Turning Inhibition of Translation Initiation into Cancer Therapy

 描述（由申请人提供）：本项目的总体目标是扩展我们对将翻译起始抑制与白细胞介素-24（IL-24）引发的细胞凋亡和肿瘤抑制联系起来的细胞和分子机制的理解。IL-24作为用于治疗实体瘤的基于基因的治疗剂的临床测试证明IL-24是有效且安全的。该提议与当前的观点一致，即现代抗癌疗法必须是靶向特异性的，以对正常细胞的最小影响抑制癌症的生长，并且低毒性的有效抗癌剂可以通过靶向负责癌症的发生、维持和/或进展的基本机制来实现。我已经表明，IL-24导致细胞内Ca++的部分耗尽和内质网（ER）应激的激活，这反过来又诱导真核起始因子2 α（eIF 2 α）的抑制性磷酸化。eIF 2 β的磷酸化和三元复合物的可用性不仅如最初所认为的那样控制总的翻译速率，而且还控制特定基因簇的表达。大多数致癌蛋白质的严格翻译控制解释了不受限制的翻译起始和恶性转化之间的联系，这在两种实验模型和许多过度表达翻译起始因子的人类癌症中已经得到了很好的证实。磷酸化eIF 2 β以高亲和力结合鸟嘌呤核苷酸交换因子eIF 2B，从而抑制eIF 2 *GDP再循环为eIF 2 *GTP，耗尽启动新一轮翻译所需的三元复合物。已经确定，ER Ca++储存的部分消耗激活eIF 2 β激酶，其磷酸化eIF 2 β，从而限制翻译起始速率。此外，我已经证明IL-24诱导结合免疫球蛋白蛋白（BiP）和C/-EBP同源蛋白（CHOP）的表达，这是三元复合物丰度的两个下游标志物。本申请的工作假设是IL-24发挥抗癌活性，因为它有助于恢复对翻译起始的生理限制。我已经证明了Sigma 1受体（Sig 1 R）与IL-24相互作用，并且这种IL-24：Sig 1 R是IL-24诱导的ER应激、钙动员、eIF 2 β磷酸化和癌细胞凋亡的关键上游信号。在本申请中，我们建议继续努力表征IL-24作为翻译起始抑制剂的癌细胞特异性作用。具体而言，我们将1）使用Cameleon钙指示蛋白直接评估IL-24处理后ER-储存的钙含量（具体目标1），2）确定三元复合物的消耗是否介导IL-24的抗癌作用（具体目标2），3）确定Sig 1 R对IL-24介导的翻译起始抑制的作用（具体目标3）。对于这些研究，我们 将与何塞·A博士合作Halperin博士，来自哈佛医学院，是癌症治疗靶向翻译起始的专家。

 DESCRIPTION (provided by applicant): The overall goal of this project is to expand our understanding of the cellular and molecular mechanisms that link translation initiation inhibition to apoptosis and tumor suppression triggered by Interleukin-24 (IL-24). Clinical testing of IL-24 as a gene-based therapeutic for the treatment of solid tumors demonstrated that IL-24 is efficacious and is safe. This proposal is consistent with the current view that modern anti-cancer therapy must be target-specific, inhibit the growth of cancer with minimal effect on normal cells, and that effective anti- cancer agents of low toxicity can be achieved by targeting the fundamental mechanisms responsible for the genesis, maintenance, and/or progression of cancer. I have shown that IL-24 causes partial depletion of intracellular Ca++ and activation of endoplasmic reticulum (ER) stress, which in turn induces inhibitory phosphorylation of eukaryotic initiation factor 2 alpha (eIF2). Phosphorylation of eIF2 and the availability of he ternary complex control not only the overall rate of translation, as initially thought, but also th expression of specific gene clusters. The tight translational control of most oncogenic proteins explains the link between unrestricted translation initiation and malignant transformation that has been well established in both experimental models and in numerous human cancers that overexpress translation initiation factors. Phosphorylated eIF2 binds with high affinity to the guanine nucleotide exchange factor eIF2B and thereby inhibits recycling of eIF2*GDP into eIF2*GTP, depleting the ternary complex necessary to initiate a new round of translation. It is well established that partial depletion of ER Ca++ stores activates the eIF2 kinases that phosphorylate eIF2 and thereby limits the rate of translation initiation. Furthermore, I have demonstrated that IL-24 induces expression of Binding immunoglobulin protein (BiP) and C/-EBP homologous protein (CHOP), two down-stream markers of ternary complex abundance. The working hypothesis of this application is that IL-24 exerts anticancer activities because it helps restore physiological restraints on translation initiation. I have demonstrated that Sigma 1 Receptor (Sig1R) interacts with IL-24 and that this IL-24:Sig1R is a critical upstream signal for IL-24-induced ER-stress, calcium mobilization, phosphorylation of eIF2 and apoptosis on cancer cells. In this application we propose to continue our efforts to characterize the cancer cell- specific actions of IL-24 as an inhibitor of translation initiation. Specifically, we will 1)use cameleon calcium indicator proteins to assess directly the calcium content of the ER-stores after IL-24 treatment (Specific Aim 1), 2) determine whether depletion of the ternary complex mediates the anticancer effect of IL-24, (Specific Aim 2), and 3) determine the role of Sig1R on IL-24-mediated inhibition of translation initiation (Specific Aim 3). For these proposed studies we will collaborate with Dr. Jose A. Halperin, from Harvard Medical School, and expert on targeting translation initiation for cancer therapy.