Project description:Chimeric antigen receptor (CAR)-T cell therapy has been clinically successful in hematologic malignancies but faces challenges in solid tumors, including limited infiltration, immunosuppressive microenvironments and antigen heterogeneity. While combining CAR-T cell therapy with chemotherapy can enhance antitumor activity, this often leads to substantial systemic toxicity. Here we show that CAR-T-drug conjugate (CAR-T-D-C), generated through click chemistry-mediated conjugation of cytotoxic payloads to antigen-specific CAR-T cells, enable localized drug delivery while preserving CAR-T cell function. CAR-T-D-Cs incorporating different CAR-T cell binders exhibit robust antitumor activity across multiple human xenografts and syngeneic tumor models. Spatial transcriptomic analyses reveal enhanced intratumoral CAR-T infiltration and activation following CAR-T-D-C treatment . Compared with conventional CAR-T therapy, CAR-T-D-C enhances immune cell infiltration, augments effector functions, promotes antigen spreading and amplifies systemic anti-tumor immunity. CAR-T-D-C represents a versatile therapeutic platform that combines the specificity of cellular immunotherapy with the potency of small-molecule therapeutics for treatment of solid tumors.
Project description:Chimeric antigen receptor (CAR)-natural killer (NK) cell therapies hold promise for solid tumors but remain limited by poor tumor infiltration, persistence, and resistance within the tumor microenvironment (TME). To identify gain-of-function (GOF) targets that enhance CAR-NK efficacy, we performed an unbiased in vivo Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) activation (CRISPRa) screen, followed by a barcoded targeted in vivo open reading frame (ORF) screen in primary human CAR-NK cells. We identified, and robustly validated OR7A10, a G protein-coupled receptor (GPCR), as the top candidate. Engineering CAR-NKs with OR7A10 cDNA, a CRISPR-independent method with simple manufacturing strategy, enhanced proliferation, activation, degranulation, cytokine production, death ligand expression, chemokine receptor expression, cytotoxicity, persistence, metabolic fitness, and TME resistance, while reducing exhaustion in primary human NK cells derived from multiple peripheral blood and cord blood donors. OR7A10-GOF CAR-NKs displayed robust in vivo efficacy across multiple solid tumor models, achieving a 100% complete response in an orthotopic breast cancer model with long term tumor control and survival benefit. These findings establish OR7A10-engineered CAR-NKs as a highly potent and scalable off-the-shelf therapeutic for solid tumors.
Project description:Chimeric antigen receptor (CAR)-T cell therapy has been clinically successful in hematologic malignancies but faces challenges in solid tumors, including limited infiltration, immunosuppressive microenvironments and antigen heterogeneity. While combining CAR-T cell therapy with chemotherapy can enhance antitumor activity, this often leads to substantial systemic toxicity. Here we show that CAR-T-drug conjugate (CAR-T-D-C), generated through click chemistry-mediated conjugation of cytotoxic payloads to antigen-specific CAR-T cells, enable localized drug delivery while preserving CAR-T cell function. CAR-T-D-Cs incorporating different CAR-T cell binders exhibit robust antitumor activity across multiple human xenografts and syngeneic tumor models. Spatial transcriptomic analyses reveal enhanced intratumoral CAR-T infiltration and activation following CAR-T-D-C treatment . Compared with conventional CAR-T therapy, CAR-T-D-C enhances immune cell infiltration, augments effector functions, promotes antigen spreading and amplifies systemic anti-tumor immunity. CAR-T-D-C represents a versatile therapeutic platform that combines the specificity of cellular immunotherapy with the potency of small-molecule therapeutics for treatment of solid tumors.
Project description:Chimeric antigen receptor (CAR) and T-cell receptor (TCR) T-cell therapies are effective in a subset of patients with solid tumors, but new approaches are needed to enhance efficacy and universally improve patient outcomes. IL-15 and IL-21 are common cytokine-receptor gamma chain family members with distinct, pleiotropic effects on T-cells and other lymphocytes. We found that self-delivery of these cytokines by CAR or TCR T-cells prevents functional exhaustion by repeated stimulation and limits the emergence of dysfunctional natural killer (NK)-like T-cells. Across different preclinical murine solid tumor models, we observe enhanced regression with each individual cytokine but the greatest anti-tumor efficacy when T-cells are armored with both. Thus, the co-expression of membrane-tethered IL-15 and IL-21 represents a technology to enhance the resilience and function of engineered T-cells against solid tumors and could be applicable to multiple therapy platforms and diseases.
