Cell-free assay technologies for the identification of active compounds

用于鉴定活性化合物的无细胞测定技术

• 批准号: 10926223
• 负责人: Barry Okeefe
• 金额: $ 68.34万
• 依托单位: DIVISION OF BASIC SCIENCES - NCI
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10926223
• 关键词: 2019-nCoV; Biochemical; Biological Assay; Biophysics; CBLB gene; Cells; Chemicals; Collaborations; Cyclic Peptides; DNA; Development; Diagnostic; Enzymes; GPC3 gene; Genes; HIV-1; Immune; Immunologics; Inositol; Manuscripts; Metabolic; Methods; Modification; Mucosa- associated lymphoid tissue lymphoma translocation protein-1; Natural Extract; Natural Products; Natural Products Chemistry; PRKACA gene; Paper; Phosphodiesterase Inhibitors; Phosphotransferases; Preparation; Primary carcinoma of the liver cells; Process; Productivity; Protease Inhibitor; Proteins; Publishing; RNA; Reading; Reporting; Role; Sampling; Source; Structure; Techniques; Technology; Viral; Work; assay development; high throughput screening; inhibitor; interest; phosphoric diester hydrolase; receptor binding; scaffold; screening; small molecule; tripolyphosphate; ubiquitin ligase

The PCMBS collaborates closely with the MTP Assay Development and Screening Section (ADSS/Henrich) to automate, validate and complete high throughput screens (HTS), with the Chemical Diversity and Development Section (CDDS/Beutler) to access new sources of chemical diversity and with the Natural Products Chemistry Section (NPCS/Grkovic, Du) to prioritize extracts for natural product isolation and biochemically evaluate pure active compounds isolated by NPCS or CDDS. Examples of PCMBS efforts during this review period, include published papers on screens of natural products for selective inhibitors of the protease MALT the ubiquitin ligase CBLB, and the phosphodiesterase TDP1. Our studies on natural product inhibitors of TDP1 also resulted in a second paper describing a new class of cyclic peptides which are the first reported allosteric inhibitors of this phosphodiesterase. We have recently also completed a screen for inhibitors of the phosphodiesterase TDP2 with a manuscript in preparation. In addition, the PCMBS has continued to increase the implementation of biophysical screening methods for the identification of specific modulators of the thermal stability of both protein and RNA structures. Initially, this work resulted in the identification of a new small molecule scaffold that altered the stability of HIV-1 TAR RNA. This technology has now been expanded upon to allow for the facile screening of pure compounds and natural product samples in a high throughput format to identify modulators of RNA and protein stability. One recently published example was the identification of a natural product that selectively modulated the thermal stability of pre-miR-21. We have recently completed two new enzymatic screens, against the chimeric kinase DNAJB1-PRKACA (PKADJ) and inositol 1,3,4-triphosphate 5/6 kinase (ITPK1), as well as four biophysical screens; against the immunologically-pertinent metabolic enzyme Immune Responsive Gene 1 (IRG1), the diagnostic target for hepatocellular carcinoma glypican-3 (GPC-3) and the SARS-CoV-2 receptor binding domain (RBD), and DNA-reading protein BRD4. The PCMBS also evaluates compounds derived from MTP screens for their specific interactions against macromolecular targets. We have used both biophysical and biochemical techniques to evaluate inhibitors of pre-miR-21 RNA, TDP1, MALT1,CBLB, and SARS-CoV-2 viral entry.

