSGGGGSGGGGSALQSVLTQPPSVSAAPGQKVTISCSGSSSNIGNNYVSWYQQ LPGTAPKLLIYDNNKRPSGIPDRFSGSKSGTSATLGITGLQAEDEADYYCQSYDSSLSGNYVF GTGTKVTVLIEVMYPPPYLDNEKSNGTIIHVKGKHLCPSPLFPGPSKPFWVLVVVGGVLACY SLLVTVAFIIFWVRSKRSRLLHSDYMNMTPRRPGPTRKHYQPYAPPRDFAAYRSRVKFSRSA DAPAYKQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLYNELQKDKM AEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 1121).

[0260] In some embodiments, the anti-CD19 CAR has the sequence EVQLVQSGAEVKKPGESLKISCKASGYRFTNYWIAWVRQRPGKGLEWMGRIDPSDSYTHY SPSFQGHVTMSTDKSISTAYLQWSSLKASDTAMYYCARPGDILTGWAMDVWGQGTLVTVS SAAASGGGGSGGGGSGGGGSALQSVLTQPPSVSAAPGQKVTISCSGSSSNIGNNYVSWYQQ LPGTAPKLLIYDNNKRPSGIPDRFSGSKSGTSATLGITGLQAEDEADYYCQSYDSSLSGNYVF GTGTKVTVLIEVMYPPPYLDNEKSNGTIIHVKGKHLCPSPLFPGPSKPFWVLVVVGGVLACY SLLVTVAFIIFWVRSKRSRLLHSDYMNMTPRRPGPTRKHYQPYAPPRDFAAYRSRVKFSRSA DAPAYKQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLYNELQKDKM AEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 1121).

[0261] In some embodiments, the anti-CD19 CAR further comprises a signal peptide.

[0262] In some embodiments, the signal peptide sequence is derived from CD8a or granulocyte macrophage colony stimulating factor receptor alpha (GMCSFRa).

[0263] In some embodiments, the signal peptide sequence is derived from CD8a and has the sequence of MALPVTALLLPLALLLHAARP (SEQ ID NO: 690).

[0264] In some embodiments, the anti-CD19 CAR comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the sequence MALPVTALLLPLALLLHAARPEVQLVQSGAEVKKPGESLKISCKASGYRFTNYWIAWVRQRPGKGLEWMGRIDPSDSYTHYSPSFQGHVTMSTDKSISTAYLQWSSLKASDTAMYYCARPGD ILTGWAMDVWGQGTLVTVSSAAASGGGGSGGGGSGGGGSALQSVLTQPPSVSAAPGQKVT ISCSGSSSNIGNNYVSWYQQLPGTAPKLLIYDNNKRPSGIPDRFSGSKSGTSATLGITGLQAED EADYYCQSYDSSLSGNYVFGTGTKVTVLIEVMYPPPYLDNEKSNGTIIHVKGKHLCPSPLFP GPSKPFWVLVVVGGVLACYSLLVTVAFIIFWVRSKRSRLLHSDYMNMTPRRPGPTRKHYQP YAPPRDFAAYRSRVKFSRSADAPAYKQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGG KPRRKNPQEGLYNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHM QALPPR (SEQ ID NO: 706).

[0265] In some embodiments, the anti-CD20 CAR binding domain comprises an anti-CD20 antibody or an antigen-binding fragment thereof.

[0266] In some embodiments, the anti-CD20 CAR binding domain is an antigen-binding fragment of an anti-CD20 antibody.

[0267] In some embodiments, the antigen-binding fragment of an anti-CD20 antibody is a single chain variable fragment (scFv).

[0268] In some embodiments, the anti-CD20 binding domain comprises a heavy chain variable region (VH) that comprises a first heavy chain complementarity determining region (HCDR1) having the sequence GFTFGDYG (SEQ ID NO: 562), a second heavy chain complementarity determining region (HCDR2) having the sequence INWNGGST (SEQ ID NO: 513), and a third heavy chain complementarity determining region (HCDR3) having the sequence ARKSYYGSGSPDVFDI (SEQ ID NO: 563).

[0269] In some embodiments, the anti-CD20 binding domain comprises a light chain variable region (VL) that comprises a first light chain complementarity determining region (LCDR1) having the sequence QSVLYSSNNKNY (SEQ ID NO: 635), a second light chain complementarity determining region (LCDR2) having the sequence WAS (SEQ ID NO: 636), and a third light chain complementarity determining region (LCDR3) having the sequence QQYYSFYQT (SEQ ID NO: 629).

[0270] In some embodiments, the anti-CD20 binding domain comprises a VH comprising a sequence having at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the sequence EVQLVESGGGVVRPGGSLRLSCTASGFTFGDYGMSWVRQAPGKGLEWVSGINWNGGSTG YADSVKGRFTISRDNAKNSLYLQMNSLRAEDTALYYCARKSYYGSGSPDVFDIWGQGTMV TVSS (SEQ ID NO: 425) and a VL comprising a sequence having at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the sequenceDIVMTQSPDSLAVSLGERATINCKSSQSVLYSSNNKNYLAWYQQKPGQPPKLLIYWASTRES GVPDRFSGSGSGSDFTLTISSLQAEDVAVYYCQQYYSFYQTFGQGTKVEIK (SEQ ID NO: 426).

[0271] In some embodiments, the anti-CD20 binding domain comprises a VH having the sequence EVQLVESGGGVVRPGGSLRLSCTASGFTFGDYGMSWVRQAPGKGLEWVSGINWNGGSTG YADSVKGRFTISRDNAKNSLYLQMNSLRAEDTALYYCARKSYYGSGSPDVFDIWGQGTMV TVSS (SEQ ID NO: 425) and a VL having the sequence DIVMTQSPDSLAVSLGERATINCKSSQSVLYSSNNKNYLAWYQQKPGQPPKLLIYWASTRES GVPDRFSGSGSGSDFTLTISSLQAEDVAVYYCQQYYSFYQTFGQGTKVEIK (SEQ ID NO: 426).

[0272] In some embodiments, the VH and VL are linked by linker comprising the sequence GGGGSGGGGSGGGGS (SEQ ID NO: 995).

[0273] In some embodiments, the anti-CD20 binding domain comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the sequenceDIVMTQSPDSLAVSLGERATINCKSSQSVLYSSNNKNYLAWYQQKPGQPPKLLIYWASTRES GVPDRFSGSGSGSDFTLTISSLQAEDVAVYYCQQYYSFYQTFGQGTKVEIKGGGGSGGGGS GGGGSEVQLVESGGGVVRPGGSLRLSCTASGFTFGDYGMSWVRQAPGKGLEWVSGINWN GGSTGYADSVKGRFTISRDNAKNSLYLQMNSLRAEDTALYYCARKSYYGSGSPDVFDIWG QGTMVTVSS (SEQ ID NO: 363).

[0274] In some embodiments, the anti-CD20 binding domain comprises an scFv having the sequence DIVMTQSPDSLAVSLGERATINCKSSQSVLYSSNNKNYLAWYQQKPGQPPKLLIYWASTRES GVPDRFSGSGSGSDFTLTISSLQAEDVAVYYCQQYYSFYQTFGQGTKVEIKGGGGSGGGGS GGGGSEVQLVESGGGVVRPGGSLRLSCTASGFTFGDYGMSWVRQAPGKGLEWVSGINWN GGSTGYADSVKGRFTISRDNAKNSLYLQMNSLRAEDTALYYCARKSYYGSGSPDVFDIWG QGTMVTVSS (SEQ ID NO: 363).

[0275] In some embodiments, the anti-CD20 CAR further comprises a hinge domain.

[0276] In some embodiments, the hinge domain of the anti-CD20 CAR is derived from CD8a or from CD28 and the transmembrane domain of the anti-CD20 CAR is derived from CD8a or from CD28.

[0277] In some embodiments, the hinge domain of the anti-CD20 CAR is derived from CD28 and the transmembrane domain is derived from CD28.

[0278] In some embodiments, the CD28 hinge domain has the sequenceIEVMYPPPYLDNEKSNGTIIHVKGKHLCPSPLFPGPSKP (SEQ ID NO: 682).

[0279] In some embodiments, the CD28 transmembrane domain has the sequence FWVLVVVGGVLACYSLLVTVAFIIFWV (SEQ ID NO: 683).

[0280] In some embodiments, the cytoplasmic domain of the anti-CD20 CAR comprises an intracellular co-stimulatory domain derived from a molecule selected from the group consisting of CD27, CD28, 4-1BB / CD137, 0X40, ICOS, and CD2.

[0281] In some embodiments, the intracellular co-stimulatory domain is derived from CD2.

[0282] In some embodiments, the CD2 intracellular co-stimulatory domain comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the sequence KRKKQRSRRNDEELETRAHRVATEERGRKPHQIPASTPQNPATSQHPPPPPGHRSQAPSHRP PPPGHRVQHQPQKRPPAPSGTQVHQQKGPPLPRPRVQPKPPHGAAENSLSPSSN (SEQ ID NO: 685).

[0283] In some embodiments, the CD2 intracellular co-stimulatory domain has the sequence KRKKQRSRRNDEELETRAHRVATEERGRKPHQIPASTPQNPATSQHPPPPPGHRSQAPSHRP PPPGHRVQHQPQKRPPAPSGTQVHQQKGPPLPRPRVQPKPPHGAAENSLSPSSN (SEQ ID NO: 685).

[0284] In some embodiments, the cytoplasmic domain of the anti-CD20 CAR comprises an intracellular signaling domain comprising one or more ITAMs derived from CD3(^.

[0285] In some embodiments, the CD3(^ intracellular signaling domain comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the sequence RVKFSRSADAPAYKQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLY NELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 686).

[0286] In some embodiments, the CD3(^ intracellular signaling domain has the sequence RVKFSRSADAPAYKQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLY NELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 686).

[0287] In some embodiments, the CD3(^ intracellular signaling domain comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the sequence RVKFSRSADAPAYQQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLY NELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 687).

[0288] In some embodiments, the CD3(^ intracellular signaling domain has the sequence RVKFSRSADAPAYQQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLYNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 687).

[0289] In some embodiments, the CD3(^ intracellular signaling domain comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the sequence RVKFSRSADAPAYQQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPQRRKNPQEGL YNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 688).

[0290] In some embodiments, the CD3(^ intracellular signaling domain has the sequence RVKFSRSADAPAYQQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPQRRKNPQEGL YNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 688).

[0291] In some embodiments, the CD3(^ intracellular signaling domain comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the sequence RVKFSRSADAPAYKQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPQRRKNPQEGL YNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 689).

[0292] In some embodiments, the CD3(^ intracellular signaling domain has the sequence RVKFSRSADAPAYKQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPQRRKNPQEGL YNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 689).

[0293] In some embodiments, the anti-CD20 CAR comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the sequence MLLLVTSLLLCELPHPAFLLIPDIVMTQSPDSLAVSLGERATINCKSSQSVLYSSNNKNYLAW YQQKPGQPPKLLIYWASTRESGVPDRFSGSGSGSDFTLTISSLQAEDVAVYYCQQYYSFYQT FGQGTKVEIKGGGGSGGGGSGGGGSEVQLVESGGGVVRPGGSLRLSCTASGFTFGDYGMS WVRQAPGKGLEWVSGINWNGGSTGYADSVKGRFTISRDNAKNSLYLQMNSLRAEDTALY YC ARKS YYGSGSPDVFDIWGQGTMVTVS SIEVMYPPPYLDNEKSNGTIIHVKGKHLCP SPLF PGPSKPFWVLVVVGGVLACYSLLVTVAFIIFWVKRKKQRSRRNDEELETRAHRVATEERGR KPHQIPASTPQNPATSQHPPPPPGHRSQAPSHRPPPPGHRVQHQPQKRPPAPSGTQVHQQKGP PLPRPRVQPKPPHGAAENSLSPSSNRVKFSRSADAPAYKQGQNQLYNELNLGRREEYDVLD KRRGRDPEMGGKPRRKNPQEGLYNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLST ATKDTYDALHMQALPPR (SEQ ID NO: 1358).

[0294] In some embodiments, the anti-CD20 CAR has the sequenceMLLLVTSLLLCELPHPAFLLIPDIVMTQSPDSLAVSLGERATINCKSSQSVLYSSNNKNYLAW YQQKPGQPPKLLIYWASTRESGVPDRFSGSGSGSDFTLTISSLQAEDVAVYYCQQYYSFYQT FGQGTKVEIKGGGGSGGGGSGGGGSEVQLVESGGGVVRPGGSLRLSCTASGFTFGDYGMS WVRQAPGKGLEWVSGINWNGGSTGYADSVKGRFTISRDNAKNSLYLQMNSLRAEDTALY YC ARKS YYGSGSPDVFDIWGQGTMVTVS SIEVMYPPPYLDNEKSNGTIIHVKGKHLCP SPLF PGPSKPFWVLVVVGGVLACYSLLVTVAFIIFWVKRKKQRSRRNDEELETRAHRVATEERGR KPHQIPASTPQNPATSQHPPPPPGHRSQAPSHRPPPPGHRVQHQPQKRPPAPSGTQVHQQKGP PLPRPRVQPKPPHGAAENSLSPSSNRVKFSRSADAPAYKQGQNQLYNELNLGRREEYDVLD KRRGRDPEMGGKPRRKNPQEGLYNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLST ATKDTYDALHMQALPPR (SEQ ID NO: 1358)

[0295] In some embodiments, the anti-CD20 CAR further comprises a signal peptide.

[0296] In some embodiments, the signal peptide sequence is derived from CD8D or granulocyte macrophage colony stimulating factor receptor alpha (GMCSFRD).

[0297] In some embodiments, the signal peptide sequence is derived from GMCSFRD and has the sequence of MLLLVTSLLLCELPHPAFLLIP (SEQ ID NO: 691).

[0298] In some embodiments, the anti-CD20 CAR comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the sequence MLLLVTSLLLCELPHPAFLLIPDYKDDDDKDIVMTQSPDSLAVSLGERATINCKSSQSVLYSS NNKNYLAWYQQKPGQPPKLLIYWASTRESGVPDRF SGSGSGSDFTLTIS SLQAED VAVYYC QQYYSFYQTFGQGTKVEIKGGGGSGGGGSGGGGSEVQLVESGGGVVRPGGSLRLSCTASGF TFGDYGMSWVRQAPGKGLEWVSGINWNGGSTGYADSVKGRFTISRDNAKNSLYLQMNSL RAEDTALYYCARKSYYGSGSPDVFDIWGQGTMVTVSSIEVMYPPPYLDNEKSNGTIIHVKG KHLCPSPLFPGPSKPFWVLVVVGGVLACYSLLVTVAFIIFWVKRKKQRSRRNDEELETRAHR VATEERGRKPHQIPASTPQNPATSQHPPPPPGHRSQAPSHRPPPPGHRVQHQPQKRPPAPSGT QVHQQKGPPLPRPRVQPKPPHGAAENSLSPSSNRVKFSRSADAPAYKQGQNQLYNELNLGR REEYDVLDKRRGRDPEMGGKPRRKNPQEGLYNELQKDKMAEAYSEIGMKGERRRGKGHD GLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 772).

[0299] In some embodiments, the one or more recombinant polynucleic acid molecules comprises a sequence encoding the amino acid sequence of

[0300] MLLLVTSLLLCELPHPAFLLIPQVQLQQSGPGLVKPSQTLSLTCAISGDSVSSNSAAW NWIRQSPSRGLEWLGRTYYRSKWYNDYAVSVKSRITINPDTSKNQFSLQLNSVTPEDTAVY YCAREVTGDLEDAFDIWGQGTMVTVSSGGGGSDIQMTQSPSSLSASVGDRVTITCRASQTIWS YLNWYQQRPGKAPNLLIYAAS SLQSGVPSRF SGRGSGTDFTLTIS SLQAEDF ATYYCQQ S YSIPQTFGQGTKLEIKAAATTTPAPRPPTPAPTIASQPLSLRPEACRPAAGGAVHTRGLDFAC DIYIWAPLAGTCGVLLLSLVITLYCKRGRKKLLYIFKQPFMRPVQTTQEEDGCSCRFPEEEEG GCELRVKFSRSADAPAYKQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQE GLYNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPRATN FSLLKQAGDVEENPGPMALPVTALLLPLALLLHAARPEVQLVQSGAEVKKPGESLKISCKAS GYRFTNYWIAWVRQRPGKGLEWMGRIDPSDSYTHYSPSFQGHVTMSTDKSISTAYLQWSSL KASDTAMYYCARPGDILTGWAMDVWGQGTLVTVSSAAASGGGGSGGGGSGGGGSALQS VLTQPPSVSAAPGQKVTISCSGSSSNIGNNYVSWYQQLPGTAPKLLIYDNNKRPSGIPDRFSG SKSGTSATLGITGLQAEDEADYYCQSYDSSLSGNYVFGTGTKVTVLIEVMYPPPYLDNEKSN GTIIHVKGKHLCPSPLFPGPSKPFWVLVVVGGVLACYSLLVTVAFIIFWVRSKRSRLLHSDY MNMTPRRPGPTRKHYQPYAPPRDFAAYRSRVKFSRSADAPAYKQGQNQLYNELNLGRREE YDVLDKRRGRDPEMGGKPRRKNPQEGLYNELQKDKMAEAYSEIGMKGERRRGKGHDGLY QGLSTATKDTYDALHMQALPPREGRGSLLTCGDVEENPGPMLLLVTSLLLCELPHPAFLLIP DIVMTQSPDSLAVSLGERATINCKSSQSVLYSSNNKNYLAWYQQKPGQPPKLLIYWASTRES GVPDRFSGSGSGSDFTLTISSLQAEDVAVYYCQQYYSFYQTFGQGTKVEIKGGGGSGGGGS GGGGSEVQLVESGGGVVRPGGSLRLSCTASGFTFGDYGMSWVRQAPGKGLEWVSGINWN GGSTGYADSVKGRFTISRDNAKNSLYLQMNSLRAEDTALYYCARKSYYGSGSPDVFDIWG QGTMVTVSSIEVMYPPPYLDNEKSNGTIIHVKGKHLCPSPLFPGPSKPFWVLVVVGGVLACY SLLVTVAFIIFWVKRKKQRSRRNDEELETRAHRVATEERGRKPHQIPASTPQNPATSQHPPPP PGHRSQAPSHRPPPPGHRVQHQPQKRPPAPSGTQVHQQKGPPLPRPRVQPKPPHGAAENSLS PSSNRVKFSRSADAPAYKQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQE GLYNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 1006)

[0301] In some embodiments, the one or more recombinant polynucleic acid molecules has the sequence atgcttctgctcgtgacaagcctgctgctgtgcgagctgccccaccctgcctttctgctgatccctCAAGTCCAGCTGCAGCAAT CTGGCCCTGGCCTGGTGAAGCCTAGCCAGACCCTGAGCCTGACATGTGCCATCAGCGGA GATTCTGTTAGCAGCAACAGCGCGGCCTGGAACTGGATCAGACAGAGCCCTAGCAGAG GACTGGAGTGGCTGGGCCGGACCTACTACAGAAGCAAGTGGTACAACGACTACGCCGT GTCCGTGAAAAGCAGAATCACCATCAACCCCGACACCAGCAAGAACCAGTTCTCCCTGC AACTGAATAGCGTCACACCCGAGGATACAGCCGTGTACTACTGCGCCAGAGAGGTTAC AGGCGACCTGGAAGATGCCTTTGACATCTGGGGCCAGGGCACGATGGTGACCGTCAGTAGCGGCGGCGGAGGAAGCGACATCCAGATGACCCAATCTCCTAGCAGCCTGTCTGCCTCTGTGGGCGACAGAGTGACCATCACCTGCCGCGCCTCTCAAACAATCTGGTCCTACCTGAACTGGTATCAGCAAAGACCTGGCAAAGCCCCAAACCTGCTGATCTACGCCGCCAGCTCCCTGCAAAGCGGAGTGCCCAGCAGATTCAGCGGCCGGGGCAGCGGCACTGATTTTACCCTGACTATTTCTTCCCTGCAAGCCGAGGACTTCGCCACATACTACTGTCAGCAGAGCTACAGCATCCCTCAGACCTTCGGCCAGGGCACCAAGCTGGAAATCAAGGCGGCCGCCACAACCACCCCTGCTCCCCGCCCACCTACCCCTGCTCCTACAATCGCCTCGCAACCTCTGAGCCTGAGACCTGAGGCCTGTAGACCCGCCGCTGGCGGCGCCGTGCACACCAGAGGCCTGGACTTCGCCTGCGACATCTACATCTGGGCCCCACTGGCCGGCACATGCGGCGTGCTGCTGCTGTCCCTGGTGATCACCCTGTATTGCAAGAGAGGCAGAAAGAAACTGCTGTACATCTTCAAGCAGCCTTTCATGCGGCCCGTGCAGACCACCCAGGAGGAAGATGGTTGCTCATGCCGGTTCCCCGAGGAAGAGGAAGGCGGATGTGAACTGAGAGTGAAGTTCAGCCGCTCCGCTGACGCACCCGCCTATAAACAGGGGCAGAATCAACTTTATAACGAACTGAACCTTGGAAGAAGGGAGGAATATGACGTGCTTGATAAGAGAAGAGGCAGAGATCCTGAAATGGGAGGGAAACCTCGGAGAAAAAATCCACAAGAGGGGCTCTACAATGAACTCCAGAAAGACAAGATGGCTGAAGCTTACTCTGAGATTGGAATGAAAGGGGAGCGGCGCCGGGGCAAAGGGCACGACGGATTGTACCAAGGACTCAGTACAGCTACTAAAGACACATATGACGCCCTGCATATGCAGGCACTGCCCCCACGGGCCACCAACTTCAGCCTGCTGAAGCAGGCCGGCGACGTGGAAGAGAACCCCGGCCCTATGGCCTTGCCCGTGACTGCTCTGCTCCTGCCCCTCGCACTGCTGCTGCACGCCGCGAGGCCCGAAGTGCAGCTGGTCCAATCTGGCGCCGAGGTGAAGAAGCCCGGCGAGAGCCTGAAGATCAGCTGTAAAGCCAGCGGATATAGATTCACCAACTACTGGATCGCCTGGGTGCGGCAGCGGCCTGGCAAGGGCCTGGAATGGATGGGCAGAATCGACCCATCTGATAGCTACACCCACTACAGCCCTAGCTTCCAAGGCCACGTGACCATGAGCACAGATAAGAGCATCAGCACCGCCTACCTGCAGTGGTCCTCCCTGAAAGCTAGCGACACCGCTATGTACTACTGCGCCAGACCTGGCGACATCCTGACCGGCTGGGCTATGGATGTGTGGGGCCAGGGAACACTGGTGACAGTGTCCAGCGCTGCAGCAAGTGGAGGCGGAGGAAGTGGAGGCGGTGGTTCAGGAGGAGGAGGATCTGCTCTGCAATCTGTGCTCACACAGCCCCCCAGCGTGTCTGCCGCTCCTGGACAGAAAGTGACCATCAGCTGCAGCGGATCTAGCTCCAACATCGGCAACAACTACGTGTCCTGGTATCAGCAGCTGCCTGGCACCGCCCCTAAGCTGCTGATCTACGACAACAACAAGCGGCCATCTGGCATCCCTGATCGCTTCAGCGGCAGCAAGTCCGGCACCAGCGCCACCCTGGGAATCACCGGCCTGCAGGCCGAGGACGAAGCCGACTACTACTGTCAGAGCTACGATAGCAGCCTGAGCGGCAATTACGTGTTCGGCACAGGCACAAAGGTCACCGTGCTGATCGAGGTGATGTACCCTCCTCCTTACCTGGATAATGAGAAGTCCAACGGAACAATCATCCACGTGAAGGGCAAGCACCTGTGCCCCTCTCCTCTGTTTCCGGGCCCTTCTAAGCCCTTCTGGGTGCTGGTCGTGGTGGGCGGAGTGCTCGCTTGTTACTCTCTGCTTGTGACCGTGGCCTTCATTATCTTCTGGGTCCGCAGCAAGCGGTCCAGACTGCTGCACAGCGACTACATGAACATGACCCCTCGGAGACCTGGACCCACCAGAAAGCACTACCAGCCTTACGCCCCTCCACGCGACTTCGCCGCTTATCGCAGCCGCGTTAAATTTTCACGCAGCGCGGACGCCCCTGCGTACAAGCAGGGACAGAACCAGCTGTACAACGAGCTCAACCTGGGCCGCCGGGAGGAGTACGACGTGCTGGACAAGCGCCGGGGGCGTGATCCAGAGATGGGCGGCAAACCCCGCCGCAAGAATCCTCAGGAGGGCTTATACAACGAGCTGCAGAAGGACAAAATGGCCGAGGCCTACAGCGAGATCGGCATGAAGGGAGAGCGCCGGCGTGGAAAGGGCCATGACGGCCTCTATCAGGGTCTGTCCACCGCCACCAAGGACACCTACGACGCTCTACACATGCAGGCCTTGCCTCCGCGCGAGGGCAGAGGCAGCCTGCTGACCTGCGGCGATGTGGAGGAGAACCCCGGACCTATGCTGTTGCTTGTTACAAGCCTCCTCCTGTGCGAATTGCCTCACCCCGCATTTCTCCTGATACCCGACATCGTGATGACCCAGTCCCCAGACAGCCTGGCTGTGTCCCTGGGCGAACGGGCCACCATCAACTGCAAGAGCAGCCAGAGCGTGCTGTACAGCAGCAACAACAAGAATTACCTGGCCTGGTACCAGCAGAAACCTGGCCAGCCTCCCAAGCTGCTGATCTACTGGGCCAGCACCAGAGAGAGCGGAGTGCCTGATAGATTCAGCGGCAGCGGATCTGGCTCTGATTTTACCCTGACAATCAGCAGCCTCCAGGCCGAGGACGTCGCCGTGTACTATTGTCAGCAATACTACTCCTTCTACCAAACATTCGGCCAGGGCACCAAAGTTGAAATCAAGGGAGGAGGAGGGTCTGGAGGTGGAGGAAGTGGCGGAGGGGGAAGTGAGGTGCAGCTGGTCGAGAGCGGCGGAGGCGTGGTGCGGCCTGGAGGATCTCTCAGACTGAGCTGTACCGCCAGCGGATTTACCTTCGGCGACTACGGCATGAGCTGGGTGCGCCAGGCCCCTGGCAAAGGCCTGGAATGGGTTTCCGGCATCAACTGGAACGGCGGCTCTACAGGCTACGCCGACAGCGTGAAGGGCAGATTCACCATCAGCCGGGACAACGCCAAGAATAGCCTGTACCTGCAAATGAACAGCCTGAGAGCTGAAGATACAGCTCTGTATTACTGCGCCAGAAAGTCCTACTACGGCTCTGGCAGCCCCGATGTGTTCGACATCTGGGGCCAGGGCACAATGGTGACCGTGTCCAGCATAGAAGTAATGTACCCTCCGCCTTATCTCGATAATGAAAAAAGCAACGGGACAATTATTCATGTGAAGGGCAAGCATCTTTGTCCCTCACCGCTTTTCCCTGGACCATCAAAACCATTTTGGGTGCTCGTCGTCGTGGGAGGGGTCCTCGCCTGCTATTCTCTGTTGGTGACTGTAGCCTTTATCATTTTTTGGGTTAAACGGAAGAAGCAGAGATCTAGACGGAACGACGAGGAACTGGAAACCCGCGCtCACAGGGTGGCCACCGAGGAAAGAGGCAGAAAGCCACACCAGATCCCCGCCTCAACACCTCAGAATCCCGCCACAAGCCAGCACCCCCCACCTCCTCCTGGCCATCGCAGCCAGGCCCCTAGCCACCGGCCTCCACCTCCCGGTCACAGAGTGCAGCACCAACCTCAAAAACGGCCCCCCGCTCCAAGCGGCACCCAAGTTCACCAGCAGAAGGGCCCTCCTCTGCCTAGACCTAGAGTCCAGCCTAAGCCTCCT CACGGCGCTGCTGAGAACAGCCTGAGCCCTTCTTCgAATCGGGTCAAATTCAGTCGCTCT GCCGATGCTCCAGCCTACAAACAAGGCCAAAACCAATTATACAATGAACTTAATCTCGG GCGCAGGGAAGAGTACGATGTTCTCGATAAAAGGCGGGGACGCGACCCCGAAATGGGT GGAAAGCCAAGAAGGAAGAACCCGCAGGAAGGACTGTATAATGAGTTACAAAAGGAT AAGATGGCAGAGGCATATTCAGAAATCGGGATGAAGGGCGAAAGAAGAAGAGGCAAG GGACACGATGGGCTTTATCAAGGCTTAAGCACTGCAACAAAGGATACTTATGATGCACT CCATATGCAAGCTCTCCCACCTAGA (SEQ ID NO: 1118)

[0302] Also provided herein is a composition comprising one or more polypeptides encoded by the one or more recombinant polynucleic acid molecules of the composition of the present disclosure.

[0303] Also provided herein is a cell comprising the one or more recombinant polynucleic acid molecules of the composition of the present disclosure or the one or more polypeptides of the present disclosure.

[0304] In some embodiments, the cell is an immune cell.

[0305] In some embodiments, the immune cell is a T cell, a B cell, a natural killer (NK) cell, a macrophage, a dendritic cell, a monocyte, or a granulocyte.

[0306] In some embodiments, the cell is a T cell.

[0307] In some embodiments, the T cell is a CD8-positive T cell, a CD4-positive T cell, a regulatory T cell (Treg), a cytotoxic T cell (CTL), a central memory T cell (TCM), an effector memory T cell (TEM), a tissue-resident memory T cell (TRM), a stem cell-like memory T cell (TSCM) or a tumor infiltrating lymphocyte (TIL).

[0308] Also provided herein is a population of cells comprising the one or more recombinant polynucleic acid molecules of the composition of the present disclosure or the one or more polypeptides the present disclosure.

[0309] Also provided herein is a pharmaceutical composition comprising the one or more recombinant polynucleic acid molecules of the composition of the present disclosure.

[0310] Also provided herein is a pharmaceutical composition comprising the one or more polypeptides of the present disclosure.

[0311] Also provided herein is a pharmaceutical composition comprising the cell of the present disclosure.

[0312] Also provided herein is a pharmaceutical composition comprising the population of cells of the present disclosure.

[0313] Also provided herein is a lentiviral expression vector comprising the one or more recombinant polynucleic acid molecules of the composition of the present disclosure.

[0314] In some embodiments, the lentiviral expression vector is the pCCL-c-MNDU2-X vector.

[0315] In some embodiments, the pCCL-cMNDU2-X vector further comprises a woodchuck hepatitis virus posttranscriptional regulatory element (WPRE).INCORPORATION BY REFERENCE

[0316] All publications, patents, and patent applications mentioned in this specification are herein incorporated by reference in their entireties to the same extent as if each individual publication, patent, or patent application was specifically and individually indicated to be incorporated by reference.BRIEF DESCRIPTION OF THE DRAWINGS

[0317] The novel features of the invention are set forth with particularity in the appended claims. A better understanding of the features and advantages of the present invention will be obtained by reference to the following detailed description that sets forth illustrative embodiments, in which the principles of the invention are utilized, and the accompanying drawings of which:

[0318] FIG. 1 shows IL-2 secretion by T cells expressing an anti-CD19 CAR. FIG. 1A shows the amount of IL-2 secretion after incubating the anti-CD19 CAR-T cells with wild type Nalm6 tumor cells. FIG. IB shows the amount of IL-2 secretion after incubating the anti-CD19 CAR-T cells with Nalm6 tumor cells expressing low level of CD 19. FIG. 1C shows the amount of IL-2 secretion after incubating the anti-CD19 CAR-T cells with Nalm6 tumor cells with CD 19 knocked out. FIG. ID shows the amount of IL-2 secretion after incubating the anti-CD19 CAR-T cells with CD 19 low Nalm6 tumor cells at a ratio of 1 : 12.

[0319] FIG. 2 shows IFN-gamma secretion by T cells expressing an anti-CD19 CAR. FIG. 2A shows the amount of IFN-gamma secretion after incubating the anti-CD19 CAR-T cells with CD 19 low Nalm6 tumor cells. FIG. 2B shows the amount of IFN-gamma secretion after incubating the anti-CD19 CAR-T cells with Nalm6 tumor cells with CD 19 knocked out. FIG. 2C shows the amount of IFN-gamma secretion by the anti-CD19 CAR-T cells without the target cells. FIG. 2D shows the amount of IFN-gamma secretion after incubating the anti-CD19 CAR-T cells with CD 19 low Nalm6 tumor cells at a ratio of 1 : 12.

[0320] FIG. 3 shows the in vivo anti-leukemia activity of anti-CD19 CAR T cells in a mouse model. FIG. 3A shows the bioluminescent flux of the tumors over time after injecting the CAR-T cells expressing the anti -CD 19 CAR with a CD28 co-stimulatory intracellular signaling domain and aCD3zeta intracellular signaling domain. FIG. 3B shows the bioluminescent flux of the tumors over time after injecting the CART cells expressing the anti-CD19 CAR with a CD2 co-stimulatory intracellular signaling domain and a CD3zeta intracellular signaling domain. The results suggested that treatment with anti-CD19 CARTs delayed tumor growth.

[0321] FIG. 4 shows IL-2 secretion by T cells expressing anti-CD20 CAR. FIG. 4A shows the amount of IL-2 secretion after incubating the anti-CD20 CAR-T cells with CD20 overexpressed Nalm6 tumor cells. FIG. 4B shows the amount of IL-2 secretion after incubating the anti-CD20 CAR-T cells with wild type Raji tumor cells. FIG. 4C shows the amount of IL-2 secretion after incubating the anti-CD20 CAR-T cells with CD20 knocked out Raji cells. FIG. 4D shows the amount of IL-2 secretion after incubating the anti-CD20 CAR-T cells with wild type Raji tumor cells.

[0322] FIG. 5 shows IFN-gamma secretion by T cells expressing an anti-CD20 CAR. FIG. 5A shows the amount of IFN-gamma secretion after incubating the anti-CD20 CAR-T cells with CD20 overexpressing Nalm6 tumor cells. FIG. 5B shows the amount of IFN-gamma secretion after incubating the anti-CD20 CAR-T cells with wild type Raji tumor cells. FIG. 5C shows the amount of IFN-gamma secretion after incubating the anti-CD20 CAR-T cells with CD20 knocked out Raji tumor cells. FIG. 5D shows the amount of IFN-gamma secretion by the anti-CD20 CAR-T cells without the target cells. FIG. 5E shows the amount of IFN-gamma secretion after incubating the anti-CD20 CAR-T cells with wild type Raji cells.

[0323] FIG. 6 shows the in vivo anti-lymphoma activity of anti-CD20 CART in a mouse model. FIG. 6A shows the bioluminescent flux of the tumors over time after injecting the CART cells expressing the anti-CD20 CAR with a CD28 co-stimulatory intracellular signaling domain and a CD3zeta intracellular signaling domain. FIG. 6B shows the bioluminescent flux of the tumors over time after injecting the CART cells expressing the anti-CD20 CAR with a CD2 co-stimulatory intracellular signaling domain and a CD3zeta intracellular signaling domain. The results suggested that treatment of the anti-CD20 CARTs inhibited tumor growth.

