Abstract 2102: Understanding the mechanism of action of rigosertib in recessive dystrophic epidermolysis bullosa-associated squamous cell carcinoma through single cellanalysis

Abstract

Abstract The allosteric inhibitor rigosertib (an experimental therapeutic) was previously identified as an effective inducer of G2M arrest and apoptosis in squamous cell carcinoma (SCC) keratinocytes isolated from patients with the rare genetic disease recessive dystrophic epidermolysis bullosa (RDEB), both in vitro and in vivo. Early clinical trial data treating RDEB-associated SCC with rigosertib has shown remarkable efficacy in a sub-set of patients and understanding the mechanism of action of rigosertib in this lethal cancer would provide rationale for patient stratification. Multiple mechanisms of action were previously attributed to rigosertib, including polo-like kinase-1 (PLK1), PI3-kinase, activated RAS and microtubule polymerization inhibition. Our published data comparing PLK1 siRNA with PI3-Kinase inhibitors, MAPK inhibitors and microtubule disruption favored PLK1 as a mechanism of action without definitive conclusion. We therefore sought to determine the mechanism of action using transcriptomic profiling. Flow cytometry experiments identified three main categories of time to increase G2M in nine separate RDEB SCC keratinocyte populations in culture ranging from 1 to 6 hours after exposure to 1uM rigosertib. Bulk-RNA sequencing of representative populations from time to G2M groupings failed to identify consistent transcriptional change after exposure to rigosertib. However, single cell sequencing was able to identify increases in sub-populations of cells associated with G2M as well as transient emergence of unique populations that were specific to individual patient populations after exposure to rigosertib in culture. Of note a transient population of RDEB SCC keratinocytes only present after 1 hour of exposure to rigosertib was characterized by increased expression of the potassium regulator KCNRG which has previously been associated with apoptosis in chronic lymphocytic leukemia. Ongoing work is determining the role this gene may play in the mechanism of apoptosis induction in RDEB SCC keratinocytes after exposure to rigosertib. Citation Format: Andrew P. South, Suhao Han, Paolo Fortina. Understanding the mechanism of action of rigosertib in recessive dystrophic epidermolysis bullosa-associated squamous cell carcinoma through single cellanalysis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 2102.