Mediation of sphingolipid metabolism on the relationship between human nitrosamines exposure and esophageal cancer risk

Although nitrosamines are known as potent carcinogenic contaminants with multisystem toxicity, the metabolic mechanisms driving esophageal carcinogenesis under multi-nitrosamine co-exposure remain poorly understood. This molecular epidemiological study sought to identify dynamic metabolic signatures mediating nitrosamine-associated esophageal squamous cell carcinoma (ESCC) risk. We quantified urinary levels of nine nitrosamines in participants across esophageal lesion stages (RE/BCH, DYS, ESCC) and employed UPLC-MS/MS metabolomics to investigate exposure-response relationships and mediation effects of key metabolites. Distinct stage-specific nitrosamine profiles were observed. Sphingolipid metabolism emerged as a critical pathway in ESCC pathogenesis: DH-SPH, DH-S1P, and S1P were positively associated with increased risk, while SPH and the SPH/S1P ratio demonstrated protective effects. Mediation analysis revealed metabolite-specific pathways, with DH-S1P acting as a shared mediator in NDphA-, NMEA-, and NPIP-induced ESCC pathways, and S1P mediating NMEA-associated progression from RE/BCH to ESCC. Sensitivity analyses indicated differential robustness across pathways. E-value assessment revealed that NDphA-related pathways exhibited high resistance to unmeasured confounding (E-value > 2). Critical ρ analysis indicated that DH-S1P-mediated pathways in ESCC exhibited high credibility (ρ thresholds ≈ 0.3), and SPH/S1P-mediated pathways yielded conservative effect estimates. ROC analysis with bootstrapping validation suggested the potential discriminative ability of sphingolipid metabolites: S1P showed promise in distinguishing across the disease spectrum, and a combined panel demonstrated improved performance for ESCC prediction and nitrosamine exposure discrimination in this study population. Collectively, these findings underscore sphingolipid dysregulation as a key associative mediator linking nitrosamine exposure to ESCC progression, and support their potential as candidate biomarkers for early screening and exposure monitoring in high-risk populations. However, due to the cross-sectional design and absence of external validation, these findings await confirmation in prospective and functional studies.