探讨BRCA缺乏的三阴性乳腺癌对复制应激的敏感性

I. Tabet, Esin Orhan, C. Velázquez, Lise Fenou, C. Theillet
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摘要

在三阴性乳腺癌(TNBC)中,化疗是唯一的全身治疗,持续缓解是罕见的。我们建议拓宽治疗选择。约30%的TNBC肿瘤存在BRCA1缺陷,表现为DNA修复缺陷和对基因毒性药物的敏感性增加。我们假设缺乏brca的TNBC对复制应激诱导药物高度敏感,从而开辟了新的治疗前景。我们的初步结果显示,BRCA1缺失的TNBC细胞系和CRISPR/Cas9 BRCA1 KO等基因模型对吉西他滨的敏感性增加。吉西他滨处理的brca1缺陷细胞的细胞周期分布以复制突变中细胞数量增加引起的亚g1分数升高为特征。80%的BRCA1缺陷细胞在没有RPA32共染色的情况下显示持续(48-72小时)gH2AX染色,而在等基因BRCA1 WT模型中,gH2AX和RPA32阳性细胞数量在24小时开始下降。有趣的是,我们注意到,除了复制突变外,吉西他滨处理的brca缺陷细胞还发生了异常的有丝分裂,微核明显增加。有趣的是,体内实验似乎可以复制体外数据。事实上,BRCA超甲基化的TNBC PDX对吉西他滨的敏感性高于BRCA1 WT。总之,我们的数据表明BRCA缺陷肿瘤对复制毒性吉西他滨更敏感。此外,这种敏感性似乎是由没有得到很好管理的复制应激反应所介导的。在吉西他滨治疗后,细胞遭受重要的DNA损伤,导致复制分叉停滞和DNA断裂。在BRCA1缺失的情况下,HR通路受损,导致分叉塌陷和单链DNA积累,从而耗尽细胞内的RPA库,诱发复制灾难。除了复制缺陷外,吉西他滨治疗brca缺陷细胞,但不是brca精通细胞,遭受有丝分裂灾难。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
Exploring sensitivity to replicative stress in BRCA deficient Triple Negative Breast Cancer
In Triple Negative Breast Cancer (TNBC), chemotherapy is the only systemic treatment and sustained remissions are rare. We propose to widen therapeutic options. About 30% TNBC tumors are BRCA1 deficient, presentoing defective DNA repair and increased sensitivity to genotoxic drugs. We hypothesized that BRCA-deficient TNBC are highly sensitive to replication stress inducing drugs, thus, opening new therapeutic perspectives. Our preliminary results shown that BRCA1-deficient TNBC cell lines and a CRISPR/Cas9 BRCA1 KO isogenic model display increased sensitivity to gemcitabine. Cell cycle distribution of gemcitabine treated BRCA1-deficient cells were characterized by an elevated Sub-G1 fraction caused by increased numbers of cells in replication catastrophe. This was illustrated by 80% of BRCA1-deficient cells showing persistent (48-72h post treatment) gH2AX staining in absence of RPA32 co-staining, whereas in the isogenic BRCA1 WT model gH2AX and RPA32 positive cell numbers started decreasing at 24h. Interestingly, we noted that in addition to replication catastrophe, BRCA-deficient cells treated with gemcitabine underwent aberrant mitosis as shown by a clear increase of micro-nuclei. Interestingly, in vivo experiments appear to reproduce in vitro data. Indeed, a BRCA hyper methylated TNBC PDX, showed a higher sensitivity to gemcitabine than the BRCA1 WT. In conclusion, our data suggest that BRCA-deficient tumors are more sensitive to the replication poison Gemcitabine. Furthermore, this sensitivity seems to be mediated by an accentuated replicative stress response that is not well managed. Upon gemcitabine treatment, the cells undergo important DNA damage that leads to stalled replication forks, and DNA breakage. In the absence of BRCA1, the HR pathway is compromised, which leads to fork collapse and accumulation of single stranded DNA, therefore exhausting the pool of RPA within the cell and inducing Replicative catastrophe. In addition to deficient replication gemcitabine treated BRCA-deficient, but not BRCA-proficient cells, are subjected to mitotic catastrophe.
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