Intratumor Lactobacillus drives ferroptosis resistance via D-lactate-STAT3 K631 lactylation in esophageal squamous cell carcinoma

Tumor-resident microbiota are increasingly recognized as active components of the gastrointestinal tumor ecosystem, yet how intratumor bacteria reshape cancer cell stress responses in esophageal squamous cell carcinoma (ESCC) remains unclear. Here, 16S rRNA gene sequencing of 102 multiregional tissue blocks from 27 patients with ESCC, integrated with untargeted metabolomics, RNA sequencing, and mass spectrometry, identified Lactobacillus, particularly Lactobacillus reuteri, as a tumor-enriched taxon associated with adverse survival. Mechanistic studies in ESCC cell lines and nude-mouse xenografts revealed a distinctive dynamic: in contrast to host-derived L-lactate, L. reuteri-derived D-lactate induced site-specific STAT3 lactylation at lysine 631, thereby promoting STAT3 dimerization and nuclear translocation. This host signaling rewiring upregulated the ferroptosis suppressors GPX4 and FTH1, reduced ferroptotic vulnerability, and enhanced ESCC growth. Disrupting bacterial D-lactate production using a ldhD-deficient L. reuteri mutant or blocking host STAT3 lactylation using STAT3-knockout cells reconstituted with wild-type STAT3 or lactylation-defective STAT3 K631R, abolished the pro-tumor and antiferroptotic effects in vitro and in vivo. Together, these findings define a tumor-resident microbe–metabolite–host signaling axis that links intratumor Lactobacillus to ferroptosis escape. By establishing bacteria-derived D-lactate as the functional driver, the study provides a novel mechanistic framework that extends beyond the classical Warburg effect for developing biomarkers and therapeutic strategies targeting STAT3 lactylation or ferroptosis sensitization. These translatable results emphasize the need for context-specific evaluation of Lactobacillus-containing probiotic supplementation in patients with ESCC.