Use of vaccines to boost CAR T efficacy and promote antigen spreading for long-term anti-tumor activity
286. Vaccine-boosted CAR T crosstalk with host immunity to reject tumors with antigen heterogeneity
Leyuan Ma (Children’s Hospital of Philadelphia, Philadelphia, PA, USA) reported results from a study of a synthetic vaccine (amph-vax) to enhance CAR T cell activity against solid tumors. Amph-vax consisted of a CAR-ligand linked to albumin-binding poly (ethylene glycol)-phospholipid and formulated with a cyclic-di-GMP (STING agonist). Fluorescently labeled amph-vax targeted lymph nodes and associated with antigen presenting cells in the draining lymph node. Amph-vax expressing tumor-specific epitope pepVIII (amph-pepVIII) was constructed and administered with EGFRvIII-targeting CAR T cells in the CT-2A mouse model of glioblastoma. Amph-pepVIII significantly increased CAR T expansion compared to mice receiving CAR T cells alone. Amph-pepVIII was also associated with significant increases in CAR T cell cytokine secretion, CAR T cell mitochondria mass, and CAR T cell cytotoxicity, indicating increased CAR T cell functionality. Amph-pepVIII was also associated with increased CAR T cell tumor infiltration and metabolic reprogramming signatures, including increased activity of oxidative phosphorylation pathways. Mice receiving amph-pepVIII and CAR T cells (CAR T-vax therapy) exhibited significant improvement in tumor control and survival. Immunological memory was also associated with amph-pepVIII: over 50% of mice previously cured with CAR T-vax therapy survived re-challenge with EGFRvIII+ and EGFRvIII - tumor cells, indicating activation of endogenous immunity through antigen spreading. Long-term tumor suppression was dependent on endogenous T cells, as Rag1 knockout mice, which do not have endogenous T cells, experienced relapse of tumors after CAR T-vax therapy. In a heterogenous tumor model with EGFRvIII+ and EGFRvIII- CT-2A cells mixed at pre-defined ratios, amph-pepVIII + CAR T cell therapy cured approximately 50% of mice bearing tumors with up to 20% EGFRvIII- cells. Antigen spreading induced by CAR T-vax therapy was also dependent on interferon gamma. The data support a model in which amph-vax activates antigen presenting cells in the lymph nodes, causing co-stimulation and cytokine support of CAR-T cells. Activated CAR T cells destroy tumor cells, and release of interferon gamma by CAR T cells promotes tumor antigen cross-presentation and T cell priming in tumor-draining lymph nodes. Results from this study provide new insights into the mechanisms underlying antigen spreading as well as mechanisms to improve CAR T cell therapy.
Immunotherapy approaches based in TCR-engineered T cell-based approaches for treatment of Merkel cell carcinoma that is refractory to immune checkpoint blockade
768. Transcriptional profiling of Merkel cell carcinoma that escaped polyomavirus-specific TCR-engineered T cells reveals actionable immunotherapy approaches.
Yuta Asano (Fred Hutchinson Cancer Center, Seattle, WA, USA) presented results from a small clinical study of engineered TCR-T cell (TCR-MCC1-T cell) therapy for Merkel cell carcinoma (MCC). Up to 50% of patients with advanced MCC fail to respond or relapse to immune checkpoint blockade (ICB). 80% of cases of MCC are driven by oncoproteins derived from Merkel cell polyomavirus (MCPyV), representing an ideal target for T cell immunotherapy. Five patients with ICB-refractory metastatic MCC received TCR-MCC1-T cell therapy with avelumab (anti-PD-L1). One patient responded to treatment, with 11 of 12 lesions responding by day 105 after treatment. All lesions from the responding patient showed evidence of T cell infiltration, with T cells expressing markers of activation and exhaustion. Although the HLA-A*02:01 gene was intact in all five patients, the non-responding patients had lost HLA expression at baseline, and the progressing lesion from the responding patient also exhibited low levels of HLA expression. HLA-high tumor cells from this lesion correlated with IFN gamma-downstream gene, suggesting IFN gamma can upregulate HLA in MCC. The trial was modified to include IFN gamma expression to upregulate HLA. Two patients received TCR-MCC1-T cell therapy with avelumab and IFN gamma, and one patient exhibited a response, marked by a rapid increase in tumor growth and then ongoing tumor regression. At 3 months, HLA expression was upregulated in the lesions of the responding patient. Single cell RNA sequencing indicated TCR T cells were present in the tumor only at early time points, which corresponding to a time period when the tumor was progressing, but TCR T cells were not detected during the time of tumor regression, suggesting endogenous immune cells were working to kill tumor. T cells and NK cells were abundant in the responding tumor during this later time point, and both cell types exhibited upregulated cytotoxicity-related genes. Results from this study suggest that engineered TCR-T cell therapy combined with ICB and interferon gamma is a promising approach for treatment of ICB-refractory MCC. Mechanisms of patient response to TCR-T cell therapy are currently being investigated.
Transcription factor FOXO1 regulates CAR T memory and is required for CAR T efficacy
247. FOX01 is a master regulator of CAR T memory programming
Alexander Doan (Stanford University, Palo Alto, CA, USA) presented a study investigating regulation of CAR T cell memory. CAR T cells with memory phenotype are associated with persistence and durable responses to therapy. TCF1 (encoded by the TCF7 gene) and FOXO1 were identified as memory-driving transcription factors in T cells. FOXO1 has been previously associated with central immunologic memory in mice, and TCF7 is a target gene of FOXO1. FOXO1 knockout CAR T cells exhibit an exhausted phenotype, decreased expression of genes associated with a naïve phenotype, and increased expression of genes associated with exhaustion. Transcriptomic profiles of CAR T cells overexpressing TCF1, FOXO1, or both TCF1 and FOXO1 were examined, and cells overexpressing FOXO1 exhibited a naïve T cell transcription signature and increased expression of genes associated with memory. Cells overexpressing TCF1 exhibited an exhausted phenotype. CD19 CAR T cells overexpressing FOXO1 exhibited increased leukemia control and prolonged survival in mouse models. FOXO1 CAR T cells were present at higher levels in the blood, and increased upon rechallenge with leukemia cells, enhancing the ability of mice to respond to rechallenge. FOXO1 overexpression enhanced solid tumor control even in a TCF7 knockout mouse. FOXO1 overexpression in HER2-targeting CAR T cells for solid tumors also led to better tumor control, increased tumor infiltrating lymphocytes, and increased cytokine secretion. Examination of datasets from patients receiving CAR T therapy for chronic lymphocytic leukemia indicated that the FOXO1 regulon, consisting of 41 putative FOX01 target genes, highly correlated with long-term persistence and positive survival outcomes in patients. Results from this study underscore the importance of memory-like phenotype for CAR T efficacy and persistence and provide a novel engineering approach to enhance CAR T anti-tumor activity.