The role of translation initiation factor eIF5B in lung cancer pathogenesis

项目总结 肺癌细胞表达高水平的PD-L1，这是T细胞上PD-1受体的配体，使肿瘤 直接抑制T细胞活性。抗PD-1/PD-L1抗体诱导强大的抗肿瘤免疫，并已被 被批准为肺癌的一线治疗方法。然而，在所有非小细胞肺癌中只有20%左右 (NSCLC)从检查站封锁中受益。进一步了解监管环境的机制 因此，肺癌的免疫检查点是必要的。为了解决这个问题，我的实验室使用了基于CRISPR的 筛选肺癌细胞中PD-L1的调节因子，揭示PD-L1对肺癌细胞的有效诱导作用 整合应激反应(ISR)途径的激活。从机制上讲，ISR激活导致 增强PD-L1翻译和抑制抗肿瘤免疫。我们进一步证明了ISR- PD-L1的依赖翻译需要替代翻译起始因子eIF5B。的典范作用 该GTP酶催化核糖体亚基连接，确保80s核糖体组装和有效的起始密码子 选择。EIF5B过度表达和/或扩增在人类肺癌中很常见，并与 预后不良。值得注意的是，eIF5B的过度表达足以诱导PD-L1蛋白水平，即使在 未激活ISR。这些发现揭示了一种新的免疫检查点激活机制和 提示eIF5B可能成为肺癌干预的新靶点。我们进一步证明了 增强表达eIF5B可加速肺癌细胞、小鼠同基因模型和 人支气管上皮细胞提示，它也以细胞自主的方式促进肿瘤生长。 我们建议通过检验以下中心假设来阐明eIF5B在肺肿瘤发生中的作用： EIF5B在人类肺癌中作为癌基因发挥作用，通过诱导PD-L1翻译和通过驱动一个 促进肿瘤的翻译计划。为了测试这一点，将追求三个具体目标 假设：在目标1中，我们将剖析eIF5B促进PD-L1和PD-L1翻译的机制 肺癌中额外的致癌基因。在目标2中，我们将对其致癌活性进行功能评估。 EIF5B使用一种新的条件性过表达eIF5B转基因小鼠模型。在《目标3》中，我们将 EIF5B功能缺失对细胞自主性肿瘤生长的影响及其对T细胞的影响 KrasLSL-G12D；TP53FL/FL小鼠模型对新产生的条件性FLOX基因敲除的反应 等位基因和杂合子生殖系基因敲除小鼠。这些目标将利用我们的专业知识，并且 以评估eIF5B的致癌活性，并阐明其作用机制 这种翻译启动因子促进肺肿瘤的发生。我们预计这些研究将提供新的 对肿瘤进展和免疫逃避中翻译控制机制的洞察。

PROJECT SUMMARY Lung cancer cells express high levels of PD-L1, a ligand of the PD-1 receptor on T-cells, allowing tumors to directly suppress T cell activity. Anti-PD-1/PD-L1 antibodies induce potent anti-tumor immunity and have been approved as a first-line therapy for lung cancer. However, only ~20% of all non-small cell lung cancers (NSCLCs) benefit from checkpoint blockade. A further understanding of the mechanisms that regulate the immune checkpoint in lung cancer is therefore needed. To address this, my laboratory used CRISPR-based screening to identify regulators of PD-L1 in lung cancer cells, revealing potent induction of PD-L1 upon activation of the integrated stress response (ISR) pathway. Mechanistically, ISR activation resulted in enhanced PD-L1 translation and suppression of anti-tumor immunity. We further demonstrated that ISR- dependent translation of PD-L1 requires the alternative translation initiation factor eIF5B. The canonical role of this GTPase is to catalyze ribosomal subunit joining, ensuring 80S ribosome assembly and efficient start codon selection. eIF5B overexpression and/or amplification is frequent in human lung cancers and is associated with poor prognosis. Remarkably, eIF5B overexpression is sufficient to induce PD-L1 protein levels even in the absence of ISR activation. These findings uncovered a new mechanism of immune checkpoint activation and suggested that eIF5B may be a novel target for lung cancer intervention. We further demonstrated that enforced expression of eIF5B accelerates proliferation in lung cancer cells, in mouse syngeneic models, and in human bronchial epithelial cells suggesting that it also promotes tumor growth in a cell-autonomous manner. We propose to elucidate the role of eIF5B in lung tumorigenesis by testing the following central hypothesis: EIF5B functions as an oncogene in human lung cancer by inducing PD-L1 translation and by driving a tumor-promoting translational program. Three Specific Aims will be pursued in order to test this hypothesis: In Aim 1, we will dissect the mechanisms through which eIF5B promotes translation of PD-L1 and additional oncogenic mRNAs in lung cancer. In Aim 2, we will functionally evaluate the oncogenic activity of eIF5B using a novel transgenic mouse model with conditional eIF5B overexpression. In Aim 3, we will characterize the effects of eIF5B loss of function on cell autonomous tumor growth versus its effect on T cell responses in the KrasLSL-G12D; Tp53 fl/fl mouse model using a newly generated conditional floxed knockout allele and a heterozygous germline knockout mouse. These aims will take advantage of our expertise, and that of our collaborators, to evaluate eIF5B oncogenic activity, and elucidate the mechanisms through which this translation initiation factor promotes lung tumorigenesis. We anticipate that these studies will provide new insights into mechanisms of translational control in tumor progression and immune evasion.