A study of tumor neoantigen burden and HLA-LOH by whole-exome sequencing to characterize immune biomarkers of lung cancer.

Abstract

8527 Background: Lung cancer is a leading cause of cancer-related death worldwide. Precision immunotherapy, such as the use of immune checkpoint inhibitors (ICIs), has been shown to improve the survival of lung cancer patients. Cancer cells with high amount of neoantigens are more likely to be captured by immune cells and thus patients with high Tumor Mutation Burden(TMB) and high Tumor Neoantigen Burden(TNB) are estimated to be prone to ICIs treatment. HLA class Ⅰ is one of the main factors affecting the prognosis of ICI in treatment. Studies have indicated that patients with highly heterozygous HLA-I are more likely to have better responses to ICI treatment. Some specific HLA-Ⅰ genotypes and somatic mutations can affect the prognosis of ICIs treatment. Methods: In this study, we performed whole-exome sequencing (WES) of 200 pairs of tumor and gDNA samples from lung cancer patients. Tumor sample comes from tumor tissue or paraffin-embedded tissue and gDNA is extracted from white blood cells from peripheral blood. We analyze the WES data to obtain the TMB and HLA types of each sample. Then the peptide affinity of MHC-I genes were quantified based on IC50(nM) by netMHCpan software, and the level of neoantigen affinity were classified as strong (IC50≤50nM), weak (50nM < IC50≤500nM), and negative (IC50 > 500nM). TNB is defined as the number of high- affinity neoantigens per megabase of interrogated genomic sequence. Then we conducted linear regression analysis to study the statistical relationship between TMB and TNB. Finally, we analyzed the LOH status at the major region of HLA (HLA-A, HLA-B, and HLA-C) and classified the individuals who have LOH at all three types of HLA as HLA-LOH strong positive, samples with one or two LOH among the three HLA types as weak positive, and samples have no LOH are classified as HLA-LOH negative. Results: Among the 200 samples, the median value of TMB and TNB is 3.25 Muts/Mb and 1.09 Neos/Mb, respectively. 8.5% of the sample have high TMB(TMB≥10). TMB and TNB are also significantly positively correlated (p-value < 0.0001). For HLA typing, 82.0% HLA-A, 96.0% HLA-B, and 92.5% HLA-C are heterozygous. The dominant HLA types are: (A*11:01), (A*02:01), (B*46:01), (B*40:01), (C*01:02), and (C*07:02). As for LOH status among the three HLA types, 14.57% of the individuals are HLA-LOH strong positive, 31.66% are HLA-LOH weak positive, and 53.77% of the samples are HLA-LOH negative. Lastly, the average numbers of detected high-affinity neoantigens are statistically significantly related to the occurrence of HLA-LOH (Analysis of Variance p-value < 0.01). Conclusions: HLA-LOH occurs in nearly half of our lung cancer samples, and it is associated with TMB-High and TNB-High. The analysis of HLA-LOH refines the prediction of neoantigen and gain understanding of drug resistance and immunotherapy.