High-Throughput Screens to Discover Novel Inhibitors of Leaky RyR2 for Heart Failure Therapy

高通量筛选发现用于心力衰竭治疗的漏性 RyR2 新型抑制剂

• 批准号: 10064096
• 负责人: Donald M Bers
• 金额: $ 75.42万
• 依托单位: UNIVERSITY OF MINNESOTA
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-01-15 至 2022-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10064096
• 关键词: Academia; Action Potentials; Acute; Adult; Adverse effects; Affinity; American; Animal Model; Animals; Arrhythmia; Back; Basic Science; Binding; Biochemical; Biological Assay; Biosensor; Calcium; Calmodulin; Cardiac; Cardiac Myocytes; Catecholaminergic Polymorphic Ventricular Tachycardia; Cell model; Chemicals; Chronic; Clinical; Collection; Confocal Microscopy; Coupled; Critical Pathways; Dantrolene; Data; Defect; Detection; Development; Diastole; Diastolic heart failure; Drug Incompatibility; Drug Targeting; Drug usage; Electrophysiology (science); Ensure; FKBP1B gene; Family suidae; Fluorescence; Fluorescence Resonance Energy Transfer; Future; Goals; Health; Heart; Heart Diseases; Heart failure; Human; Industrialization; Industry; Label; Lead; Letters; Libraries; Link; Malignant hyperpyrexia due to anesthesia; Measures; Mediating; Membrane; Methods; Modeling; Molecular; Molecular Conformation; Molecular Target; Muscle Cells; Outcome; Oxidative Stress; Pathologic; Pathology; Patients; Peptides; Pharmaceutical Chemistry; Pharmaceutical Preparations; Phenotype; Proteins; Reader; Research; RyR1; RyR2; Ryanodine; Ryanodine Receptor Calcium Release Channel; Sarcoplasmic Reticulum; Signal Transduction; Skeletal Muscle; Structure; System; Tacrolimus Binding Proteins; Technology; Testing; Therapeutic; Toxic effect; Translating; Translations; Treatment Failure; Ventricular; Vesicle; analog; base; calmodulin-dependent protein kinase II; drug development; drug discovery; drug repurposing; experience; high throughput screening; in vivo; induced pluripotent stem cell; inhibitor/antagonist; insight; notch protein; novel; novel therapeutics; screening; tool; uptake

Project Summary Our long-term goal is to develop drugs that target the cardiac sarcoplasmic reticulum (SR) Ca release channel (ryanodine receptor, RyR2) for heart failure (HF) and arrhythmia therapy. RyR2 Ca release is a key player in regulating cardiac contraction, electrophysiology, energetics and signaling. Abnormally high diastolic SR Ca “leak” via RyR2, and reduced SR Ca uptake, conspire to reduce SR Ca content and elevate diastolic [Ca]i, hallmarks of both systolic and diastolic HF. Inappropriately timed SR Ca leak is also arrhythmogenic. Thus RyR2 is widely recognized as a molecular target with excellent therapeutic potential for HF and some arrhythmias. Indeed, some repurposed drugs provide proof-of-principle for this concept. To greatly accelerate discovery of drugs that target the RyR2 leak, we propose the first high-throughput screening (HTS) methods using our well-established FRET-based RyR-targeting system and extensive supporting basic research. Pathology-associated RyR leak is associated with two phenotypic features that are sensitive to RyR conformation – reduced calmodulin (CaM) binding and increased binding of a biosensor peptide (DPc10). We find that SR Ca leak can be reversed by either forced CaM binding or dantrolene (a drug used for acute RyR1 leak in malignant hyperthermia). Dantrolene is unsuitable for chronic use, so we seek novel drugs that restore normal CaM and DPc10 affinity (RyR conformation) and thus inhibit pathological SR Ca leak. We have established direct FRET-based assays of CaM and DPc10 binding to RyR2, and a novel fluorescence lifetime plate-reader enables the translation of these FRET tools into ultrasensitive assays of RyR conformation and interactions with binding partners, in HTS format. Results from pilot screens demonstrate that we are poised to carry out an explicit drug-discovery campaign to detect pathophysiological RyR2 conformations and identify compounds that restore normal RyR2 conformation and function, thus translating our mechanistic research into therapies. Identification of lead compounds from this HTS platform and medicinal chemistry development of analogues will be focused through secondary screens that measure RyR activity in SR membranes, and cellular toxicity and Ca leak in patient-derived iPSC cardiomyocytes and in animal- derived adult ventricular myocytes (normal and HF). Feasibility is ensured by: (1) a robust and sensitive FRET system to specifically resolve RyR structural changes, (2) demonstrated experience applying this FRET system for RyR1-targeted HTS, (3) a novel high-precision FLT-PR, (4) functional insight from parallel hypothesis- driven mechanistic myocyte and animal studies, and (5) top-notch team of MPIs and collaborators. The central hypothesis – that binding of CaM and DPc10 to RyR2 are key markers of RyR2 pathology – will be tested in the following Specific Aims: (1) Screen a collection of 50k-350k compounds, and (2) Determine Hit effects on RyR2-mediated calcium leak in control and HF myocytes. Outcomes will include new HTS assays, a model for large-scale HTS campaigns, and novel compounds that may be developed into new drugs or RyR2 probes.

