Deciphering the immunological determinants of response to neoadjuvant chemo-radiation in esophageal adenocarcinoma

Background The standard therapy for esophageal adenocarcinoma (EAC) is pre-operative neoadjuvant chemo-radiation (NACR) and surgery. Less than 30% of patients achieve a pathological complete response (pCR) and increased 5-year survival after NACR. Understanding the mechanisms of response to NACR is pivotal to better stratify patients and inform the design of more efficacious therapies. Cancer progression partly depends on defective immune surveillance, and the presence of tumor infiltrating T lymphocytes (TILs) is associated with better prognosis of several malignancies. We hypothesize that NACR induces immunogenic cell death that stimulates immune responses against tumor neoantigens (TNAs), derived from cancer somatic mutations, in the responders. We thus aim at comparing responders and non-responders for tumor immune contexture, immune molecular pathways, circulating immune markers and TNAs-specific T cells responses. Methods Prospective pre-NACR tumor biopsies from EAC patients were subjected to DNA exome sequencing (WES) and RNAseq, high dimensional (HD) multiparametric flow cytometry and histological immune contexture analysis, while their blood collected pre- and post- NACR was banked for immunological assays and in part analyzed by HD flow cytometry. Results In silico analysis revealed higher potential TNA load and enriched hallmark immune signatures in treatment-naïve EACs of NACR responders compared with non-responders. Immunohistochemical and HD flow cytometry analysis showed augmented effector CD8+ T cells infiltration and reduced CD25+CD127low T regulatory lymphocytes and M2-like pro-tumoral macrophages in pre-treatment EACs of responders. These data were corroborated by in silico CIBERSORT analysis on bulk RNAseq. In addition to reduced response to NACR, high M2-like macrophage score was associated with lower overall survival. Furthermore, increased frequencies of circulating myeloid suppressor populations were detected both at baseline and after NACR in non-responder patients by high-dimensional flow cytometry. We are validating our observations on a larger patient cohort with other tumor and circulating markers for lymphocytes, macrophages and immune activation. Conclusions Collectively, these preliminary results support pre-existing immune-mediated mechanisms of anti-tumor response stimulated by NACR in the fraction of responder EAC patients, suggesting possible ways to stratify the patients and direct them to more tailored neoadjuvant and/or adjuvant treatments, possibly in combination with personalized immunotherapy approaches. Legal entity responsible for the study The authors. Funding Marie Curie Individual Fellowship GA 752698 EMBO Long-term fellowship ALTF 1358-2016. Disclosure All authors have declared no conflicts of interest.