Abstract 3411: High-throughput drug screening combined with a druggable target CRISPR drop-out screen identifies therapeutic vulnerabilities of BCOR-CCNB3 fusion positive sarcomas

Abstract

Abstract Undifferentiated round cell sarcomas, which carry the BCOR-CCNB3 fusion gene, have traditionally been called “Ewing-like sarcomas” and have been treated with Ewing sarcoma treatment regimens. While there has been emerging evidence that these BCOR-CCNB3 sarcomas (BCS) are molecularly distinct from Ewing sarcomas, the biology of these tumors is poorly understood, and patients have few options when they do not respond to Ewing sarcoma-based therapies. Here, we report the establishment of a novel BCS cell line, which we utilized to identify a unique BCS-specific therapeutic vulnerability using a hybrid screening approach consisting of a high-throughput drug screen combined with a CRISPR/Cas9-based knock out genetic screen. The BCOR-CCNB3-positive sarcoma cell line NBC-1 was established from the tumor tissue obtained from an 18-year-old male with a soft tissue tumor of the buttock with lymph node metastasis, and the expression of the fusion BCOR-CCNB3 transcript was confirmed by RT-PCR. An in vitro high-throughput screening was performed using the L1300 library of 2570 drugs, where the cells were incubated for 48 hours with drug before cell viability was measured. This drug screen identified 148 drugs, which included 84 FDA-approved drugs. While these drugs predictably included a significant number of conventional cytotoxic chemotherapy drugs currently used to treat BCS, such as vincristine and doxorubicin, several other drug classes emerged, identifying twenty tyrosine kinase inhibitors, proteasome inhibitors such as Carfizomib, Ixazomib, and Bortezomib, and a histone deacetylase inhibitor, Panobinostat, all of which had distinctly higher cytotoxicity compared to the Ewing sarcoma cell controls. Next, we utilized a custom-made CRISPR panel consisting of 9771 sgRNAs targeting 1221 genes where we were able to identify a specific pharmacologic inhibitor in the literature. Using this library, we carried out a genetic loss-of-function screen to identify genes that are required for the proliferation of BCS. Based on these initial findings, we were able to demonstrate that BCS tumors have distinct vulnerabilities from Ewing sarcoma, suggesting that a BCS-specific therapeutic approach is necessary. Citation Format: Naoki Oike, Hiroyuki Kawashima, Akira Ogose, Takashi Ariizumi, Yudai Murayama, Hajime Umezu, Molishree Joshi, Colin Sempeck, Deandra Warker, Avery Bodlak, Naoto Endo, Masanori Hayashi. High-throughput drug screening combined with a druggable target CRISPR drop-out screen identifies therapeutic vulnerabilities of BCOR-CCNB3 fusion positive sarcomas [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 3411.