Abstract 5356: RIDR-PI-103, ROS-activated PI3K inhibitor prodrug inhibits cell growth and impairs the PI3K/Akt pathway in BRAF and MEK inhibitor resistant BRAF-mutant melanoma cells
Authors: Patel Hima , Mishra Rosalin , Wier Adam , Mokhtarpour Nazanin , Merino Edward J. , Garrett Joan T. 
Published: June 2022

The five-year survival rate for patients diagnosed with metastatic melanoma is only 27%. About half of melanomas have the presence of the BRAFV600E oncoprotein, which leads to hyperactivation of the mitogen activated protein kinase (MAPK) pathway. The MAPK pathway is known to crosstalk with the PI3K/Akt pathway. Treatment with the BRAF inhibitor, Dabrafenib and the MEK inhibitor, Trametinib is limited as patient responses begin to drop due to acquired resistance to these kinase inhibitors. High concentrations of reactive oxygen species (ROS) accompany this resistance. ROS-induced drug release (RIDR)-PI-103 is a novel prodrug with a self-cyclizing moiety linked to PI-103, a PI3K inhibitor. Under high ROS conditions, RIDR-PI-103 releases PI-103, which inhibits conversion of PIP2 to PIP3. We have generated Trametinib and Dabrafenib resistant (TDR) cells for 3 BRAF-mutant cell lines: A375, WM115 and WM983B from parental (drug sensitive) cell lines. Studies in our lab demonstrate that TDR cells maintain p-Akt levels compared to parental counterparts and have significantly higher ROS. This is a rationale to explore the efficacy RIDR-PI-103 in TDR cells. We tested the effect of RIDR-PI-103 on cell viability for 1789C melanocytes, 1788B melanocytes, A375 TDR, WM983B TDR, and WM115 TDR using an MTT proliferation assay. RIDR-PI-103 exhibited less toxicity as compared to PI-103 at 5 µM in normal melanocytes. While RIDR-PI-103 significantly inhibited TDR cell proliferation at 5 and 10 µM compared to cells treated with vehicle DMSO and melanocytes. This effect was consistent when TDR cells were treated with RIDR-PI-103 in long-term crystal violet cell proliferation assays. WM115 TDR and WM983B TDR cells were highly sensitive to RIDR-PI-103 at 5 µM, while A375 TDR cells had only ~50% inhibition at 5 µM in both cell proliferation assays. 24 hour treatment with RIDR-PI-103 inhibited p-Akt (S473) along with downstream p-S6 (S240/244) and p-S6 (235/236). To examine the mechanism of action of RIDR-PI-103 and to test that the drug release is ROS induced, we assessed the effect of glutathione (GSH), an antioxidant on the TDR cells in the presence or absence of RIDR-PI-103. GSH alone did not affect proliferation of TDR cells. Addition of GSH to RIDR-PI-103 significantly rescued the cell proliferation in all three TDR cell lines. We next tested the effect of t-butyl hydrogen peroxide (TBHP), a ROS inducer on TDR cells with lower concentrations of RIDR-PI-103. Addition of TBHP to RIDR-PI-103 significantly induced proliferation in TDR cells. Ongoing experiments are focused on assessing the effect of RIDR-PI-103 on the mTOR pathway. Examining the efficacy of RIDR-PI-103 on BRAF and MEK inhibitor resistant cells will expand possible treatment options and open up avenues for the development of new treatment therapies for BRAF-mutant melanoma patients.

Citation Format: Hima Patel, Rosalin Mishra, Adam Wier, Nazanin Mokhtarpour, Edward J. Merino, Joan T. Garrett. RIDR-PI-103, ROS-activated PI3K inhibitor prodrug inhibits cell growth and impairs the PI3K/Akt pathway in BRAF and MEK inhibitor resistant BRAF-mutant melanoma cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 5356.