Abstract PR-002: Activation of the integrated stress response by NXP800, an orally available, clinical-stage, investigational agent in ARID1A-mutated, platinum resistant ovarian cancer
Authors: Powers Marissa V. , Swap Swee Y. , te Poele Robert , Lampraki Eirini-Maria , Roe Toby , Pellegrino Loredana , Taskinen Maria , Eccles Suzenne , Raynaud Florence , Cheeseman Matthew , Jones Keith , Clarke Paul A. , Workman Paul 
Published: March 2024

Abstract
Using innovative phenotypic screening, targeting the Heat Shock Factor 1 (HSF1) pathway, followed by multiparameter medicinal chemistry optimization, we discovered NXP800, an orally active, potent inhibitor of cell proliferation. Evaluation in a mini-panel of human cancer cell lines and tumor xenografts revealed high sensitivity in ARIDIA-deficient human ovarian cancer models, confirmed in the large Sanger panel and isogenic systems. By RNAseq we identified overlapping gene expression changes in human cancer cell lines exposed to NXP800, including expected changes in HSF1-regulated genes plus alterations in ATF4-regulated gene expression associated with activation of the integrated stress response (ISR). This did not indicate a global stress response to NXP800 as we saw no activation of the unfolded protein response.
Consistent with activation of the ISR, NXP800 induced phosphorylation of EIF2A and increased expression of downstream ISR markers/effectors ATF4, CHAC1 and CHOP both in human ovarian cells in vitro and corresponding tumor xenograft models in vivo. Using an siRNA approach, we found that blocking the induction of ATF4 reduced the response of sensitive, ARID1A mutant SK-OV-3 human ovarian carcinoma cells to NXP800 treatment. Phosphorylation of EIF2A is tightly regulated by four stress-controlled kinases, GCN2, HRI, PKR and PERK. Using either systematic siRNA knockdown or inhibition by two small-molecule tool compounds from different chemotypes, we discovered that GCN2 alone was required for ISR activation by NXP800. Also, inactivation of GCN2 markedly reduced the antiproliferative activity of NXP800. Global phospho-proteome analysis demonstrated defined changes in response to NXP800 which were reversed on co-treatment with a GCN2 inhibitor. Furthermore, ISR induction inhibited HSF1 activation, confirming the mechanistic link between ISR activation and inhibition of HSF1-mediated transcription. In summary, we discovered the mechanistically novel drug NXP800 which acts on cancer cells to stimulate GCN2 and thereby activate the ISR pathway, leading to inhibition of cap-dependent protein translation. NXP800 shows highly promising activity in human ovarian cancer, including tumor regression of ARID1A-deficient ovarian cancer xenografts. Studies are currently underway to determine precisely how NXP800 stimulates GCN2 activity and the role of ARID1A deficiency. With Nuvectis Pharma, the Phase 1a dose escalation study is completing and the multicentre Phase 1b expansion cohort study in platinum-resistant ARID1A-mutated ovarian cancer is now initiated (NCT05226507) in collaboration with the GOG Foundation and the European Network of Gynecological Oncological Trial Group (ENGOT). FDA has issued a Fast Track designation to NXP800 in this setting.
Citation Format: Marissa V. Powers, Swee Y. Swap, Robert te Poele, Eirini-Maria Lampraki, Toby Roe, Loredana Pellegrino, Maria Taskinen, Suzenne Eccles, Florence Raynaud, Matthew Cheeseman, Keith Jones, Paul A. Clarke, Paul Workman. Activation of the integrated stress response by NXP800, an orally available, clinical-stage, investigational agent in ARID1A-mutated, platinum resistant ovarian cancer [abstract]. In: Proceedings of the AACR Special Conference on Ovarian Cancer; 2023 Oct 5-7; Boston, Massachusetts. Philadelphia (PA): AACR; Cancer Res 2024;84(5 Suppl_2):Abstract nr PR-002.