Abstract 1722: Role of tumor infiltrating lymphocytes and spatial immune heterogeneity in sensitivity to PD-1 axis blockers in non-small cell lung cancer (NSCLC)
Authors: Gregorio Miguel Lopez de Rodas , Nagineni Venkata , Ravi Arvind , Datar Ila J. , Mino-Kenudson Mari , Corredor German , Barrera Cristian , Behlman Lindsey , Rimm David L. , Herbst Roy S. , Madabhushi Anant , Riess Jonathan W. , Velcheti Vamsidhar , Hellmann Matthew , Gainor Justin F. , Schalper Kurt A. 
Published: June 2022

Abstract
The rapid clinical expansion of immune checkpoint blockers (ICB) has transformed the therapeutic arsenal for non-small cell lung cancer (NSCLC). However, only a relatively small fraction of patients benefits from these treatments and those who respond can develop acquired resistance. This underlines the need to identify factors or biomarkers associated with treatment sensitivity and resistance to guide clinical decisions. It has been proposed that elevated levels of tumor infiltrating lymphocytes (TIL) in the tumor microenvironment could be associated with favorable clinical outcomes after treatment with ICB. However, few studies have explored this relationship and they used semi-quantitative scoring methods and/or analyzed small tumor areas. Here, we used multiplexed quantitative immunofluorescence (QIF) panels to study major TIL subsets (DAPI, CK, CD4, CD8 and CD20) and T-cell exhaustion markers (DAPI, CD3, LAG-3, PD-1, TIM-3) in full-face baseline whole-tumor slides from a large multi-institutional patient cohort (n=179). The analysis of numerous fields of view per case allowed us to examine the spatial distribution of immune cells and spatial immune heterogeneity. Our results demonstrate that CD8+ effector T-cells are the largest TIL subpopulation and that they are preferentially located in the stromal compartment. We also show that tertiary lymphoid structures have limited association with survival and their detection depends on the size of the tissue area analyzed. Additionally, higher levels of baseline CD8+ T-cells in the stroma were significantly associated with better survival in PD-L1 positive patients but not in PD-L1 negative. To validate these results, we used whole-exome sequencing to analyze the TCR-burden in an independent cohort of ICI-treated NSCLC patients. Increased expression of the exhaustion markers LAG-3 and TIM-3 in CD3 positive T-cells was associated with reduced survival. Finally, we used a novel multiparametric heterogeneity metric termed Rao’s quadratic entropy that enabled us to measure the spatial immune heterogeneity and was associated with worse survival. In summary, we quantitatively measured the density, functional properties, and spatial distribution of major TIL subsets in a large cohort of NSCLC patients treated with ICB. Our results highlight the prominent role of TILs in NSCLC and their potential role as a biomarker. These results could be easily translated into the clinic and used to guide optimal treatment options.
Citation Format: Miguel Lopez de Rodas Gregorio, Venkata Nagineni, Arvind Ravi, Ila J. Datar, Mari Mino-Kenudson, German Corredor, Cristian Barrera, Lindsey Behlman, David L. Rimm, Roy S. Herbst, Anant Madabhushi, Jonathan W. Riess, Vamsidhar Velcheti, Matthew Hellmann, Justin F. Gainor, Kurt A. Schalper. Role of tumor infiltrating lymphocytes and spatial immune heterogeneity in sensitivity to PD-1 axis blockers in non-small cell lung cancer (NSCLC) [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1722.