RP1: Targeting Beclin 1 complexes for broad-spectrum anti-infective therapeutics

项目摘要-RP 1：细胞内病原体的自噬传递到溶酶体进行破坏， 先天免疫的核心机制我们的目标是制定成功的战略， 感染细胞内病原体的患者的细胞/组织中的自噬，以治疗优先病原体， 基孔肯雅病毒（CHIKV）、西尼罗河病毒（WNV）、寨卡病毒（ZIKV）和其他传染病。我们 发现（1）第一个在自噬中起作用的哺乳动物基因，beclin 1;（2）一个保守的作用， 抗菌宿主防御中的自噬基因;（3）细胞穿透自噬诱导肽（Tat-Beclin 1)在体内具有广谱抗感染活性;（4）调节Beclin 1的分子机制 自噬活性，包括Bcl-2结合的抑制。在我们目前的CETR计划中，我们（1）优化了 Tat-Beclin 1肽（目前正在与我们的合作伙伴Casma Therapeutics进行IND研究）;（2） 鉴定了破坏Bcl-2/Beclin 1结合并诱导自噬的纳摩尔效力化学实体;以及 (3)使用全基因组siRNA筛选来鉴定在病毒感染期间触发自噬的蛋白质， SNX 5是一种新发现的Beclin 1结合蛋白，对抗病毒免疫很重要。 此外，我们产生了敲入Beclin 1突变的小鼠，该突变破坏了Bcl-2的结合并增加了Bcl-2的表达。 基础自噬;这些小鼠具有延长的寿命和健康寿命，并且显示出降低的M.结核 复制和对致死性CHIKV感染的易感性。这提供了重要的遗传学原理证明， 增加的自噬是安全的，并验证了Bcl-2/Beclin结合的破坏作为自噬诱导的靶点。 治疗学为了实现我们的目标，我们将利用在当前CETR期间取得的发现 计划研究，以（1）推进我们目前的半优化化合物（Tat-Beclin 1，Bcl-2/Beclin 1结合 干扰物），和（2）发现新的化学实体，通过模拟 Tat-Beclin 1的作用机制，增加Beclin 1 III类磷脂酰肌醇3-激酶的活性 （PI 3 K）自噬复合物，或增强SNX 5的自噬诱导活性。这将是完成 由UTSW的多学科团队在高通量化学筛选，药物 化学，结构生物学和临床前药理学，以及与其他领域的化学家合作， CETR项目（RP 2、RP 5）。我们将评估我们新工作中产生的新化学实体的功效 和其他项目在这个CETR计划（RP 2-RP 5）在我们建立的新生儿动物模型的西尼罗河病毒， CHIKV和ZIKV。我们预计，这些方法（旨在激活Beclin 1自噬功能）将 为未来的IND研究鉴定出先导化合物，从而开发新的自噬途径- 定向广谱抗感染药

Project Summary – RP1: The autophagic delivery of intracellular pathogens to the lysosome for destruction is a central mechanism of innate immunity. Our objective is to develop successful strategies to increase autophagy in cells/tissues of patients infected with intracellular pathogens to treat priority pathogens such as chikungunya virus (CHIKV), West Nile virus (WNV), Zika virus (ZIKV) and other infectious diseases. We discovered (1) the first mammalian gene shown to function in autophagy, beclin 1; (2) a conserved role for autophagy genes in antimicrobial host defense; (3) a cell penetrating autophagy-inducing peptide (Tat-Beclin 1) with broad-spectrum anti-infective activity in vivo; and (4) molecular mechanisms that regulate Beclin 1 autophagy activity, including inhibition by Bcl-2 binding. In our current CETR program, we (1) optimized the Tat-Beclin 1 peptide (now moving into IND-enabling studies with our partner Casma Therapeutics); (2) identified nanomolar potency chemical entities that disrupt Bcl-2/Beclin 1 binding and induce autophagy; and (3) used a genome-wide siRNA screen to identify proteins that trigger autophagy during viral infection, leading to the discovery of SNX5 as a newly-identified Beclin 1 binding protein that is important for antiviral immunity. Furthermore, we generated mice with a knock-in Beclin 1 mutation that disrupts Bcl-2 binding and increases basal autophagy; these mice have extended lifespan and healthspan, and show decreased M. tuberculosis replication and susceptibility to lethal CHIKV infection. This provides important genetic proof-of-principle that increased autophagy is safe and validates disruption of Bcl-2/Beclin binding as a target for autophagy-inducing therapeutics. To accomplish our objective, we will leverage discoveries made during our current CETR Program research to (1) advance our current semi-optimized compounds (Tat-Beclin 1, Bcl-2/Beclin 1 binding disruptors), and (2) discover new chemical entities that induce anti-infective autophagy by mimicking the mechanism of action of Tat-Beclin 1, increasing the activity of Beclin 1 Class III phosphatidylinositol 3-kinase (PI3K) autophagy complexes, or enhancing the autophagy-inducing activity of SNX5. This will be accomplished by a multidisciplinary team at UTSW with expertise in high-throughput chemical screening, medicinal chemistry, structural biology, and pre-clinical pharmacology, as well as in collaboration with chemists in other CETR Projects (RP2, RP5). We will evaluate the efficacy of new chemical entities that arise from our new work and other projects in this CETR Program (RP2-RP5) in our established neonatal animal models of WNV, CHIKV, and ZIKV. We anticipate that these approaches (aimed at activating Beclin 1 autophagy function) will result in identification of lead compounds for future IND-enabling studies to develop new autophagy pathway- directed broad-spectrum anti-infectives.