Activation mechanisms of Class F GPCRs – from structural understanding to novel therapeutics

生物体的细胞必须能够感知和适应周围环境的变化。这些过程是由膜嵌入的生物分子介导的，G蛋白偶联受体（gpcr）家族在真核细胞中起着重要作用。目前批准的所有药物中约有30%针对gpcr，这类细胞表面受体将继续成为未来药物发现计划的重点。除了开发新型GPCR定向药物外，现代药物发现的一个主要策略是利用尚未开发的GPCR家族作为新的药物靶点。其中之一是GPCR家族F类，包括10个卷曲受体（FZD1-10）和平滑受体（SMO）。FZDs参与多种病理生理过程，因此是治疗不同肿瘤、炎症过程和神经退行性疾病的有吸引力的靶点。不幸的是，由于几个原因，我们目前对这个GPCR家族的了解相当有限。FZDs作为分泌的无翼/int1 （WNT）脂蛋白的细胞表面受体，需要经过艰苦的生物技术程序纯化。因此，wnt不能作为理想的药理学工具来探索F类gpcr的结构和功能。此外，FZD激活和随后的受体介导的信号传导的许多方面仍然未知，例如，不同的FZD介导的信号级联与不同疾病发病机制的相关性尚存疑问。此外，尽管在过去几年中取得了重大进展，但在WNT/FZD群体中，关于g蛋白介导的FZD信号的（病理）生理相关性仍存在强烈分歧。缺乏合适的FZD配体和先进的生化以及细胞生物学方法进一步阻碍了对这些基本FZD功能的研究，并最终减缓了FZD导向治疗方法的发展。该项目的主要目标是开发先进的技术来研究F类gpcr，并利用这些方法对创新的FZD配体进行药理学表征。在本项目中确定的高效FZD配体，将评估其稳定特定受体构象的FZD的能力，并通过冷冻电子显微镜对卷曲的gpcr进行后续结构研究。申请人具有多年GPCR检测开发经验。此外，与FZD药理学领域的先驱研究员，以及与卡罗林斯卡研究所优秀的研究基础设施的合作，将实现有希望的协同效应，并使FZD研究取得重大突破。这项研究的见解不仅将影响这些受体的后续结构研究，而且还将指导未来以fzd为导向的药物发现计划。

The cells of living organisms have to be able to perceive and adopt to changes of their surrounding environment. These processes are mediated by membrane-embedded biomolecules and the family of G protein-coupled receptors (GPCRs) plays a major role within eukaryotic cells. Approximately 30% of all currently approved drugs address GPCRs and this class of cell surface receptors will continue to be in the focus of future drug discovery programs. Besides developing novel types of GPCR-directed drugs, one major strategy in modern drug discovery is to exploit yet untapped GPCR families as novel drug targets. One of these is the GPCR family class F, encompassing the ten Frizzled receptors (FZD1-10) and Smoothened (SMO). FZDs are involved in diverse pathophysiological processes and therefore represent attractive targets for the treatment of different tumours, inflammatory processes and neurodegenerative diseases. Unfortunately, our current understanding of this GPCR family is quite limited for several reasons. FZDs function as cell surface receptors for secreted Wingless/int1 (WNT) lipoglycoproteins which need to be purified under laborious biotechnological procedures. Thus, WNTs cannot serve as ideal pharmacological tools to explore structure and function of class F GPCRs. Furthermore, many aspects of FZD activation and subsequent receptor-mediated signalling remain unknown, leaving for instance the relevance of distinct FZD-mediated signalling cascades for the pathogenesis of different diseases in doubt. Moreover, despite significant progress over the last years, there is yet a strong disagreement in the WNT/FZD community on the (patho-)physiological relevance of G-protein-mediated FZD signalling. The lack of appropriate FZD ligands and advanced biochemical, as well as cell biological methods further hampers the investigation of such essential FZD functions and ultimately slows down the development of FZD-directed therapeutics. It’s the major goal of this project to develop advanced technologies to study class F GPCRs and employ these methods to pharmacologically characterize innovative FZD ligands. High potent FZD ligands, identified as such in the course of this project, will be evaluated for their ability to stabilise FZDs in specific receptor conformations and to make them accessible for subsequent structural studies of frizzled GPCRs through cryo-electronmicroscopy. The applicant has multi-year experience in GPCR assay development. Furthermore, the collaboration with the host-scientist, who is a pioneering researcher in the field of FZD pharmacology, as well as the integration into the excellent research infrastructure at the Karolinska Institute will enable promising synergies and allow for significant breakthroughs in FZD research. The insights of this study will not only affect subsequent structural studies of these receptors but also guide future FZD-directed drug discovery programs.