DEVELOPMENT OF NOVEL CHEMOPRVENTIVE AGENTS

描述(由申请人提供)： 为了实现在开发癌症控制的化学预防方法方面日益增长的兴趣和希望的目标，需要新的药物(Sporn和Liby，2005，Smith等人，2005；Lippman和Lee，2006；Francis等人，2006)。在寻找核苷转运抑制剂作为潜在的化学预防药物的过程中，我们发现双嘧达莫是一种有效的化学预防药物，可以对抗12-O-佛波醇-13-肉豆蔻酸酯(TPA)对JB6 P+细胞的促肿瘤作用。双嘧达莫的IC50为10 NM，远高于已知的化学预防药物脱氢表雄酮(DHEA)，后者在同一JB6 P+细胞致癌实验中的IC50为1.0？M。因此，这项应用计划利用这一新发现来进行结构-活性关系(SAR)研究，并利用它来选择将在活体动物模型中作为新的癌症化学预防药物进行评估的化合物。申请人的实验室已经建立了合成方法的新化合物将被合成，并对12-O-佛波醇-13-肉豆蔻酸乙酯(TPA)在JB6 P+细胞中诱导的肿瘤促进作用进行评估。该应用程序与及时的计划公告(PAR-06-313)非常吻合，该计划有两个目标，即“化学预防新方法的试验测试和开发……”以及“开发和评估分子靶标，以通过天然的、合成的、化学预防药物来预防、逆转或延缓癌前病变(以及癌症过程)的进展。”我们正在开发合成化学防腐剂。我们试图在这一应用程序中实现两个具体目标。具体目的1.对本实验室合成的新型核苷转运抑制剂作为抗肿瘤促进剂进行体外评价，筛选有效的、类药物的化合物进行体内动物实验。特异性目的2.研究特异性目的1化合物在体内小鼠皮肤癌变模型中的化学预防能力。申请人实验室开发的一种新的AP-1-SEAP JB6 P+细胞报告试验将与锚定非依赖性克隆形成试验结合使用，以评估目标化合物的体外化学预防活性。还将进行辅助研究，为化学预防药物的开发建立合理的基础。因此，在JB6P+小鼠表皮癌变模型中，将对参与肿瘤促进信号转导通路的丝裂原活化蛋白激酶(MAPKs)，如ERKs、p38和JNK的水平和激活进行测定。Sencar小鼠DMBA-发起-TPA促进皮肤癌模型将用于特定目标2的体内研究，以确定潜在的临床前开发作为化学预防药物的候选药物。肿瘤的发病率和多样性将进行组织病理学和免疫组织化学分析，以及体内研究中将诱导的肿瘤的生物标志物分析。该项目的成功将扩大我们的化学预防药物的武器库，并发现新的化学预防分子靶点。

DESCRIPTION (provided by applicant): Novel agents are needed for achieving the goals of the growing interest and promise in developing chemoprevention approaches to cancer control (Sporn and Liby, 2005, Smith et al., 2005; Lippman and Lee, 2006; Francis et al., 2006). In pursuing nucleoside transport inhibitors as potential chemopreventive agents, we have discovered that dipyridamole is a potent chemopreventive agent against 12-O-phorbol-13-myristylacetate (TPA) tumor promotion in JB6 P+ cells. With an IC50 of 10 nM, dipyridamole is a much more potent than the known chemopreventive agent dehydroepiandrosterone (DHEA), which exhibited an IC50 of 1.0 ¿M in the same JB6 P+ cell carcinogenesis assay. This application is thus planned to capitalize on this novel finding to conduct a structure-activity relationship (SAR) study, and use it to select compounds that will be evaluated as novel cancer chemoprevention agents in an in vivo animal model. New compounds for which synthetic methods have already been established in the applicant's laboratory will be synthesized and evaluated against 12-O-phorbol-13- myristylacetate (TPA)-induced tumor promotion in JB6 P+ cells. This application fits well with the timely program announcement (PAR-06-313), which has two of its objectives as the "pilot testing and development of new methods of chemoprevention..." and the "development and evaluation of molecular targets to prevent, reverse, or retard progression of precancerous lesions (and, hence, the cancer process) by natural, synthetic, chemopreventive agents." We are developing synthetic chemopreventive agents. We seek to achieve two specific aims in this application. Specific Aim 1. To evaluate novel nucleoside transport inhibitors synthesized in the applicant's laboratory as antitumor promotion agents in vitro, and select potent and "drug-like" compounds for in vivo animal testing. Specific Aim 2. To investigate the chemopreventive ability of compounds selected from Specific Aim 1 in an in vivo mouse skin carcinogenesis model. A new AP-1-SEAP JB6 P+ cell reporter assay developed in the applicant's laboratory will be used in conjunction with an anchorage independent clonogenic assay to assess the in vitro chemopreventive activities of target compounds. Ancillary studies will also be undertaken to establish a rational basis for chemopreventive agent development. Thus, the determination of the levels and of activation of mitogen activated protein kinases (MAPKs) such as ERKs, p38 and JNK, which have been shown to be involved in tumor promotion signal transduction pathways in the JB6 P+ mouse epidermal carcinogenesis model, will be undertaken. The SENCAR mouse DMBA-initiation-TPA-promotion skin carcinogenesis model will be used for the in vivo studies in Specific Aim 2 to identify potential candidates for preclinical development as chemopreventive agents. Tumor incidence and multiplicity will be Histopathological and immunohistochemical analysis will be performed, as well as biomarker analysis of tumors that will be induced in the in vivo studies. The success of this project will expand our armamentarium of chemopreventive agents and uncover novel chemoprevention molecular targets.