Role of AMP-18 in oral mucositis

Oral mucositis (OM) is a devasting toxicity associated with cytotoxic cancer therapy. Antrum mucosal protein (AMP)-18 and a synthetic peptide surrogate, exhibit cell protective and mitogenic properties in in vitro and in vivo models of gastrointestinal epithelial cell injury. The mucosal barrier-protective effects may be mediated by AMP-18's capacity to increase accumulation of specific tight junction (TJ) and adherens junction proteins, and also protect against their loss after injury. Here we asked if AMP peptide could protect the oral mucosa and speed healing from radiation-induced injury. We found AMP peptide prevented radiation-induced OM in a murine model. The peptide also stimulated HaCaT cell growth used to model the oral mucosa. Binding of recombinant human (rh) AMP-18 to the plasma membrane of keratinocytes in normal human oral mucosal tissue suggested that its effects may be receptor mediated. Using an immobilized His-tagged rhAMP-18 fusion protein the receptor was identified as the cholecystokinin-B/gastrin receptor (CCKBR) by affinity purification and mass spectrometry analysis. CCKBR was expressed and co-immunoprecipitated with exogenous rhAMP-18 in diverse epithelial cell lines. Immunofluorescence staining revealed that rhAMP-18 colocalized with CCKBR on the surface of CCKBR-transfected cells. Furthermore, rhAMP-18-stimulated signaling pathways were blocked by a CCKBR-specific antagonist, YM022. rhAMP-18 enhanced viability and growth of CCKBR-transfected, but not empty vector-transfected cells. These results suggest the importance of epithelial junctional integrity in the pathogenesis of OM and demonstrate that AMP-18, by targeting TJ proteins through the activation of CCKBR, could provide a novel strategy for the prevention and treatment of OM.