Expression of human-Torpedo hybrid acetylcholine receptor (AChR) for analysing the subunit specificity of antibodies in sera from patients with myasthenia gravis (MG).

Abstract

The nicotinic AChR, a pentamer composed of alpha2betagamma(or epsilon)delta subunits, is the autoantigen in the human autoimmune disease MG. Anti-AChR antibodies in MG sera bind mainly to conformational epitopes, therefore determination of their specificities requires the use of intact AChR. Indirect antibody competition studies have suggested that most MG antibodies are inhibited from binding to AChR by MoAb to the main immunogenic region (MIR) on the alpha-subunits. More recently, based on the knowledge that MG antibodies show little detectable cross-reaction with Torpedo AChR, we have shown, using mouse-Torpedo hybrid AChR, that most MG antibodies that detectably cross-react with the mouse AChR bind to the alpha-subunit. To analyse the whole anti-AChR antibody repertoire in MG sera, we expressed on stably transfected fibroblasts a novel human alpha+ Torpedo betagammadelta AChR and compared the antibody titres against human, Torpedo, and the hybrid AChR. Direct information was provided for the subunit specificity of several MoAbs and sera from 50 MG patients. On average, at least 48% of the anti-AChR antibodies in the sera were directed against the alpha-subunit. Interestingly, the anti-alpha-subunit antibodies predominated in low titre (0.6-7.4 nM) but not in high titre (10-386 nM) sera, where they comprised on average 68% versus 23% of the antibodies, respectively. Finally, the directly determined anti-alpha-subunit antibodies and the anti-MIR antibodies defined by antibody competition were significantly correlated, thus suggesting that at least a significant fraction of the anti-MIR antibodies in MG sera bind to the alpha-subunit.