Abstract 895: Tumor-wide neoantigen-specific T-cells infiltrating mutant IDH1 low-grade gliomas and persisting in peripheral blood allow for personalized TCR-based immunotherapies

BACKGROUND: The low mutational burden and immunologically “cold” microenvironment of mutant IDH1 low-grade gliomas (LGG) are considerable challenges facing immunotherapy against these tumor types. However, we hypothesize that LGG-targeting T-cells may exist at low frequency and with limited regional infiltration within the tumor. Multi-region tumor sampling coupled with high-throughput T-cell receptor (TCR) profiling across the LGG landscape detected neoantigen-specific T-cells that persisted in peripheral blood. TCR-engineered T-cells transduced with these TCRs demonstrated neoantigen-specific immunogenicity. METHODS: Maximal-anatomical sampling of at least 10 distinct tumor regions were collected at the initial resection for three WHO Grade II diffuse astrocytoma patients for exome-based prediction of clonally and subclonally expressed neoantigens, RNAseq analysis of regional immune cell composition, and TCR beta deep sequencing. We used these predictions to generate a barcoded library of patient-specific peptide-HLA multimers loaded with predicted neoepitopes. With this library, neoantigen-specific CD8+ T-cells were captured and isolated from patient peripheral blood. Single cell TCR sequencing allowed us to identify the neoantigen-reactive TCR clonotypes which were transduced subsequently into Jurkat76 cell lines for functional validation. RESULTS: We screened patient-derived peripheral blood drawn two years after initial resection in 3 mutant IDH1 LGG patients and detected a total of 20 TCR clonotypes recognizing neoepitopes derived from truncal, tumor-wide mutations in CNTNAP1 (n=8), TP53 (n=3), and MRPL46 (n=2) as well as subclonal mutations in PRMT5 (n=1) and ZDHHC5 (n=6). Multi-sampling RNAseq analysis indicated varying degrees of interpatient and intratumoral immune infiltration as well as distally located populations of neoantigen-reactive T-cells within the tumor, suggesting widespread migration of neoantigen-specific T-cells across the glioma landscape. We proceeded with TCR functional analysis for one patient (P375) with 5 detected TCR clonotypes recognizing neoantigens derived from mutations in PRMT5, MRPL46, and TP53. Jurkat76 cells transduced with the mutant-PRMT5-specific TCR demonstrated a neoantigen-specific immune response when co-cultured with mutant-PRMT5 pulsed-antigen presenting cells expressing HLA-A*0201 (T2 cells). CONCLUSION: Our study demonstrates the existence and persistence of neoantigen-targeting T-cells within the blood a