Disruption of AKT Pathway for Cancer Intervention

描述(申请人提供)：AKT是诱导细胞存活、生长和转化的主要途径。目前已从AKT家族中鉴定出三种亚型：AKT1、AKT2和AKT3。在人类恶性肿瘤中发现了AKT，尤其是AKT2的频繁改变。AKT的异位表达诱导化疗耐药，而显性阴性AKT使细胞对化疗药物诱导的细胞凋亡敏感。通过筛选来自140,000种化合物的NCI多样性集，我们最近获得了30多种化合物，它们显著抑制了AKT2转化的NIH3T3细胞的生长，但在pcDNA3转基因的NIH3T3细胞中却没有。进一步分析表明，其中两个直接抑制了结构性活性的AKT2激酶活性，另外两个直接降低了AKT的磷酸化。所有四(4)种化合物都能显著抑制AKT途径改变的三种人类癌细胞系的生长。基于这些发现，我们假设AKT是癌症干预的关键治疗靶点，AKT信号的特异性抑制剂(S)将降低AKT活性升高的肿瘤的致瘤性。由于抑制AKT可诱导一系列哺乳动物细胞的凋亡，因此AKT抑制剂S与其他抗癌药物联合应用可有效治疗其他基因突变的肿瘤。随着越来越多的研究表明AKT三种亚型之间的生物学/生理功能差异明显，针对每一种亚型寻找抑制剂(S)将提高疗效并减少副作用。这项研究的基本原理是，AKT在人类恶性肿瘤中经常发生改变，抑制AKT会诱导细胞凋亡和细胞生长停滞。因此，开发特异性的AKT抑制剂(S)具有很大的潜力，可以改善癌症的治疗，并为AKT信号转导途径的研究提供新的手段。本项目的目标是开发特异性的AKT抑制剂，并评估它们通过阻断AKT途径而无肉眼毒性逆转人类肿瘤恶性转化的能力。具体目标是：(1)验证AKT1、AKT2和AKT3作为人类癌症的治疗靶点，并通过高通量筛选合成AKT底物模拟物和NCI多样性集的化合物来确定先导化合物是AKT的潜在干扰物。(2)测定先导化合物抑制AKT途径的特异性。(3)检测有希望的AKT抑制剂S对AKT1、AKT2和AKT3野生型和活性AKT1、AKT2和AKT3AKT信号转导的阻断作用和对细胞转化的抑制作用。(4)评价活性化合物在细胞培养和PI3K/PTEN/AKT通路改变的人肿瘤动物模型中的抗肿瘤作用。

DESCRIPTION (provided by applicant): AKT is a major pathway to induce cell survival, growth and transformation. Three isoforms, AKT1, AKT2 and AKT3, have been identified from the AKT family. Frequent alterations of AKT, especially AKT2, have been detected in human malignancies. Ectopic expression of AKT induces chemo-resistance, whereas, dominant negative AKT sensitizes cells to chemotherapeutic drug-induced apoptosis. By screening the NCI diversity set that was derived from 140,000 compounds, we have recently obtained more than 30 compounds that significantly inhibited growth in AKT2-transformed, but not in pcDNA3-transfected NIH 3T3 cells. Further analyses show that two of them directly inhibited constitutively active AKT2 kinase activity, and another two directly decreased AKT phosphorylation. All four (4) compounds significantly inhibit growth in three (3) human cancer cell lines where the AKT pathway is altered. Based on these findings, we hypothesize that AKT is a critical therapeutic target for cancer intervention and that specific inhibitor(s) of AKT signaling will reduce tumorigenicity in tumors with elevated AKT activity. Since inhibition of AKT induces apoptosis in a range of mammalian cells, AKT inhibitor(s) could be effective, in combination with other anticancer drugs, for the treatment of tumors with other gene alterations. As accumulated studies show clearly biological/physiological function differences between three (3) isoforms of AKT, identification of inhibitor(s) for each isoform of AKT will enhance therapeutic efficacy and reduce side effects. The rationale for the proposed research is that AKT is frequently altered in human malignancy and that inhibition of AKT induces apoptosis and cell growth arrest. Therefore, development of specific AKT inhibitor(s) has great potential to improve cancer treatment and provide additional means to characterize the AKT signal transduction pathway. The objective of this project is to develop specific AKT inhibitors and evaluate their abilities to reverse malignant transformation of human tumors by disruption of the AKT pathway without gross toxicity. The specific aims are: (1) Validate AKT1, AKT2, and AKT3 as therapeutic targets in human cancer and identify lead compounds as potential disruptors of AKT by high-throughput screening of compounds from synthetic AKT substrate mimics and the NCI diversity set. (2) Determine the specificity of lead compounds in the inhibition of AKT pathway. (3) Examine the abilities of promising AKT inhibitor(s) to disrupt AKT signaling in intact cells and inhibit cell transformation by wild type and constitutively active AKT 1, AKT2 and AKT3. (4) Evaluate anti-tumor efficacy of active compounds in cell culture and animal models of human cancers where the PI3K/PTEN/AKT pathway is altered.