Background: Gastric adenocarcinoma (GA) represents a leading cause of cancer death worldwide. For localized and locally advanced (LA) GA, accounting for 30% of new diagnoses, surgery, with or without peri-operative chemotherapy (CTX), remains the cornerstone of treatment. Nevertheless, the cure rate remains unsatisfactory. Genomic biomarkers have been tested to tailor anticancer treatments, but no one is able to guide the treatment choice in GA. Gut microbiota represents an emerging area of investigation in cancer, as a key modulator of host immune response. However, its role in GA on treatment tolerability and outcome is not unraveled. Additionally, radiomics, which can perform massive data mining to increase diagnostic power, and extensive dietary assessment are fast-growing tools. Our aim is to set an innovative approach to mapping the interaction among nutrition, microbiome, genomics, and radiomics and correlate them with clinical outcomes. Methods: We are conducting a prospective observational trial in GA patients (pts), candidates to receive peri-operative CTX or upfront surgery. For each patient, we longitudinally collect blood, fecal and salivary samples, alongside clinical and nutritional information. Additionally, past dietary consumption is measured using the food frequency questionnaire. At baseline, CT scan for staging, radiomic analysis, upper digestive endoscopy with biopsy, and molecular biomarkers are performed. Additional samples are taken from tumor and surrounding normal mucosa (1 to 3 cm) for microbiome analysis. Genomic DNA from stool, buccal and gastric tissue samples will be extracted and subjected to 16S metagenomic sequencing. Taxonomic and functional features within and between anatomical compartments will be correlated with clinical and radiomic data. Results: Here, we are presenting preliminary data of fecal samples from 35 GA pts in comparison with a cohort of healthy subjects (HCs) collected at our Institution. Pts’ characteristics are outlined here. We observed that the structure of the gut microbiota of GA pts is distinct from HCs in terms of beta diversity, and this difference is maintained after CTX. In particular, GA’s microbiota was enriched in S. anginosus, among other taxa. Conclusions: Our preliminary data support the feasibility of the study. The differences in structure and composition of the gut microbiota of GA pts compared to HCs confirm previous reports while providing the rationale for developing gut microbiota profiling into a non-invasive biomarker, to implement early diagnosis and prevention. The study is ongoing and actively recruiting.