Chronic inflammation is implicated in the pathogenesis of esophageal squamous cell carcinoma (ESCC). The causes of inflammation in ESCC, however, are undefined. Dietary zinc (Zn)-deficiency (ZD) increases the risk of ESCC. We have previously shown that short-term ZD (6 weeks) in rats induces overexpression of the proinflammatory mediators S100a8 and S100a9 in the esophageal mucosa with accompanying esophageal epithelial hyperplasia. Here we report that prolonged ZD (21 weeks) in rats amplified this inflammation that when combined with non-carcinogenic low doses of the environmental carcinogen, N-nitrosomethylbenzylamine (NMBA) elicited a 66.7% (16/24) incidence of ESCC. With Zn-sufficiency, NMBA produced no cancers (0/21) (P<0.001). At tumor endpoint, the neoplastic ZD esophagus, as compared with Zn-sufficient esophagus, had an inflammatory gene signature with upregulation of numerous cancer-related inflammation genes (CXC and CC chemokines, chemokine receptors, cytokines and Cox-2) in addition to S100a8 and S100a9. This signature was already activated in the earlier dysplastic stage. Additionally, time-course bioinformatics analysis of expression profiles at tumor endpoint and before NMBA exposure revealed that this sustained inflammation was due to ZD rather than carcinogen exposure. Importantly, Zn replenishment reversed this inflammatory signature at both the dysplastic and neoplastic stages of ESCC development, and prevented cancer formation. Thus, the molecular definition of ZD-induced inflammation as a critical factor in ESCC development has important clinical implications with regard to development and prevention of this deadly disease.