Mechanism of chemoresistance mediated by TGF-beta

描述(申请人提供)：许多化疗药物通过破坏DNA来对抗癌细胞。化疗耐药性是癌症治疗的主要临床障碍。癌症患者化疗耐药的机制尚不完全清楚，迫切需要确定控制药物反应的因素，并开发新的治疗方法来提高治疗效果。转化生长因子(TGF)是正常细胞中的一种肿瘤抑制因子，它的信号在癌症中被劫持，以促进疾病的进展。在乳腺癌中，转化生长因子β与不良的临床结果和化疗耐药有关，其机制仍然很不清楚。我们先前的研究表明，在乳腺癌细胞中，转化生长因子β诱导针对DNA损伤传感器ATM和MSH2的microRNAs(miR-21和miR-181)，因此可能调节癌症对基因毒性化疗的反应。本研究的目的是剖析转化生长因子-2介导的化疗耐药的分子机制，并探索提高药物疗效的潜在治疗方法。在目标1中，将利用已建立的分子和细胞生物学方法，在具有不同P53状态的乳腺癌细胞中，确定转化生长因子β对细胞对各种DNA损伤处理和对聚(ADP-核糖)聚合酶(PARP)抑制的反应的作用。受转化生长因子β调控的miRNAs和ATM/MSH2通路的作用将通过基因敲除和过度表达策略来确定。在目标2中，将检验这样一种假设，即增强Smad2/3与其RNA靶标的结合介导转化生长因子-βS在癌细胞中的功能转向诱导miRNA调节和化疗耐药。表达不同水平的Smad2/3辅因子(即Smad4、DROSHA和p68)的乳腺癌细胞将被检测其对Smad2/3功能和转化生长因子-2作用的动态调节。在目标3中，将在动物肿瘤模型中评估转化生长因子-β对化疗反应的影响以及前两个目标中确定的机制。将探索从治疗上抑制这种转化生长因子-β功能并提高治疗效果的新策略。这项研究将有助于更好地理解耐药和转化生长因子-β信号作为癌症治疗的标志物和靶点。虽然本文中确定的机制可能在了解癌症和确定治疗方法方面具有普遍应用，但我们的研究对临床侵袭性、难以治疗的基底细胞样癌(主要是三阴性)乳腺癌增加了意义，这些乳腺癌经常经历活跃的转化生长因子-β信号。这项研究将为Smad2/3介导的miRNA处理在肿瘤中转化生长因子-β的功能转换提供新的见解。了解转化生长因子-2介导的化疗耐药可能会揭示新的治疗靶点和策略，从而提高缺乏系统治疗靶点的癌症的化疗效果。我们的长期目标是在原发癌中验证这一机制，并建立标准的方法来确定适合于针对转化生长因子-S耐药效应的治疗的患者，并了解转化生长因子-β介导的miRNA失调在人类癌症中的全球影响。

DESCRIPTION (provided by applicant): Many chemotherapy drugs act against cancer cells by causing damage to the DNA. Resistance to chemotherapy is a major clinical obstacle in cancer treatment. The mechanisms of chemoresistance in cancer patients are not fully understood, leading to urgent needs for determining factors that control drug response and developing novel therapies to enhance the treatment efficacy. Signaling from transforming growth factor (TGF) ¿, a tumor suppressor in normal cells, is hijacked in cancer to promote disease progression. In breast cancer, TGF- ¿ is linked to poor clinical outcomes and chemoresistance through mechanisms that remain largely unknown. Our previous studies indicate that in breast cancer cells, TGF- ¿ induces microRNAs (miR-21 and miR-181) that target the DNA damage sensors ATM and MSH2, and may therefore regulate cancer response to genotoxic chemotherapy. The goals of this study are to dissect the molecular mechanism of TGF-2-mediated chemoresistance, and to explore potential therapies to enhance drug efficacy. In Aim 1, TGF- ¿ action on cell response to various DNA-damaging treatments and to inhibition of poly(ADP-ribose) polymerase (PARP) will be determined in breast cancer cells with different p53 status using established molecular and cellular biology assays. The role of the TGF- ¿ -regulated miRNAs and the ATM/MSH2 pathways will be determined using gene knockdown and overexpression strategies. In Aim 2, the hypothesis that enhanced SMAD2/3 binding to their RNA targets mediates TGF- ¿ 's functional shift in cancer cells towards inducing miRNA regulation and chemoresistance will be examined. Breast cancer cells expressing various levels of the SMAD2/3 cofactors (i.e., SMAD4, Drosha and p68) will be examined for their dynamic regulation of SMAD2/3 function and TGF-2 effect. In Aim 3, the effect of TGF- ¿ on chemotherapy response and the mechanism identified in the first two aims will be evaluated in animal tumor models. Novel strategies to therapeutically suppress this TGF- ¿ function and enhance the treatment efficacy will be explored. This study will enable better understandings of drug resistance and of TGF- ¿ signaling as both a marker and a target in cancer treatment. Although the mechanism identified herein may have a general application to understanding cancer and defining treatments, our study has added significance for clinically aggressive, hard-to-treat basal- like (mainly triple-negative) breast cancer that often experience active TGF- ¿ signaling. This study will provide novel insight into the functional switch of TGF- ¿ in cancer via SMAD2/3-mediated miRNA processing. Understanding TGF-2-mediated chemoresistance may reveal novel therapeutic targets and strategies that will enhance the chemotherapy efficacy for cancers that lack targets for systemic treatments. Our long-term objectives are to validate this mechanism in primary cancers and establish standard approaches to identify patients suitable for therapies targeting TGF- ¿ 's drug resistant effect, and to understand the global effect of TGF- ¿ -mediated miRNA dysregulation in human cancer.