TGF-β介导乳腺癌化疗耐药及PARP抑制剂逆转耐药的现象及其机制的研究

Transforming growth factor beta (TGF-β) proteins are multitasking cytokines, in which high levels at tumor sites generally correlate with poor prognosis in human patients with cancer. Previously, we first report that TGF-β downregulates the expression of ataxia telangiectasia–mutated (ATM), mutS homolog 2 (MSH2), and breast cancer 1 early onset (BRCA1) of DNA damage repair gene in breast cancer cells through upregulating the miR-21 and miR-181. TGF-β induces resistance to DNA-damaging agents and sensitivity to poly (ADP-ribose) polymerase (PARP) inhibitor in breast cancer cells. Xenograft tumors with active TGF-β signaling exhibited resistance to the DNA-damaging agent doxorubicin but increased sensitivity to the PARP inhibitor ABT-888. Combination of doxorubicin with ABT-888 significantly improved the treatment efficacy in TGF-β-active tumors. We therefore hypothesize that TGF-β induces resistance to genotoxic chemotherapy by disrupting DNA damage sensing, an effect mediated by downregulation of MSH2, ATM and BRCA1 via miRNAs whose maturation by SMAD2/3 is dysregulated in cancer. Thus, TGF-β intervention, miR-21/miR-181 depletion and PARP inhibition could be effective therapeutic approaches for cancer exhibiting TGF-β activation. In this project, we future investigate the mechanism of chemoresistance and PARP inhibitor reversal chemoresistance through DNA-repair gene regulated by TGF-β. Our future goals are to validate this mechanism in primary breast cancer and establish standard approaches to identify patients suitable for therapies targeting TGF-β-mediated chemoresistance, and to understand the global effect of TGF-β-mediated dysregulation of miRNA processing in cancer.

TGF-β是一种有多功能的细胞因子，肿瘤中高表达TGF-β的患者通常预后不良。前期工作中，我们首先报道乳腺癌中TGF-β能通过非经典的Smad2/3-Drosha/p68途径上调miR-21和miR-181，它们能下调MSH2、ATM、BRCA1 DNA修复基因；TGF-β能提高乳腺癌对DNA损伤剂的耐药性，但可增强乳腺癌对PARP抑制剂的敏感性。由此推测：TGF-β通过非经典Smad2/3- Drosha/p68-miRNAs途径下调多种DNA修复基因来介导乳腺癌化疗耐药；在“协同致死”效应作用下，能使PARP抑制剂逆转化疗耐药。本课题是在前期工作基础上，进一步深入探讨TGF-β通过非经典途径调控DNA修复基因介导化疗耐药及PARP抑制剂逆转耐药的现象及其机制；体内验证靶向拮抗miR-21和miR-181逆转肿瘤耐药的效应。本课题为寻找肿瘤新的靶向治疗的靶点及药物提供理论基础和临床依据。

研究背景：TGF-β是一种有多功能的细胞因子，肿瘤中高表达TGF-β的患者通常预后不良。前期工作中，我们首先报道乳腺癌中TGF-β能通过非经典的Smad2/3-Drosha/p68途径上调miR-21和miR-181，它们能下调MSH2、ATM、BRCA1等DNA修复基因；TGF-β能提高乳腺癌对DNA损伤剂的耐药性，但可增强乳腺癌对PARP抑制剂的敏感性。由此推测：TGF-β通过非经典Smad2/3- Drosha/p68-miRNAs途径下调多种DNA修复基因来介导乳腺癌化疗耐药；在“协同致死”效应作用下，能使PARP抑制剂逆转化疗耐药。本课题是在前期工作基础上，进一步深入探讨TGF-β通过非经典途径调控DNA修复基因介导化疗耐药及PARP抑制剂逆转耐药的现象及其机制；体内验证靶向拮抗miR-21和miR-181逆转肿瘤耐药的效应。本课题为寻找肿瘤新的靶向治疗的靶点及药物提供理论基础和临床依据。. 主要研究内容：1）研究TGF-β介导化疗耐药和PARP抑制剂逆转耐药的现象及机制；2）采用pHIV7-p(CAGA)12-Fluc/Rluc-(miR-21/miR-181)双报告基团载体，来研究肿瘤中TGF-β调控miRNAs的机制；3）建立小鼠乳腺癌移植瘤模型，体内验证上述机制和结论；4）利用临床标本进一步验证上述结论。. 重要结果：1）通过体外实验证明肿瘤中TGF-β通过非经典Smad2/3-Drosha/p68-miRNAs途径促进miRNAs成熟（如miR-21和miR-181），miRNAs能下调其靶基因BRCA1、ATM、MSH2，这些DNA损伤修复基因表达量明显降低，DNA损伤修复能力受损，导致肿瘤细胞对DNA损伤剂耐药；并阐明由于“协同致死”效应，PARP酶抑制剂能逆转DNA损伤剂的耐药性。2）通过小鼠实验，证明体内PARP抑制剂联合DNA损伤剂可逆转化疗耐药，证明靶向拮抗miR-21和miR-181可逆转肿瘤耐药，有明显抗肿瘤作用。. 科学意义：首次证明高表达TGF-β的乳腺癌患者可优先选择PARP抑制剂进行个体化治疗；PARP抑制剂联合化疗，PARP抑制剂能逆转化疗耐药，抗肿瘤效果更好。