Targeting telomerase for melanoma therapeutics

靶向端粒酶用于黑色素瘤治疗

• 批准号: 9981663
• 负责人: Emmanuel Skordalakes
• 金额: $ 47.14万
• 依托单位: WISTAR INSTITUTE
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-08-01 至 2022-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9981663
• 关键词: Age; Allosteric Site; Apoptosis; Area; Binding; Binding Sites; Biochemical; Biological; Biological Assay; Boronic Acids; CDKN2A gene; Cancer cell line; Cell Aging; Cell Cycle Arrest; Cell Line; Cell Proliferation; Cells; Cessation of life; Chemicals; Chromosome abnormality; Chromosomes; Crystallization; Cutaneous Melanoma; Data; Development; Diagnosis; Drug Kinetics; Evaluation; Excretory function; Foxes; Functional disorder; Genetic Transcription; Growth; Human; Immunocompetent; Immunotherapy; In Vitro; Lead; Length; Libraries; Ligands; Malignant Neoplasms; Melanoma Cell; Metabolism; Metaphase; Methods; Modernization; Mus; Mutation; Oncolytic; PTEN gene; Pathway interactions; Pharmaceutical Chemistry; Pharmacology; Pharmacotherapy; Positioning Attribute; Prevalence; Property; Proteins; Public Health; Reagent; Recurrence; Reporting; Research; Research Personnel; Resolution; Roentgen Rays; Signal Transduction; Site; Skin Cancer; Specificity; Structure; Structure-Activity Relationship; Surface; TERT gene; Telomerase; Telomerase Inhibitor; Telomerase RNA Component; Telomerase inhibition; Testing; The Wistar Institute; Therapeutic; Therapeutic Index; Thumb structure; Time; United States; Woman; Work; X-Ray Crystallography; Xenograft Model; absorption; analog; base; cancer cell; cancer therapy; cell growth; cell killing; computational chemistry; cytotoxicity; design; experience; improved; in silico; in vivo; insight; melanoma; men; metabolic abnormality assessment; novel; novel strategies; novel therapeutics; promoter; public health relevance; scaffold; small molecule; small molecule inhibitor; statistics; structural biology; structured data; telomere

PROJECT SUMMARY Highly recurrent TERT (the catalytic subunit of telomerase) promoter mutations in human familial and sporadic melanoma lead to a 2-4 fold increase in TERT transcription and telomerase activation, making telomerase an attractive target for melanoma cancer therapies. The development of effective small-molecule inhibitors of telomerase has been hindered by the lack of high-resolution structural data on telomerase. We used the TERT structure determined by X-ray crystallography to screen >500,000 compounds by in silico methods. This approach allowed us to identify a set of small molecules containing a similar scaffold that inhibited the enzymatic activity of telomerase. We have obtained the X-ray co-crystal structure of TERT bound to one of these compounds, revealing a novel and unexpected allosteric binding site, namely the FVYL pocket, located on the surface of the TERT thumb domain. This is the first-ever ligand-TERT co-crystal structure that has been solved to date. Using biochemical assays, we showed that the FVYL pocket binds telomerase RNA (TER), and therefore our initial lead acts by inhibiting the TERT – TER association and telomerase RNP assembly. We have also found compounds that can selectively inhibit the growth of telomerase-positive human melanoma cell lines, but show little growth inhibition of telomerase negative cancer and non-transformed cells in culture. The previous telomerase inhibitor BIBR-1532 was also examined. We determined the BIBR-1532- TERT co-crystal structure, showing that it binds to the same general area, but in a slightly different manner, as do the ligands that we identified by computational prescreening. We also found that BIBR-1532 had substantially less efficacy with a delayed action at killing melanoma cells than does the top hit from our own studies. The aims of this proposal are to (A) use structure-based design methods combined with medicinal chemistry to improve potency for the inhibition of telomerase, (B) evaluate the biochemical activity and specificity for telomerase while investigating the cellular pathways perturbed on telomerase inhibition, and (C) probe the in vivo melanoma oncolytic activity of the telomerase inhibitors. We are uniquely positioned to accomplish the work described because of our ability to iteratively obtain additional ligand-TERT co-crystal structures, and our expertise in the rapid parallel synthesis of new compound libraries, taking advantage of >20,000 reagents and starting materials available onsite (>800 boronic acids). Our research may provide a new approach for the structure-based design of small-molecule inhibitors of telomerase, and may reinvigorate research in the area of small-molecule telomerase inhibitors, eventually resulting in new drugs to treat melanoma alone or in combination with existing chemo- or immunotherapies. The research team is led by an expert in telomerase (Skordalakes) at The Wistar Institute, along with experienced pharma researchers in medicinal (Reitz) and computational chemistry (Reynolds) at the Fox Chase Chemical Diversity Center, Inc.

项目摘要 人类和零星的高度复发性TERT（端粒酶的催化亚基）启动子突变 黑色素瘤导致TERT转录和端粒酶激活增加2-4倍，使端粒酶A 黑色素瘤疗法的有吸引力的靶标。开发有效的小分子抑制剂 端粒酶缺乏高分辨率的结构数据，从而阻碍了端粒酶。我们使用了tert 通过X射线晶体学确定的结构，通过硅方法中的屏幕> 500,000种化合物。这 方法使我们能够确定一组包含类似支架的小分子，该分子抑制了 端粒酶的酶活性。我们已经获得了与其中一个的X射线共晶结构 这些化合物揭示了一个新颖且意外的变构结合位点，即位于FVYL袋 在Tert拇指域的表面上。这是有史以来的第一个配体共晶结构 迄今为止解决了。使用生化测定，我们表明FVYL袋结合了端粒酶RNA（TER）， 因此，我们的初始铅通过抑制TERT - TER关联和端粒酶RNP组装而起作用。 我们还发现可以选择性抑制端粒酶阳性人的生长的化合物 黑色素瘤细胞系，但几乎没有生长抑制端粒酶阴性癌和未转化的细胞 在文化中。还检查了先前的端粒酶抑制剂BIBR-1532。我们确定了Bibr-1532- TERT共晶结构，表明它与同一一般区域结合，但以稍有不同的方式 做我们通过计算预筛选确定的配体。我们还发现Bibr-1532有 杀死黑色素瘤细胞的延迟作用的效率要低得多，而不是我们自己的顶部 研究。该建议的目的是（a）使用基于结构的设计方法与医学结合 化学以提高抑制端粒酶的效力，（b）评估生化活性和 端粒酶的特异性在研究端粒酶抑制作用的细胞途径时，（c） 探测端粒酶抑制剂的体内黑色素瘤溶瘤活性。我们是独特的位置 完成所描述的工作是因为我们有能力获得额外的配体共晶体 结构以及我们在新化合物库的快速并行合成方面的专业知识，利用 > 20,000种试剂和起始材料（> 800个硼酸）。我们的研究可能会提供 用于基于结构的小分子抑制剂的新方法，并可能重振 在小分子端粒酶抑制剂领域的研究，最终导致新药治疗 单独或与现有的化学疗法或免疫疗法结合使用黑色素瘤。研究团队由 Wistar Institute的端粒酶（Skordalakes）专家以及经验丰富的药物研究人员 Fox Chase化学多样性中心的药用（Reitz）和计算化学（Reynolds）