CDK4/6 inhibition to resensitize BRAF/EGFR inhibitor in patient-derived BRAF/PTEN-mutant colon cancer cells.

Background: In v-raf murine sarcoma viral oncogene homolog B1 (BRAF)-mutant colorectal cancer (CRC), encorafenib-cetuximab has been established as standard second-line therapy, but not all patients respond and the duration of response is relatively short. Overcoming intrinsic or acquired resistance to BRAF/EGFR inhibitors is crucial for enhancing treatment outcomes in metastatic BRAF-mutated CRC. The aim of the study is to investigate the resistance mechanisms in BRAF-mutant CRC patient refractory to BRAF/EGFR targeted therapy. Methods: We established patient-derived cells (PDCs) from a patient with BRAF/PTEN-mutant metastatic colon cancer who progressed rapidly on encorafenib plus cetuximab. To explore potential treatment options for inherent resistance caused by simultaneous PTEN mutation in BRAF-mutated CRC, we conducted cell viability assays using PDCs treated with encorafenib-cetuximab in combination with a cyclin-dependent kinase-4 and 6 (CDK4/6) inhibitor. Results: The patient's tumor had concurrent PTEN loss-of-function alteration at diagnosis and PDCs were generated from ascites after resistance to the BRAF/EGFR inhibitor. The PDCs were resistant to the encorafenib-cetuximab combination even at a high concentration of cetuximab (up to 500 µg/mL). Adding the CDK4/6 inhibitor, ribociclib, to encorafenib-cetuximab showed a synergistic effect in a proliferation assay. Ribociclib plus encorafenib-cetuximab represented a significantly lower expression of Ki-67 compared to the dual combination alone. An MTS assay showed that triplet therapy with ribociclib, encorafenib, and cetuximab suppressed cell viability more efficiently than the two-drug combinations. Investigating the combined effect of triplet therapy using the calculated combination index (CI) showed that ribociclib had a synergistic effect with encorafenib-cetuximab when applied to PDCs with a concurrent BRAF/PTEN mutation. Conclusions: Our results suggest that combining the CDK4/6 inhibitor with the BRAF/EGFR inhibitor might be a novel treatment strategy for concomitant BRAF and PTEN-mutant CRC.