Abstract 6348: Cholecystokinin receptor blockade modulates in vitro fibrosis gene expression
Authors: Martha Gay, Hong Cao, Jill P. Smith
Journal: Cancer Research (2025)

Abstract Introduction: Cholecystokinin (CCK) receptors, G-protein coupled receptors, play crucial roles in gastrointestinal physiology, but their overexpression in cancer and fibrosis is implicated in disease progression and therapeutic resistance. Recent evidence suggests that activation of CCK receptors in hepatic stellate cells (HSCs) induces fibrosis-associated gene expression, contributing to liver pathology in conditions like metabolic dysfunction-associated steatohepatitis (MASH) and hepatocellular carcinoma (HCC). We have shown that a diet high in saturated fat induces the expression of hepatic CCK- B receptors in a murine model of nonalcoholic steatohepatitis. The CCK receptor antagonist, proglumide, reversed inflammation, fibrosis, and steatosis prevented the development of HCC in the murine Metabolic dysfunction-associated steatohepatitis (MASH) model. Hypothesis: The blockade of CCK-B receptor signaling using proglumide will reduce the activation of fibrosis-related genes and prevent the progression of liver injury and carcinogenesis in both murine and human in vitro models. Methods/ murine cells: Using C57BL/6 mice we will follow the standard protocol for isolation of murine HSC. Stellate cells will be isolated and cultured then stimulated with CCK in vitro (1-100nM). Another group of stellate cells will be treated with CCK and proglumide to determine if proglumide can prevent activation in fibrosis genes. Stellate cellular protein will be also extracted and subjected to Western analysis for fibrosis/collagen associated proteins. Macrophages release cytokines/chemokines that activate stellate cells. For our co-culture experiments we will culture murine RAW264.7 macrophages with the mouse HSC to activate fibrosis genes and proteins by cytokine /chemokine activation of stellate cells. Methods/ human cells: We will use the immortalized LX-2 human HSC line to evaluate the activation of the fibrosis genes with addition of CCK to the growth media and examine if proglumide prevents the activation of the LX-2 cells and perform independently CCK-B or CCK-A receptor CRISPR to determine if CCK-induced fibrosis activation is mediated by the CCK-B or CCK-A receptor. Clonal CRISPR CCK-receptor KO stellate cells will also be activated by co- culture and /or with spent media from our human U937 macrophages.Anticipated outcomes: When stellate cells are grown on plastic in vitro, they become activated. We know the cytokines / chemokines released from macrophages will stimulate stellate cells, but the stimulation and collagen synthesis may not be as pronounced if the stellate cells were not already in an activated state. Ongoing data collection. Conclusion: This investigation aims to clarify the molecular mechanisms by which CCK receptor signaling drives hepatic fibrosis and explore the therapeutic potential of proglumide in reversing fibrosis-related gene expression in liver disease. Citation Format: Martha Gay, Hong Cao, Jill P. Smith. Cholecystokinin receptor blockade modulates in vitro fibrosis gene expression [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 6348.