Abstract 5213: Drug repurposing to treat triple negative breast cancer (TNBC) based on a novel ER beta-p53-p73 signaling axis
Authors: Oturkar Chetan C. , Adams Christina , Mukhopadhyay Utpal K. , Caradori Alexander , Oshi Manasori , Tokumaru Yoshihisa , Creighton Chad J. , Park Jun H. , Gandhi Nishant , Takabe Kazuaki , Attwood Kristopher M. , Kaipparettu Benny A. , Das Gokul M. 
Published: September 2020

Abstract
That endocrine therapy or HER2 targeted therapies are not effective against TNBC makes it the most difficult breast cancer subtype to treat. Although these tumors initially respond to chemotherapy, there is high risk of relapse. Therefore, there is urgent need to develop new therapeutic strategies against these aggressive cancers. Reports including the Cancer Genome Atlas (TCGA) show that about 80% of TNBC express mutant p53 and it is a major driver of these cancers. Furthermore, 60-80% of TNBC express estrogen receptor-beta (ERβ). We have shown that tumor suppressor protein p53 status (wild type versus mutant) is a determinant of pro-versus anti-tumorigenic roles of ERβin breast cancer (JNCI, 2019, 111:1202-1215). These observations, along with our novel discovery that Tamoxifen (Tam) increases interaction between ERβ and mutant p53 leading to decreased binding of mutant p53 to tumor suppressor p73 resulting in reactivation of p73 in TNBC cells, led us to hypothesize that Tam can be repurposed to treat TNBC stratified based on combined expression of mutant p53 and ERβ. Experimental procedures to test this hypothesis include use of cell culture, TNBC cell line-derived xenograft (CDX), and patient tumor-derived organoid (PDO) models, proximity ligation assay (PLA), co-immunoprecipitation (Co-IP), quantitative chromatin immunoprecipitation (qChIP), quantitative real time PCR (qPCR), RNAi-mediated knockdown, gene knockout with CRISPR, RNA-seq, and bioinformatics data analysis. Our data show that Tam when combined with the widely used chemotherapeutic agent doxorubicin (Doxo) reduces several fold the IC50 of Doxo in TNBC cells. Importantly, Tam was unable to decrease the IC50 of Doxo in isogenic TNBC cells where mutant p53 was knocked out demonstrating that the effect of Tam mediated by ERβ and p73 is directed against the oncogenic gain-of-functions of mutant p53. Consistent with this, the combination treatment elicited robust antitumor effect on CDX tumors in vivo. The drug combination also increases apoptosis as a consequence of restored p73 activity. Conversely, PHTPP (an ERβ-specific antagonist) disrupts the ERβ-mutant p53 interaction enabling mutant p53 to bind and inactivate p73. Our finding that Tam in combination with Doxo activates p73 leading to robust apoptotic response and inhibition of tumor growth suggest that Tam, a drug that is well tolerated with relatively less side effects, can be repurposed in combination with Doxo to treat TNBCs expressing mutant p53 and ERβ and this approach can reach the clinic relatively fast as both the drugs are FDA-approved and have been in the clinic for several years to treat other forms of cancer. Furthermore, based on our data that Tam decreases the IC50 of Doxo considerably, there is potential for using Doxo at a dose much lower than what is currently used in the management of TNBC, thereby reducing major side effects of Doxo.
Citation Format: Chetan C. Oturkar, Christina Adams, Utpal K. Mukhopadhyay, Alexander Caradori, Manasori Oshi, Yoshihisa Tokumaru, Chad J. Creighton, Jun H. Park, Nishant Gandhi, Kazuaki Takabe, Kristopher M. Attwood, Benny A. Kaipparettu, Gokul M. Das. Drug repurposing to treat triple negative breast cancer (TNBC) based on a novel ER beta-p53-p73 signaling axis [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 5213.