High-throughput Molecular Analysis of Pseudohypoparathyroidism 1b Patients Reveals Novel Genetic and Epigenetic Defects

Context: Patients with pseudohypoparathyroidism type 1b (PHP1b) show disordered imprinting of the maternal GNAS allele or paternal uniparental disomy (UPD). Genetic deletions in STX16 or in upstream exons of GNAS are present in many familial but not sporadic cases. Objective: Characterization of epigenetic and genetic defects in patients with PHP1b. Design and Patients: DNA from 84 subjects, including 26 subjects with sporadic PHP1b, 27 affected subjects and 17 unaffected and/or obligate gene carriers from 12 PHP1b families, 11 healthy individuals, and 3 subjects with PHP1a was subjected to quantitative pyrosequencing of GNAS differentially methylated regions (DMRs), microarray analysis, and microsatellite haplotype analysis. Setting: Academic medical center. Main Outcome Measurements: Molecular pathology of PHP1b. Results: Healthy subjects, unaffected family members and obligate carriers of paternal PHP1b alleles, and subjects with PHP1a showed normal methylation of all DMRs. All PHP1b subjects showed loss of methylation (LOM) at the exon A/B DMR. Affected members of 9 PHP1b kindreds showed LOM only at the exon A/B DMR, which was associated with a 3-kb deletion of STX16 exons 4 through 6 in 7 families and a novel deletion of STX16 and adjacent NEPEPL1 in 1 family. A novel NESP deletion was found in 1 of 2 other families with more extensive methylation defects. One sporadic PHP1b had UPD of 20q, 2 had 3-kb STX16 deletions, and 5 had apparent epigenetic mosaicism. Conclusions: We found diverse patterns of defective methylation and identified novel or previously known mutations in 9 of 12 PHP1b families.