[0324] FIG. 7 depicts the mechanism of treating CD58 downregulated tumors with CARs comprising a CD2 co-stimulatory intracellular signaling domain.

[0325] FIG. 8 illustrates that CART cells with a CD2 co-stimulatory intracellular signaling domain can improve cell killing against tumor cells that lack CD58 expression. FIG. 8A depicts an exemplary structure of anti-CD22 CAR and anti-CD19 CAR with a CD2 co-stimulatory intracellular signaling domain. FIG. 8B shows the tumor killing activity of T-cells expressing the anti-CD22 CAR and / or anti-CD19 CAR.

[0326] FIG. 9 illustrates the technology of creating a cell population with a high percentage of cells expressing multiple CARs. FIG. 9A depicts the process of creating and purifying the cells expressing multiple CARs in a cell population. FIG. 9B lists the advantages of this technology.

[0327] FIG. 10 is a flow cytometry plot demonstrating that after the process shown in FIG. 9, more than 80% cells in the cell population expressed three CARs.

[0328] FIG. 11A depicts an exemplary first polynucleotide molecule encoding a first polypeptide containing a cytokine receptor switch polypeptide and a first chimeric antigen receptor separated by a sequence encoding a P2A self-cleaving peptide, an exemplary second polynucleotide molecule encoding a second polypeptide containing a transmembrane domain and a second chimeric antigen receptor separated by a sequence encoding a P2A self-cleaving peptide, and an exemplary third polynucleotide molecule encoding a third polypeptide containing a transmembrane domain and a third chimeric antigen receptor separated by a sequence encoding a P2A self-cleaving peptide. FIG. 11B depicts an exemplary cytokine receptor switch polypeptide expressed at the cell-surface when the second polypeptide and the third polypeptide of FIG. 11A are expressed in the same cell (which form a multimer with the cytokine receptor switch polypeptide). FIG. 11C depicts an exemplary cytokine receptor switch polypeptide expressed at the cell-surface when the second polypeptide of FIG. 11A is expressed in the same cell.

[0329] FIG. 12A depicts an exemplary first polynucleotide molecule encoding a cytokine receptor switch polypeptide with a protease cleavage site and a retention tag and a first chimeric antigen receptor separated by a sequence encoding a P2A self-cleaving peptide, an exemplary second polynucleotide molecule encoding a C-terminal portion of a protease and a second chimeric antigen receptor separated by a sequence encoding a P2A self-cleaving peptide, and an exemplary third polynucleotide molecule encoding a N-terminal portion of the protease and a third chimeric antigen receptor separated by a sequence encoding a P2A self-cleaving peptide. FIG. 12B depicts an exemplary cytokine receptor switch polypeptide expressed at the cell-surface when the C-terminal portion of the protease and the N-terminal portion of the protease are expressed in the same cell and form a functional protease. As depicted, the C-terminal portion and N-terminal portion of the protease come together to form a functional protease that cleaves the cytokine receptor switch polypeptide at the protease cleavage site in the ER or Golgi, releasing the retention tag and promoting cell-surface expression of the cytokine receptor switch polypeptide. FIG. 12C depicts an exemplary cytokine receptor switch polypeptide expressed at the cell-surface when the C-terminal portion of the protease and the N-terminal portion of the protease are expressed in the same cell and form a functional protease. As depicted, the C-terminal portion and N-terminal portion of theprotease come together to form a functional protease that cleaves the cytokine receptor switch polypeptide at the protease cleavage site in the ER or Golgi, releasing the retention tag and promoting cell-surface expression of the cytokine receptor switch polypeptide.

[0330] FIG. 13 shows IL-2 secretion by monospecific or trispecific CAR-T cells after incubating with Nalm6 tumor cells. FIG. 13A shows IL-2 secretion by CAR-T cells incubated with Nalm6 CD19+CD20'CD22‘ cells. FIG. 13B shows IL-2 secretion by CAR-T cells incubated with Nalm6 CD19'CD20+CD22‘ cells. FIG. 13C shows IL-2 secretion by CAR-T cells incubated with Nalm6 CD19 CD20 CD22+cells. The result suggested that each CAR in the trispecific CAR-T cells was able to induce the T-cells to secret IL-2 in response to the antigen at levels similar to the mono- specific CAR-T cells.

[0331] FIG. 14 shows the in vivo anti-lymphoma activity of the monospecific CAR-T and the trispecific CAR-T in a mouse model.

[0332] FIG. 15 shows the in vivo anti-lymphoma activity of the monospecific CAR-T and the trispecific CAR-T in an antigen knocked-out tumor mouse model.

[0333] FIG. 16A depicts an exemplary polynucleotide molecule encoding a polypeptide containing an anti-CD20 scFv, an anti-CD19 scFv, a hinge domain, a transmembrane domain, a co-stimulatory signaling domain, and an activation domain. FIG. 16B depicts an exemplary first polynucleotide molecule encoding a first polypeptide containing a first scFv, a first hinge domain, a first transmembrane domain, a first co-stimulatory signaling domain, and a first activation domain, and an exemplary second polynucleotide molecule encoding a second polypeptide containing a second scFv, a second hinge domain, a second transmembrane domain, a second co-stimulatory signaling domain, and a second activation domain. FIG. 16C depicts an exemplary first polynucleotide molecule encoding a first polypeptide containing a first scFv, a first hinge domain, a first transmembrane domain, a first co-stimulatory signaling domain, and a first activation domain, and an exemplary second polynucleotide molecule encoding a second polypeptide containing a second scFv, a third scFv, a second hinge domain, a second transmembrane domain, a second co-stimulatory signaling domain, and a second activation domain. FIG. 16D depicts an exemplary first polynucleotide molecule encoding a first polypeptide containing a first scFv, a first hinge domain, a first transmembrane domain, a first co-stimulatory signaling domain, and a first activation domain, an exemplary second polynucleotide molecule encoding a second polypeptide containing a second scFv, a second hinge domain, a second transmembrane domain, a second co-stimulatory signaling domain, and a second activation domain, and an exemplary third polynucleotide molecule encoding a thirdpolypeptide containing a third scFv, a third hinge domain, a third transmembrane domain, a third costimulatory signaling domain, and a third activation domain.

[0334] FIG. 17 depicts exemplary polynucleotide molecules encoding a first CAR, a second CAR, and a third CAR, separated by a sequence encoding a P2A self-cleaving peptide.

[0335] FIG. 18A shows the amount of IL-2 secretion after incubating the anti-CD19 / anti-CD20 bispecific CAR-T cells with Raji WT cells, Raji CD19KO cells, Raji CD20KO cells, Raji CD19 / CD20 double KO cells, or Nalm6 clone 11 cells. The anti-CD19 CAR and the anti-CD20 CAR were expressed in a bicistronic vector as shown in FIG. 16B. FIG. 18B shows the amount of IFN-gamma secretion after incubating the anti-CD19 / anti-CD20 bispecific CAR-T cells with Raji WT cells, Raji CD19KO cells, Raji CD20KO cells, Raji CD19 / CD20 double KO cells, or Nalm6 clone 11 cells for 24 hours. FIG. 18C shows the in vivo anti-lymphoma activity of the antiCD 19 / anti-CD20 bispecific CAR-T cells in an antigen knocked-out tumor mouse model.

[0336] FIG. 19A shows the amount of IL-2 secretion after incubating the anti-CD19 / anti-CD20 bispecific CAR-T cells with Raji WT cells, Raji CD19KO cells, Raji CD20KO cells, Raji CD19 / CD20 double KO cells, or Nalm6 clone 11 cells. The bispecific CAR-T cells targeting CD19 and CD20 consisted of a single CAR with two antigen binding moi eties as shown in FIG. 16A. FIG. 19B shows the amount of IFN-gamma secretion after incubating the anti-CD19 / anti-CD20 bispecific CAR-T cells with Raji WT cells, Raji CD19KO cells, Raji CD20KO cells, Raji CD19 / CD20 double KO cells, or Nalm6 clone 11 cells for 24 hours. FIG. 19C shows the in vivo anti-lymphoma activity of the anti-CD19 / anti-CD20 bispecific CAR-T cells in an antigen knocked-out tumor mouse model.

[0337] FIG. 20A shows the amount of IL-2 secretion after incubating the anti-CD19 / anti-CD20 bispecific CAR-T cells or the anti-CD20 / anti-CD22 bispecific CAR-T cells with Raji WT cells, Raji CD19 / CD20 double KO cells, Raji CD19 / CD22 double KO cells, Raji CD20 / CD22 double KO cells, or Raji CD19 / CD20 / CD22 triple KO cells. The bispecific CAR-T cells consisted of two independent CARs expressed in a bicistronic vector. FIG. 20B shows the amount of IFN-gamma secretion after incubating the anti-CD19 / anti-CD20 bispecific CAR-T cells or the anti-CD20 / anti-CD22 bispecific CAR-T cells with Raji WT cells, Raji CD19 / CD20 double KO cells, Raji CD19 / CD22 double KO cells, Raji CD20 / CD22 double KO cells, or Raji CD19 / CD20 / CD22 triple KO cells for 24 hours.

[0338] FIG. 21 A shows the amount of IL-2 secretion after incubating the anti-CD19 / anti-CD20 / anti- CD22 trispecific CAR-T cells with Raji WT cells, Raji CD58KO cells, Raji CD19 / CD20 double KO cells, or Raji CD19 / CD20 / CD22 triple KO cells. The trispecific CAR-T cells targeting CD19, CD20 and CD22 consisted of a tandem anti-CD19 / anti-CD20 CAR and a monospecific antiCD22 CAR encoded on the same polynucleotide molecule. FIG. 21B shows the amount of IFN-gammasecretion after incubating the anti-CD19 / anti-CD20 / anti-CD22 trispecific CAR-T cells with Raji WT cells, Raji CD58KO cells, Raji CD19 / CD20 double KO cells, or Raji CD19 / CD20 / CD22 triple KO cells for 24 hours. FIG. 21C shows the in vivo anti -lymphoma activity of the anti-CD19 / anti- CD20 / anti-CD22 trispecific CAR-T cells in an antigen knocked-out tumor mouse model.

[0339] FIG. 22A shows the amount of IL-2 secretion after incubating the anti-CD19 / anti-CD20 / anti- CD22 trispecific CAR-T cells with Raji WT cells, Raji CD58KO cells, Raji CD19 / CD20 double KO cells, or Raji CD19 / CD20 / CD22 triple KO cells. The trispecific CAR-T cells targeting CD19, CD20 and CD22 consisted of three independent CARs. FIG. 22B shows the amount of IFN-gamma secretion after incubating the anti-CD19 / anti-CD20 / anti-CD22 trispecific CAR-T cells with Raji WT cells, Raji CD58KO cells, Raji CD19 / CD20 double KO cells, or Raji CD19 / CD20 / CD22 triple KO cells for 24 hours. FIG. 22C shows the in vivo anti -lymphoma activity of the anti-CD19 / anti- CD20 / anti-CD22 trispecific CAR-T cells in an antigen knocked-out tumor mouse model.

[0340] FIG. 23A shows the amount of IL-2 secretion after incubating the anti-CD19 / anti-CD20 / anti- CD22 trispecific CAR-T cells with Raji WT cells. The trispecific CAR-T cells targeting CD19, CD20 and CD22 consisted of three independent CARs encoded in one polynucleotide molecule as shown in FIG. 17. FIG. 23B shows the amount of IFN-gamma secretion after incubating the anti- CD19 / anti-CD20 / anti-CD22 trispecific CAR-T cells with Nalm6 WT cells for 24 hours. FIG. 23C shows the in vivo anti -lymphoma activity of the anti-CD19 / anti-CD20 / anti-CD22 trispecific CAR-T cells in an antigen knocked-out tumor mouse model.

[0341] FIG. 24A shows the amount of IL-2 secretion after incubating the anti-CD19 / anti-CD20 / anti- CD22 trispecific CAR-T cells with Nalm6 WT cells, Nalm6 CD22KO cells, Nalm6 CD19 / CD22 double KO cells, or Nalm6 CD19 / CD20 / CD22 triple KO cells. The trispecific CAR-T cells targeting CD 19, CD20 and CD22 consisted of three independent CARs. FIG. 24B shows the amount of IFN- gamma secretion after incubating the anti-CD19 / anti-CD20 / anti-CD22 trispecific CAR-T cells with Nalm6 WT cells, Nalm6 CD22KO cells, Nalm6 CD19 / CD22 double KO cells, or Nalm6 CD19 / CD20 / CD22 triple KO cells for 24 hours. FIG. 24C shows the in vivo anti -lymphoma activity of the anti-CD19 / anti-CD20 / anti-CD22 trispecific CAR-T cells in an antigen knocked-out tumor mouse model.

[0342] FIG. 25A shows the amount of IL-2 secretion after incubating the anti-CD19 / anti-CD20 / anti- CD22 trispecific CAR-T cells with Nalm6 WT cells, Nalm6 CD19KO cells, Nalm6 CD22KO cells, Nalm6 cells with high level of CD20, Nalm6 CD19 / CD22 double KO cells, or Nalm6 CD19 / CD20 / CD22 triple KO cells. The trispecific CAR-T cells targeting CD 19, CD20 and CD22 consisted of three independent CARs with either a CD28 or 4-1BB co-stimulatory signaling domainon the CD 19 CAR. FIG. 25B shows the amount of IFN-gamma secretion after incubating the anti- CD19 / anti-CD20 / anti-CD22 trispecific CAR-T cells with Nalm6 WT cells, Nalm6 CD19K0 cells, Nalm6 CD22KO cells, Nalm6 cells with high level of CD20, Nalm6 CD19 / CD22 double KO cells, or Nalm6 CD19 / CD20 / CD22 triple KO cells for 24 hours. FIG. 25C shows the in vivo antilymphoma activity of the anti-CD19 / anti-CD20 / anti-CD22 trispecific CAR-T cells in a Nalm6 with low level of CD 19 tumor mouse model.

[0343] FIG. 26A shows the amount of IL-2 secretion after incubating the anti-CD19 / anti-CD20 / anti- CD22 trispecific CAR-T cells with Nalm6 WT cells, Nalm6 CD58KO cells, Nalm6 CD19KO cells, Nalm6 CD22KO cells, Nalm6 cells with high level of CD20, Nalm6 CD19 / CD22 double KO cells with high level of CD20, or Nalm6 CD19 / CD20 / CD22 triple KO cells. The trispecific CAR-T cells targeting CD 19, CD20 and CD22 consisted of three independent CARs encoded in one polynucleotide molecule as shown in FIG. 17. FIG. 26B shows the amount of IFN-gamma secretion after incubating the anti-CD19 / anti-CD20 / anti-CD22 trispecific CAR-T cells with Nalm6 WT cells, Nalm6 CD58KO cells, Nalm6 CD19KO cells, Nalm6 CD22KO cells, Nalm6 cells with high level of CD20, Nalm6 CD19 / CD22 double KO cells with high level of CD20, or Nalm6 CD19 / CD20 / CD22 triple KO cells for 24 hours.

[0344] FIG. 27A depicts two exemplary polynucleotide constructs. One polynucleotide encodes Tag-TREMl, an anti-CD19 CAR, and an anti-CD20 CAR. The other polynucleotide construct encodes DAP 12 and an anti-CD22 CAR.

[0345] FIG. 27B shows the percentage of cells with cell surface expression of CD22 CAR and a EGFRt or CD34t tag before and after MACS beads purification.

[0346] FIG. 27C shows the percentage of cells with cell surface expression of CD22 CAR and CD 19 CAR prior to MACS beads purification and after MACS beads purification.

[0347] FIG. 28A shows the percentage of cells with cell surface expression of the CD34t tag and CD22 CAR prior to MACS separation, in the flow through, and after MACS selection. The cells were transduced with Vector 2883 and Vector 3050, 3051, 3056, or 3057.

[0348] FIG. 28B shows the percentage of cells with cell surface expression of the CD34t tag and CD22 CAR prior to MACS separation, in the flow through, or after MACS selection. The cells were transduced with Vector 2884 and Vector 3050, 3051, 3056, or 3057.

[0349] FIG. 29A shows the percentage of cells with cell surface expression of the CD34t tag and CD22 CAR prior to MACS separation, in the flow through, or after MACS selection. The cells were transduced with Vector 2884 and Vector 3137, 3138, or 3139.

[0350] FIG. 29B shows the percentage of cells with cell surface expression of the CD34t tag and CD22 CAR prior to MACS separation, in the flow through, or after MACS selection. The cells were transduced with Vector 2884 and Vector 3140, 3141, or 3142.

[0351] FIG. 30A shows the percentage of cells expressing the CD34t tag and CD22 CAR prior to MACS purification. The cells were transduced with a vector expressing CD34t-TREMl and CD20 CAR and a vector expressing DAP 12, CD 19 CAR, and CD22 CAR.

[0352] FIG. 30B shows the percentage of cells expressing the CD34t tag and CD22 CAR after MACS purification. The cells were transduced with a vector expressing CD34t-TREMl and CD20 CAR and a vector expressing DAP 12, CD 19 CAR, and CD22 CAR.

[0353] FIG. 30C shows the percentage of cells expressing CD 19 CAR and CD22 CAR after MACS purification. The cells were transduced with a vector expressing CD34t-TREMl and CD20 CAR and a vector expressing DAP 12, CD 19 CAR, and CD22 CAR.

[0354] FIG. 31A depicts three exemplary polynucleotide constructs. One polynucleotide construct encodes a CD22 CAR and CD79a linked by a P2A sequence. One polynucleotide construct encodes a CD 19 CAR and CD79b linked by a P2A sequence.

[0355] FIG. 31B shows the percentage of cells expressing the CD34t tag and CD22 CAR prior to MACS purification. The cells were transduced with a vector expressing CD22 CAR and CD79a, a vector expressing CD 19 CAR and CD79b, and a vector expressing CD20 CAR and mlgM with a truncated CD34 tag.

[0356] FIG. 31C shows the percentage of cells expressing the CD34t tag and CD22 CAR after MACS purification. The cells were transduced with a vector expressing CD22 CAR and CD79a, a vector expressing CD 19 CAR and CD79b, and a vector expressing CD20 CAR and mlgM with a CD34t tag.

[0357] FIG. 31D shows the percentage of cells expressing CD 19 CAR and CD22 CAR after MACS purification. The cells were transduced with a vector expressing CD22 CAR and CD79a, a vector expressing CD 19 CAR and CD79b, and a vector expressing CD20 CAR and mlgM with a CD34t tag.

[0358] FIG. 31E shows the percentage of cells expressing CD20 CAR and CD22 CAR prior to MACS purification. The cells were transduced with a vector expressing CD22 CAR and CD79a, a vector expressing CD 19 CAR and CD79b, and a vector expressing CD20 CAR and mlgM with a CD34t tag.

[0359] FIG. 32 shows the in vivo anti-lymphoma activity of the anti -CD 19 19c493 monospecific CAR-T, anti-CD22 m971 monospecific CAR-T, anti-CD20 20c27 monospecific CAR-T and the trispecific CAR-T in a mouse model.

[0360] FIGs. 33A-33B show quantification of cytokine secretion by trispecific pSync3811 CAR-T cells incubated with single antigen-expressing target cells.

[0361] FIGs. 34A-34B show quantification of cytotoxicity by trispecific pSync3811 CAR-T cells incubated with single antigen-expressing target cells.

[0362] FIGs. 35A-35F show repeated killing of single antigen-expressing Raji tumor cell variants by the trispecific CAR-T cell encoded by the plasmid construct designated 3811 or other designated investigational multi-specific CAR-T cells.

[0363] FIGs. 36A-36C show repeated killing of single antigen-expressing Raji tumor cell variants by the trispecific CAR-T cell encoded by the plasmid construct designated 3811 or other multispecific CAR-T cells.

[0364] FIG. 37 shows repeated killing of Raji wild-type cells by various CAR-T cells at an effector- to-target ratio of 1 :3.

[0365] FIG. 38 shows a dose response of the trispecific CAR-T cell encoded by the plasmid construct designated 3811 against Raji tumor cells in NCG mice for Donor 1.

[0366] FIG. 39 shows a dose response of the trispecific CAR-T cell encoded by the plasmid construct designated 3811 against Raji tumor cells in NCG mice for Donor 2.

[0367] FIG. 40 shows a dose response of the trispecific CAR-T cell encoded by the plasmid construct designated 3811 against Raji tumor cells in NCG mice for Donor 3.

[0368] FIG. 41 shows clearance of tumor cells in antigen loss animal model by the trispecific CAR- T cell encoded by the plasmid construct designated 3811.

[0369] FIG. 42 shows clearance of Raji tumor cells in NCG mice by the trispecific CAR-T cell encoded by the plasmid construct designated 3811 or other mono- and multi-specific CAR-T cells.

[0370] FIG. 43 shows low dose CRG-023 effectively controls tumor growth.

[0371] FIGs. 44A-44C shows comparison of m971 BBz CAR with the trispecific CAR-T cell encoded by the plasmid construct designated 3811.DETAILED DESCRIPTION

[0372] Autologous cell therapy with T cells expressing synthetic chimeric antigen receptors (CARs) targeting one of the B cell antigens CD 19, CD20, and CD22 has revolutionized treatment of B cell malignancies since the FDA first approved the sale of Kymriah®, a CD19-specific CAR-T cell therapy, in August 2017. Although autologous CAR-T cells have induced beneficial therapeuticresponses in many of the patients receiving such treatment, as many as 60% of those treated relapse, and 10-20% of those who relapse after CD 19 CAR-T cell treatment experience CD 19-negative relapse. In addition to down-regulation or loss of target antigen expression, relapse can be caused by mutation of the epitope recognized by the antigen-binding domain of the CAR, poor CAR-T cell persistence, transient B cell aplasia, or down-regulation or elimination of co-stimulatory signaling pathways necessary for T cell activation, among other things. Thus, there remains a substantial unmet clinical need for improved autologous CAR-T cell therapies for the treatment of B cell cancers.

[0373] The complex nature of the interaction between a CAR-T cell and the target tumor cell - particularly with respect to the binding interaction between the antigen-specific domain(s) and its cognate antigen(s) - renders it difficult to predict what combination of physical parameters will result in an effective therapeutic. For example, optimal antigen affinity and avidity for a CAR binder must be sufficiently high to initiate the T cell activation signaling cascade but not high enough to result in activation-induced death of the CAR T cell. However, it has also been shown that even antigen-binding fragments (i.e., single chain variable fragments, or scFvs) with similar binding affinities for the same target antigen can differentially impact CAR T cell function. Thus, empirical testing of these parameters alone and in combination is essential to developing an effective mono-, bi- or multi-specific CAR-T cell.

[0374] The various CAR constructs disclosed herein employ novel fully human antigen-binding domains for CD 19 and CD20; the CD 19 binder was selected for its ability to bind CD 19 at low antigen density and at a lower binding affinity than FMC63, the standard CD19 scFv commonly used in the field. Coupled with effective hinge and transmembrane domains and a combination of three different co-stimulatory domains capable of providing co-stimulation along the CD28, 4-1BB, and CD2 signaling axes, the tri-specific CAR-T cell embodiments disclosed herein comprise three mono- specific CARs that are each robustly expressed from a single lentiviral expression vector, each sufficient to induce a durable and independent anti-tumor response when only one of the three cognate B cell antigens is present on the target cells, and capable of clearing tumor cells in vivo at substantially lower doses than other mono-, bi-, and tri-specific CARs under development. Thus, provided herein are novel, unexpectedly potent tri-specific CAR-T cell therapies poised to reduce the likelihood of relapse in first-line patients and to offer hope to the patient population having relapsed or refractory disease following treatment with a mono-specific CAR-T cell therapy (e.g., CD19- specific CAR-T cell therapy).

[0375] Thus, in general, the present disclosure relates, inter alia, to compositions and methods for improving autologous CAR-T cell therapy. The inventors have discovered ways to improve the therapeutic efficacy of T cells by addressing loss or down-regulation of target antigen expression, loss of co-stimulation and limited CAR-T-cell persistence.

[0376] In particular, some embodiments of the disclosure provide novel antigen binding domains to CD 19 and CD20, which have a lower binding affinity for their target antigens as compared to known anti-CD19 and anti-CD20 binders and exhibit superior properties when incorporated into CARs. Also provided herein are exemplary chimeric receptors that incorporate the novel antigen binding domains to CD 19 and CD20, as well as the previously characterized anti-CD22 binder m971, and possess differing costimulatory domains and intracellular domains of a CAR, for example, hinge and transmembrane domains derived from CD8alpha and CD28, and intracellular signaling domains derived from CD3zeta, CD28, CD2, and 4- IBB. In a composition comprising the combination of three binders targeting the antigens CD 19, CD20, and CD22, these trispecific CARs outperform other monospecific, bispecific, and trispecific CARs both in vitro and in vivo. In particular, the new CARs and compositions provided herein can enhance both cytokine production and elimination of tumors at lower doses than CARs of comparable specificity.

[0377] Disclosed herein include compositions and methods for immunotherapies. In one aspect, the present disclosure describes a composition comprising one or more recombinant polynucleic acid molecules comprising: (a) a sequence encoding an anti-CD20 CAR, wherein the anti-CD20 CAR comprises: (i) an extracellular domain comprising an anti-CD20 binding domain, (ii) a transmembrane domain, and (iii) a cytoplasmic domain comprising (A) a co-stimulatory signaling domain from CD2 and (B) an intracellular signaling domain; and (b) a sequence encoding an anti- CD22 CAR, wherein the anti-CD22 CAR comprises: (i) an extracellular domain comprising an anti- CD22 binding domain, (ii) a transmembrane domain, and (iii) a cytoplasmic domain comprising an intracellular signaling domain.

[0378] In another aspect, provided herein is a composition comprising one or more recombinant polynucleic acid molecules comprising: (a) a sequence encoding an anti-CD19 CAR, wherein the anti-CD19 CAR comprises: (i) an extracellular domain comprising an anti-CD19 binding domain, (ii) a transmembrane domain, and (iii) a cytoplasmic domain comprising (A) a co-stimulatory signaling domain from CD2, CD28, or 4-1BB and (B) an intracellular signaling domain; and (b) a sequence encoding an anti-CD22 CAR, wherein the anti-CD22 CAR comprises: (i) an extracellular domain comprising an anti-CD22 binding domain, (ii) a transmembrane domain, and (iii) a cytoplasmic domain comprising an intracellular signaling domain.

[0379] In another aspect, provided herein is a composition comprising one or more recombinant polynucleic acid molecules comprising: (a)a sequence encoding an anti-CD20 CAR, wherein the anti-CD20 CAR comprises: (i) an extracellular domain comprising an anti-CD20 binding domain,(ii) a transmembrane domain, and (iii) a cytoplasmic domain comprising an intracellular signaling domain; and (b) a sequence encoding an anti-CD19 CAR, wherein the anti-CD19 CAR comprises: (i) an extracellular domain comprising an anti-CD19 binding domain, (ii) a transmembrane domain, and (iii) a cytoplasmic domain comprising an intracellular signaling domain wherein the cytoplasmic domain of the anti-CD19 CAR and / or the cytoplasmic domain of the anti-CD20 CAR further comprises a co-stimulatory signaling domain from CD2.

[0380] In another aspect, provided herein is a composition comprising one or more recombinant polynucleic acid molecules comprising: (a) a sequence encoding an anti-CD19 chimeric antigen receptor (CAR), wherein the anti-CD19 CAR comprises: (i) an extracellular domain comprising an anti-CD19 binding domain, (ii) a transmembrane domain, and (iii) a cytoplasmic domain comprising an intracellular signaling domain; and (b) a sequence encoding an anti-CD20 CAR, wherein the anti- CD20 CAR comprises: (i) an extracellular domain comprising an anti-CD20 binding domain, (ii) a transmembrane domain, and (iii) a cytoplasmic domain comprising an intracellular signaling domain; and (c) a sequence encoding an anti-CD22 CAR, wherein the anti-CD22 CAR comprises: (i) an extracellular domain comprising an anti-CD22 binding domain, (ii) a transmembrane domain, and(iii) a cytoplasmic domain comprising an intracellular signaling domain; wherein the cytoplasmic domain of the anti-CD19 CAR and / or the cytoplasmic domain of the anti-CD20 CAR further comprises a co-stimulatory signaling domain from CD2. In some embodiments, the cytoplasmic domain of the anti-CD20 CAR comprises a co-stimulatory signaling domain from CD2. In some embodiments, the cytoplasmic domain of the anti-CD19 CAR comprises a co-stimulatory signaling domain from CD2, CD28, 4-1BB or CD3zeta.

[0381] Also provided herein is a composition comprising a recombinant polynucleic acid molecule comprising a sequence encoding polypeptide with an anti-CD19 binding domain, wherein the anti- CD19 binding domain comprises a heavy chain variable region (VH) that comprises a heavy chain CDR3 (HCDR3) sequence selected from the group consisting of HCDR3 sequences in tables 5-7.

[0382] In another aspect, provided herein is a composition comprising a recombinant polynucleic acid molecule comprising a sequence encoding polypeptide with an anti-CD20 binding domain, wherein the anti-CD20 binding domain comprises a heavy chain variable region (VH) that comprises a heavy chain CDR3 (HCDR3) sequence selected from the group consisting of HCDR3 sequences in tables 13-15.

[0383] Also provided herein is a composition comprising one or more recombinant polynucleic acid molecules comprising: (a) a sequence encoding a bispecific chimeric antigen receptor (CAR), wherein the bispecific CAR comprises: (i) an extracellular domain comprising an anti-CD19 binding domain and an anti-CD20 binding domain, (ii) a transmembrane domain, and (iii) a cytoplasmic domain comprising an intracellular signaling domain; and wherein the cytoplasmic domain further comprises a co-stimulatory signaling domain from CD2.

[0384] Also provided herein is a composition comprising one or more recombinant polynucleic acid molecules comprising: (a) a sequence encoding a bispecific CAR, wherein the bispecific CAR comprises: (i) an extracellular domain comprising an anti-CD19 binding domain and an anti-CD22 binding domain, (ii) a transmembrane domain; and (iii) a cytoplasmic domain comprising (A) a costimulatory signaling domain from CD2, CD28, or 4-1BB and (B) an intracellular signaling domain.

[0385] Also provided herein is a composition comprising one or more recombinant polynucleic acid molecules comprising: (a) a sequence encoding a bispecific CAR, wherein the bispecific CAR comprises: (i) an extracellular domain comprising an anti-CD20 binding domain and anti-CD22 binding domain, (ii) a transmembrane domain, and (iii) a cytoplasmic domain comprising (A) a costimulatory signaling domain from CD2 and (B) an intracellular signaling domain.

[0386] Also provided herein is a composition comprising one or more recombinant polynucleic acid molecules comprising: (a) a sequence encoding a bispecific chimeric antigen receptor (CAR), wherein the bispecific CAR comprises: (i) an extracellular domain comprising an anti-CD19 binding domain and an anti-CD20 binding domain, (ii) a transmembrane domain, and (iii) a cytoplasmic domain comprising an intracellular signaling domain; and (b) a sequence encoding an anti-CD22 CAR, wherein the anti-CD22 CAR comprises: (i) an extracellular domain comprising an anti-CD22 binding domain, (ii) a transmembrane domain, and (iii) a cytoplasmic domain comprising an intracellular signaling domain; wherein the cytoplasmic domain of the bispecific CAR further comprises a co-stimulatory signaling domain from CD2.

[0387] Also provided herein is a cell comprising one or more recombinant polynucleic acid molecules of the composition disclosed herein. In another aspect, provided herein is a pharmaceutical composition comprising one or more of the recombinant polynucleic acid molecules of the compositions disclosed herein. In another aspect, provided herein is a method of making a cell comprising introducing one or more recombinant polynucleic acid molecules of the compositions disclosed herein or any one or more of the polypeptides disclosed herein into a cell. In another aspect, provided herein is a method of treating a disease or condition in a subject in need thereofcomprising administering a therapeutically effective amount of a pharmaceutical composition disclosed herein.

[0388] In another aspect, provided herein is a viral vector comprising a polynucleic acid sequence that is at least 3000 nucleotides in length, wherein the polynucleic acid sequence encodes a polypeptide that is at least 1000 amino acids in length, and wherein the polypeptide is expressed in at least 50% of the cells in a cell population when transduced into the cell population.

[0389] In yet another aspect, provided herein is a viral vector comprising a polynucleic acid sequence that is at least 3000 nucleotides in length, wherein the polynucleic acid sequence encodes at least two polypeptides, wherein the sum of the lengths of the at least two polypeptides is at least 1000 amino acids in length, and wherein each of the at least two polypeptides is expressed in at least 50% of the cells in a cell population when transduced into the cell population.

[0390] In another aspect, provided herein is a population of at least 1 x 10A3 cells expressing a polypeptide encoded by a polynucleic acid sequence of a single viral vector that is at least 3000 nucleotides in length, wherein the polypeptide is at least 1000 amino acids in length.Definitions

[0391] The singular form “a”, “an”, and “the” include plural references unless the context clearly dictates otherwise. For example, the term “a cell” includes one or more cells, including mixtures thereof. “A and / or B” is used herein to include all of the following alternatives: “A”, “B”, “A or B”, and “A and B.”

[0392] Where a range of values is provided, it is understood that each intervening value, to the tenth of the unit of the lower limit unless the context clearly dictates otherwise, between the upper and lower limit of that range and any other stated or intervening value in that stated range, is encompassed within the disclosure. The upper and lower limits of these smaller ranges may independently be included in the smaller ranges, and are also encompassed within the disclosure, subject to any specifically excluded limit in the stated range. Where the stated range includes one or both of the limits, ranges excluding either or both of those included limits are also included in the disclosure.

[0393] Certain ranges are presented herein with numerical values being preceded by the term “about.” The term “about” is used herein to provide literal support for the exact number that it precedes, as well as a number that is near to or approximately the number that the term precedes. In determining whether a number is near to or approximately a specifically recited number, the near orapproximating unrecited number may be a number which, in the context in which it is presented, provides the substantial equivalent of the specifically recited number.

[0394] “Percent (%) identity” with respect to the nucleic acid or amino acid sequences identified herein is defined as the percentage of nucleic acid or amino acid residues in a candidate sequence that are identical with the nucleotide residues in the polynucleic acid being compared or the amino acid residues in the polypeptide being compared, after aligning the sequences considering any conservative substitutions as part of the sequence identity.

[0395] All ranges disclosed herein also encompass any and all possible sub-ranges and combinations of sub-ranges thereof. Any listed range can be recognized as sufficiently describing and enabling the same range being broken down into at least equal halves, thirds, quarters, fifths, tenths, and so forth. As a non-limiting example, each range discussed herein can be readily broken down into a lower third, middle third and upper third, and the like. As will also be understood by one skilled in the art all language such as “up to,” “at least,” “greater than,” “less than,” and the like include the number recited and refer to ranges which can be subsequently broken down into sub-ranges as discussed above. Finally, as will be understood by one skilled in the art, a range includes each individual member. Thus, for example, a group having 1-3 articles refers to groups having 1, 2, or 3 articles. Similarly, a group having 1-5 articles refers to groups having 1, 2, 3, 4, or 5 articles, and so forth.

