The Role of Epithelial IKKβ in the Transition from Normal to Precancerous Tissue in Esophageal Epithelium

Esophageal cancer is the 6th leading cause of cancer related death with a 5-year survival rate of under 20%. Esophageal squamous cell carcinoma (ESCC) is associated with chronic inflammation and injury. Currently, little is understood about the molecular mechanisms regulating the early stages of esophageal carcinogenesis. The NFB/IKK pathway plays a central role in inflammation and has been implicated in ESCC. However, its functional role in the transition to esophageal precancer is unknown. To investigate this, we utilized ED-L2/Cre mice to knockout IKKβ (IKKβEEC-KO) in the esophagus. These mice and littermate controls were treated with the carcinogen 4-nitroquinoline-1-oxide (4-NQO) or a vehicle for one month. Esophagi were harvested and examined through histological, protein, flow cytometry, and RNA analyses. Histological analysis showed that 4-NQO treatment led to increased inflammation and intraepithelial CD45+ immune cells which was largely attenuated in IKKβEEC-KO mice. Total T cells, Tregs, CD4+, and CD8+ T cells showed a similar trend, while macrophages showed a reverse trend. RNAseq data showed upregulation of inflammatory pathways and downregulation in wound healing related pathways in 4-NQO treated control mice versus knockout. We also saw increased expression of the chemokine CXCL9, a T cell chemoattractant. This increase was blocked in 4-NQO treated IKKβEEC-KO mice or in mice treated with anti-IFN. Additionally, we demonstrated in vitro that IFN upregulates CXCL9 in a NF-ĸB dependent manner in esophageal keratinocytes. So epithelial IKK regulates the immune landscape in the transition to precancer through IFN/CXCL9 signaling and recruitment of T cells to the esophagus.