Hepatocellular Carcinomas With Mutational Activation of Beta-Catenin Require Choline and Can Be Detected by Positron Emission Tomography

Abstract

Background & aims: In one-third of hepatocellular carcinomas (HCCs), cancer cells have mutations that activate β-catenin pathway. These cells have alterations in glutamine, bile, and lipid metabolism. We investigated whether positron emission tomography (PET) imaging allows identification of altered metabolic pathways that might be targeted therapeutically.

Methods: We studied mice with activation of β-catenin in liver (Apc^ko-liv mice) and male C57Bl/6 mice given injections of diethylnitrosamine, which each develop HCCs. Mice were fed a conventional or a methionine- and choline-deficient diet or a choline-deficient (CD) diet. Choline uptake and metabolism in HCCs were analyzed by micro-PET imaging of mice; livers were collected and analyzed by histologic, metabolomic, messenger RNA quantification, and RNA-sequencing analyses. Fifty-two patients with HCC underwent PET imaging with ¹⁸F-fluorodeoxyglucose, followed by ¹⁸F-fluorocholine tracer metabolites. Human HCC specimens were analyzed by immunohistochemistry, quantitative polymerase chain reaction, and DNA sequencing. We used hepatocytes and mouse tumor explants for studies of incorporation of radiolabeled choline into phospholipids and its contribution to DNA methylation. We analyzed HCC progression in mice fed a CD diet.

Results: Livers and tumors from Apc^ko-liv mice had increased uptake of dietary choline, which contributes to phospholipid formation and DNA methylation in hepatocytes. In patients and in mice, HCCs with activated β-catenin were positive in ¹⁸F-fluorocholine PET, but not ¹⁸F-fluorodeoxyglucose PET, and they overexpressed the choline transporter organic cation transporter 3. The HCC cells from Apc^ko-liv mice incorporated radiolabeled methyl groups of choline into phospholipids and DNA. In Apc^ko-liv mice, the methionine- and choline-deficient diet reduced proliferation and DNA hypermethylation of hepatocytes and HCC cells, and the CD diet reduced long-term progression of tumors.

Conclusions: In mice and humans, HCCs with mutations that activate β-catenin are characterized by increased uptake of a fluorocholine tracer, but not ¹⁸F-fluorodeoxyglucose, revealed by PET. The increased uptake of choline by HCCs promotes phospholipid formation, DNA hypermethylation, and hepatocyte proliferation. In mice, the CD diet reverses these effects and promotes regression of HCCs that overexpress β-catenin.

Keywords: CTNNB1; Liver Cancer; OCT3; Wnt Pathway.