Tumors form complex ecosystems that include normal epithelial cells, fibroblasts, blood and lymphatic vessels as well as structural components, and importantly, immune cells. The density, location, and organization of various tumor-infiltrating hematopoietic cells of myeloid and lymphoid origin can impact tumor growth, spreading, and clinical outcome of various cancers, including non-small cell lung cancer (NSCLC). Here, we propose to characterize the microenvironment of lung tumors using a unique set of matched fresh tissue single cell suspensions of tumor and normal lung, paraffin-embedded tissues, and peripheral blood plasma and cells obtained at time of surgery in the same patient. We aim to query mechanisms for local immunogenicity and immunosuppression of NSCLC, by defining interactions between tumor cells and various immune cell subsets in the local microenvironment, and by correlating these findings with presence of tumor-associated antigens (TAAs) and their capacity to elicit spontaneous local and systemic immune responses, as well as immune checkpoints known to impair immune responses. We applied multiplexed immunohistochemistry for formalin-fixed paraffin embedded cancer tissues (MICSSS method; n = 40), serological assays for immunogenicity of tumor antigens from plasma samples and supernatants of B cell cultures from the same patients (ELISA; n = 40), as well as mass cytometry on cell suspensions obtained from matched fresh specimens (CYTOF, n = 20) to (1) characterize the composition and organization of the immune microenvironment, (2) identify drivers of immunosuppression in NSCLC and (3) identify the expression profile of TAAs and B cell specificity in NSCLC. We found that TAAs, such as p53, NY-ESO-1, or ERG, were expressed by lung cancer cells in a subset of patients and that protein expression at the tumor site correlated with the antibody (Ab) response found in plasma of matched patients. While circulating plasma Ab titers did not appear to be correlated with the density of infiltrating B cells, potential associations with presence of tertiary-lymphoid structure, plasma cells (CD138+ cytokeratin− cells) or T follicular helper cells (CXCR5+ CD3+ CD8−) are currently being assessed. Association between molecular alterations and immune cell composition and location will also be discussed. We will also quantify expression of checkpoint molecules (PD-L1, VISTA and HHLA-2) on immune and tumor cells. The correlation between checkpoint inhibitor expression and immune cell densities and presence of immune responses will be presented, to assess whether immune cells contribute to the modulation of T cell effector functions. In conclusion, this study is the stepping-stone to a comprehensive understanding of the complex interactions between lung tumors and their immune microenvironment. By using novel multiplexed approaches, we intend to map the immune and antigenic landscape of human NSCLC, to identify mechanisms of early disease immunogenicity and immunosuppression, and to find new prognostic and predictive markers for the development of novel therapeutic strategies and immunotherapy trials in NSCLC.
Citation Format: Romain Remark, Yonit Lavin, Adeeb Rahman, Camille Bigenwald, Soma Kobayashi, Christian Becker, Raja Flores, Sacha Gnjatic, Miriam Merad. Comprehensive multidimensional analysis of the immune contexture in non-small cell lung cancer [abstract]. In: Proceedings of the Second CRI-CIMT-EATI-AACR International Cancer Immunotherapy Conference: Translating Science into Survival; 2016 Sept 25-28; New York, NY. Philadelphia (PA): AACR; Cancer Immunol Res 2016;4(11 Suppl):Abstract nr A092.
- ©2016 American Association for Cancer Research.