Quantitative analysis of methylation defects and correlation with clinical characteristics in patients with pseudohypoparathyroidism type I and GNAS epigenetic alterations

Pseudohypoparathyroidism type I (PHP-I) includes two main subtypes, PHP-Ia and -Ib. About 70% of PHP-Ia patients, who show Albright hereditary osteodystrophy (AHO) associated with resistance toward multiple hormones (PTH/TSH/GHRH/gonadotropins), carry heterozygous mutations in the α-subunit of the stimulatory G protein (Gsα) exons 1-13, encoded by the guanine nucleotide binding-protein α-stimulating activity polypeptide 1 (GNAS), whereas the majority of PHP-Ib patients, who classically display hormone resistance limited to PTH and TSH with no AHO sign, have methylation defects in the imprinted GNAS cluster. Recently methylation defects have been detected also in patients with PHP and different degrees of AHO, indicating a molecular overlap between the two forms.

The objectives of the study were to collect patients with the following characteristics: clinical PHP-I (with or without AHO), no mutation in Gsα coding sequence, but the presence of GNAS methylation alterations and to investigate the existence of correlations between the degree of the epigenetic defect and the severity of the disease.

We quantified GNAS methylation alterations by both PCR-pyrosequencing and methylation specific-multiplex ligation-dependent probe amplification assay in genomic DNA from 63 patients with PHP-I and correlated these findings with clinical parameters (age at diagnosis; calcium, phosphorus, PTH, TSH levels; presence or absence of each AHO sign).

By both approaches, the degree of the imprinting defect did not correlate with the onset of the disease, the severity of endocrine resistances, or with the presence/absence of specific AHO signs.

Similar molecular alterations may lead to a broad spectrum of diseases, from isolated PTH resistance to complete PHP-Ia, and the degree of methylation alterations does not reflect or anticipate the severity and the type of different PHP/AHO manifestations.