초록

The present invention is concerned with epitope regions of a thyrotrophin (TSH) receptor, uses thereof and antibodies thereto.

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1. A method of screening a sample of body fluid for TSH receptor autoantibodies wherein said sample is from a subject suspected of suffering from, susceptible to, having or recovering from autoimmune disease associated with an immune reaction to a TSH receptor, said method comprising: (a) contacting

1. A method of screening a sample of body fluid for TSH receptor autoantibodies wherein said sample is from a subject suspected of suffering from, susceptible to, having or recovering from autoimmune disease associated with an immune reaction to a TSH receptor, said method comprising: (a) contacting said sample with: a polypeptide comprising a full length TSH receptor so as to permit said polypeptide to bind with TSH receptor autoantibodies present in said sample; andone or more murine monoclonal antibodies capable of competing with autoantibodies to TSH receptor in the binding of said full length TSH receptor, wherein said one or more monoclonal antibodies is positive for thyroid stimulating activity at a concentration of 20 μg/ml in a cyclic AMP thyroid cell assay and inhibits binding of TSH to TSH receptor, said one or more monoclonal antibodies being classified as positive if the thyroid stimulating activity of said one or more monoclonal antibodies is greater than 180 percent in the cyclic AMP thyroid cell assay wherein said percent thyroid stimulating activity comprises 100 X ratio of cyclic AMP produced in the presence of said one or more monoclonal antibodies to cyclic AMP produced in the presence of sera pooled from healthy blood donors; and(b) detecting binding of said polypeptide with said autoantibodies thereby providing an indication of the presence of said autoantibodies in said sample. 2. A method of diagnosing the likely onset or presence of autoimmune disease associated with an immune reaction to a TSH receptor in a subject suspected of suffering from, susceptible to, having or recovering from, autoimmune disease associated with an immune reaction to a TSH receptor, the method comprising detecting autoantibodies produced in response to a TSH receptor in a sample of body fluid from the subject according to claim 1, and whereby the detected autoantibodies can provide a diagnosis of the likely onset or presence of autoimmune disease associated with an immune reaction to a TSH receptor in the subject. 3. A method according to claim 1, wherein said one or more monoclonal antibodies comprises a binding affinity of at least 108 molar−1 for TSH receptor. 4. A method according to claim 1, wherein said one or more monoclonal antibodies is positive for thyroid stimulating activity at a concentration of 2 μg/ml in the cyclic AMP thyroid cell assay. 5. A method according to claim 1, wherein said one or more monoclonal antibodies includes a monoclonal antibody comprising: i) a VH domain wherein CDR1 comprises amino acid residues 31 to 35 of SEQ ID NO: 67, CDR2 comprises amino acid residues 50 to 66 of SEQ ID NO:67, and CDR3 comprises amino acid residues 99 to 109 of SEQ ID NO:67; and a VL domain wherein CDR1 comprises amino acid residues 24 to 34 of SEQ ID NO:68, CDR2 comprises amino acid residues 50 to 56 of SEQ ID NO:68, and CDR3 comprises amino acid residues 89 to 97 of SEQ ID NO: 68;ii) a VH domain wherein CDR1 comprises amino acid residues 31 to 35 of SEQ ID NO:69, CDR 2 comprises amino acid residues 50 to 66 of SEQ ID NO:69, and CDR 3 comprises amino acid residues 99 to 109 of SEQ ID NO:69, and a VL domain wherein CDR1 comprises amino acid residues 24 to 34 of SEQ ID NO:70, CDR2 comprises amino acid residues 50 to 56 of SEQ ID NO:70, and CDR3 comprises amino acid residues 89 to 97 of SEQ ID NO: 70; oriii) a VH domain wherein CDR1 comprises amino acid residues 31 to 35 of SEQ ID NO: 71, CDR2 comprises amino acid residues 50 to 66 of SEQ ID NO:71, and CDR 3 comprises amino acid residues 99 to 109 of SEQ ID NO:71, and a VL domain wherein CDR1 comprises amino acid residues 24 to 34 of SEQ ID NO:72, CDR2 comprises amino acid residues 50 to 56 of SEQ ID NO:72, and CDR3 comprises amino acid residues 89 to 97 of SEQ ID NO: 72. 