PROTACs as a novel Approach to target Protein Tyrosine Kinases for degradation in human Platelets

身体已经发展出一种精细调节的机制来阻止血管损伤部位的失血，同时防止血管因过度凝块形成而堵塞。它取决于一系列涉及血小板和血浆凝血因子的复杂的相互关联的事件。血小板是一种小的细胞碎片，通常处于非活性状态。然而，当血管受伤时，它们会迅速粘在受伤部位并相互粘在一起止血。血小板反应如此有效的原因是，它们表面有一种叫做“受体”的蛋白质，可以识别受损血管中的分子。当这些分子与血小板受体结合时，细胞内的通讯信号（=信号转导）就会受到刺激，这些信号会告诉血小板聚集在一起并止血。然而，有时刺激信号会导致过多的血栓形成，从而导致心脏病发作或中风。许多研究已经完成，以调查哪些分子/蛋白质参与这些信号转导途径。然而，实验研究这些信号转导途径并不简单，因为血小板不像其他细胞那样具有DNA，因此不能进行基因修饰。因此，大部分研究都是在转基因小鼠模型和/或使用药理学药物上进行的。这些都有物种差异的相关并发症（老鼠和人不一样），而且药物通常不能区分不同的分子/蛋白质。因此，尽管动物模型显示一组称为酪氨酸激酶的蛋白质有助于血小板功能，但尚不清楚它们如何促进人类血小板功能。因此，在这项拨款申请中，我们建议开发和优化一种名为PROTAC（靶向嵌合体蛋白水解）的新型实验技术，用于人类血小板。PROTACs是一种小分子，可以添加到细胞中，劫持它们的内部系统，专门分解特定的蛋白质。因此，这将有助于我们研究这种特殊蛋白质在人类血小板功能和止血中的作用。我们将集中精力研究酪氨酸激酶，并将利用现有的PROTACs和进一步设计、开发和优化新的PROTACs来研究酪氨酸激酶在人类血小板中的作用。

The body has developed a finely tuned mechanism to stop blood loss at sites of vascular injury while preventing vessel blockage by excessive clot formation. It depends on a complicated series of interrelated events involving platelets and plasma coagulation factors. Platelets are small cell fragments that are normally circulating in an inactive state. However, when a blood vessel becomes injured, they rapidly stick to the site of injury and to each other to stop the bleeding. The reason that platelets can respond so efficiently, is that they have proteins on their surface called 'receptors' that can recognize molecules in the damaged blood vessel. When these molecules bind to the platelet receptors, communication signals are stimulated in the cell (=signal transduction) that tell the platelets to clump together and stop the bleeding. However, sometimes signals are stimulated that lead to too much clot formation which can lead to a heart attack or a stroke.A lot of research has been done to investigate which molecules/proteins are involved in these signal transduction pathways. However, to study these signal transduction pathways experimentally is not straight forward as platelets do not have DNA like other cells, and therefore cannot be genetically modified. Much of the research has therefore been performed on genetically modified mouse models and/or the use of pharmacological drugs. These have the associated complication of species differences (mice are just not the same as people) and the drugs can often not discriminate between different molecules/proteins. Therefore, despite animal models showing that a set of proteins called tyrosine kinases contribute to platelet function, it is not yet clear how they contribute to platelet function in humans. In this grant application, we therefore propose to develop and optimise a novel experimental technique called PROTAC (PROteolysis TArgeting Chimera) for use in human platelets. PROTACs are small molecules that can be added to cells and hijack their internal system to specifically break down a particular protein. This thus will help us to study the role of that particular protein in human platelet function and the stopping of a bleed. We will focus our efforts on proteins called tyrosine kinases and will use existing PROTACs and further design, develop and optimise novel PROTACs to study the role of tyrosine kinases in human platelets.