Methods and compositions for modulating CAR-T activity
Inventors
Schrepfer, Sonja • Weng, Lindong • Fry, Terry J.
Assignees
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Publication Number
US-11802157-B2
Publication Date
2023-10-31
Expiration Date
Abstract
Disclosed herein are engineered cells and/or hypoimmunogenic cells including engineered and/or hypoimmunogenic stem cells, engineered and/or hypoimmunogenic cells differentiated therefrom, engineered and/or hypoimmunogenic CAR-T cells (primary or differentiated from engineered and/or hypoimmunogenic stem cells) and related methods of their use and generation. Provided herein are engineered and/or hypoimmunogenic cells exhibiting reduced expression of MHC class I and/or MHC class II human leukocyte antigens and T-cell receptors. In some embodiments, such cells also exogenously express one or more tolerogenic factors such as CD47 and one or more chimeric antigen receptors (CAR)s.
Core Innovation
The invention relates to engineered T cells derived from a primary T cell, where immune-activation related features are reduced and immune-evasion features are increased while retaining antigen targeting capability. The engineered T cell reduces expression of B2M, CIITA, and/or T cell receptor (TCR)-alpha relative to a control T cell, and increases expression of CD47 encoded by a first exogenous polynucleotide relative to the control T cell. The engineered T cell also expresses a CD19-specific chimeric antigen receptor (CAR) encoded by a second exogenous polynucleotide, with insertion into at least one allele.
The engineered CAR is defined by an antigen binding domain that specifies light chain and heavy chain complementarity determining regions. The light chain includes CDR1, CDR2, and CDR3 sequences set forth in SEQ ID NO: 21, SEQ ID NO: 22, and SEQ ID NO: 23, respectively. The heavy chain includes CDR1, CDR2, and CDR3 sequences set forth in SEQ ID NO: 26, SEQ ID NO: 27, and SEQ ID NO: 28, respectively.
The provided content also describes hypoimmunogenic primary T cells engineered with reduced endogenous TCR/MHC components and increased CD47 and CAR expression, including genomic modifications in B2M/CIITA/TRAC/TRB loci and lentiviral vector-based polynucleotide insertion. The content further describes assessment and assay contexts, including decreased or absent immune activation, reduced cytotoxicity and reduced complement-dependent cytotoxicity (CDC), and therapeutic use contexts including cancer indications.
Claims Coverage
The consolidated claim coverage includes one independent claim family with three core inventive features: reduced expression of B2M, CIITA, and/or TCR-alpha, increased CD47 expression via a first exogenous polynucleotide, and CD19-specific CAR expression via a second exogenous polynucleotide. The CAR antigen binding domain is defined by specific light-chain and heavy-chain CDR sequences, and both exogenous polynucleotides are inserted into at least one allele.
Reduced B2M, CIITA, and/or TCR-alpha expression in a primary-derived engineered T cell
The engineered T cell is derived from a primary T cell and comprises reduced expression of B2M, CIITA, and/or T cell receptor (TCR)-alpha relative to a control T cell.
Increased CD47 expression encoded by a first exogenous polynucleotide
The engineered T cell comprises increased expression of CD47 encoded by a first exogenous polynucleotide relative to the control T cell, wherein the first exogenous polynucleotide is inserted into a locus of at least one allele of the engineered T cell.
CD19-specific CAR expression encoded by a second exogenous polynucleotide with defined CDRs
The engineered T cell expresses a CD19-specific chimeric antigen receptor (CAR) encoded by a second exogenous polynucleotide, wherein the second exogenous polynucleotide is inserted into a locus of at least one allele of the engineered T cell and the CAR comprises an antigen binding domain defined by light-chain CDR1/CDR2/CDR3 sequences (SEQ ID NO: 21/22/23) and heavy-chain CDR1/CDR2/CDR3 sequences (SEQ ID NO: 26/27/28).
Genomic insertion of both exogenous polynucleotides into at least one allele
The first exogenous polynucleotide is inserted into a locus of at least one allele of the engineered T cell, and the second exogenous polynucleotide is inserted into a locus of at least one allele of the engineered T cell.
The inventive concept is an engineered, primary-derived T cell that combines reduced B2M/CIITA/TCR-alpha expression with increased CD47 expression and CD19-specific CAR expression, where the CAR antigen binding domain is defined by specified CDR sequences and both exogenous polynucleotides are inserted into at least one allele.
Stated Advantages
decreased or absent immune activation including T cell, NK cell, macrophage engulfment, TH1 activation, and PBMC activation
reduced cytotoxicity and reduced complement-dependent cytotoxicity (CDC)
reduced donor-specific antibodies, including reduced IgG and reduced IgM
Reduced immune-recognition and functional responses associated with B2M/CIITA/TCR-alpha and CD47 modulation.
Reduced T/NK/macrophage responses, reduced TH1 activation, and reduced NK killing.
Immune evasion, including CDC/complement killing/evasion.
Preserved cytotoxicity.
Lack of IL-2-independent proliferation or GvHR in mouse models.
Documented Applications
therapeutic cancers, including B-ALL, lymphomas, lung and other solid tumors
Therapeutic use of engineered hypoimmunogenic CAR-T and related engineered/hypoimmunogenic cells, including pharmaceutical compositions and cryopreserved formulations for dosing regimens.
Tumor-killing efficacy described in relation to in vitro and in vivo contexts [procedural detail omitted for safety].
In vitro and in vivo immune-evasion and functional assessment of hypoimmunogenic CD19-CAR-T cells, including complement/CDC killing assays and mouse xenograft models.
Treatment framework for administering CD19-specific CAR-T cells, including optional immunosuppressive/immunomodulatory agents with timing relative to lymphodepletion agents and potential co-administration with agents interacting with NK receptors.
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