Investigation of ATV-Based Heterobifunctional Degraders to Combat Growing HIV-1 PR Inhibitor Resistance

研究基于 ATV 的异双功能降解剂以对抗日益增长的 HIV-1 PR 抑制剂耐药性

• 批准号: 10676954
• 负责人: CHRISTINA OCHSENBAUER
• 金额: $ 19.64万
• 依托单位: UNIVERSITY OF ALABAMA IN TUSCALOOSA
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-08-04 至 2025-07-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10676954
• 关键词: Affinity; Antiviral Agents; Atazanavir; Autoimmune Diseases; Binding; Binding Proteins; Binding Sites; Biological; Biological Assay; Biological Availability; Biology of HIV Infection; Cell membrane; Cells; Clinic; Clinical Trials; Complex; Computing Methodologies; Development; Dimerization; Drug resistance; Epidemic; Event; Exhibits; Failure; Foundations; Future; Goals; HIV; HIV Infections; HIV Protease; HIV Protease Inhibitors; HIV resistance; HIV-1; HIV-1 protease; Impairment; Infection; Investigation; Length; Ligands; Ligase; Mediating; Membrane; Modality; Modeling; Molecular Conformation; Multi-Drug Resistance; Mutation; Nuclear; Organic Synthesis; Patients; Peptide Hydrolases; Pharmaceutical Chemistry; Pharmaceutical Preparations; Predisposition; Probability; Process; Property; Protease Inhibitor; Proteins; Reagent; Regimen; Resistance; Resistance development; Series; Site; Structure; Testing; Therapeutic; Therapeutic Intervention; Ubiquitination; Variant; Viral; Viral Physiology; Virion; Visit; Work; analog; antimicrobial drug; antiretroviral therapy; assay development; combat; design; dimer; drug development; drug discovery; experience; high reward; high risk; in vitro activity; in vivo; inhibitor; lead optimization; member; membrane assembly; next generation; novel; novel strategies; novel therapeutic intervention; particle; pol Gene Products; pre-exposure prophylaxis; prevent; prospective; prototype; recruit; refractory cancer; resistant strain; screening; simulation; small molecule inhibitor; trend; ubiquitin-protein ligase; viral resistance

PROJECT SUMMARY/ABSTRACT Heterobifunctional targeted protein degraders (TPDs) offer advantages over traditional occupancy-based inhibitors including a unique catalytic mechanism of action (MOA), greater target selectivity, and a reduced probability for resistance development. TPDs are known to effectively target not only cytosolic proteins, but also nuclear and membrane-bound proteins. This therapeutic modality shows great promise for treating drug-resistant cancers and autoimmune diseases but has seen only limited application in antiviral drug discovery. HIV-1 protease (PR) is essential for proteolytic cleavage of Gag and Gag-Pol polyproteins and, thus, virion maturation and infectivity. Gag-Pol forms dynamic dimers at plasma membrane assembly/budding sites, allowing the embedded precursor PR to dimerize as a prerequisite for auto-processing. We identified precursor PR/Gag-Pol as a promising target for protease inhibitor (PI)-based TPDs. The widely used HIV-1 PI, Atazanavir (ATV), is amenable to conjugation with linkers and ubiquitin E3 ligase recruiting ligands to serve as prototype HIV-1 TPDs. Importantly, ATV inhibits activity of mature PR as well as autoprocessing of precursor PR/Gag-Pol during the assembly and budding processes. The OBJECTIVE of this study is to show proof-of-concept that HIV-1 Gag- Pol assembling on the inner leaflet of the plasma membrane can be targeted for aberrant ubiquitination, degradation, and/or inhibition, thereby impairing HIV infectivity. Importantly, due to TPDs’ established MOA, even low-affinity Gag-Pol/TPD interactions are likely to lead to impaired Gag-Pol function. Thus, we pose the HYPOTHESIS that novel ATV-based TPDs will not only augment the inhibition of ATV-sensitive HIV-1 strains compared to ATV, but also exhibit increased and broader biological activity against drug-resistant variants. The objective of AIM 1 is to design and synthesize ATV-based TPDs built on state-of-the-art computational methods and predictive physicochemical properties currently accepted for in vivo active TPDs. Our approach will feature a modular TPD design to establish the appropriate ATV attachment point, E3 ligase recruiter, and length and composition of linker required for HIV-1 Gag-Pol ubiquitination/proteasomal degradation. The goal of AIM 2 is to provide proof-of-concept that ATV-TPDs exert superior activity versus ATV against wild type and PI-resistant HIV-1 strains. We will systematically screen four series of novel ATV-TPD for activity in vitro, in single-round infectivity, and in multi-round replication assays to identify the most promising ATV-TPD candidates. The latter will be tested in complementary limited-scope mechanistic studies, including comparing ATV-TPDs with analogs bearing inactive E3-ligase ligands, to probe if antiviral activity is consistent with the proposed MOA. The IMPACT will be TPDs targeting HIV-1 Gag-Pol and precursor PR that limit infectivity and replication through a mechanism distinct from occupancy-based HIV-1 PI. This will spur the future development of efficacious regimens against PI-resistant HIV strains with reduced susceptibility for resistance development.

