Article ; Online: Preomic profile of BxPC-3 cells after treatment with BRC4
Journal of Proteomics. 2023 Sept., v. 288 p.104983-
2023
Abstract: BRCA2 and RAD51 are two proteins that play a central role in homologous recombination (HR) and DNA double strand break (DSB) repair. BRCA2 assists RAD51 fibrillation and defibrillation through binding with its eight BRC repeats, with BRC4 being one of ... ...
| Abstract | BRCA2 and RAD51 are two proteins that play a central role in homologous recombination (HR) and DNA double strand break (DSB) repair. BRCA2 assists RAD51 fibrillation and defibrillation through binding with its eight BRC repeats, with BRC4 being one of the most efficient and best characterized. RAD51 inactivation by small molecules has been proposed as a strategy to impair BRCA2/RAD51 binding and, ultimately, the HR pathway, with the aim of making cancer cells more sensitive to PARP inhibitors (PARPi). This strategy, which mimics a synthetic lethality (SL) approach, has been successfully performed in vitro by using the myristoylated derivative of BRC4 (myr-BRC4), designed for a more efficient cell entry. The present study applies a method to obtain a proteomic fingerprint after cellular treatment with the myr-BRC4 peptide using a mass spectroscopy (MS) proteomic approach. (Data are available via ProteomeXchange with identifier PXD042696.) We performed a comparative proteomic profiling of the myr-BRC4 treated vs. untreated BxPC-3 pancreatic cancer cells and evaluated the differential expression of proteins. Among the identified proteins, we focused our attention on proteins shared by both the RAD51 and the BRCA2 interactomes, and on those whose reduction showed high statistical significance. Three downregulated proteins were identified (FANCI, FANCD2, and RPA3), and protein downregulation was confirmed through immunoblotting analysis, validating the MS approach. Our results suggest that, being a direct consequence of myr-BRC4 treatment, the detection of FANCD2, FANCI, and RPA3 downregulation could be used as an indicator for monitoring HR impairment. RAD51's inhibition has gained increasing attention because of its possible implications in personalized medicine through the SL approach. Chemical disruption of protein-protein interactions (PPIs) between RAD51 and BRCA2, or some of its partner proteins, could potentiate PARPi DNA damage-induced cell death. This could have application for difficult to treat cancers, such as BRCA-competent and olaparib (PARPi) resistant pancreatic adenocarcinoma. Despite RAD51 being a widely studied target, researchers still lack detailed mechanistic information. This has stifled progress in the field with only a few RAD51 inhibitors having been identified, none of which have gained regulatory approval. Nevertheless, the peptide BRC4 is one of the most specific and best characterized RAD51 binder and inhibitor reported to date. Our study is the first to report the proteomic fingerprint consequent to cellular treatment of myr-BRC4, to offer a reference for the discovery of specific protein/pathway alterations within DNA damage repair. Our results suggest that, being a direct consequence of myr-BRC4 treatment, and ultimately ofBRCA2/RAD51 disruption, the detection of FANCD2, FANCI, and RPA3 downregulation could be used as an indicator for monitoring DNA damage repair impairment and therefore be used to potentiate the development of new effective therapeutic strategies. |
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| Keywords | DNA ; DNA repair ; adenocarcinoma ; cell death ; death ; gene expression regulation ; homologous recombination ; immunoblotting ; mass spectrometry ; pancreatic neoplasms ; peptides ; precision medicine ; proteomics ; tumor suppressor proteins ; RAD51 ; RAD51/BRCA2 disruptors ; Synthetic lethality |
| Language | English |
| Dates of publication | 2023-09 |
| Publishing place | Elsevier B.V. |
| Document type | Article ; Online |
| ZDB-ID | 2400835-7 |
| ISSN | 1876-7737 ; 1874-3919 |
| ISSN (online) | 1876-7737 |
| ISSN | 1874-3919 |
| DOI | 10.1016/j.jprot.2023.104983 |
| Database | NAL-Catalogue (AGRICOLA) |
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