Abstract 5523: Inhibition of FASN postpones development of resistance to PLX8394 in colorectal cancer

Mariah E. Geisen; Josiane Weber Tessmann; Courtney O. Kelson; Ellen Beswick; Daheng He; Chi Wang; Abu Saleh Mosa Faisal; Jill M. Kolesar; Yekaterina Zaytseva

doi:10.1158/1538-7445.AM2025-5523

Background: Mutations in proto-oncogene B-Raf (BRAF) occur in about 10% of colorectal cancer (CRC) patients and BRAFV600E is the most common type. This subset of CRC is associated with a reduced response to chemotherapy and poor prognosis compared to BRAF wild-type CRC. While the FDA-approved encorafenib plus cetuximab therapy for BRAF-mutant CRC provides a significant benefit, only 22% of patients respond to this therapeutic approach and resistance ultimately develops in the majority of patients. Therefore, development of new efficacious strategies based on further characterization of resistance mechanisms is needed to improve outcomes for BRAFV600E CRC. We found that development of resistance to PLX8394, a second-generation BRAF inhibitor, is associated with an increase in cellular proliferation, invasion, and lipid metabolism. We identified fatty acid synthase (FASN), a key enzyme in lipid synthesis, as a major lipogenic enzyme overexpressed in PLX8394-resistant cells. Therefore, the goal of this study is to test if inhibition of FASN will enhance the efficacy of PLX8394 in BRAF-mutant CRC. Methods: HT29, PT130, and PT24249pt, parental and PLX8394-resistant, CRC cells were utilized. The effect of PLX8394 alone and in combination with FASN inhibitors (TVB3664 and C75) was tested using cell viability and colony formation assays. The synergy score was determine using a BLISS synergy model (SynergyFinder 3.0). The development of resistance to PLX8394 in the presence or absence of TVB3664 was evaluated using IC50 analysis, flow cytometry, and western blot. Results: We found that PLX8394-resistant cells exhibit an increase in FASN expression, oxidative phosphorylation, and triglyceride storage. Combination of PLX8394 and FASN inhibitors led to a significant decrease in cell viability and colony formation as compared to each drug alone. Consistently, we found a significant synergy between PLX8394 and C75 using the BLISS synergy model. FASN-knockdown cells exhibited higher sensitivity to PLX8394 as compared to control cells supporting the role of FASN in resistance to PLX8394. Importantly, an addition of TVB3664 to PLX8394 treatment postpones the development of resistance to PLX8394. Analysis of cell cycle by flow cytometry confirmed a suppression of cell cycle progression via a decrease in a number of cells entering the S phase when cells are treated with combination of PLX8394 and TVB3664. Consistently, we observed a decrease in expression of proteins associated with the cell cycle such as cyclin D, pRB, and E2F expression in this group. Conclusion: Collectively, these data show that combination of PLX8394 with FASN-targeted therapy at treatment initiation reduces BRAFV600E CRC cell proliferation via inhibition of their cell cycle progression. These findings suggest that targeting FASN could enhance the efficacy and delay the development of resistance to PLX8394 in BRAF-mutant CRC.

Abstract 5523: Inhibition of FASN postpones development of resistance to PLX8394 in colorectal cancer

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