Targetted sequencing of a gastric cancer panel for multiple primary and metastatic sites within Gastro-Oesophageal adenocarcinomas identified intra tumour heterogeneity at both the mutational and copy number level.. Gastric Intratumour Heterogeneity Study

Gastro-esophageal adenocarcinomas (GEAs) are aggressive cancers and multiple trials of targeted therapies recently failed to improve survival in these tumors. Intratumor heterogeneity (ITH) is suspected to contribute to poor outcomes. We investigated ITH in multiple regions from nine GEA primary tumors and associated lymph node metastases. Somatic mutation and DNA copy number analysis identified ITH in all tumors. Eight chromosomally instable GEAs predominantly evolved through DNA copy number aberrations whereas a microsatellite instable tumor exclusively evolved through point mutations and showed evidence for parallel evolution. ITH increased significantly with lymph node metastasis formation and subclonal aberrations activating the Mitogen Activated Protein Kinase (MAPK)-pathway were significantly enriched in nodal metastases. This shows that selection pressures in the lymph node ecosystem differ from those in the primary tumor, leading to evolutionary convergence of distinct tumors when they spread to lymph nodes. Assessment of the functional relevance of MAPK activating aberrations showed that To assess the functional impact of MAPK-activating amp evolution, we analysed published primary CIN subtype GEAs. Cytolytic activity (CYT), estimating tumor immune recognition from RNA expression data, correlated with the mutation load in GEAs harboring EGFR, ERBB2 or MET amp (p=0.04). In contrast, there was no correlation between CYT and mutation load in GEAs with KRAS or ERBB3 amp (p=0.22, NRAS/ERK2: insufficient data), implying that the specific alterations recurrently identified in LN may enable cancer cells to evade immune recognition. Downregulation of TAP and Class I MHC genes (p<0.05) in KRAS or ERBB3 amp GEAs suggested impaired antigen processing and presentation as the mechanisms driving T cell immune evasion in LN.increased pathway activity downregulates peptide antigen processing and presentation, potentially enabling cancer cells to evade immune recognition by T cells in the lymph node. These candidate metastasis drivers are also predicted to confer resistance to upstream tyrosine kinase receptor-targeting drugs. Lymph node metastasis evolution may hence foster the expansion of subclones which are pre-adapted to future targeted therapy. However, a panRAF/SRC inhibitor successfully inhibited GEA cells harboring multiple distinct upstream activating alterations, suggesting that this therapy can effectively intercept GEAs with heterogeneous subclones and the outgrowth of resistant cells. Taken together, this study identified ITH in GEAs, characterized by parallel and convergent evolution, and provides evidence that varying selection pressures in distinct ecosystems foster the evolution of intratumoral subclones with decreased immunogenicity and increased resistance to targeted therapies.