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Copy Number Variations in Candidate Genomic Regions Confirm Genetic Heterogeneity and Parental Bias in Hirschsprung Disease


Copy Number Variations in Candidate Genomic Regions Confirm Genetic Heterogeneity and Parental Bias in Hirschsprung Disease

Francesca Lantieri et al. Orphanet J Rare Dis.


Background: Hirschsprung Disease (HSCR) is a congenital defect of the intestinal innervations characterized by complex inheritance. Many susceptibility genes including RET, the major HSCR gene, and several linked regions and associated loci have been shown to contribute to disease pathogenesis. Nonetheless, a proportion of patients still remains unexplained. Copy Number Variations (CNVs) have already been involved in HSCR, and for this reason we performed Comparative Genomic Hybridization (CGH), using a custom array with high density probes.

Results: A total of 20 HSCR candidate regions/genes was tested in 55 sporadic patients and four patients with already known chromosomal aberrations. Among 83 calls, 12 variants were experimentally validated, three of which involving the HSCR crucial genes SEMA3A/3D, NRG1, and PHOX2B. Conversely RET involvement in HSCR does not seem to rely on the presence of CNVs while, interestingly, several gains and losses did co-occur with another RET defect, thus confirming that more than one predisposing event is necessary for HSCR to develop. New loci were also shown to be involved, such as ALDH1A2, already found to play a major role in the enteric nervous system. Finally, all the inherited CNVs were of maternal origin.

Conclusions: Our results confirm a wide genetic heterogeneity in HSCR occurrence and support a role of candidate genes in expression regulation and cell signaling, thus contributing to depict further the molecular complexity of the genomic regions involved in the Enteric Nervous System development. The observed maternal transmission bias for HSCR associated CNVs supports the hypothesis that in females these variants might be more tolerated, requiring additional alterations to develop HSCR disease.

Keywords: Candidate genes and regions; Comparative genomic hybridization; Copy number variations; Custom array; Hirschsprung disease.

Conflict of interest statement

The authors declare that they have no competing interests.


Fig. 1
Fig. 1
Diagram of the study design. Flowchart of the analysis performed on the complete panel of 59 patients, including the Agilent informatic method (on the left) and the visual inspection (on the right), that have led to the detection of 83 CNVs, together with the tables and files generated at each step. In particular, excluding the six already known control CNVs, the remaining 77 are further distinguished based on the DGV database (25 CNVs with a frequency higher than 5%, and thus considered true, and 52 CNVs novel or very rare on DGV) and on a visual classification. Fifteen likely true CNVs and three CNVs located on known HSCR genes have been validated by a different approach, confirming a total of 12 novel “true” CNVs in addition to the 25 already described on DGV. Numbers shown on the top of the diagram (above the red line) refer to samples, while those shown below the red line refer to CNVs (not coincident with the number of samples carrying the CNVs)
Fig. 2
Fig. 2
Profiles for some validated CNVs. Copy Number Variation (CNVs) detected at 9q31 (a), 15q21 (b), 16q23 (c) and PHOX2B (d) are shown. On the left of each panel there is the chromosomal view, on the middle the detailed region view with genes reported, and on the right the possible presence of CNVs

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