CB-RAF600E-1 exerts efficacy in vemurafenib-resistant and non-resistant-melanoma cells via dual inhibition of RAS/RAF/MEK/ERK and PI3K/Akt signaling pathways

Mesfer Al Shahrani; Prasanna Rajagopalan; Mohammad Abohassan; Mohammad Alshahrani; Yasser Alraey

doi:10.1016/j.sjbs.2022.103285

CB-RAF600E-1 exerts efficacy in vemurafenib-resistant and non-resistant-melanoma cells via dual inhibition of RAS/RAF/MEK/ERK and PI3K/Akt signaling pathways

Saudi J Biol Sci. 2022 Jun;29(6):103285. doi: 10.1016/j.sjbs.2022.103285. Epub 2022 Apr 23.

Authors

Mesfer Al Shahrani¹, Prasanna Rajagopalan^{1

2}, Mohammad Abohassan¹, Mohammad Alshahrani¹, Yasser Alraey¹

Affiliations

¹ Department of Clinical Laboratory Sciences, College of Applied Medical Sciences, King Khalid University, Abha, Saudi Arabia.
² Central Research Laboratory, College of Applied Medical Sciences, King Khalid University, Abha, Saudi Arabia.

Abstract

Background and aim: Predicting novel dual inhibitors to combat adverse effects such as the development of resistance to vemurafenib in melanoma treatment due to the reactivation of MAPK and PI3K/AKT signaling pathways is studied to help in reversal of cancer symptoms.Reversal of cancer symptoms in melanoma associated with vemurafenib resistance is driven by reactivation of MAPK and PI3K/Akt signaling pathways. Novel dual inhibitors targeting these proteins would be beneficial to combat resistance.

Methods: High-throughput virtual screening of the ChemBridge library against B-RAFV600E and Akt was performed using an automated protocol with the AutoDock VINA program. Luminescence and time-resolved fluorescence kits were used to measure enzyme activities. The MTT assay was used to determine proliferation in normal and vemurafenib-resistant A375 cells. Flow cytometry was used to examine apoptosis, cell cycle, and phosphorylation of ERK/Akt signaling pathway.

Results: High-throughput screening from the ChemBridge library identified 15 compounds with high binding energy towards B-RAFV600E; among these, CB-RAF600E-1 had the highest ΔG_binding score -11.9 kcal/mol. The compound also had a high affinity towards Akt, with a ΔG_binding score of -11.5 kcal/mol. CB-RAF600E-1 dose-dependently inhibited both B-RAFV600E and Akt with IC₅₀ values of 635 nM and 154.3 nM, respectively. The compound effectively controlled the proliferations of normal and vemurafenib-resistant A375 cells, with GI₅₀ values of 222.3 nM and 230.5 nM, respectively. A dose-dependent increase in the sub G₀/G₁ phase of the cell cycle and total apoptosis was observed following compound treatment in both normal and vemurafenib-resistant melanoma cells. Treatment with CB-RAF600E-1 decreased the pERK/pAkt dual-positive populations in normal and vemurafenib-resistant A375 cells.

Conclusion: CB-RAF600E-1, identified as a novel dual inhibitor effective against normal and vemurafenib-resistant melanoma cells, requires further attention for development as an effective chemotherapeutic agent for melanoma management.

Keywords: Akt; BRAFV600E; ERK; High throughput; Melanoma; Resistance; Vemurafenib.