泛素特异性蛋白酶14与聚（ADP-核糖）糖水解酶共同抑制在BRCA1突变、聚（ADP-核糖）聚合酶抑制剂耐药三阴性乳腺癌细胞中的协同作用
Synergistic Effect of Ubiquitin-Specific Protease 14 and Poly(ADP-Ribose) Glycohydrolase Co-Inhibition in BRCA1-Mutant, Poly(ADP-Ribose) Polymerase Inhibitor-Resistant Triple-Negative Breast Cancer Cells.
作者信息
Li Pisong, Zhu Xiaoyu, Qu Hui, Han Zhongbin, Yao Xingyu, Wei Yuan, Li Baijun, Chen Hongshen
机构信息
Department of Breast and Thyroid Surgery, Affiliated Zhongshan Hospital of Dalian University, Dalian, 116001, People's Republic of China.
出版信息
Onco Targets Ther. 2024 Sep 6;17:741-753. doi: 10.2147/OTT.S463217. eCollection 2024.
PURPOSE
The clinical benefits of poly(ADP-ribose) polymerase (PARP) inhibitors are limited to triple-negative breast cancer (TNBC) with BRCA deficiency due to primary and acquired resistance. Thus, there is a pressing need to develop alternative treatment regimens to target BRCA-mutated TNBC tumors that are resistant to PARP inhibition. Similar to PARP, poly(ADP-ribose) glycohydrolase (PARG) plays a role in DNA replication and repair. However, there are conflicting reports on the vulnerability of BRCA1-deficient tumor cells to PARG inhibition. This study aims to investigate the synergistically lethal effect of the PARG inhibitor COH34 and the ubiquitin-specific protease (USP) 14 inhibitor IU1-248 and the underlying mechanisms in BRCA1-mutant, PARP inhibitor-resistant TNBC cells.
METHODS
The cytotoxicity of PARG inhibition alone or in combination with USP14 inhibition in the BRCA-mutant, PARP inhibitor-resistant TNBC cell lines, HCC1937 and SUM149PT, was analyzed using cell viability and proliferation assays and flow cytometry. The molecular mechanisms underlying the synergistic effects of IU1-248 and COH34 were evaluated by immunofluorescence staining, DNA repair reporter assays and Western blot analysis.
RESULTS
It was found that HCC1937 and SUM149PT cells exhibited moderate responsiveness to PARG inhibition alone. To the best of our knowledge, this research is the first to demonstrate that the combination of IU1-248 and COH34 produces synergistic effects against TNBC cells in the same setting. Mechanistically, the blockade of USP14 by IU1-248 was shown to increase DNA damage and promote error-prone non-homologous end joining (NHEJ), as evidenced by the accumulation of γH2AX and 53BP1 in the nucleus and the activation of a reporter assay. Additionally, it was demonstrated that the inhibition of NHEJ repair activity attenuates the synergistic effects of concomitant PARG and USP14 inhibition. IU1-248 promotes NHEJ repair through the downregulation of the expression of c-Myc.
CONCLUSION
USP14 inhibition may be a plausible strategy for expanding the utility of PARG inhibitors in TNBC in BRCA-mutant, PARP inhibitor-resistant settings.
目的
聚（ADP - 核糖）聚合酶（PARP）抑制剂的临床益处仅限于因原发性和获得性耐药而存在BRCA缺陷的三阴性乳腺癌（TNBC）。因此，迫切需要开发替代治疗方案，以靶向对PARP抑制耐药的BRCA突变TNBC肿瘤。与PARP类似，聚（ADP - 核糖）糖苷水解酶（PARG）在DNA复制和修复中起作用。然而，关于BRCA1缺陷肿瘤细胞对PARG抑制的敏感性存在相互矛盾的报道。本研究旨在探讨PARG抑制剂COH34和泛素特异性蛋白酶（USP）14抑制剂IU1 - 248在BRCA1突变、PARP抑制剂耐药的TNBC细胞中的协同致死作用及其潜在机制。
方法
使用细胞活力和增殖测定以及流式细胞术分析PARG抑制单独或与USP14抑制联合在BRCA突变、PARP抑制剂耐药的TNBC细胞系HCC1937和SUM149PT中的细胞毒性。通过免疫荧光染色、DNA修复报告基因测定和蛋白质印迹分析评估IU1 - 248和COH34协同作用的分子机制。
结果
发现HCC1937和SUM149PT细胞对单独的PARG抑制表现出中等反应性。据我们所知，本研究首次证明IU1 - 248和COH34的组合在相同环境下对TNBC细胞产生协同作用。从机制上讲，IU1 - 248对USP14 的阻断显示会增加DNA损伤并促进易出错的非同源末端连接（NHEJ），细胞核中γH2AX和53BP1的积累以及报告基因测定的激活证明了这一点。此外，还证明抑制NHEJ修复活性会减弱PARG和USP14联合抑制的协同作用。IU1 - 248通过下调c - Myc的表达促进NHEJ修复。
结论
在BRCA突变、PARP抑制剂耐药的情况下，抑制USP14可能是扩大PARG抑制剂在TNBC中应用的合理策略。
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