Project description:Chimeric antigen receptor (CAR)-T cell therapies have shown great success in treating hematologic malignancies. Nonetheless, their therapeutic effect on solid tumors remains to be improved. Recently, macrophages have attracted great attention, given their ability to infiltrate solid tumors, phagocytize tumor cells as well as their immunomodulatory capacities. The first generation of CD3ζ-based CAR-macrophages demonstrated that the CAR could stimulate macrophage phagocytosis in a tumor antigen-dependent way. Here, we genetically engineered induced pluripotent stem cell (iPSC)-derived macrophages (iMACs) with TLR4 intracellular TIR domain-containing CARs against EGFRvIII and GPC3, which yielded markedly enhanced antitumor effect in two different solid tumor models including glioblastoma, and hepatocellular carcinoma in which complete remission was achieved with CAR-iMACs alone or in combination with CD47 antibody. Moreover, the tandem CD3ζ-TIR-CAR, or the “second-generation” design of TIR-based dual signaling CAR, endowed iMACs with both target engulfment/efferocytosis capacity against antigen-expressing solid tumor cells, and potency of antigen-dependent M1 state polarization and M2 state resistance in an NF-κB dependent manner. We also illustrated a surprising mechanism of tumor cell elimination by CAR-induced efferocytosis against tumor cell apoptotic bodies. Taken together, we established the next generation CAR-iMACs equipped with orthogonal phagocytosis and polarization capacity for better antitumor functions in treating solid tumors.
Project description:Chimeric antigen receptor (CAR)-natural killer (NK) cell therapies hold promise for solid tumors but remain limited by poor tumor infiltration, persistence, and resistance within the tumor microenvironment (TME). To identify gain-of-function (GOF) targets that enhance CAR-NK efficacy, we performed an unbiased in vivo Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) activation (CRISPRa) screen, followed by a barcoded targeted in vivo open reading frame (ORF) screen in primary human CAR-NK cells. We identified, and robustly validated OR7A10, a G protein-coupled receptor (GPCR), as the top candidate. Engineering CAR-NKs with OR7A10 cDNA, a CRISPR-independent method with simple manufacturing strategy, enhanced proliferation, activation, degranulation, cytokine production, death ligand expression, chemokine receptor expression, cytotoxicity, persistence, metabolic fitness, and TME resistance, while reducing exhaustion in primary human NK cells derived from multiple peripheral blood and cord blood donors. OR7A10-GOF CAR-NKs displayed robust in vivo efficacy across multiple solid tumor models, achieving a 100% complete response in an orthotopic breast cancer model with long term tumor control and survival benefit. These findings establish OR7A10-engineered CAR-NKs as a highly potent and scalable off-the-shelf therapeutic for solid tumors.
Project description:Chimeric antigen receptor (CAR)-natural killer (NK) cell therapies hold promise for solid tumors but remain limited by poor tumor infiltration, persistence, and resistance within the tumor microenvironment (TME). To identify gain-of-function (GOF) targets that enhance CAR-NK efficacy, we performed an unbiased in vivo Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) activation (CRISPRa) screen, followed by a barcoded targeted in vivo open reading frame (ORF) screen in primary human CAR-NK cells. We identified, and robustly validated OR7A10, a G protein-coupled receptor (GPCR), as the top candidate. Engineering CAR-NKs with OR7A10 cDNA, a CRISPR-independent method with simple manufacturing strategy, enhanced proliferation, activation, degranulation, cytokine production, death ligand expression, chemokine receptor expression, cytotoxicity, persistence, metabolic fitness, and TME resistance, while reducing exhaustion in primary human NK cells derived from multiple peripheral blood and cord blood donors. OR7A10-GOF CAR-NKs displayed robust in vivo efficacy across multiple solid tumor models, achieving a 100% complete response in an orthotopic breast cancer model with long term tumor control and survival benefit. These findings establish OR7A10-engineered CAR-NKs as a highly potent and scalable off-the-shelf therapeutic for solid tumors.
Project description:Chimeric antigen receptor (CAR)-natural killer (NK) cell therapies hold promise for solid tumors but remain limited by poor tumor infiltration, persistence, and resistance within the tumor microenvironment (TME). To identify gain-of-function (GOF) targets that enhance CAR-NK efficacy, we performed an unbiased in vivo Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) activation (CRISPRa) screen, followed by a barcoded targeted in vivo open reading frame (ORF) screen in primary human CAR-NK cells. We identified, and robustly validated OR7A10, a G protein-coupled receptor (GPCR), as the top candidate. Engineering CAR-NKs with OR7A10 cDNA, a CRISPR-independent method with simple manufacturing strategy, enhanced proliferation, activation, degranulation, cytokine production, death ligand expression, chemokine receptor expression, cytotoxicity, persistence, metabolic fitness, and TME resistance, while reducing exhaustion in primary human NK cells derived from multiple peripheral blood and cord blood donors. OR7A10-GOF CAR-NKs displayed robust in vivo efficacy across multiple solid tumor models, achieving a 100% complete response in an orthotopic breast cancer model with long term tumor control and survival benefit. These findings establish OR7A10-engineered CAR-NKs as a highly potent and scalable off-the-shelf therapeutic for solid tumors.