Heterodimerization of cholecystokinin 1 and cholecystokinin 2 receptors in gallbladder cancer: a new mechanism for carcinogenesis.

PURPOSE: Cholecystokinin is present in abundance in gallbladder tissue and mediates function through two structurally related receptors, CCK1R and CCK2R. Heterodimerization of these receptors is known to impact cell growth in vitro. However, the significance of these heterodimers in gallbladder carcinogenesis is relatively unknown. METHODS: Therefore, we evaluated the expression and the dimerization status of CCK1 and CCK2 receptors in human gallbladder carcinoma cell line (GBC-SD) and resected gallbladder tissue from normal (n = 10), cholelithiasis (n = 25) and gallbladder cancer (n = 25) by immunofluorescence/immunohistochemistry and western blot. The dimerization status of CCK1R and CCK2R was evaluated by co-immunoprecipitation. To understand the effect of heterodimerization of these receptors on growth-related signaling pathways, the expression of p-AKT, rictor, raptor and p-ERK was evaluated by western blot. RESULTS: We demonstrated the expression and heterodimerization of CCK1 and CCK2 receptor in GBC-SD gall bladder carcinoma cell line. Knockdown of CCK1R and CCK2R in the cell line led to significant reduction in p-AKT (P = 0.005; P = 0.0001) and rictor (P < 0.001; P < 0.001) levels. In tissue samples, significantly higher expression of CCK1R and CCK2R was observed in gallbladder cancer when compared to other groups both by immunohistochemistry (P = 0.008 and P = 0.013) and western blot (P = 0.009 and P = 0.003). An increase in heterodimer formation of CCK1R with CCK2R was observed in gallbladder cancer when compared to normal and cholelithiasis tissues. No significant difference in the expression of p-AKT and p-ERK was observed between the three groups. CONCLUSION: Our results provide the first evidence of heterodimerization of CCK1R and CCK2R in gallbladder tissue, and its association with development of gallbladder cancer. This finding has potential clinical and therapeutic significance.