Screen and Design p18 Chemical Probes for Hematopoietic Stem Cell Self-Renewal

用于造血干细胞自我更新的 p18 化学探针的筛选和设计

基本信息

批准号：
8174548
负责人：
Xiang-Qun Xie
金额：
$ 22.53万
依托单位：
UNIVERSITY OF PITTSBURGH AT PITTSBURGH
依托单位国家：
美国
项目类别：
财政年份：
2011
资助国家：
美国
起止时间：
2011-07-01 至 2013-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8174548
关键词：
Affect Behavior Binding Biochemical Biological Assay Bone Marrow CDK4 gene CDK6-associated protein p18 Cell Cycle Cell Therapy Cells Chemicals Chemistry Clinic Collaborations Colony-Forming Units Assay Commit Computer Simulation Cyclin-Dependent Kinase Inhibitor Data Databases Disease Drug Delivery Systems Ectopic Expression Eligibility Determination Equilibrium Figs - dietary Future G1 Phase Goals Grant Harvest Hematological Disease Hematopoietic Stem Cell Transplantation Hematopoietic stem cells Human Immune system In Vitro Laboratories Lead Libraries Malignant - descriptor Malignant Neoplasms Mediating Modification Molecular Target Mus Nature Outcome Patients Pharmaceutical Chemistry Pharmaceutical Preparations Protein Binding Proteins Protocols documentation Published Comment Publishing Recommendation Regenerative Medicine Research Research Design Research Proposals Retroviridae Screening procedure Signal Transduction Site-Directed Mutagenesis Staging Stem Cell Research Stem cell transplant Stem cells Stimulus Study models Synthesis Chemistry Techniques Testing Therapeutic Therapeutic Uses Toxic effect Umbilical Cord Blood Work adult stem cell base design high risk human stem cells improved in vitro Bioassay in vivo inhibitor/antagonist innovation leukemogenesis meetings member novel novel strategies research and development research study response self-renewal small molecule stem cell biology stem cell therapy virtual

项目摘要

DESCRIPTION (provided by applicant): Human hematopoietic stem cells (HSC) transplantation is currently being used as regenerative medicine for the treatment of congenital deficiencies and malignant diseases as well as cancers and other disorders of the blood and immune systems. However, despite all the enthusiasm surrounding HSC biology and therapeutics, the potential of HSC-based therapies has yet to be fully realized. A major roadblock of broader use of the adult stem cells is the limited number of HSC per harvest for therapeutic benefit and their poorly understood expansion and differentiation behavior in response to proliferative stimuli. In vitro expansion of HSC remains a major challenge for wide applications of HSC transplantation for patients. Our studies show that p18, a member of the cyclin-dependent kinase (CDK) inhibitors (CKI), is a potent negative regulator of HSC self- renewal. Thus, we hypothesize that p18 is a unique drug target, and that small molecules capable of blocking p18 function and interfering with p18/CDK6 interactions are likely to be potent drugs for activating HSC self- renewal. Our objective is to screen/identify p18 inhibitors that act by disrupting p18/CDK6 interactions, thus activating HSC self-renewal and increasing the quantity of active stem cells, and to use them as chemical probes for mechanism studies of HSC self-renewal. The feasibility of the proposed innovative research is supported by the proof-of-principle pilot data obtained by well-established research teams that have complementary expertise for the proposed research. Considering the limited throughput capacity of the current HSC bone-marrow culture protocol, we propose first to use our established in silico screening approach for initial screening to generate p18-focused lead sublibraries (Aim 1). We also apply NMR assays to screen/validate and characterize the p18 hits and their binding interactions with the protein in order to generate p18-active subsets (Aim 2A). Also, the small subsets of validated p18-targeting compounds will then be confirmed by extensive HSC functional assays (Aim 2B). Through these, the compounds that are capable of increasing the number of active stem cells in the bone-marrow culture will be identified as leads. The discovered leads are then used as specific chemical probes for studies of p18/CDK6 interactions and signaling mechanisms of the G1-phase of the cell cycle. As a future plan, the identified leads will be further optimized by chemistry modification and SAR medicinal chemistry studies to improve the potency and cell toxicity. Our long- term goal is to identify/design CKI p18-specific small molecule effectors that can either maintain/stimulate self- renewal of hematopoietic stem cells in a predictable manner, and ultimately to develop new drugs for HSC therapies. Achieving this goal will have a significant impact on stem cell drug research development in general. PUBLIC HEALTH RELEVANCE: Narrative: We propose to screen and identify specific small molecule effectors that can stimulate self-renewal of hematopoietic stem cells. The proposed research will offer a promising approach to design new drugs for hematopoietic stem cell therapies and have a significant impact on stem cell drug research and development in the future.

描述（由申请人提供）：人类造血干细胞（HSC）移植目前正在用作治疗先天性缺陷和恶性疾病以及血液和免疫系统的癌症和其他疾病的再生医学。然而，尽管HSC生物学和治疗剂的热情充满了热情，但基于HSC的疗法的潜力尚未充分实现。成人干细胞的广泛使用的主要障碍是，每收获的HSC数量有限，以供治疗益处，并且对增殖性刺激的响应，其较少理解的扩张和分化行为。 HSC的体外扩张仍然是HSC移植对患者的广泛应用的主要挑战。我们的研究表明，P18是细胞周期蛋白依赖性激酶（CDK）抑制剂（CKI）的成员，是HSC自我更新的有效负调节剂。因此，我们假设p18是一个独特的药物靶标，并且能够阻断p18功能并干扰p18/cdk6相互作用的小分子可能是激活HSC自我更新的有效药物。我们的目标是筛选/识别通过破坏p18/cdk6相互作用来起作用的p18抑制剂，从而激活HSC自我更新并增加活性干细胞的数量，并将其用作HSC自我更新机制研究的化学探针。拟议的创新研究的可行性得到了由公认的研究团队获得的原理证明的试点数据，这些研究团队具有互补的专业知识，可用于拟议的研究。考虑到当前的HSC骨骼培养方案的吞吐量有限，我们首先建议将我们在计算机筛选方法中建立的初始筛选以生成以P18为中心的铅sublibraries（AIM 1）。我们还将NMR分析应用于筛选/验证并表征p18命中及其与蛋白质的结合相互作用，以生成p18活性子集（AIM 2A）。同样，经过验证的P18靶向化合物的小亚集将通过广泛的HSC功能测定（AIM 2B）确认。通过这些，能够增加骨髓培养中活性干细胞数量的化合物将被鉴定为铅。然后，发现的铅用作细胞周期G1期P18/CDK6相互作用的研究和信号传导机理的特定化学探针。作为未来的计划，化学修饰和SAR药物化学研究将进一步优化所确定的铅，以提高效力和细胞毒性。我们的长期目标是识别/设计CKI P18特异性的小分子效应子，这些效应子可以以可预测的方式维持/刺激造血干细胞的自我更新，并最终开发用于HSC疗法的新药。实现这一目标将对整个干细胞药物研究开发产生重大影响。公共卫生相关性：叙述：我们建议筛选和识别可以刺激造血干细胞自我更新的特定小分子效应子。拟议的研究将提供一种有前途的方法来设计用于造血干细胞疗法的新药，并对未来的干细胞药物研究和发育产生重大影响。