Monoclonal Thyroid Stimulating Antibodies

描述（由申请人提供）：促甲状腺激素（TSH）受体（TSHR）是Graves病的主要抗原，也是此类患者血清中促甲状腺激素抗体（TSHR- ab）的靶标。TSHR-Ab的特点是其作用时间长，最初被称为长效甲状腺刺激剂（LATS）。在这个应用中，我们寻求继续成功克隆和鉴定TSH受体单克隆抗体，这导致了第一个具有刺激活性的单克隆抗体。特别是，单克隆抗体MS-1在稳定表达hTSHR的CHO细胞中显示出强大的TSHR刺激活性，活性低至2ng/ml的IgG。这一竞争性更新的具体目的是：目的1：鉴定额外的单克隆促甲状腺抗体。我们将首先在成功的基础上继续开发具有促甲状腺活性的TSH受体单克隆抗体。目的2：确定tshrr - ab表位。我们将对衍生的TSH受体抗体的线性和非线性抗原表位进行表征，以确定TSH受体上的主要免疫原区（mir）。目的3：定义抗体介导的TSHR翻译后修饰调控。我们将在CHO细胞中共同表达TSHRcpp - TSHRmyc构建体，并检查tshrp - abs对受体分子的占领是否会像我们在TSH配体中看到的那样调节FRET指数。此外，我们最近开发了一种独特的测定TSHR切割的方法，并将研究TSHR- abs对受体切割的影响，目的是深入了解这种切割在甲状腺自身免疫中的功能作用。目的4：研究tshr -抗体的生物学功能和信号转导。我们将研究TSHR-Abs在刺激/抑制生长潜能和激活/抑制分化甲状腺细胞功能方面的特异性。分离和鉴定与Graves病患者血清中发现的TSH抗体特征相似的TSH抗体，将为其作用机制提供重要的新见解，并为控制甲状腺功能障碍提供新方法。

DESCRIPTION (provided by applicant): The thyrotropin (TSH) receptor (TSHR) is the primary antigen of Graves' disease and the target for thyroid stimulating 'TSHR antibodies (TSHR-Ab) found in the serum of such patients. TSHR-Ab are characterized by their prolonged action and were originally called Long Acting Thyroid Stimulators (LATS). In this application, we seek to continue our successful cloning and characterization of monoclonal antibodies to the TSH receptor which have resulted in the first monoclonal antibody with stimulating activity. In particular, monoclonal antibody MS-1 has shown potent TSHR stimulating activity when tested with CHO cells stably expressing hTSHR with activity evident down to 2ng/ml of IgG. The specific aims of this competitive renewal are: Aim 1: Identify additional monoclonal thyroid-stimulating antibodies. We will first continue to build on our success by developing an expanded panel of monoclonal antibodies to the TSH receptor with thyroid stimulating activity. Aim 2: Define TSHR-AB epitopes. We will characterize the linear and non-linear antigenic epitopes for the derived TSH receptor antibodies to define the major immunogenic 'regions (MIRs) on the TSH receptor. Aim 3: Defining antibody-mediated regulation of TSHR post-translational modifications. We will co-express TSHRcpp TSHRmyc constructs in CHO cells and examine whether occupation of the receptor molecule by TSHR-Abs will modulate the FRET index as we have seen for TSH ligand. In addition, we have recently developed a unique assay for measuring TSHR cleavage and will examine the influence of TSHR-Abs on receptor cleavage with the aim of gaining insight into the functional role of such cleavage in thyroid autoimmunity. Aim 4: Examine the biologic function and signal transduction induced by TSHR-Abs. We will examine the specificity of TSHR-Abs with regard to their growth stimulating/inhibiting potential and their activarion/inacrivation of differentiated thyroid cell function. The isolation and characterization of antibodies to the TSH with similar characteristics to the TSHR-Abs found in the serum of Graves' disease patients will provide major new insight into their mechanisms of action and offer novel approaches to the control of thyroid dysfunction.