Pathogenic and likely pathogenic genetic alterations and polymorphisms in growth hormone gene (GH1) and growth hormone releasing hormone receptor gene (GHRHR) in a cohort of isolated growth hormone deficient (IGHD) children in Sri Lanka

Objective: Genetic alterations in GH1 and GHRHR genes are known to cause isolated growth hormone deficiency (IGHD). Of these, GHRHR codon 72 mutation has been reported to be highly prevalent in the Indian subcontinent, but among Sri Lankans its prevalence was low compared to reports from neighboring countries. The present study was therefore carried out to identify genetic alterations in the GH1 gene and rest of the GHRHR gene in a cohort of Sri Lankan IGHD patients who tested negative for GHRHR codon 72 mutation.

Methods: Fifty five IGHD children negative for codon 72 (GHRHR) mutation were screened for gross GH1 gene deletion by polymerase chain reaction (PCR) and restriction fragment length polymorphism technique. The coding, intronic and promoter regions of the GH1 gene were sequenced in children who were negative for GH1 deletion (N=53). In a subset (N=40), coding, flanking intronic and promoter regions of the GHRHR gene were screened by single strand conformation polymorphism/sequencing. Identified coding region and intronic variants were subjected to in silico analysis to ascertain pathogenicity. Family members available were screened for the significant variants observed in the index child.

Results: Gross GH1 gene deletions, 6.7kb and 7.0kb were observed in one child each. One novel and 24 reported single nucleotide variants (SNVs) were observed in the GH1 gene and its promoter. These included one reported pathogenic splice site mutation (c.172-2A>T) and one reported likely pathogenic missense mutation (c.406G>T). One large novel deletion of 5875 base pairs that included exon 1, one likely pathogenic novel SNV (c.211G>T) and 18 reported SNVs were observed in the GHRHR gene. Fourteen variants observed were of uncertain significance (8 in GH1 and 6 in GHRHR), twenty three variants were likely benign (11 in GH1 and 12 in GHRHR) and four variants were benign (4 in GH1 and none in GHRHR).

Conclusion: In a cohort of IGHD children, six pathogenic or likely pathogenic genetic alterations of either GH1 gene or GHRHR gene were found. These affected a total of six children. Pathogenic status of four of these had been reported in the literature. Novel SNV in the GHRHR gene was predicted to be pathogenic through in silico analysis. The large novel deletion is likely to be pathogenic as it included exon 1 of GHRHR gene. Analysis of other genes will be needed to ascertain the genetic cause of IGHD in the remaining children.