Abstract 1629: Targeting HSBP1 Cys137 residue by 4-Iodobutyl acetate selectively kills triple-negative breast cancer

Abstract Background: Triple-negative breast cancer (TNBC) accounts for 15-20% of breast cancers and has a higher mortality rate because of metastatic character as compared to the non-TNBC subtypes. Chemotherapy is a non-selective anti-cancer treatment and currently remains as a first-line regimen for TNBC patients. As a result, identifying a more effective and selective drug target is urgently needed for TNBC. The aim of this study was made to discover the iodoacetamide derivative-targetable cysteine (Cys) residues of protein which is involved in the specific protein-protein interactions (PPIs) that are critical for TNBC survival. Methods: MTT assay was used to discover the iodoacetamide derivative that has a cytotoxic effectiveness on TNBC cell line HCC1806 better than normal mammary cell line MCF10A. Mass spectrometric analysis was utilized to identify the iodoacetamide derivative-targetable proteins and Cys residues. Site-directed mutagenesis was employed to validate the specific incorporation of iodoacetamide derivative candidate into the identified protein Cys residue. Cell-based assays and mouse models were established to determine the anti-TNBC effectiveness of iodoacetamide derivative candidate. Pulldown assay was performed to determine the specific PPIs of iodoacetamide derivative candidate-targeted protein in TNBC cells. Results: MTT assays revealed that 4-Iodobutyl Acetate (IBA) is able to selectively kills HCC1806 cells with half maximal inhibitory concentration (IC50) at 0.01 µM which is much lower than that of MCF10A cells (IC50 = 7.91 µM). Mass spectrometric analysis indicated that IBA incorporates into the Cys137 residue of HSPB1 protein and thereby induces cell apoptosis in HCC1806 cells. Either the removal of iodo-group from IBA or the mutation of Cys137 to Ser significantly abolished its cytotoxic effects on HCC1806 cells both in vitro and in vivo. Pulldown assay demonstrated that the incorporation of IBA into Cys 137 leads to the disruption of HSPB1 PPIs with Akt, Cytochrome c and Caspase-3, which ultimately results in the loss of metastatic ability and induction of cell apoptosis in HCC1806 cells. Conclusion: Our results provide a novel, selective and even more safe therapeutic strategy for combating TNBC via targeting the pocket surrounding the Cys137 of HSPB1 protein. Citation Format: Ruei-Chen Hung, Chiou-Feng Lin, Yuan-Feng Lin. Targeting HSBP1 Cys137 residue by 4-Iodobutyl acetate selectively kills triple-negative breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 1629.