Systematic Targeting of Oncogene Interacting Proteins to Reveal New Therapeutic Strategies

项目总结/摘要 癌症是我们亲密的一部分，招募一系列复杂的内源性细胞过程来驱动它的健康。 癌症的发生和发展通常是多种信号协调失调的结果 关键癌基因的通路，战略性地拉拢癌症信号网络。这些致癌基因很少起作用 而是形成复杂的高阶复合体，参与蛋白质的多面网络， 蛋白质相互作用所有人类癌症中最常见的突变蛋白质之一是 磷脂酰肌醇3-激酶（PI 3 K）癌基因，一种可以利用多种细胞程序来驱动 疾病，包括增加的增殖、存活、运动性、细胞生长和代谢活性。广泛抑制 已知PI 3 K产生全身毒性，尤其是代谢毒性，这限制了其临床开发。然而，在这方面， 靶向PI 3 K相互作用蛋白可以通过中断PI 3 K致癌活性， 最小化代谢失调。本提案的长期目标是深化和完善我们的 利用系统遗传学、蛋白质组学和 数学建模方法。本提案的总体目标是确定PI 3 K相互作用的作用。 本发明涉及蛋白质在调节PI 3 K活性和下游细胞过程的募集中的作用，以及使用这种蛋白质， 了解以确定替代治疗靶点。这一目标将通过测试中央 假设PI 3 K相互作用蛋白或下游信号通路可以被调节以调节PI 3 K 活性和特异性，具有同时降低恶性肿瘤和全身毒性的潜力。为了验证这一 假设，将追求以下三个目标。(Aim 1）揭示PI 3 K活性和癌症的调节 PIK 3CA相互作用蛋白的表型。这一目标将利用CRISPR/Cas9基因敲除技术， 系统性缺失和过表达对应于PIK 3CA相互作用蛋白基因，并使用活细胞 显微镜，结合生物化学测量，以评估对细胞增殖的影响， 存活、生长、运动和代谢。(Aim 2）阐明由PI 3 K介导的蛋白质组学开发， PIK 3CA相互作用蛋白。这里，PIK 3CA相互作用物优先结合常见的H1047 R突变体，如 以及目标1中确定的命中，将进行全球蛋白质组学和磷酸蛋白质组学分析，以确定 信号通路和生物过程由每个PIK 3CA相互作用蛋白调节。(Aim 3）描绘 PI 3 K介导的促癌信号转导的操纵机制。在这里，一个新的计算框架 将通过将数据驱动的网络传播技术与机械常微分相结合来开发 方程（ODE）建模，以描绘将每个PIK 3CA相互作用蛋白质与其 下游效应。成功完成拟议的研究将大大提高我们的机械 了解癌症中的癌基因调控。这将是一个重大的贡献，因为它将揭示新的 治疗策略，以对抗癌症，同时最大限度地减少全身毒性。

PROJECT SUMMARY/ABSTRACT Cancer is an intimate part of us, recruiting a complex array of endogenous cellular processes to drive its fitness. Cancer initiation and progression is frequently the result of coordinated dysregulation of multiple signaling pathways by key oncogenes that strategically coopt the cancer signaling network. These oncogenes rarely work in isolation but instead form intricate higher-order complexes and participate in multifaceted networks of protein- protein interactions. One of the most commonly mutated proteins across all of human cancers is the phosphoinositide 3-kinase (PI3K) oncogene, a lipid kinase that can exploit diverse cellular programs to drive disease, including increased proliferation, survival, motility, cell growth and metabolic activity. Broad inhibition of PI3K is known to generate systemic toxicities, especially metabolic, which limit its clinical development. However, targeting PI3K interacting proteins may enable a safer alternative, by interrupting PI3K oncogenic activity while minimizing metabolic dysregulation. The long-term goal of this proposal is to deepen and refine our understanding of oncogene regulation of cancer signaling networks using systematic genetic, proteomic, and mathematical modeling approaches. The overall objective of this proposal is to identify the role of PI3K interacting proteins in modulating PI3K activity and recruitment of downstream cellular processes and to use this understanding to identify alternative therapeutic targets. This objective will be reached by testing the central hypothesis that PI3K interacting proteins, or downstream signaling pathways, can be modulated to tune PI3K activity and specificity, with the potential to simultaneously reduce malignancy and systemic toxicity. To test this hypothesis, the following three aims will be pursued. (Aim 1) Reveal Regulation of PI3K Activity and Cancer Phenotypes by PIK3CA Interacting Proteins. This aim will use CRISPR/Cas9 gene knockout technology to systematically delete and overexpress genes corresponding to PIK3CA interacting proteins and use live cell microscopy, combined with biochemical measurements, to assess the resulting impact on cell proliferation, survival, growth, motility, and metabolism. (Aim 2) Elucidate Proteomic Exploitation by PI3K as Mediated by PIK3CA Interacting Proteins. Here, PIK3CA interactors that preferentially bind the common H1047R mutant, as well as hits identified from Aim 1, will be subject to global proteomics and phosphoproteomics profiling to identify signaling pathways and biological processes regulated by each PIK3CA interacting protein. (Aim 3) Delineate Mechanisms of PI3K-mediated Manipulation of Pro-Cancer Signaling. Here, a novel computational framework will be developed by uniting data-driven network propagation techniques with mechanistic ordinary differential equation (ODE) modeling to delineate mechanistic pathways linking each PIK3CA interacting protein to its downstream effect. Successful completion of the proposed research will greatly enhance our mechanistic understanding of oncogene regulation in cancer. This will be a significant contribution as it will reveal novel therapeutic strategies to fight cancer while minimizing systemic toxicities.