A Path Towards Determining Tumor Mutation Burden And Identifying Neoantigens Using Next-Generation Sequencing (Ngs)

Abstract Introduction: The recent clinical demonstrations that cancer immunotherapy is effective for some patients, but not others has led to widespread interest in identifying cancer genetic factors that can predict positive response. Importantly, it has been shown by various studies that tumor mutation burden (TMB) and predicted neoantigen load correlate with patient response to checkpoint inhibitors. This project aims to develop a clinical next-generation sequencing (NGS) assay that utilizes TMB and neoantigen information to guide cancer immunotherapy selection. Experimental Procedures: Whole exome sequencing (WES) and whole transcriptome sequencing (WTS) libraries were generated using Illumina’s TruSeq® Exome and RNA Access library preparation kits, respectively. The samples were pair-end sequenced (2x75bp) using HiSeq® 2000 and 2500 instruments. Data Summary: Here, we demonstrate a workflow using tumor/normal WES and tumor-only WTS to determine expressed TMB. Using the WES data, we were also able to accurately identify human leukocyte antigen (HLA) major histocompatibility complex (MHC) Class I genes at four-digit resolution. We then identified putative neoantigens, defined as peptides with mutated amino acids, that were expressed in the tumor samples and predicted to bind to the respective HLA sequences. In addition, as a path towards developing a clinical assay for TMB and neoantigen determination, we tested the ability of reagents developed for Illumina’s Trusight® Tumor 170 panel to be used for an exome panel. With workflow modifications, we achieved comparable quality of sequencing metrics as compared to the TruSeq® Exome kit. Conclusion: Our data demonstrate a path towards developing a clinical assay that can be used to assess TMB and neoantigen candidates. Citation Format: Alex So, Shannon Kaplan, Nai-yu Wang, Shile Zhang, Aaron Wise, Kristina Kruglyak, Karen Gutekunst. A path towards determining tumor mutation burden and identifying neoantigens using next-generation sequencing (NGS) [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 427. doi:10.1158/1538-7445.AM2017-427