Discovery of the Anti-Angiogenesis Effect of Eltrombopag in Breast Cancer Through Targeting of HuR Protein

Abstract

Background: HuR, an mRNA-binding protein responsible for poor prognosis in nearly all kinds of malignancies, is a potential anti-tumor target for drug development. Eltrombopag is a medication for chronic immune thrombocytopenia and also shows effectiveness in managing thrombocytopenia associated with cancer chemotherapy. However, the mechanism through which eltrombopag inhibits tumor via targeting HuR is poorly understood. Methods: Eltrombopag was screened out from a fluorescence polarization-based screening system. The binding activity of eltrombopag was analyzed by FP, EMSA and simulation docking. In vitro and in vivo anti-tumor effect of eltrombopag was detected with MTT, tumor allograft experiment and immunohistochemistry. The HuR-dependent mechanism of eltrombopag for anti-angiogenesis effect was studied by qRT-PCR, RNA stability assays, luciferase assays, RNA immunoprecipitation, cell scratch assay and in vitro Matrigel angiogenesis assay. Findings: We showed that eltrombopag could disrupt HuR binding to its target ARE sequence as a RNA anologue, and exhibit anti-tumor effect in vitro and in vivo. The in vivo data showed eltrombopag was efficient in reducing microvessels in tumor tissues, which was subsequently explained by results that eltrombopag could reduce angiogenesis-related factors in HuR-dependent manner in both 4T1 cells and RAW 264·7 macrophages, and results that in vitro anti-angiogenesis effect of eltrombopag mediated by macrophages. Interpretation: This study reveals the HuR-dependent anti-angiogenesis effect of eltrombopag in tumors, suggesting that the existing drug eltrombopag may be used as an anti-cancer drug. Funding: This work was supported by National Natural Science Foundation of China (81573579), the New Interdisciplinary Subject Funding Program for Shanghai Traditional Chinese Medicine (E2-F18003), Shanghai Municipal Education Commission (2019-01-07-00-10-E00072), Science and Technology Commission of Shanghai Municipality (18401933500), Postdoctoral Science Foundation of China (A2-X1802408) and Shanghai Municipal Commission of Health and Family Planning (2018YQ003). Declaration of Interest: The authors declare that they have no competing interests. Ethical Approval: All animal protocols and procedures were approved by the Shanghai University of Traditional Chinese Medicine Institutional Animal Care and Use Committees and were carried out in accordance with the guidelines outlined in the Guide for the Care and Use of Laboratory Animals published by National Institute of Health. All experiments were carried out in compliance with ARRIVE guidelines.