Identification of small molecule inhibitors of the DDI2 protease

DDI2 蛋白酶小分子抑制剂的鉴定

• 批准号: 10638837
• 负责人: Gary L. Kleiger
• 金额: $ 73.1万
• 依托单位: VIRGINIA COMMONWEALTH UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-04-10 至 2027-03-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10638837
• 关键词: Achievement; Acquired Immunodeficiency Syndrome; Address; Affect; Apoptosis; Applications Grants; Area; Binding; Biological Assay; Biophysics; Cancer Patient; Catalytic Domain; Cell Survival; Cells; Cellular Assay; Clinic; Clinical Trials; Data; Detection; Development; Disease remission; Dose; Drug Targeting; Dryness; Enzymes; Event; Exhibits; FDA approved; Fruit; Funding; Future; Genes; Goals; Grant; HIV Protease; Human; Immune system; Length; Libraries; Marketing; Measurement; Modality; Multiple Myeloma; Mus; Normal Cell; Pathway interactions; Patient-Focused Outcomes; Patients; Peptide Hydrolases; Play; Powder dose form; Predisposition; Production; Protease Inhibitor; Proteasome Inhibition; Proteasome Inhibitor; Protein Kinase; Proteins; Protocols documentation; Recovery; Reproducibility; Risk; Role; Signal Transduction Pathway; System; Testing; Therapeutic; Transcript; Ubiquitin; Up-Regulation; Validation; Work; Xenograft Model; cancer cell; cancer therapy; cheminformatics; counterscreen; expectation; high throughput screening; immune modulating agents; inhibitor; liquid chromatography mass spectrometry; malignant breast neoplasm; multicatalytic endopeptidase complex; programs; protein degradation; proteostasis; response; scaffold; scale up; screening; side effect; small molecule inhibitor; synergism; targeted cancer therapy; transcription factor; tumor; tumor growth

PROJECT SUMMARY Proteasome inhibitors are currently being used in the clinic against Multiple myeloma. This approach is thought to work at least in part because cancer cells appear to rely more heavily on proteasomes than do normal cells. However, our previous studies showed that proteasome inhibition invokes an adaptive program driven by the transcription factor NRF1 which upregulates proteasome genes resulting in the recovery of proteasome activity, thus limiting the efficacy of this approach. Consistent with this notion, our preliminary data suggest that depletion of NRF1 potentiates the action of proteasome inhibition therapy in a breast cancer xenograft model. Taking advantage of the fact that a protease DDI2 is essential for NRF1 activation, here we propose to identify small molecule inhibitors of this protease that could be used to enhance the efficacy of proteasome inhibitors. Moreover, based on our preliminary data that showed depleting DDI2 in itself retards tumor growth, DDI2 inhibitors could also find use as a single-agent. Based on our pilot screen that demonstrates feasibility and consistent with the goals of PAR-20-271, this 4-year project will pursue the following specific aims. In AIM 1, we will express and purify DDI2 protein and perform high-throughput screens using a protein thermal shift (PTS) assay to identify compounds that bind DDI2. In AIM 2, we will perform hit selection, confirmation and profiling using a panel of secondary assays that includes orthogonal and counter-screen assays as well as biophysical assays. In AIM 3, we will perform hit validation, hit expansion, probe selection and profile the mechanism of action of hits, followed by cellular assays to assess DDI2 target engagement, impact on the DDI2-NRF1 axis and cancer cell apoptosis.

项目摘要 蛋白酶体抑制剂目前用于临床治疗多发性骨髓瘤。这 这种方法被认为至少部分有效，因为癌细胞似乎更依赖于 蛋白酶体比正常细胞。然而，我们以前的研究表明，蛋白酶体 抑制引起由转录因子NRF 1驱动的适应性程序， 上调蛋白酶体基因，导致蛋白酶体活性的恢复，从而限制 这种方法的有效性。与这一观点一致，我们的初步数据表明， NRF 1的缺失增强蛋白酶体抑制疗法在乳腺癌中的作用 异种移植模型利用蛋白酶DDI 2对于NRF 1是必需的这一事实， 激活，在这里，我们建议确定这种蛋白酶的小分子抑制剂， 用于增强蛋白酶体抑制剂的功效。而且根据我们的初步数据 这表明消耗DDI 2本身可以延缓肿瘤生长，DDI 2抑制剂也可以用作 一个特工根据我们的试点屏幕，证明可行性和一致性， 根据PAR-20-271的目标，这个为期四年的项目将追求以下具体目标。在AIM 1中，我们 将表达和纯化DDI 2蛋白，并使用蛋白质进行高通量筛选 热位移（PTS）测定以鉴定结合DDI 2的化合物。在AIM 2中，我们将执行命中 使用一组二级检测进行选择、确认和分析， 和反筛选测定以及生物物理测定。在AIM 3中，我们将执行命中 验证，命中扩展，探针选择和剖析命中的作用机制，随后 通过细胞测定评估DDI 2靶结合、对DDI 2-NRF 1轴的影响以及 癌细胞凋亡