[0396] Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which the recombinant polypeptides, methods and other aspects belong. Although any recombinant polypeptides, methods and other aspects similar or equivalent to those described herein can also be used in the practice or testing of the recombinant polypeptides, methods and other aspects, representative illustrative recombinant polypeptides, methods and other aspects are now described.

[0397] The term “binding domain”, as used herein, refers to a protein, or polypeptide sequence, which specifically binds to a target.

[0398] The term “humanized” antibody, as used herein, refers to an antibody in which all or substantially all CDR amino acid residues are derived from non-human CDRs and all or substantially all FR amino acid residues are derived from human FRs. A humanized antibody optionally may include at least a portion of an antibody constant region derived from a human antibody. A “humanized form” of a non-human antibody refers to a variant of the non-human antibody that has undergone humanization, typically to reduce immunogenicity to humans, while retaining the specificity and affinity of the parental non-human antibody. In some embodiments, some FR residues in a humanized antibody are substituted with corresponding residues from a non-human antibody(e.g., the antibody from which the CDR residues are derived), e.g., to restore or improve antibody specificity or affinity.

[0399] As used herein, the term “CAR-T cells” means a T cell or population thereof, which has been modified through molecular biological methods to express a chimeric antigen receptor (CAR) on the T cell surface. The CAR is a polypeptide having a pre-defined binding specificity to a desired target which is operably connected to the intracellular part of a T cell activation domain. By bypassing MHC-class I and class II restriction, CAR engineered T cells of both CD8+ and CD4+ subsets can be recruited for redirected target cell recognition. For examples, some CARs are fusions of immunoglobulin binding functionality e.g, as a single-chain variable fragment (scFv) derived from a monoclonal antibody) to CD3-zeta (CD3Q transmembrane and endodomain. Such molecules result in the transmission of a zeta signal in response to recognition by the immunoglobulin binding functionality of its target.

[0400] As used herein, a subject is “in need of’ a treatment if such subject would benefit biologically, medically or in quality of life from such treatment.

[0401] As used herein, the term “operably connected” or “operably linked” refers to positioning of components such that they function in their intended manner. For example, the components can be operably connected by a linker and / or a spacer.

[0402] As used herein, “specifically binds” means that the binding domain preferentially binds the corresponding target over other proteins. In some embodiments, “specifically binds” means that the binding domains have a higher affinity for the target than for other proteins. In some embodiments, the binding domain is a cytokine binding domain and the target is the corresponding cytokine.

[0403] As used herein, a “therapeutically effective amount” of an agent is an amount sufficient to provide a therapeutic benefit in the treatment or management of a disease or disorder, or to delay or minimize one or more symptoms associated with the disease or disorder. A therapeutically effective amount of an agent means an amount of therapeutic agent, alone or in combination with other therapeutic agents, which provides a therapeutic benefit in the treatment or management of the cancer. The term “therapeutically effective amount” can encompass an amount that improves overall therapy, reduces or avoids symptoms or causes of the disease or disorder, or enhances the therapeutic efficacy of another therapeutic agent. An example of an “effective amount” is an amount sufficient to contribute to the treatment, prevention, or reduction of a symptom or symptoms of a disease, which could also be referred to as a “therapeutically effective amount.” A “reduction” of a symptom means decreasing of the severity or frequency of the symptom(s), or elimination of the symptom(s). Theexact amount of a composition including a “therapeutically effective amount” will depend on the purpose of the treatment and will be ascertainable by one skilled in the art using known techniques.

[0404] As used herein, the term “treat,” “treating” or “treatment” of any disease or disorder refers, in one instance, to ameliorating the disease or disorder (z.e., slowing or arresting or reducing the development of the disease or at least one of the clinical symptoms thereof). In another instance, “treat”, “treating” or “treatment” refers to alleviating or ameliorating at least one physical parameter including those which may not be discernible by the patient. In yet another instance, “treat”, “treating” or “treatment” refers to modulating the disease or disorder, either physically, e.g., stabilization of a discernible symptom), physiologically, (e.g., stabilization of a physical parameter), or both.

[0405] It is appreciated that certain features of the recombinant polypeptides, and / or recombinant nucleic acids encoding the recombinant polypeptides, methods and other aspects, which are, for clarity, described in the context of separate embodiments, may also be provided in combination in a single embodiment. Conversely, various features of the recombinant polypeptides, and / or recombinant nucleic acids encoding the recombinant polypeptides, methods and other aspects, which are, for brevity, described in the context of a single embodiment, may also be provided separately or in any suitable sub-combination. All combinations of the embodiments are specifically embraced by the present disclosure and are disclosed herein just as if each and every combination was individually and explicitly disclosed, to the extent that such combinations embrace operable processes and / or compositions. In addition, all sub-combinations listed in the embodiments describing such variables are also specifically embraced by the present recombinant polypeptides, methods and other aspects and are disclosed herein just as if each and every such sub-combination was individually and explicitly disclosed herein.Recombinant Polynucleic Acid Molecules

[0406] As described above, provided herein are recombinant polynucleic acid molecules.

[0407] In some embodiments, the present disclosure describes a recombinant polynucleic acid molecule comprising a sequence encoding a polypeptide with an antigen binding domain. In another aspect, the present disclosure describes a recombinant polynucleic acid molecule comprising a sequence encoding a polypeptide with an anti-CD19 binding domain. In another aspect, the present disclosure describes a recombinant polynucleic acid molecule comprising a sequence encoding a polypeptide with an anti-CD20 binding domain. In another aspect, the present disclosure describes arecombinant polynucleic acid molecule comprising a sequence encoding a polypeptide with an anti- CD22 binding domain.

[0408] In another aspect, the present disclosure describes a recombinant polynucleic acid molecule comprising a sequence encoding a polypeptide of a chimeric antigen receptor (CAR). In another aspect, the present disclosure describes a recombinant polynucleic acid molecule comprising a sequence encoding a polypeptide of a first CAR and a second CAR. In some embodiments, the first CAR and the second CAR are separated by a protease cleavage site. In another aspect, the present disclosure describes a recombinant polynucleic acid molecule comprising a sequence encoding a polypeptide of three or more CARs. In some embodiments, the three or more CAR are separated by a protease cleavage site. In some embodiments, the protease cleavage site is a P2A or T2A selfcleaving peptide (also referred to as “viral ribosome skipping peptide”). As used herein, the terms “self-cleaving peptide” and “viral ribosome skipping peptide” refer to the same virus-derived peptides and the terms are used interchangeably herein. In another aspect, the present disclosure describes a recombinant polynucleic acid molecule comprising a sequence encoding a polypeptide of an anti-CD19 chimeric antigen receptor (CAR). In another aspect, the present disclosure describes a recombinant polynucleic acid molecule comprising a sequence encoding a polypeptide of an anti- CD20 chimeric antigen receptor (CAR). In another aspect, the present disclosure describes a recombinant polynucleic acid molecule comprising a sequence encoding a polypeptide of an anti- CD22 chimeric antigen receptor (CAR).

[0409] In some embodiments, the recombinant polynucleic acid molecule comprises a sequence encoding a polypeptide of an anti -CD 19 CAR and an anti-CD22 CAR. In some embodiments, the recombinant polynucleic acid molecule comprises a sequence encoding a polypeptide of an anti- CD20 CAR and an anti-CD22 CAR. In some embodiments, the recombinant polynucleic acid molecule comprises a sequence encoding a polypeptide of an anti-CD19 CAR and an anti-CD20 CAR. In some embodiments, the recombinant polynucleic acid molecule comprises a sequence encoding a polypeptide of an anti -CD 19 CAR, an anti-CD20 CAR, and an anti-CD22 CAR.

[0410] In some embodiments, the recombinant polynucleic acid molecule comprises a sequence from 5’ to 3’ encoding an anti-CD19 CAR and an anti-CD22 CAR. In some embodiments, the recombinant polynucleic acid molecule comprises a sequence from 5’ to 3’ encoding an anti-CD22 CAR and an anti-CD19 CAR. In some embodiments, the recombinant polynucleic acid molecule comprises a sequence from 5’ to 3’ encoding an anti-CD20 CAR and an anti-CD22 CAR. In some embodiments, the recombinant polynucleic acid molecule comprises a sequence from 5’ to 3’ encoding an anti-CD22 CAR and an anti-CD20 CAR. In some embodiments, the recombinantpolynucleic acid molecule comprises a sequence from 5’ to 3’ encoding an anti-CD19 CAR and an anti-CD20 CAR. In some embodiments, the recombinant polynucleic acid molecule comprises a sequence from 5’ to 3’ encoding an anti-CD20 CAR and an anti-CD19 CAR.

[0411] In some embodiments, the recombinant polynucleic acid molecule comprises a sequence from 5’ to 3’ encoding an anti-CD19 CAR, an anti-CD20 CAR and an anti-CD22 CAR. In some embodiments, the recombinant polynucleic acid molecule comprises a sequence from 5’ to 3’ encoding an anti-CD19 CAR, an anti-CD22 CAR and an anti-CD20 CAR. In some embodiments, the recombinant polynucleic acid molecule comprises a sequence from 5’ to 3’ encoding an anti- CD20 CAR, an anti-CD19 CAR and an anti-CD22 CAR. In some embodiments, the recombinant polynucleic acid molecule comprises a sequence from 5’ to 3’ encoding an anti-CD20 CAR, an anti- CD22 CAR and an anti-CD19 CAR. In some embodiments, the recombinant polynucleic acid molecule comprises a sequence from 5’ to 3’ encoding an anti-CD22 CAR, an anti-CD19 CAR and an anti-CD20 CAR. In some embodiments, the recombinant polynucleic acid molecule comprises a sequence from 5’ to 3’ encoding an anti-CD22 CAR, an anti-CD20 CAR and an anti-CD19 CAR.

[0412] In some embodiments, the one or more recombinant polynucleic acid molecules comprises a recombinant polynucleic acid molecule comprising a sequence encoding an anti-CD22 CAR and a sequence encoding an anti-CD19 CAR.

[0413] In some embodiments, the one or more recombinant polynucleic acid molecules comprises a recombinant polynucleic acid molecule comprising a sequence encoding an anti-CD22 CAR and a sequence encoding an anti-CD20 CAR.

[0414] In some embodiments, the one or more recombinant polynucleic acid molecules comprises a recombinant polynucleic acid molecule comprising a sequence encoding an anti-CD19 CAR and a sequence encoding an anti-CD20 CAR.

[0415] In some embodiments, the one or more recombinant polynucleic acid molecules comprises a recombinant polynucleic acid molecule comprising a sequence encoding an anti-CD22 CAR, a sequence encoding an anti-CD20 CAR, and a sequence encoding an anti-CD19 CAR.

[0416] In some embodiments, the sequence encoding an anti-CD19 CAR, the sequence encoding an anti-CD20 CAR, and the sequence encoding an anti-CD22 CAR are separated by a sequence encoding a P2A site, a T2A self-cleaving peptide, an E2A self-cleaving peptide or an F2A selfcleaving peptide.

[0417] In some embodiments, the recombinant polynucleic acid molecule comprises from 5’ to 3’ a sequence encoding an anti-CD22 CAR, a sequence encoding a P2A self-cleaving peptide, a T2A self-cleaving peptide, an E2A self-cleaving peptide or an F2A self-cleaving peptide, a sequenceencoding an anti-CD20 CAR, a sequence encoding a P2A self-cleaving peptide, a T2A self-cleaving peptide, an E2A self-cleaving peptide or an F2A self-cleaving peptide, and a sequence encoding an anti-CD19 CAR.

[0418] In some embodiments, the recombinant polynucleic acid molecule comprises from 5’ to 3’ a sequence encoding an anti-CD22 CAR, a sequence encoding a P2A self-cleaving peptide, a sequence encoding an anti-CD20 CAR, a sequence encoding a T2A self-cleaving peptide, and a sequence encoding an anti-CD19 CAR.

[0419] In some embodiments, the recombinant polynucleic acid molecule comprises from 5’ to 3’ a sequence encoding an anti-CD22 CAR, a sequence encoding a P2A self-cleaving peptide, a T2A self-cleaving peptide, an E2A self-cleaving peptide or an F2A self-cleaving peptide, a sequence encoding an anti-CD19 CAR, a sequence encoding the P2A self-cleaving peptide, and a sequence encoding an anti-CD20 CAR.

[0420] In some embodiments, the recombinant polynucleic acid molecule comprises from 5’ to 3’ a sequence encoding an anti-CD22 CAR, a sequence encoding a T2A self-cleaving peptide, a sequence encoding the anti-CD19 CAR, a sequence encoding the P2A self-cleaving peptide, and a sequence encoding an anti-CD20 CAR.

[0421] In some embodiments, the recombinant polynucleic acid molecule comprises from 5’ to 3’ a sequence encoding an anti-CD22 CAR, a sequence encoding a P2A self-cleaving peptide, a sequence encoding the anti-CD19 CAR, a sequence encoding the T2A self-cleaving peptide, and a sequence encoding an anti-CD20 CAR.

[0422] In some embodiments, the recombinant polynucleic acid molecule comprises from 5’ to 3’ a sequence encoding an anti-CD19 CAR, a sequence encoding a P2A self-cleaving peptide, a T2A self-cleaving peptide, an E2A self-cleaving peptide or an F2A self-cleaving peptide, a sequence encoding an anti-CD20 CAR, a sequence encoding a P2A self-cleaving peptide, a T2A self-cleaving peptide, an E2A self-cleaving peptide or an F2A self-cleaving peptide, and a sequence encoding an anti-CD22 CAR.

[0423] In some embodiments, the recombinant polynucleic acid molecule comprises from 5’ to 3’ a sequence encoding an anti-CD19 CAR, a sequence encoding a P2A self-cleaving peptide, a sequence encoding an anti-CD20 CAR, a sequence encoding a T2A self-cleaving peptide, and a sequence encoding an anti-CD22 CAR.

[0424] In some embodiments, the recombinant polynucleic acid molecule comprises from 5’ to 3’ a sequence encoding an anti-CD19 CAR, a sequence encoding ta P2A self-cleaving peptide, a T2A self-cleaving peptide, an E2A self-cleaving peptide or an F2A self-cleaving peptide, a sequenceencoding an anti-CD22 CAR, a sequence encoding a P2A self-cleaving peptide, a T2A self-cleaving peptide, an E2A self-cleaving peptide or an F2A self-cleaving peptide, and a sequence encoding an anti-CD20 CAR.

[0425] In some embodiments, the recombinant polynucleic acid molecule comprises from 5’ to 3’ a sequence encoding an anti-CD19 CAR, a sequence encoding a P2A self-cleaving peptide, a sequence encoding an anti-CD22 CAR, a sequence encoding a T2A self-cleaving peptide, and a sequence encoding an anti-CD20 CAR.

[0426] In some embodiments, the recombinant polynucleic acid molecule comprises from 5’ to 3’ a sequence encoding an anti-CD20 CAR, a sequence a P2A self-cleaving peptide, a T2A self-cleaving peptide, an E2A self-cleaving peptide or an F2A self-cleaving peptide, a sequence encoding an antiCD 19 CAR, a sequence encoding a P2A self-cleaving peptide, a T2A self-cleaving peptide, an E2A self-cleaving peptide or an F2A self-cleaving peptide, and a sequence encoding an anti-CD22 CAR.

[0427] In some embodiments, the recombinant polynucleic acid molecule comprises from 5’ to 3’ a sequence encoding an anti-CD20 CAR, a sequence a P2A self-cleaving peptide, a sequence encoding an anti-CD19 CAR, a sequence encoding a T2A self-cleaving peptide, and a sequence encoding an anti-CD22 CAR.

[0428] In some embodiments, the recombinant polynucleic acid molecule comprises from 5’ to 3’ a sequence encoding an anti-CD20 CAR, a sequence encoding a P2A self-cleaving peptide, a T2A self-cleaving peptide, an E2A self-cleaving peptide or an F2A self-cleaving peptide, a sequence encoding an anti-CD19 CAR, a sequence encoding a P2A self-cleaving peptide, a T2A self-cleaving peptide, an E2A self-cleaving peptide or an F2A self-cleaving peptide, and a sequence encoding an anti-CD22 CAR.

[0429] In some embodiments, the recombinant polynucleic acid molecule comprises from 5’ to 3’ a sequence encoding an anti-CD20 CAR, a sequence encoding a P2A self-cleaving peptide, a T2A self-cleaving peptide, an E2A self-cleaving peptide or an F2A self-cleaving peptide, a sequence encoding an anti-CD22 CAR, a sequence encoding a P2A self-cleaving peptide, a T2A self-cleaving peptide, an E2A self-cleaving peptide or an F2A self-cleaving peptide, and a sequence encoding an anti-CD19 CAR.

[0430] In some embodiments, the recombinant polynucleic acid molecule comprises from 5’ to 3’ a sequence encoding an anti-CD20 CAR, a sequence encoding a P2A self-cleaving peptide, a sequence encoding an anti-CD22 CAR, a sequence encoding a T2A self-cleaving peptide, and a sequence encoding an anti-CD19 CAR.

[0431] In some embodiments, the recombinant polynucleic acid molecule is a viral vector. In some embodiments, the viral vector is a lentiviral vector.Anti-CD19 binding domain

[0432] The anti-CD19 binding domain can be any molecule that binds to CD 19 with sufficient affinity and specificity, and is often an antibody or an antibody derivative, such as an scFv, single domain antibody (sdAb), Fab' fragment, (Fab')2 fragment, nanobody, diabody, or the like. As used herein, “sufficient affinity and specificity” can refer to the property that, under designated conditions, binding domain binds preferentially to its particular target protein (i.e., CD19) and does not bind in a significant amount to other proteins in a sample or subject. In certain embodiments, the anti-CD19 binding domain will have a sufficiently high binding affinity for CD 19, for example, the antibody may bind CD19 with a Kd value of between IE-07 to 10E-10 M. Antibody affinities may be determined, e.g., by a surface plasmon resonance based assay (such as the BIAcore® assay as described in PCT Application Publication No. W02005 / 012359); enzyme-linked immunoabsorbent assay (ELISA); and competition assays (e.g. radioimmunoassays (RIAs)). Alternatively, the anti- CD19 binding domain can be a receptor or a receptor fragment that binds specifically to CD 19. The anti-CD19 binding domain can be attached to the rest of the receptor directly (covalently) or indirectly (for example, through the noncovalent binding of two or more binding partners). Antibody derivatives are molecules that resemble antibodies in their mechanism of ligand binding, and include, for example, nanobodies, duobodies, diabodies, triabodies, minibodies, F(ab')2 fragments, Fab fragments, single chain variable fragments (scFv), single domain antibodies (sdAb), and functional fragments thereof. See for example, D.L. Porter et al., N Engl J Med ( 2011) 365(8): 725-33 (scFv); E.L. Smith et al, Mol Ther (2018)26(6): 1447-56 (scFv); S.R. Banihashemi et al., Iran J Basic Med Sci (2018) 21 (5):455-64 (CD19 nanobody); F. Rahbarizadeh et al Adv Drug Deliv Rev (2019) 141 :41-46 (sdAb);S.M. Kipriyanov et al., Int J Cancer (1998) 77(5):763-72 (diabody); F. Le Gall et al., FEBS Lett (1999) 453(1-2): 164-68 (triabody); M.A. Ghetie et al., Blood (1994) 83(5): 1329-36 (F(ab')2); and M.A. Ghetie et al., Clin Cancer Res (1999) 5(12):3920-27 (F(ab')2 and Fab').Antibody derivatives can also be prepared from therapeutic antibodies, for example without limitation, by preparing a nanobody, duobody, diabody, triabody, minibody, F(ab')2 fragment, Fab fragment, single chain variable fragment (scFv), or single domain antibody (sdAb) based on a therapeutic antibody. Antibody derivatives can also be designed using phage display techniques (see, e.g., E. Romao et al., Curr Pharm Des (2016) 22(43):6500- 18).

[0433] In some embodiments, the anti-CD19 binding domain comprises a heavy chain variable region (VH) that comprises a heavy chain CDR1 (HCDR1) sequence with at least 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to a sequence in Tables 5-7. In some embodiments, the anti-CD19 binding domain comprises a heavy chain variable region (VH) that comprises a heavy chain CDR1 (HCDR1) sequence in Tables 5-7. In some embodiments, the anti-CD19 binding domain comprises a heavy chain variable region (VH) that comprises a heavy chain CDR2 (HCDR2) sequence with at least 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to a sequence in Tables 5-7. In some embodiments, the anti-CD19 binding domain comprises a heavy chain variable region (VH) that comprises a heavy chain CDR2 (HCDR2) sequence in Tables 5-7. In some embodiments, the anti-CD19 binding domain comprises a heavy chain variable region (VH) that comprises a heavy chain CDR3 (HCDR3) sequence with at least 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to a sequence in Tables 5-7. In some embodiments, the anti-CD19 binding domain comprises a heavy chain variable region (VH) that comprises a heavy chain CDR3 (HCDR3) sequence in Tables 5-7.

[0434] In some embodiments, the anti-CD19 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 152, 153, and 154, respectively.

[0435] In some embodiments, the anti-CD19 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 155, 156, and 157, respectively.

[0436] In some embodiments, the anti-CD19 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 158, 159, and 160, respectively.

[0437] In some embodiments, the anti-CD19 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 161, 162, and 163, respectively.

[0438] In some embodiments, the anti-CD19 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 164, 165, and 166, respectively.

[0439] In some embodiments, the anti-CD19 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 167, 168, and 169, respectively.

[0440] In some embodiments, the anti-CD19 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 170, 171, and 172, respectively.

[0441] In some embodiments, the anti-CD19 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 173, 174, and 175, respectively.

[0442] In some embodiments, the anti-CD19 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 176, 165, and 177, respectively.

[0443] In some embodiments, the anti-CD19 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 178, 179, and 180, respectively.

[0444] In some embodiments, the anti-CD19 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 164, 165, and 181, respectively.

[0445] In some embodiments, the anti-CD19 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 176, 165, and 182, respectively.

[0446] In some embodiments, the anti-CD19 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 183, 184, and 185, respectively.

[0447] In some embodiments, the anti-CD19 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 186, 187, and 188, respectively.

[0448] In some embodiments, the anti-CD19 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 189, 190, and 191, respectively.

[0449] In some embodiments, the anti-CD19 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 192, 193, and 194, respectively.

[0450] In some embodiments, the anti-CD19 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 186, 195, and 188, respectively.

[0451] In some embodiments, the anti-CD19 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 196, 197, and 198, respectively.

[0452] In some embodiments, the anti-CD19 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 199, 200, and 201, respectively.

[0453] In some embodiments, the anti-CD19 binding domain comprises a light chain variable region (VL) that comprises a light chain CDR1 (LCDR1) sequence with at least 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to a sequence in Tables 8-10. In some embodiments, the antiCD 19 binding domain comprises a light chain variable region (VL) that comprises a light chain CDR1 (LCDR1) sequence in Tables 8-10. In some embodiments, the anti-CD19 binding domain comprises a light chain variable region (VL) that comprises a light chain CDR2 (LCDR2) sequence with at least 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to a sequence in Tables 8- 10. In some embodiments, the anti-CD19 binding domain comprises a light chain variable region (VL) that comprises a light chain CDR2 (LCDR2) sequence in Tables 8-10. In some embodiments, the anti-CD19 binding domain comprises a light chain variable region (VL) that comprises a light chain CDR3 (LCDR3) sequence with at least 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to a sequence in Tables 8-10. In some embodiments, the anti-CD19 binding domaincomprises a light chain variable region (VL) that comprises a light chain CDR3 (LCDR3) sequence in Tables 8-10.

[0454] In some embodiments, the anti-CD19 binding domain comprises an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 270, 271, and 272, respectively.

[0455] In some embodiments, the anti-CD19 binding domain comprises an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 273, 274, and 275, respectively.

[0456] In some embodiments, the anti-CD19 binding domain comprises an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 276, 277, and 278, respectively.

[0457] In some embodiments, the anti-CD19 binding domain comprises an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 279, 280, and 281, respectively.

[0458] In some embodiments, the anti-CD19 binding domain comprises an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 282, 280, and 283, respectively.

[0459] In some embodiments, the anti-CD19 binding domain comprises an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 284, 285, and 286, respectively.

[0460] In some embodiments, the anti-CD19 binding domain comprises an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 287, 274, and 288, respectively.

[0461] In some embodiments, the anti-CD19 binding domain comprises an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 289, 280, and 290, respectively.

[0462] In some embodiments, the anti-CD19 binding domain comprises an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 291, 292, and 293, respectively.

[0463] In some embodiments, the anti-CD19 binding domain comprises an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 294, 295, and 296, respectively.

[0464] In some embodiments, the anti-CD19 binding domain comprises an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 297, 298, and 299, respectively.

[0465] In some embodiments, the anti-CD19 binding domain comprises an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 300, 301, and 302, respectively.

[0466] In some embodiments, the anti-CD19 binding domain comprises an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 303, 304, and 305, respectively.

[0467] In some embodiments, the anti-CD19 binding domain comprises an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 306, 307, and 308, respectively.

[0468] In some embodiments, the anti-CD19 binding domain comprises an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 309, 292, and 310, respectively.

[0469] In some embodiments, the anti-CD19 binding domain comprises an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 311, 312, and 313, respectively.

[0470] In some embodiments, the anti-CD19 binding domain comprises an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 314, 315, and 316, respectively.

[0471] In some embodiments, the anti-CD19 binding domain comprises an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 317, 318, and 319, respectively.

[0472] In some embodiments, the anti-CD19 binding domain comprises an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 320, 321, and 322, respectively.

[0473] In some embodiments, the anti-CD19 binding domain comprises an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 323, 292, and 324, respectively.

[0474] In some embodiments, the anti-CD19 binding domain comprises an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 325, 315, and 326, respectively.

[0475] In some embodiments, the anti-CD19 binding domain comprises an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 327, 292, and 328, respectively.

[0476] In some embodiments, the anti-CD19 binding domain comprises an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 329, 330, and 331, respectively.

[0477] In some embodiments, the anti-CD19 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 152, 153, and 154, respectively, and an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 270, 271, and 272, respectively.

[0478] In some embodiments, the anti-CD19 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 155, 156, and 157, respectively, and an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 273, 274, and 275, respectively.

[0479] In some embodiments, the anti-CD19 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 158, 159, and 160, respectively, and an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 276, 277, and 278, respectively.

[0480] In some embodiments, the anti-CD19 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 161, 162, and 163, respectively, and an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 279, 280, and 281, respectively.

[0481] In some embodiments, the anti-CD19 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 161, 162, and 163, respectively, and an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 282, 280, and 283, respectively.

[0482] In some embodiments, the anti-CD19 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 161, 162, and 163, respectively, and an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 282, 280, and 283, respectively.

[0483] In some embodiments, the anti-CD19 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 161, 162, and 163, respectively, and an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 284, 285, and 286, respectively.

[0484] In some embodiments, the anti-CD19 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 164, 165, and 166, respectively, and an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 287, 274, and 288, respectively.

[0485] In some embodiments, the anti-CD19 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 161, 162, and 163, respectively, and an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 289, 280, and 290, respectively.

[0486] In some embodiments, the anti-CD19 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 167, 168, and 169, respectively, and an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 291, 292, and 293, respectively.

[0487] In some embodiments, the anti-CD19 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 170, 171, and 172, respectively, and an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 294, 295, and 296, respectively.

[0488] In some embodiments, the anti-CD19 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 173, 174, and 175, respectively, and an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 297, 298, and 299, respectively.

[0489] In some embodiments, the anti-CD19 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 176, 165, and 177, respectively, and an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 300, 301, and 302, respectively.

[0490] In some embodiments, the anti-CD19 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 178, 179, and 180, respectively, and an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 303, 304, and 305, respectively.

[0491] In some embodiments, the anti-CD19 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 164, 165, and 181, respectively, and an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 306, 307, and 308, respectively.

[0492] In some embodiments, the anti-CD19 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 176, 165, and 182, respectively, and an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 309, 292, and 310, respectively.

[0493] In some embodiments, the anti-CD19 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 183, 184, and 185, respectively, and an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 311, 312, and 313, respectively.

[0494] In some embodiments, the anti-CD19 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 186, 187, and 188, respectively, and an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 314, 315, and 316, respectively.

[0495] In some embodiments, the anti-CD19 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 189, 190, and 191, respectively, and an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 317, 318, and 319, respectively.

[0496] In some embodiments, the anti-CD19 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 192, 193, and 194, respectively and an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 320, 321, and 322, respectively.

[0497] In some embodiments, the anti-CD19 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 170, 171, and 172, respectively, and an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 323, 292, and 324, respectively.

[0498] In some embodiments, the anti-CD19 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 186, 195, and 188, respectively, and an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 325, 315, and 326, respectively.

[0499] In some embodiments, the anti-CD19 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 196, 197, and 198, respectively, and an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 327, 292, and 328, respectively.

[0500] In some embodiments, the anti-CD19 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 199, 200, and 201, respectively, and an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 329, 330, and 331, respectively.

[0501] In some embodiments, the anti-CD19 binding domain comprises a heavy chain variable region (VH) with a sequence having at least about 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 sequence set forth in Table 4. In some embodiments, the anti-CD19 binding domain comprises a heavy chain variable region (VH) with a sequence set forth in Table 4.

[0502] In some embodiments, the anti-CD19 binding domain comprises a light chain variable region (VL) with a sequence having at least about 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 sequenceset forth in Table 4. In some embodiments, the anti-CD19 binding domain comprises a light chain variable region (VL) with a sequence set forth in Table 4.

[0503] In some embodiments, the anti-CD19 binding domain comprises a VH of SEQ ID NO: 29 and a VL of SEQ ID NO: 30. In some embodiments, the anti-CD19 binding domain comprises a VH of SEQ ID NO: 31 and a VL of SEQ ID NO: 32. In some embodiments, the anti-CD19 binding domain comprises a VH of SEQ ID NO: 33 and a VL of SEQ ID NO: 34. In some embodiments, the anti-CD19 binding domain comprises a VH of SEQ ID NO: 35 and a VL of SEQ ID NO: 36. In some embodiments, the anti-CD19 binding domain comprises a VH of SEQ ID NO: 37 and a VL of SEQ ID NO: 38. In some embodiments, the anti-CD19 binding domain comprises a VH of SEQ ID NO: 37 and a VL of SEQ ID NO: 39. In some embodiments, the anti-CD19 binding domain comprises a VH of SEQ ID NO: 40 and a VL of SEQ ID NO: 41. In some embodiments, the anti- CD19 binding domain comprises a VH of SEQ ID NO: 37 and a VL of SEQ ID NO: 42. In some embodiments, the anti-CD19 binding domain comprises a VH of SEQ ID NO: 43 and a VL of SEQ ID NO: 44. In some embodiments, the anti-CD19 binding domain comprises a VH of SEQ ID NO: 45 and a VL of SEQ ID NO: 46. In some embodiments, the anti-CD19 binding domain comprises a VH of SEQ ID NO: 47 and a VL of SEQ ID NO: 48. In some embodiments, the anti-CD19 binding domain comprises a VH of SEQ ID NO: 49 and a VL of SEQ ID NO: 50. In some embodiments, the anti-CD19 binding domain comprises a VH of SEQ ID NO: 51 and a VL of SEQ ID NO: 52. In some embodiments, the anti-CD19 binding domain comprises a VH of SEQ ID NO: 53 and a VL of SEQ ID NO: 54. In some embodiments, the anti-CD19 binding domain comprises a VH of SEQ ID NO: 55 and a VL of SEQ ID NO: 56. In some embodiments, the anti-CD19 binding domain comprises a VH of SEQ ID NO: 57 and a VL of SEQ ID NO: 58. In some embodiments, the anti- CD19 binding domain comprises a VH of SEQ ID NO: 59 and a VL of SEQ ID NO: 60. In some embodiments, the anti-CD19 binding domain comprises a VH of SEQ ID NO: 61 and a VL of SEQ ID NO: 62. In some embodiments, the anti-CD19 binding domain comprises a VH of SEQ ID NO: 63 and a VL of SEQ ID NO: 64. In some embodiments, the anti-CD19 binding domain comprises a VH of SEQ ID NO: 45 and a VL of SEQ ID NO: 65. In some embodiments, the anti-CD19 binding domain comprises a VH of SEQ ID NO: 66 and a VL of SEQ ID NO: 67. In some embodiments, the anti-CD19 binding domain comprises a VH of SEQ ID NO: 68 and a VL of SEQ ID NO: 69. In some embodiments, the anti-CD19 binding domain comprises a VH of SEQ ID NO: 70 and a VL of SEQ ID NO: 71. In some embodiments, the anti-CD19 binding domain comprises a VH of SEQ ID NO: 72 and a VL of SEQ ID NO: 73.

[0504] In some embodiments, the anti-CD19 binding domain is an scFv. In some embodiments, the scFv of the anti-CD19 binding domain comprises a linker between the VH and the VL. In some embodiments, the linker is a (G4S)nlinker, where n is an integer of from 1 to 10. In some embodiments, the linker is a (G4S)s linker (z.e., wherein n=3). In some embodiments, the linker is a (G4S)e linker (z.e., wherein n=6). In some embodiments, the linker is GGGGS (SEQ ID NO: 1007; i.e., where n=l). In some embodiments, the linker is GGGGSGGGGSGGGGSGGGGSGGGGSGGGGS (SEQ ID NO: 1122; i.e., where n=6). In some embodiments, the linker comprises the sequence AAASGGGGSGGGGSGGGGSAL (SEQ ID NO: 994). In some embodiments, the VH and VL position can be switched. In some embodiments, the anti-CD19 binding domain comprises from N-terminus to C-terminus a sequence of VH-linker-VL. In some embodiments, the anti-CD19 binding domain comprises from N-terminus to C-terminus a sequence of VL-linker-VH. In some embodiments, the anti-CD19 binding domain comprises a sequence having at least about 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 sequence set forth in Table 3. In some embodiments, the anti-CD19 binding domain comprises a sequence set forth in Table 3. In some embodiments, the anti-CD19 binding domain comprises a sequence having at least about 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 the amino acid sequence of SEQ ID NO: 11. In some embodiments, the anti-CD19 binding domain comprises the amino acid sequence of SEQ ID NO: 11.

