Specific binding sites for synthetic growth hormone secretagogues in non-tumoral and neoplastic human thyroid tissue

Abstract

The presence of specific receptors for synthetic growth hormone secretagognes (GHSs) has been investigated in non-tumoral and neoplastic human thyroid tissue using a radio-iodinated peptidyl GHS (125I-labelled Tyr-Ala-hexarelin) as ligand. Specific binding sites for Tyr-Ala-hexarelin were detected in membranes from non-tumoral and follicular-derived neoplastic thyroid tissue, but not in thyroid tumours (medullary carcinomas) of parafollicular (C cell) origin. The binding activity was greatest in well differentiated neoplasms (papillary and follicular carcinomas), followed by poorly differentiated carcinomas, non-tumoral thyroid parenchyma, follicular adenomas and anaplastic carcinomas. Both peptidyl (Tyr-Ala-hexarelin, hexarelin, growth hormone releasing peptide (GHRP6) and non-peptidyl (MK 0677) GHSs completely displaced the radioligand from binding sites of non- tumoral thyroid gland, but MK 0677 was significantly less potent. The IC50 values were (1.9±0.3) x 10-8 mol/l for Tyr-Ala-hexarelin, (2.1 ± 0.2) x 10-8 mol/l for hexarelin, (2.4± 0.3) x 10-8 mol/l for GHRP6 and only (1.5 ± 0.4) x 10-7 mol/l for MK 0677. Similar IC50 values were found in neoplastic thyroid tissue. Scatchard analysis of the binding revealed a finite number of binding sites in non-tumoral (B(max): 1232 ± 32 fmol/mg protein, n=3) and neoplastic (papillary carcinomas) thyroid tissue (B(max): 2483 ± 380 fmol/mg protein, n=5), with dissociation constants (K(d)) of (3.8 ± 0.3) x 10-9 and (4.4 ± 0.6) x 10-9 mol/l, respectively. On the basis of this evidence, we investigated the effects of some GHS on the proliferation of three different human follicular thyroid carcinoma cell lines (NPA, WRO and ARO) in which the presence of specific GHS receptors was also demonstrated. Tyr-Ala-hexarelin, GHRP6 and MK 0677 were able to inhibit serum-stimulated [3H]thymidine incorporation in NPA cells at concentrations close to their binding affinity. These substances also caused a significant inhibition of cell proliferation, which was evident at the earliest time of treatment (24 h) in all the cell lines, and at the latest time (96 h) in NPA cells only. In conclusion, this paper confirms the existence of specific binding sites for GHS in normal thyroid tissue and demonstrates, for the first time, that these binding sites are present in papillary and follicular carcinomas, low in anaplastic carcinomas and absent in medullary carcinomas of the thyroid. This work also provides evidence of a growth-inhibitory effect of GHS on cell lines derived from follicular thyroid cancers.