Isolation of Monoclonal Antibodies against Membrane Proteins

描述(申请人提供)：由于它们在细胞信号和人类疾病中的重要性，G蛋白偶联受体(GPCRs)和离子通道等完整的膜蛋白构成了现有药物靶标的40%以上。识别膜蛋白上构象相关表位的单抗通常是最有价值的抗体类型，因为它们通常与受体的关键结构结合，可用于检测或抑制受体。然而，开发构象依赖的、对膜蛋白具有抑制作用的单抗尤其困难，因为与可溶性蛋白不同，大多数膜蛋白依赖于脂质环境来维持其天然的三级和四级(寡聚体)结构。膜蛋白通常难以纯化，通常在细胞表面低浓度表达，并且通常不能被线型多肽所代表。需要新的方法来开发用于治疗、诊断和研究应用的单抗。在这里，我们建议使用一种新的技术，脂肪制品，以一种可免疫的形式捕获和浓缩结构完整的膜蛋白。使用脂肪颗粒开发针对细胞膜蛋白的抗体的概念建立在将病毒颗粒用作成功疫苗的历史上。 公共卫生相关性：这项提议将产生针对重要膜蛋白靶标的单抗，用于治疗开发、诊断和生物医学研究。优化为免疫原的脂肪颗粒将被开发为商业化产品。

DESCRIPTION (provided by applicant): Because of their importance in cell signaling and human disease, integral membrane proteins such as G protein-coupled receptors (GPCRs) and ion channels comprise over 40% of existing drug targets. Monoclonal antibodies (MAbs) that recognize conformation-dependent epitopes on membrane proteins are usually the most valuable type of antibody because they often bind to critical structures of the receptor that can be exploited for its detection or inhibition. However, the development of conformation-dependent, inhibitory MAbs against membrane proteins is especially difficult because, unlike soluble proteins, most membrane proteins are dependent on a lipid environment to maintain their native tertiary and quaternary (oligomeric) structures. Membrane proteins are often difficult to purify, often express at low concentrations on the cell surface, and are usually poorly represented by linear peptides. New approaches are needed to develop such MAbs for therapeutic, diagnostic, and research applications. Here we propose to use a novel technology, the Lipoparticle, to capture and concentrate structurally intact membrane proteins in a format amenable to immunization. The concept of using Lipoparticles to develop antibodies against cellular membrane proteins builds on the historic use of viral particles as successful vaccines. PUBLIC HEALTH RELEVANCE: This proposal will result in monoclonal antibodies against important membrane protein targets for therapeutic development, diagnostics, and biomedical research. Lipoparticles optimized as immunogens will be developed as commercial products.