Design of de novo peptides and electrophysiological testing for voltage-gated sodium channel 1.7 inhibition related to chronic pain treatment

与慢性疼痛治疗相关的电压门控钠通道 1.7 抑制的从头肽设计和电生理学测试

• 批准号: 10660928
• 负责人: Brandon John Harris
• 金额: $ 4.04万
• 依托单位: UNIVERSITY OF CALIFORNIA AT DAVIS
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-02-03 至 2024-12-02
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10660928
• 关键词: Action Potentials; Adult; Affinity; American; Benchmarking; Binding; Biological Assay; Biological Products; Biological Response Modifier Therapy; Cardiac; Cells; Charge; Clinical; Communication; Complex; Computational Science; Cryoelectron Microscopy; Data; Development; Disulfides; Drug Design; Electrophysiology (science); Family; Genetic; Heart Arrest; Human; Human Genetics; Hydrophobicity; Industry Collaboration; Integral Membrane Protein; Investigation; Ion Channel; Knowledge; Learning; Libraries; Manuals; Methodology; Methods; Modification; Mutagenesis; Mutate; Neurobiology; Neurosciences; Neurotoxins; Nociception; Pain management; Paralysed; Pathology; Peptides; Peripheral Nervous System; Peruvian; Pharmaceutical Preparations; Positioning Attribute; Protein Engineering; Proteins; Publishing; Research; Resolution; Safety; Scorpions; Seizures; Side; Sodium; Sodium Channel; Software Design; Spiders; Structure; Testing; Therapeutic; Time; Toxin; Training; Translational Research; Validation; Variant; Work; academic preparation; addiction liability; career; chemical stability; chronic pain; chronic pain management; clinical efficacy; cytokine; design; drug candidate; drug discovery; effective therapy; experience; experimental study; falls; improved; knowledgebase; mimetics; nanomolar; neuronal excitability; non-opioid analgesic; novel; pain signal; patch clamp; peptidomimetics; pre-clinical; prescription opioid; preservation; protein structure prediction; rational design; screening; skeletal; skills; small molecule; structural imaging; targeted treatment; thermostability; validation studies; virtual screening; voltage

Project Summary/Abstract Current drug discovery efforts for non-addictive, chronic pain management are targeting human voltage-gated sodium (hNaV) channels: pore-forming transmembrane proteins that evoke the fast action potential in excitable neuronal, cardiac, and skeletal cells. Genetic and preclinical target validation studies have identified hNaV1.7, hNaV1.8, hNaV1.9 channel subtypes as key proteins in pain signaling with emphasis on hNaV1.7 for its predominant expression in the peripheral nervous system. Attempts at selectively targeting hNaV1.7 with pre- clinical small-molecule drugs fall short due to non-selective binding to other hNaV channel subtypes and other ion channel families; non-selective binding can lead to cardiac arrest, paralysis and seizure. Peptide toxins originating from tarantula, spider, and scorpion have been identified as potent hNaV-inhibiting biologics. Notably, Protoxin-II, the neurotoxin from the Peruvian green velvet tarantula, has approximately 1 nM half- maximal inhibitory concentration (IC50) to hNav1.7. However, these peptide toxins also have non-selective binding to hNav1.7. With recent cryo-EM structural images of Protoxin-II bound to voltage-sensing domain II of hNaV1.7, it is now possible to design peptides mimicking Protoxin-II that are selective to hNaV1.7. Advances in de novo protein design methods using Rosetta – a protein structure prediction and design software suite – enable a new avenue of drug design and virtual screening prior to experimental validation. Thus, the project aims to create new peptides inspired from Protoxin-II to selectively target hNaV1.7. The methodology designs new peptide topologies that incorporate the Protoxin-II motif structurally shown to bind to hNaV1.7. The peptides are further optimized to bind to residues unique to hNaV1.7: residues that are not targeted by natural peptide toxins. Upon synthesis of candidate peptides, they will be validated to selectively target hNaV1.7 using whole-cell patch-clamp electrophysiology. The results will improve our mechanistic understanding needed for selective, yet potent hNaV1.7 inhibition. Further, this research will be the first attempt at using a combination of Rosetta de novo protein design methods to create unique peptides selective for hNaV1.7. The applicant upon completion of this project will have a refined expertise of computational protein design methods and receive training in electrophysiology for the first time. Throughout this project, the sponsor and co-sponsor will implement a training plan to strengthen the applicant’s knowledge of neuroscience and drug discovery, while broadening their collaborative network, and sharpening their scientific communication skills. This plan is tailored such that the applicant is prepared for an academic career designing peptide biologics for the treatment and investigation of ion channel pathologies.

项目总结/文摘