Abstract LB088: Targeting ATR enhances the antitumor efficacy of patritumab deruxtecan (HER3-DXd) in tamoxifen-resistant ER+ breast cancer cells by reprogramming cell cycle progression
Authors: Xie Xuemei , Lee Jangsoon , Fuson Jon A. , Liu Huey , Gi Young Jin , Fan Pang-Dian , Koyama Kumiko , Tripathy Debu , Ueno Naoto T. 
Published: June 2022

Abstract
Background: HER3, a member of the ErbB family of receptor tyrosine kinases that activate multiple oncogenic signaling pathways, is overexpressed in 50-70% of breast cancers (BC). HER3 mRNA levels are higher in ER+ breast tumors than in other molecular subtypes. Inhibition of ER activity using an antagonist increased HER3 protein expression and activation, which was essential for growth and survival of ER+ BC cells resistant to the ER antagonist. Therefore, therapeutically targeting HER3 with HER3-DXd, an antibody-drug conjugate (ADC) composed of a fully human anti-HER3 monoclonal antibody (patritumab) conjugated to a topoisomerase I inhibitor payload (an exatecan derivative) via a tetrapeptide-based cleavable linker, can be an effective treatment for tamoxifen-resistant (TMR) ER+ BC. After first assessing HER3-DXd as a single agent, we sought to identify a synergistic partner to maximize HER3-DXd’s antitumor activity in HER3+/ER+ TMR BC.
Methods: Whole-genome high-throughput siRNA screening was performed to identify targets whose inhibition enhances HER3-DXd’s antitumor efficacy. The antitumor effects of HER3-DXd plus the synergistic partners were assessed using a soft agar colony formation assay and a clonogenic assay in TMR HER3+/ER+ MCF7 and T47D BC cells. Treatment effects on cell cycle distribution, apoptosis, and expression of proteins that regulate cell cycle progression were assessed by flow cytometry, annexin V-PE and 7-AAD staining, and Western blotting, respectively.
Results: HER3-DXd inhibited the anchorage-independent growth of HER3+/ER+ cells by &gt;50% at 5 nM and their colony formation at 5-25 nM (P&lt;0.05). To maximize HER3-DXd’s antitumor efficacy, we performed high-throughput siRNA screening and identified ATR, CD247, RAB7A, UPK3A, ROCK2, SLC29A1, and WNT7A as potential synergistic targets. Among these targets, inhibiting ATR with siRNA or BAY1895344 showed the most synergistic effect with HER3-DXd in HER3+/ER+ BC cells. In contrast, no synergistic effect was observed with the combination of BAY1895344 plus patritumab or control ADC (IgG-DXd), suggesting its dependence on HER3-DXd-mediated delivery of DXd. The combination of HER3-DXd plus BAY1895344 reprogrammed cell cycle progression from G2/M arrest to G1 arrest by inhibiting both ATR/Chk1/cyclin A2/CDK2 and ATR/Chk1/cyclin E/CDK2 signaling. The combination also reduced expression of H2AX, an ATR substrate that contributes to DNA repair, but increased that of γH2AX, indicating the induction of DNA damage. HER3-DXd and BAY1895344 synergistically inhibited growth of HER3+/ER+ BC cells by inducing apoptosis.
Conclusion: The combination of HER3-DXd plus ATR inhibitors has therapeutic potential for overcoming tamoxifen resistance in HER3+/ER+ BC. We are currently validating the synergy in endocrine-resistant ER+ BC xenograft models.
Citation Format: Xuemei Xie, Jangsoon Lee, Jon A. Fuson, Huey Liu, Young Jin Gi, Pang-Dian Fan, Kumiko Koyama, Debu Tripathy, Naoto T. Ueno. Targeting ATR enhances the antitumor efficacy of patritumab deruxtecan (HER3-DXd) in tamoxifen-resistant ER+ breast cancer cells by reprogramming cell cycle progression [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr LB088.