New probes for matrix metalloproteinase 13

描述（由申请人提供）：骨关节炎（OA）是关节炎的最常见形式，其特征在于关节软骨的破坏。关节或关节软骨的主要成分是II型胶原和各种蛋白聚糖，如聚集蛋白聚糖、硫酸软骨素和透明质酸。基质金属蛋白酶13（MMP-13）是OA关节软骨降解的主要胶原酶。开发基于MMP-13的药物的多次尝试失败主要是由于统称为肌肉骨骼综合征（MSS）的剂量限制性副作用。虽然MSS的确切原因尚不清楚，但据信是由于候选药物对金属蛋白酶家族的其他代表缺乏选择性。蛋白酶二级结合位点（外切位点），即促进或调节蛋白酶活性的非活性位点区域的鉴定可用于设计蛋白酶家族内的选择性抑制剂。我们的实验室已经开发了三螺旋肽（THP）荧光共振能量转移（FRET）底物的高通量筛选（HTS）的胶原蛋白溶解MMP和鉴定推定的外来结合化合物。使用HTS技术与FRET THP测定，我们以前确定了几个选择性低微摩尔抑制剂MMP-13。一种抑制剂（化合物Q/4，PubChem CID 2047223）表现出表明其作用模式不是通过Zn螯合的性质。化合物Q/4具有比先前描述的MMP-13抑制剂显著更小的分子支架，但提供优异的选择性。我们希望进一步开发化合物Q/4及其类似物作为体内MMP-13探针。实现该目标的具体目的如下：（1）MMP-13外位点探针的X射线晶体学引导的药物化学;和（2）MMP-13外位点探针的体外和体内测试。目前的工作将导致在体内的探针，可以评估的影响，选择性抑制剂对细胞外蛋白水解活性在OA和奠定了基础，开发新的MMP成像剂。此外，将鉴定MMP抑制的独特模式。

DESCRIPTION (provided by applicant): Osteoarthritis (OA), the most common form of arthritis, is characterized by the destruction of articular cartilage. The main constituents of articular or joint cartilage are type II collagen and various proteoglycans, such as aggrecan, chondroitin sulfate, and hyaluronan. Matrix metalloproteinase 13 (MMP-13) has been shown to be main collagenase responsible for degradation of articular cartilage during OA. Multiple attempts to develop an MMP-13 inhibitor-based drug failed mostly due to the dose limiting side effects collectively known as musculoskeletal syndrome (MSS). While the exact cause of MSS is not known it is believed to be due to the lack of selectivity of drug candidates towards other representatives of metalloproteinase families. Identification of protease secondary binding sites (exosites), i.e. non-active site regions that facilitate or modulate protease activity, could be utilized for the design of selective inhibitors within protease families. Our laboratory has developed triple-helical peptide (THP) fluorescence resonance energy transfer (FRET) substrates for high throughput screening (HTS) of collagenolytic MMPs and identification of putative exosite-binding compounds. Using HTS techniques with FRET THP assays, we previously identified several selective low micromolar inhibitors of MMP-13. One inhibitor (compound Q/4, PubChem CID 2047223) exhibited properties suggesting that its mode of action was not via Zn chelation. Compound Q/4 possessed a significantly smaller molecular scaffold than previously described MMP-13 inhibitors, yet offered excellent selectivity. We wish to further develop compound Q/4 and its analogs as in vivo MMP-13 probes. The specific aims to achieve this goal are as follows: (1) X-ray crystallography-guided medicinal chemistry of MMP-13 exosite probes; and (2) in vitro and in vivo testing of MMP-13 exosite probes. The present work will lead to in vivo probes that can evaluate the effects of selective inhibitors on extracellular proteolytic activity in OA and lay the groundwork for the development of novel MMP imaging agents. In addition, a unique mode of MMP inhibition will be identified.