High Throughput Screening (HTS) to Discover Graft-Versus-Host Disease Inhibitors

高通量筛选 (HTS) 发现移植物抗宿主疾病抑制剂

• 批准号: 8474927
• 负责人: Sophie Paczesny
• 金额: $ 33.76万
• 依托单位: INDIANA UNIV-PURDUE UNIV AT INDIANAPOLIS
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-04-08 至 2016-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8474927
• 关键词: Acute Graft Versus Host Disease; Affect; Allogenic; Asthma; Automobile Driving; Binding; Binding Sites; Biological; Biological Assay; Biological Factors; Biological Markers; Biological Models; Blood; Chemicals; Clinic; Clinical; Collection; Complication; Computer Simulation; Data; Detection; Development; Disease; Docking; Drug Targeting; Effector Cell; Enzyme-Linked Immunosorbent Assay; Equilibrium; Evaluation; Exhibits; Foundations; Functional disorder; Goals; Heart failure; Hematological Disease; Hematopoietic Stem Cell Transplantation; Human; Immune response; Immunotherapeutic agent; In Vitro; Lead; Libraries; Life; Ligands; Malignant - descriptor; Malignant Neoplasms; Mediating; Modeling; Mus; Outcome; Pathology; Pathway interactions; Patients; Pharmaceutical Preparations; Phenotype; Plasma; Play; Pre-Clinical Model; Property; Proteins; Publishing; Research; Role; Signal Transduction; Steroid therapy; Steroids; Structure; Symptoms; T-Lymphocyte; Technology; Testing; Th2 Cells; Therapeutic; Therapeutic Intervention; Toxic effect; Tumorigenicity; United States National Institutes of Health; Validation; Work; base; cancer therapy; extracellular; graft vs host disease; high throughput screening; in vitro Model; in vivo; inhibitor/antagonist; innovation; insight; molecular dynamics; mortality; new therapeutic target; novel; overexpression; pharmacophore; public health relevance; receptor; response; simulation; small molecule; small molecule libraries; tandem mass spectrometry; therapeutic development; therapeutic evaluation; therapeutic target

DESCRIPTION (provided by applicant): Allogeneic hematopoietic stem cell transplantation (HSCT) is a potentially curative therapy for many malignant diseases but its clinical utility has been impeded by graft-versus-host disease (GVHD). Unfortunately, there are currently no therapies that target proteins that are elevated in the blood of patients presenting with GVHD, and no effort has been made to develop drugs specific for GVHD. This represents an important problem, because currently therapies are limited to the nonspecific targeting of effector cells. Our long-term goal is to develop drugs targeting proteins that are dysregulated in patients with GVHD as potential novel GVHD therapies. Our objective in this application is to discover chemical probes that can inhibit these elevated proteins through high throughput screening (HTS) of small molecules libraries and be evaluated in the biological model systems. These compounds will allow for discovery of potential GVHD-specific drugs and therefore better harnessing of treatment in many patients with hematological cancers. Our central hypothesis is that the lead plasma biomarker of GVHD, suppression of tumorigenicity 2 (ST2) can be targeted with small molecule compounds to alleviate GVHD. This hypothesis was formed based on our published data characterizing the soluble form of ST2 (sST2) as the most significant biomarker to predict the non-response to GVHD therapy. sST2 acts as a decoy receptor for IL-33, the only known ligand for ST2, driving Th2 cells toward a Th1 phenotype. This phenotypic change leads to GVHD, and an inhibitor of the binding association between sST2 and IL-33 could alleviate GVHD. We therefore embark on a pilot HTS campaign using the AlphaScreen detection assay to discover chemical probes that inhibit the ST2 and IL33 interaction. Our optimized assay condition was robust giving z'-factors e 0.7 and permitted discovery of 17 hits. The rationale for this study is that the likelihood for better probes than those in the pilot screen warrants a large screen. In addition, a platform including in silico enrichment of potential hits from broader compound libraries and testing of the chemical probes in biological models of GVHD will be relevant. The three specific aims are: 1) HTS implementation to discover chemical probes that inhibit the ST2/IL33 pathway that is overexpressed in graft-versus-host disease. 2) Enrich hits using focus compound libraries generated from pharmacophore models based on confirmed hits from HTS and identification of putative binding sites in ST2. 3) Hits validation with in vitro huma assays and in vivo murine models of GVHD. This integrative approach is innovative because it applies HTS technology of 176,000 diverse small molecules with in silico enrichment of hits and target binding site evaluation yielding the identification and development of chemical probes against biomarkers of GVHD refractoriness to therapy. The proposed research is significant because discovery of GVHD-specific inhibitors will provide the foundation for therapeutic evaluation of GVHD biomarkers as druggable targets. The application of these drugs will drive the phenotypic changes in appropriate effector T cells with increased efficacy and lowered toxicity.

描述（由申请人提供）：同种异体造血干细胞移植（HSCT）是许多恶性疾病的潜在治疗方法，但其临床应用受到移植物抗宿主病（GVHD）的阻碍。不幸的是，目前还没有针对 GVHD 患者血液中升高的蛋白质的治疗方法，也没有努力开发针对 GVHD 的特异性药物。这是一个重要的问题，因为目前的治疗仅限于效应细胞的非特异性靶向。我们的长期目标是开发针对 GVHD 患者失调蛋白的药物，作为潜在的新型 GVHD 疗法。我们在此应用中的目标是通过小分子文库的高通量筛选 (HTS) 发现可以抑制这些升高的蛋白质的化学探针，并在生物模型系统中进行评估。这些化合物将有助于发现潜在的 GVHD 特异性药物，从而更好地利用许多血液癌患者的治疗。我们的中心假设是，GVHD 的主要血浆生物标志物——致瘤性 2 (ST2) 的抑制可以用小分子化合物来缓解 GVHD。这一假设是基于我们发表的数据形成的，该数据将可溶性 ST2 (sST2) 描述为预测 GVHD 治疗无反应的最重要的生物标志物。 sST2 充当 IL-33（唯一已知的 ST2 配体）的诱饵受体，驱动 Th2 细胞向 Th1 表型发展。这种表型变化会导致 GVHD，而 sST2 和 IL-33 之间结合的抑制剂可以减轻 GVHD。因此，我们开始使用 AlphaScreen 检测分析进行 HTS 试点活动，以发现抑制 ST2 和 IL33 相互作用的化学探针。我们优化的检测条件非常稳健，z' 因子 e 为 0.7，并允许发现 17 个匹配项。这项研究的基本原理是，比试点屏幕中的探头更好的探头可能需要大屏幕。此外，一个平台包括从更广泛的化合物库中进行潜在命中的计算机富集以及 GVHD 生物模型中化学探针的测试也将是相关的。这三个具体目标是：1）实施 HTS 来发现抑制移植物抗宿主病中过度表达的 ST2/IL33 途径的化学探针。 2) 使用基于 HTS 确认命中的药效团模型生成的焦点化合物库来丰富命中，并识别 ST2 中推定的结合位点。 3) 通过体外人类试验和 GVHD 体内小鼠模型进行命中验证。这种综合方法具有创新性，因为它应用了 176,000 种不同小分子的 HTS 技术，通过计算机模拟富集命中和靶标结合位点评估，从而识别和开发针对 GVHD 治疗难治性生物标志物的化学探针。拟议的研究意义重大，因为 GVHD 特异性抑制剂的发现将为 GVHD 生物标志物作为药物靶点的治疗评估奠定基础。这些药物的应用将驱动适当效应T细胞的表型变化，从而提高疗效并降低毒性。