[0505] In some embodiments, the antigen binding domain is a mouse anti-human CD 19 binding domain. In some embodiments, the antigen binding domain is a chimeric mouse anti -human CD 19 binding domain. In some embodiments, the antigen binding domain is a humanized anti-human CD 19 binding domain. In some embodiments, the antigen binding domain is a fully human antihuman CD 19 binding domain.Anti-CD20 binding domain

[0506] The anti-CD20 binding domain can be any molecule that binds to CD20 with sufficient affinity and specificity, and is often an antibody or an antibody derivative, such as an scFv, single domain antibody (sdAb), Fab' fragment, (Fab')2 fragment, nanobody, diabody, or the like. As used herein, the phrase “sufficient affinity and specificity” refers to the property that, under designated conditions, binding domain binds preferentially to its particular target protein (i.e., CD20) and does not bind in a significant amount to other proteins in a sample or subject. In certain embodiments, theanti-CD19 binding domain will have a sufficiently high binding affinity for CD 19, for example, the antibody may bind CD19 with a Kd value of between IxlO'7to 6xlO'10M. Antibody affinities may be determined, e.g., by a surface plasmon resonance based assay (such as the BIAcore® assay described in PCT Application Publication No. W02005 / 012359); enzyme-linked immunoabsorbent assay (ELISA); and competition assays (e.g., radioimmunoassays (RIAs)). Alternatively, the anti- CD20 binding domain can be a receptor or a receptor fragment that binds specifically to CD20. The anti-CD20 binding domain can be attached to the rest of the receptor directly (covalently) or indirectly (for example, through the noncovalent binding of two or more binding partners). Antibody derivatives are molecules that resemble antibodies in their mechanism of ligand binding, and include, for example, nanobodies, duobodies, diabodies, triabodies, minibodies, F(ab')2 fragments, Fab fragments, single chain variable fragments (scFv), single domain antibodies (sdAb), and functional fragments thereof. See for example, D.L. Porter et al., N Engl J Med ( 2011) 365(8):725-33 (scFv); E.L. Smith et al, Mol Ther (2018)26(6): 1447-56 (scFv); S.R. Banihashemi et al., Iran J Basic Med Sci (2018) 21(5):455-64 (CD19 nanobody); F. Rahbarizadeh et al Adv Drug Deliv Rev (2019) 141 :41-46 (sdAb);S.M. Kipriyanov et al., Int J Cancer (1998) 77(5):763-72 (diabody); F. Le Gall et al., FEBS Lett (1999) 453(1-2): 164-68 (triabody); M.A. Ghetie et al., Blood (1994) 83(5): 1329-36 (F(ab')2); and M.A. Ghetie et al., Clin Cancer Res (1999) 5(12):3920-27 (F(ab')2 and Fab'). Antibody derivatives can also be prepared from therapeutic antibodies, for example without limitation, by preparing a nanobody, duobody, diabody, triabody, minibody, F(ab')2 fragment, Fab fragment, single chain variable fragment (scFv), or single domain antibody (sdAb) based on a therapeutic antibody. Antibody derivatives can also be designed using phage display techniques (see, e.g., E. Romao et al., Curr Pharm Des (2016) 22(43):6500-18).

[0507] In some embodiments, the anti-CD20 binding domain comprises a heavy chain variable region (VH) that comprises a heavy chain CDR1 (HCDR1) sequence with at least 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to a sequence in Tables 13-15. In some embodiments, the anti-CD20 binding domain comprises a heavy chain variable region (VH) that comprises a heavy chain CDR1 (HCDR1) sequence in Tables 13-15. In some embodiments, the anti-CD20 binding domain comprises a heavy chain variable region (VH) that comprises a heavy chain CDR2 (HCDR2) sequence with at least 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to a sequence in Tables 13-15. In some embodiments, the anti-CD20 binding domain comprises a heavy chain variable region (VH) that comprises a heavy chain CDR2 (HCDR2) sequence in Tables 13-15. In some embodiments, the anti-CD20 binding domain comprises a heavy chain variable region (VH) that comprises a heavy chain CDR3 (HCDR3) sequence with at least 90%, 95%, 96%, 97%, 98%,99% or 100% sequence identity to a sequence in Tables 13-15. In some embodiments, the anti-CD20 binding domain comprises a heavy chain variable region (VH) that comprises a heavy chain CDR3 (HCDR3) sequence in Tables 13-15.

[0508] In some embodiments, the anti-CD20 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 512, 513, and 514, respectively.

[0509] In some embodiments, the anti-CD20 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 515, 516, and 517, respectively.

[0510] In some embodiments, the anti-CD20 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 518, 519, and 520, respectively.

[0511] In some embodiments, the anti-CD20 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 521, 522, and 523, respectively.

[0512] In some embodiments, the anti-CD20 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 524, 525, and 526, respectively.

[0513] In some embodiments, the anti-CD20 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 527, 528, and 529, respectively.

[0514] In some embodiments, the anti-CD20 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 527, 530, and 531, respectively.

[0515] In some embodiments, the anti-CD20 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 532, 531, 1116, and 517, respectively.

[0516] In some embodiments, the anti-CD20 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 524, 533, and 534, respectively.

[0517] In some embodiments, the anti-CD20 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 527, 522, and 535, respectively.

[0518] In some embodiments, the anti-CD20 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 536, 538, and 539, respectively.

[0519] In some embodiments, the anti-CD20 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 537, 540, and 541, respectively.

[0520] In some embodiments, the anti-CD20 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 542, 538, and 539, respectively.

[0521] In some embodiments, the anti-CD20 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 543, 544, and 545, respectively.

[0522] In some embodiments, the anti-CD20 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 546, 547, and 548, respectively.

[0523] In some embodiments, the anti-CD20 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 537, 540, and 549, respectively.

[0524] In some embodiments, the anti-CD20 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 532, 550, and 551, respectively.

[0525] In some embodiments, the anti-CD20 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 543, 552, and 553, respectively.

[0526] In some embodiments, the anti-CD20 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 532, 516, and 517, respectively.

[0527] In some embodiments, the anti-CD20 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 532, 554, and 555, respectively.

[0528] In some embodiments, the anti-CD20 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 556, 557, and 558, respectively.

[0529] In some embodiments, the anti-CD20 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 518, 513, and 520, respectively.

[0530] In some embodiments, the anti-CD20 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 559, 560, and 561, respectively.

[0531] In some embodiments, the anti-CD20 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 524, 533, and 534, respectively.

[0532] In some embodiments, the anti-CD20 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 562, 513, and 563, respectively.

[0533] In some embodiments, the anti-CD20 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 536, 538, and 539, respectively.

[0534] In some embodiments, the anti-CD20 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 532, 516, and 564, respectively.

[0535] In some embodiments, the anti-CD20 binding domain comprises a light chain variable region (VL) that comprises a light chain CDR1 (LCDR1) sequence with at least 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to a sequence in Tables 16-18. In some embodiments, the anti- CD20 binding domain comprises a light chain variable region (VL) that comprises a light chain CDR1 (LCDR1) sequence in Tables 16-18. In some embodiments, the anti-CD20 binding domain comprises a light chain variable region (VL) that comprises a light chain CDR2 (LCDR2) sequence with at least 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to a sequence in Tables 16-18. In some embodiments, the anti-CD20 binding domain comprises a light chain variable region (VL) that comprises a light chain CDR2 (LCDR2) sequence in Tables 16-18. In some embodiments,-n-the anti-CD20 binding domain comprises a light chain variable region (VL) that comprises a light chain CDR3 (LCDR3) sequence with at least 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to a sequence in Tables 16-18. In some embodiments, the anti-CD20 binding domain comprises a light chain variable region (VL) that comprises a light chain CDR3 (LCDR3) sequence in Tables 16-18.

[0536] In some embodiments, the anti-CD20 binding domain comprises an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 631, 632, and 567, respectively.

[0537] In some embodiments, the anti-CD20 binding domain comprises an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 631, 632, and 567, respectively.

[0538] In some embodiments, the anti-CD20 binding domain comprises an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 633, 634, and 570, respectively.

[0539] In some embodiments, the anti-CD20 binding domain comprises an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 635, 636, and 573, respectively.

[0540] In some embodiments, the anti-CD20 binding domain comprises an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 631, 632, and 575, respectively.

[0541] In some embodiments, the anti-CD20 binding domain comprises an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 637, 636, and 577, respectively.

[0542] In some embodiments, the anti-CD20 binding domain comprises an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 639, 636, and 584, respectively.

[0543] In some embodiments, the anti-CD20 binding domain comprises an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 640, 641, and 587, respectively.

[0544] In some embodiments, the anti-CD20 binding domain comprises an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 642, 634, and 570, respectively.

[0545] In some embodiments, the anti-CD20 binding domain comprises an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 643, 641, and 590, respectively.

[0546] In some embodiments, the anti-CD20 binding domain comprises an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 644, 636, and 592, respectively.

[0547] In some embodiments, the anti-CD20 binding domain comprises an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 645, 634, and 595, respectively.

[0548] In some embodiments, the anti-CD20 binding domain comprises an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 640, 641, and 587, respectively.

[0549] In some embodiments, the anti-CD20 binding domain comprises an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 646, 647, and 599, respectively.

[0550] In some embodiments, the anti-CD20 binding domain comprises an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 648, 649, and 602, respectively.

[0551] In some embodiments, the anti-CD20 binding domain comprises an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 645, 634, and 603, respectively.

[0552] In some embodiments, the anti-CD20 binding domain comprises an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 650, 649, and 602, respectively.

[0553] In some embodiments, the anti-CD20 binding domain comprises an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 653, 647, and 609, respectively.

[0554] In some embodiments, the anti-CD20 binding domain comprises an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 654, 634, and 611, respectively.

[0555] In some embodiments, the anti-CD20 binding domain comprises an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 655, 634, and 613, respectively.

[0556] In some embodiments, the anti-CD20 binding domain comprises an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 656, 641, and 616, respectively.

[0557] In some embodiments, the anti-CD20 binding domain comprises an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 651, 652, and 618, respectively.

[0558] In some embodiments, the anti-CD20 binding domain comprises an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 642, 634, and 580, respectively.

[0559] In some embodiments, the anti-CD20 binding domain comprises an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 657, 658, and 621, respectively.

[0560] In some embodiments, the anti-CD20 binding domain comprises an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 659, 660, and 624, respectively.

[0561] In some embodiments, the anti-CD20 binding domain comprises an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 635, 636, and 575 respectively.

[0562] In some embodiments, the anti-CD20 binding domain comprises an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 661, 662, and 627, respectively.

[0563] In some embodiments, the anti-CD20 binding domain comprises an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 663, 636, and 577, respectively.

[0564] In some embodiments, the anti-CD20 binding domain comprises an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 635, 636, and 629, respectively.

[0565] In some embodiments, the anti-CD20 binding domain comprises an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 648, 649, and 602, respectively.

[0566] In some embodiments, the anti-CD20 binding domain comprises an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 663, 636, and 577, respectively.

[0567] In some embodiments, the anti-CD20 binding domain comprises an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 648, 649, and 630, respectively.

[0568] In some embodiments, the anti-CD20 binding domain comprises an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 633, 634, and 570, respectively.

[0569] In some embodiments, the anti-CD20 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 512, 513, and 514, respectively, and an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 631, 632, and 567, respectively.

[0570] In some embodiments, the anti-CD20 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 515, 516, and 517, respectively, and an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 633, 634, and 570, respectively.

[0571] In some embodiments, the anti-CD20 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 518, 519, and 520, respectively, and an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 635, 636, and 573, respectively.

[0572] In some embodiments, the anti-CD20 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 518, 513, and 520, respectively, and an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 635, 636, and 575, respectively.

[0573] In some embodiments, the anti-CD20 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 521, 522, and 523, respectively, and an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 637, 636, and 577, respectively.

[0574] In some embodiments, the anti-CD20 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 524, 525, and 526, respectively, and an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 638, 636, and 579, respectively.

[0575] In some embodiments, the anti-CD20 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 527, 528, and 529, respectively, and an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 637, 636, and 577, respectively.

[0576] In some embodiments, the anti-CD20 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 527, 530, and 531, respectively, and an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 639, 636, and 584, respectively.

[0577] In some embodiments, the anti-CD20 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 527, 530, and 531, respectively, and an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 640, 641, and 587, respectively.

[0578] In some embodiments, the anti-CD20 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 532, 531, 1116, and 517, respectively, and an LCDRl, LCDR2, and LCDR3 of SEQ ID NOs: 642, 634, and 570, respectively.

[0579] In some embodiments, the anti-CD20 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 532, 531, 1116, and 517, respectively, and an LCDRl, LCDR2, and LCDR3 of SEQ ID NOs: 643, 641, and 590, respectively.

[0580] In some embodiments, the anti-CD20 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 524, 533, and 534, respectively, and an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 644, 636, and 592, respectively.

[0581] In some embodiments, the anti-CD20 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 527, 522, and 535, respectively, and an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 645, 634, and 595, respectively.

[0582] In some embodiments, the anti-CD20 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 527, 522, and 535, respectively, and an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 640, 641, and 587, respectively.

[0583] In some embodiments, the anti-CD20 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 527, 522, and 535, respectively, and an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 646, 647, and 599, respectively.

[0584] In some embodiments, the anti-CD20 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 536, 538, and 539, respectively, and an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 648, 649, and 602, respectively.

[0585] In some embodiments, the anti-CD20 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 537, 540, and 541, respectively, and an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 648, 649, and 602, respectively.

[0586] In some embodiments, the anti-CD20 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 527, 530, and 531, respectively, and an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 640, 641, and 587, respectively.

[0587] In some embodiments, the anti-CD20 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 542, 538, and 539, respectively, and an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 645, 634, and 603, respectively.

[0588] In some embodiments, the anti-CD20 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 542, 538, and 539, respectively, and an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 650, 649, and 602, respectively.

[0589] In some embodiments, the anti-CD20 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 543, 544, and 545, respectively, and an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 651, 652, and 607, respectively.

[0590] In some embodiments, the anti-CD20 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 546, 547, and 548, respectively, and an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 653, 647, and 609, respectively.

[0591] In some embodiments, the anti-CD20 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 537, 540, and 549, respectively, and an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 654, 634, and 611, respectively.

[0592] In some embodiments, the anti-CD20 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 537, 540, and 549, respectively, and an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 655, 634, and 613, respectively.

[0593] In some embodiments, the anti-CD20 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 532, 550, and 551, respectively, and an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 656, 641, and 616, respectively.

[0594] In some embodiments, the anti-CD20 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 543, 552, and 553, respectively, and an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 651, 652, and 618, respectively.

[0595] In some embodiments, the anti-CD20 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 532, 516, and 517, respectively, and an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 642, 634, and 580, respectively.

[0596] In some embodiments, the anti-CD20 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 532, 554, and 555, respectively, and an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 657, 658, and 621, respectively.

[0597] In some embodiments, the anti-CD20 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 556, 557, and 558, respectively, and an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 659, 660, and 624, respectively.

[0598] In some embodiments, the anti-CD20 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 518, 513, and 520, respectively, and an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 635, 636, and 575, respectively.

[0599] In some embodiments, the anti-CD20 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 559, 560, and 561, respectively, and an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 661, 662, and 627, respectively.

[0600] In some embodiments, the anti-CD20 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 524, 533, and 534, respectively, and an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 663, 636, and 577, respectively.

[0601] In some embodiments, the anti-CD20 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 562, 513, and 563, respectively, and an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 635, 636, and 629, respectively.

[0602] In some embodiments, the anti-CD20 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 536, 538, and 539, respectively, and an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 648, 649, and 602, respectively.

[0603] In some embodiments, the anti-CD20 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 524, 533, and 534, respectively, and an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 663, 636, and 577, respectively.

[0604] In some embodiments, the anti-CD20 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 536, 538, and 539, respectively, and an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 648, 649, and 630, respectively.

[0605] In some embodiments, the anti-CD20 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 532, 516, and 564, respectively, and an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 633, 634, and 570, respectively.

[0606] In some embodiments, the anti-CD20 binding domain comprises a heavy chain variable region (VH) with a sequence having at least about 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 sequence set forth in Table 12. In some embodiments, the anti-CD20 binding domain comprises a heavy chain variable region (VH) with a sequence set forth in Table 12.

[0607] In some embodiments, the anti-CD20 binding domain comprises a light chain variable region (VL) with a sequence having at least about 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 sequence set forth in Table 12. In some embodiments, the anti-CD20 binding domain comprises a light chain variable region (VL) with a sequence set forth in Table 12.

[0608] In some embodiments, the anti-CD20 binding domain comprises a VH of SEQ ID NO: 373 and a VL of SEQ ID NO: 374. In some embodiments, the anti-CD20 binding domain comprises a VH of SEQ ID NO: 375 and a VL of SEQ ID NO: 376. In some embodiments, the anti-CD20 binding domain comprises a VH of SEQ ID NO: 377 and a VL of SEQ ID NO: 378. In some embodiments, the anti-CD20 binding domain comprises a VH of SEQ ID NO: 379 and a VL of SEQID NO: 380. In some embodiments, the anti-CD20 binding domain comprises a VH of SEQ ID NO: 381 and a VL of SEQ ID NO: 382. In some embodiments, the anti-CD20 binding domain comprises a VH of SEQ ID NO: 383 and a VL of SEQ ID NO: 384. In some embodiments, the anti-CD20 binding domain comprises a VH of SEQ ID NO: 385 and a VL of SEQ ID NO: 386. In some embodiments, the anti-CD20 binding domain comprises a VH of SEQ ID NO: 387 and a VL of SEQ ID NO: 388. In some embodiments, the anti-CD20 binding domain comprises a VH of SEQ ID NO: 387 and a VL of SEQ ID NO: 389. In some embodiments, the anti-CD20 binding domain comprises a VH of SEQ ID NO: 390 and a VL of SEQ ID NO: 391. In some embodiments, the anti-CD20 binding domain comprises a VH of SEQ ID NO: 390 and a VL of SEQ ID NO: 392. In some embodiments, the anti-CD20 binding domain comprises a VH of SEQ ID NO: 393 and a VL of SEQ ID NO: 394. In some embodiments, the anti-CD20 binding domain comprises a VH of SEQ ID NO: 395 and a VL of SEQ ID NO: 396. In some embodiments, the anti-CD20 binding domain comprises a VH of SEQ ID NO: 395 and a VL of SEQ ID NO: 397. In some embodiments, the anti-CD20 binding domain comprises a VH of SEQ ID NO: 395 and a VL of SEQ ID NO: 398. In some embodiments, the anti-CD20 binding domain comprises a VH of SEQ ID NO: 399 and a VL of SEQ ID NO: 400. In some embodiments, the anti-CD20 binding domain comprises a VH of SEQ ID NO: 401 and a VL of SEQ ID NO: 400. In some embodiments, the anti-CD20 binding domain comprises a VH of SEQ ID NO: 387 and a VL of SEQ ID NO: 389. In some embodiments, the anti-CD20 binding domain comprises a VH of SEQ ID NO: 402 and a VL of SEQ ID NO: 403. In some embodiments, the anti-CD20 binding domain comprises a VH of SEQ ID NO: 404 and a VL of SEQ ID NO: 405. In some embodiments, the anti-CD20 binding domain comprises a VH of SEQ ID NO: 406 and a VL of SEQ ID NO: 407. In some embodiments, the anti-CD20 binding domain comprises a VH of SEQ ID NO: 408 and a VL of SEQ ID NO: 409. In some embodiments, the anti-CD20 binding domain comprises a VH of SEQ ID NO: 408 and a VL of SEQ ID NO: 410. In some embodiments, the anti-CD20 binding domain comprises a VH of SEQ ID NO: 411 and a VL of SEQ ID NO: 412. In some embodiments, the anti-CD20 binding domain comprises a VH of SEQ ID NO: 413 and a VL of SEQ ID NO: 414. In some embodiments, the anti-CD20 binding domain comprises a VH of SEQ ID NO: 415 and a VL of SEQ ID NO: 416. In some embodiments, the anti-CD20 binding domain comprises a VH of SEQ ID NO: 417 and a VL of SEQ ID NO: 418. In some embodiments, the anti-CD20 binding domain comprises a VH of SEQ ID NO: 419 and a VL of SEQ ID NO: 420. In some embodiments, the anti-CD20 binding domain comprises a VH of SEQ ID NO: 379 and a VL of SEQ ID NO: 380. In some embodiments, the anti-CD20 binding domain comprises a VH of SEQ ID NO: 421 and a VL of SEQ ID NO: 422. In some embodiments, the anti-CD20binding domain comprises a VH of SEQ ID NO: 423 and a VL of SEQ ID NO: 424. In some embodiments, the anti-CD20 binding domain comprises a VH of SEQ ID NO: 425 and a VL of SEQ ID NO: 426. In some embodiments, the anti-CD20 binding domain comprises a VH of SEQ ID NO: 399 and a VL of SEQ ID NO: 427. In some embodiments, the anti-CD20 binding domain comprises a VH of SEQ ID NO: 399 and a VL of SEQ ID NO: 428. In some embodiments, the anti-CD20 binding domain comprises a VH of SEQ ID NO: 429 and a VL of SEQ ID NO: 430.

[0609] In some embodiments, the anti-CD20 binding domain is an scFv. In some embodiments, the scFv of the anti-CD20 binding domain comprises a linker between the VH and the VL. In some embodiments, the linker is a (G4S)nlinker, where n is an integer of from 1 to 10. In some embodiments, the linker is a (G4S)s linker. In some embodiments, the linker is a (G4S)e linker. In some embodiments, the linker is GGGGS (SEQ ID NO: 1007). In some embodiments, the linker is GGGGSGGGGSGGGGSGGGGSGGGGSGGGGS (SEQ ID NO: 1122). In some embodiments, the linker comprises the sequence GGGGS GGGGS GGGGS (i.e., n=3; SEQ ID NO: 995). The VH and VL position can be switched. In some embodiment, the anti-CD20 binding domain comprises from N-terminus to C-terminus a sequence of VH-linker-VL. In some embodiment, the anti-CD20 binding domain comprises from N-terminus to C-terminus a sequence of VL-linker- VH.

[0610] In some embodiments, the anti-CD20 binding domain comprises a sequence having at least about 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 sequence set forth in Table 11. In some embodiments, the anti-CD20 binding domain comprises a sequence set forth in Table 11. In some embodiments, the anti-CD20 binding domain comprises a sequence having at least about 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 the amino acid sequence of SEQ ID NO: 363. In some embodiments, the anti-CD20 binding domain comprises the amino acid sequence of SEQ ID NO: 363.

[0611] In some embodiments, the antigen binding domain is a mouse anti-human CD20 binding domain. In some embodiments, the antigen binding domain is a chimeric mouse anti-human CD20 binding domain. In some embodiments, the antigen binding domain is a humanized anti-human CD20 binding domain. In some embodiments, the antigen binding domain is a fully human antihuman CD20 binding domain.Anti-CD22 binding domain

[0612] The anti-CD22 binding domain can be any molecule that binds to CD22 with sufficient affinity and specificity, and is often an antibody or an antibody derivative, such as an scFv, single domain antibody (sdAb), Fab' fragment, (Fab')2 fragment, nanobody, diabody, or the like. As used herein, “sufficient affinity and specificity” can refer to the property that, under designated conditions, binding domain binds preferentially to its particular target protein (i.e., CD22) and does not bind in a significant amount to other proteins in a sample or subject. In certain embodiments, the anti-CD22 binding domain will have a sufficiently high binding affinity for CD22, for example, the antibody may bind CD22 with a Kd value of between IxlO'7M to 10xl0'10M. Antibody affinities may be determined, e.g., by a surface plasmon resonance based assay (such as the BIAcore® assay as described in PCT Application Publication No. W02005 / 012359); enzyme-linked immunoabsorbent assay (ELISA); and competition assays (e.g. radioimmunoassays (RIAs)). Alternatively, the anti- CD22 binding domain can be a receptor or a receptor fragment that binds specifically to CD22. The anti-CD22 binding domain can be attached to the rest of the receptor directly (covalently) or indirectly (for example, through the noncovalent binding of two or more binding partners). Antibody derivatives are molecules that resemble antibodies in their mechanism of ligand binding, and include, for example, nanobodies, duobodies, diabodies, triabodies, minibodies, F(ab')2 fragments, Fab fragments, single chain variable fragments (scFv), single domain antibodies (sdAb), and functional fragments thereof. See for example, D.L. Porter et al., N Engl J Med ( 2011) 365(8): 725-33 (scFv); E.L. Smith et al, Mol Ther (2018)26(6): 1447-56 (scFv); S.R. Banihashemi et al., Iran J Basic Med Sci (2018) 21 (5):455-64 (CD19 nanobody); F. Rahbarizadeh et al Adv Drug Deliv Rev (2019) 141 :41-46 (sdAb);S.M. Kipriyanov et al., Int J Cancer (1998) 77(5):763-72 (diabody); F. Le Gall et al., FEBS Lett (1999) 453(1-2): 164-68 (triabody); M.A. Ghetie et al., Blood (1994) 83(5): 1329-36 (F(ab')2); and M.A. Ghetie et al., Clin Cancer Res (1999) 5(12):3920-27 (F(ab')2 and Fab').Antibody derivatives can also be prepared from therapeutic antibodies, for example without limitation, by preparing a nanobody, duobody, diabody, triabody, minibody, F(ab')2 fragment, Fab fragment, single chain variable fragment (scFv), or single domain antibody (sdAb) based on a therapeutic antibody. Antibody derivatives can also be designed using phage display techniques (see, e.g., E. Romao et al., Curr Pharm Des (2016) 22(43):6500- 18).

[0613] In some embodiments, the anti-CD22 binding domain comprises a heavy chain variable region (VH) that comprises a heavy chain CDR1 (HCDR1) sequence with at least 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to a sequence in Table 19. In some embodiments, the anti-CD22 binding domain comprises a heavy chain variable region (VH) that comprises a heavy chain CDR1 (HCDR1) sequence in Table 19. In some embodiments, the anti-CD22 binding domaincomprises a heavy chain variable region (VH) that comprises a heavy chain CDR2 (HCDR2) sequence with at least 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to a sequence in Tables 19. In some embodiments, the anti-CD22 binding domain comprises a heavy chain variable region (VH) that comprises a heavy chain CDR2 (HCDR2) sequence in Table 19. In some embodiments, the anti-CD22 binding domain comprises a heavy chain variable region (VH) that comprises a heavy chain CDR3 (HCDR3) sequence with at least 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to a sequence in Table 19. In some embodiments, the anti-CD22 binding domain comprises a heavy chain variable region (VH) that comprises a heavy chain CDR3 (HCDR3) sequence in Table 19.

[0614] In some embodiments, the anti-CD22 binding domain comprises a light chain variable region (VL) that comprises a light chain CDR3 (LCDR1) sequence with at least 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to a sequence in Table 19. In some embodiments, the anti- CD22 binding domain comprises a light chain variable region (VL) that comprises a heavy chain CDR1 (LCDR1) sequence in Tables 19. In some embodiments, the anti-CD22 binding domain comprises a light chain variable region (VL) that comprises a light chain CDR2 (LCDR2) sequence with at least 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to a sequence in Table 19. In some embodiments, the anti-CD22 binding domain comprises a light chain variable region (VL) that comprises a heavy chain CDR2 (LCDR2) sequence in Table 19. In some embodiments, the anti- CD22 binding domain comprises a light chain variable region (VL) that comprises a light chain CDR3 (LCDR3) sequence with at least 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to a sequence in Table 19. In some embodiments, the anti-CD22 binding domain comprises a light chain variable region (VL) that comprises a heavy chain CDR3 (LCDR3) sequence in Table 19.

[0615] In some embodiments, the anti-CD20 binding domain comprises an HCDR1, HCDR2, and HCDR3 of SEQ ID NOs: 667, 675, and 676, respectively, and an LCDR1, LCDR2, and LCDR3 of SEQ ID NOs: 677, 678, and 672, respectively.

[0616] In some embodiments, the anti-CD22 binding domain comprises a heavy chain variable region (VH) with a sequence having at least about 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 the sequence QVQLQQSGPGLVKPSQTLSLTCAISGDSVSSNSAAWNWIRQSPSRGLEWLGRTYYRSKWYN DYAVSVKSRITINPDTSKNQFSLQLNSVTPEDTAVYYCAREVTGDLEDAFDIWGQGTMVTV SS (SEQ ID NO: 665). In some embodiments, the anti-CD22 binding domain comprises a heavy chain variable region (VH) with the sequenceQVQLQQSGPGLVKPSQTLSLTCAISGDSVSSNSAAWNWIRQSPSRGLEWLGRTYYRSKWYN DYAVSVKSRITINPDTSKNQFSLQLNSVTPEDTAVYYCAREVTGDLEDAFDIWGQGTMVTV SS (SEQ ID NO: 665).

[0617] In some embodiments, the anti-CD22 binding domain comprises a light chain variable region (VL) with a sequence having at least about 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 the sequence DIQMTQSPSSLSASVGDRVTITCRASQTIWSYLNWYQQRPGKAPNLLIYAASSLQSGVPSRFS GRGSGTDFTLTISSLQAEDFATYYCQQSYSIPQTFGQGTKLEIK (SEQ ID NO: 666). In some embodiments, the anti-CD22 binding domain comprises a light chain variable region (VL) with the sequenceDIQMTQSPSSLSASVGDRVTITCRASQTIWSYLNWYQQRPGKAPNLLIYAASSLQSGVPSRFS GRGSGTDFTLTISSLQAEDFATYYCQQSYSIPQTFGQGTKLEIK (SEQ ID NO: 666).

[0618] In some embodiments, the anti-CD22 binding domain is an scFv. In some embodiments, the scFv of the anti-CD22 binding domain comprises a linker between the VH and the VL and comprises the sequence GGGGS. In some embodiments, the anti-CD22 binding domain comprises a sequence having at least about 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 the sequenceQVQLQQSGPGLVKPSQTLSLTCAISGDSVSSNSAAWNWIRQSPSRGLEWLGRTYYRSKWYN DYAVSVKSRITINPDTSKNQFSLQLNSVTPEDTAVYYCAREVTGDLEDAFDIWGQGTMVTV SSGGGGSDIQMTQSPSSLSASVGDRVTITCRASQTIWSYLNWYQQRPGKAPNLLIYAASSLQ SGVPSRFSGRGSGTDFTLTISSLQAEDFATYYCQQSYSIPQTFGQGTKLEIK (SEQ ID NO: 664). In some embodiments, the anti-CD22 binding domain comprises the sequenceQVQLQQSGPGLVKPSQTLSLTCAISGDSVSSNSAAWNWIRQSPSRGLEWLGRTYYRSKWYN DYAVSVKSRITINPDTSKNQFSLQLNSVTPEDTAVYYCAREVTGDLEDAFDIWGQGTMVTV SSGGGGSDIQMTQSPSSLSASVGDRVTITCRASQTIWSYLNWYQQRPGKAPNLLIYAASSLQ SGVPSRFSGRGSGTDFTLTISSLQAEDFATYYCQQSYSIPQTFGQGTKLEIK (SEQ ID NO: 664).

[0619] In some embodiments, the antigen binding domain is a mouse anti-human CD22 binding domain. In some embodiments, the antigen binding domain is a chimeric mouse anti-human CD22 binding domain. In some embodiments, the antigen binding domain is a humanized anti-human CD22 binding domain. In some embodiments, the antigen binding domain is a fully human antihuman CD22 binding domain.Chimeric Antisen Receptors

[0620] The anti-CD19, anti-CD20, and anti-CD22 binding domains as described herein can be provided in the form of a chimeric antigen receptor (CAR). CARs comprising the binding domains of the present disclosure can include, without limitation, a hinge domain, a spacer domain, a transmembrane domain, and a cytoplasmic domain. In some embodiments, such CARs optionally further comprise one or more spacer or linker domains disposed between certain of the other domains comprising the CAR. In some embodiments, when a hinge domain, a spacer domain, a transmembrane domain, and / or a cytoplasmic domain is “derived from” a recited protein or sequence, it can mean that the hinge domain, spacer domain, transmembrane domain, and / or cytoplasmic domain is the corresponding domain from the recited protein or sequence.Hinge and Spacer Domains

[0621] In some embodiments, the antigen binding domain of a CAR provided herein is operatively linked to a transmembrane domain by a hinge or a spacer domain. The hinge domain is generally a flexible polypeptide connector region disposed between the targeting moiety (z.e., the antigen binding scFv domain) and the transmembrane domain. In certain embodiments, the hinge or spacer domain is a portion of an immunoglobulin, including, but not limited to, one or more heavy chain constant regions, e.g., CH2 and CH3. The spacer domain may include the amino acid sequence of a naturally occurring immunoglobulin hinge region or an altered immunoglobulin hinge region. In some embodiments, the spacer domain includes the CH2 and / or CH3 of IgGl, lgG4, or IgD. Illustrative spacer domains suitable for use in the CARs described herein include the hinge domain derived from the extracellular portion of type 1 membrane proteins such as CD8a and CD28, which may be wild-type hinge domains from these molecules or variants thereof. In some embodiments, the term “derived from” can mean that the hinge domain is the extracellular portion of type 1 membrane proteins such as CD8a and CD28, for example. In certain aspects, the hinge domain comprises a CD8a or CD28 hinge domain. In some embodiments, the hinge is a PD-1 hinge or CD 152 hinge.

[0622] In some embodiments, the CAR further includes an extracellular spacer domain, which may include a hinge domain. The hinge domain is generally a flexible polypeptide connector domain comprising a short sequence of amino acid sequences disposed between the targeting moiety and the transmembrane domain. Exemplary hinge domain sequences include those from IgG subclasses (such as IgGl and IgG4), IgD, CD28, and CD8a domains. The hinge domain provides structural flexibility to flanking polypeptide regions. In some embodiments, the hinge domain may consist of natural or synthetic polypeptides. It will be appreciated by those skilled in the art that hinge domains may improve the function of the CAR by promoting optimal positioning of the antigen bindingdomain in relation to the portion of the antigen recognized by it. In some embodiments, a hinge domain may not be required for optimal CAR activity. In some embodiments, a hinge domain comprising a short sequence of amino acids that promotes CAR activity by facilitating antigenbinding, for example, by relieving steric constraints that could otherwise alter antibody binding kinetics. In some embodiments, the hinge domain is linked downstream of the antigen-binding domain of a CAR and upstream of the transmembrane domain of a CAR.

[0623] Non-limiting examples of suitable hinge domains include those derived from CD8a, CD28, CTLA4, CD4, PD1, IgGl, PGK, IgG4, or IgD. In some embodiments, the term “derived from” can mean that the hinge domain is from CD8a, CD28, CTLA4, CD4, PD1, IgGl, PGK, IgG4, or IgD. In some embodiments, the hinge domain can include regions derived from a human CD8a molecule, a CD28 molecule, and any other receptors that provide a similar function in providing flexibility to flanking regions. In some embodiments, the CAR disclosed herein includes a hinge domain derived from a CD8a hinge domain. In some embodiments, the CAR disclosed herein includes a hinge domain derived from a CD28 hinge domain. In some embodiments, the hinge domain has about 70, 75, 80, 85, 90, 92, 93, 94, 95, 96, 97, 98, 99 or about 100% sequence identity to a CD8a (SEQ ID NO:679), CD28 (SEQ ID NO: 682), CTLA4 (SEQ ID NO: 1347), CD4 (SEQ ID NO: 1349), PD1 (SEQ ID NO: 1349), IgGl (SEQ ID NO: 1350), PGK, IgG4 (SEQ ID NO: 1352), or IgD hinge domain (SEQ ID NO: 1354).

