Evolution of immune and stromal cell states and ecotypes during gastric adenocarcinoma progression

Understanding how the tumor microenvironment (TME) is orchestrated along the continuum of gastric adenocarcinoma (GAC) may uncover novel therapeutic strategies. Here, we performed comprehensive single-cell profiling of precancerous lesions, localized and metastatic GACs, identifying alterations in TME cell states and phenotypes during tumorigenesis and progression. IgA+ plasma cells appear to be the most active regulators in the premalignant microenvironment, whereas immunosuppressive myeloid and stromal subsets dominated the late-stage GACs. We discover six TME ecotypes, four of which are unique to the premalignant/progression in the continuum, with two ecotypes, identified in primary GACs, showing strong associations with histopathologic, genomic characteristics and prognosis. Extensive stromal remodeling is evident with GAC progression. High SDC2 expression in cancer-associated fibroblasts (CAFs) correlates with aggressive phenotypes and poor survival and SDC2 overexpression in CAFs contributes to tumor growth. This study illustrates a roadmap of the GAC TME and highlights potential targets for further investigation. Six patients (Pt1-5, Pt9) were enrolled and diagnosed at Zhejiang Cancer Hospital. Diagnosis was confirmed by experienced gastrointestinal pathologists and radiologists and fresh biopsies (primary tumors, ovarian or liver metastases, n = 17) were obtained from each patient, with adjacent normal tissue or peripheral blood as controls. The primary tumor tissues and adjacent non-neoplastic tissues were collected by gastroscopy. The ascites samples were collected when patients required a therapeutic paracentesis and cells were isolated for scRNA-seq. The liver metastatic tissues were collected when patients required needle aspiration biopsy during the diagnostic procedure and the ovary metastatic tissues were collected when patients required cytoreductive surgery. None of these six patients were treated with chemotherapy or radiotherapy prior to sample collection. **Raw data not provided due to patient privacy concerns**