title: Spontaneous production of anti-IFN-alpha and anti-IL-12 autoantibodies by thymoma cells from myasthenia gravis patients suggests autoimmunization in the tumor.
abstract: Myasthenia gravis (MG) is mediated by autoantibodies to the acetylcholine receptor (AChR), expressed in muscle and rare thymic myoid cells. Most early-onset cases show thymic lymph node-type infiltrates, including pre-activated plasma cells spontaneously producing anti-AChR antibodies. Since these are not evident in the associated thymomas found in another 10% of MG patients, AChR-specific B cells must be autosensitized elsewhere. Unexpectedly, at diagnosis, >70% of MG/thymoma patients also have high-titer neutralizing autoantibodies to IFN-alpha, and >50% to IL-12; moreover, titers increase strikingly if the thymomas recur, indicating a closer tumor relationship than for anti-AChR. To investigate this, we have measured autoantibody production by cells cultured from thymomas, any available thymic remnants and blood, with or without the B cell stimulant pokeweed mitogen (PWM). To check autoantibody specificity and clonal origins, we isolated Fabs from two combinatorial libraries from producer thymus/thymoma cells. Surprisingly, thymoma cells spontaneously produced antibodies to IFN-alpha and/or IL-12 in >40% of seropositive cases, showing typical plasma cell behavior, whereas they produced anti-AChR only after PWM stimulation. We isolated 15 combinatorial Fabs to IFN-alpha (versus only one to AChR). Their strong binding in radio-immunoprecipitation and Western blots implies high affinities. The four Fabs tested neutralized anti-viral actions of IFN-alpha. The diverse V genes clearly showed ongoing antigen-driven selection. These results imply pre-activation in situ by native IFN-alpha/IL-12 expressed within a 'dangerous' tumor microenvironment. With these molecules, it should be easier to identify provoking cell type(s) that may give novel additional clues to autoimmunization against T-cell epitopes from the more complex AChR.
authors.affiliation: Neuroscience Group, Weatherall Institute for Molecular Medicine, University of Oxford, Oxford OX3 9DS, UK.
authors.forename: Hiroyuki
authors.initials: H
authors.lastname: Shiono
authors.forename: Yat Lei
authors.initials: YL
authors.lastname: Wong
authors.forename: Ian
authors.initials: I
authors.lastname: Matthews
authors.forename: Jian-Li
authors.initials: JL
authors.lastname: Liu
authors.forename: Wei
authors.initials: W
authors.lastname: Zhang
authors.forename: Gary
authors.initials: G
authors.lastname: Sims
authors.forename: Anthony
authors.initials: A
authors.lastname: Meager
authors.forename: David
authors.initials: D
authors.lastname: Beeson
authors.forename: Angela
authors.initials: A
authors.lastname: Vincent
authors.forename: Nicholas
authors.initials: N
authors.lastname: Willcox
mesh_terms: D000328:Adult; D000368:Aged; D000906:Antibodies / Q000276:immunology; D001323:Autoantibodies / Q000276:immunology*; D015551:Autoimmunity / Q000235:genetics / Q000276:immunology; D015153:Blotting, Western; D064211:Connectin; D005260:Female; D006801:Humans; D007140:Immunoglobulin Fab Fragments / Q000737:chemistry / Q000235:genetics / Q000276:immunology; D007135:Immunoglobulin Variable Region / Q000235:genetics / Q000276:immunology; D016898:Interferon-alpha / Q000097:blood / Q000276:immunology*; D018664:Interleukin-12 / Q000097:blood / Q000276:immunology*; D008297:Male; D008875:Middle Aged; D008969:Molecular Sequence Data; D009124:Muscle Proteins / Q000276:immunology; D009157:Myasthenia Gravis / Q000276:immunology* / Q000503:physiopathology; D009500:Neutralization Tests; D019151:Peptide Library; D011494:Protein Kinases / Q000276:immunology; D011863:Radioimmunoassay; D011950:Receptors, Cholinergic / Q000276:immunology; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013945:Thymoma / Q000276:immunology* / Q000473:pathology; D013950:Thymus Gland / Q000166:cytology / Q000276:immunology; D014407:Tumor Cells, Cultured / Q000276:immunology
chemical_list: D000906:Antibodies; D001323:Autoantibodies; D064211:Connectin; D007140:Immunoglobulin Fab Fragments; D007135:Immunoglobulin Variable Region; D016898:Interferon-alpha; D009124:Muscle Proteins; D019151:Peptide Library; D011950:Receptors, Cholinergic; C577285:TTN protein, human; D018664:Interleukin-12; D011494:Protein Kinases