[0624] In some embodiments, the anti-CD22 CAR comprises a hinge domain derived from CD8a. In some embodiments, the anti-CD19 CAR comprises a hinge domain derived from CD28. In some embodiments, the anti-CD20 CAR comprises a hinge domain derived from CD28.

[0625] In some embodiments, the spacer domain further comprises a linker including one or more intervening amino acid residues that are positioned between the antigen binding domain and the extracellular hinge domain. In some embodiments, the linker is positioned downstream from the antigen binding domain and upstream from the hinge domain. In principle, there are no particular limitations to the length and / or amino acid composition of the linker. In some embodiments, any arbitrary single-chain peptide comprising about one to about 300 amino acid residues (e.g., 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, or more amino acid residues) can be used as a linker. In some embodiments, the linker includes at least about 5, 6, 7, 8, 9, 10, 11, 12, 15, 20, 25, 30, 35, 40, 45, or 50 amino acids. In some embodiments, the linker includes no more than about 300, 250, 200, 150, 140, 130, 120, 110, 100, 90, 85, 80, 75, 70, 65, 60, 55, 50, 45, 40, 35, or 30 amino acid residues. In some embodiments, the length and amino acid composition of the extracellular spacer can be optimized to vary the orientation and / or proximity of the antigen binding domain andthe extracellular hinge domain to one another to achieve a desired activity of the CAR. In some embodiments, the orientation and / or proximity of the antigen binding domain and the extracellular hinge domain to one another can be varied and / or optimized as a “tuning” tool or effect to enhance or reduce the efficacy of the CAR. In some embodiments, the orientation and / or proximity of the antigen binding domain and the hinge domain to one another can be varied and / or optimized to create a partially functional version of the CAR. In some embodiments, the extracellular spacer domain includes an amino acid sequence corresponding to an IgG4 hinge domain and an IgG4 CH2- CH3 domain (SEQ ID NOs: 1351 and 1353).

[0626] Alternatively, the spacer domain can be a synthetic polypeptide spacer, such as a spacer having a random sequence, a (gly-gly-ser)n(“GGSn”) sequence, or a variation thereof such as (SGG)n, (GGGS)n, (GGGGS)n, (SGGG)n, (GSGGG)n, and the like, where n is an integer ranging from about 1 to about 15. The synthetic polypeptide spacer domain can also include a naturally occurring sequence, such as a hinge domain derived from CD8a, IgG, and the like.Transmembrane Domains

[0627] In some embodiments, the extracellular domain of the CAR is operably connected to the transmembrane domain. In some embodiments, the extracellular domain is connected to the transmembrane domain by a spacer or a hinge domain or a spacer and a hinge domain. The transmembrane domain of the CAR serves to transduce the external signal received by the extracellular domain to the cytoplasmic domain. The transmembrane domain can be any transmembrane domain known in the art, including but not limited to, a CD3(^ transmembrane domain, a CD28 transmembrane domain, a CD8a transmembrane domain, a CD8a hinge / transmembrane domain (CD8H / TM), and a CD28 hinge / CD28 transmembrane domain (CD28H / TM). The transmembrane domain can be selected from a transmembrane region of a transmembrane protein such as, for example, Type I transmembrane proteins, an artificial hydrophobic sequence or a combination thereof. Examples of the transmembrane domain include the transmembrane regions of the alpha, beta or zeta chain of the T cell receptor, CD28, CD3 epsilon, CD45, CD4, CD5, CD8a, CD9, CD16, CD22, CD33, CD37, CD64, CD80, CD86, CD134, CD137, CD 154. Synthetic transmembrane domains may comprise a triplet of phenylalanine, tryptophan and valine. Optionally, a short oligo- or polypeptide linker, preferably between 2 and 10 amino acids in length, may form the linkage between the transmembrane domain and the intracellular signaling domain of the CAR. A glycine-serine doublet provides a particularly suitable linker between the transmembrane domain and the intracellular signaling domain.

[0628] In some embodiments, the CAR comprises a transmembrane domain derived from a polypeptide selected from the group consisting of: CD4, CD8a, CD28, CD154, and PD-1. In some embodiments, the term “derived from” can mean that the transmembrane domain is from the group consisting of: CD4, CD8a, CD28, CD154, and PD-1. Such a CAR may further include a spacer domain between the antigen-binding portion and the transmembrane domain, e.g., a CD8a hinge.

[0629] In some embodiments, the CAR comprises a transmembrane domain from CD28. In some embodiments, the CAR comprises a transmembrane domain with the sequence FWVLVVVGGVLACYSLLVTVAFIIFWV (SEQ ID NO: 683).

[0630] In some embodiments, the CAR comprises a transmembrane domain from CD8a. In some embodiments, the CAR comprises a transmembrane domain with the sequence IWAPLAGTCGVLLLSLVITLYC (SEQ ID NO: 691). In some embodiments, the CAR comprises a transmembrane domain from CD8a. In some embodiments, the CAR comprises a transmembrane domain with the sequence IYIWAPLAGTCGVLLLSLVITLYC (SEQ ID NO: 1117).

[0631] The transmembrane domain may be derived either from a natural, synthetic, semi -synthetic, or recombinant source. In some embodiments, the transmembrane domain is derived from (e.g., includes at least the transmembrane region(s) or a functional portion thereof) of the alpha or beta chain of the T cell receptor, CD3y, CD35, CD3s, CD35, CD3< CD4, CD5, CD8a, CD9, CD16, CD22, CD27, CD28, CD33, CD37, CD45, CD64, CD80, CD86, CD134, CD137, CD152, CD154, and / or PD-1. In some embodiments, the term “derived from” can mean that the transmembrane domain is from the alpha or beta chain of the T cell receptor, CD3y, CD35, CD3s, CD35, CD3(^, CD4, CD5, CD8a, CD9, CD16, CD22, CD27, CD28, CD33, CD37, CD45, CD64, CD80, CD86, CD134, CD137, CD152, CD154, and / or PD-1..

[0632] The transmembrane domain may include, for example without limitation, all or part of the transmembrane domain of the CD3(^ chain), CD28, CD2, CD4, 0X40, 4-1BB (CD137), ICOS (CD278), ILRB (CD122), IL-2RG (CD132), CTLA-4, PD-1, or CD40, or a sequence derived from such a transmembrane domain. The cytoplasmic signaling domain in general comprises a domain that transduces the event of ligand binding into an intracellular signal that activates the T cell. The CD3(^ cytoplasmic domain / activating domain is frequently used, although others such as MyD88 can be used. In an embodiment, the transmembrane domain is the transmembrane domain from CD3(^, CD2, CD8a, or CD28. In an embodiment, the transmembrane domain is derived from the transmembrane domain from CD2 or CD28. In some embodiments, the transmembrane domain has about 70, 75, 80, 85, 90, 92, 93, 94, 95, 96, 97, 98, 99 or about 100% sequence identity to a CD3(^ (SEQ ID NO: 1337), CD28 (SEQ ID NO: 683), CD2 (SEQ ID NO: 1338), CD4 (SEQ ID NO: 1339),0X40 (SEQ ID NO: 1340), 4-1BB (CD137) (SEQ ID NO: 1341), FcERIy (SEQ ID NO: 1342), ICOS (CD278) (SEQ ID NO: 1343), ILRB (CD122) (SEQ ID NO: 1344), IL-2RG (CD132) (SEQ ID NO: 1345), or CD40 transmembrane domain (SEQ ID NO: 1346).

[0633] According to some embodiments, a CAR includes a transmembrane domain derived from CD8a or CD28 and a short polypeptide linker, e.g., between 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acids in length, that links the transmembrane domain and the intracellular signaling domain of the CAR. A glycine-serine linker may be employed as such a linker, for example.

[0634] In some embodiments, the CAR comprises a CD8a hinge domain and a CD28 transmembrane domain. In some embodiments, the CAR comprises a CD8a hinge domain and a CD8a transmembrane domain. In some embodiments, the CAR comprises a CD28 hinge domain and a CD28 transmembrane domain. In some embodiments, the CAR comprises a CD28 hinge domain and a CD8a transmembrane domain. In some embodiments, the CAR comprises an IgG4 hinge domain and a CD28 transmembrane domain. In some embodiments, the CAR comprises an IgG4 hinge domain and a CD2 transmembrane domain.Cytoplasmic Domains

[0635] In some embodiments, the transmembrane domain of the CAR is operably connected to the cytoplasmic domain. The cytoplasmic domain transduces the received external signal to initiate the downstream signaling cascade. In some embodiments, the cytoplasmic domain comprises an intracellular signaling domain. Signals generated through the T cell receptor (TCR) alone may be insufficient for full activation of the T cell and a secondary or costimulatory signal may also be required. Full T cell activation is mediated by two distinct classes of intracellular signaling domains: primary signaling domains that in native T cells (z.e., non-CAR-T cells) initiate antigen-dependent primary T cell 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. In some embodiments, the CAR may include an intracellular signaling domain comprising a primary signaling domain and one or more costimulatory signaling domains. In some embodiments, the cytoplasmic domain comprises one or more intracellular signaling domains selected from the group consisting of CD3< 4-1BB (CD137) CD28, ICOS, FcyRI, FcRy, FcR, CD3y, CD35, CD3s, CD35, CD22, CD79a, CD79b, CD665, TLR1, TLR2, TLR3, TLR4, TLR5, TLR6, TLR7, TLR8, TLR9, TLR10, CARD11, CD2, CD7, CD27, CD30, CD40, CD54 (ICAM), CD83, CD134 (0X40), CD137 (4-1BB), CD278 (ICOS), DAP10, LAT, KD2C, SLP76, TRIM, and ZAP70.

[0636] Primary signaling domains regulate primary activation of the TCR complex either in a stimulatory manner, or in an inhibitory manner. Primary signaling domains that act in a stimulatorymanner may contain signaling motifs which are known as immunoreceptor tyrosine-based activation motifs (or “IT AMs”). Non-limiting examples of ITAM-containing primary signaling domains suitable for use in a CAR of the present disclosure include those derived from FcyRI, FcRy, FcR, CD3y, CD35, CD3s, CD3< CD35, CD22, CD79a, CD79b, and CD665. In some embodiments, the term “derived from” can mean that the transmembrane domain is from FcyRI, FcRy, FcR, CD3y, CD35, CD3s, CD3< CD35, CD22, CD79a, CD79b, and CD665. In some embodiments, the intracellular signaling domain comprises one or more ITAM-containing primary signaling domains. In some embodiments, the one or more ITAM-containing primary signaling domains comprises a CD3(^ signaling domain.

[0637] In some embodiments, the CAR comprises a cytoplasmic domain comprising an intracellular signaling domain from CD3(^. In some embodiments, the CAR comprises a cytoplasmic domain comprising an intracellular signaling domain from CD3(^ having the sequence RVKFSRSADAPAYKQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLY NELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 686). In some embodiments, the CAR comprises a cytoplasmic domain comprising an intracellular signaling domain from CD3(^ having the sequence RVKFSRSADAPAYQQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLY NELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 687).

[0638] In some embodiments, the CAR comprises a cytoplasmic domain comprising an intracellular signaling domain from CD3(^. In some embodiments, the CAR comprises a cytoplasmic domain comprising an intracellular signaling domain from CD3(^ having the sequence RVKFSRSADAPAYQQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPQRRKNPQEGL YNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 688). In some embodiments, the CAR comprises a cytoplasmic domain comprising an intracellular signaling domain from CD3(^ having the sequence RVKFSRSADAPAYKQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPQRRKNPQEGL YNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 689).

[0639] In some embodiments, the CAR comprises a cytoplasmic domain comprising an intracellular signaling domain from CD3s. In some embodiments, the CAR comprises a cytoplasmic domain comprising an intracellular signaling domain from CD3s having the sequenceRPPPVPNPDYEPIRKGQRDLYSGLNQRRI (SEQ ID NO: 1003). In some embodiments, the CAR comprises a cytoplasmic domain comprising a truncated CD3s cytoplasmic domain.

[0640] In some embodiments, the CAR comprises one or more costimulatory intracellular signaling domains to enhance the efficacy, persistence, and / or expansion of T cells expressing the CAR. Costimulatory proteins are found in at least the following protein families: TNF receptor proteins, immunoglobulin-like proteins, cytokine receptors, integrins, signaling lymphocytic activation molecules (SLAM proteins), and activating NK cell receptors. Examples of such molecules include CD27, CD28, 4-1BB (CD137), 0X40, GITR, CD30, CD40, ICOS, BAFFR, HVEM, lymphocyte function-associated antigen- 1 (LFA-1), CD2, CD7, LIGHT, NKG2C, SLAMF7, NKp80, CD 160, B7-H3, and CD83 ligand (CD83L), a ligand that specifically binds with CD83. Exemplary costimulatory molecules comprising costimulatory intracellular signaling domains suitable for use in CARs contemplated in particular embodiments include TLR1, TLR2, TLR3, TLR4, TLR5, TLR6, TLR7, TLR8, TLR9, TLR10, CARD11, CD2, CD7, CD27, CD28, CD30, CD40, CD54 (ICAM), CD83, CD134 (0X40), CD137 (4-1BB), CD278 (ICOS), DAP10, LAT, KD2C, SLP76, TRIM, and / or ZAP70. In some embodiments, the costimulatory signaling domain has at least about 70, 75, 80, 85, 90, 92, 93, 94, 95, 96, 97, 98, 99 or 100% sequence identity to a costimulatory signaling domain from TLR1 (SEQ ID NO: 1316), TLR2 (SEQ ID NO: 1317), TLR3 (SEQ ID NO: 1318), TLR4 (SEQ ID NO: 1319), TLR5 (SEQ ID NO: 1320), TLR6 (SEQ ID NO: 1321), TLR7 (SEQ ID NO: 1322), TLR8 (SEQ ID NO: 1323), TLR9 (SEQ ID NO: 1324), TLR10 (SEQ ID NO: 1325), CARD1 1 (SEQ ID NO: 1326), CD2 (SEQ ID NO: 685), CD7 (SEQ ID NO: 1327), CD27 (SEQ ID NO: 1356), CD28 (SEQ ID NO: 684), CD30 (SEQ ID NO: 1328), CD40 (SEQ ID NO: 1329), CD54 (ICAM) (SEQ ID NO: 1330), CD83 (SEQ ID NO: 1331), CD134 (0X40), CD137 (4-1BB), CD278 (ICOS) (SEQ ID NO: 1357), DAP10 (SEQ ID NO: 1332), LAT (SEQ ID NO: 1333), KD2C, SLP76 (SEQ ID NO: 1334), TCRIM (SEQ ID NO: 1335), and / or ZAP70 (SEQ ID NO: 1336) domain. In some embodiments, a CAR includes one or more co-stimulatory signaling domains selected from the group consisting of CD2, 4-1BB, CD28, CD137, and CD134, and a CD3(^ primary signaling domain. In some embodiments, a CAR includes one or more co-stimulatory signaling domains selected from the group consisting of CD27, CD28, 4-1BB / CD137, 0X40, ICOS, and CD2.

[0641] Tumor cells can escape CAR-T cell destruction through the downregulation of signaling molecules that serve as co-stimulators of T-cell activation. These include CD58, the ligand of the CD2 costimulatory receptor. CD58 alteration is associated with poor prognosis in large B cell lymphoma (LBCL) and leads to lack of response to treatment with CD 19 CAR-T cells. As shown in Figure 7, incorporating a CD2 co-stimulatory signaling domain in a CAR construct may improveCAR-T cell efficacy, especially in treating CD58 downregulated tumors, which may be found in patients having LBCL, including relapsed or refractory LBCL following treatment with CD 19- specific CAR-T cells. In some embodiments, the CAR comprises a cytoplasmic domain comprising an intracellular signaling domain from CD2.

[0642] In some embodiments, the CAR comprises a cytoplasmic domain comprising an intracellular signaling domain comprising an intracellular signaling domain from CD3^ (SEQ ID NOs: 686-689) and a co-stimulatory intracellular signaling domain from CD2 having the sequence KRKKQRSRRNDEELETRAHRVATEERGRKPHQIPASTPQNPATSQHPPPPPGHRSQAPSHRP PPPGHRVQHQPQKRPPAPSGTQVHQQKGPPLPRPRVQPKPPHGAAENSLSPSSN (SEQ ID NO: 685). In some embodiments, the CAR comprises a cytoplasmic domain comprising an intracellular signaling domain from CD3^ and a costimulatory intracellular signaling domain from CD2 having the sequence KRKKQRSRRNDEELETRAHRVATEERGRKPHQIPASTPQNPAT (SEQ ID NO: 999). PATSQHPPPPPGHRSQAPSHRPPPPGHRVQH (SEQ ID NO: 1002)

[0643] In some embodiments, the CAR comprises a cytoplasmic domain comprising an intracellular signaling domain from CD3(^ and an intracellular costimulatory signaling domain from 4-1BB (CD137). In some embodiments, the CAR comprises a cytoplasmic domain comprising an intracellular costimulatory signaling domain from 4-1BB (CD137). In some embodiments, the CAR comprises a cytoplasmic domain comprising an intracellular signaling domain comprising a costimulatory intracellular signaling domain from 4-1BB (CD137) having the sequence KRGRKKLLYIFKQPFMRPVQTTQEEDGCSCRFPEEEEGGCEL (SEQ ID NO: 998).

[0644] CARs of the disclosure may comprise a CD2 co-stimulatory signaling domain, and one or more additional co-stimulatory signaling domains to increase cytokine production or sensitivity, reduce or prevent anergy, and / or to increase proliferation and cytotoxic activity. These additional co- stimulatory signaling domains can be derived from co-stimulatory proteins such as B7-1 (CD80), B7-2 (CD86), CTLA-4, PD-1, CD278, CD122, CD132, B7- H2, B7-H3, PD-L1, PD-L2, B7-H4, PDCD6, BTLA, 41BB (CD137), FcERTy, CD40L, 4- 1BBL, GITR, BAFF, GITR-L, BAFF-R, HVEM, CD27, LIGHT, CD27L, 0X40, OX40L, CD30, CD30L, TAC1, CD40, CD244, CD84, BLAME, CD229, CRACC, CD2F-10, NTB-A, CD48, SLAM (CD150), CD58, ikaros, CD53, integrin a4, CD82, integrin a4bl, CD90, integrin a4b7, CD96, LAG-3, CD 160, LMIR, CRTAM, TCL1A, DAP12; TIM-1, Dectin-1, TIM-4, TSLP, EphB6, TSLP-R, and / or HL A-DR. In some embodiments, the cytoplasmic signaling domain has about 70, 75, 80, 85, 90, 92, 93, 94, 95, 96, 97, 98, 99 or about 100% sequence identity to an B7-1 (CD80), B7-2 (CD86), CTLA-4, PD-1, CD278, CD122, CD132, B7- H2, B7-H3, PD-L1, PD-L2, B7-H4, PDCD6, BTLA, 41BB (CD137), FcERTy,CD40L, 4- 1BBL, GITR, BAFF, GITR-L, BAFF-R, HVEM, CD27, LIGHT, CD27L, 0X40, OX40L, CD30, CD30L, TAC1, CD40, CD244, CD84, BLAME, CD229, CRACC, CD2F-10, NTB- A, CD48, SLAM (CD150), CD58, ikaros, CD53, integrin a4, CD82, integrin a4bl, CD90, integrin a4b7, CD96, LAG-3, CD 160, LMIR, CRTAM, TCL1A, DAP 12; TIM-1, Dectin- 1, TIM-4, TSLP, EphB6, TSLP-R, and / or HLA-DR domains. In some embodiments, the co-stimulatory protein is from one of the above listed domains.

[0645] In certain embodiments, the CAR comprises two or more intracellular signaling domains. For example, the CAR may comprise a first signaling domain and a second signaling domain or fragments thereof independently selected from the group consisting of a CD3(^ intracellular signaling domain, a CD28 costimulatory intracellular signaling domain, a 4- IBB costimulatory intracellular signaling domain, an OX-40 costimulatory intracellular signaling domain, an inducible co-stimulator (ICOS) costimulatory intracellular signaling domain, a CD27 costimulatory intracellular signaling domain, and a MyD88 / CD40 intracellular signaling domain. In some embodiments, a CAR may include a first intracellular signaling domain or fragment thereof that is a CD3(^ intracellular signaling domain and a second intracellular signaling domain or fragment thereof that is a CD28 costimulatory intracellular signaling domain. In some embodiments, a CAR may include a first intracellular signaling domain or fragment thereof that is a CD3(^ intracellular signaling domain and a second intracellular signaling domain or fragment thereof that is a 4- IBB costimulatory intracellular signaling domain. In some embodiments, a CAR may include a first intracellular signaling domain or fragment thereof that is a CD3(^ intracellular signaling domain and a second intracellular signaling domain or fragment thereof that is a CD2 costimulatory intracellular signaling domain In some embodiments, a CAR may include a first intracellular signaling domain or fragment thereof that is a CD3(^ intracellular signaling domain, a second intracellular signaling domain or fragment thereof that is a 4-1BB costimulatory intracellular signaling domain, and a third intracellular signaling domain or fragment thereof that is a CD3epsilon intracellular signaling domain.

[0646] In certain embodiments, a CAR includes a CD3(^ primary signaling domain and one or more costimulatory signaling domains. In certain embodiments, a CAR includes a 4-1BB costimulatory intracellular signaling domain. As described herein, the intracellular domain comprises a primary signaling domain and a costimulatory domain, and the intracellular domain is operably linked to the C-terminus of the transmembrane domain. In certain embodiments, a CAR lacks a CD2 intracellular signaling domain.

[0647] In some embodiments, the anti-CD22 CAR comprises an intracellular costimulatory signaling domain from 4-1BB and a CD3(^ intracellular signaling domain. In some embodiments, the anti-CD19 CAR comprises an intracellular costimulatory signaling domain from CD28 and a CD3(^ intracellular signaling domain. In some embodiments, the anti-CD20 CAR comprises an intracellular costimulatory signaling domain from CD2 and a CD3(^ intracellular signaling domain.

[0648] CARs of the disclosure may comprise a CD3< 4-1BB (CD137), CD28, ICOS, FcyRI, FcRy, FcR, CD3y, CD35, CD3s, CD35, CD22, CD79a, CD79b, CD665, TLR1, TLR2, TLR3, TLR4, TLR5, TLR6, TLR7, TLR8, TLR9, TLR10, CARD11, CD2, CD7, CD27, CD30, CD40, CD54 (ICAM), CD83, CD134 (0X40), CD137 (4-1BB), CD278 (ICOS), DAP10, LAT, KD2C, SLP76, TRIM, and / or ZAP70 cytoplasmic signaling domain. In some embodiments, the cytoplasmic signaling domain has about 70, 75, 80, 85, 90, 92, 93, 94, 95, 96, 97, 98, 99 or about 100% sequence identity to a CD3< 4-1BB (CD137), CD28, ICOS, FcyRI, FcRy, FcR, CD3y, CD35, CD3s, CD35, CD22, CD79a, CD79b, CD665, TLR1, TLR2, TLR3, TLR4, TLR5, TLR6, TLR7, TLR8, TLR9, TLR10, CARD11, CD2, CD7, CD27, CD30, CD40, CD54 (ICAM), CD83, CD134 (0X40), CD137 (4-1BB), CD278 (ICOS), DAP10, LAT, KD2C, SLP76, TRIM, and / or ZAP70 cytoplasmic signaling domain.Exemplary Anti-CD19 CARs

[0649] In another aspect, the present disclosure describes recombinant polynucleic acid molecules comprising sequences encoding one, two, or three different chimeric antigen receptor (CAR) polypeptides.

[0650] In some embodiments, the recombinant polynucleic acid comprises a sequence encoding a polypeptide encoding an anti-CD19 CAR comprising: (i) an extracellular domain comprising an anti- CD19 binding domain, (ii) a transmembrane domain, and (iii) a cytoplasmic domain comprising an intracellular signaling domain. In some embodiments, the polypeptide is an anti-CD19 CAR comprising: (i) an extracellular domain comprising an anti -CD 19 binding domain and a hinge domain from CD8a or CD28, (ii) a transmembrane domain from CD8a or CD28, and (iii) a cytoplasmic domain comprising an intracellular co-stimulatory signaling domain from CD28 and an intracellular signaling domain from CD3<^. In some embodiments, the polypeptide is an anti-CD19 CAR comprising: (i) an extracellular domain comprising an anti-CD19 binding domain and a hinge domain from CD28, (ii) a transmembrane domain from CD28, and (iii) a cytoplasmic domain comprising an intracellular co-stimulatory signaling domain from CD28 and an intracellular signaling domain from CD3<^. The anti-CD19 binding domain can be any of the anti-CD19 binding domains provided herein. In some embodiments, the hinge domain sequence of the anti-CD19 CAR comprises a hinge domain derived from CD28. In some embodiments, the CD28 hinge domain ofthe anti-CD19 CAR comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to the sequence lEVMYPPPYLDNEKSNGTIIHVKGKHLCPSPLFPGPSKP (SEQ ID NO: 682). In some embodiments, the CD28 hinge domain of the anti-CD19 CAR comprises the sequence lEVMYPPPYLDNEKSNGTIIHVKGKHLCPSPLFPGPSKP (SEQ ID NO: 682).

[0651] In some embodiments, the transmembrane domain sequence of the anti-CD19 CAR comprises a transmembrane domain derived from CD28. In some embodiments, the CD28 transmembrane domain of the anti-CD19 CAR comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to the sequence FWVLVVVGGVLACYSLLVTVAFIIFWV (SEQ ID NO: 683). In some embodiments, the CD28 transmembrane domain of the anti-CD19 CAR comprises the sequence FWVLVVVGGVLACYSLLVTVAFIIFWV (SEQ ID NO: 683).

[0652] In some embodiments, the intracellular co-stimulatory signaling domain of the anti-CD19 CAR is a CD28 intracellular co-stimulatory signaling domain. In some embodiments, the CD28 intracellular co-stimulatory signaling domain of the anti-CD19 CAR comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to the sequence RSKRSRLLHSDYMNMTPRRPGPTRKHYQPYAPPRDFAAYRS (SEQ ID NO: 684). In some embodiments, the CD28 intracellular co-stimulatory signaling domain of the anti-CD19 CAR comprises the sequence RSKRSRLLHSDYMNMTPRRPGPTRKHYQPYAPPRDFAAYRS (SEQ ID NO: 684).

[0653] In some embodiments, the anti-CD19 CAR comprises an intracellular signaling domain comprising an ITAM-containing signaling domain. In some embodiments, the ITAM-containing signaling domain is a CD3^ intracellular signaling domain. In some embodiments, the CD3^ intracellular signaling domain of the anti-CD19 CAR comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to the sequence RVKFSRSADAPAYKQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLY NELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 686). In some embodiments, the CD3^ intracellular signaling domain of the anti-CD19 CAR comprises the sequenceRVKFSRSADAPAYKQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLY NELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 686).. In some embodiments, the CD3^ intracellular signaling domain of the anti-CD19 CAR comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity toany one of the CD3^ sequences in Table 20. In some embodiments, the CD3^ intracellular signaling domain of the anti-CD19 CAR comprises any one of the CD3^ sequences in Table 20.

[0654] In some embodiments, the anti-CD19 CAR comprises a signal peptide sequence derived from CD8a or granulocyte-macrophage colony-stimulating factor receptor (GMCSFRa). In some embodiments, the anti-CD19 CAR comprises a signal peptide sequence derived from CD8a. In some embodiments, the CD8a signal peptide sequence comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to the sequence MALPVTALLLPLALLLHAARP (SEQ ID NO: 1000). In some embodiments, the CD8a signal peptide sequence comprises the sequence MALPVTALLLPLALLLHAARP (SEQ ID NO: 1000).

[0655] In some embodiments, the anti-CD19 CAR comprises a signal peptide sequence derived from GMCSFRa. In some embodiments, the GMCSFRa signal peptide sequence comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to the sequence MLLLVTSLLLCELPHPAFLLIP (SEQ ID NO: 1001). In some embodiments, the GMCSFRa signal peptide sequence comprises the sequence MLLLVTSLLLCELPHPAFLLIP (SEQ ID NO: 1001).

[0656] In some embodiments, the anti-CD19 CAR comprises a CD28 hinge domain, a CD28 transmembrane domain, a CD28 intracellular costimulatory signaling domain and a CD3^ intracellular signaling domain comprising a sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to the sequence IEVMYPPPYLDNEKSNGTIIHVKGKHLCPSPLFPGPSKPFWVLVVVGGVLACYSLLVTVAFII FWVRSKRSRLLHSDYMNMTPRRPGPTRKHYQPYAPPRDFAAYRSRVKFSRSADAPAYKQG QNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLYNELQKDKMAEAYSEIG MKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 1004). In some embodiments, the anti-CD19 CAR comprises a CD28 hinge domain, a CD28 transmembrane domain, a CD28 intracellular costimulatory signaling domain and a CD3^ intracellular signaling domain comprising the sequence lEVMYPPPYLDNEKSNGTIIHVKGKHLCPSPLFPGPSKPFWVLVVVGGVLACYSLLVTVAFII FWVRSKRSRLLHSDYMNMTPRRPGPTRKHYQPYAPPRDFAAYRSRVKFSRSADAPAYKQG QNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLYNELQKDKMAEAYSEIG MKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 1004).

[0657] In some embodiments, the anti-CD19 CAR comprises a sequence having at least about 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 sequence selected from the group consisting of the anti-CD19 CAR sequences in Table 21. In some embodiments, the anti-CD19 CAR comprises any one of the anti-CD19 CAR sequences in Table 21.

[0658] In some embodiments, the anti-CD19 CAR comprises an scFv derived from a human CD 19 antibody designated 19c493, a CD28 hinge domain, a CD28 transmembrane domain, a CD28 intracellular costimulatory signaling domain, and a CD3^ intracellular signaling domain together having the sequenceEVQLVQSGAEVKKPGESLKISCKASGYRFTNYWIAWVRQRPGKGLEWMGRIDPSDSYTHY SPSFQGHVTMSTDKSISTAYLQWSSLKASDTAMYYCARPGDILTGWAMDVWGQGTLVTVS SAAASGGGGSGGGGSGGGGSALQSVLTQPPSVSAAPGQKVTISCSGSSSNIGNNYVSWYQQ LPGTAPKLLIYDNNKRPSGIPDRFSGSKSGTSATLGITGLQAEDEADYYCQSYDSSLSGNYVF GTGTKVTVLIEVMYPPPYLDNEKSNGTIIHVKGKHLCPSPLFPGPSKPFWVLVVVGGVLACY SLLVTVAFIIFWVRSKRSRLLHSDYMNMTPRRPGPTRKHYQPYAPPRDFAAYRSRVKFSRSA DAPAYKQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLYNELQKDKM AEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 1121)

[0659] In some embodiments, the anti-CD19 CAR comprises a signal sequence derived from CD8a, an scFv derived from a human CD 19 antibody designated 19c493, a CD28 hinge domain, a CD28 transmembrane domain, a CD28 intracellular costimulatory signaling domain, and a CD3^ intracellular signaling domain together having the sequenceMALPVTALLLPLALLLHAARPEVQLVQSGAEVKKPGESLKISCKASGYRFTNYWIAWVRQR PGKGLEWMGRIDPSDSYTHYSPSFQGHVTMSTDKSISTAYLQWSSLKASDTAMYYCARPGD ILTGWAMDVWGQGTLVTVSSAAASGGGGSGGGGSGGGGSALQSVLTQPPSVSAAPGQKVT ISCSGSSSNIGNNYVSWYQQLPGTAPKLLIYDNNKRPSGIPDRFSGSKSGTSATLGITGLQAED EADYYCQSYDSSLSGNYVFGTGTKVTVLIEVMYPPPYLDNEKSNGTIIHVKGKHLCPSPLFP GPSKPFWVLVVVGGVLACYSLLVTVAFIIFWVRSKRSRLLHSDYMNMTPRRPGPTRKHYQP YAPPRDFAAYRSRVKFSRSADAPAYKQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGG KPRRKNPQEGLYNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHM QALPPR (SEQ ID NO: 706).Exemplary Anti-CD20 CAR

[0660] In some embodiments, the recombinant polynucleic acid comprises a sequence encoding a polypeptide encoding an anti-CD20 CAR comprising: (i) an extracellular domain comprising an anti- CD20 binding domain, (ii) a transmembrane domain, and (iii) a cytoplasmic domain comprising an intracellular signaling domain. In some embodiments, the polypeptide is an anti-CD20 CAR comprising: (i) an extracellular domain comprising an anti-CD20 binding domain and a hingedomain from CD28, (ii) a transmembrane domain from CD28, and (iii) a cytoplasmic domain comprising an intracellular co-stimulatory signaling domain from CD2 and an intracellular signaling domain from CD3^. The anti-CD20 binding domain can be any of the anti-CD20 binding domains provided herein.

[0661] In some embodiments, the hinge domain sequence of the anti-CD20 CAR comprises a hinge domain derived from CD28. In some embodiments, the CD28 hinge domain of the anti-CD20 CAR comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to the sequence lEVMYPPPYLDNEKSNGTIIHVKGKHLCPSPLFPGPSKP (SEQ ID NO: 682). In some embodiments, the CD28 hinge domain of the anti-CD20 CAR comprises the sequence lEVMYPPPYLDNEKSNGTIIHVKGKHLCPSPLFPGPSKP (SEQ ID NO: 682).

[0662] In some embodiments, the transmembrane domain sequence of the anti-CD20 CAR comprises a transmembrane domain derived from CD28. In some embodiments, the CD28 transmembrane domain of the anti-CD20 CAR comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to the sequence FWVLVVVGGVLACYSLLVTVAFIIFWV (SEQ ID NO: 683). In some embodiments, the transmembrane domain of the anti-CD20 CAR comprises the sequence FWVLVVVGGVLACYSLLVTVAFIIFWV (SEQ ID NO: 683).

[0663] In some embodiments, the intracellular co-stimulatory signaling domain of the anti-CD20 CAR is derived from CD2. In some embodiments, the CD2 intracellular co-stimulatory signaling domain of the anti-CD20 CAR comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to the sequence KRKKQRSRRNDEELETRAHRVATEERGRKPHQIPASTPQNPATSQHPPPPPGHRSQAPSHRP PPPGHRVQHQPQKRPPAPSGTQVHQQKGPPLPRPRVQPKPPHGAAENSLSPSSN (SEQ ID NO: 685). In some embodiments, the CD2 intracellular co-stimulatory signaling domain of the anti- CD20 CAR comprises the sequenceKRKKQRSRRNDEELETRAHRVATEERGRKPHQIPASTPQNPATSQHPPPPPGHRSQAPSHRP PPPGHRVQHQPQKRPPAPSGTQVHQQKGPPLPRPRVQPKPPHGAAENSLSPSSN (SEQ ID NO: 685).

