Optimizing a Stapled-Peptide That Specifically Targets HSV-1 to Treat Herpes Ocular Keratitis

优化专门针对 HSV-1 的钉合肽以治疗疱疹性眼角膜炎

• 批准号: 10650858
• 负责人: ROBERT Paul RICCIARDI
• 金额: $ 99万
• 依托单位: FOX CHASE CHEMICAL DIVERSITY CENTER, INC
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-05-01 至 2025-05-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10650858
• 关键词: 3-Dimensional; Acyclovir; Address; Anatomy; Antiviral Agents; Area; Binding; Biological Assay; Blindness; Cells; Chemicals; Clinical; Cornea; Corneal Ulcer; Crystallization; Cyclic Peptides; DNA; DNA biosynthesis; Development; Disease; Dissociation; Dose; Drug Kinetics; Engineering; Ensure; Epithelial Cells; Evaluation; Exhibits; Exposure to; Eye; Eye Infections; Face; Failure; Formulation; Gene Mutation; Goals; Herpes Labialis; Herpesviridae; Herpesviridae Infections; Herpesvirus 1; Herpetic Keratitis; Histopathology; Human; In Vitro; Individual; Infection; Keratitis; Lead; Liposomes; Measures; Medical; Modeling; Mutagenicity Tests; Oryctolagus cuniculus; Patients; Penetration; Peptide Hydrolases; Peptides; Permeability; Persons; Pharmaceutical Preparations; Physiological; Polymerase; Population; Preclinical Drug Development; Property; Proteins; Publishing; Recurrence; Research; Resistance; Risk; Running; Safety; Series; Solubility; Solvents; Surface; Testing; Therapeutic; Time; Tissues; Toxic effect; Vascularization; Viral; Viral Load result; Virus; Virus Replication; Vision; alpha helix; analog; anti-viral efficacy; corneal epithelium; corneal scar; crosslink; drug resistant virus; efficacy study; efficacy testing; experience; genital herpes; genotoxicity; in vivo; mutant; novel; ocular surface; peptide A; pre-IND studies; preclinical development; prevent; protein protein interaction; residence; risk mitigation; safety testing; scale up; small molecule; standard care; stapled peptide; three dimensional cell culture; thymidine kinase 1; uptake; viral DNA

ABSTRACT Infection of the eye by Herpes Simplex Virus-1 (HSV-1) can result in Herpes Keratitis (HK), which is the leading cause of infectious corneal blindness worldwide. In the U.S., nearly 500,000 individuals experience ocular herpes infections that are often recurrent and culminate in progressive corneal scarring and loss of vision. The gold standard treatment is Acyclovir (ACV) that targets HSV-1 thymidine kinase (TK). However, emergence of ACV viral resistant mutants in 7-14% of ocular HK patients has created the urgent need to discover a second drug directed against a different HSV-1 target. A new class of antiviral targets are processivity factors (PFs) that are essential for tethering their cognate polymerases (Pols) to the template to enable continuous DNA synthesis. During HSV-1 replication, the DNA-anchored PF binds to the extreme C-terminus of Pol (C-Pol) to keep Pol from dissociating from the viral template. When co-crystallized with its PF, the C-terminus of the viral Pol (C-Pol) forms an α-helix, with one face making multiple bonds with several residues of PF while the other face is solvent exposed. Our goal is to develop a novel antiviral drug that specifically targets herpes PF for the purpose of treating ACV resistant HK. Stapled a-helical peptides have emerged for use in targeting protein-protein interactions that often display as long flat surfaces which are difficult for small molecules to bind efficiently. Peptide a-helices are ideal structural motifs for incorporating chemical staples to provide rigidity to make natural bonds with residues on the target protein. Moreover, staples can create a protease shield to prolong residence time. We have now engineered a stapled peptide of C-Pol (SPep7B) that blocks the mechanism of processive DNA synthesis in vitro and HSV-1 infection in human organotypic (3D) corneal epithelial cells; the unstapled peptide is non-inhibitory. SPep7B was shown to eliminate HSV-1 viral DNA in infected cells while failing to block a different virus. Importantly, SPep7B exhibits undetectable toxicity (CC50>100µM) by two different assays in human ocular epithelial cells. Our GOAL is to enter SPep7B into preclinical development as a Topical to treat HK. This will be accomplished by conducting a series of AIMS that include scaleup of SPep7B and evaluating two formulations for delivery of SPep7B into human 3D cultures and then ex-vivo excised rabbit cornea for permeability and toxicity. These studies are essential to confront the physiological and anatomical barriers to topical ocular delivery. In vivo studies will evaluate SPep7B for tolerability, genotoxicity, PK, and repeat dosing. An efficacy study will evaluate SPep7B for clinical cure and viral clearing. Backup analogs of SPep7B will be prepared and evaluated for risk mitigation. A model for new antiviral drugs: Closely aligned with this approach is the potential for using PFs and stapled peptides of other viruses for treating unmet medical needs caused by infections that lead to deleterious complications.

