Abstract 1844: Anti-tumor activity of a dual BET/CBP/EP300 inhibitor, NEO2734, in undifferentiated pleomorphic sarcomas and identification of genes involved in resistance

Introduction: Undifferentiated pleomorphic sarcoma is the most frequent and the most aggressive sarcoma subtype. Despite adequate locoregional treatment, up to 40% will develop metastatic disease. Doxorubicin represents the standard 1st line of treatment for patient with advanced disease. However, its activity is limited with a response rate of only 10% and a progression-free survival of less than 6-months. Identification of new therapeutic strategies is therefore an important medical need. Bromodomain and extra-terminal domain (BET) proteins, cyclic adenosine monophosphate response element-binding protein (CBP), and the E1A-binding protein of p300 (EP300) are important players in histone acetylation. BET inhibitors have already shown pre-clinical activity in translocation-related sarcomas such as Ewing tumors. However, BET and dual BET/P300 inhibitors could have also relevant anti-tumor activity if MYC driven tumors. We previously found that at least a subgroup of UPS is characterized by a strong expression of MYC-targets pathway (Toulmonde et al, EBIOMEDICINE, 2020) Methods: The anti-tumor activity of three compounds which inhibits either CBP/P300 only (CPI-637) or dual inhibitors of both BET and CBP/P300 proteins (NEO1132 and NEO2437) was investigated in four UPS cell lines and two patient-derived xenografts (PDX) established at Institut Bergonié (Bordeaux, France). A CRISPR-KO screen was used to identify genes that mediate resistance to NEO2734. Results: We found that the 3 compounds act mainly by inhibiting the cell cycle with a concomitant reduction of MYC expression. NEO2734 was the most potent one with IC50 range between 0.2 to 1 µM in vitro and an antitumor activity in vivo in the two PDX models of UPS. To investigate the mechanism of action, we performed a transcriptomic analysis by RNA-sequencing. Gene set enrichment analysis revealed that NEO2734 induced a downregulation of G2M checkpoint and E2F targets hallmarks. We confirmed these results at the protein level showing a downregulation of several proteins involved in these pathways such as PLK1, AURKA or CCNB1. Interestingly, two of our four UPS cells lines were resistant to NEO2734 (IC50 27µM and 73µM). To elucidate the genetic factors involved in this resistance, we performed a CRISPR-KO screen. We identified more than 200 genes as potential candidates involved in drug resistance and decided to focus on hnRNPU which is an interactor of P300 and play a role in cell cycle regulation through th