Design and synthesis of novel fluorescent ligands for the Beta-3 adrenoceptor

β-3肾上腺素能受体(B3AR)是治疗肥胖、膀胱过度活动症(OAB)和2型糖尿病的有效靶点，属于G蛋白偶联受体(GPCR)超家族。GPCRs是一种复杂而灵活的膜结合蛋白，由约800个家族成员组成。B3AR被内源性儿茶酚胺激活，如肾上腺素/去甲肾上腺素，并介导关键的动态平衡过程，如脂解和产热。尽管B3AR是在40年前被发现的，但关于它的药理还有很多需要了解的地方，到目前为止，只有少数几种治疗OAB的疗法获得了临床批准。特别是，最理想的B3AR药物是激动剂(激活受体)，这些配体的实时结合行为是它们刺激目标细胞类型(如脂肪细胞和平滑肌)以提供减肥或膀胱松弛效果的关键组件。最近，与OAB药物mirabegron结合的犬B3AR的结构已被发表(Nagiri等，2021，PDB：7DH5)。虽然人类和狗的蛋白质序列总体上有83%的相似性，但它们的差异可能会在它们的三维形状和配体相互作用方面产生显著的差异。因此，有必要开发一个可靠的人类B3AR模型来为设计新的B3AR选择性配体提供信息。建模工作将允许设计和合成一个将进行药理学表征的荧光配体文库。此外，有效的荧光配体将允许建立强大的基于荧光的B3AR分析，将更详细的配体结合动力学分析映射到所需的功能反应。该项目的具体目标将是：1.生成优化的人类B3AR计算模型，并使用这些模型来设计、合成和表征新型荧光配体的文库。对所有β-肾上腺素能受体亚型的新荧光配体进行药理学表征。利用人类受体的计算模型对Leads进行虚拟筛选，为B3ARs开发新型的选择性配体支架。将新的多学科知识(结构、化学、药理学)应用于将选定的铅开发成有效的和亚型选择性的B3AR配体。这个专注于化学生物学的项目将跨越合成化学、建模和药理学的学科，以增加我们对B3AR生物学的理解。肾上腺素能受体家族作为典型的GPCRs和关键的治疗靶点的重要性意味着该项目将与未来的药物发现工作直接相关。

The Beta-3 adrenoceptor (B3AR) is an attractive therapeutic target for obesity, overactive bladder (OAB) and type-2 diabetes and belongs to the G protein-coupled receptor (GPCR) superfamily. GPCRs are complex and flexible membrane-bound proteins comprising ~800 family members. The B3AR is activated by endogenous catecholamines, such as epinephrine/norephinephrine and mediates critical homeostatic processes such as lipolysis and thermogenesis. Though the B3AR was identified >40 years ago, there is much to be learnt about its pharmacology and to-date only a handful of therapeutics for OAB have been clinically approved. In particular, most desired B3AR drugs are agonists (activating the receptors), and the real time binding behaviour of these ligands is a key component of how well they stimulate target cell types such as adipocytes and smooth muscle to provide an anti-obesity or bladder relaxation effect.Recently, the structure of the canine B3AR bound to the OAB drug mirabegron has been published (Nagiri et al, 2021, PDB: 7DH5). Whilst the human and canine protein sequences share ~83% overall similarity, their differences may confer significant variation in their three-dimensional shape and ligand interaction. Therefore, there is a need to develop a reliable human B3AR model to inform the design of new B3AR-selective ligands.The modelling work will allow the design and synthesis of a library of fluorescent ligands which will undergo pharmacological characterisation. Furthermore, effective fluorescent ligands will allow powerful fluorescence-based B3AR assays to be established, mapping more detailed analysis of ligand-binding kinetics to the desired functional responses.Specific aims of the project will be:1. To generate optimised computational models of the human B3AR and use these to design, synthesise and characterise a library of novel fluorescent ligands.2. To pharmacologically characterise the new fluorescent ligands at all beta-adrenoceptor subtypes3. To use the computational model of the human receptor to conduct a virtual screen for leads to develop novel, selective ligand scaffolds for the B3AR.4. To apply the new multidisciplinary knowledge (structures, chemistry, pharmacology) to the development of selected leads into potent and subtype-selective ligands for the B3AR.This chemical biology-focused project will span the disciplines of synthetic chemistry, modelling and pharmacology to increase our understanding of B3AR biology. The importance of the adrenoceptor family as prototypical GPCRs and key therapeutic targets means this project will be of direct relevance to future drug discovery efforts.