Targeting multiple signaling steps to achieve synergy

7.摘要 发展癌症治疗领域正处于一个令人兴奋的时刻。主要进展 了解导致癌症进展和化疗耐药性的分子缺陷，以及 可利用的分子靶向药物急剧增加，为开发 可能有用的联合疗法。最近FDA决定进一步帮助科学追求， 允许对涉及一种以上未经批准的药物的组合进行临床试验。 新的药物靶向控制基因表达的信号通路步骤，包括抑制剂 生长因子与其受体的结合以及抑制剂如酪氨酸激酶抑制剂。越 最近引入的RNAi试剂提供了额外的机会， 转录水平。这个项目的目标是发展概念（数学框架）和方法 用于组合调节信号级联中不同步骤的试剂，目的是实现真正的 协同或临床协同。我们提出了三个目标，使用Wnt/<$-catenin/survivin信号作为模型， 通路该通路在癌症发展和化学抗性中起着多种重要作用。 第一个目标是建立药物组合的相互作用药效学模型 针对转录前和转录后信号事件。这些模型用于单代理PD 参数值（斜率、Emax、EC 50，选自文献值和目标3中的实验数据）， 模拟不同组合的结果。使用在中开发的方法分析计算机模拟结果。 目的2鉴定协同、相加或拮抗组合，用于目的3中的实验验证。 第二个目标是开发一种方法，以统计确定性量化交互性 在具有平行或非平行浓度效应曲线的药物中。第三个目标是获得 模型预测的实验验证。第一个目标是在培养的细胞中获得PD参数 针对Wnt/β-连环蛋白/生存素中几个转录前和转录后步骤的单一药物的值 信令（例如，抗Frizzled抗体、siRNA（抗Disheveled、PP 2A、生存素）和小分子 药物OHL（被发现可以减少<$-catenin/survivin信号）。效应子包括野生型生存素 mRNA和蛋白质水平、总的和活性非磷酸化的<$-连环蛋白水平以及Tcf/Lef 转录活性第二个目标是评估模型预测的交互性是否在 体外和体内。 潜在的收益是三倍。首先，研究结果将有助于开发组合靶向 信号通路中的多个步骤。第二，鉴于Wnt/β-catenin/survivin的关键作用， 通过研究肿瘤发展中的信号传导，结果将进一步了解其对治疗收益的调节。 第三，克服生存素介导的化疗耐药性，这是治疗失败的主要原因， 化疗的效果。

7. Abstract The field of developmental cancer therapeutics is at an exciting juncture. The major advances in the understanding of molecular defects leading to cancer progression and chemoresistance, together with the dramatic increase in the available molecular targeting drugs, provide unprecedented opportunities to develop potentially useful combination therapies. The scientific pursuit is further aided by the recent FDA decision to allow clinical testing of combinations involving more than one unapproved drugs. The newer agents target steps in signaling pathways that control gene expression, including inhibitors of binding of growth factors to their receptors and inhibitors of such as tyrosine kinase inhibitors. The more recent introduction of RNAi agents provides additional opportunity for gene silencing on the post- transcriptional level. The goal of this project is to develop the concept (mathematical framework) and methods for combining agents that modulate different steps in a signaling cascade, with the goal of achieving true synergy or clinical synergy. We propose three aims, using Wnt/¿-catenin/survivin signaling as the model pathway. This pathway plays multiple important roles in cancer development and chemoresistance. The first aim is to establish interactive pharmacodynamic models (IPM) on combinations of agents targeting pre- and the post-transcriptional signaling events. These models are used with the single agent PD parameter values (slope, Emax, EC50, selected from literature values and from experimental data in Aim 3) to simulate the results of different combinations. The in silico results are analyzed with the method developed in Aim 2 to identify synergistic, additive or antagonistic combinations for experimental validation in Aim 3. The second aim is to develop a method that quantifies, with statistical certainty, the interactivity among drugs with parallel or non-parallel concentration-effect curves. The third aim is to obtain experimental validation of model predictions. The first objective is obtain in cultured cells the PD parameter values for single agents directed at several pre- and post-transcriptional steps in Wnt/¿-catenin/survivin signaling (e.g., antibody against Frizzled, siRNAs (against Disheveled, PP2A, survivin), and a small molecule drug OHL (which was found to reduce ¿-catenin/survivin signaling). The effectors include wild type survivin mRNA and protein levels, levels of total and the active non-phosphorylated ¿-catenin, and Tcf/Lef transcriptional activity. The second objective is to evaluate if the model-predicted interactivity is achieved in vitro and in vivo. Potential gains are three-fold. First, the results will assist the development of combinations targeting multiple steps in a signaling pathway. Second, in view of the critical roles of the Wnt/¿-catenin/survivin signaling in tumor development, the results will further yield insights on its modulations for therapeutic gains. Third, overcoming the survivin-mediated chemoresistance, a major cause of treatment failures, will improve the efficacy of chemotherapy.