Molecular Imaging of G-Protein-Coupled Receptors for Drug Development

描述(申请人提供)：G蛋白偶联受体(GPCRs)是人类最重要的蛋白质之一，因为它们是许多信号通路的“门户”，包括那些导致疾病的通路，它们的功能可以通过药物干预。通过识别和结合特定的化学物质，GPCRs将信号从细胞外传递到细胞内，在那里它们触发了一系列生物事件。因此，GPCRs是用于结构确定的最多产的一类蛋白质。它们是最大的一类膜蛋白；大约800个GPCR编码在人类基因组中，其中大约一半是潜在的药物靶点。然而，目前只有约60种药物被用作小分子药物的受体，其中约25种被用于基于天然配体的生物治疗；然而，与GPCRs结合的药物约占所有治疗药物的三分之一。到2014年，全球与GPCRs相互作用的药物的年度市场预计将达到1180亿美元。另外80多个GPCR潜在地服从于抗体疗法，这是一种正在受到越来越多关注的新的攻击途径，我们将探索这一途径。从大分子膜蛋白制备X射线结晶学和溶液核磁共振样品的困难一直是基于结构的药物发现的主要障碍。为了克服这一障碍，我们同时开发了固体核磁共振波谱的新方法，并将其应用于膜蛋白的结构测定。我们的主要目标是趋化因子受体CXCR1，在该奖项的前五年，我们在确定其结构方面取得了实质性进展。我们对IL-8与CXCR1相互作用的研究为发现可能通过与CXCR1结合而影响炎症、癌症转移和其他疾病的药物提供了一个框架。为了加速设计和发现与CXCR1和其他GPCRs相互作用的药物，我们正在开发一种通用的方法来确定GPCRs在生理条件下天然磷脂环境中的三维结构。这项研究是多学科的，涉及分子生物学、生物化学、结构生物学、核磁共振波谱和计算。它在培训能够跨越化学和生物边界工作和互动的科学家方面非常有效。我们认识到，它需要现有的最高水平的技术；因此，它是加州大学圣地亚哥分校(UCSD)和两家生物技术公司(位于马萨诸塞州安多弗的剑桥同位素实验室(CIL)和位于加利福尼亚州圣地亚哥的膜受体技术公司(MRT))之间的生物工程研究伙伴关系。CIL的关键技术是同位素标记氨基酸和前体的合成，以及独特的同位素标记细菌生长介质的制造；他们的工作由Joel Bradley博士领导。MRT拥有表达、纯化和复性生物活性GPCRs的技术，最初由M-Fold Biotech的Hans Kiefer博士开发，他仍然参与这项研究。

DESCRIPTION (provided by applicant): G-protein coupled receptors (GPCRs) are among the most important proteins in humans because they are the 'gateway' for many signally pathways, including those that contribute to diseases, and their functions are amenable to intervention by drugs. By recognizing and binding specific chemicals, GPCRs transduce signals from the outside of cell to the inside where they trigger a cascade of biological events. Consequently, GPCRs are the most fecund class of proteins for structure determination. They are the largest class of membrane proteins; ~800 GPCRs are encoded in the human genome, about half of which are potential drug targets. However, only ~60 of them are currently used as receptors for small molecule drugs and ~25 of them for bio- therapeutics based on the natural ligand; nonetheless, drugs that bind to GPCRs account for about one-third of all therapeutic drugs. The annual worldwide market for drugs that interact with GPCRs is predicted to be $118 Billion by 2014. More than 80 additional GPCRs are potentially amenable to antibody therapeutics, a novel avenue of attack that is receiving increasing attention, and one that we will explore. The difficulty in preparing samples for X-ray crystallography and Solution NMR from large membrane proteins in liquid crystalline phospholipid bilayers has been the principal roadblock to structure-based drug discovery. In order to overcome this roadblock we have simultaneously developed new methods for solid-state NMR spectroscopy and applied to structure determination of membrane proteins. Our principal target is the chemokine receptor CXCR1, and we have made substantial progress towards determining its structure during the first five years of this award. Our studies of interleukin-8 (IL-8) interacting with CXCR1 provide a framework for discovery of drugs that may affect inflammation, cancer metastasis, and other diseases by binding to CXCR1. In order to accelerate the design and discovery of drugs that interact with CXCR1 and other GPCRs, we are developing a general method for determining the three-dimensional structures of GPCRs in their native phospholipid environment under physiological conditions. This research is multidisciplinary, involving molecular biology, biochemistry, structural biology, NMR spectroscopy, and computation. It is highly effective at training scientists who can work and interact across chemical and biological boundaries. We recognize that it requires the highest levels of technology available; as a result, it is organized as a Bioengineering Research Partnership among the University of California, San Diego (UCSD) and two biotechnology companies (Cambridge Isotope Laboratories (CIL), Andover, Massachusetts and Membrane Receptor Technologies (MRT), San Diego, California. CIL's key technology is the synthesis of isotopically labeled amino acids and precursors, and the manufacture of unique isotopically labeled bacterial growth media; their effort is led by Joel Bradley, Ph.D. MRT has the technology for the expression, purification, and refolding of biologically active GPCRs developed originally by Hans Kiefer, Ph.D., at M-Fold Biotech and he remains involved in the research.