[0664] In some embodiments, the intracellular signaling domain can be derived from an ITAM- containing domain. In some embodiments, the ITAM-containing domain is derived from CD3^. In some embodiments, the intracellular signaling domain of the anti-CD20 CAR is derived from CD3^. In some embodiments, the CD3^ intracellular signaling domain of the anti-CD20 CAR comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to thesequence RVKFSRSADAPAYKQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLY NELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 686). In some embodiments, the CD3^ intracellular signaling domain of the anti-CD20 CAR comprises the sequence RVKFSRSADAPAYKQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLY NELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 686). In some embodiments, the intracellular signaling domain of the anti-CD20 CAR comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to any of the CD3^ sequences in Table 20. In some embodiments, the intracellular signaling domain of the anti-CD20 CAR comprises a CD3^ sequence in Table 20.

[0665] In some embodiments, the anti-CD20 CAR comprises a signal peptide sequence derived from CD8a or granulocyte-macrophage colony-stimulating factor receptor (GMCSFRa). In some embodiments, the anti-CD20 CAR comprises a signal peptide sequence derived from CD8a. In some embodiments, the signal peptide sequence derived from CD8a comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to the sequence MALPVTALLLPLALLLHAARP (SEQ ID NO: 1000). In some embodiments, the signal peptide sequence derived from CD8a comprises the sequence MALPVTALLLPLALLLHAARP (SEQ ID NO: 1000).

[0666] In some embodiments, the anti-CD20 CAR comprises a signal peptide sequence derived from granulocyte-macrophage colony-stimulating factor receptor (GMCSFRa). In some embodiments, the anti-CD20 CAR comprises a signal peptide sequence derived from GMCSFRa. In some embodiments, the signal peptide sequence derived from GMCSFRa comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the sequence MLLLVTSLLLCELPHPAFLLIP (SEQ ID NO: 1001). In some embodiments, the signal peptide sequence derived from GMSCFRa comprises the sequence MLLLVTSLLLCELPHPAFLLIP (SEQ ID NO: 1001).

[0667] In some embodiments, the anti-CD20 CAR comprises a CD28 hinge domain, a CD28 transmembrane domain, a CD2 intracellular costimulatory signaling domain, and a CD3^ intracellular signaling domain comprising a sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to the sequence lEVMYPPPYLDNEKSNGTIIHVKGKHLCPSPLFPGPSKPFWVLVVVGGVLACYSLLVTVAFIIFWVKRKKQRSRRNDEELETRAHRVATEERGRKPHQIPASTPQNPATSQHPPPPPGHRSQAPSHRPPPPGHRVQHQPQKRPPAPSGTQVHQQKGPPLPRPRVQPKPPHGAAENSLSPSSNRVKFS RSADAPAYKQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLYNELQK DKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 1005).

[0668] In some embodiments, the anti-CD20 CAR comprises a CD28 hinge domain, a CD28 transmembrane domain, a CD2 intracellular costimulatory signaling domain, and a CD3^ intracellular signaling domain comprising the sequence IEVMYPPPYLDNEKSNGTIIHVKGKHLCPSPLFPGPSKPFWVLVVVGGVLACYSLLVTVAFII FWVKRKKQRSRRNDEELETRAHRVATEERGRKPHQIPASTPQNPATSQHPPPPPGHRSQAPS HRPPPPGHRVQHQPQKRPPAPSGTQVHQQKGPPLPRPRVQPKPPHGAAENSLSPSSNRVKFS RSADAPAYKQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLYNELQK DKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 1005).

[0669] In some embodiments, the anti-CD20 CAR comprises a sequence having at least about 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 sequence selected from the group consisting of any of the anti-CD20 CAR sequences in Table 21. In some embodiments, the anti-CD20 CAR comprises any of the anti-CD20 CAR sequences in Table 21.

[0670] In some embodiments, the anti-CD20 CAR comprises a Flag tag, an scFv derived from a human CD20 antibody designated 20c27, a CD28 hinge domain, a CD28 transmembrane domain, a CD2 intracellular costimulatory signaling domain, and a CD3^ intracellular signaling domain together having the sequenceDYKDDDDKDIVMTQSPDSLAVSLGERATINCKSSQSVLYSSNNKNYLAWYQQKPGQPPKLL IYWASTRESGVPDRFSGSGSGSDFTLTISSLQAEDVAVYYCQQYYSFYQTFGQGTKVEIKGG GGSGGGGSGGGGSEVQLVESGGGVVRPGGSLRLSCTASGFTFGDYGMSWVRQAPGKGLE WVSGINWNGGSTGYADSVKGRFTISRDNAKNSLYLQMNSLRAEDTALYYCARKSYYGSGS PDVFDIWGQGTMVTVSSIEVMYPPPYLDNEKSNGTIIHVKGKHLCPSPLFPGPSKPFWVLVV VGGVLACYSLLVTVAFIIFWVKRKKQRSRRNDEELETRAHRVATEERGRKPHQIPASTPQNP ATSQHPPPPPGHRSQAPSHRPPPPGHRVQHQPQKRPPAPSGTQVHQQKGPPLPRPRVQPKPP HGAAENSLSPSSNRVKFSRSADAPAYKQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGG KPRRKNPQEGLYNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHM QALPPR (SEQ ID NO: 1119)

[0671] In some embodiments, the anti-CD20 CAR comprises an scFv derived from a human CD20 antibody designated 20c27, a CD28 hinge domain, a CD28 transmembrane domain, a CD2 intracellular costimulatory signaling domain, and a CD3^ intracellular signaling domain together having the sequenceDIVMTQSPDSLAVSLGERATINCKSSQSVLYSSNNKNYLAWYQQKPGQPPKLLIYWASTRES GVPDRFSGSGSGSDFTLTISSLQAEDVAVYYCQQYYSFYQTFGQGTKVEIKGGGGSGGGGS GGGGSEVQLVESGGGVVRPGGSLRLSCTASGFTFGDYGMSWVRQAPGKGLEWVSGINWN GGSTGYADSVKGRFTISRDNAKNSLYLQMNSLRAEDTALYYCARKSYYGSGSPDVFDIWG QGTMVTVSSIEVMYPPPYLDNEKSNGTIIHVKGKHLCPSPLFPGPSKPFWVLVVVGGVLACY SLLVTVAFIIFWVKRKKQRSRRNDEELETRAHRVATEERGRKPHQIPASTPQNPATSQHPPPP PGHRSQAPSHRPPPPGHRVQHQPQKRPPAPSGTQVHQQKGPPLPRPRVQPKPPHGAAENSLS PSSNRVKFSRSADAPAYKQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQE GLYNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 1359)

[0672] In some embodiments, the anti-CD20 CAR comprises a signal sequence derived from GMCSFRa, a FLAG tag, an scFv derived from a human CD20 antibody designated 20c27, a CD28 hinge domain, a CD28 transmembrane domain, a CD2 intracellular costimulatory signaling domain, and a CD3^ intracellular signaling domain together having the sequenceMLLLVTSLLLCELPHPAFLLIPDYKDDDDKDIVMTQSPDSLAVSLGERATINCKSSQSVLYSS NNKNYLAWYQQKPGQPPKLLIYWASTRESGVPDRF SGSGSGSDFTLTIS SLQAED VAVYYC QQYYSFYQTFGQGTKVEIKGGGGSGGGGSGGGGSEVQLVESGGGVVRPGGSLRLSCTASGF TFGDYGMSWVRQAPGKGLEWVSGINWNGGSTGYADSVKGRFTISRDNAKNSLYLQMNSL RAEDTALYYCARKSYYGSGSPDVFDIWGQGTMVTVSSIEVMYPPPYLDNEKSNGTIIHVKG KHLCPSPLFPGPSKPFWVLVVVGGVLACYSLLVTVAFIIFWVKRKKQRSRRNDEELETRAHR VATEERGRKPHQIPASTPQNPATSQHPPPPPGHRSQAPSHRPPPPGHRVQHQPQKRPPAPSGT QVHQQKGPPLPRPRVQPKPPHGAAENSLSPSSNRVKFSRSADAPAYKQGQNQLYNELNLGR REEYDVLDKRRGRDPEMGGKPRRKNPQEGLYNELQKDKMAEAYSEIGMKGERRRGKGHD GLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 772)

[0673] In some embodiments, the anti-CD20 CAR comprises a signal sequence derived from GMCSFRa, an scFv derived from a human CD20 antibody designated 20c27, a CD28 hinge domain, a CD28 transmembrane domain, a CD2 intracellular costimulatory signaling domain, and a CD3^ intracellular signaling domain together having the sequenceMLLLVTSLLLCELPHPAFLLIPDIVMTQSPDSLAVSLGERATINCKSSQSVLYSSNNKNYLAW YQQKPGQPPKLLIYWASTRESGVPDRFSGSGSGSDFTLTISSLQAEDVAVYYCQQYYSFYQT FGQGTKVEIKGGGGSGGGGSGGGGSEVQLVESGGGVVRPGGSLRLSCTASGFTFGDYGMS WVRQAPGKGLEWVSGINWNGGSTGYADSVKGRFTISRDNAKNSLYLQMNSLRAEDTALY YC ARKS YYGSGSPDVFDIWGQGTMVTVS SIEVMYPPPYLDNEKSNGTIIHVKGKHLCP SPLF PGPSKPFWVLVVVGGVLACYSLLVTVAFIIFWVKRKKQRSRRNDEELETRAHRVATEERGR KPHQIPASTPQNPATSQHPPPPPGHRSQAPSHRPPPPGHRVQHQPQKRPPAPSGTQVHQQKGP PLPRPRVQPKPPHGAAENSLSPSSNRVKFSRSADAPAYKQGQNQLYNELNLGRREEYDVLD KRRGRDPEMGGKPRRKNPQEGLYNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLST ATKDTYDALHMQALPPR (SEQ ID NO: 1358)Exemplary Anti-CD22 CAR

[0674] In some embodiments, the polypeptide is an anti-CD22 CAR comprising: (i) an extracellular domain comprising an anti-CD22 binding domain, (ii) a transmembrane domain, and (iii) a cytoplasmic domain comprising an intracellular signaling domain. In some embodiments, the polypeptide is an anti-CD22 CAR comprising: (i) an extracellular domain comprising an anti-CD22 binding domain and a hinge domain from CD8a or CD28, (ii) a transmembrane domain from CD8a or CD28, and (iii) a cytoplasmic domain comprising an intracellular co- stimulatory signaling domain from 4- IBB and an intracellular signaling domain from CD3^. In some embodiments, the polypeptide is an anti-CD22 CAR comprising: (i) an extracellular domain comprising an anti-CD22 binding domain and a hinge domain from CD8a, (ii) a transmembrane domain from CD8a, and (iii) a cytoplasmic domain comprising an intracellular co-stimulatory signaling domain from 4- IBB and an intracellular signaling domain from CD3^. The anti-CD22 binding domain can be any anti-CD22 binding domain provided herein. In some embodiments, the anti-CD22 binding domain comprises the m971 scFv (SEQ ID NO:664).

[0675] In some embodiments, the linker between the anti-CD22 binding domain and the hinge domain from CD8a or CD28 comprises the sequence AAA.

[0676] In some embodiments, the hinge domain of the anti-CD22 CAR is derived from CD8a. In some embodiments, the CD8a hinge domain of the anti-CD22 CAR comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to the sequence TTTPAPRPPTPAPTIASQPLSLRPEACRPAAGGAVHTRGLDFACD (SEQ ID NO: 679). In some embodiments, the CD8a hinge domain of the anti-CD22 CAR comprises the sequence TTTPAPRPPTPAPTIASQPLSLRPEACRPAAGGAVHTRGLDFACD (SEQ ID NO: 679).

[0677] In some embodiments, the transmembrane domain of the anti-CD22 CAR is from CD8a. In some embodiments, the CD8a transmembrane domain of the anti-CD22 CAR comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to the sequence IYIWAPLAGTCGVLLLSLVIT (SEQ ID NO: 680). In some embodiments, the transmembrane domain of the anti-CD22 CAR comprises the sequence IYIWAPLAGTCGVLLLSLVIT (SEQ ID NO: 680).

[0678] In some embodiments, the anti-CD22 CAR comprises a portion of a CD8a cytoplasmic domain. In some embodiments, the portion of a CD8a cytoplasmic domain of the anti-CD22 CAR comprises the sequence LYC.

[0679] In some embodiments, the intracellular co-stimulatory domain of the anti-CD22 CAR is derived from 4-1BB. In some embodiments, the 4-1BB co-stimulatory signaling domain of the anti- CD22 CAR comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to the sequence KRGRKKLLYIFKQPFMRPVQTTQEEDGCSCRFPEEEEGGCEL (SEQ ID NO: 998). In some embodiments, the 4- IBB co-stimulatory signaling domain of the anti-CD22 CAR comprises the sequence KRGRKKLLYIFKQPFMRPVQTTQEEDGCSCRFPEEEEGGCEL (SEQ ID NO: 998).

[0680] In some embodiments, the intracellular signaling domain can be an ITAM-containing domain. In some embodiments, the ITAM-containing domain is derived from CD3^. In some embodiments, the intracellular signaling domain of the anti-CD22 CAR is derived from CD3^. In some embodiments, the CD3^ intracellular signaling domain of the anti-CD22 CAR comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to the sequence RVKFSRSADAPAYKQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLY NELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 686). In some embodiments, the CD3^ intracellular signaling domain of the anti-CD22 CAR comprises the sequence RVKFSRSADAPAYKQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLY NELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 686). In some embodiments, the CD3^ intracellular signaling domain of the anti-CD22 CAR comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to any of the CD3^ sequences in Table 20. In some embodiments, the CD3^ intracellular signaling domain of the anti-CD22 CAR comprises any of the CD3^ sequences in Table 20.

[0681] In some embodiments, the anti-CD22 CAR comprises a signal peptide sequence derived from CD8a or GMCSFRa. In some embodiments, the signal peptide sequence is derived from GMCSFRa and comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to the sequence MLLLVTSLLLCELPHPAFLLIP (SEQ ID NO: 1001).

[0682] In some embodiments, the signal peptide sequence is derived from GMCSFRa and comprises the sequence MLLLVTSLLLCELPHPAFLLIP (SEQ ID NO: 1001).

[0683] In some embodiments, the signal peptide sequence is derived from CD8a and comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to the sequence MALPVTALLLPLALLLHAARP (SEQ ID NO: 1000) (SEQ ID NO: 1000).

[0684] In some embodiments, the signal peptide sequence is derived from CD8a and comprises the sequence MALPVTALLLPLALLLHAARP (SEQ ID NO: 1000).

[0685] In some embodiments, the anti-CD22 CAR comprises a sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to the sequenceQVQLQQSGPGLVKPSQTLSLTCAISGDSVSSNSAAWNWIRQSPSRGLEWLGRTYYRSKWYN DYAVSVKSRITINPDTSKNQFSLQLNSVTPEDTAVYYCAREVTGDLEDAFDIWGQGTMVTV SSGGGGSDIQMTQSPSSLSASVGDRVTITCRASQTIWSYLNWYQQRPGKAPNLLIYAASSLQ SGVPSRFSGRGSGTDFTLTISSLQAEDFATYYCQQSYSIPQTFGQGTKLEIKAAATTTPAPRPP TPAPTIASQPLSLRPEACRPAAGGAVHTRGLDFACDIYIWAPLAGTCGVLLLSLVITLYCKRG RKKLLYIFKQPFMRP VQTTQEEDGC SCRFPEEEEGGCELRVKF SRS ADAP AYKQGQNQLYN ELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLYNELQKDKMAEAYSEIGMKGERR RGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 1008).

[0686] In some embodiments, the anti-CD22 CAR comprises the sequenceQVQLQQSGPGLVKPSQTLSLTCAISGDSVSSNSAAWNWIRQSPSRGLEWLGRTYYRSKWYN DYAVSVKSRITINPDTSKNQFSLQLNSVTPEDTAVYYCAREVTGDLEDAFDIWGQGTMVTV SSGGGGSDIQMTQSPSSLSASVGDRVTITCRASQTIWSYLNWYQQRPGKAPNLLIYAASSLQ SGVPSRFSGRGSGTDFTLTISSLQAEDFATYYCQQSYSIPQTFGQGTKLEIKAAATTTPAPRPP TPAPTIASQPLSLRPEACRPAAGGAVHTRGLDFACDIYIWAPLAGTCGVLLLSLVITLYCKRG RKKLLYIFKQPFMRP VQTTQEEDGC SCRFPEEEEGGCELRVKF SRS ADAP AYKQGQNQLYN ELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLYNELQKDKMAEAYSEIGMKGERR RGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 1008).

[0687] In some embodiments, the anti-CD22 CAR comprises a sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to the sequenceMLLLVTSLLLCELPHPAFLLIPQVQLQQSGPGLVKPSQTLSLTCAISGDSVSSNSAAWNWIRQSPSRGLEWLGRTYYRSKWYNDYAVSVKSRITINPDTSKNQFSLQLNSVTPEDTAVYYCARE VTGDLEDAFDIWGQGTMVTVSSGGGGSDIQMTQSPSSLSASVGDRVTITCRASQTIWSYLN WYQQRPGKAPNLLIYAASSLQSGVPSRFSGRGSGTDFTLTISSLQAEDFATYYCQQSYSIPQT FGQGTKLEIKAAATTTPAPRPPTPAPTIASQPLSLRPEACRPAAGGAVHTRGLDFACDIYIWA PLAGTCGVLLLSLVITLYCKRGRKKLLYIFKQPFMRPVQTTQEEDGCSCRFPEEEEGGCELR VKFSRSADAPAYKQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLYN ELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 996).

[0688] In some embodiments, the anti-CD22 CAR comprises the sequenceMLLLVTSLLLCELPHPAFLLIPQVQLQQSGPGLVKPSQTLSLTCAISGDSVSSNSAAWNWIRQ SPSRGLEWLGRTYYRSKWYNDYAVSVKSRITINPDTSKNQFSLQLNSVTPEDTAVYYCARE VTGDLEDAFDIWGQGTMVTVSSGGGGSDIQMTQSPSSLSASVGDRVTITCRASQTIWSYLN WYQQRPGKAPNLLIYAASSLQSGVPSRFSGRGSGTDFTLTISSLQAEDFATYYCQQSYSIPQT FGQGTKLEIKAAATTTPAPRPPTPAPTIASQPLSLRPEACRPAAGGAVHTRGLDFACDIYIWA PLAGTCGVLLLSLVITLYCKRGRKKLLYIFKQPFMRPVQTTQEEDGCSCRFPEEEEGGCELR VKFSRSADAPAYKQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLYN ELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 996).

[0689] In some embodiments, the anti-CD22 CAR comprises a sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to the sequence MALPVTALLLPLALLLHAARPEVQLVESGGGWRPGGSLRLSCTASGFTFGDYGMSWVRQ APGKGLEWVSGINWNGGSTGYADSVKGRFTISRDNAKNSLYLQMNSLRAEDTALYYCARK SYYGSGSPDVFDIWGQGTMVTVSSGGGGSGGGGSGGGGSDIVMTQSPDSLAVSLGERATIN CKSSQSVLYSSNNKNYLAWYQQKPGQPPKLLIYWASTRESGVPDRFSGSGSGSDFTLTISSL QAEDVAVYYCQQYYSFYQTFGQGTKVEIKIEVMYPPPYLDNEKSNGTIIHVKGKHLCPSPLF PGPSKPFWVLVVVGGVLACYSLLVTVAFIIFWVKRKKQRSRRNDEELETRAHRVATEERGR KPHQIPASTPQNPATSQHPPPPPGHRSQAPSHRPPPPGHRVQHQPQKRPPAPSGTQVHQQKGP PLPRPRVQPKPPHGAAENSLSPSSNRVKFSRSADAPAYKQGQNQLYNELNLGRREEYDVLD KRRGRDPEMGGKPRRKNPQEGLYNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLST ATKDTYDALHMQALPPR (SEQ ID NO: 1314).

[0690] In some embodiments, the anti-CD22 CAR comprises a sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to the sequenceMLLLVTSLLLCELPHPAFLLIPEVQLVESGGGVVRPGGSLRLSCTASGFTFGDYGMSWVRQAPGKGLEWVSGINWNGGSTGYADSVKGRFTISRDNAKNSLYLQMNSLRAEDTALYYCARKS YYGSGSPDVFDIWGQGTMVTVSSGGGGSGGGGSGGGGSDIVMTQSPDSLAVSLGERATINC KSSQSVLYSSNNKNYLAWYQQKPGQPPKLLIYWASTRESGVPDRFSGSGSGSDFTLTISSLQ AEDVAVYYCQQYYSFYQTFGQGTKVEIKIEVMYPPPYLDNEKSNGTIIHVKGKHLCPSPLFP GPSKPFWVLVVVGGVLACYSLLVTVAFIIFWVKRKKQRSRRNDEELETRAHRVATEERGRK PHQIPASTPQNPATSQHPPPPPGHRSQAPSHRPPPPGHRVQHQPQKRPPAPSGTQVHQQKGPP LPRPRVQPKPPHGAAENSLSPSSNRVKFSRSADAPAYKQGQNQLYNELNLGRREEYDVLDK RRGRDPEMGGKPRRKNPQEGLYNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTA TKDTYDALHMQALPPR (SEQ ID NO: 997).Bispecific Chimeric Antisen Receptors

[0691] In another aspect, the present disclosure describes a recombinant polynucleic acid molecule comprising a sequence encoding a polypeptide of a bispecific chimeric antigen receptor (CAR). In some embodiments, the bispecific CAR comprises: (i) an extracellular domain comprising an anti- CD19 binding domain and an anti-CD20 binding domain, (ii) a transmembrane domain, and (iii) a cytoplasmic domain comprising an intracellular signaling domain. In some embodiments, the cytoplasmic domain further comprises a co-stimulatory signaling domain from CD2.

[0692] In some embodiments, the bispecific CAR comprises: (i) an extracellular domain comprising an anti-CD19 binding domain and an anti-CD22 binding domain, (ii) a transmembrane domain; and (iii) a cytoplasmic domain comprising (A) a co-stimulatory signaling domain from CD2, CD28, or 4- 1BB and (B) an intracellular signaling domain.

[0693] In some embodiments, the bispecific CAR comprises: (i) an extracellular domain comprising an anti-CD20 binding domain and anti-CD22 binding domain, (ii) a transmembrane domain, and (iii) a cytoplasmic domain comprising (A) a co-stimulatory signaling domain from CD2 and (B) an intracellular signaling domain.

[0694] The anti-CD19 binding domain can be any anti-CD19 binding domain provided herein. The anti-CD20 binding domain can be any anti-CD20 binding domain provided herein. The anti-CD22 binding domain can be any anti-CD22 binding domain provided herein.

[0695] In some embodiments, the extracellular domain of the bispecific CAR further comprises a hinge domain, wherein the hinge domain is from CD8a or CD28.

[0696] In some embodiments, the transmembrane domain of the bispecific CAR is from CD8a or CD28.

[0697] In some embodiments, the intracellular signaling domain of the bispecific CAR is from CD3zeta.

[0698] In some embodiments, the hinge domain from CD8a or CD28 comprises the sequence with at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity toIEVMYPPPYLDNEKSNGTIIHVKGKHLCPSPLFPGPSKP (SEQ ID NO: 682).

[0699] In some embodiments, the transmembrane domain from CD28 comprises the sequence with at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity toFWVLVVVGGVLACYSLLVTVAFIIFWV (SEQ ID NO: 683).

[0700] In some embodiments, the wherein the cytoplasmic domain of the bispecific CAR or the anti- CD20 CAR comprises a co-stimulatory signaling domain from CD2, and wherein the co-stimulatory signaling domain from CD2 comprises a sequence with at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the sequenceKRKKQRSRRNDEELETRAHRVATEERGRKPHQIPASTPQNPATSQHPPPPPGHRSQAPSHRP PPPGHRVQHQPQKRPPAPSGTQVHQQKGPPLPRPRVQPKPPHGAAENSLSPSSN (SEQ ID NO: 685).

[0701] In some embodiments, the intracellular signaling domain from CD3zeta of the bispecific CAR comprises a sequence with at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the sequenceRVKFSRSADAPAYKQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLY NELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 686).

[0702] In some embodiments, the bispecific CAR comprises a sequence with at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to a sequence selected from the group of sequences in Table 23. In some embodiments, the bispecific CAR comprises a sequence selected from the group of sequences in Table 23.Compositions

[0703] In one aspect, the present disclosure provides compositions comprising any of the recombinant polynucleic acids disclosed herein, either alone or in combination. In other aspects, the present disclosure describes compositions comprising any of the antigen binding domains disclosed herein. In another aspects, the composition comprises any of the CARs disclosed herein.

[0704] In one aspect, provided herein is a composition comprising one or more recombinant polynucleic acid molecules comprising: (a) a sequence encoding an anti-CD20 CAR, wherein the anti-CD20 CAR comprises: (i) an extracellular domain comprising an anti-CD20 binding domain, (ii) a transmembrane domain, and (iii) a cytoplasmic domain comprising (A) a co-stimulatorysignaling domain from CD2 and (B) an intracellular signaling domain; and (b) a sequence encoding an anti-CD22 CAR, wherein the anti-CD22 CAR comprises: (i) an extracellular domain comprising an anti-CD22 binding domain, (ii) a transmembrane domain, and (iii) a cytoplasmic domain comprising an intracellular signaling domain. In some embodiments, the one or more recombinant polynucleic acid molecules further comprises a sequence encoding an anti-CD19 CAR. In some embodiments, the anti-CD19 CAR comprises (i) an extracellular domain comprising an anti-CD19 binding domain, (ii) a transmembrane domain, and (iii) a cytoplasmic domain comprising an intracellular signaling domain. In some embodiments, the cytoplasmic domain of the anti-CD19 CAR further comprises a co-stimulatory signaling domain from CD2, CD28, 4- IBB or CD3zeta. In some embodiments, the cytoplasmic domain of the anti-CD19 CAR comprises a co-stimulatory signaling domain CD28.

[0705] In another aspect, provided herein is a composition comprising one or more recombinant polynucleic acid molecules comprising: (a) a sequence encoding an anti-CD19 CAR, wherein the anti-CD19 CAR comprises: (i) an extracellular domain comprising an anti-CD19 binding domain, (ii) a transmembrane domain, and (iii) a cytoplasmic domain comprising (A) a co-stimulatory signaling domain from CD2, CD28, or 4-1BB and (B) an intracellular signaling domain; and (b) a sequence encoding an anti-CD22 CAR, wherein the anti-CD22 CAR comprises: (i) an extracellular domain comprising an anti-CD22 binding domain, (ii) a transmembrane domain, and (iii) a cytoplasmic domain comprising an intracellular signaling domain.

[0706] In another aspect, provided herein is a composition comprising one or more recombinant polynucleic acid molecules comprising: (a) a sequence encoding a bispecific chimeric antigen receptor (CAR), wherein the bispecific CAR comprises: (i) an extracellular domain comprising an anti-CD19 binding domain and an anti-CD20 binding domain, (ii) a transmembrane domain, and (iii) a cytoplasmic domain comprising an intracellular signaling domain; and wherein the cytoplasmic domain further comprises a co-stimulatory signaling domain from CD2.

[0707] In another aspect, provided herein is a composition comprising one or more recombinant polynucleic acid molecules comprising: (a) a sequence encoding a bispecific CAR, wherein the bispecific CAR comprises: (i) an extracellular domain comprising an anti-CD19 binding domain and an anti-CD22 binding domain, (ii) a transmembrane domain; and (iii) a cytoplasmic domain comprising (A) a co-stimulatory signaling domain from CD2, CD28, or 4- IBB and (B) an intracellular signaling domain.

[0708] In another aspect, provided herein is a composition comprising one or more recombinant polynucleic acid molecules comprising: (a) a sequence encoding a bispecific CAR, wherein thebispecific CAR comprises: (i) an extracellular domain comprising an anti-CD20 binding domain and anti-CD22 binding domain, (ii) a transmembrane domain, and (iii) a cytoplasmic domain comprising (A) a co- stimulatory signaling domain from CD2 and (B) an intracellular signaling domain.

[0709] In another aspect, provided herein is a composition comprising one or more recombinant polynucleic acid molecules comprising: (a) a sequence encoding a bispecific chimeric antigen receptor (CAR), wherein the bispecific CAR comprises: (i) an extracellular domain comprising an anti-CD19 binding domain and an anti-CD20 binding domain, (ii) a transmembrane domain, and (iii) a cytoplasmic domain comprising an intracellular signaling domain; and (b) a sequence encoding an anti-CD22 CAR, wherein the anti-CD22 CAR comprises: (i) an extracellular domain comprising an anti-CD22 binding domain, (ii) a transmembrane domain, and (iii) a cytoplasmic domain comprising an intracellular signaling domain; wherein the cytoplasmic domain of the bispecific CAR further comprises a co-stimulatory signaling domain from CD2.

[0710] In some embodiments, the one or more recombinant polynucleic acid molecules of the composition further comprises a sequence encoding an anti-CD20 CAR, wherein the anti-CD20 CAR comprises: (i) an extracellular domain comprising an anti-CD20 binding domain, (ii) a transmembrane domain, and (iii) a cytoplasmic domain comprising an intracellular signaling domain. In some embodiments, the cytoplasmic domain of the anti-CD20 CAR further comprises a costimulatory signaling domain from CD2.

[0711] In another aspect, provided herein is a composition comprising one or more recombinant polynucleic acid molecules comprising: (a) a sequence encoding an anti-CD20 CAR, wherein the anti-CD20 CAR comprises: (i) an extracellular domain comprising an anti-CD20 binding domain, (ii) a transmembrane domain, and (iii) a cytoplasmic domain comprising an intracellular signaling domain; and (b) a sequence encoding an anti-CD19 CAR, wherein the anti-CD19 CAR comprises: (i) an extracellular domain comprising an anti-CD19 binding domain, (ii) a transmembrane domain, and (iii) a cytoplasmic domain comprising an intracellular signaling domain wherein the cytoplasmic domain of the anti-CD19 CAR and / or the cytoplasmic domain of the anti-CD20 CAR further comprises a co-stimulatory signaling domain from CD2.

[0712] In some embodiments, the one or more recombinant polynucleic acid molecules of the composition further comprises a sequence encoding an anti-CD22 CAR, wherein the anti-CD22 CAR comprises: (i) an extracellular domain comprising an anti-CD22 binding domain, (ii) a transmembrane domain, and (iii) a cytoplasmic domain comprising an intracellular signaling domain.

[0713] In another aspect, provided herein is a composition comprising one or more recombinant polynucleic acid molecules comprising: (a) a sequence encoding an anti-CD19 chimeric antigenreceptor (CAR), wherein the anti-CD19 CAR comprises: (i) an extracellular domain comprising an anti-CD19 binding domain, (ii) a transmembrane domain, and (iii) a cytoplasmic domain comprising an intracellular signaling domain; and (b) a sequence encoding an anti-CD20 CAR, wherein the anti- CD20 CAR comprises: (i) an extracellular domain comprising an anti-CD20 binding domain, (ii) a transmembrane domain, and (iii) a cytoplasmic domain comprising an intracellular signaling domain; and (c) a sequence encoding an anti-CD22 CAR, wherein the anti-CD22 CAR comprises: (i) an extracellular domain comprising an anti-CD22 binding domain, (ii) a transmembrane domain, and (iii) a cytoplasmic domain comprising an intracellular signaling domain; wherein the cytoplasmic domain of the anti-CD19 CAR and / or the cytoplasmic domain of the anti-CD20 CAR further comprises a co-stimulatory signaling domain from CD2.

[0714] In some embodiments, the cytoplasmic domain of the anti-CD20 CAR comprises a costimulatory signaling domain from CD2. In some embodiments, the cytoplasmic domain of the anti- CD19 CAR comprises a co-stimulatory signaling domain from CD2, CD28, or 4- IBB.

[0715] In some embodiments, the cytoplasmic domain of the anti-CD19 CAR comprises a co- stimulatory signaling domain from CD28.

[0716] In some embodiments, the cytoplasmic domain of the anti-CD22 CAR further comprises a co-stimulatory signaling domain from 4-1BB.

[0717] In some embodiments, the composition comprises one or more recombinant polynucleic acid molecules comprising: (a) a sequence encoding an anti-CD19 chimeric antigen receptor (CAR), wherein the anti-CD19 CAR comprises: (i) an extracellular domain comprising an anti-CD19 binding domain, (ii) a CD28 hinge domain and a CD28 transmembrane domain, and (iii) a cytoplasmic domain comprising an intracellular costimulatory signaling domain derived from CD28 and an intracellular signaling domain derived from CD3 and (b) a sequence encoding an anti-CD20 CAR, wherein the anti-CD20 CAR comprises: (i) an extracellular domain comprising an anti-CD20 binding domain, (ii) a CD28 hinge domain and CD28 transmembrane domain, and (iii) a cytoplasmic domain comprising an intracellular costimulatory signaling domain derived from CD2 and an intracellular signaling domain derived from CD3 and (c) a sequence encoding an anti-CD22 CAR, wherein the anti-CD22 CAR comprises: (i) an extracellular domain comprising an anti-CD22 binding domain, (ii) a CD8a hinge domain and CD8a transmembrane domain, and (iii) a cytoplasmic domain comprising an intracellular costimulatory signaling domain derived from 4- IBB and an intracellular signaling domain derived from CD3^.

[0718] In some embodiments, the one or more recombinant polynucleic acid molecules of the composition comprises a first recombinant polynucleic acid molecule comprising a sequenceencoding an anti-CD22 CAR and a second recombinant polynucleic acid molecule comprising a sequence encoding an anti-CD19 CAR.

[0719] In some embodiments, the one or more recombinant polynucleic acid molecules of the composition comprises a first recombinant polynucleic acid molecule comprising a sequence encoding an anti-CD22 CAR and a second recombinant polynucleic acid molecule comprising a sequence encoding an anti-CD20 CAR.

[0720] In some embodiments, the one or more recombinant polynucleic acid molecules of the composition comprises a first recombinant polynucleic acid molecule comprising a sequence encoding an anti-CD19 CAR and a second recombinant polynucleic acid molecule comprising a sequence encoding an anti-CD20 CAR.

[0721] In some embodiments, the one or more recombinant polynucleic acid molecules of the composition comprises a first recombinant polynucleic acid molecule comprising a sequence encoding an anti-CD22 CAR, a second recombinant polynucleic acid molecule comprising a sequence encoding an anti-CD20 CAR, and a third recombinant polynucleic acid molecule comprising a sequence encoding an anti-CD19 CAR.

[0722] In some embodiments, the first recombinant polynucleic acid molecule of the composition is encoded by a first viral vector, the second recombinant polynucleic acid molecule of the composition is encoded by a second viral vector, and a third recombinant polynucleic acid molecule of the composition is encoded by a third viral vector. In some embodiments, the first viral vector, the second viral vector and the third viral vector are each lentiviral vectors. In some embodiments, the first recombinant polynucleic acid molecule, the second recombinant polynucleic acid molecule, and the third recombinant polynucleic acid molecule of the composition are in one viral vector.