6. A method according to claim 1, wherein said one or more monoclonal antibodies includes a monoclonal antibody comprising: i) a VH domain comprising amino acid residues 9 to 120 of SEQ ID NO:67 and a VL domain comprising amino acid residues 9 to 107 of SEQ ID NO:68;ii) a VH domain comprising amino acid residues 9 to 120 of SEQ ID NO:69 and a VL domain comprising amino acid residues 9 to 107 of SEQ ID NO:70; oriii) a VH domain comprising amino acid residues 9 to 120 of SEQ ID NO:71 and a VL domain comprising amino acid residues 9 to 107 of SEQ ID NO:72. 7. A method of screening a sample of body fluid for TSH receptor autoantibodies wherein said sample is from a subject suspected of suffering from, susceptible to, having or recovering from autoimmune disease associated with an immune reaction to a TSH receptor, said method comprising: (a) providing said sample of body fluid from said subject;(b) contacting said sample with (i) a full length TSH receptor comprising amino acid residues 246-260, wherein the amino acid sequence of residues 246-260 comprises amino acid residues 47 to 61 of SEQ ID NO:17 or 22, and(ii) one or more murine monoclonal antibodies capable of competing with autoantibodies to a TSH receptor in the binding of said full length TSH receptor, wherein said one or more competitor monoclonal antibodies is positive for thyroid stimulating activity at a concentration of 20 μg/ml in a cyclic AMP thyroid cell assay and inhibits binding of TSH to TSH receptor, said one or more monoclonal antibodies being classified as positive if the thyroid stimulating activity of said one or more monoclonal antibodies is greater than 180 percent in the cyclic AMP thyroid cell assay, said percent thyroid stimulating activity comprising 100× ratio of cyclic AMP produced in the presence of said one or more monoclonal competitor antibodies to cyclic AMP produced in the presence of sera pooled from healthy blood donors,so as to permit said full length TSH receptor with either autoantibodies to TSH receptor present in said sample, or said one or more competitor monoclonal antibodies; and(c) detecting the binding of said full length TSH receptor with said autoantibodies present in said sample, thereby providing an indication of the presence of said autoantibodies to a TSH receptor in said sample. 8. A method according to claim 7, wherein said one or more competitor monoclonal antibodies comprises a binding affinity of at least 108 molar−1 for TSH receptor. 9. A method according to claim 7, wherein said one or more monoclonal antibodies is positive for thyroid stimulating activity at a concentration of 2 μg/ml in the cyclic AMP thyroid cell assay. 10. A method according to claim 7, wherein said one or more monoclonal antibodies includes a monoclonal antibody comprising: i) a VH domain wherein CDR1 comprises amino acid residues 31 to 35 of SEQ ID NO: 67, CDR2 comprises amino acid residues 50 to 66 of SEQ ID NO:67, and CDR3 comprises amino acid residues 99 to 109 of SEQ ID NO:67; and a VL domain wherein CDR1 comprises amino acid residues 24 to 34 of SEQ ID NO:68, CDR2 comprises amino acid residues 50 to 56 of SEQ ID NO:68, and CDR3 comprises amino acid residues 89 to 97 of SEQ ID NO: 68;ii) a VH domain wherein CDR1 comprises amino acid residues 31 to 35 of SEQ ID NO:69, CDR 2 comprises amino acid residues 50 to 66 of SEQ ID NO:69, and CDR 3 comprises amino acid residues 99 to 109 of SEQ ID NO:69, and a VL domain wherein CDR1 comprises amino acid residues 24 to 34 of SEQ ID NO:70, CDR2 comprises amino acid residues 50 to 56 of SEQ ID NO:70, and CDR3 comprises amino acid residues 89 to 97 of SEQ ID NO: 70; oriii) a VH domain wherein CDR1 comprises amino acid residues 31 to 35 of SEQ ID NO: 71, CDR2 comprises amino acid residues 50 to 66 of SEQ ID NO:71, and CDR 3 comprises amino acid residues 99 to 109 of SEQ ID NO:71, and a VL domain wherein CDR1 comprises amino acid residues 24 to 34 of SEQ ID NO:72, CDR2 comprises amino acid residues 50 to 56 of SEQ ID NO:72, and CDR3 comprises amino acid residues 89 to 97 of SEQ ID NO: 72. 