title: A human monoclonal autoantibody to the thyrotropin receptor with thyroid-stimulating blocking activity.
abstract: 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 125I-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.
  
  
  RESULTS
  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.
authors.affiliation: FIRS Laboratories, RSR Ltd, Llanishen, Cardiff, United Kingdom.
authors.forename: Jane
authors.initials: J
authors.lastname: Sanders
authors.forename: Michele
authors.initials: M
authors.lastname: Evans
authors.forename: Corrado
authors.initials: C
authors.lastname: Betterle
authors.forename: Paul
authors.initials: P
authors.lastname: Sanders
authors.forename: Anshu
authors.initials: A
authors.lastname: Bhardwaja
authors.forename: Stuart
authors.initials: S
authors.lastname: Young
authors.forename: Emma
authors.initials: E
authors.lastname: Roberts
authors.forename: Jane
authors.initials: J
authors.lastname: Wilmot
authors.forename: Tonya
authors.initials: T
authors.lastname: Richards
authors.forename: Angela
authors.initials: A
authors.lastname: Kiddie
authors.forename: Kasemsri
authors.initials: K
authors.lastname: Small
authors.forename: Hayley
authors.initials: H
authors.lastname: Platt
authors.forename: Sara
authors.initials: S
authors.lastname: Summerhayes
authors.forename: Rebecca
authors.initials: R
authors.lastname: Harris
authors.forename: Magnus
authors.initials: M
authors.lastname: Reeve
authors.forename: Graziella
authors.initials: G
authors.lastname: Coco
authors.forename: Renato
authors.initials: R
authors.lastname: Zanchetta
authors.forename: Shu
authors.initials: S
authors.lastname: Chen
authors.forename: Jadwiga
authors.initials: J
authors.lastname: Furmaniak
authors.forename: Bernard Rees
authors.initials: BR
authors.lastname: Smith
mesh_terms: D000328:Adult; D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology* / Q000494:pharmacology* / Q000627:therapeutic use; D001323:Autoantibodies / Q000097:blood; D016466:CHO Cells; D002460:Cell Line; D006224:Cricetinae; D003412:Cricetulus; D000242:Cyclic AMP / Q000378:metabolism; D005260:Female; D049970:Graves Ophthalmopathy / Q000188:drug therapy; D006801:Humans; D007037:Hypothyroidism / Q000378:metabolism; D009154:Mutation / Q000235:genetics; D010053:Ovary / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D011989:Receptors, Thyrotropin / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D013961:Thyroid Gland / Q000187:drug effects* / Q000378:metabolism; D013964:Thyroid Neoplasms / Q000188:drug therapy
chemical_list: D000911:Antibodies, Monoclonal; D001323:Autoantibodies; D011989:Receptors, Thyrotropin; D000242:Cyclic AMP