Next Generation Helix Mimetics - Probes for Target Selectivity

蛋白质是人体的机器，它们相互作用的方式对它们的正常运作至关重要。这些蛋白质-蛋白质相互作用(PPI)不仅控制细胞中的关键过程，还可能参与疾病的发展。例如，在许多癌症中，通常用来调节细胞生存或死亡的PPI被用来促进肿瘤的生长。因此，已经作出了相当大的努力，试图防止这些有害的相互作用发生。然而，针对PPI并不是一目了然的。蛋白质通常使用非常大的，通常是平坦的表面相互作用。这使得小得多的药物分子要破坏/抑制这些相互作用是具有挑战性的。克服这一挑战的一种策略是以一种方式构建分子，使其模仿参与PPI的一种蛋白质相互作用表面的关键特征。通过这种方式，分子可以复制在天然PPI中发生的相同的相互作用，而不是另一个相互作用的伙伴，后者现在可以自由地执行其正常的生物功能。这种拟态策略已经成功地用于抑制癌细胞中有害的PPI。然而，目前尚不清楚的是，除了它们预定的目标外，这些分子还能与多少蛋白质相互作用，这种分子被称为拟态。如果这些分子要在药物开发过程中取得进一步的进展，了解这种选择性分布是至关重要的。在这个项目中，我们将表征一系列针对相同PPI的领先模拟物的选择性分布。这不仅将提供关于任何有问题的目标外目标的关键信息，而且还将突出可能提高其效力的任何有益的额外目标。这项研究还将首次全面比较领先的模拟学，以使最有希望的候选者的开发成为优先事项。我们将通过向模拟结构添加两个额外的功能来实现这一点。首先，我们将添加一个化学基团，当它被紫外光激活时，将在模拟物和它与之相互作用的任何蛋白质之间形成永久的联系。其次，我们将添加一个分子钓鱼钩，这将使我们能够从细胞内复杂的蛋白质混合物中分离出与模拟物相连的蛋白质。然后，我们将识别与每个模拟物结合的所有蛋白质的质量，并使用替代的、以单一蛋白质为重点的方法来验证结果。这个项目不仅可以显著提高我们对主要模拟分子在复杂生物环境中的活性的了解，而且还将提供访问未来模拟物靶标特征的通用方法，从而可以准确地预测它们的完整生物学活性，从而为确定蛋白质模拟物作为治疗的潜力向前迈进了重要的一步。

Proteins are the machines of the body and the way they interact with one another is critical for their correct functioning. These protein-protein interactions (PPIs) not only control key processes in cells but can also be involved in the development of disease. For example, in many cancers PPIs which are normally used to regulate whether a cell lives or dies are exploited to enable tumour growth. Therefore, considerable effort has been placed on trying to prevent these harmful interactions from occurring. However, targeting PPIs is not straightforward. Proteins commonly interact with one another using very large, and often mostly flat, surfaces. This makes it challenging for a much smaller drug molecule to disrupt/inhibit these interactions. One strategy to overcome this challenge is to construct molecules in such a way that they mimic a key feature of the interacting surface of one of the proteins involved in the PPI. This way the molecule can replicate the same interactions which occur in the native PPI in place of the other interacting partner which is now free to carry out its normal biological function. This strategy of mimicry has been used successfully to inhibit harmful PPIs in cancer cells. However, what is not currently clear is how many proteins these molecules, known as mimetics, can interact with aside from their intended target. It is critical to understand this selectivity profile if these molecules are to progress further in the drug development process.In this project, we will characterise the selectivity profile of a series of leading mimetics targeted against the same PPI. This will not only provide critical information on any problematic off-targets but also will highlight any beneficial additional targets that may enhance their effectiveness. This study will also provide the first comprehensive comparison of leading mimetics to enable development of the most promising candidates to be prioritised. We will achieve this by adding two additional features to structures of the mimetics. Firstly, we will add a chemical group which, when activated by UV light, will form a permanent linkage between the mimetic and any protein it is interacting with. Secondly, we will add a molecular fishing hook which will enable us to isolate only the proteins that are linked to the mimetic from the complex mixture of proteins inside a cell. We will then identify all the proteins bound to each mimetic by their mass and validate the results using alternative, single protein focused methods.This project will not only allow significantly enhance our understanding of the activity of leading mimetic molecules in complex biological environments but it will also provide access to general methods for the target profile characterisation of future mimetics allowing their complete biological activity to be accurately predicted providing a vital step forward in establishing the potential of protein mimetics as therapeutics.