A human monoclonal autoantibody to the thyrotropin receptor with thyroid-stimulating blocking activity
A human monoclonal autoantibody to the thyrotropin receptor with thyroid-stimulating blocking activity
复制标题
DOI：
10.1089/thy.2007.0327
复制
发表时间：
2008-07-01
期刊：
影响因子：
6.6
通讯作者：
Smith, Bernard Rees
中科院分区：
文献类型：
--
作者：
Sanders, Jane;Evans, Michele;Smith, Bernard Rees
背景：人单克隆自身抗体（MAb）是研究自身免疫反应的重要工具。迄今为止，只有一个人单克隆抗体的促甲状腺激素（TSH）受体（TSHR）具有刺激活性。我们现在描述的详细特征的阻断型人单克隆抗体的TSHR.Methods：一个单一的异源杂交瘤细胞系分离的患者的严重甲状腺功能减退症（TSH 278 mU/L），使用标准技术的外周血淋巴细胞。该细胞系稳定表达TSHR自身抗体（5C 9; IgG 1/κ）。使用涂有溶解的TSHR的管测试5C 9结合I-125-TSH或TSHR抗体并与I-125-TSH或TSHR抗体竞争结合TSHR的能力。此外，使用表达野生型人类TSHR或具有氨基酸突变的TSHR的中国仓鼠卵巢（CHO）细胞评估5C 9对刺激环AMP产生的阻断作用。主要结果：5C 9 IgG以高亲和力（4 × 10（10）L/mol）结合TSHR，并抑制TSH和甲状腺刺激性人类单克隆自身抗体（M22）与受体的结合。5C 9 IgG制剂抑制TSH、M22、血清TSHR自身抗体和促甲状腺激素小鼠单克隆抗体的环AMP刺激活性。此外，5C 9降低了野生型TSHR和具有一些激活突变的TSHR的组成型活性。TSHR中不同氨基酸突变对5C 9生物活性的影响进行了研究，TSHR Lys 129 Ala或Asp 203 Ala完全消除了5C 9阻断TSH介导的刺激环AMP production.Conclusions的能力：5C 9的可用性提供了新的机会，研究TSHR阻断型自身抗体的结合和生物活性，包括在分子水平上的研究。此外，单克隆抗体，如5C 9，很可能提供新的药物控制TSHR活性，包括在甲状腺癌和Graves眼病的应用的基础。
Background: Human monoclonal autoantibodies (MAbs) are valuable tools to study autoimmune responses. To date only one human MAb to the thyrotropin (TSH) receptor (TSHR) with stimulating activity has been available. We now describe the detailed characterization of a blocking type human MAb to the TSHR.Methods: A single heterohybridoma cell line was isolated from the peripheral blood lymphocytes of a patient with severe hypothyroidism (TSH 278 mU/L) using standard techniques. The line stably expresses a TSHR autoantibody (5C9; IgG1/kappa). Ability of 5C9 to bind and compete with I-125-TSH or TSHR antibodies binding to the TSHR was tested using tubes coated with solubilized TSHR. Furthermore, the blocking effects of 5C9 on stimulation of cyclic AMP production was assessed using Chinese hamster ovary (CHO) cells expressing the wild-type human TSHR or TSHRs with amino acid mutations.Main outcome: 5C9 IgG bound to the TSHR with high affinity (4 x 10(10) L/mol) and inhibited binding of TSH and a thyroid-stimulating human monoclonal autoantibody (M22) to the receptor. 5C9 IgG preparations inhibited the cyclic AMP-stimulating activities of TSH, M22, serum TSHR autoantibodies and thyroid-stimulating mouse monoclonal antibodies. Furthermore 5C9 reduced the constitutive activity of wild-type TSHR and TSHR with some activating mutations. The effect of different amino acid mutations in the TSHR on 5C9 biological activity was studied and TSHR Lys129Ala or Asp203Ala completely abolished the ability of 5C9 to block TSH-mediated stimulation of cyclic AMP production.Conclusions: The availability of 5C9 provides new opportunities to investigate the binding and biological activity of TSHR blocking type autoantibodies including studies at the molecular level. Furthermore, monoclonal antibodies such as 5C9 may well provide the basis of new drugs to control TSHR activity including applications in thyroid cancer and Graves' ophthalmopathy.