PCMBS与MTP检测开发和筛选部门密切合作（ADSS/Henrich）与化学品多样性和开发科合作，自动化、验证和完成高通量筛选（HTS）（CDDS/Beutler）以获得化学多样性的新来源，并与天然产品化学科合作，（NPCS/Grkovic，Du）对天然产物分离的提取物进行优先排序，并对NPCS或CDDS分离的纯活性化合物进行生物化学评价。PCMBS在此审查期间的努力的例子，包括已发表的论文筛选天然产物的蛋白酶MALT的泛素连接酶CBLB和磷酸二酯酶TDP 1的选择性抑制剂。我们对天然产物TDP 1抑制剂的研究也导致了第二篇论文，描述了一类新的环肽，这是首次报道的这种磷酸二酯酶的变构抑制剂。我们最近还完成了磷酸二酯酶TDP 2抑制剂的筛选，手稿正在准备中。此外，PCMBS继续增加生物物理筛选方法的实施，以鉴定蛋白质和RNA结构热稳定性的特定调节剂。最初，这项工作导致了一种新的小分子支架的鉴定，改变了HIV-1 TAR RNA的稳定性。该技术现在已经扩展到允许以高通量形式简单筛选纯化合物和天然产物样品，以鉴定RNA和蛋白质稳定性的调节剂。最近发表的一个例子是鉴定了一种天然产物，该产物选择性地调节前体miR-21的热稳定性。我们最近完成了针对嵌合激酶DNAJB 1-PRKACA（PKADJ）和肌醇1，3，4-三磷酸5/6激酶（ITPK 1）的两个新的酶筛选，以及四个生物物理筛选;针对免疫相关代谢酶免疫应答基因1（IRG 1）、肝细胞癌磷脂酰肌醇蛋白聚糖-3（GPC-3）的诊断靶点和SARS-CoV-2受体结合结构域（RBD），和DNA阅读蛋白BRD 4。PCMBS还评估了来自MTP筛选的化合物对大分子靶标的特异性相互作用。我们使用生物物理和生物化学技术来评估前体miR-21 RNA、TDP 1、MALT 1、CBLB和SARS-CoV-2病毒进入的抑制剂。

项目成果

Antiviral lectins: Selective inhibitors of viral entry.

• DOI: 10.1016/j.antiviral.2017.03.007
• 发表时间: 2017-06
• 期刊: Antiviral research
• 影响因子: 7.6
• 作者: Mitchell CA;Ramessar K;O'Keefe BR
• 通讯作者: O'Keefe BR

Dentithecamides A-H, Diacylated Zoanthoxanthin Derivatives with PAX3-FOXO1 Inhibitory Activity from the Hydroid Dentitheca habereri.

Dentithecamides A-H，来自水螅 Dentitheca habereri 的具有 PAX3-FOXO1 抑制活性的二酰化 Zoanthoxanthin 衍生物。

• DOI: 10.1021/acs.jnatprod.2c00246
• 发表时间: 2022
• 期刊: Journal of natural products
• 影响因子: 5.1
• 作者: Jiang,Wei;Tian,Xiangrong;Wang,Dongdong;Bokesch,HeidiR;Thomas,CherylL;Woldemichael,GirmaM;Gryder,BerkleyE;Wei,JunS;Song,YoungK;Chou,Hsien-Chao;Khan,Javed;O'Keefe,BarryR;Gustafson,KirkR
• 通讯作者: Gustafson,KirkR

Suberitamides A-C, Aryl Alkaloids from a Pseudosuberites sp. Marine Sponge that Inhibit Cbl-b Ubiquitin Ligase Activity.

• DOI: 10.3390/md18110536
• 发表时间: 2020-10-28
• 期刊: Marine drugs
• 影响因子: 5.4
• 作者: Kim CK;Wang D;Wilson BAP;Saurí J;Voeller D;Lipkowitz S;O'Keefe BR;Gustafson KR
• 通讯作者: Gustafson KR

Sinularamides A-G, Terpenoid-Derived Spermidine and Spermine Conjugates with Casitas B-Lineage Lymphoma Proto-Oncogene B (Cbl-b) Inhibitory Activities from a Sinularia sp. Soft Coral.

• DOI: 10.1021/acs.jnatprod.1c00367
• 发表时间: 2021-06-25
• 期刊: JOURNAL OF NATURAL PRODUCTS
• 影响因子: 5.1
• 作者: Jiang, Wei;Wang, Dongdong;Wilson, Brice A. P.;Voeller, Donna;Bokesch, Heidi R.;Smith, Emily A.;Lipkowitz, Stanley;O'Keefe, Barry R.;Gustafson, Kirk R.
• 通讯作者: Gustafson, Kirk R.

Rare Caulamidine Hexacyclic Alkaloids from the Marine Ascidian Polyandrocarpa sp.

• DOI: 10.1021/acs.jnatprod.3c00393
• 发表时间: 2023-06-27
• 期刊: JOURNAL OF NATURAL PRODUCTS
• 影响因子: 5.1
• 作者: Tian，Xiangrong;Wang，Dongdong;Gustafson，Kirk R.
• 通讯作者: Gustafson，Kirk R.