HMG-CoA Reductase and Polyamine Inhibitors for Prevention of Colorectal Cancer

描述（由申请人提供）：本提案的总体目标是制定有效的化学预防策略，并了解他汀类药物（3-羟基-3甲基戊二酰辅酶a还原酶（HMG-R）抑制剂）和多胺抑制剂DFMO联合抑制结肠癌的分子机制。结直肠癌是美国男性和女性中最常见的恶性肿瘤之一。在制定翻译策略时，优选的方法是靶向选择性地促进肿瘤生长的多种信号通路，从而提供协同/加性功效。结肠肿瘤细胞通过激活鸟氨酸脱羧酶（ODC）内源性产生高水平的多胺。DFMO选择性地抑制ODC活性，从而抑制内源性多胺合成和结肠肿瘤生长。通过小窝蛋白-1 （cav-1）和SLC3A2介导的途径对肿瘤细胞中多胺转运的新见解表明，需要新的途径来有效调节肿瘤细胞中的多胺。尽管DFMO抑制内源性多胺合成，但肿瘤细胞仍然可以通过依赖于cav-1的内吞机制摄取细胞外多胺。此外，Cav-1在一氧化氮（NO）的生成以及AKT和rho信号的激活中发挥重要作用，从而增强肿瘤细胞的增殖和侵袭。我们的初步结果表明，瑞舒伐他汀与低剂量DFMO联合使用可抑制偶氮甲烷（AOM）诱导的结肠异常隐窝灶（ACF）和异常隐窝细胞增殖。我们的体外和体内实验结果表明，cav-1在结肠癌中起重要作用，他汀类药物部分通过调节cav-1抑制结肠癌肿瘤。因此，我们希望开发他汀类药物和DFMO联合用于结肠癌的预防/治疗，并了解cav-1在结肠癌中的作用及其可能通过miRNAs进行的调控。具体目的是：1)确定瑞舒伐他汀（Crestor）在F344大鼠结肠癌模型中的体内疗效(最大耐受/最佳剂量选择；剂量反应效应；以及促进/进展阶段（早期/晚期干预）的有效性。2)确定阿托伐他汀/瑞舒伐他汀联合DFMO对大鼠结肠ACF（起始后/早期）或腺瘤（进展/晚期）发展到结肠腺癌形成的影响。我们还想确定他汀类药物和DFMO对结肠黏膜和肿瘤组织中cav-1、多胺、ODC活性、NO （iNOS和eNOS活性）、SLC3A2、AKT和Rho信号蛋白、miRNA修饰水平的影响，并将这些结果与结肠肿瘤抑制联系起来。3)利用cav-1-/-敲除小鼠模型，明确cav-1在结肠癌发生中的作用，确定他汀类药物加或不加DFMO对cav-1-/-和cav-1+/+小鼠致癌物诱导的结肠肿瘤形成的影响。此外，拟开展的研究将阐明选择性miRNA在结肠癌细胞中对cav-1的调控作用，并评估他汀类药物和DFMO对cav-1 miRNA调控的可能作用。

DESCRIPTION (provided by applicant): The overall goal of this proposal is to develop effective chemopreventive strategies and understand the molecular mechanism of colon tumor inhibition by a combination of statins (3- hydroxy-3methyl glutaryl CoA reductase (HMG-R) inhibitors) and polyamine inhibitor, DFMO against colon cancer. Colorectal cancer is one of the most common malignancies in both men and women in the US. In developing translational strategies, a preferable approach is to target multiple signaling pathways that selectively contribute towards tumor growth, so that it provides a synergistic/additive efficacy. Colon tumor cells produce high levels of polyamines endogenously through the activation of ornithine decarboxylase (ODC). DFMO selectively inhibits ODC activity and thereby endogenous polyamine synthesis and colon tumor growth. New insights into the transport of polyamines in tumor cells by caveolin-1 (cav-1) and SLC3A2 mediated pathways suggest that new approaches are needed for effective polyamine regulation in tumor cells. In spite of the fact that DFMO inhibits endogenous polyamine synthesis, tumor cells can still uptake extracellular polyamines through a cav-1 dependent endocytic mechanism. Further, Cav-1 plays an important role in the generation of nitric oxide (NO) and activation of AKT and Rho-signaling, leading to enhanced tumor cell proliferation and invasion. Our preliminary results suggest, that a combination of rosuvastatin with low-dose DFMO suppresses azoxymethane (AOM)-induced colonic aberrant crypt foci (ACF), and aberrant crypt cell proliferation. Our in vitro and in vivo results suggest that cav-1 plays an important role in colon cancer, and statins exhibit colon tumor inhibition in part by modulating cav-1. Therefore, we want to develop a combination of statin and DFMO for colon cancer prevention/treatment and understand the role of cav-1 in colon cancer and its possible regulation by miRNAs. Specific aims are 1) Determine the in vivo efficacy of Rosuvastatin (Crestor) in a F344 rat colon carcinogenesis model (maximum tolerated/optimal dose selection; dose- response effects; and effectiveness during promotion/progression stages (early/late interventions). 2) Determine the combinational efficacy of atorvastatin/rosuvastatin and DFMO on the progression of colonic ACF (post-initiation/early stage) or adenoma (progression/late stage) to colon adenocarcinoma formation in rats. We also want to determine the effect of statins and DFMO on the levels of cav-1, polyamines, ODC activity, NO (iNOS and eNOS activities), SLC3A2, AKT and Rho signaling proteins, miRNA modifications in colonic mucosa and tumor tissues and correlate these results with colon tumor inhibition. 3) Define the role of cav-1 in colon carcinogenesis using cav-1-/- knockout mouse model and determine the effect of statins with or without DFMO on carcinogen induced colon tumor formation in cav- 1-/- and cav-1+/+ mice. In addition, proposed research will elucidate the role of selective miRNAs on the regulation of cav-1 in colon cancer cells, and assess the possible modulatory role of statins and DFMO on cav-1 miRNA regulation. PUBLIC HEALTH RELEVANCE: Colorectal cancer is one of the most common malignancies in both men and women in the US accounting for 50,000 deaths annually. Developing translational strategies to prevent the malignant progression of colon tumor growth is an ideal approach. The proposed studies will produce a detailed preclinical evaluation, thus setting the stage for the eventual clinical assessment of statins either alone or in combination with low-dose ODC inhibitor for colon cancer prevention and treatment. In addition, these studies will also provide valuable information on the most widely used statin, rosuvastatin which exerts multiple health benefits in humans and especially for colon cancer prevention, when it is specifically administered during the adenoma stage of colon carcinogenesis. Furthermore, the proposed studies will provide mechanistic insight into how a combination of statins and ODC inhibitors provides enhanced suppression of cellular polyamine levels leading to synergistic colon tumor growth inhibition and finally elucidate the role of cav-1 in colon cancer/prevention.