Abstract 1175: Identification of novel combination therapies with the LSD1 inhibitor SP-2577 (Seclidemstat) in Ewing's Sarcoma using a complete druggable CRISPR library approach

Abstract

Abstract Introduction: Ewing's Sarcoma (ES) is a rare and aggressive pediatric bone tumor that is driven by a chromosomal translocation and fusion between EWS and FLI1 genes resulting in a chimeric protein. Nearly 85% of ES cases express the EWS/FLI protein which functions as a transcription factor and drives oncogenesis. It has been previously observed in ES patient samples have high levels of the histone lysine specific demethylase 1 (LSD1). LSD1 is recruited by EWS/FLI as a member of the nucleosome remodeling and histone deacetylase (NuRD) complex to repress the critical EWS/FLI targets LOX and TGFBR2. Treatment with the potent LSD1 reversible inhibitor, SP-2577 (Seclidemstat), causes de-repression of these genes and impaired tissue culture cell viability in multiple ES cell lines. In this study we investigated new targeted therapies to use in combination with SP-2577 to help increase the cytotoxicity of the drug within ES cell types. Methods: Synthego® arrayed CRISPR screening platform was used to knock down genes associated with known druggable interactions in the ES cell line SK-N-MC. Genes were knocked down singularly either in the presence/absence of SP-2577. Viability screening was used to determine genes that increased the cytotoxicity of SP-2577. Statistical analysis, z-score, and IPA analysis was performed to investigate correlations between loss of gene effect and LSD1 pathway. Hit-gene validation was performed using siRNA and/or sgRNA knockdown/knockout, and other methods such as qPCR, and western blot assay for protein expression. Results: Arrayed CRISPR screening identified 103 gene deletions that resulted in increased SP-2577 cytotoxicity in SK-N-MC cell line. Among those, the top 19 hits were selected by identifying a FDA approved drug-gene interaction. Two high- interest hits were seen to have significant interaction with the LSD1 pathway: ABL2 and SH2B3. Drug combination treatment of SP-2577 with either Ruxolitinib or Dasatinib in ES cell lines showed high synergy at low dose ratio. Downstream pathways analysis confirmed the efficacy of the drugs combination. Detailed data will be presented. Conclusion: Our data support the validity of the arrayed CRISPR screening to identify new therapeutic strategies in ES, and highlights the potential for Ruxolitinib or Dasatinib treatment in combination with SP-2577 in ES patients. Citation Format: Samuel Sampson, Trason Thode, Mohan Kaadige, Alexis Weston, Serina Ng, Kevin Drenner, Raffaella Soldi, Sunil Sharma. Identification of novel combination therapies with the LSD1 inhibitor SP-2577 (Seclidemstat) in Ewing's Sarcoma using a complete druggable CRISPR library approach [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1175.