Chemoprevention of urinary bladder carcinogenesis by flavokawain A

描述（由申请人提供）： 膀胱癌是一个主要的公共卫生负担。膀胱癌患者从诊断到死亡的成本是所有癌症中最高的（美国每位患者96，000至187，000美元）。我们的长期目标是开发新的化学预防剂，以减少膀胱癌的负担。由于暴露于致癌物是膀胱癌的主要危险因素，并且植物源性食品中含有的许多潜在保护化合物集中在尿液中，因此植物中的生物活性剂可能在膀胱癌化学预防中发挥作用。卡瓦黄素A是从卡瓦根提取物中鉴定的主要查耳酮。flavokawain A的前景是基于以下数据：1）一项流行病学研究报告说，尽管南太平洋岛屿的人口中有许多吸烟者，但高卡瓦消费与癌症发病率较低有关;（二）体外细胞培养研究表明，卡瓦提取物中的卡瓦黄素而不是卡瓦内酯对人膀胱癌细胞表现出强的抗增殖和凋亡作用，低级和高级; 3）卡瓦胡椒素A在浅表性和浸润性膀胱癌的异种移植模型中抑制肿瘤生长，而没有任何明显的副作用;以及4）卡瓦胡椒素A的化学结构与一些其他查耳酮类似，使我们预测它不会对DNA造成直接损伤，与β-微管蛋白结合，并诱导II相酶以防止致癌。我们推测卡瓦胡椒素A通过增强细胞周期调节和增加凋亡机制的敏感性对膀胱上皮细胞具有化学预防作用。将使用三种小鼠模型来检验这一假设：致癌物[4-羟丁基（丁基）亚硝胺（OH-BBN）]诱导的小鼠膀胱致癌模型和两种自发性小鼠膀胱转基因致癌模型，每种模型都将解决不同的问题。小鼠中的OH-BBN模型将用于检查黄素卡瓦因A在一级预防中的效用;用于预防具有吸烟和工业暴露等风险因素的人首次发生膀胱癌。转基因模型各自诱导一种单独的晚期膀胱癌前体状态：乳头状肿瘤或原位癌，如在临床膀胱癌的早期阶段所见。在这些转基因模型中检查黄素卡瓦因A将提供关于其在二级预防中的有用性的信息，这将在临床上解决预防膀胱癌复发和进展的问题。我们的微阵列分析已经确定了一些潜在的转录靶点（即BIM，BID，生存素，Plk 1，14-3-3gamma，和SKP-2）的黄素卡瓦菌素A时，诱导细胞凋亡和细胞周期停滞。我们将验证这些目标，通过检查它们在这些测试的动物模型中的体内表达，并通过使用RNA干扰和过表达载体，以确定它们的贡献，在体外的黄素卡瓦菌素A的行动，以及通过研究这些目标的下游事件。总之，这些研究将提供一个坚定的答案flavokawin A作为化学预防和/或化疗剂的承诺。项目叙述 虽然膀胱癌是化学预防的理想候选者，但目前可用的化学预防剂的数量有限。我们将研究一种新的化学预防活性剂，黄卡瓦因A，在致癌物质[4-羟丁基（丁基）亚硝胺（OH-BBN）]诱导的小鼠膀胱癌模型，和两个新的膀胱转基因小鼠模型，每个模型诱导一个单独的前体状态，以先进的膀胱癌：无论是乳头状移行细胞癌或原位癌。此外，我们将验证潜在的转录靶点（即，BIM、BID、生存素、Plk 1、14-3- 3 γ和SKP-2）对卡瓦胡椒素A诱导的细胞凋亡和细胞周期停滞的作用，通过在这些测试的动物模型中检查它们的体内表达，并通过使用RNA干扰和过表达载体来确定它们对卡瓦胡椒素A的体外作用的贡献。

DESCRIPTION (provided by applicant): Bladder cancer represents a major public health burden. The cost per bladder cancer patient from diagnosis to death is the highest among all cancers ($96,000 to $187, 000 per patient in the US). Our long-term goal is to develop new chemopreventive agents to reduce the burden of bladder cancer. Because exposure to carcinogens is the major risk factor for bladder cancer and many potentially protective compounds contained in plant-derived food are concentrated in the urine, it is plausible that bioactive agents in plants may play a role in bladder cancer chemoprevention. Flavokawain A is the predominant chalcone identified from kava root extract. The promise of flavokawain A is based on the following data : 1) an epidemiological study reported that high kava consumption correlated with lower incidences of cancers despite the presence of many smokers in the populations of the South Pacific Islands; 2) In vitro cell culture studies showed that flavokawains but not kavalactones in kava extracts exhibited strong antiproliferative and apoptotic effects on human bladder cancer cells characterized as low- and high- grades; 3) Flavokawain A inhibited tumor growth in xenograft models of superficial and invasive bladder cancer without any noticeable side effects; and 4) The chemical structure of flavokawain A, similar to some other chalcones, leads us to predict that it would cause no direct damage to DNA, bind to beta-tubulin, and induce phase II enzymes to protect against carcinogenesis. We hypothesize flavokawain A has chemopreventive effects on bladder epithelium via enhancing cell cycle regulation and increasing the sensitivity of apoptotic mechanisms. Three mouse models will be used to test this hypothesis: a carcinogen [4-hydroxybutyl (butyl) nitrosamine (OH-BBN)]-induced carcinogenesis model in mice bladders, and two spontaneous mouse bladder transgenic carcinogenesis models, each of which will address distinct issues. The OH-BBN model in mice will be used to examine flavokawain A's utility in primary prevention; for preventing the first occurrence of bladder cancer in those with risk factors such as cigarette smoking and industrial exposures. The transgenic models each induce a separate precursor state to advanced bladder cancer: either papillary tumors or carcinoma in situ, as seen in early stages of clinical bladder cancer. Examining flavokawain A in these transgenic models will provide information about its usefulness in secondary prevention, which clinically will address preventing bladder cancer recurrence and progression. Our microarray analysis has identified a few potential transcriptional targets (i.e. BIM, BID, survivin, Plk1, 14-3-3gamma, and SKP-2) for flavokawain A when inducing apoptosis and cell cycle arrests. We will validate these targets by examining their expression in vivo in these tested animal models and by using RNA interference and overexpression vectors to determine their contributions to flavokawain A's action in vitro, as well as by studying the down-stream events of these targets. Together, these studies will provide a firm answer to the promise of flavokawin A as a chemoprventative and/or chemotherapeutic agent.Project Narrative Although bladder cancer is an ideal candidate for chemoprevention, the number of currently available chemopreventive agents is limited. We will examine the chemopreventive activities of a novel agent, flavokawain A, in a carcinogen [4-hydroxybutyl (butyl) nitrosamine (OH-BBN)]-induced mouse bladder carcinogenesis model, and two novel bladder transgenic mouse models each which induces a separate precursor state to advanced bladder cancer: either papillary transitional cell carcinoma or carcinoma in situ. In addition, we will validate potential transcriptional targets (i.e., BIM, BID, survivin, Plk1, 14-3-3gamma, and SKP-2) for flavokawain A-induced apoptosis and cell cycle arrests by examining their expression in vivo in these tested animal models and by using RNA interference and overexpression vectors to determine their contributions to flavokawain A's action in vitro.