Reliable screening for a pain-protective haplotype in the GTP cyclohydrolase 1 gene (GCH1) through the use of 3 or fewer single nucleotide polymorphisms.

Background: A haplotype in the GTP cyclohydrolase 1 (dopa-responsive dystonia) gene (GCH1) is associated with decreased persistent pain. The aim of the present study was to develop a screening method for the pain-protective haplotype.

Methods: Complete genetic information for all 15 GCH1 DNA positions constituting the pain-protective GCH1 haplotype was available from 278 patients. In silico analyses, including discriminant analysis of the most frequent haplotypes, identified distinctive DNA positions that allow detection of the pain-protective haplotype at high sensitivity and specificity with the smallest possible number of DNA positions. Pyrosequencing(trade mark) assays were subsequently developed for these DNA positions, established with 662 DNA samples from healthy volunteers, and prospectively validated with a random selection of DNA samples genotyped for all 15 DNA positions.

Results: Diagnosis of the pain-protective GCH1 haplotype was possible with 100% sensitivity and specificity by screening for just 3 GCH1 genetic variants that span the entire DNA range of the haplotype: c.-9610G>A (dbSNP rs8007267G>A) in the 5' untranslated region, c.343 + 8900A>T (dbSNP rs3783641A>T) in intron 1, and c.*4279 (dbSNP rs10483639C>G) in the 3' untranslated region. Test sensitivity and specificity were still >95% with 2 or even just 1 of these GCH1 DNA positions.

Conclusions: In silico analysis of complex GCH1 gene haplotypes reduced the requisite number of tested DNA positions from 15 to 3 while maintaining the reliability, specificity, and sensitivity of the genetic diagnosis. This screening method could reduce laboratory diagnostic efforts and facilitate investigations of the pain-protective GCH1 haplotype.