Salmonella Abundance Associated with Disease Progression in Stomach Adenocarcinoma and Decreased Intratumoral Natural Killer Cell Infiltrate

Introduction: Gut microbiota dysbiosis has been implicated in the biology of stomach adenocarcinoma (STAD). We hypothesize that alterations in the abundance of Salmonella, among the most commonly found in the human gut microbiome, may affect infiltrating immune cell density in the intratumor microbiome of stomach adenocarcinoma. In this report, we assess the associations of intratumoral Salmonella RNA signatures in STAD with overall, disease-specific, disease-free, progression-free survival and with intratumoral Natural Killer (NK) cell fraction. Methods: Intratumoral microbial RNA sequencing (RNA-seq) data was procured as described previously by Poore et al. (Nature, 2020) and used to calculate average log2 counts per million (CPM) of Salmonella. Abundance of intratumoral immune cell types was estimated via the quanTIseq method described previously by Finotello et al. (Genome Medicine, 2019). The Cancer Genome Atlas STAD clinical dataset was downloaded from cBioPortal.org. Relative hazard ratios (HRs) for progression-free, disease-specific, and disease-free survival were estimated using multivariate cox proportional hazards models within the lifelines python package. Age, gender, and stage were included as covariates. Kaplan-Meier survival analysis was conducted using python Microbial abundance was correlated with immune cell abundance using Spearman correlation analysis using the python scipy package. The P-value level of significance was set at < 0.05 for independent tests. Results: Our study included 406 patients with STAD and Salmonella RNA-seq data. Comparison of the rate of survival for patients in the top and bottom halves of Salmonella log CPM showed higher Salmonella abundance was associated with significantly decreased progression-free (P < 0.0034), disease-specific (P < 0.0063), and disease-free survival (P < 0.0452) (Figure 1). A multivariate Cox proportional hazards model demonstrated that individuals with increased Salmonella abundance in STAD had increased risk of disease progression (HR 1.34; p = 0.0007) (Table 1). Spearman correlation analysis showed that intratumoral Salmonella abundance is significantly negatively correlated with NK Cell abundance (correlation coefficient = -0.109; P = 0.0279). Conclusion: Our results suggest that patients with increased Salmonella genetic signatures in STAD tumor samples have worsening survival outcomes. This is partially explained by our data demonstrating a decrease in Natural Killer cell infiltrate, suggesting lowered immune response.Figure 1.: Increased intratumoral Salmonella is associated with decreased survival in Stomach Adenocarcinoma A) Disease-free Survival (p = 0.045) B) Disease-specific Survival (P < 0.006) C) Progression-free Survival (P < 0.003). Table 1. - Associations of Salmonella abundance with survival in Stomach Adenocarcinoma using Cox Proportional Hazards Model Malignancy Survival Outcome Hazard Ratio P-value Stomach adenocarcinoma Disease-specific Survival 1.34 0.0063 Stomach adenocarcinoma Progression-free Survival 1.19 0.0034 Stomach adenocarcinoma Disease-free Survival 1.1 0.0452