[0723] In some embodiments, the one or more recombinant polynucleic acid molecules of the composition comprises a recombinant polynucleic acid molecule comprising a sequence encoding an anti-CD22 CAR and a sequence encoding an anti-CD19 CAR.

[0724] In some embodiments, the one or more recombinant polynucleic acid molecules of the composition comprises a recombinant polynucleic acid molecule comprising a sequence encoding an anti-CD22 CAR and a sequence encoding an anti-CD20 CAR.

[0725] In some embodiments, the one or more recombinant polynucleic acid molecules of the composition comprises a recombinant polynucleic acid molecule comprising a sequence encoding an anti-CD19 CAR and a sequence encoding an anti-CD20 CAR.

[0726] In some embodiments, the one or more recombinant polynucleic acid molecules of the composition comprises a recombinant polynucleic acid molecule comprising a sequence encoding ananti-CD22 CAR, a sequence encoding an anti-CD20 CAR, and a sequence encoding an anti-CD19 CAR.

[0727] In some embodiments, the sequence encoding an anti-CD19 CAR, the sequence encoding an anti-CD20 CAR, and the sequence encoding an anti-CD22 CAR are separated by a sequence encoding a P2A self-cleaving peptide, a T2A self-cleaving peptide, an E2A self-cleaving peptide or an F2A self-cleaving peptide.

[0728] In some embodiments, the recombinant polynucleic acid molecule of the composition comprises from 5’ to 3’ a sequence encoding an anti-CD22 CAR, a sequence encoding a P2A selfcleaving peptide, a T2A self-cleaving peptide, an E2A self-cleaving peptide or an F2A self-cleaving peptide, a sequence encoding an anti-CD20 CAR, a sequence encoding a P2A self-cleaving peptide, a T2A self-cleaving peptide, an E2A self-cleaving peptide or an F2A self-cleaving peptide, and a sequence encoding an anti-CD19 CAR.

[0729] In some embodiments, the recombinant polynucleic acid molecule of the composition comprises from 5’ to 3’ a sequence encoding an anti-CD22 CAR, a sequence encoding a P2A selfcleaving peptide, a sequence encoding an anti-CD20 CAR, a sequence encoding a T2A self-cleaving peptide, and a sequence encoding an anti-CD19 CAR.

[0730] In some embodiments, the recombinant polynucleic acid molecule of the composition comprises from 5’ to 3’ a sequence encoding an anti-CD22 CAR, a sequence encoding a P2A selfcleaving peptide, a T2A self-cleaving peptide, an E2A self-cleaving peptide or an F2A self-cleaving peptide, a sequence encoding an anti-CD19 CAR, a sequence encoding a P2A self-cleaving peptide, and a sequence encoding an e anti-CD20 CAR.

[0731] In some embodiments, the recombinant polynucleic acid molecule of the composition comprises from 5’ to 3’ a sequence encoding an anti-CD22 CAR, a sequence encoding a T2A selfcleaving peptide, a sequence encoding an anti-CD19 CAR, a sequence encoding a P2A self-cleaving peptide, and a sequence encoding an anti-CD20 CAR.

[0732] In some embodiments, the recombinant polynucleic acid molecule of the composition comprises from 5’ to 3’ a sequence encoding an anti-CD22 CAR, a sequence encoding a P2A selfcleaving peptide, a sequence encoding an anti-CD19 CAR, a sequence encoding a T2A self-cleaving peptide, and a sequence encoding an anti-CD20 CAR.

[0733] In some embodiments, the recombinant polynucleic acid molecule of the composition comprises a single polynucleic acid comprising a sequence encoding a CD22 CAR (SEQ ID NO: 996), a P2A ribosome skipping peptide, a CD19 CAR (SEQ ID NO: 706), a T2A ribosome skipping peptide, and a CD20 CAR (SEQ ID NO: 1358) together having the sequenceMLLLVTSLLLCELPHPAFLLIPQVQLQQSGPGLVKPSQTLSLTCAISGDSVSSNSAAWNWIRQ SPSRGLEWLGRTYYRSKWYNDYAVSVKSRITINPDTSKNQFSLQLNSVTPEDTAVYYCARE VTGDLEDAFDIWGQGTMVTVSSGGGGSDIQMTQSPSSLSASVGDRVTITCRASQTIWSYLN WYQQRPGKAPNLLIYAASSLQSGVPSRFSGRGSGTDFTLTISSLQAEDFATYYCQQSYSIPQT FGQGTKLEIKAAATTTPAPRPPTPAPTIASQPLSLRPEACRPAAGGAVHTRGLDFACDIYIWA PLAGTCGVLLLSLVITLYCKRGRKKLLYIFKQPFMRPVQTTQEEDGCSCRFPEEEEGGCELR VKFSRSADAPAYKQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLYN ELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPRATNFSLLK QAGDVEENPGPMALPVTALLLPLALLLHAARPEVQLVQSGAEVKKPGESLKISCKASGYRF TNYWIAWVRQRPGKGLEWMGRIDPSDSYTHYSPSFQGHVTMSTDKSISTAYLQWSSLKAS DTAMYYCARPGDILTGWAMDVWGQGTLVTVSSAAASGGGGSGGGGSGGGGSALQSVLTQ PPS VS AAPGQKVTISC SGS S SNIGNNYVSWYQQLPGTAPKLLIYDNNKRPSGIPDRF SGSKSG TSATLGITGLQAEDEADYYCQSYDSSLSGNYVFGTGTKVTVLIEVMYPPPYLDNEKSNGTII HVKGKHLCPSPLFPGPSKPFWVLVVVGGVLACYSLLVTVAFIIFWVRSKRSRLLHSDYMNM TPRRPGPTRKHYQPYAPPRDFAAYRSRVKFSRSADAPAYKQGQNQLYNELNLGRREEYDVL DKRRGRDPEMGGKPRRKNPQEGLYNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLS TATKDTYDALHMQALPPREGRGSLLTCGDVEENPGPMLLLVTSLLLCELPHPAFLLIPDIVM TQSPDSLAVSLGERATINCKSSQSVLYSSNNKNYLAWYQQKPGQPPKLLIYWASTRESGVPD RFSGSGSGSDFTLTISSLQAEDVAVYYCQQYYSFYQTFGQGTKVEIKGGGGSGGGGSGGGG SEVQLVESGGGVVRPGGSLRLSCTASGFTFGDYGMSWVRQAPGKGLEWVSGINWNGGSTG YADSVKGRFTISRDNAKNSLYLQMNSLRAEDTALYYCARKSYYGSGSPDVFDIWGQGTMV TVSSIEVMYPPPYLDNEKSNGTIIHVKGKHLCPSPLFPGPSKPFWVLVVVGGVLACYSLLVT VAFIIFWVKRKKQRSRRNDEELETRAHRVATEERGRKPHQIPASTPQNPATSQHPPPPPGHRS QAPSHRPPPPGHRVQHQPQKRPPAPSGTQVHQQKGPPLPRPRVQPKPPHGAAENSLSPSSNR VKFSRSADAPAYKQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLYN ELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 1006).

[0734] In some embodiments, the recombinant polynucleic acid molecule of the composition comprises a single polynucleic acid comprising the sequence atgcttctgctcgtgacaagcctgctgctgtgcgagctgccccaccctgcctttctgctgatccctCAAGTCCAGCTGCAGCAAT CTGGCCCTGGCCTGGTGAAGCCTAGCCAGACCCTGAGCCTGACATGTGCCATCAGCGGA GATTCTGTTAGCAGCAACAGCGCGGCCTGGAACTGGATCAGACAGAGCCCTAGCAGAG GACTGGAGTGGCTGGGCCGGACCTACTACAGAAGCAAGTGGTACAACGACTACGCCGTGTCCGTGAAAAGCAGAATCACCATCAACCCCGACACCAGCAAGAACCAGTTCTCCCTGCAACTGAATAGCGTCACACCCGAGGATACAGCCGTGTACTACTGCGCCAGAGAGGTTACAGGCGACCTGGAAGATGCCTTTGACATCTGGGGCCAGGGCACGATGGTGACCGTCAGTAGCGGCGGCGGAGGAAGCGACATCCAGATGACCCAATCTCCTAGCAGCCTGTCTGCCTCTGTGGGCGACAGAGTGACCATCACCTGCCGCGCCTCTCAAACAATCTGGTCCTACCTGAACTGGTATCAGCAAAGACCTGGCAAAGCCCCAAACCTGCTGATCTACGCCGCCAGCTCCCTGCAAAGCGGAGTGCCCAGCAGATTCAGCGGCCGGGGCAGCGGCACTGATTTTACCCTGACTATTTCTTCCCTGCAAGCCGAGGACTTCGCCACATACTACTGTCAGCAGAGCTACAGCATCCCTCAGACCTTCGGCCAGGGCACCAAGCTGGAAATCAAGGCGGCCGCCACAACCACCCCTGCTCCCCGCCCACCTACCCCTGCTCCTACAATCGCCTCGCAACCTCTGAGCCTGAGACCTGAGGCCTGTAGACCCGCCGCTGGCGGCGCCGTGCACACCAGAGGCCTGGACTTCGCCTGCGACATCTACATCTGGGCCCCACTGGCCGGCACATGCGGCGTGCTGCTGCTGTCCCTGGTGATCACCCTGTATTGCAAGAGAGGCAGAAAGAAACTGCTGTACATCTTCAAGCAGCCTTTCATGCGGCCCGTGCAGACCACCCAGGAGGAAGATGGTTGCTCATGCCGGTTCCCCGAGGAAGAGGAAGGCGGATGTGAACTGAGAGTGAAGTTCAGCCGCTCCGCTGACGCACCCGCCTATAAACAGGGGCAGAATCAACTTTATAACGAACTGAACCTTGGAAGAAGGGAGGAATATGACGTGCTTGATAAGAGAAGAGGCAGAGATCCTGAAATGGGAGGGAAACCTCGGAGAAAAAATCCACAAGAGGGGCTCTACAATGAACTCCAGAAAGACAAGATGGCTGAAGCTTACTCTGAGATTGGAATGAAAGGGGAGCGGCGCCGGGGCAAAGGGCACGACGGATTGTACCAAGGACTCAGTACAGCTACTAAAGACACATATGACGCCCTGCATATGCAGGCACTGCCCCCACGGGCCACCAACTTCAGCCTGCTGAAGCAGGCCGGCGACGTGGAAGAGAACCCCGGCCCTATGGCCTTGCCCGTGACTGCTCTGCTCCTGCCCCTCGCACTGCTGCTGCACGCCGCGAGGCCCGAAGTGCAGCTGGTCCAATCTGGCGCCGAGGTGAAGAAGCCCGGCGAGAGCCTGAAGATCAGCTGTAAAGCCAGCGGATATAGATTCACCAACTACTGGATCGCCTGGGTGCGGCAGCGGCCTGGCAAGGGCCTGGAATGGATGGGCAGAATCGACCCATCTGATAGCTACACCCACTACAGCCCTAGCTTCCAAGGCCACGTGACCATGAGCACAGATAAGAGCATCAGCACCGCCTACCTGCAGTGGTCCTCCCTGAAAGCTAGCGACACCGCTATGTACTACTGCGCCAGACCTGGCGACATCCTGACCGGCTGGGCTATGGATGTGTGGGGCCAGGGAACACTGGTGACAGTGTCCAGCGCTGCAGCAAGTGGAGGCGGAGGAAGTGGAGGCGGTGGTTCAGGAGGAGGAGGATCTGCTCTGCAATCTGTGCTCACACAGCCCCCCAGCGTGTCTGCCGCTCCTGGACAGAAAGTGACCATCAGCTGCAGCGGATCTAGCTCCAACATCGGCAACAACTACGTGTCCTGGTATCAGCAGCTGCCTGGCACCGCCCCTAAGCTGCTGATCTACGACAACAACAAGCGGCCATCTGGCATCCCTGATCGCTTCAGCGGCAGCAAGTCCGGCACCAGCGCCACCCTGGGAATCACCGGCCTGCAGGCCGAGGACGAAGCCGACTACTACTGTCAGAGCTACGATAGCAGCCTGAGCGGCAATTACGTGTTCGGCACAGGCACAAAGGTCACCGTGCTGATCGAGGTGATGTACCCTCCTCCTTACCTGGATAATGAGAAGTCCAACGGAACAATCATCCACGTGAAGGGCAAGCACCTGTGCCCCTCTCCTCTGTTTCCGGGCCCTTCTAAGCCCTTCTGGGTGCTGGTCGTGGTGGGCGGAGTGCTCGCTTGTTACTCTCTGCTTGTGACCGTGGCCTTCATTATCTTCTGGGTCCGCAGCAAGCGGTCCAGACTGCTGCACAGCGACTACATGAACATGACCCCTCGGAGACCTGGACCCACCAGAAAGCACTACCAGCCTTACGCCCCTCCACGCGACTTCGCCGCTTATCGCAGCCGCGTTAAATTTTCACGCAGCGCGGACGCCCCTGCGTACAAGCAGGGACAGAACCAGCTGTACAACGAGCTCAACCTGGGCCGCCGGGAGGAGTACGACGTGCTGGACAAGCGCCGGGGGCGTGATCCAGAGATGGGCGGCAAACCCCGCCGCAAGAATCCTCAGGAGGGCTTATACAACGAGCTGCAGAAGGACAAAATGGCCGAGGCCTACAGCGAGATCGGCATGAAGGGAGAGCGCCGGCGTGGAAAGGGCCATGACGGCCTCTATCAGGGTCTGTCCACCGCCACCAAGGACACCTACGACGCTCTACACATGCAGGCCTTGCCTCCGCGCGAGGGCAGAGGCAGCCTGCTGACCTGCGGCGATGTGGAGGAGAACCCCGGACCTATGCTGTTGCTTGTTACAAGCCTCCTCCTGTGCGAATTGCCTCACCCCGCATTTCTCCTGATACCCGACATCGTGATGACCCAGTCCCCAGACAGCCTGGCTGTGTCCCTGGGCGAACGGGCCACCATCAACTGCAAGAGCAGCCAGAGCGTGCTGTACAGCAGCAACAACAAGAATTACCTGGCCTGGTACCAGCAGAAACCTGGCCAGCCTCCCAAGCTGCTGATCTACTGGGCCAGCACCAGAGAGAGCGGAGTGCCTGATAGATTCAGCGGCAGCGGATCTGGCTCTGATTTTACCCTGACAATCAGCAGCCTCCAGGCCGAGGACGTCGCCGTGTACTATTGTCAGCAATACTACTCCTTCTACCAAACATTCGGCCAGGGCACCAAAGTTGAAATCAAGGGAGGAGGAGGGTCTGGAGGTGGAGGAAGTGGCGGAGGGGGAAGTGAGGTGCAGCTGGTCGAGAGCGGCGGAGGCGTGGTGCGGCCTGGAGGATCTCTCAGACTGAGCTGTACCGCCAGCGGATTTACCTTCGGCGACTACGGCATGAGCTGGGTGCGCCAGGCCCCTGGCAAAGGCCTGGAATGGGTTTCCGGCATCAACTGGAACGGCGGCTCTACAGGCTACGCCGACAGCGTGAAGGGCAGATTCACCATCAGCCGGGACAACGCCAAGAATAGCCTGTACCTGCAAATGAACAGCCTGAGAGCTGAAGATACAGCTCTGTATTACTGCGCCAGAAAGTCCTACTACGGCTCTGGCAGCCCCGATGTGTTCGACATCTGGGGCCAGGGCACAATGGTGACCGTGTCCAG...

Claims

CLAIMSWHAT IS CLAIMED IS:

1. A composition comprising one or more recombinant polynucleic acid molecules comprising:(a) a sequence encoding an anti-CD19 chimeric antigen receptor (CAR), the anti-CD19CAR comprising:(i) an extracellular domain comprising an anti-CD19 binding domain;(ii) a transmembrane domain; and(iii) a cytoplasmic domain comprising an intracellular signaling domain comprising one or more immunoreceptor tyrosine-based activation motifs (IT AMs) and optionally an intracellular costimulatory signaling domain; and(b) a sequence encoding an anti-CD20 CAR, the anti-CD20 CAR comprising:(i) an extracellular domain comprising an anti-CD20 binding domain;(ii) a transmembrane domain; and(iii) a cytoplasmic domain comprising an intracellular signaling domain comprising one or more IT AMs and optionally an intracellular costimulatory signaling domain derived from CD2; and(c) a sequence encoding an anti-CD22 CAR, the anti-CD22 CAR comprising:(i) an extracellular domain comprising an anti-CD22 binding domain;(ii) a transmembrane domain; and(iii) a cytoplasmic domain comprising an intracellular signaling domain comprising one or more IT AMs and optionally an intracellular costimulatory signaling domain.2 The composition of claim 1, wherein the sequence encoding an anti-CD19 CAR, the sequence encoding an anti-CD20 CAR, and the sequence encoding an anti-CD22 CAR are encoded on the same polynucleic acid molecule.3 The composition of any one of claims 1 to 2, wherein the sequence encoding the anti-CD19 CAR, the sequence encoding the anti-CD20 CAR, and the sequence encoding the anti-CD22 CAR are each separated from the sequences encoding the other CARs by a sequence encoding a viral ribosome skipping peptide selected from the group consisting of a P2A peptide, a T2A peptide, an E2A peptide, and an F2A peptide.The composition of any one of claims 1 to 3, wherein the single recombinant polynucleic acid comprises, from 5’ to 3’, a sequence encoding the anti-CD22 CAR; a sequence encoding a viral ribosome skipping peptide selected from the group consisting of a P2A peptide, a T2A peptide, an E2A peptide, and an F2A peptide; a sequence encoding the anti -CD 19 CAR; a sequence encoding a viral ribosome skipping peptide selected from the group consisting of a P2A peptide, a T2A peptide, an E2A peptide, and an F2A peptide; and a sequence encoding the anti-CD20 CAR. The composition of any one of claims 1 to 4, wherein the single recombinant polynucleic acid comprises, from 5’ to 3’, a sequence encoding the anti-CD22 CAR; a sequence encoding a viral P2A ribosome skipping peptide; a sequence encoding the anti-CD19 CAR; a sequence encoding a viral T2A ribosome skipping peptide; and a sequence encoding the anti-CD20 CAR. The composition of any one of claims 1 to 5, wherein the anti-CD22 CAR binding domain comprises an anti-CD22 antibody or an antigen-binding fragment thereof. The composition of any one of claims 1 to 6, wherein the anti-CD22 CAR binding domain is an antigen-binding fragment of an anti-CD22 antibody. The composition of any one of claims 1 to 7, wherein the antigen-binding fragment of an anti-CD22 antibody is a single chain variable fragment (scFv). The composition of any one of claims 1 to 8, wherein the anti-CD22 binding domain comprises a heavy chain variable region (VH) that comprises a first heavy chain complementarity determining region (HCDR1) having the sequence GDSVSSNSA (SEQ ID NO: 667), a second heavy chain complementarity determining region (HCDR2) having the sequence TYYRSKWYN (SEQ ID NO: 675), and a third heavy chain complementarity determining region (HCDR3) having the sequence AREVTGDLEDAFDI (SEQ ID NO: 676). The composition of one of claims 1 to 9, wherein anti-CD22 binding domain comprises a light chain variable region (VL) that comprises a first light chain complementarity determining region (LCDR1) having the sequence QTIWSY (SEQ ID NO: 677), a secondlight chain complementarity determining region (LCDR2) having the sequence AASSLQS (SEQ ID NO: 671), and a third light chain complementarity determining region (LCDR3) having the sequence AAS (SEQ ID NO: 678).

11. The composition of any one of claims 1 to 10, wherein the anti-CD22 binding domain comprises a VH comprising a sequence having at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the sequence QVQLQQSGPGLVKPSQTLSLTCAISGDSVSSNSAAWNWIRQSPSRGLEWLGRTYYRS KWYNDYAVSVKSRITINPDTSKNQFSLQLNSVTPEDTAVYYCAREVTGDLEDAFDIW GQGTMVTVSS (SEQ ID NO: 665) and a VL comprising a sequence having at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the sequence DIQMTQSPSSLSASVGDRVTITCRASQTIWSYLNWYQQRPGKAPNLLIYAASSLQSGV PSRFSGRGSGTDFTLTISSLQAEDFATYYCQQSYSIPQTFGQGTKLEIK (SEQ ID NO: 666).

12. The composition of claim 11, wherein the anti-CD22 binding domain comprises a VH having the sequence QVQLQQSGPGLVKPSQTLSLTCAISGDSVSSNSAAWNWIRQSPSRGLEWLGRTYYRS KWYNDYAVSVKSRITINPDTSKNQFSLQLNSVTPEDTAVYYCAREVTGDLEDAFDIW GQGTMVTVSS (SEQ ID NO: 665) and a VL having the sequence DIQMTQSPSSLSASVGDRVTITCRASQTIWSYLNWYQQRPGKAPNLLIYAASSLQSGV PSRFSGRGSGTDFTLTISSLQAEDFATYYCQQSYSIPQTFGQGTKLEIK (SEQ ID NO: 666).

13. The composition of claims 11 or 12, wherein the VH and VL are linked by a linker comprising the sequence GGGGS (SEQ ID NO: 1007).

14. The composition of any one of claims 1 to 13, wherein the anti-CD22 binding domain comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the sequence QVQLQQSGPGLVKPSQTLSLTCAISGDSVSSNSAAWNWIRQSPSRGLEWLGRTYYRS KWYNDYAVSVKSRITINPDTSKNQFSLQLNSVTPEDTAVYYCAREVTGDLEDAFDIW GQGTMVTVSSGGGGSDIQMTQSPSSLSASVGDRVTITCRASQTIWSYLNWYQQRPGKAPNLLIYAASSLQSGVPSRFSGRGSGTDFTLTISSLQAEDFATYYCQQSYSIPQTFGQ GTKLEIK (SEQ ID NO: 664).

15. The composition of claim 14, wherein the anti-CD22 binding domain comprises an scFv having the sequence QVQLQQSGPGLVKPSQTLSLTCAISGDSVSSNSAAWNWIRQSPSRGLEWLGRTYYRS KWYNDYAVSVKSRITINPDTSKNQFSLQLNSVTPEDTAVYYCAREVTGDLEDAFDIW GQGTMVTVSSGGGGSDIQMTQSPSSLSASVGDRVTITCRASQTIWSYLNWYQQRPG KAPNLLIYAASSLQSGVPSRFSGRGSGTDFTLTISSLQAEDFATYYCQQSYSIPQTFGQ GTKLEIK (SEQ ID NO: 664).

16. The composition of any one of claims 1 to 15, wherein the anti-CD22 CAR further comprises a hinge domain.

17. The composition of claim 16, wherein the hinge domain of the anti-CD22 CAR is derived from CD8a or from CD28 and the transmembrane domain of the anti-CD22 CAR is derived from CD8a or from CD28.

18. The composition of claim 17, wherein the hinge domain of the anti-CD22 CAR is derived from CD8a and the transmembrane domain is derived from CD8a.

19. The composition of any one of claims 16 to 18, wherein the CD8a hinge domain has the sequence TTTPAPRPPTPAPTIASQPLSLRPEACRPAAGGAVHTRGLDFACD (SEQ ID NO: 679).

20. The composition of any one of claims 16 to 19, wherein the CD8a transmembrane domain has the sequence IYIWAPLAGTCGVLLLSLVIT (SEQ ID NO: 680).

21. The composition of any one of claims 16 to 20, wherein the CD8a transmembrane domain further comprises a portion of the CD8a cytoplasmic domain having the sequence LYC (SEQ ID NO: 681).

22. The composition of any one of claims 1 to 21, wherein the cytoplasmic domain of the anti- CD22 CAR comprises an intracellular co-stimulatory domain derived from a molecule selected from the group consisting of CD27, CD28, 4-1BB / CD137, 0X40, ICOS, and CD2.

23. The composition of claim 22, wherein the intracellular co-stimulatory domain is derived from 4-1BB / CD137.

24. The composition of claim 23, wherein the 4-1BB / CD137 intracellular co-stimulatory domain comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the sequence KRGRKKLLYIFKQPFMRPVQTTQEEDGCSCRFPEEEEGGCEL (SEQ ID NO: 998).

25. The composition of claim 24, wherein the 4-1BB / CD137 intracellular co-stimulatory domain has the sequence KRGRKKLLYIFKQPFMRPVQTTQEEDGCSCRFPEEEEGGCEL (SEQ ID NO: 998).

26. The composition of any one of claims 1 to 25, wherein the cytoplasmic domain of the anti- CD22 CAR comprises an intracellular signaling domain comprising one or more ITAMs derived from CD3^.

27. The composition of claim 26, wherein the CD3^ intracellular signaling domain comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the sequence RVKFSRSADAPAYKQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQ EGLYNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQAL PPR (SEQ ID NO: 686).

28. The composition of claim 27, wherein the CD3^ intracellular signaling domain has the sequence RVKFSRSADAPAYKQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQ EGLYNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQAL PPR (SEQ ID NO: 686).

29. The composition of claim 26, wherein the CD3^ intracellular signaling domain comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the sequence RVKFSRSADAPAYQQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQ EGLYNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQAL PPR (SEQ ID NO: 687).

30. The composition of claim 29, wherein the CD3^ intracellular signaling domain has the sequence RVKFSRSADAPAYQQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQ EGLYNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQAL PPR (SEQ ID NO: 687).

31. The composition of claim 26, wherein the CD3^ intracellular signaling domain comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the sequence RVKFSRSADAPAYQQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPQRRKNP QEGLYNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQA LPPR (SEQ ID NO: 688).

32. The composition of claim 31, wherein the CD3^ intracellular signaling domain has the sequence RVKFSRSADAPAYQQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPQRRKNP QEGLYNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQA LPPR (SEQ ID NO: 688).

33. The composition of claim 26, wherein the CD3^ intracellular signaling domain comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the sequence RVKFSRSADAPAYKQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPQRRKNP QEGLYNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQA LPPR (SEQ ID NO: 689).

34. The composition of claim 33, wherein the CD3^ intracellular signaling domain has the sequence RVI<FSRSADAPAYI<QGQNQLYNELNLGRREEYDVLDI<RRGRDPEMGGI<PQRRI<NP QEGLYNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQA LPPR (SEQ ID NO: 689).

35. The composition of any one of claims 1 to 34, wherein the anti-CD22 CAR comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the sequence QVQLQQSGPGLVKPSQTLSLTCAISGDSVSSNSAAWNWIRQSPSRGLEWLGRTYYRS KWYNDYAVSVKSRITINPDTSKNQFSLQLNSVTPEDTAVYYCAREVTGDLEDAFDIW GQGTMVTVSSGGGGSDIQMTQSPSSLSASVGDRVTITCRASQTIWSYLNWYQQRPG KAPNLLIYAASSLQSGVPSRFSGRGSGTDFTLTISSLQAEDFATYYCQQSYSIPQTFGQ GTKLEIKAAATTTPAPRPPTPAPTIASQPLSLRPEACRPAAGGAVHTRGLDFACDIYIW APLAGTCGVLLLSLVITLYCKRGRKKLLYIFKQPFMRPVQTTQEEDGCSCRFPEEEEG GCELRVKFSRSADAPAYKQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRR KNPQEGLYNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALH MQALPPR (SEQ ID NO: 1008).

36. The composition of claim 35, wherein the anti-CD22 CAR has the sequence QVQLQQSGPGLVKPSQTLSLTCAISGDSVSSNSAAWNWIRQSPSRGLEWLGRTYYRS KWYNDYAVSVKSRITINPDTSKNQFSLQLNSVTPEDTAVYYCAREVTGDLEDAFDIW GQGTMVTVSSGGGGSDIQMTQSPSSLSASVGDRVTITCRASQTIWSYLNWYQQRPG KAPNLLIYAASSLQSGVPSRFSGRGSGTDFTLTISSLQAEDFATYYCQQSYSIPQTFGQ GTKLEIKAAATTTPAPRPPTPAPTIASQPLSLRPEACRPAAGGAVHTRGLDFACDIYIW APLAGTCGVLLLSLVITLYCKRGRKKLLYIFKQPFMRPVQTTQEEDGCSCRFPEEEEG GCELRVKFSRSADAPAYKQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRR KNPQEGLYNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALH MQALPPR (SEQ ID NO: 1008).

37. The composition of any one of claims 1 to 36, wherein the anti-CD22 CAR further comprises a signal peptide.

38. The composition of claim 37, wherein the signal peptide sequence is derived from CD8a or granulocyte macrophage colony stimulating factor receptor alpha (GMCSFRa).

39. The composition of claim 38, wherein the signal peptide sequence is derived from GMCSFRa and has the sequence of MLLLVTSLLLCELPHPAFLLIP (SEQ ID NO: 691).

40. The composition of any one of claims 1 to 39, wherein the anti-CD22 CAR comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the sequence MLLLVTSLLLCELPHPAFLLIPQVQLQQSGPGLVKPSQTLSLTCAISGDSVSSNSAAW NWIRQSPSRGLEWLGRTYYRSKWYNDYAVSVKSRITINPDTSKNQFSLQLNSVTPED TAVYYCAREVTGDLEDAFDIWGQGTMVTVSSGGGGSDIQMTQSPSSLSASVGDRVT ITCRASQTIWSYLNWYQQRPGKAPNLLIYAASSLQSGVPSRFSGRGSGTDFTLTISSLQ AEDFATYYCQQSYSIPQTFGQGTKLEIKAAATTTPAPRPPTPAPTIASQPLSLRPEACR PAAGGAVHTRGLDFACDIYIWAPLAGTCGVLLLSLVITLYCKRGRKKLLYIFKQPFM RPVQTTQEEDGCSCRFPEEEEGGCELRVKFSRSADAPAYKQGQNQLYNELNLGRREE YDVLDKRRGRDPEMGGKPRRKNPQEGLYNELQKDKMAEAYSEIGMKGERRRGKG HDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 996).

41. The composition of any one of claims 1 to 40, wherein the anti-CD19 CAR binding domain comprises an anti-CD19 antibody or an antigen-binding fragment thereof.

42. The composition of any one of claims 1 to 41, wherein the anti-CD19 CAR binding domain is an antigen-binding fragment of an anti-CD19 antibody.

43. The composition of any one of claims 1 to 42, wherein the antigen-binding fragment of an anti-CD19 antibody is a single chain variable fragment (scFv).

44. The composition of any one of claims 1 to 43, wherein the anti-CD19 binding domain comprises a heavy chain variable region (VH) that comprises a first heavy chain complementarity determining region (HCDR1) having the sequence GYRFTNYW (SEQ ID NO: 173), a second heavy chain complementarity determining region (HCDR2) having the sequence IDPSDSYT (SEQ ID NO: 174), and a third heavy chain complementaritydetermining region (HCDR3) having the sequence ARPGDILTGWAMDV (SEQ ID NO: 175).

45. The composition of one of claims 1 to 44, wherein anti-CD19 binding domain comprises a light chain variable region (VL) that comprises a first light chain complementarity determining region (LCDR1) having the sequence SSNIGNNY (SEQ ID NO: 297), a second light chain complementarity determining region (LCDR2) having the sequence DNN (SEQ ID NO: 298), and a third light chain complementarity determining region (LCDR3) having the sequence QSYDSSLSGNYV (SEQ ID NO: 299).

46. The composition of any one of claims 1 to 45, wherein the anti-CD19 binding domain comprises a VH comprising a sequence having at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the sequence EVQLVQSGAEVKKPGESLKISCKASGYRFTNYWIAWVRQRPGKGLEWMGRIDPSDS YTHYSPSFQGHVTMSTDKSISTAYLQWSSLKASDTAMYYCARPGDILTGWAMDVW GQGTLVTVSS (SEQ ID NO: 47) and a VL comprising a sequence having at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the sequence QSVLTQPPSVSAAPGQKVTISCSGSSSNIGNNYVSWYQQLPGTAPKLLIYDNNKRPSG IPDRFSGSKSGTSATLGITGLQAEDEADYYCQSYDSSLSGNYVFGTGTKVTVLK (SEQ ID NO: 48).

47. The composition of claim 46, wherein the anti-CD19 binding domain comprises a VH having the sequence EVQLVQSGAEVKKPGESLKISCKASGYRFTNYWIAWVRQRPGKGLEWMGRIDPSDS YTHYSPSFQGHVTMSTDKSISTAYLQWSSLKASDTAMYYCARPGDILTGWAMDVW GQGTLVTVSS (SEQ ID NO: 47) and a VL having the sequence QSVLTQPPSVSAAPGQKVTISCSGSSSNIGNNYVSWYQQLPGTAPKLLIYDNNKRPSG IPDRFSGSKSGTSATLGITGLQAEDEADYYCQSYDSSLSGNYVFGTGTKVTVLK (SEQ ID NO: 48).

48. The composition of claim 46 or 47, wherein the VH and VL are linked by a linker comprising the sequence GGGGSGGGGSGGGGS (SEQ ID NO: 995).

49. The composition of any one of claims 1 to 48, wherein the anti-CD19 binding domain comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the sequence EVQLVQSGAEVKKPGESLKISCKASGYRFTNYWIAWVRQRPGKGLEWMGRIDPSDS YTHYSPSFQGHVTMSTDKSISTAYLQWSSLKASDTAMYYCARPGDILTGWAMDVW GQGTLVTVSSAAASGGGGSGGGGSGGGGSALQSVLTQPPSVSAAPGQKVTISCSGSS SNIGNNYVSWYQQLPGTAPKLLIYDNNKRPSGIPDRFSGSKSGTSATLGITGLQAEDE ADYYCQSYDSSLSGNYVFGTGTKVTVL (SEQ ID NO: 11).

50. The composition of claim 49, wherein the anti-CD19 binding domain comprises an scFv having the sequence EVQLVQSGAEVKKPGESLKISCKASGYRFTNYWIAWVRQRPGKGLEWMGRIDPSDS YTHYSPSFQGHVTMSTDKSISTAYLQWSSLKASDTAMYYCARPGDILTGWAMDVW GQGTLVTVSSAAASGGGGSGGGGSGGGGSALQSVLTQPPSVSAAPGQKVTISCSGSS SNIGNNYVSWYQQLPGTAPKLLIYDNNKRPSGIPDRFSGSKSGTSATLGITGLQAEDE ADYYCQSYDSSLSGNYVFGTGTKVTVL (SEQ ID NO: 11).

51. The composition of any one of claims 1 to 50, wherein the anti-CD19 CAR further comprises a hinge domain.

52. The composition of claim 51, wherein the hinge domain of the anti-CD19 CAR is derived from CD8a or from CD28 and the transmembrane domain of the anti-CD19 CAR is derived from CD8a or from CD28.

53. The composition of claim 52, wherein the hinge domain of the anti-CD19 CAR is derived from CD28 and the transmembrane domain is derived from CD28.

54. The composition of any one of claims 51 to 53, wherein the CD28 hinge domain has the sequence IEVMYPPPYLDNEKSNGTIIHVKGKHLCPSPLFPGPSKP (SEQ ID NO: 682).

