Is there loss or qualitative changes in the expression of thyroid peroxidase protein in thyroid epithelial cancer?

There is disagreement concerning the expression of thyroid peroxidase (TPO) in thyroid cancer, some studies finding qualitative as well as quantitative differences compared to normal tissue. To investigate TPO protein expression and its antigenic properties, TPO was captured from a solubilizate of thyroid microsomes by a panel of murine anti-TPO monoclonal antibodies and detected with a panel of anti-human TPO IgGkappa Fab. TPO protein expression in 30 samples of malignant thyroid tissue was compared with TPO from adjacent normal tissues. Virtual absence of TPO expression was observed in 8 cases. In the remaining 22 malignant thyroid tumours the TPO protein level varied considerably from normal to nearly absent when compared to normal thyroid tissue or tissues from patients with Graves' disease (range less than 0.5 to more than 12.5 microg mg(-1) of protein). When expressed TPO displayed similar epitopes, to that of TPO from Graves' disease tissue. The results obtained by the TPO capturing method were confirmed by SDS-PAGE and Western blot analysis with both microsomes and their solubilizates. The present results show that in about two-thirds of differentiated thyroid carcinomas, TPO protein is expressed, albeit to a more variable extent than normal; when present, TPO in malignant tissues is immunologically normal.

In old age the majority of thyroid peroxidase autoantibodies are directed to a single TPO domain irrespective of thyroid function and iodine intake.

OBJECTIVE: We have examined (1) which epitopes on thyroid peroxidase (TPO) are recognized by TPO autoantibodies (TPO-Aab) in old age and to what extent? (2) Does the TPO-Aab pattern differ in euthyroid and hypothyroid elderly subjects or does it depend on their iodine intake? DESIGN: TPO-Aab positive sera obtained from a screening study of nursing-home residents living in areas of varying iodine intake were tested by competition studies with monoclonal antibodies (mAbs) recognizing different epitopes on TPO. SUBJECTS: The nursing-home residents with TPO-Aab positivity were from (A) an iodine abundant area (Eastern Hungary, median iodine excretion -MIE-: 0.462 mumol/mmol creatinine, N = 13); (B) an area of obligatory iodinated salt prophylaxis since the 1950s (Slovakia, MIE: 0.090 mumol/mmol creatinine, N = 11); (C) a moderately iodine-deficient area (Northern Hungary, MIE: 0.065 mumol/mmol creatinine, N = 13). MEASUREMENTS: Thirteen murine TPO antibodies generated against several epitopes of the four (A, B, C, D) antigenic domains on the TPO were co-incubated with the TPO-Aab positive sera on TPO coated microtitre plates. The amount of mAb bound was estimated after further incubation with goat anti-mouse antibodies, conjugated with horseradish peroxidase and tetramethylbenzidine as chromogen. The TPO-Aab positive sera were characterized by the pattern of percentage of inhibition of mAb binding caused by the TPO-Aabs. RESULTS: TPO-Aabs inhibited only the binding of mAbs raised against the antigenic domains A (mAb9, mAb2, mAb60) and B (mAb64, mAb59, mAb18, mAb15). The extent of inhibition depended upon the TPO-Aab titre but in all cases the binding of mAb9 was inhibited to the highest degree. The percentage inhibition of mAb9 was (a) 34 +/- 17% (M +/- SD) caused by sera (N = 8) with TPO-Aab titre 1/100-1/200 (higher than that of all mAbs recognizing domain B, P < 0.01-P < 0.001), (b) 76 +/- 18% caused by sera (N = 14) with TPO-Aab titre 1/1000 (higher than that of all other mAbs -P < 0.01-P < 0.001, except mAb64), (c) 99 +/- 4% caused by sera (N = 15) with TPO-Aab titre 1/4000-1/16,000 (higher than that of all other mAbs, P < 0.01-P < 0.001). Thus, only mAb9 was inhibited completely by high titres of TPO-Aabs. The qualitative and quantitative distribution pattern of mAb inhibition was similar in the subgroups of elderly hypothyroid and euthyroid subjects with comparable TPO-Aab levels, as well as in the subgroups with varying iodine intake. CONCLUSIONS: (1) In old age, there is a polyclonal TPO autoantibody response but the majority of the autoantibodies are directed to the TPO region mapped by or close to mAb9 (domain A); (2) the autoantibody response does not differ in elderly subjects with or without the clinical manifestations of autoimmune thyroid disease and does not depend on the iodine supply of the elderly subjects.

Majority of thyroid peroxidase autoantibodies in patients with autoimmune thyroid disease are directed to a single TPO domain.

Murine monoclonal antibodies (mAb) produced against native human thyroid peroxidase (TPO) are powerful tools for analyzing the autoantibody (Aab) epitopes on TPO. Binding sites of thirteen mAbs cover all or most antigenic regions on TPO. We determined the competition between Aabs from 75 AITD patients and 13 mAbs in binding to TPO. Autoantibodies recognize predominantly the TPO area close or identical to mAb#9 epitope. All sera tested inhibited this mAb binding by 92.9 +/- 14.8 (mean +/- SD), range from 69-100%. AITD patients' sera with low Aabs titer up to 1/2,000 inhibited mAb#9 binding to TP0 by 85 +/- 11.5% (mean +/- SD) and did not influence remaining mAbs binding to TPO. With elevated Aab levels the inhibition of other mAbs binding was higher, but never exceeded 35%. The amount of Aabs yielding 50% inhibition of mAbs binding was lowest for mAb#9. In order to obtain this degree of inhibition for other mAbs 5 to 25 times more Aabs were needed. Our results demonstrate that the majority of autoantibodies in sera of patients with AITD recognize a single immunodominant region on the TPO mapped by mAb#9. They account for about 80-90% of serum TPO autoantibodies. The autoimmune response to other regions on TPO molecule is directed to several other epitopes, but represents quantitatively a minority of autoantibodies. This response intensifies with increasing Aabs level in the serum.