11. A method according to claim 7, wherein said one or more monoclonal antibodies includes a monoclonal antibody comprising: i) a VH domain comprising amino acid residues 9 to 120 of SEQ ID NO:67 and a VL domain comprising amino acid residues 9 to 107 of SEQ ID NO:68;ii) a VH domain comprising amino acid residues 9 to 120 of SEQ ID NO:69 and a VL domain comprising amino acid residues 9 to 107 of SEQ ID NO:70; oriii) a VH domain comprising amino acid residues 9 to 120 of SEQ ID NO:71 and a VL domain comprising amino acid residues 9 to 107 of SEQ ID NO:72. 12. A method of screening a sample of body fluid for autoantibodies to a TSH receptor wherein said sample is from a subject suspected of suffering from, susceptible to, having or recovering from autoimmune disease associated with an immune reaction to a TSH receptor, said method comprising: (a) contacting said sample with (i) a full length TSH receptor, one or more epitopes thereof or a and(ii) one or more murine monoclonal antibodies for the TSH receptor, wherein said one or more monoclonal antibodies: does not comprise naturally produced autoantibodies to the TSH receptor,inhibits binding of TSH to TSH receptor, andis positive for thyroid stimulating activity at a concentration of 20 μg/ml in a cyclic AMP thyroid cell assay, said one or more monoclonal antibodies being classified as positive if the thyroid stimulating activity of said one or more monoclonal antibodies is greater than 180 percent in the cyclic AMP thyroid cell assay, wherein said percent thyroid stimulating activity comprises 100× ratio of cyclic AMP produced in the presence of said one or more monoclonal antibodies to cyclic AMP produced in the presence of sera pooled from healthy blood donors;so as to permit said TSH receptor to bind with either autoantibodies to TSH receptor present in said sample, or said one or more monoclonal antibodies for the TSH receptor; and(b) detecting binding of said TSH receptor, with said autoantibodies present in said sample, thereby providing an indication of the presence of said autoantibodies to TSH receptor in said sample. 13. A method according to claim 12, which comprises providing labeling means for said one or more monoclonal antibodies for a TSH receptor. 14. A method according to claim 12, wherein said one or more monoclonal antibodies comprises a binding affinity of at least 108 molar−1 for TSH receptor. 15. A method according to claim 12, wherein said one or more monoclonal antibodies is positive for thyroid stimulating activity at a concentration of 2 μg/ml in the cyclic AMP thyroid cell assay. 16. A method according to claim 12, wherein said one or more monoclonal antibodies includes a monoclonal antibody comprising: i) a VH domain wherein CDR1 comprises amino acid residues 31 to 35 of SEQ ID NO: 67, CDR2 comprises amino acid residues 50 to 66 of SEQ ID NO:67, and CDR3 comprises amino acid residues 99 to 109 of SEQ ID NO:67; and a VL domain wherein CDR1 comprises amino acid residues 24 to 34 of SEQ ID NO:68, CDR2 comprises amino acid residues 50 to 56 of SEQ ID NO:68, and CDR3 comprises amino acid residues 89 to 97 of SEQ ID NO: 68;ii) a VH domain wherein CDR1 comprises amino acid residues 31 to 35 of SEQ ID NO:69, CDR 2 comprises amino acid residues 50 to 66 of SEQ ID NO:69, and CDR3 comprises amino acid residues 99 to 109 of SEQ ID NO:69, and a VL domain wherein CDR1 comprises amino acid residues 24 to 34 of SEQ ID NO:70, CDR2 comprises amino acid residues 50 to 56 of SEQ ID NO:70, and CDR3 comprises amino acid residues 89 to 97 of SEQ ID NO: 70; oriii) a VH domain wherein CDR1 comprises amino acid residues 31 to 35 of SEQ ID NO: 71, CDR2 comprises amino acid residues 50 to 66 of SEQ ID NO:71, and CDR3 comprises amino acid residues 99 to 109 of SEQ ID NO:71, anda VL domain wherein CDR1 comprises amino acid residues 24 to 34 of SEQ ID NO:72, CDR2 comprises amino acid residues 50 to 56 of SEQ ID NO:72, and CDR3 comprises amino acid residues 89 to 97 of SEQ ID NO: 72. 17. A method according to claim 12, wherein said one or more monoclonal antibodies includes a monoclonal antibody comprising: i) a VH domain comprising amino acid residues 9 to 120 of SEQ ID NO:67 and a VL domain comprising amino acid residues 9 to 107 of SEQ ID NO:68;ii) a VH domain comprising amino acid residues 9 to 120 of SEQ ID NO:69 and a VL domain comprising amino acid residues 9 to 107 of SEQ ID NO:70; oriii) a VH domain comprising amino acid residues 9 to 120 of SEQ ID NO:71 and a VL domain comprising amino acid residues 9 to 107 of SEQ ID NO:72. 