Development and characterization of HIV-1 Tat degraders

HIV-1 Tat 降解剂的开发和表征

• 批准号: 10483950
• 负责人: Susana T Valente
• 金额: $ 30万
• 依托单位: THIMBLE THERAPEUTICS, INC.
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-05-01 至 2023-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10483950
• 关键词: Affinity; Animal Model; Anti-HIV Agents; Anti-Retroviral Agents; Apical; Automobile Driving; Binding Sites; Biochemical; Biological Assay; CD4 Positive T Lymphocytes; Calorimetry; Cell Survival; Cell model; Cells; Chemicals; Chimeric Proteins; Clinical; Clinical Trials; Collaborations; Complement; Complex; Confocal Microscopy; DNA; Development; Disease Progression; Docking; Electrophoretic Mobility Shift Assay; Epigenetic Process; Feedback; Florida; Future; Gene Silencing; Genetic Transcription; Genome; Goals; HIV; HIV tat Protein; HIV-1; Hela Cells; Human; In Vitro; Individual; Interruption; Laboratories; Lead; Libraries; Luciferases; Mediating; Metabolic; Modification; Molecular; Molecular Computations; Names; Nuclear; Parents; Pathway interactions; Persons; Pharmaceutical Preparations; Pharmacology; Production; Property; Proviruses; RNA; Research; Research Institute; Role; Shock; Specificity; Structure; Structure-Activity Relationship; System; TNF gene; Techniques; Testing; Tetanus Helper Peptide; Therapeutic; Therapeutic Index; Therapeutic Intervention; Tissues; Titrations; Toxic effect; Transactivation; Transcript; Validation; Viral; Viral reservoir; Virus; Virus Activation; Virus Replication; analog; antiretroviral therapy; base; blood-brain barrier disruption; chemical property; chromatin immunoprecipitation; clinical candidate; commercialization; cortistatin; cost; cost effective; cytotoxicity; detection limit; disease transmission; epigenetic silencing; follow-up; high throughput screening; high-throughput drug screening; humanized mouse; improved; inhibitor; interest; molecular dynamics; mouse model; mutant; neuroinflammation; neurotoxicity; novel; novel drug class; novel strategies; particle; pre-clinical; prevent; promoter; small molecule; targeted treatment; tat Protein; viral rebound; viral resistance

Abstract Despite effective antiretroviral therapy (ART), latent proviruses can reinitiate viral production upon cell stimulation or treatment interruption. The viral Tat protein enhances transcript elongation from the HIV-1 promoter, controlling the switch between latency and active viral production. The block-and-lock functional cure aims at the transcriptional silencing of the viral reservoir, rendering suppressed HIV promoters extremely difficult to reactivate from latency. The Tat inhibitor, didehydro-cortistatin A (dCA) was used to prove this concept. Combining dCA with ART inhibits transcription and blocks viral rebound upon treatment interruption, as the promoter becomes epigenetically repressed. dCA defines a novel class of drugs that can silence and maintain a transcriptionally inactive HIV promoter, offering a novel approach in the treatment of HIV. Tat is very attractive target for therapeutic intervention because: 1) is expressed early during virus replication; 2) no cellular homologs; 3) Tat inhibitors block the feedback loop necessary for viral amplification; 4) epigenetic modifications accumulate at the HIV promoter rendering reactivation less likely. Tat is also known for its role in neurotoxicity, blood-brain barrier disruption, and neuroinflammation. Thus, the immense interest in the development of Tat inhibitors to complement ART. The major hurdle towards advancing dCA into clinical trials is the cost of producing large quantities of this molecule due to its complex structure. Additional clinical candidates, structurally distinct from dCA, that embody equivalent bioactivity are needed in the pre-clinical pipeline. We optimized a cell-based Tat transactivation assay to use in high throughput screening (HTS), with dCA as control. We combined appropriate counter-screens and a wealth of techniques to quickly discard small molecules that are not Tat specific. The HTS of 369,203 compounds was completed by Southern Research (SR), yielding three compounds, TT-44951, TT-44881 and TT-44863, with therapeutic index (TI) varying from 51.2 to 181.1 and good chemical properties. In this application, we propose to perform hit validation and characterization of chemically synthesized analogs generated by Thimble Therapeutics, during the compound progression pathway (CPP), to expand our panel of Tat inhibitors to commercialize this novel class of compounds. We propose the following aims: Specific Aim 1. Validate Tat inhibitors based on disruption of Tat HIV-1 LTR transactivation. Specific Aim 2. Characterize the mechanism of action of selected hits. At the end of this study we expect to: (a) have identified small molecules that will specifically inhibit Tat in cell-based assays; (b) have adequate metabolic stability and PK properties for future pharmacological assessment in animal models and eventually in human clinical trials.

摘要 尽管有效的抗逆转录病毒疗法(ART)，潜伏的前病毒可以在细胞上重新启动病毒的产生 刺激或治疗中断。病毒Tat蛋白增强HIV-1转录本的延长 启动子，控制潜伏和活跃的病毒生产之间的切换。阻止和锁定功能曲线 旨在抑制病毒库的转录沉默，使被抑制的艾滋病毒启动子变得极其困难 从延迟中重新激活。TAT抑制剂二氢皮质酮A(DCA)被用来证明这一概念。 DCA与ART结合可抑制转录，并在治疗中断时阻止病毒反弹，因为 启动子受到表观遗传的抑制。DCA定义了一类新的药物，可以沉默和维持 转录失活的HIV启动子，为HIV的治疗提供了一种新的方法。 TAT是非常有吸引力的治疗干预靶点，因为：1)在病毒复制早期表达； 2)没有细胞同源物；3)TAT抑制剂阻断了病毒扩增所需的反馈回路；4)表观遗传学 修饰累积在HIV启动子上，使得重新激活的可能性降低。TAT还因其在 神经毒性、血脑屏障破坏和神经炎症。因此，人们对世界银行的巨大兴趣 补充抗逆转录病毒疗法的TAT抑制剂的开发。 推动DCA进入临床试验的主要障碍是大量生产这种药物的成本 分子由于其复杂的结构。其他临床候选，在结构上与DCA不同，体现了 在临床前的管道中需要同等的生物活性。 我们优化了一种基于细胞的TAT反式激活试验，用于高通量筛查(HTS)，以DCA为 控制力。我们结合了适当的反筛选和丰富的技术来快速丢弃小分子 不是特定于纹身的。南方研究(SR)完成了369,203个化合物的HTS，产生了 三个化合物，TT-44951，TT-44881和TT-44863，治疗指数(TI)在51.2到181.1之间 和良好的化学性质。在本应用程序中，我们建议执行HIT验证和表征 Thimble Treateutics在复合进展途径中产生的化学合成类似物 (CPP)，以扩大我们的TAT抑制剂产品阵容，使这类新型化合物商业化。我们建议 以下目标： 具体目的1.基于对TAT HIV-1 LTR反式激活的干扰来验证TAT抑制剂。 具体目标2.描述选定命中的作用机制。 在这项研究结束时，我们预计：(A)已经确定了将专门抑制 TAT在基于细胞的分析中；(B)具有足够的代谢稳定性和未来的PK特性 在动物模型中以及最终在人类临床试验中进行的药理学评估。