In addition, case-control analysis demonstrated that RETrs2435357 is significantly correlated with HSCR (P = 2.2 × 10(-13)) with an odds ratio of 5.1 (95% confidence interval = 3.2-8.1).
We now show that a common non-coding RET variant within a conserved enhancer-like sequence in intron 1 is significantly associated with HSCR susceptibility and makes a 20-fold greater contribution to risk than rare alleles do.
Inactivating mutations of the RET proto-oncogene and of one of its soluble ligand molecules, glial cell line derived neurotrophic factor (GDNF), have been found in a subset of patients with Hirschsprung disease (HSCR).
Both null and missense mutations impaired the interaction of TCF4 with ASCL1 from the PHOX-RET pathway in transactivating an E box-containing reporter construct; therefore, hyperventilation and Hirschsprung disease in patients with Pitt-Hopkins syndrome might be explained by altered development of noradrenergic derivatives.
The full application of the proposed protocol in most of the unexplained HSCR cases will allow us to establish the precise role of RET not only in causing but also in predisposing to HSCR pathogenesis.
Taken together, the low penetrance of the mutant gene, the lack of genotype-phenotype correlation, the sex-dependent effect of RET mutations and the variable clinical expression of the disease support the existence of one or more modifier genes in familial HSCR.
Although RET is a well-established risk factor, a recent genome-wide association study (GWAS) of HSCR has identified NRG1 as an additional susceptibility locus.
Germline and somatic mutations in RET that produce constitutively activated receptors cause the cancer syndrome multiple endocrine neoplasia type 2 and several endocrine and neural-crest-derived tumors, whereas mutations resulting in nonfunctional RET or lower expression of RET are found in individuals affected with Hirschsprung disease.
Germline RET mutations in Hirschsprung disease are mainly inactivating, and have been reported to account for up to 20 and 50% of sporadic and familial cases, respectively.
Such a functional effect of this common RET variant explains the under-representation of the whole haplotype and its role as a modifying factor in HSCR pathogenesis.
MEN2A and MEN2B have recently been mapped within the centromeric region closely linked to RBP3 and D10S15 (which are located inside the deletion) and cosegregate with HSCR in at least two different pedigrees.
The germline mutation was detected because of the systematic genetic screening of the RET proto-oncogene, which is useful for genetic counseling of potential risk of HSCR and MTC in other family members.
RET is a candidate gene for IHPS due to its role in the development of the intrinsic innervation and ganglia of the smooth musculature and the association of RET variants with another motility disorder (Hirschsprung's disease).
This distinctive feature of RET mutations associated with Hirschsprung's disease and of the RET mutations associated with thyroid cancer indicates a basic biological difference between the mutational events leading to the different phenotypes.
In addition, there was strong evidence against linkage to two Hirschsprung disease susceptibility genes (RET and EDNRB), a condition that can cosegregate with neuroblastoma as in one of the kindreds tested here.
Although the picture is complex, recent associations between specific RET proto-oncogene variations have been shown to be significant in Down syndrome patients with Hirschsprung disease, as they probably interfere with vital RET functions in the development of the autonomic and enteric nervous systems, increasing the risk of disturbed normal function.
We conclude that genomic rearrangements in RET are rare and were not responsible for the HSCR phenotype in individuals without identifiable germline RET variants in our group of patients, yet this possibility cannot be excluded altogether because the confidence to identify variation in at least two percent of the individuals was only 95%.