18. A method of screening a sample of body fluid for TSH receptor autoantibodies wherein said sample is from a subject suspected of suffering from, susceptible to, having or recovering from autoimmune disease associated with an immune reaction to a TSH receptor, said method comprising: (a) contacting said sample with (i) a full length TSH receptor, and(ii) one or more murine monoclonal antibodies for said TSH receptor, wherein said one or more monoclonal antibodies is positive for thyroid stimulating activity at a concentration of 20 μg/ml in a cyclic AMP thyroid cell assay and inhibits binding of TSH to TSH receptor, said one or more monoclonal antibodies being classified as positive if the thyroid stimulating activity of said one or more monoclonal antibodies is greater than 180 percent in the cyclic AMP thyroid cell assay, said percent thyroid stimulating activity comprising 100× ratio of cyclic AMP produced in the presence of said one or more monoclonal antibodies to cyclic AMP produced in the presence of sera pooled from healthy blood donors;so as to permit said TSH receptor to bind with either autoantibodies to said TSH receptor present in said sample, or said one or more monoclonal antibodies; and(b) detecting binding of said TSH receptor with said autoantibodies present in said sample, thereby providing an indication of the presence of said autoantibodies to TSH receptor in said sample; wherein said one or more monoclonal antibodies for said TSH receptor are directly or indirectly immobilised to a surface either prior to, or after step (a). 19. A method according to claim 18, wherein said one or more monoclonal antibodies does not comprise naturally produced autoantibodies to the TSH receptor. 20. A method according to claim 18, which comprises providing labeling means for said TSH receptor. 21. A method according to claim 18, wherein said one or more monoclonal antibodies comprises a binding affinity of at least 108 molar−1 for TSH receptor. 22. A method according to claim 18, wherein said one or more monoclonal antibodies is positive for thyroid stimulating activity at a concentration of 2 μg/ml in the cyclic AMP thyroid cell assay. 23. A method according to claim 18, wherein said one or more monoclonal antibodies includes a monoclonal antibody comprising: i) a VH domain wherein CDR1 comprises amino acid residues 31 to 35 of SEQ ID NO: 67, CDR2 comprises amino acid residues 50 to 66 of SEQ ID NO:67, and CDR3 comprises amino acid residues 99 to 109 of SEQ ID NO:67; and a VL domain wherein CDR1 comprises amino acid residues 24 to 34 of SEQ ID NO:68, CDR2 comprises amino acid residues 50 to 56 of SEQ ID NO:68, and CDR3 comprises amino acid residues 89 to 97 of SEQ ID NO: 68;ii) a VH domain wherein CDR1 comprises amino acid residues 31 to 35 of SEQ ID NO:69, CDR 2 comprises amino acid residues 50 to 66 of SEQ ID NO:69, and CDR3 comprises amino acid residues 99 to 109 of SEQ ID NO:69, and a VL domain wherein CDR1 comprises amino acid residues 24 to 34 of SEQ ID NO:70, CDR2 comprises amino acid residues 50 to 56 of SEQ ID NO:70, and CDR3 comprises amino acid residues 89 to 97 of SEQ ID NO: 70; oriii) a VH domain wherein CDR1 comprises amino acid residues 31 to 35 of SEQ ID NO: 71, CDR2 comprises amino acid residues 50 to 66 of SEQ ID NO:71, and CDR3 comprises amino acid residues 99 to 109 of SEQ ID NO:71, and a VL domain wherein CDR1 comprises amino acid residues 24 to 34 of SEQ ID NO:72, CDR2 comprises amino acid residues 50 to 56 of SEQ ID NO:72, and CDR3 comprises amino acid residues 89 to 97 of SEQ ID NO: 72. 24. A method according to claim 18, wherein said one or more monoclonal antibodies includes a monoclonal antibody comprising: i) a VH domain comprising amino acid residues 9 to 120 of SEQ ID NO:67 and a VL domain comprising amino acid residues 9 to 107 of SEQ ID NO:68;ii) a VH domain comprising amino acid residues 9 to 120 of SEQ ID NO:69 and a VL domain comprising amino acid residues 9 to 107 of SEQ ID NO:70; oriii) a VH domain comprising amino acid residues 9 to 120 of SEQ ID NO:71 and a VL domain comprising amino acid residues 9 to 107 of SEQ ID NO:72.