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Co(III) Schiff base complexes as selective and irreversible inhibitors of MMP-2

Co(III) 希夫碱配合物作为 MMP-2 的选择性和不可逆抑制剂

基本信息

批准号：
8512054
负责人：
Thomas J Meade
金额：
$ 7.73万
依托单位：
NORTHWESTERN UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2013
资助国家：
美国
起止时间：
2013-04-01 至 2015-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8512054
关键词：
3-Dimensional Active Sites Adverse effects Antimetastatic Agent Antineoplastic Agents Behavior Binding Biological Biological Assay Cancer cell line Cells Cleaved cell Clinical Trials Cobalt Complex Development Disseminated Malignant Neoplasm Effectiveness Electron Transport Complex III Endopeptidases Enzyme Inhibitor Drugs Enzyme Inhibitors Enzymes Fluorescence Microscopy Gelatinase A Goals Histidine Human Imidazole Immunofluorescence Immunologic In Situ In Vitro Inhibition of Matrix Metalloproteinases Pathway Kinetics Ligands Marimastat Matrix Metalloproteinase Inhibitor Matrix Metalloproteinases Modeling Monitor Musculoskeletal Neoplasm Metastasis Peptide Hydrolases Peptides Proteins Sampling Schiff Bases Specificity Testing Thermodynamics Tissues Transition Elements Vertebral column Zinc absorption analog biological systems cancer cell cytotoxicity improved in vivo inhibitor/antagonist loss of function metastasis prevention migration monolayer novel peptide A protein structure public health relevance screening small molecule tumor tumor microenvironment

项目摘要

DESCRIPTION (provided by applicant): Our goal is to develop a transition metal agent that specifically inhibits matrix metalloproteinase (MMP)-2 by a unique mechanism. Such an agent may have potential use in antimetastatic therapy. A MMP-2 specific peptide substrate will be tethered to a Co(III)-Schiff base (Co(III)-sb) complex that has been previously shown to irreversibly inhibit histidine (His)-containing enzymes. The MMP-2 targeting peptide will impart specificity for MMP-2 over other enzymes and bring the Co(III)-sb complex to close proximity of the target protein such that selective inhibition of MMP-2 occurs. The peptide conjugate is expected to show low levels of off-target effects while maintaining effective inhibition of the target enzyme. Peptides known to be selectively cleaved by MMP-2 over other MMPs will be synthesized and tethered to Co(III)-sb (Co(III)-pep), and fluorescent analogues (Co(III)-flupep) will be synthesized to allow monitoring of their action and localization in biological systems usin fluorescence microscopy. The effectiveness of Co(III)-pep and Co(III)-flupep as a MMP-2-specific inhibitor will be evaluated by investigating kinetic and thermodynamic interactions between the inhibitor and enzyme. In addition, the specificity of Co(III)-pep and Co(III)-flupep fo MMP-2 over other enzymes will be investigated by screening them against a range of MMPs and non-MMP enzymes using inhibition assays and zymography. The binding and inhibitory activity of Co(III)-pep and Co(III)-flupep will be compared to the non-targeted Co(III)-sb complex to evaluate the suitability of targeted Co(III) complexes as MMP-2 specific inhibitors. Further, the effect of Co(III)-pep on the invasion and migration of various cancer cell lines will be validated by testing in biological systems. Migration and invasion assays on cell monolayers and 3-dimensional tumor models such as spheroids will be carried out to evaluate the effect of the peptide conjugates in inhibiting metastatic activity of the cell. Immunofluorescence assays will be performed to ascertain the extent of MMP-2 inhibition in spheroid sections. Upon completion of the proposed in vitro biological studies and if successful results are obtained, we believe this novel inhibitor can be utilized as an antimetastatic agent in vivo.

描述(申请人提供)：我们的目标是开发一种过渡金属试剂，通过一种独特的机制特异性地抑制基质金属蛋白酶(MMPs)-2。这种药物可能在抗转移治疗中有潜在的用途。基质金属蛋白酶-2的特异性多肽底物将被连接到钴(III)-席夫碱(Co(III)-Sb)络合物上，该络合物先前已被证明不可逆转地抑制含有组氨酸(His)的酶。MMP2靶向多肽将赋予MMP2比其他酶更高的特异性，并使Co(III)-Sb络合物更接近目标蛋白，从而选择性地抑制MMP2。这种多肽结合物有望在保持对靶酶的有效抑制的同时，表现出低水平的非靶标效应。已知的被基质金属蛋白酶-2选择性切割的多肽将被合成并连接到Co(III)-Sb(Co(III)-PEP)上，以及荧光类似物(Co(III)-flpep)将被合成，以便利用荧光显微镜监测它们在生物体系中的作用和定位。Co(III)-PEP和Co(III)-FLUPEP作为基质金属蛋白酶-2特异性抑制剂的有效性将通过研究抑制剂与酶之间的动力学和热力学相互作用来评估。此外，Co(III)-PEP和Co(III)-FLUPEP对基质金属蛋白酶-2的特异性将通过抑制试验和酶谱分析对它们进行筛选，以研究它们相对于其他酶的特异性。将Co(III)-PEP和Co(III)-FLUPEP的结合和抑制活性与非靶向的Co(III)-Sb络合物进行比较，以评估靶向Co(III)络合物作为基质金属蛋白酶-2特异性抑制剂的适用性。此外，Co(III)-PEP对各种癌细胞系侵袭和迁移的影响将通过生物系统的测试来验证。将在细胞单层和三维肿瘤模型(如球体)上进行迁移和侵袭分析，以评估多肽结合物在抑制细胞转移活性方面的效果。免疫荧光分析将确定球体切片中基质金属蛋白酶-2的抑制程度。在完成拟议的体外生物学研究后，如果获得成功的结果，我们相信新型抑制剂可作为体内的抗肿瘤药物。