项目总结/摘要 异双功能靶向蛋白降解剂（TPD）提供了优于传统的基于占位性的优势， 抑制剂，包括独特的催化作用机制（MOA），更大的目标选择性，和减少 抗性发展的可能性。已知TPD不仅有效地靶向胞质蛋白， 核和膜结合蛋白。这种治疗方式显示出治疗耐药的巨大希望。 癌症和自身免疫性疾病，但在抗病毒药物发现中的应用有限。HIV-1 蛋白酶（PR）对于Gag和Gag-Pol多蛋白的蛋白水解切割以及病毒粒子成熟至关重要 和传染性。Gag-Pol在质膜组装/出芽位点处形成动态二聚体，允许细胞在质膜组装/出芽位点处形成动态二聚体。 嵌入的前体PR二聚作为自动处理的先决条件。我们鉴定了PR/Gag-Pol的前体 作为基于蛋白酶抑制剂（PI）的TPD的有前途的靶标。广泛使用的HIV-1 PI，阿扎那韦（ATV）， 适合与接头和泛素E3连接酶募集配体缀合以用作原型HIV-1 TPD。 重要的是，ATV抑制成熟PR的活性以及前体PR/Gag-Pol的自加工。 组装和出芽过程。本研究的目的是证明HIV-1 Gag- 在质膜的内小叶上组装的Pol可以作为异常泛素化的靶点， 降解和/或抑制，从而损害HIV感染性。重要的是，由于TPD的既定MOA，即使 低亲和力Gag-Pol/TPD相互作用可能导致Gag-Pol功能受损。因此，我们提出 假设基于ATV的新型TPD不仅能增强对ATV敏感的HIV-1毒株的抑制作用， 此外，与ATV相比，它还表现出针对耐药变体的增加的和更广泛的生物活性。 AIM 1的目标是设计和合成基于ATV的TPD， 方法和预测的物理化学性质目前接受的体内活性TPD。我们的方法 将采用模块化TPD设计，以建立适当的ATV附着点，E3连接酶募集， HIV-1 Gag-Pol泛素化/蛋白酶体降解所需接头的长度和组成。目标 目的是提供ATV-TPD相对于ATV对野生型发挥上级活性的概念验证， 抗PI的HIV-1病毒株。我们将系统地筛选四个系列的新型ATV-TPD的体外活性， 单轮感染性，并在多轮复制测定中鉴定最有希望的ATV-TPD候选物。 后者将在补充的有限范围机制研究中进行测试，包括比较ATV-TPD 与带有无活性E3-连接酶配体的类似物，以探测抗病毒活性是否与所提出的MOA一致。 IMPACT将是靶向HIV-1 Gag-Pol和前体PR的TPD，通过以下方式限制感染性和复制： 这是一种不同于基于占有的HIV-1 PI的机制。这将刺激未来的发展， 针对PI耐药HIV毒株的方案，耐药性发展的敏感性降低。