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: 10370428
• 负责人: John S Chorba
• 金额: $ 36.34万
• 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/10370428
• 关键词: Address; Affect; Antibodies; Anticoagulants; Atherosclerosis; Binding; Biological; Biology; Blood Circulation; CRISPR interference; Cardiovascular system; Cell Surface Receptors; Cell surface; Cells; Chemicals; Cholesterol; Clinic; Clinical; Coronary heart disease; Development; Drug Targeting; Engineering; Event; Exhibits; Foundations; Genetic; Goals; Health; Health Services Accessibility; Heparan Sulfate Proteoglycan; Heparin; Hepatic; Hepatocyte; Human; Human Genetics; Impairment; In Vitro; Insurance Carriers; Lead; Liver; Long-Term Effects; Low Density Lipoprotein Receptor; Low-Density Lipoproteins; Lysosomes; Maps; Mediating; Mediator of activation protein; Mission; Molecular; Molecular Chaperones; Mutation; Oral; Outcome; Pathway interactions; Patients; Peptide Hydrolases; Pharmaceutical Preparations; Phenocopy; Phenotype; Physiological; Price; Problem Solving; Proprotein Convertases; Proteolysis; Regulation; Research Design; Resistance; Role; Safety; Serum; Small Interfering RNA; Subtilisins; Therapeutic; Tissues; United States National Institutes of Health; Variant; analog; biochemical tools; blood lipoprotein; cardioprotection; clinical efficacy; cost; cost effective; gain of function; genetic manipulation; genetic variant; genome-wide; improved; improved outcome; in vivo; in vivo Model; inhibitor; innovation; insight; loss of function; new therapeutic target; novel; novel therapeutics; public health relevance; receptor downregulation; receptor expression; scaffold; screening; small molecule; small molecule therapeutics; therapeutic target; tissue tropism; tool; trafficking; uptake

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.

项目摘要/摘要 血清低密度脂蛋白（LDL）引起动脉粥样硬化心脏病。作为LDL受体（LDLR） 肝脏清除血液中的LDL，上调肝LDLR可减少LDL和心血管事件。这 自切割的蛋白酶PCSK9（前蛋白转化酶枯草蛋白/Kexin型9）是经过验证的治疗靶标。它 伴侣的LDLR用于溶酶体降解，下调其功能。针对PCSK9较低的抗体 LDL并改善临床结果，但成本和管理要求说明了替代方案的需求。 以肝靶向的siRNA也可以稳健地降低LDL，但与耐受性良好的遗传变体不同，它去除了所有 细胞中的PCSK9，引起了安全问题。虽然是PCSK9对肝的影响的机械基础 LDLR很了解，其在此规范途径之外的功能还不太清楚。因此，整体 我们建议的目标是开发生化工具，以机械地剖析PCSK9的生物学，以便 1）预期当前抗PCSK9疗法的长期影响和2）开发原则证明 新颖的PCSK9靶向策略。为此，我们确定了三种有希望的铅化合物 特别针对细胞内PCSK9。这些化合物均抑制PCSK9自蛋白解析，也抑制 上调LDLR表达。在AIM 1中，我们将化学优化这些化合物，确认机械 其效果的基础，并在体内模型中验证其对PCSK9和LDLR的功能。这会 建立新的化学探针，可用于研究细胞内PCSK9的鲜为人知的功能， 并用作替代小分子治疗的起点，以上调LDLR。我们 还发现，包括硫酸乙酰肝素蛋白聚糖（HSPG）在内的其他因素与PCSK9相互作用 为了调节其运输到溶酶体，这表明该途径中还有其他药物靶标。在 AIM 2，我们将评估HSPG作为PCSK9内吞贩运的介体的贡献 LDLR下调，无论是在体外还是在人类中。这些结果将阐明是否是肝素 已经批准的疗法可以重新使用以抑制PCSK9功能。在AIM 3中，我们将确定并 通过公正的，全基因组CRISPR验证PCSK9内吞贩运的未知调节剂 干扰屏幕。这些结果将有助于机械地定义PCSK9如何运输到溶酶体，， 提供了为什么PCSK9仅影响某些组织并提供潜在的新型治疗靶标的答案 预计与PCSK9本身具有类似的治疗窗口。总体而言，我们希望我们的研究能 提供有关长期PCSK9抑制安全性的关键信息，为新型治疗奠定基础， 并确定针对动脉粥样硬化的新治疗靶标。