Novel AMPK Activators in Breast Cancer Prevention

描述（申请人提供）：越来越多的证据表明，一磷酸腺苷激活的蛋白激酶（AMPK）（一种众所周知的能量稳态和胰岛素敏感性的调节剂）与癌细胞生长和存活之间的联系是通过调节mTOR信号传导途径，抑制炎性细胞因子，特别是白细胞介素（IL）-6，通过蛋白磷酸酶（PP）2A激活刺激Akt去磷酸化，以及调节多种癌基因和肿瘤抑制因子的表达。我们对内部的基于噻唑烷二酮的聚焦化合物文库的筛选鉴定了一种先导剂Cpd 53，尽管其缺乏PPAR激动剂活性，但在THP-1巨噬细胞中以低微M效力激活AMPK并抑制LPS诱导的IL-6分泌，并通过调节AMPK相关的各种生物标志物的功能/表达状态抑制人乳腺癌细胞的增殖，IC 50为1 - 2 μ M，mTOR-p70 S6 K信号传导、Akt和IL-6产生。我们假设化合物53可以在结构上优化，以产生具有亚微摩尔效力的新型AMPK激活剂，其可以在临床可行的浓度和不存在剂量限制性毒性的情况下阻断乳腺肿瘤发生。以下具体目标将被执行，以测试我们的假设。具体目标1。继续对化合物53进行结构修饰，并对最佳药物进行机理验证。化合物53将通过组合合成进行进一步修饰，目的是产生在不同乳腺癌细胞系中维斯维斯正常乳腺上皮细胞具有低于5 M效力和癌细胞选择性的衍生物。最佳候选物将经受与AMPK活化相关的信号传导靶标的机制表征。具体目标2。评估两种优化的AMPK激活剂在MCF-7和MDA-MB-231异种移植肿瘤模型中的体内功效。我们假设，靶向AMPK激活与小分子药物构成了一个潜在的成功的策略，以抑制乳腺肿瘤的生长，而不管ER和/或其他遗传异常的功能状态。将测试两种结构优化和机械验证的衍生物与化合物53和二甲双胍相比的体内功效。将确定每种药物的最大耐受剂量，然后评价肿瘤抑制活性、肿瘤内生物标志物调节和毒性。具体目标3。评估最佳AMPK激活剂在致癌物诱导的乳腺肿瘤模型中阻断乳腺肿瘤发生的体内疗效。我们假设，目标2中确定的最活跃的AMPK激活剂将在临床可达到的浓度和没有限制毒性的情况下阻断体内乳腺肿瘤的发生。还将在未接受致癌物治疗的动物中检查该药物长期经口给药的安全性。由于AMPK代表了治疗代谢综合征和癌症的治疗相关靶标，因此对开发这种燃料敏感酶的新型药理学活化剂的兴趣越来越大。化合物53通过AMPK活化和mTOR信号传导的调节靶向肿瘤代谢和存活信号传导的能力对于开发乳腺癌的有效化学预防策略具有高转化价值。

DESCRIPTION (provided by applicant): Accumulating evidence suggests a link between adenosine monophosphate-activated protein kinase (AMPK), a well known regulator of energy homeostasis and insulin sensitivity, and cancer cell growth and survival through the regulation of mTOR signaling pathways, the suppression of inflammatory cytokines, especially interleukin (IL)-6, the stimulation of Akt dephosphorylation through protein phosphatase (PP)2A activation, and modulation of the expression of multiple oncogenes and tumor suppressors. Our screening of an in-house, thiazolidinedione-based focused compound library identified a lead agent, Cpd 53, that despite lacking PPAR agonist activity, activates AMPK and inhibits LPS-induced IL-6 secretion in THP-1 macrophages with low microM potency, and inhibits the proliferation of human breast cancer cells with IC50 of 1 - 2 microM by modulating the functional/expression status of various biomarkers related to AMPK, mTOR-p70S6K signaling, Akt, and IL-6 production. We hypothesize that Cpd 53 can be structurally optimized to generate novel AMPK activators with sub-microM potency that can block mammary tumorigenesis at clinically feasible concentrations and in the absence of dose-limiting toxicities. The following Specific Aims will be carried out to test our hypothesis. Specific Aim 1. To continue the structural modification of Cpd 53 and to conduct mechanistic validation of optimal agents. Cpd 53 will undergo further modifications via combinatorial synthesis with the goal of generating derivatives with sub-5M potency and cancer cell selectivity in different breast cancer cell lines vis-`- vis normal breast epithelial cells. The optimal candidate(s) will be subjected to mechanistic characterizations of signaling targets pertinent to AMPK activation. Specific Aim 2. To assess the in vivo efficacy of two optimized AMPK activators in MCF-7 and MDA-MB-231 xenograft tumor models. We hypothesize that targeting AMPK activation with a small-molecule agent constitutes a potentially successful strategy to suppress breast tumor growth irrespective of the functional status of ER and/or other genetic abnormalities. Two structurally optimized and mechanistically validated derivatives will be tested for in vivo efficacy in comparison to Cpd 53 and metformin. Maximum tolerated doses for each agent will be determined, followed by evaluation of tumor suppressive activity, intratumoral biomarker modulation, and toxicity. Specific Aim 3. To assess the in vivo efficacy of the optimal AMPK activator to block mammary tumorigenesis in a carcinogen-induced mammary tumor model. We hypothesize that the most active AMPK activator identified in Aim 2 will block mammary tumorigenesis in vivo at clinically attainable concentrations and in the absence of limiting toxicity. The safety of chronic oral treatment with this agent will also be examined in carcinogen-untreated animals. As AMPK represents a therapeutically relevant target for the treatment of the metabolic syndrome and cancer, there is a growing interest in the development of novel pharmacological activators for this fuel-sensing enzyme. Cpd 53's ability to target tumor metabolism, and survival signaling via AMPK activation and modulation of mTOR signaling is of high translational value to develop an effective chemopreventive strategy for breast cancer.