Identification of (S)-10-Hydroxycamptothecin as a potent BRD4 inhibitor for treating triple-negative breast cancer

Objective: The purpose of this study was to find a new potent BRD4 inhibitor for the treatment of TNBC. Methods: HTRF based assay was applied to identify BRD4 inhibitors. Then, molecular docking was used to characterize the potential binding mechanism of the compound. After that, dialysis assay and ultra-centrifuge assay were applied to test the reversible binding of 10-HCPT on BRD4(BD1) and BRD4(BD1), respectively. CESTA was used to test the binding of 10-HCPT to BRD4 in cellular level. Finally, CCK8 assay, flow cytometry, western blotting and xenograft model bearing MDA-MB-231 cells were utilized to test the activity and mechanism of 10-HCPT on MDA-MB-231 cells in vitro and in vivo. Results: 10-HCPT was identified to be a strong inhibitor of BRD4 with IC50 = 928.1 nM against the BRD4(BD1) recombinant and IC50 = 865.8 nM versus the BRD4(BD2) recombinant, adopting a drug repositioning approach. Moreover, we found that 10-HCPT can inhibit BRD4 reversibly with the aid of dilution assay and dialysis experiments. At the cellular level, 10-HCPT binds to BRD4 and inhibits the growth of MDA-MB-231, a triple-negative breast cancer (TNBC) cell line that is partly dependent on the level of BRD4. Meanwhile, 10-HCPT can induce apoptosis as well as the decreasing amount of C-MYC, a substrate of BRD4, Ki-67, BCL-XL, and BCl-2, as well as induce the cleavage of caspase 3. In vivo xenograft model bearing MDA-MB-231 cells indicated that 10-HCPT can strongly inhibit MDA-MB-231 proliferation in BRD4 partially dependent behavior with minimal toxicity. Conclusion: BRD4 may be a new potential target for 10-HCPT and provide new insight into the future 10-HCPT-based TNBC therapies. (C) 2022 Elsevier B.V. All rights reserved.

Automated summary

In this study, a potent BRD4 inhibitor, 10-HCPT, was identified for the treatment of triple-negative breast cancer (TNBC). The results showed that 10-HCPT can inhibit the growth of TNBC cells by binding to BRD4 and inducing apoptosis. The study also provided new insights into the potential use of 10-HCPT as a therapy for TNBC.