Targeting stromal reaction in pancreatic cancer to overcome chemo-resistance

描述(由申请人提供)： 美国癌症协会的最新估计显示，2010年有43,140例新发胰腺导管腺癌(PDAC)病例，36,800例死于PDAC。这种令人沮丧的预后是因为胰腺癌不仅具有侵袭性，而且是一种高度耐化疗的恶性肿瘤。具体地说，在原发肿瘤和远处转移瘤中都存在明显的纤维化反应，限制了化疗的效果和效果。PDAC肿瘤表现为胶原酶膜型1-基质金属蛋白酶(MT1-MMP1，MMP14)和高迁移率族A2(HMGA2)的表达增加，HMGA2是一种参与调节染色质结构的结构性DNA结合蛋白。因此，这项应用的目的是阐明纤维化反应导致化疗耐药的机制。我们的中心假设是，胶原微环境通过诱导常染色质的形成和促进DNA修复来促进化疗耐药。我们进一步假设，体内靶向MT1-MMPs不仅通过改变染色质结构，而且通过延缓纤维化进展本身，从而提高化疗敏感性。这一假说是基于我们的初步数据，即在3D胶原凝胶中生长的PDAC细胞增加了组蛋白H3的乙酰化，减少了异染色质蛋白11(HP11)的表达，并防止了吉西他滨诱导的检查点停滞。我们的初步数据还表明，胶原增加了PDAC细胞中HMGA2和MT1-MMP的表达，而siRNA降低了HMGA2或MT1-MMP的水平，从而减弱了胶原微环境中的吉西他滨抵抗。令人惊讶的是，在转基因和异种移植的胰腺癌小鼠模型中，人MT1-MMPs的表达矛盾地导致了与星状细胞激活相关的显著纤维化。目标1中提出的实验旨在确定3D胶原诱导常染色质促进化疗耐药的机制。具体地说，我们将研究HMGA2在增强3D胶原中常染色质和促进DNA修复方面的作用。Aim 2中提出的实验旨在验证这样的假设，即靶向MT1-MMP会通过诱导异染色质和减少DNA修复来增加化疗敏感性。MT1-MMP靶向靶点为功能封闭抗体，金纳米粒子为MT1-MMP特异性siRNA功能化靶点。Aim 3中的实验旨在验证这样一种假设，即靶向MT1-MMP除了增强化疗敏感性外，还将减轻纤维化。这种方法是创新的，因为它将利用胰腺癌的体外器官类型和体内原位和转基因小鼠模型，以及带有siRNA功能的金纳米颗粒，以证明靶向MT1-MMP使PDAC细胞对化疗药物的效果敏感。这项拟议的研究具有重要意义，因为它是连续研究的关键的第一步，该研究不仅将解决显着的纤维化反应如何导致化疗耐药，而且还将导致确定MT1-MMP作为提高PDAC肿瘤化疗反应的治疗靶点。

DESCRIPTION (provided by applicant): The American Cancer Society's most recent estimates showed that there were 43,140 new cases of pancreatic ductal adenocarcinoma (PDAC) and 36,800 deaths from PDAC in 2010. This dismal prognosis results from the fact that pancreatic cancer is not only aggressive but also a highly chemo-resistant malignancy. Specifically, the pronounced fibrotic reaction present in both the primary tumor and distant metastases limits the delivery and efficacy of chemotherapy. PDAC tumors demonstrate increased expression of the collagenase membrane type 1-matrix metalloproteinase (MT1-MMP, MMP-14) and high mobility group A2 (HMGA2), an architectural DNA-binding protein involved in regulating chromatin structure. The objective of this application is thus to de- lineate the mechanisms by which the fibrotic reaction contributes to chemotherapy resistance. Our central hy- pothesis is that the collagen microenvironment promotes chemo-resistance by inducing euchromatin formation and facilitating DNA repair. We further hypothesize that targeting MT1-MMP in vivo will consequently increase chemo-sensitivity not only through altering the chromatin structure but also by attenuating fibrosis progression itself. This hypothesis is based on our preliminary data demonstrating that PDAC cels grown in 3D collagen gels have increased histone H3 acetylation; decreased expression of heterochromatin protein 11 (HP11); and are protected from gemcitabine-induced checkpoint arrest. Our preliminary data also show that collagen in- creases HMGA2 and MT1-MMP in PDAC cells, and decreasing HMGA2 or MT1-MMP levels with siRNA at- tenuates the gemcitabine resistance seen in the collagen microenvironment. Surprisingly, expression of human MT1-MMP in transgenic and xenograft mouse models of pancreatic cancer paradoxically results in pronounced fibrosis associated with stellate cell activation. Experiments proposed in Aim 1 have been designed to deter- mine the mechanism by which 3D collagen induces euchromatin to promote chemo-resistance. Specifically, we will examine the role of HMGA2 in enhancing euchromatin in 3D collagen and in facilitating DNA repair. Ex- periments proposed in Aim 2 have been designed to test the hypothesis that targeting MT1-MMP will increase chemo-sensitivity by inducing heterochromatin and reducing DNA repair. MT1-MMP will be targeted with func- tion blocking antibody and gold nanoparticles functionalized with MT1-MMP-specific siRNA. Experiments in Aim 3 have been designed to test the hypothesis that targeting MT1-MMP will also attenuate fibrosis in addi- tion to enhancing chemo-sensitivity. The approach is innovative because it will utilize in vitro organotypic and in vivo orthotopic and transgenic mouse models of pancreatic cancer, together with gold nanoparticles functional- ized with siRNA, to demonstrate that targeting MT1-MMP sensitizes PDAC cells to the effects of chemother- apy. The proposed research is significant because it is a critical first step in the continuum of research that will not only address how the pronounced fibrotic reaction contributes to chemo-resistance but will also lead to identification of MT1-MMP as a therapeutic target to increase chemotherapy response of PDAC tumors.