Class III PI3K as an Autophagy Reactivation Switch in Malignant Transformation

项目摘要 巨噬细胞自噬(以下简称自噬)对细胞内环境的稳定至关重要。在健康细胞中，高 基础自噬水平对于正常的动态平衡和抗癌是必需的。 免疫监控。当自噬受损时，可能会发生恶性转化。自噬 在肿瘤超过其可用营养供应之前，活动是低的。随着肿瘤的生长超出了它的血液 供应，环境变得缺氧，自噬重新激活对于肿瘤是必要的 扩散和侵袭。抗疟药氯喹先导化合物抑制自噬的药理学研究 然而，在肿瘤消退方面，这种药物并不是针对癌细胞的，而且有许多非靶向途径。 自噬在癌症中的双重作用使我们在治疗上针对它的能力变得更加复杂。这是未知的 如何恢复自噬功能，这是该领域的中心问题。 所有自噬激活机制都涉及磷脂酰肌醇3-激酶。 含有肿瘤抑制蛋白BECN1的复合体I(PI3KC3-C1)。PI3KC3-C1是中心引发剂， 然而，它的激活机制尚不清楚，这使得它成为一个具有挑战性的靶向酶。 从治疗上讲。在原子水平上发现这种中枢脂蛋白激酶的激活机制 通过冷冻电子显微镜的复杂将极大地有助于合理设计治疗方法的能力 在癌症转化或增殖的不同阶段选择性地激活或抑制PI3KC3-C1。 鉴定SILAC(稳定)自噬再激活过程中蛋白质组的大规模变化 细胞培养中氨基酸的同位素标记)与质谱学相结合 已执行。这里的目标是确定在增殖的肿瘤中是否有一个独特的靶点， 依赖于特定的自噬激活机制或选择性的自噬途径。 理想情况下，这项研究将产生新的候选蛋白质用于治疗，这将 比羟基氯喹治疗胰腺癌更具特异性，毒性更小。

Project Summary Macroautophagy (hereafter autophagy) is crucial to cellular homeostasis. In healthy cells, high levels of basal autophagy are necessary for proper homeostasis and are required for anticancer immunosurveillance. Malignant transformations can arise when autophagy is impaired. Autophagic activity is low until the tumor outpaces its available nutrient supply. As the tumor outgrows its blood supply, the environment become hypoxic, and autophagy re-activation is necessary for tumor proliferation and invasion. Pharmological inhibition of autophagy by antimalarial drug chloroquine leads to tumor regression, however, this drug is not specific to cancerous cells and has many off-target paths. This dual role of autophagy in cancer complicates our ability to target it therapeutically. It is unknown how autophagic function is restored, and this is a central question in the field. All autophagy activation mechanisms involve the lipid kinase class III phosphatidylinositol 3-kinase complex I (PI3KC3-C1) containing tumor suppressor protein BECN1. PI3KC3-C1 is a central initiator, however, its activation mechanism is unknown and this makes it a challenging enzyme to target therapeutically. Discovery of the activation mechanism, at the atomic level, of this central lipid kinase complex through cryo-electron microscopy will greatly aid the ability to rationally design therapeutics to selectively activate or inhibit PI3KC3-C1 at various stages in cancer transformation or proliferation. To identify large-scale changes in the proteome during autophagy reactivation in SILAC (Stable Isotope Labeling with Amino acids in Cell culture) coupled with mass spectrometry will be performed. The objective here is to determine if there is a unique target in proliferating tumors, dependent on a specific autophagy activation mechanism or on a selective autophagy pathway. Ideally, this study would produce new protein candidates to target therapeutically, which would be more specific and less toxic than hydroxychloroquine to treat pancreatic cancers.