靶向非经典配体结合口袋的选择性PPARγ调节剂的药理机制研究

Insulin resistance is an important risk factor for Type 2 diabetes. The agonists of peroxisome proliferator-activated receptor-γ (PPARγ) are expected to reverse insulin resistance in patients with type 2 diabetes mellitus. The full PPARγ agonists such as TZDs have an unattractive side effect profile including weight gain, edema, neutropenia and hemodilution. Recent studies have indicated that selective PPARγ modulators (partial PPARγ agonist) exhibited improved safety margins compared to full PPARγ agonists and consequently much effort has been put in promoting these modulators for clinical development. Crystallographic studies showed that most of the selective PPARγ modulators bound to the classic ligand binding domain of the PPARγ-LBD. In our previous work, a novel non-thiazolidinedione selective PPARγ modulator MDCCCL1636 was investigated. The compound displayed partial agonist activity and caused reversal of insulin resistance. MDCCCL1636 can also reduce FSG and reverse the dyslipidemia and damage of the pancreas without significantly increasing body weight. Furthermore, MDCCCL1636 had lower toxicity compared to pioglitazone in vivo and in vitro. Interestingly, we found that MDCCCL1636 bound to the non-classical ligand binding sites of PPARγ-LBD in the crystal structure of their complex. In this project, we will investigate the molecular mechanisms of this selective modulator with non-classical ligand binding mode. The ligand binding mode of MDCCCL1636 will be further validated, and the downstream signaling pathways and related target gene affected by MDCCCL1636 will be elucidated. Based on the molecular mechanism research, a series of selective PPARγ modulators possessing this kind of non-classical ligand binding domain will be designed using the computer-aided drug design method. The druggability and structure-activity relationship of this kind of modulators will be evaluated. Based on the above research, the feasibility of this non-classical ligand binding “pocket” as a potential drug “target” will be elucidated. This work will provide new ideas for selective PPARγ modulators development.

胰岛素抵抗是II型糖尿病的重要致病因素，选择性PPARγ调节剂可以在改善胰岛素抵抗的同时，避免现有TZDs类药物的副作用，是目前的研究热点。晶体学研究表明，绝大多数PPARγ选择性调节剂均结合在其经典的Y型配体结合口袋中。本项目前期研究发现了选择性PPARγ调节剂MDCCCL1636，可以显著改善糖尿病大鼠的胰岛素抵抗、血糖及血脂参数且副作用小。晶体学数据表明MDCCCL1636结合在PPARγ-LBD的非经典配体结合位点，其背后的分子机制值得深入研究。本项目拟进一步研究靶向PPARγ非经典配体结合口袋的小分子MDCCCL1636发挥选择性调节作用的药理机制，并对以此为基础设计的PPARγ调节剂的成药性和构效关系进行研究，评价该非经典配体结合“口袋”作为潜在药物开发“靶点”的可行性，为新型选择性PPARγ调节剂的开发提供理论基础。

本课题的研究目标是阐明小分子药物MDCCCL1636（HMPA）调节糖脂代谢发挥T2DM治疗作用的分子机制，为该药物的进一步开发打下理论基础。并且基于HMPA的分子结构，设计合成新的小分子药物，以期寻找到具有更好的药理活性的小分子。本课题前期通过体外研究工作发现HMPA对PPARγ有选择性调节作用，HMPA结合在PPARγ非经典配体结合口袋位置。我们在初期也对这一新型的结合方式进行了一些探索工作。然而我们在后期进行药物给药方式的研究及药代动力学研究时发现：HMPA的最佳给药方式为灌胃给药，注射给药及腹腔给药的药效均较弱。药代动力学研究结果也表明：HMPA的血药浓度较低，清除速率快。这些提示我们：HMPA在体内发挥功能可能有跟我们的体外研究不一样的分子机制。由于其灌胃给药对T2DM大鼠的降糖效果优于腹腔注射和尾静脉给药，我们推测肠道菌群可能介导了HMPA的降糖作用。因此我们对HMPA的药理作用特点进行了深入的研究。伪无菌小鼠及菌群移植实验结果表明，肠道菌群在HMPA调节T2DM的过程中发挥着重要的介导作用，HMPA可以调节肠道菌群的结构，调节不同群落之间的平衡，改善肠道的屏障功能，促进短链脂肪酸（SCFAs）的生成。分子机制研究结果显示，HMPA可以通过靶向GlnR蛋白，改善肠道菌群碳源利用过程中的碳源代谢阻遏（CCR）效应，促进菌群对多种碳源的利用，从而增加肠道菌群发酵产物SCFAs的含量，发挥T2DM调节作用。这一新颖的分子机制的发现，为T2DM治疗药物提供了新的研发思路，可以通过调控肠道菌群的功能，提高发酵产物短链脂肪酸的含量，来发挥改善T2DM的功能。并且我们也对HMPA进行了成药性评价，发现其成药性良好。根据其分子结构，也进行了一些化学修饰，也发现了几个同样具有较好药理活性的小分子，为后期的进一步药物开发打下了基础。