Abstract P3-10-12: Targeting a common drug compensation pathway using NUPR1 inhibition in triple negative breast cancer
Authors: Solzak Jeffrey P , Wang Chao , Hancock Brad , Radovich Milan 
Published: March 2020

Abstract
Introduction: Triple negative breast cancer (TNBC) is a devastating disease responsible for a higher rate of morbidity and mortality compared to all other breast cancer subtypes. With few effective treatments, many have moved towards combination therapies. Using a genomically driven methodology, we show that Nuclear Protein 1 (NUPR1) is upregulated and activated in response to several targeted therapies and to standard chemotherapy. In this work, we show that targeting NUPR1 in conjunction with targeted therapies or paclitaxel, results in synergistic combinations.
Methods: To evaluate a cohort of individuals with TNBC, 58 tumors were analyzed pre- versus post-chemotherapy using Ingenuity Pathway Analysis (IPA, Qiagen) after whole transcriptome sequencing. Six TNBC cell lines were utilized for in vitro work to reflect the heterogeneity of disease presentation. Four targeted therapies (Dasatinib, Erlotinib, Everolimus, and Trametinib) were chosen based on prior data suggesting activation of SRC, EGFR, mTOR, and MEK1/2 in TNBC respectively. Paclitaxel was used as the standard chemotherapy. IC50s were calculated using standard log dose escalations and Prism 7 (GraphPad). Next, whole transcriptome sequencing and pathway analysis using IPA was performed on treated cells at their designated IC50s for each drug and compared to vehicle controls. Trifluoperazine (TFP, a small molecule inhibitor of NUPR1) combined with targeted therapies and paclitaxel experiments were completed on TNBC cell lines and CI values were determined using the Chou-Talalay method. To observe the effects of NUPR1 inhibition, a NUPR1 siRNA was utilized in combination with paclitaxel at increasing doses with mock transfected cells for control.
Results: We observed NUPR1 activation and RNA overexpression in all six cell lines treated with four targeted therapies. Combining the RNA sequencing analysis of these cell lines and focusing on common responses, NUPR1 was in the top three responses across all treatments. Our analysis of TNBC patient tumors (N=58) before and after chemotherapy resulted in observing an activation of NUPR1 (p=5.32E-10). In experiments on TFP combined with targeted therapies or paclitaxel, we observed CI values across cell lines ranging from .45-.90 denoting synergy. Experiments performed that knocked down NUPR1 with a siRNA displayed similar significant synergy with paclitaxel. Further, targeted therapy experiments where TFP was kept at its respective IC50 for each cell line, we saw a significant decrease in the IC50 of the primary therapeutic.
Conclusion: Heterogeneous cell lines treated with targeted therapies as well as patients treated with chemotherapy demonstrate upregulation of the somatic pathway governing NUPR1. Targeting NUPR1’s ability to communicate using TFP in combination with targeted therapies or paclitaxel increased the potency of both treatment strategies. Further in vivo experiments testing these combinations, utilizing a precision medicine philosophy, are an obligate next step.
Citation Format: Jeffrey P Solzak, Chao Wang, Brad Hancock, Milan Radovich. Targeting a common drug compensation pathway using NUPR1 inhibition in triple negative breast cancer [abstract]. In: Proceedings of the 2019 San Antonio Breast Cancer Symposium; 2019 Dec 10-14; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2020;80(4 Suppl):Abstract nr P3-10-12.