In multiple biopsy samples from primary tumors of 15 newly diagnosed NSCLC patients, integrated data from whole-exome sequencing, RNAseq, and TCR sequencing showed spatial heterogeneity in total mutational burden (TMB) and neoantigen load. Local TMB correlated with local CD8+ T cell clonal expansion, but not with cytolytic activity or PD-L1 expression within the TME. An “immune map” created with 278 input variables further demonstrated that neither local TMB nor neoantigen load predicted local immunogenicity. Spatially heterogeneous immune cell infiltration was observed, and it appeared to be related to certain mutations found at different tumor loci.

Combining whole exome sequencing, transcriptome profiling, and T cell repertoire analysis, we investigate the spatial features of surgically-removed biopsies from multiple loci in tumor masses of 15 patients with non-small cell lung cancer (NSCLC). This revealed that the immune microenvironment has high spatial heterogeneity such that intratumoral regional variation is as large as inter-personal variation. While the local total mutational burden (TMB) is associated with local T-cell clonal expansion, local anti-tumor cytotoxicity does not directly correlate with neoantigen abundance. Together, these findings caution against that immunological signatures can be predicted solely from TMB or microenvironmental analysis from a single locus biopsy.

Author Info: (1) Institute of Cancer, Xinqiao Hospital, Third Military Medical University, Chongqing, 400037, China. Chongqing Key Laboratory of Tumor Immunotherapy, Chongqing, 400037, China. (2) Department of

Author Info: (1) Institute of Cancer, Xinqiao Hospital, Third Military Medical University, Chongqing, 400037, China. Chongqing Key Laboratory of Tumor Immunotherapy, Chongqing, 400037, China. (2) Department of Cardiothorathic Surgery, Southwest Hospital, Third Military Medical University, Chongqing, 400038, China. (3) Geneplus-Beijing Institute, Beijing, 102206, China. (4) Department of Immunology, Duke University Medical Center, Durham, 27710, NC, USA. (5) Institute of Cancer, Xinqiao Hospital, Third Military Medical University, Chongqing, 400037, China. Chongqing Key Laboratory of Tumor Immunotherapy, Chongqing, 400037, China. (6) Institute of Cancer, Xinqiao Hospital, Third Military Medical University, Chongqing, 400037, China. Chongqing Key Laboratory of Tumor Immunotherapy, Chongqing, 400037, China. (7) Institute of Cancer, Xinqiao Hospital, Third Military Medical University, Chongqing, 400037, China. Chongqing Key Laboratory of Tumor Immunotherapy, Chongqing, 400037, China. Biomedical Analysis Center, Third Military Medical University, Chongqing, 400038, China. (8) Geneplus-Beijing Institute, Beijing, 102206, China. (9) Geneplus-Beijing Institute, Beijing, 102206, China. (10) Geneplus-Beijing Institute, Beijing, 102206, China. Houston Methodist Research Institute, Houston, 77030, TX, USA. (11) Geneplus-Beijing Institute, Beijing, 102206, China. (12) Geneplus-Beijing Institute, Beijing, 102206, China. (13) Department of Microbiology and Immunology, Lineberger Comprehensive Cancer Center, University of North Carolina at Chapel Hill, Chapel Hill, 27514, NC, USA. (14) Department of Cardiothorathic Surgery, Southwest Hospital, Third Military Medical University, Chongqing, 400038, China. (15) GeneCast Biotechnology Co., Ltd, Beijing, 102206, China. (16) Department of Genomic Medicine, The University of Texas MD Anderson Cancer Center, Houston, TX, USA. (17) Department of Immunology, Duke University Medical Center, Durham, 27710, NC, USA. Qi-Jing.Li@Duke.edu. Biomedical Analysis Center, Third Military Medical University, Chongqing, 400038, China. Qi-Jing.Li@Duke.edu. (18) Institute of Cancer, Xinqiao Hospital, Third Military Medical University, Chongqing, 400037, China. bo.zhu@tmmu.edu.cn. Chongqing Key Laboratory of Tumor Immunotherapy, Chongqing, 400037, China. bo.zhu@tmmu.edu.cn.

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