Endocrine Abstracts

In Graves' disease (GD), it is recognised that isolation of the rare pathogenic autoantibodies that stimulate the thyrotropin receptor (TSHR) would assist in understanding disease pathogenesis. To date, monoclonal antibodies (mabs) to the TSHR with thyroid stimulating activity (TSAbs) have been isolated from experimental models of GD which are 100 fold weaker than the reported single human mab with strong TSAb activity. We describe two novel IgG mabs from a murine model of GD induced with recombinant adenovirus expressing TSHR, with TSAb potencies comparable to the human mab. In assessing their ability to stimulate cAMP in JP09 cells, the mabs KSAb1 and KSAb2 demonstrated EC50s of 0.12 nanomolar plus/minus 0.04 and 1.1 nanomolar plus/minus 0.04 respectively. Notably, they exhibited full agonistic activity to the human TSHR under physiological ionic conditions; the maximal cAMP ceiling response was obtained with 30 and 3000 nanograms per millilitre respectively. Moreover, a 3 fold stimulation of cAMP over basal levels was obtained with 1.2 nanograms per millilitre plus/minus 0.5 and 2.7 nanograms per millilitre plus/minus 0.5 respectively. Interestingly, they displayed differential activities in their ability to block TSH mediated stimulation of cAMP with KSAb2 demonstrating >20% blocking activity. In TSH binding inhibition (TRAK) assays, nanogram doses of the mabs led to complete (>98%) inhibition of labelled TSH binding. Neither mab reacted with a panel of overlapping synthetic peptides representing the entire extra-cellular region of TSHR providing compelling evidence for recognition of conformational dependent epitopes on the receptor. In summary, KSAb1 and KSAb2 represent the first description of mabs from an experimental model reported to date with potencies matching those arising spontaneously in human disease. They emphasise the closeness of the experimental model in reproducing the human disorder and provide improved tools to explore the immune mechanisms implicated in the pathogenesis of autoimmune hyperthyroidism.