Discovery of ligands for directed-differentiation of stem cells

干细胞定向分化配体的发现

• 批准号: 8240375
• 负责人: KIT S LAM
• 金额: $ 19.21万
• 依托单位: UNIVERSITY OF CALIFORNIA AT DAVIS
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-12-15 至 2013-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8240375
• 关键词: Amino Acids; Apoptotic; Binding; Biochemical; Biological; Bone Marrow; Cancer Cell Growth; Cancer cell line; Cell Adhesion; Cell Surface Receptors; Cell surface; Cells; Chemicals; Collagen; Differentiation and Growth; Engineering; Extracellular Matrix; Extracellular Matrix Proteins; Fibronectins; Flow Cytometry; Focal Adhesion Kinase 1; Glycopeptides; Goals; Growth; Growth Factor; Homing; Human; Hydrogels; Image; Integrin Binding; Knock-in Mouse; Label; Laminin; Libraries; Life; Ligands; Maintenance; Mediating; Methods; Mitogen-Activated Protein Kinases; Mus; Nature; Neurons; Oligodendroglia; Peptide Fragments; Peptide Phage Display Library; Peptides; Play; Property; Proteins; Proteolysis; RGD (sequence); Reporter; Research; Resistance; Role; Screening procedure; Signal Transduction; Solid; Stem Cell Research; Stem cells; Stromal Cells; Structure; Surface; Therapeutic Human Experimentation; base; cancer cell; cell behavior; cell growth; cell type; combinatorial; combinatorial chemistry; design; human embryonic stem cell; in vivo; induced pluripotent stem cell; nerve stem cell; nervous system disorder; novel; oligodendrocyte precursor; osteogenic; peptidomimetics; pluripotency; precursor cell; protein aminoacid sequence; receptor; research study; scaffold; small molecule; stem; stem cell differentiation; tool

DESCRIPTION (provided by applicant): In addition to discovering soluble small molecule for hESC/iPSC maintenance and directed differentiation, combinatorial chemistry can also serve as an excellent tool for the identification of extracellular matrix (ECM) mimics that can further facilitate the maintenance or specific- differentiation of hESC/iPSCs. The one-bead-one-compound (OBOC) combinatorial chemistry invented by Dr. Lam is particularly suited for the discovery of ECM mimics or other functional ligands against unique cell surface receptors of stem cells. We have recently developed a one- bead-two-compound (OB2C) library method by incorporating a known cell adhesion peptide (e.g. an integrin binding ligand) on the outer layer of every bead of the OBOC library. As a result, every bound cell will be exposed to the library compound displayed on the surface of each bead. We have generated genetically-labeled Olig2-GFP knock-in hESC/iPSC reporter lines that can be conveniently used as readouts in the proposed combinatorial library screening for directed-differentiation toward various lineages. Specific aims of this proposal are: 1. To design and synthesize several OBOC and OB2C combinatorial libraries for the discovery of cell surface acting molecules that can induce directed-differentiation of hESCs or iPSCs. 2. To use live Olig2-GFP knock-in reporter hESC/iPSC lines in conjunction with immunocytochemical methods to screen various OB2C libraries (from aim 1) for directed-differentiation into oligodendroglial progenitor cells. 3. To fully characterize effects of the ligands and molecules identified in aim 2 on hESC/iPSC growth and differentiation using a range of biological and biochemical approaches. To optimize and use these ligands in a novel chemically-defined self- assemble PVA-based hydrogel as artificial ECM to support stem cell growth and directed- differentiation into oligodendroglial progenitor cells [OPCs]. Impact The proposed research will enable us to discover synthetic molecules that bind to unique receptors on the surface of hESC/iPSCs. Some of these molecules may support the growth of hESC/iPSCs while maintaining its "stem-ness". Others may induce specific cell signaling and oligodendrocyte differentiation. Our long term goal is to direct-differentiate oligodendrocyte precursor cells further for the treatment of neurological diseases. PUBLIC HEALTH RELEVANCE: In addition to discovering soluble small molecule for hESC/iPSC maintenance and directed differentiation, combinatorial chemistry can also serve as an excellent tool for the identification of extracellular matrix (ECM) mimics that can further facilitate the maintenance or specific- differentiation of hESC/iPSCs. The one-bead-one-compound (OBOC) and one-bead-two- compound (OB2C) combinatorial chemistry invented by Dr. Lam is particularly suited for the discovery of ECM mimics or other functional ligands against unique cell surface receptors of stem cells. Genetically-labeled Olig2-GFP knock-in hESC/iPSC reporter lines will be used as readouts in the proposed combinatorial library screening for directed-differentiation into oligodendroglial progenitor cells.

描述（由申请人提供）：除了发现用于HESC/IPSC维护和定向分化的可溶性小分子外，组合化学还可以作为鉴定细胞外基质（ECM）模拟物的绝佳工具，可以进一步促进HESC/IPSC的维持或特定差异。 LAM博士发明的单珠一个组合（OBOC）组合化学特别适合发现ECM模拟物或其他功能性配体针对干细胞的独特细胞表面受体。我们最近通过在OBOC库的每个珠子的外层上掺入已知的细胞粘附肽（例如整联蛋白结合配体），开发了一种单珠 - 两种化合物（OB2C）库方法。结果，每个绑定的单元将暴露于每个珠子表面上显示的库化合物。我们已经生成了遗传标记的Olig2-GFP敲入HESC/IPSC报告基因线，可以方便地用作建议的组合库筛选中的读数，以定向划分各种谱系。该提案的具体目的是：1。设计和合成几个OBOC和OB2C组合库，以发现可以诱导hESCS或IPSC的指示分化的细胞表面作用分子。 2。将活的Olig2-GFP敲入报告基因hESC/IPSC系列与免疫细胞化学方法结合使用，以筛选各种OB2C文库（来自AIM 1），将定位分化为寡头祖细胞。 3。用一系列生物学和生化方法完全表征AIM 2中鉴定的配体和分子对HESC/IPSC生长和分化的影响。为了在一种新型的化学定义的基于PVA的水凝胶作为人工ECM中优化和使用这些配体，以支持干细胞的生长并将其定向 - 分化为寡胶质祖细胞[OPC]。影响拟议的研究将使我们能够发现与HESC/IPSC表面上独特受体结合的合成分子。这些分子中的一些可以支持hESC/IPSC的生长，同时保持其“干性”。其他可能诱导特定的细胞信号传导和少突胶质细胞分化。 我们的长期目标是将少突胶质细胞前体细胞进一步指导以治疗神经系统疾病。 公共卫生相关性：除了发现用于hESC/IPSC维护和定向分化的可溶性小分子外，组合化学还可以作为鉴定细胞外基质（ECM）模拟的绝佳工具，可以进一步促进HESC/IPSC的特定差异。 Lam博士发明的单珠单螺旋（OBOC）和一孔 - 两种化合物（OB2C）组合化学反应特别适合于发现ECM模拟物或其他功能性配体针对干细胞的独特细胞表面受体。遗传标记的Olig2-GFP敲入HESC/IPSC报告基因将用作拟议组合库筛选中的读数，以将二分化为寡头胚胎祖细胞进行定位分化。