Chemical Biology to Modulate PCSK9 and Treat Atherosclerosis

调节 PCSK9 和治疗动脉粥样硬化的化学生物学

• 批准号: 10598882
• 负责人: John S Chorba
• 金额: $ 9.34万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-07-01 至 2026-06-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10598882
• 关键词: Address; Affect; Antibodies; Anticoagulants; Atherosclerosis; Biology; Blood Circulation; CRISPR interference; Cardiovascular system; Cells; Chemicals; Cholesterol; Clinical; Coronary heart disease; Drug Targeting; Event; Goals; Guidelines; Health; Heparan Sulfate Proteoglycan; Heparin; Hepatic; Human; In Vitro; Lead; Liver; Long-Term Effects; Low Density Lipoprotein Receptor; Low-Density Lipoproteins; Lysosomes; Mediator of activation protein; Mission; Molecular Chaperones; National Heart, Lung, and Blood Institute; Outcome; Pathway interactions; Peptide Hydrolases; Pharmaceutical Preparations; Proprotein Convertases; Proteolysis; Research; Research Design; Safety; Serum; Small Interfering RNA; Subtilisins; Therapeutic; Tissues; United States National Institutes of Health; biochemical tools; blood lipoprotein; cost; genetic variant; genome-wide; improved; in vivo Model; innovation; new therapeutic target; novel; novel therapeutics; parent grant; public health relevance; receptor downregulation; receptor expression; small molecule; small molecule therapeutics; therapeutic target; trafficking

PARENT GRANT PROJECT SUMMARY/ABSTRACT Serum low-density lipoprotein (LDL) causes atherosclerotic heart disease. As the LDL receptor (LDLR) on the liver clears LDL from the blood, upregulating hepatic LDLR reduces both LDL and cardiovascular events. The self-cleaving protease PCSK9 (proprotein convertase subtilisin/kexin type 9) is a validated therapeutic target; it chaperones the LDLR for lysosomal degradation, downregulating its function. Antibodies against PCSK9 lower LDL and improve clinical outcomes, but cost and administration requirements illustrate a need for alternatives. Liver-targeted siRNA also robustly lowers LDL, but unlike the well-tolerated genetic variants, it removes all PCSK9 from the cell, raising safety concerns. Though the mechanistic basis for PCSK9's effect on the hepatic LDLR is well understand, its functions outside this canonical pathway are less clear. Therefore, the overall goal of our proposal is to develop biochemical tools to mechanistically dissect the biology of PCSK9, so as to 1) anticipate the long-term effects of current anti-PCSK9 therapies and 2) develop the proofs-of-principle for novel PCSK9-targeting strategies. To this end, we have identified three promising lead compounds that specifically target intracellular PCSK9. These compounds both inhibit PCSK9 self-proteolysis and also upregulate LDLR expression. In Aim 1, we will chemically optimize these compounds, confirm the mechanistic basis of their effects, and validate their function on PCSK9 and the LDLR in an in vivo model. This will establish novel chemical probes which can be used to study the poorly known function of intracellular PCSK9, as well as serve as the starting point for an alternative small molecule therapeutic to upregulate the LDLR. We have also found that additional factors, including heparan sulfate proteoglycans (HSPGs), interact with PCSK9 to modulate its trafficking to the lysosome, suggesting that there are additional drug targets in this pathway. In Aim 2, we will evaluate the contribution of HSPGs as mediators of the PCSK9 endocytic trafficking required for LDLR downregulation, both in vitro and in the human. These results will elucidate whether heparins, which are already approved therapies, could be repurposed to inhibit PCSK9 function. In Aim 3, we will identify and validate as yet unknown regulators of PCSK9 endocytic trafficking via an unbiased, genome-wide CRISPR interference screen. These results will help mechanistically define how PCSK9 is trafficked to the lysosome, provide answers to why PCSK9 only affects certain tissues, and offer potential novel therapeutic targets which would be anticipated to have similar therapeutic windows to PCSK9 itself. Overall, we expect our study to provide key information on the safety of long-term PCSK9 inhibition, lay the groundwork for novel treatments, and identify new therapeutic targets against atherosclerosis.

家长资助项目摘要/摘要 血清低密度脂蛋白（LDL）会导致动脉粥样硬化性心脏病。由于 LDL 受体 (LDLR) 肝脏清除血液中的低密度脂蛋白，上调肝脏低密度脂蛋白受体可减少低密度脂蛋白和心血管事件。这 自裂解蛋白酶 PCSK9（前蛋白转化酶枯草杆菌蛋白酶/kexin 9 型）是经过验证的治疗靶点；它 陪伴 LDLR 进行溶酶体降解，下调其功能。 PCSK9 抗体较低 LDL 可以改善临床结果，但成本和管理要求表明需要替代方案。 肝脏靶向 siRNA 也能显着降低 LDL，但与耐受性良好的基因变异不同，它消除了所有 PCSK9 来自细胞，引发安全担忧。 PCSK9 对肝脏影响的机制基础 LDLR 已广为人知，但其在该经典途径之外的功能尚不清楚。因此，总体 我们提案的目标是开发生化工具来机械地剖析 PCSK9 的生物学，以便 1) 预测当前抗 PCSK9 疗法的长期效果，2) 开发原理验证 新颖的 PCSK9 靶向策略。为此，我们确定了三种有前途的先导化合物 特异性靶向细胞内 PCSK9。这些化合物既抑制 PCSK9 自身蛋白水解，又 上调 LDLR 表达。在目标 1 中，我们将对这些化合物进行化学优化，确认其机理 确定它们的作用，并在体内模型中验证它们对 PCSK9 和 LDLR 的功能。这将 建立新的化学探针，可用于研究鲜为人知的细胞内 PCSK9 功能， 以及作为上调 LDLR 的替代小分子疗法的起点。我们 还发现其他因素，包括硫酸乙酰肝素蛋白聚糖 (HSPG)，与 PCSK9 相互作用 调节其向溶酶体的运输，表明该途径中还有其他药物靶标。在 目标 2，我们将评估 HSPG 作为 PCSK9 内吞转运调节剂的贡献 LDLR 下调，无论是在体外还是在人体中。这些结果将阐明肝素是否 已经批准的疗法可以重新用于抑制 PCSK9 功能。在目标 3 中，我们将确定并 通过公正的全基因组 CRISPR 验证 PCSK9 内吞运输的未知调节因子 干扰屏。这些结果将有助于机械地定义 PCSK9 如何运输到溶酶体， 提供为什么 PCSK9 只影响某些组织的答案，并提供潜在的新治疗靶点 预计与 PCSK9 本身具有相似的治疗窗口。总的来说，我们期望我们的研究 提供有关长期 PCSK9 抑制的安全性的关键信息，为新疗法奠定基础， 并确定抗动脉粥样硬化的新治疗靶点。