Characterization of the immunoreactivity of anti-interferon alpha antibodies in myasthenia gravis patients. Epitope mapping

Abstract

Cytokines, such as interferons (IFN), underlie many immunological functions and are increasingly implicated in disease-related symptoms and pathology. In order to study the potential roles of IFN alpha and its antagonists in autoimmune phenomena, the sera from 89 patients (aged 15-95 years, 65 females) diagnosed as having myasthenia gravis (MG) (2 months to 34 years duration) were tested for the presence of natural anti-IFN alpha-2b auto-antibodies. Sera were screened for anti-IFN alpha-2b by a sandwich-type enzyme immunoassay system. Ten (11.2%) and 6 (6.7%) sera were identified that contained positive-competing and non-competing anti-IFN alpha-2b auto-antibodies, respectively. The MG sera were further analyzed by immunobloting against reduced IFN alpha-2b and for neutralizing anti-IFN alpha activity in an antiviral assay cells system. From tested EIA positive-competing sera, 5 were shown to be positive by immunoblot and 6 sera were found to contain neutralizing anti-IFN alpha-2b. Four of the 6 neutralizing anti-IFN alpha-2b sera came from patients with thymoma-associated MG. The sera were studied for linear epitope recognition on the IFN alpha-2b molecule by a solid phase binding assay, in which overlapping peptides homologous with the entire IFN alpha-2b sequence were separately synthesized on a nitrocellulose sheet. Peptides number 2 (residues 8-21), 3 (15-28), 6 (33-46), 10 (63-76), 15 (98-112), and 21 (141-154) were immunoreactive. Peptide 21 was apparently associated with antiviral activity, although peptide 21 has not been previously described as an immunogenic determinant on the IFN alpha-2b molecule. These results indicate that neutralizing anti-IFN alpha-2b is often present in MG, particularly in cases of thymoma-associated MG, and recognize a variety of epitopes on the IFN alpha-2b molecule, including those involved in its biological activity. Two groups of IFN epitopes were described associated with patient's age but not with diseases evolution.