Future therapeutic strategies in autoimmune myasthenia gravis.
Antibodies against muscle acetylcholine receptor (AChR) undoubtedly play a critical role in the pathology of most myasthenia gravis (MG) cases. Selective elimination of the majority of these antibodies should result in a considerable improvement of the MG symptoms. Such a specific elimination could be achieved by AChR-based immunoadsorbents. However, sufficient quantities of native human AChR are not available while bacterially expressed recombinant domains of the AChR are unable to bind satisfactorily MG antibodies. We have undertaken the production of the extracellular domains of human AChR subunits in eukaryotic systems, in native-like conformation, for their use as potent immunoadsorbents. The N-terminal extracellular domain (amino acids 1-210; alpha(1-210)) of the alpha(1) subunit of the human muscle AChR was expressed in the yeast Pichia pastoris. The polypeptide was water-soluble, glycosylated, and in monomer form. The alpha(1-210) bound 125I-alpha-bungarotoxin (125I-alpha-BTX) with a high affinity (Kd = 5.1 +/- 2.4 nM), and this binding was blocked by unlabeled d-tubocurarine and gallamine. Several conformation-dependent anti-AChR antibodies were able to bind alpha(1-210) as did antibodies from a large proportion of MG patients. The purified protein was subsequently immobilized on Sepharose-CNBr and was used to immunoadsorb anti-AChR antibodies from 64 MG sera. It eliminated more than 50% (50-94%) of the anti-AChR antibodies in 20% of the sera, whereas from another 30% of the sera it eliminated 20-60% of their anti-AChR antibodies. Work is in progress for the expression of the extracellular domain of all other muscle AChR subunits. It is expected that their combined use may eliminate the great majority of the anti-AChR antibodies from most MG patients.
Psaridi-Linardaki
Loukia
L
Department of Biochemistry, Hellenic Pasteur Institute, 11521 Athens, Greece.
Mamalaki
Avgi
A
Tzartos
Socrates J
SJ
D000818:Animals; D000888:Antibodies, Anti-Idiotypic; D000911:Antibodies, Monoclonal / Q000378:metabolism; D000918:Antibody Specificity; D001323:Autoantibodies / Q000276:immunology* / Q000378:metabolism; D001667:Binding, Competitive; D015153:Blotting, Western; D002038:Bungarotoxins / Q000378:metabolism; D002217:Carbachol / Q000493:pharmacokinetics; D018678:Cholinergic Agents / Q000378:metabolism; D018679:Cholinergic Agonists / Q000493:pharmacokinetics; D002846:Chromatography, Affinity; D004305:Dose-Response Relationship, Drug; D004797:Enzyme-Linked Immunosorbent Assay; D005703:Gallamine Triethiodide / Q000493:pharmacokinetics; D006031:Glycosylation; D006801:Humans; D007163:Immunosorbent Techniques; D008024:Ligands; D009157:Myasthenia Gravis / Q000097:blood / Q000276:immunology / Q000628:therapy*; D009538:Nicotine / Q000493:pharmacokinetics; D018733:Nicotinic Antagonists / Q000493:pharmacokinetics; D011233:Precipitin Tests; D011487:Protein Conformation; D011950:Receptors, Cholinergic / Q000276:immunology; D011994:Recombinant Proteins / Q000276:immunology; D014403:Tubocurarine / Q000493:pharmacokinetics
D000888:Antibodies, Anti-Idiotypic; D000911:Antibodies, Monoclonal; D001323:Autoantibodies; D002038:Bungarotoxins; D018678:Cholinergic Agents; D018679:Cholinergic Agonists; D008024:Ligands; D018733:Nicotinic Antagonists; D011950:Receptors, Cholinergic; D011994:Recombinant Proteins; D009538:Nicotine; D002217:Carbachol; D005703:Gallamine Triethiodide; D014403:Tubocurarine