55. The composition of any one of claims 52 to 54, wherein the CD28 transmembrane domain has the sequence FWVLVVVGGVLACYSLLVTVAFIIFWV (SEQ ID NO: 683).

56. The composition of any one of claims 1 to 55, wherein the cytoplasmic domain of the antiCD 19 CAR comprises an intracellular co-stimulatory domain derived from a molecule selected from the group consisting of CD27, CD28, 4-1BB / CD137, 0X40, ICOS, and CD2.

57. The composition of claim 56, wherein the intracellular co-stimulatory domain is derived from CD28.

58. The composition of claim 57, wherein the CD28 intracellular co-stimulatory domain comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the sequence RSKRSRLLHSDYMNMTPRRPGPTRKHYQPYAPPRDFAAYRS (SEQ ID NO: 684).

59. The composition of claim 58, wherein the CD28 intracellular co-stimulatory domain has the sequence RSKRSRLLHSDYMNMTPRRPGPTRKHYQPYAPPRDFAAYRS (SEQ ID NO: 684).

60. The composition of any one of claims 1 to 59, wherein the cytoplasmic domain of the anti- CD19 CAR comprises an intracellular signaling domain comprising one or more IT AMs derived from CD3^.

61. The composition of claim 60, wherein the CD3^ intracellular signaling domain comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the sequence RVKFSRSADAPAYKQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQ EGLYNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQAL PPR (SEQ ID NO: 686).

62. The composition of claim 61, wherein the CD3^ intracellular signaling domain has the sequence RVKFSRSADAPAYKQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQ EGLYNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQAL PPR (SEQ ID NO: 686).

63. The composition of claim 60, wherein the CD3^ intracellular signaling domain comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the sequence RVKFSRSADAPAYQQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQ EGLYNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQAL PPR (SEQ ID NO: 687).

64. The composition of claim 63, wherein the CD3^ intracellular signaling domain has the sequence RVKFSRSADAPAYQQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQ EGLYNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQAL PPR (SEQ ID NO: 687).

65. The composition of claim 60, wherein the CD3^ intracellular signaling domain comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the sequence RVKFSRSADAPAYQQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPQRRKNP QEGLYNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQA LPPR (SEQ ID NO: 688).

66. The composition of claim 65, wherein the CD3^ intracellular signaling domain has the sequence RVKFSRSADAPAYQQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPQRRKNP QEGLYNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQA LPPR (SEQ ID NO: 688).

67. The composition of claim 60, wherein the CD3^ intracellular signaling domain comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the sequence RVKFSRSADAPAYKQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPQRRKNP QEGLYNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQA LPPR (SEQ ID NO: 689).

68. The composition of claim 67, wherein the CD3^ intracellular signaling domain has the sequence RVI<FSRSADAPAYI<QGQNQLYNELNLGRREEYDVLDI<RRGRDPEMGGI<PQRRI<NP QEGLYNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQA LPPR (SEQ ID NO: 689).

69. The composition of any one of claims 1 to 68, wherein the anti-CD19 CAR comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the sequence EVQLVQSGAEVKKPGESLKISCKASGYRFTNYWIAWVRQRPGKGLEWMGRIDPSDS YTHYSPSFQGHVTMSTDKSISTAYLQWSSLKASDTAMYYCARPGDILTGWAMDVW GQGTLVTVSSAAASGGGGSGGGGSGGGGSALQSVLTQPPSVSAAPGQKVTISCSGSS SNIGNNYVSWYQQLPGTAPKLLIYDNNKRPSGIPDRFSGSKSGTSATLGITGLQAEDE ADYYCQSYDSSLSGNYVFGTGTKVTVLIEVMYPPPYLDNEKSNGTIIHVKGKHLCPS PLFPGPSKPFWVLVVVGGVLACYSLLVTVAFIIFWVRSKRSRLLHSDYMNMTPRRPG PTRI<HYQPYAPPRDFAAYRSRVI<FSRSADAPAYI<QGQNQLYNELNLGRREEYDVLD KRRGRDPEMGGKPRRKNPQEGLYNELQKDKMAEAYSEIGMKGERRRGKGHDGLY QGLSTATKDTYDALHMQALPPR (SEQ ID NO: 1121).

70. The composition of claim 69, wherein the anti-CD19 CAR has the sequence EVQLVQSGAEVKKPGESLKISCKASGYRFTNYWIAWVRQRPGKGLEWMGRIDPSDS YTHYSPSFQGHVTMSTDKSISTAYLQWSSLKASDTAMYYCARPGDILTGWAMDVW GQGTLVTVSSAAASGGGGSGGGGSGGGGSALQSVLTQPPSVSAAPGQKVTISCSGSS SNIGNNYVSWYQQLPGTAPKLLIYDNNKRPSGIPDRFSGSKSGTSATLGITGLQAEDE ADYYCQSYDSSLSGNYVFGTGTKVTVLIEVMYPPPYLDNEKSNGTIIHVKGKHLCPS PLFPGPSKPFWVLVVVGGVLACYSLLVTVAFIIFWVRSKRSRLLHSDYMNMTPRRPG PTRI<HYQPYAPPRDFAAYRSRVI<FSRSADAPAYI<QGQNQLYNELNLGRREEYDVLD KRRGRDPEMGGKPRRKNPQEGLYNELQKDKMAEAYSEIGMKGERRRGKGHDGLY QGLSTATKDTYDALHMQALPPR (SEQ ID NO: 1121).

71. The composition of any one of claims 1 to 70, wherein the anti-CD19 CAR further comprises a signal peptide.

72. The composition of claim 71, wherein the signal peptide sequence is derived from CD8a or granulocyte macrophage colony stimulating factor receptor alpha (GMCSFRa).

73. The composition of claim 72, wherein the signal peptide sequence is derived from CD8a and has the sequence of MALPVTALLLPLALLLHAARP (SEQ ID NO: 690).

74. The composition of any one of claims 1 to 73, wherein the anti-CD19 CAR comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the sequence MALPVTALLLPLALLLHAARPEVQLVQSGAEVKKPGESLKISCKASGYRFTNYWIA WVRQRPGKGLEWMGRIDPSDSYTHYSPSFQGHVTMSTDKSISTAYLQWSSLKASDT AMYYCARPGDILTGWAMDVWGQGTLVTVSSAAASGGGGSGGGGSGGGGSALQSV LTQPPSVSAAPGQKVTISCSGSSSNIGNNYVSWYQQLPGTAPKLLIYDNNKRPSGIPD RFSGSKSGTSATLGITGLQAEDEADYYCQSYDSSLSGNYVFGTGTKVTVLIEVMYPPP YLDNEKSNGTIIHVKGKHLCPSPLFPGPSKPFWVLVVVGGVLACYSLLVTVAFIIFWV RSKRSRLLHSDYMNMTPRRPGPTRKHYQPYAPPRDFAAYRSRVKFSRSADAPAYKQ GQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLYNELQKDKMAE AYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 706).

75. The composition of any one of claims 1 to 74, wherein the anti-CD20 CAR binding domain comprises an anti-CD20 antibody or an antigen-binding fragment thereof.

76. The composition of any one of claims 1 to 75, wherein the anti-CD20 CAR binding domain is an antigen-binding fragment of an anti-CD20 antibody.

77. The composition of any one of claims 1 to 76, wherein the antigen-binding fragment of an anti-CD20 antibody is a single chain variable fragment (scFv).

78. The composition of any one of claims 1 to 77, wherein the anti-CD20 binding domain comprises a heavy chain variable region (VH) that comprises a first heavy chain complementarity determining region (HCDR1) having the sequence GFTFGDYG (SEQ ID NO: 562), a second heavy chain complementarity determining region (HCDR2) having the sequence INWNGGST (SEQ ID NO: 513), and a third heavy chain complementaritydetermining region (HCDR3) having the sequence ARKSYYGSGSPDVFDI (SEQ ID NO: 563).

79. The composition of one of claims 1 to 78, wherein anti-CD20 binding domain comprises a light chain variable region (VL) that comprises a first light chain complementarity determining region (LCDR1) having the sequence QSVLYSSNNKNY (SEQ ID NO: 635), a second light chain complementarity determining region (LCDR2) having the sequence WAS (SEQ ID NO: 636), and a third light chain complementarity determining region (LCDR3) having the sequence QQYYSFYQT (SEQ ID NO: 629).

80. The composition of any one of claims 1 to 79, wherein the anti-CD20 binding domain comprises a VH comprising a sequence having at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the sequence EVQLVESGGGVVRPGGSLRLSCTASGFTFGDYGMSWVRQAPGKGLEWVSGINWNG GSTGYADSVKGRFTISRDNAKNSLYLQMNSLRAEDTALYYCARKSYYGSGSPDVFDI WGQGTMVTVSS (SEQ ID NO: 425) and a VL comprising a sequence having at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the sequence DIVMTQSPDSLAVSLGERATINCKSSQSVLYSSNNKNYLAWYQQKPGQPPKLLIYWA STRESGVPDRFSGSGSGSDFTLTISSLQAEDVAVYYCQQYYSFYQTFGQGTKVEIK (SEQ ID NO: 426).

81. The composition of claim 78, wherein the anti-CD20 binding domain comprises a VH having the sequence EVQLVESGGGVVRPGGSLRLSCTASGFTFGDYGMSWVRQAPGKGLEWVSGINWNG GSTGYADSVKGRFTISRDNAKNSLYLQMNSLRAEDTALYYCARKSYYGSGSPDVFDI WGQGTMVTVSS (SEQ ID NO: 425) and a VL having the sequence DIVMTQSPDSLAVSLGERATINCKSSQSVLYSSNNKNYLAWYQQKPGQPPKLLIYWA STRESGVPDRFSGSGSGSDFTLTISSLQAEDVAVYYCQQYYSFYQTFGQGTKVEIK (SEQ ID NO: 426).

82. The composition of claim 80 or 81, wherein the VH and VL are linked by linker comprising the sequence GGGGSGGGGSGGGGS (SEQ ID NO: 995).

83. The composition of any one of claims 1 to 82, wherein the anti-CD20 binding domain comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the sequence DIVMTQSPDSLAVSLGERATINCKSSQSVLYSSNNKNYLAWYQQKPGQPPKLLIYWA STRESGVPDRFSGSGSGSDFTLTISSLQAEDVAVYYCQQYYSFYQTFGQGTKVEIKGG GGSGGGGSGGGGSEVQLVESGGGVVRPGGSLRLSCTASGFTFGDYGMSWVRQAPG KGLEWVSGINWNGGSTGYADSVKGRFTISRDNAKNSLYLQMNSLRAEDTALYYCA RKSYYGSGSPDVFDIWGQGTMVTVSS (SEQ ID NO: 363).

84. The composition of claim 83, wherein the anti-CD20 binding domain comprises an scFv having the sequence DIVMTQSPDSLAVSLGERATINCKSSQSVLYSSNNKNYLAWYQQKPGQPPKLLIYWA STRESGVPDRFSGSGSGSDFTLTISSLQAEDVAVYYCQQYYSFYQTFGQGTKVEIKGG GGSGGGGSGGGGSEVQLVESGGGVVRPGGSLRLSCTASGFTFGDYGMSWVRQAPG KGLEWVSGINWNGGSTGYADSVKGRFTISRDNAKNSLYLQMNSLRAEDTALYYCA RKSYYGSGSPDVFDIWGQGTMVTVSS (SEQ ID NO: 363).

85. The composition of any one of claims 1 to 84, wherein the anti-CD20 CAR further comprises a hinge domain.

86. The composition of claim 85, wherein the hinge domain of the anti-CD20 CAR is derived from CD8a or from CD28 and the transmembrane domain of the anti-CD20 CAR is derived from CD8a or from CD28.

87. The composition of claim 86, wherein the hinge domain of the anti-CD20 CAR is derived from CD28 and the transmembrane domain is derived from CD28.

88. The composition of any one of claims 85 to 87, wherein the CD28 hinge domain has the sequence lEVMYPPPYLDNEKSNGTIIHVKGKHLCPSPLFPGPSKP (SEQ ID NO: 682).

89. The composition of any one of claims 86 to 88, wherein the CD28 transmembrane domain has the sequence FWVLVVVGGVLACYSLLVTVAFIIFWV (SEQ ID NO: 683).

90. The composition of any one of claims 1 to 89, wherein the cytoplasmic domain of the anti- CD20 CAR comprises an intracellular co-stimulatory domain derived from a molecule selected from the group consisting of CD27, CD28, 4-1BB / CD137, 0X40, ICOS, and CD2.

91. The composition of claim 90, wherein the intracellular co-stimulatory domain is derived from CD2.

92. The composition of claim 91, wherein the CD2 intracellular co-stimulatory domain comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the sequence KRKKQRSRRNDEELETRAHRVATEERGRKPHQIPASTPQNPATSQHPPPPPGHRSQAP SHRPPPPGHRVQHQPQKRPPAPSGTQVHQQKGPPLPRPRVQPKPPHGAAENSLSPSSN (SEQ ID NO: 685).

93. The composition of claim 92, wherein the CD2 intracellular co-stimulatory domain has the sequence KRKKQRSRRNDEELETRAHRVATEERGRKPHQIPASTPQNPATSQHPPPPPGHRSQAP SHRPPPPGHRVQHQPQKRPPAPSGTQVHQQKGPPLPRPRVQPKPPHGAAENSLSPSSN (SEQ ID NO: 685).

94. The composition of any one of claims 1 to 93, wherein the cytoplasmic domain of the anti- CD20 CAR comprises an intracellular signaling domain comprising one or more ITAMs derived from CD3^.

95. The composition of claim 94, wherein the CD3^ intracellular signaling domain comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the sequence RVKFSRSADAPAYKQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQ EGLYNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQAL PPR (SEQ ID NO: 686).

96. The composition of claim 95, wherein the CD3^ intracellular signaling domain has the sequenceRVKFSRSADAPAYKQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLYNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQAL PPR (SEQ ID NO: 686).

97. The composition of claim 94, wherein the CD3^ intracellular signaling domain comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the sequence RVKFSRSADAPAYQQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQ EGLYNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQAL PPR (SEQ ID NO: 687).

98. The composition of claim 97, wherein the CD3^ intracellular signaling domain has the sequence RVKFSRSADAPAYQQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQ EGLYNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQAL PPR (SEQ ID NO: 687).

99. The composition of claim 94, wherein the CD3^ intracellular signaling domain comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the sequence RVKFSRSADAPAYQQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPQRRKNP QEGLYNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQA LPPR (SEQ ID NO: 688).

100. The composition of claim 99, wherein the CD3^ intracellular signaling domain has the sequence RVKFSRSADAPAYQQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPQRRKNP QEGLYNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQA LPPR (SEQ ID NO: 688).

101. The composition of claim 94, wherein the CD3^ intracellular signaling domain comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the sequenceRVKFSRSADAPAYKQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPQRRKNP QEGLYNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQA LPPR (SEQ ID NO: 689).

102. The composition of claim 101, wherein the CD3^ intracellular signaling domain has the sequence RVKFSRSADAPAYKQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPQRRKNP QEGLYNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQA LPPR (SEQ ID NO: 689).

103. The composition of any one of claims 1 to 102, wherein the anti-CD20 CAR comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the sequence MLLLVTSLLLCELPHPAFLLIPDIVMTQSPDSLAVSLGERATINCKSSQSVLYSSNNKN YLAWYQQKPGQPPKLLIYWASTRESGVPDRFSGSGSGSDFTLTISSLQAEDVAVYYC QQYYSFYQTFGQGTKVEIKGGGGSGGGGSGGGGSEVQLVESGGGVVRPGGSLRLSC TASGFTFGDYGMSWVRQAPGKGLEWVSGINWNGGSTGYADSVKGRFTISRDNAKN SLYLQMNSLRAEDTALYYCARKSYYGSGSPDVFDIWGQGTMVTVSSIEVMYPPPYL DNEKSNGTIIHVKGKHLCPSPLFPGPSKPFWVLVVVGGVLACYSLLVTVAFIIFWVKR KKQRSRRNDEELETRAHRVATEERGRKPHQIPASTPQNPATSQHPPPPPGHRSQAPSH RPPPPGHRVQHQPQKRPPAPSGTQVHQQKGPPLPRPRVQPKPPHGAAENSLSPSSNR VKFSRSADAPAYKQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQE GLYNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPP R (SEQ ID NO: 1358).

104. The composition of claim 103, wherein the anti-CD20 CAR has the sequence MLLLVTSLLLCELPHPAFLLIPDIVMTQSPDSLAVSLGERATINCKSSQSVLYSSNNKN YLAWYQQKPGQPPKLLIYWASTRESGVPDRFSGSGSGSDFTLTISSLQAEDVAVYYC QQYYSFYQTFGQGTKVEIKGGGGSGGGGSGGGGSEVQLVESGGGVVRPGGSLRLSC TASGFTFGDYGMSWVRQAPGKGLEWVSGINWNGGSTGYADSVKGRFTISRDNAKN SLYLQMNSLRAEDTALYYCARKSYYGSGSPDVFDIWGQGTMVTVSSIEVMYPPPYL DNEKSNGTIIHVKGKHLCPSPLFPGPSKPFWVLVVVGGVLACYSLLVTVAFIIFWVKR KKQRSRRNDEELETRAHRVATEERGRKPHQIPASTPQNPATSQHPPPPPGHRSQAPSH- TI-RPPPPGHRVQHQPQKRPPAPSGTQVHQQKGPPLPRPRVQPKPPHGAAENSLSPSSNR VKFSRSADAPAYKQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQE GLYNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPP R (SEQ ID NO: 1358).

105. The composition of any one of claims 1 to 104, wherein the anti-CD20 CAR further comprises a signal peptide.

106. The composition of claim 105, wherein the signal peptide sequence is derived from CD8a or granulocyte macrophage colony stimulating factor receptor alpha (GMCSFRa).

107. The composition of claim 106, wherein the signal peptide sequence is derived from GMCSFRa and has the sequence of MLLLVTSLLLCELPHPAFLLIP (SEQ ID NO: 691).

108. The composition of any one of claims 1 to 107, wherein the anti-CD20 CAR comprises a sequence having at least 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the sequence MLLLVTSLLLCELPHPAFLLIPDYKDDDDKDIVMTQSPDSLAVSLGERATINCKSSQS VLYSSNNKNYLAWYQQKPGQPPKLLIYWASTRESGVPDRFSGSGSGSDFTLTISSLQ AEDVAVYYCQQYYSFYQTFGQGTKVEIKGGGGSGGGGSGGGGSEVQLVESGGGVV RPGGSLRLSCTASGFTFGDYGMSWVRQAPGKGLEWVSGINWNGGSTGYADSVKGR FTISRDNAKNSLYLQMNSLRAEDTALYYCARKSYYGSGSPDVFDIWGQGTMVTVSSI EVMYPPPYLDNEKSNGTIIHVKGKHLCPSPLFPGPSKPFWVLVVVGGVLACYSLLVT VAFIIFWVKRKKQRSRRNDEELETRAHRVATEERGRKPHQIPASTPQNPATSQHPPPP PGHRSQAPSHRPPPPGHRVQHQPQKRPPAPSGTQVHQQKGPPLPRPRVQPKPPHGAA ENSLSPSSNRVKFSRSADAPAYKQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGG KPRRKNPQEGLYNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYD ALHMQALPPR (SEQ ID NO: 772).

109. The composition of any one of claims 1 to 107, wherein the one or more recombinant polynucleic acid molecules comprises a sequence encoding the amino acid sequence of MLLLVTSLLLCELPHPAFLLIPQVQLQQSGPGLVKPSQTLSLTCAISGDSVSSNSAAW NWIRQSPSRGLEWLGRTYYRSKWYNDYAVSVKSRITINPDTSKNQFSLQLNSVTPEDTAVYYCAREVTGDLEDAFDIWGQGTMVTVSSGGGGSDIQMTQSPSSLSASVGDRVT ITCRASQTIWSYLNWYQQRPGKAPNLLIYAASSLQSGVPSRFSGRGSGTDFTLTISSLQ AEDFATYYCQQSYSIPQTFGQGTKLEIKAAATTTPAPRPPTPAPTIASQPLSLRPEACR PAAGGAVHTRGLDFACDIYIWAPLAGTCGVLLLSLVITLYCKRGRKKLLYIFKQPFM RPVQTTQEEDGCSCRFPEEEEGGCELRVKFSRSADAPAYKQGQNQLYNELNLGRREE YDVLDKRRGRDPEMGGKPRRKNPQEGLYNELQKDKMAEAYSEIGMKGERRRGKG HDGLYQGLSTATKDTYDALHMQALPPRATNFSLLKQAGDVEENPGPMALPVTALLL PLALLLHAARPEVQLVQSGAEVKKPGESLKISCKASGYRFTNYWIAWVRQRPGKGL EWMGRIDPSDSYTHYSPSFQGHVTMSTDKSISTAYLQWSSLKASDTAMYYCARPGDI LTGWAMDVWGQGTLVTVSSAAASGGGGSGGGGSGGGGSALQSVLTQPPSVSAAPG QKVTISCSGSSSNIGNNYVSWYQQLPGTAPKLLIYDNNKRPSGIPDRFSGSKSGTSATL GITGLQAEDEADYYCQSYDSSLSGNYVFGTGTKVTVLIEVMYPPPYLDNEKSNGTIIH VKGKHLCPSPLFPGPSKPFWVLVVVGGVLACYSLLVTVAFIIFWVRSKRSRLLHSDY MNMTPRRPGPTRKHYQPYAPPRDFAAYRSRVKFSRSADAPAYKQGQNQLYNELNL GRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLYNELQKDKMAEAYSEIGMKGERR RGKGHDGLYQGLSTATKDTYDALHMQALPPREGRGSLLTCGDVEENPGPMLLLVTS LLLCELPHPAFLLIPDIVMTQSPDSLAVSLGERATINCKSSQSVLYSSNNKNYLAWYQ QKPGQPPKLLIYWASTRESGVPDRFSGSGSGSDFTLTISSLQAEDVAVYYCQQYYSFY QTFGQGTKVEIKGGGGSGGGGSGGGGSEVQLVESGGGVVRPGGSLRLSCTASGFTF GDYGMSWVRQAPGKGLEWVSGINWNGGSTGYADSVKGRFTISRDNAKNSLYLQM NSLRAEDTALYYCARKSYYGSGSPDVFDIWGQGTMVTVSSIEVMYPPPYLDNEKSN GTIIHVKGKHLCPSPLFPGPSKPFWVLVVVGGVLACYSLLVTVAFIIFWVKRKKQRSR RNDEELETRAHRVATEERGRKPHQIPASTPQNPATSQHPPPPPGHRSQAPSHRPPPPG HRVQHQPQKRPPAPSGTQVHQQKGPPLPRPRVQPKPPHGAAENSLSPSSNRVKFSRS ADAPAYKQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLYNEL QKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO: 1006).

10. The composition of any one of claims 1 to 107, wherein the one or more recombinant polynucleic acid molecules has the sequence atgcttctgctcgtgacaagcctgctgctgtgcgagctgccccaccctgcctttctgctgatccctCAAGTCCAGCTGCAG CAATCTGGCCCTGGCCTGGTGAAGCCTAGCCAGACCCTGAGCCTGACATGTGCCA TCAGCGGAGATTCTGTTAGCAGCAACAGCGCGGCCTGGAACTGGATCAGACAGAGCCCTAGCAGAGGACTGGAGTGGCTGGGCCGGACCTACTACAGAAGCAAGTGGTACAACGACTACGCCGTGTCCGTGAAAAGCAGAATCACCATCAACCCCGACACCAGCAAGAACCAGTTCTCCCTGCAACTGAATAGCGTCACACCCGAGGATACAGCCGTGTACTACTGCGCCAGAGAGGTTACAGGCGACCTGGAAGATGCCTTTGACATCTGGGGCCAGGGCACGATGGTGACCGTCAGTAGCGGCGGCGGAGGAAGCGACATCCAGATGACCCAATCTCCTAGCAGCCTGTCTGCCTCTGTGGGCGACAGAGTGACCATCACCTGCCGCGCCTCTCAAACAATCTGGTCCTACCTGAACTGGTATCAGCAAAGACCTGGCAAAGCCCCAAACCTGCTGATCTACGCCGCCAGCTCCCTGCAAAGCGGAGTGCCCAGCAGATTCAGCGGCCGGGGCAGCGGCACTGATTTTACCCTGACTATTTCTTCCCTGCAAGCCGAGGACTTCGCCACATACTACTGTCAGCAGAGCTACAGCATCCCTCAGACCTTCGGCCAGGGCACCAAGCTGGAAATCAAGGCGGCCGCCACAACCACCCCTGCTCCCCGCCCACCTACCCCTGCTCCTACAATCGCCTCGCAACCTCTGAGCCTGAGACCTGAGGCCTGTAGACCCGCCGCTGGCGGCGCCGTGCACACCAGAGGCCTGGACTTCGCCTGCGACATCTACATCTGGGCCCCACTGGCCGGCACATGCGGCGTGCTGCTGCTGTCCCTGGTGATCACCCTGTATTGCAAGAGAGGCAGAAAGAAACTGCTGTACATCTTCAAGCAGCCTTTCATGCGGCCCGTGCAGACCACCCAGGAGGAAGATGGTTGCTCATGCCGGTTCCCCGAGGAAGAGGAAGGCGGATGTGAACTGAGAGTGAAGTTCAGCCGCTCCGCTGACGCACCCGCCTATAAACAGGGGCAGAATCAACTTTATAACGAACTGAACCTTGGAAGAAGGGAGGAATATGACGTGCTTGATAAGAGAAGAGGCAGAGATCCTGAAATGGGAGGGAAACCTCGGAGAAAAAATCCACAAGAGGGGCTCTACAATGAACTCCAGAAAGACAAGATGGCTGAAGCTTACTCTGAGATTGGAATGAAAGGGGAGCGGCGCCGGGGCAAAGGGCACGACGGATTGTACCAAGGACTCAGTACAGCTACTAAAGACACATATGACGCCCTGCATATGCAGGCACTGCCCCCACGGGCCACCAACTTCAGCCTGCTGAAGCAGGCCGGCGACGTGGAAGAGAACCCCGGCCCTATGGCCTTGCCCGTGACTGCTCTGCTCCTGCCCCTCGCACTGCTGCTGCACGCCGCGAGGCCCGAAGTGCAGCTGGTCCAATCTGGCGCCGAGGTGAAGAAGCCCGGCGAGAGCCTGAAGATCAGCTGTAAAGCCAGCGGATATAGATTCACCAACTACTGGATCGCCTGGGTGCGGCAGCGGCCTGGCAAGGGCCTGGAATGGATGGGCAGAATCGACCCATCTGATAGCTACACCCACTACAGCCCTAGCTTCCAAGGCCACGTGACCATGAGCACAGATAAGAGCATCAGCACCGCCTACCTGCAGTGGTCCTCCCTGAAAGCTAGCGACACCGCTATGTACTACTGCGCCAGACCTGGCGACATCCTGACCGGCTGGGCTATGGATGTGTGGGGCCAGGGAACACTGGTGACAGTGTCCAGCGCTGCAGCAAGTGGAGGCGGAGGAAGTGGAGGCGGTGGTTCAGGAGGAGGAGGATCTGCTCTGCAATCTGTGCTCACACAGCCCCCCAGCGTGTCTGCCGCTCCTGGACAGAAAGTGACCATCAGCTGCAGCGGATCTAGCTCCAACATCGGCAACAACTACGTGTCCTGGTATCAGCAGCTGCCTGGCACCGCCCCTAAGCTGCTGATCTACGACAACAACAAGCGGCCATCTGGCATCCCTGATCGCTTCAGCGGCAGCAAGTCCGGCACCAGCGCCACCCTGGGAATCACCGGCCTGCAGGCCGAGGACGAAGCCGACTACTACTGTCAGAGCTACGATAGCAGCCTGAGCGGCAATTACGTGTTCGGCACAGGCACAAAGGTCACCGTGCTGATCGAGGTGATGTACCCTCCTCCTTACCTGGATAATGAGAAGTCCAACGGAACAATCATCCACGTGAAGGGCAAGCACCTGTGCCCCTCTCCTCTGTTTCCGGGCCCTTCTAAGCCCTTCTGGGTGCTGGTCGTGGTGGGCGGAGTGCTCGCTTGTTACTCTCTGCTTGTGACCGTGGCCTTCATTATCTTCTGGGTCCGCAGCAAGCGGTCCAGACTGCTGCACAGCGACTACATGAACATGACCCCTCGGAGACCTGGACCCACCAGAAAGCACTACCAGCCTTACGCCCCTCCACGCGACTTCGCCGCTTATCGCAGCCGCGTTAAATTTTCACGCAGCGCGGACGCCCCTGCGTACAAGCAGGGACAGAACCAGCTGTACAACGAGCTCAACCTGGGCCGCCGGGAGGAGTACGACGTGCTGGACAAGCGCCGGGGGCGTGATCCAGAGATGGGCGGCAAACCCCGCCGCAAGAATCCTCAGGAGGGCTTATACAACGAGCTGCAGAAGGACAAAATGGCCGAGGCCTACAGCGAGATCGGCATGAAGGGAGAGCGCCGGCGTGGAAAGGGCCATGACGGCCTCTATCAGGGTCTGTCCACCGCCACCAAGGACACCTACGACGCTCTACACATGCAGGCCTTGCCTCCGCGCGAGGGCAGAGGCAGCCTGCTGACCTGCGGCGATGTGGAGGAGAACCCCGGACCTATGCTGTTGCTTGTTACAAGCCTCCTCCTGTGCGAATTGCCTCACCCCGCATTTCTCCTGATACCCGACATCGTGATGACCCAGTCCCCAGACAGCCTGGCTGTGTCCCTGGGCGAACGGGCCACCATCAACTGCAAGAGCAGCCAGAGCGTGCTGTACAGCAGCAACAACAAGAATTACCTGGCCTGGTACCAGCAGAAACCTGGCCAGCCTCCCAAGCTGCTGATCTACTGGGCCAGCACCAGAGAGAGCGGAGTGCCTGATAGATTCAGCGGCAGCGGATCTGGCTCTGATTTTACCCTGACAATCAGCAGCCTCCAGGCCGAGGACGTCGCCGTGTACTATTGTCAGCAATACTACTCCTTCTACCAAACATTCGGCCAGGGCACCAAAGTTGAAATCAAGGGAGGAGGAGGGTCTGGAGGTGGAGGAAGTGGCGGAGGGGGAAGTGAGGTGCAGCTGGTCGAGAGCGGCGGAGGCGTGGTGCGGCCTGGAGGATCTCTCAGACTGAGCTGTACCGCCAGCGGATTTACCTTCGGCGACTACGGCATGAGCTGGGTGCGCCAGGCCCCTGGCAAAGGCCTGGAATGGGTTTCCGGCATCAACTGGAACGGCGGCTCTACAGGCTACGCCGACAGCGTGAAGGGCAGATTCACCATCAGCCGGGACAACGCCAAGAATAGCCTGTACCTGCAAATGAACAGCCTGAGAGCTGAAGATACAGCTCTGTATTACTGCGCCAGAAAGTCCTACTACGGCTCTGGCAGCCCCGATGTGTTCGA CATCTGGGGCCAGGGCACAATGGTGACCGTGTCCAGCATAGAAGTAATGTACCC TCCGCCTTATCTCGATAATGAAAAAAGCAACGGGACAATTATTCATGTGAAGGG CAAGCATCTTTGTCCCTCACCGCTTTTCCCTGGACCATCAAAACCATTTTGGGTGC TCGTCGTCGTGGGAGGGGTCCTCGCCTGCTATTCTCTGTTGGTGACTGTAGCCTTT ATCATTTTTTGGGTTAAACGGAAGAAGCAGAGATCTAGACGGAACGACGAGGAA CTGGAAACCCGCGCtCACAGGGTGGCCACCGAGGAAAGAGGCAGAAAGCCACAC CAGATCCCCGCCTCAACACCTCAGAATCCCGCCACAAGCCAGCACCCCCCACCTC CTCCTGGCCATCGCAGCCAGGCCCCTAGCCACCGGCCTCCACCTCCCGGTCACAG AGTGCAGCACCAACCTCAAAAACGGCCCCCCGCTCCAAGCGGCACCCAAGTTCA CCAGCAGAAGGGCCCTCCTCTGCCTAGACCTAGAGTCCAGCCTAAGCCTCCTCAC GGCGCTGCTGAGAACAGCCTGAGCCCTTCTTCgAATCGGGTCAAATTCAGTCGCT CTGCCGATGCTCCAGCCTACAAACAAGGCCAAAACCAATTATACAATGAACTTA ATCTCGGGCGCAGGGAAGAGTACGATGTTCTCGATAAAAGGCGGGGACGCGACC CCGAAATGGGTGGAAAGCCAAGAAGGAAGAACCCGCAGGAAGGACTGTATAAT GAGTTACAAAAGGATAAGATGGCAGAGGCATATTCAGAAATCGGGATGAAGGG CGAAAGAAGAAGAGGCAAGGGACACGATGGGCTTTATCAAGGCTTAAGCACTGC AACAAAGGATACTTATGATGCACTCCATATGCAAGCTCTCCCACCTAGA (SEQ ID NO: 1118).

111. A composition comprising one or more polypeptides encoded by the one or more recombinant polynucleic acid molecules of the composition of any one of claims 1 to 108.

112. A cell comprising the one or more recombinant polynucleic acid molecules of the composition of any one of claims 1 to 108 or the one or more polypeptides of the composition of claim 111.

113. The cell of claim 112, wherein the cell is an immune cell.

114. The cell of claim 113, wherein the immune cell is a T cell, a B cell, a natural killer (NK) cell, a macrophage, a dendritic cell, a monocyte, or a granulocyte.

115. The cell of claim 114, wherein the cell is a T cell.

116. The cell of claim 115, wherein the T cell is a CD8-positive T cell, a CD4-positive T cell, a regulatory T cell (Treg), a cytotoxic T cell (CTL), a central memory T cell (TCM), an effector memory T cell (TEM), a tissue-resident memory T cell (TRM), a stem cell-like memory T cell (TSCM) or a tumor infiltrating lymphocyte (TIL).

117. A population of cells comprising the one or more recombinant polynucleic acid molecules of the composition of any one of claims 1 to 108 or the one or more polypeptides the composition of claim 111.

118. A pharmaceutical composition comprising the one or more recombinant polynucleic acid molecules of the composition of any one of claims 1 to 108.

119. A pharmaceutical composition comprising the one or more polypeptides of claim 111.

120. A pharmaceutical composition comprising the cell of any one of claims 112 to 116.

121. A pharmaceutical composition comprising the population of cells of claim 117.

122. A lentiviral expression vector comprising the one or more recombinant polynucleic acid molecules of the composition of any one of claims 1 to 108.

123. The lentiviral expression vector of claim 122, wherein the lentiviral expression vector is the pCCL-c-MNDU2-X vector.

124. The lentiviral expression vector of claim 123, wherein the pCCL-cMNDU2-X vector further comprises a woodchuck hepatitis virus posttranscriptional regulatory element (WPRE).

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