LETTER TO JMG Ancestral RET haplotype associated with Hirschsprung’s disease shows linkage disequilibrium breakpoint at 21249

T he RET proto-oncogene, located at 10q11.2, encodes receptor tyrosine kinase expressed during neural crest development and is involved in different human neurocristopathies such as the multiple endocrine neoplasia type 2 (MEN 2) syndromes and Hirschsprung’s disease (HSCR). HSCR (OMIM 142623), or aganglionic megacolon, is a common developmental disorder characterised by the absence of enteric neurones in variable lengths of the distal gastrointestinal tract, resulting in functional intestinal obstruction. Although RET is considered to be the major gene involved in HSCR, only a subset of HSCR patients can be attributable to traditional germline RET mutations (50% of familial HSCR and 10–15% of sporadic cases in selected series, 3 or 30% of familial and 3% of sporadic cases in a population based study). In an even smaller subset of HSCR patients (5–10%), germline variants are present in other genes, generally related to the developmental programme of neural crest cells, such as the glial cell line derived neurotrophic factor (GDNF), neurturin (NTN), endothelin 3 (EDN3), endothelin receptor b (EDNRB), endothelin converting enzyme 1 (ECE1), transcriptional factors SOX10 and PHOX2B, and Smad interacting protein 1 (SIP1). In addition, RET modifying gene loci at 9q31, 3p21, 10q11, and 19q12 have been described associated with an HSCR phenotype. 9 Because traditional germline mutations accounted for such a small subset of HSCR, we sought to determine whether there are more common susceptibility factors which predispose to a majority of HSCR cases. The silent variant A45A was described by our group in the only member from a MEN 2A/HSCR family co-segregating both phenotypes. When this variant in codon 45 (exon 2) and the RET haplotypes carrying it were found to be highly associated with a large subset of non-familial HSCR, 12 it prompted us to propose that this was the associated variant itself, or that A45A was in linkage disequilibrium with the putative low penetrance susceptibility locus that would account for the majority of HSCR cases. Subsequently, we have shown that A45A anchors ancestral haplotypes in linkage disequilibrium, with the putative common founding susceptibility locus estimated to be 22 to 50 kb upstream. These observations have been confirmed in other populations, suggesting that this founder locus dates to the Stone Age. In this study we have undertaken systematic screening of the region upstream of the anchoring A45A single nucleotide polymorphism (SNP)—comprising RET intron 1, exon 1, and promoter—in 117 population based HSCR cases to find this putative founding locus.