CYP eicosanoid pathway mediates colon cancer-promoting effects of dietary linoleic acid

Abstract

Human and animal studies support that consuming a high level of linoleic acid (LA, 18:2ω-6), an essential fatty acid and key component of the human diet, increases the risk of colon cancer. However, results from human studies have been inconsistent, making it challenging to establish dietary recommendations for optimal LA intake. Given the importance of LA in the human diet, it is crucial to better understand the molecular mechanisms underlying its potential colon cancer-promoting effects. Using LC-MS/MS-based targeted lipidomics, we find that the cytochrome P450 (CYP) monooxygenase pathway is a major pathway for LA metabolism in vivo. Furthermore, CYP monooxygenase is required for the colon cancer-promoting effects of LA, since the LA-rich diet fails to exacerbate colon cancer in CYP monooxygenase-deficient mice. Finally, CYP monooxygenase mediates the pro-cancer effects of LA by converting LA to epoxy octadecenoic acids (EpOMEs), which have potent effects on promoting colon tumorigenesis via gut microbiota-dependent mechanisms. Overall, these results support that CYP monooxygenase-mediated conversion of LA to EpOMEs plays a crucial role in the health effects of LA, establishing a unique mechanistic link between dietary fatty acid intake and cancer risk. These results could help in developing more effective dietary guidelines for optimal LA intake and identifying subpopulations that may be especially vulnerable to LA's negative effects.

Keywords: colon cancer; cytochrome P450 monooxygenase; epoxy octadecenoic acids (EpOMEs); linoleic acid; lipidomics; precision health.

Figure 1.. LA-rich diet increases development of colon tumorigenesis in mice.

A. C57BL/6 mice were treated with Ctrl or LA-rich diet, then stimulated with AOM/DSS to induce colon tumorigenesis. B. Colon tumorigenesis in mice. C. Representative images of H&E and immunohistochemical staining of PCNA and β-catenin, and quantification of the immunohistochemical staining. D. FACS analysis of immune cells in colon. E. Gene expression in colon. The data are mean ± SEM, n = 9-11 mice per group.

Figure 2.. Cytochrome P450 (CYP) monooxygenase pathway is a major pathway to metabolize LA in vivo.

A. Volcano plot of the metabolites detected in the plasma of AOM/DSS mice treated with Ctrl diet or LA-rich diet. Each plotted point represents a lipid metabolite. The X-axis shows the log₂ of the fold of change of the metabolite detected in the LA-rich group vs. that of the Ctrl group. The Y-axis shows the −log₁₀ of the P value calculated by using Welch’s T-test. B. Metabolism of LA by cytochrome P450 monooxygenases leads to the formation of 9,10- and 12,13-EpOME, which are further metabolized by epoxide hydrolase to generate 9,10- and 12,13-DiHOME, respectively. C. Concentrations of EpOMEs and DiHOMEs in plasma. D. Volcano plot of the metabolites detected in the colon of the AOM/DSS mice treated with Ctrl diet or LA-rich diet. The X-axis shows the log₂ of the fold of change of the metabolite detected in the LA-rich group vs. that of the Ctrl group. The Y-axis shows the −log₁₀ of the P value calculated by using Welch’s T-test. E. Concentrations of EpOMEs and DiHOMEs in the colon. The data are mean ± SEM, n = 9-10 mice per group.

Figure 3.. The colon cancer-enhancing effects of dietary LA are mediated by the CYP monooxygenase pathway.

A. WT (Cyp2c^+/+) mice or Cyp2c^+/− mice were treated with Ctrl or LA-rich diet, then the development of AOM/DSS-induced colon tumorigenesis was examined. B-C. Colon tumorigenesis in mice. D. Histological and immunochemistry staining of the colon. Left panel: representative H&E and immunohistochemical images of the colon, right panel: quantification of PCNA and β-catenin staining. E. Gene expression in colon. The data are mean ± SEM, n = 19-24 mice per group.

Figure 4.. EpOME, but not its downstream metabolite DiHOME, increases the development of colon tumorigenesis in mice.

A. C57BL/6 mice were stimulated with AOM/DSS to induce colon tumorigenesis, then treated with 12,13-EpOME, 12,13-DiHOME, or vehicle via osmotic mini-pumps. B. Colon tumorigenesis in mice. C. Representative H&E and immunohistochemical images of the colon, and quantification of immunohistochemical staining. D. Gene expression in colon. The data are mean ± SEM, n = 10-11 mice per group.

Figure 5.. EpOME alters the diversity and composition of colon cancer-associated gut microbiota in mice.

A. C57BL/6 mice were stimulated with AOM/DSS to induce colon tumorigenesis, then treated with 12,13-EpOME via osmotic mini-pumps. At t = week 0 and 9, fecal samples were collected for sequencing. B. α diversity of gut microbiota. C. β diversity of the microbiota. D. Composition of the microbiota at phylum levels. E. Composition of the microbiota at genus levels. n = 10-11 mice per group.

Figure 6.. EpOME promotes colon tumorigenesis via gut microbiota-dependent mechanisms.

A. C57BL/6 mice were stimulated with AOM/DSS to induce colon tumorigenesis, then treated with 12,13-EpOME or vehicle via osmotic mini-pumps, with or without co-administration of an antibiotic cocktail in drinking water. B. Colon tumorigenesis. C. Representative H&E and immunohistochemical images of the colon, and quantification of immunohistochemical staining. D. Gene expression in colon. The data are mean ± SEM, n = 11-22 mice per group.

Figure 7.. Proposed model: CYP monooxygenase-mediated conversion of LA to EpOMEs contributes to the colon cancer-enhancing effects of dietary LA.

After a high intake of dietary LA, part of the LA is incorporated into membrane phospholipids of the colon tissues. The membrane-incorporated LA is further metabolized by CYP monooxygenases (largely CYP2C and CYP2J isoforms), which are upregulated in colon tumors, leading to the formation of EpOMEs. EpOMEs have potent effects to induce colon tumorigenesis via gut microbiota-dependent mechanisms. In contrast, DiHOMEs, which are downstream metabolites of EpOMEs, don’t have such effects. Overall, these results support that CYP monooxygenase-mediated conversion of LA to EpOMEs plays a critical role in mediating the colon cancer-enhancing effects of dietary LA.