Automated Trb Locus Haplotype Analysis By Long-Amplicon Tcrb Chain Sequencing For Immune-Related Adverse Events Biomarker Research

Introduction: Identifying predictive biomarkers for immune related adverse events (irAEs) following checkpoint blockade inhibition (CPI) remains an outstanding goal of immune-oncology translational research. Polymorphism within the T cell receptor variable gene (TRBV) has been proposed as a potential risk factor for irAEs owing to the potential link between TRBV polymorphism and chronic autoimmune disease. Efforts to interrogate the potential biomarker utility of TRBV polymorphism have been hampered by the repetitive nature of the TRB locus. We previously demonstrated a method for inferring TRB locus haplotypes from long amplicon TCRB chain sequencing data, which we used to identify major haplotype groups in a set of 81 Caucasians. Here we present an automated method for haplotype group assignment from TCRB chain sequencing data. Methods: Rearranged TCRB chains from 10 blood samples were amplified and sequenced from 25ng peripheral blood total RNA via the Oncomine™ TCRB-LR assay using the Genexus™ Integrated Sequencer. 12 samples were run per chip with 4 samples run in each lane. TCRB clonotyping and repertoire feature analysis was performed using the Genexus™ software v5.107.14. Automated haplotype group assignment was performed by generation and comparison of TRBV allele profiles to those presented in DOI: 10.4103/JIPO.JIPO_16_19. For context, TCR evenness, convergence, and haplotype group assignment were compared to values obtained following analysis of the same samples via the GeneStudio™ S5 platform and Ion Reporter™ 5.12 software. Results: TCR Evenness and convergence values were highly correlated across replicates run on the Genexus™ Integrated Sequencer (Spearman correlation \u003e0.95 and \u003e.70, respectively). Evenness at equivalent clone count and convergence at equivalent sequencing depth were not significantly different across platforms (Spearman correlation \u003e0.88). Haplotype group assignments demonstrated 100% agreement across replicates on both platforms. Conclusions: We demonstrate an automated and reproducible research method for TRB haplotype group assignment via the Oncomine™ TCR-Beta LR Assay, GX run on the Genexus™ Integrated Sequencer. Future studies will evaluate the potential biomarker utility of TRB haplotyping for the prediction of irAEs. Citation Format: Jennifer Burke, Frances Chan, Jiajie Huang, Marina Sedova, Timothy Looney, Fiona Hyland. Automated TRB locus haplotype analysis by long-amplicon TCRB chain sequencing for immune-related adverse events biomarker research [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 4279.