Abstract 3986: Combination of tamoxifen and doxorubicin targets estrogen receptor beta-mutant p53-p73 axis: A novel therapeutic strategy for triple negative breast cancer
Authors: Das Gokul M. , Oturkar Chetan C. , Adams Christina , Park Jung H. , Dolan Melissa , Mastri Michalis , Oshi Manasori , Tokumaru Yoshihisa , Mukhopadhyay Utpal K. , Abha Kalyani , Jung Kwang H. , Yang Sukjin , Kim Suna , Ebos John , Takabe Kazuaki , Kaipparettu Benny A. 
Published: June 2022

Abstract
Triple-negative breast cancer (TNBC) is an aggressive subtype without any effective targeted therapies and rapidly become resistant to generic chemotherapy. Therefore, there is an urgent need to develop new therapeutic strategies. Estrogen receptor beta (ER beta) levels are high in about 60-70% of TNBCs. Recent reports including the Cancer Genome Atlas (TCGA) show that about 80% of TNBC express mutant p53 and it is the most predominant driver in these cancers. We have previously reported (JNCI, 2019, 111:1202-1215) that ER-beta binds p53 and exerts proliferative versus anti-proliferative/tumor suppressive functions depending on the wild type and mutant p53 status in TNBC, respectively. In the current work we used multiple approaches such as immunoprecipitation, in situ proximity ligation assay (PLA), and gene expression analysis by quantitative real-time PCR (qRT-PCR) to show that tamoxifen (Tam) increases ER beta-p53 interaction resulting in decrease of mutant p53 binding to p73 leading to cell cycle arrest, increased apoptosis, decreased proliferation, and increased expression of anti-proliferation genes. Importantly, ER beta antagonist PHTPP decreases the ER beta-p53 interaction in TNBC cells, whereas ER beta agonist DPN did not have any effect. Importantly, Tam synergized with doxorubicin (Dox) to decrease the IC50 of Dox more than 3-fold. This synergism was absent in an isogenic cell line where TP53 gene was knocked out. The fact that mutant p53 expression was necessary for Tam to synergize with Dox, along with our observation that upregulation of anti-proliferation gene expression was dependent on both ER beta and p73, strongly suggests that ER beta-mutant p53-p73 axis is the target of the novel effect of Tam. RNA-seq and reverse phase protein array (RPPA) analysis of isogenic TNBC cells differing in p53 mutational status without and with ER beta depletion revealed important cellular pathways impacted by the synergistic effect of Tam plus Dox combination treatment. To test the effect of Tam plus Dox combination therapy in vivo, we used isogenic MDA-MB-231 cell line-derived xenograft (CDX) and TNBC patient-derived xenograft (PDX) tumors. Consistent with our observations in the TNBC cell models, combination therapy inhibited progression of both CDX and PDX tumors more effectively compared to monotherapies. Furthermore, the antitumor effect was dependent on expression of mutant p53 in tumors. Our study has revealed a novel ER beta-mutant p53-p73 axis that could be targeted by Tam in combination with chemotherapy, raising the possibility of repurposing Tam to treat molecularly stratified TNBC that expresses both ERβ and mutant p53. Besides the potential for relatively faster entry of a safe and less expensive therapy to the clinic, our discovery can be exploited to reduce toxic adverse effects by reducing the dose of Dox in treatment regimens.
Citation Format: Gokul M. Das, Chetan C. Oturkar, Christina Adams, Jung H. Park, Melissa Dolan, Michalis Mastri, Manasori Oshi, Yoshihisa Tokumaru, Utpal K. Mukhopadhyay, Kalyani Abha, Kwang H. Jung, Sukjin Yang, Suna Kim, John Ebos, Kazuaki Takabe, Benny A. Kaipparettu. Combination of tamoxifen and doxorubicin targets estrogen receptor beta-mutant p53-p73 axis: A novel therapeutic strategy for triple negative breast cancer [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 3986.