项目总结

项目成果

Regulation of cardiac calcium signaling by newly identified calcium pump modulators.

新发现的钙泵调节剂对心脏钙信号的调节。

• DOI: 10.1016/j.bbrc.2023.149136
• 发表时间: 2023
• 期刊: Biochemical and biophysical research communications
• 影响因子: 3.1
• 作者: Bovo,Elisa;Rebbeck,RobynT;Roopnarine,Osha;Cornea,RazvanL;Thomas,DavidD;Zima,AlekseyV
• 通讯作者: Zima,AlekseyV

Human iPSC modeling of heart disease for drug development.

• DOI: 10.1016/j.chembiol.2021.02.016
• 发表时间: 2021-03-18
• 期刊: Cell chemical biology
• 影响因子: 8.6
• 作者: Hnatiuk AP;Briganti F;Staudt DW;Mercola M
• 通讯作者: Mercola M

ent-Verticilide B1 Inhibits Type 2 Ryanodine Receptor Channels and is Antiarrhythmic in Casq2 -/- Mice.

ent-Verticilide B1 抑制 Casq2 -/- 小鼠的 2 型 Ryanodine 受体通道并具有抗心律失常作用。

• DOI: 10.1124/molpharm.123.000752
• 发表时间: 2024
• 期刊: Molecular pharmacology
• 影响因子: 3.6
• 作者: Gochman,Aaron;Do,TriQ;Kim,Kyungsoo;Schwarz,JacobA;Thorpe,MadelaineP;Blackwell,DanielJ;Ritschel,PaxtonA;Smith,AbigailN;Rebbeck,RobynT;Akers,WendellS;Cornea,RazvanL;Laver,DerekR;Johnston,JeffreyN;Knollmann,BjornC
• 通讯作者: Knollmann,BjornC

New N-aryl-N-alkyl-thiophene-2-carboxamide compound enhances intracellular Ca2+ dynamics by increasing SERCA2a Ca2+ pumping.

新型 N-芳基-N-烷基-噻吩-2-甲酰胺化合物通过增加 SERCA2a Ca2 泵送来增强细胞内 Ca2 动力学。

• DOI: 10.1016/j.bpj.2022.12.002
• 发表时间: 2023
• 期刊: Biophysical journal
• 影响因子: 3.4
• 作者: Nikolaienko,Roman;Bovo,Elisa;Yuen,SamanthaL;Treinen,LevyM;Berg,Kaja;Aldrich,CourtneyC;Thomas,DavidD;Cornea,RazvanL;Zima,AlekseyV
• 通讯作者: Zima,AlekseyV

The selective RyR2 inhibitor ent-verticilide suppresses atrial fibrillation susceptibility caused by Pitx2 deficiency.

• DOI: 10.1016/j.yjmcc.2023.04.005
• 发表时间: 2023-04
• 期刊: Journal of molecular and cellular cardiology
• 影响因子: 5
• 作者: Kyungsoo Kim;Daniel J. Blackwell;Samantha L. Yuen;Madelaine P Thorpe;Jeff N Johnston;Răzvan L. Cornea;B. Knollmann