抽象的 单纯疱疹病毒 1 (HSV-1) 感染眼睛可导致疱疹性角膜炎 (HK)，这是 全世界传染性角膜失明的主要原因。在美国，近 50 万人 经历眼部疱疹感染，这种感染通常会反复发作，最终导致进行性角膜疤痕 和视力丧失。金标准治疗是针对 HSV-1 胸苷激酶的阿昔洛韦 (ACV) （传统知识）。然而，7-14% 的 HK 眼病患者出现了 ACV 病毒抗性突变体，这导致了 迫切需要发现另一种针对不同 HSV-1 靶点的药物。一类新的抗病毒药物 目标是持续性因子 (PF)，对于将同源聚合酶 (Pols) 束缚到 模板以实现连续的 DNA 合成。在 HSV-1 复制期间，DNA 锚定的 PF 结合 到 Pol (C-Pol) 的最末端，以防止 Pol 从病毒模板上解离。什么时候 病毒 Pol (C-Pol) 与其 PF 共结晶，C 端形成 α 螺旋，其中一个面形成 与多个 PF 残基形成多重键，而另一面暴露于溶剂中。我们的目标是发展 一种新型抗病毒药物，专门针对疱疹 PF，用于治疗 ACV 耐药性 HK。 钉合的α-螺旋肽已出现用于靶向蛋白质-蛋白质相互作用，这些相互作用通常 显示为长的平坦表面，小分子难以有效结合。肽α螺旋 是结合化学主钉的理想结构基序，以提供形成自然键的刚性 目标蛋白上有残基。此外，钉书钉可以形成蛋白酶屏障以延长停留时间 时间。我们现在已经设计了一种 C-Pol (SPep7B) 的钉合肽，它可以阻断 体外持续性 DNA 合成和人器官型 (3D) 角膜上皮细胞中的 HSV-1 感染； 未钉合的肽是非抑制性的。 SPep7B 被证明可以消除感染者中的 HSV-1 病毒 DNA 细胞同时无法阻止另一种病毒。重要的是，SPep7B 表现出不可检测的毒性 (CC50>100μM) 通过在人眼上皮细胞中进行两种不同的测定。我们的目标是进入 SPep7B 作为治疗 HK 的外用药物进入临床前开发。这将通过进行 系列 AIMS，包括 SPep7B 的放大和评估两种将 SPep7B 递送到体内的制剂 人类 3D 培养物，然后离体切除兔角膜以测定通透性和毒性。这些研究是 对于应对局部眼部给药的生理和解剖学障碍至关重要。体内研究 将评估 SPep7B 的耐受性、遗传毒性、PK 和重复给药。功效研究将评估 SPep7B 用于临床治愈和病毒清除。 SPep7B 的备用类似物将被准备和评估 用于缓解风险。新型抗病毒药物的模型：与这种方法密切相关的是 使用 PF 和其他病毒的钉合肽来治疗未满足的医疗需求的潜力 导致有害并发症的感染。