Preliminary study shows novel variant detected in the screening of RET gene in Malaysian patients with Hirschsprung’s disease

Nor Azian Abdul Murad, Sue-Mian Then, Mohd Ridhwan Abd Razak, Conjeevaram Rajendrarao Thambidorai, Sri Noraima Othman, Rosniza Mohamad Hussain, Mohan Nallusamy, Syed Zulkifli Syed Zakaria, Isa Mohammad Rose, Zarina Latiff, Rahman Jamal


Hirschsprung’s disease (HSCR) is a disorder associated with congenital absence of ganglion cells in the gastrointestinal tract.  Molecular analyses have identified variants in various genes including RET, GDNF, EDN3 and EDNRB that are involved in the development, migration and survival of neural cells. Variants in the receptor tyrosine kinase (RET) are most common and have been identified in 10-20% of sporadic HSCR patients. The objective of this study was to screen for RET gene variants in Malaysian patients with HSCR. Thirty-two patients with HSCR and 30 normal controls were recruited for this study. Mutations were screened using the Polymerase Chain Reaction – Denaturing High Performance Liquid Chromatography (PCR-dHPLC) approach. Mutations identified were then confirmed using Sanger sequencing. We identified one novel rare variant in exon 4 (A268A c807 G>C) in one patient. We also identified the common coding sequence variantsA45A (c135G>A), A432A (c1296A>G), L769L (c2307 T>G) and the G691S in our cohort of patients.  In conclusion, our Malaysian patients with HSCR diseases showed the presence of similar RET gene common variants which have been described in other populations. We have also identified a novel variant in exon 4 (A268A).

Full Text:



Whitehouse F, Kernohan J. (1948) Myenteric plexuses in congenial megacolon; study of 11 cases. Arch Int Med82:75-111.

Kenny SE, Tam PK, Garcia-Barcelo M. (2010) Hirschsprung's disease. Semin Pediatr Surg 19(3):194-200.

Badner JA, Sieber WK, Garver KL, et al (1990) A genetic study of Hirschsprung disease. Am J Hum Genet 46(3):568-580.

Emison ES, Garcia-Barcelo M, Grice EA, et al (2010) Differential contributions of rare and common, coding and noncoding Ret mutations to multifactorial Hirschsprung disease liability. Am J Hum Genet 9;87(1):60-74.

Amiel J, Sproat-Emison E, Garcia-Barcelo M (2008) Hirschsprung disease, associated syndromes and genetics: a review. J Med Genet 45:1-14.

Angrist M, Kauffman E, Slaugenhaupt SA, (1993) A gene for Hirschsprung disease (megacolon) in the pericentromeric region of human chromosome 10. Nat Genet 4(4):351-356.

Lyonnet S, Bolino A, Pelet A, et al (1993) A gene for Hirschsprung disease maps to the proximal long arm of chromosome 10. Nat Genet4(4):346-350.

Wu TT, Tsai TW, Chang H, et al (2010) Polymorphisms of the RET gene in Hirschsprung disease, anorectal malformation and intestinal pseudo-obstruction in Taiwan. J Formos Med Assoc 109(1):32-38.

Nunez-Torres R, Fernandez RM, Acosta MJ, et al (2011) Comprehensive analysis of RET common and rare variants in a series of Spanish Hirschsprung patients confirms a synergistic effect of both kinds of events. BMC Med Genet 12:138-144.

Ruiz-Ferrer M, Torroglosa A, Luzon-Toro B, et al (2011) Novel mutations at RET ligand genes preventing receptor activation are associated to Hirschsprung's disease. J Mol Med (Berl) 89(5):471-480.

So MT, Leon TY, Cheng G, et al (2011) RET mutational spectrum in Hirschsprung disease: evaluation of 601 Chinese patients. PLoS One 6(12):e28986.

Cecherini I, Bocciardi R, Luo Y, et al (1993) Exon structure and flanking intronic sequences of the human RET proto-oncogene. Biochem Biophys Res Commun 196:1288-1295.

Burzynski GM, Nolte IM, Bronda A, Bos KK (2005) Identifying candidate Hirschsprung disease-associated RET variants. Am J Hum Genet 76(5):850-858.

Chin TW, Chiu CY, Tsai HL, et al (2008) Analysis of the RET gene in subjects with sporadic Hirschsprung's disease. J Chin Med Assoc 71(8):406-410.

Kim JH, Yoon KO, Kim JK, et al (2006) Novel mutations of RET gene in Korean patients with sporadic Hirschsprung's disease. J Pediatr Surg 41(7):1250-154.

Leon TY, So MT, Lui VC, et al (2012) Functional analyses of RET mutations in Chinese Hirschsprung disease patients. Birth Defects Res A Clin Mol Teratol 94(1):47-51.;jsessionid=4DEDBB3A355C6E83A3294CD507304852.f04t04

Ngo DN, So MT, Gui H, et al (2012) Screening of the RET gene of Vietnamese Hirschsprung patients identifies 2 novel missense mutations. J Pediatr Surg 47(10):1859-1864.

Wu T-T, Tsai T-W, Chu C-T et al (2005) Low RET mutation frequency and polymorphism analysis of the RET and EDNRB genes in patients with Hirschsprung disease in Taiwan. J Hum Genet 50:168-174.

Cornes BK, Tang CS, Leon TY, et al (2010) Haplotype analysis reveals a possible founder effect of RET mutation R114H for Hirschsprung's disease in the Chinese population. PLoS One 5(6):e10918.

Liu CP, Tang QQ, Lou JT et al (2010) Association analysis of the RET proto-oncogene with Hirschsprung disease in the Han Chinese population of southeastern China. Biochem Genet 48(5-6):496-503.

Moore SW, Zaahl MG (2008) A review of genetic mutation in familial Hirschsprung's disease in South Africa: towards genetic counseling. J Pediatr Surg 43(2):325-329.

Phusantisampan T, Sangkhathat S, Phongdara A, et al (2012) Association of genetic polymorphisms in the RET-protooncogene and NRG1 with Hirschsprung disease in Thai patients. J Hum Genet 57(5):286-293.

Miller SA, Dykes DD, Polesky HF (1988) A simple salting out procedure for extracting DNA from human nucleated cells. Nucleic Acid Res 16:1251.

Ceccherini I, Bocciardi R, Luo Y, et al (1993) Exon structure and flanking intronic sequences of the human RET proto-oncogene. Biochem Biophys Res Commun 196:1288-95.

Ceccherini I, Hofstra RMW, Luo Y, et al (1994) DNA polymorphism and conditions for SSCP analysis of the 20 exons of the ret proto-oncogene. Oncogene 9:3025-3029.

Sakai T, Nirasawa Y, Itoh Y, et al (2000) Japanese patients with sporadic Hirschsprung: mutation analysis of the receptor tyrosine kinase proto-oncogene, endothelin-B receptor, endothelin-3, glial cell line-derived neurotrophic factor and neurturin genes: a comparison with similar studies. Eur J Pediatr 159:160-167.

Romero P, Niesler B, Schmitz-Winnenthal H, et al (2012) Is there a link between the calcium sensing receptor and Hirschsprung's disease? A mutational analysis. J Pediatr Surg 47(3):551-555.

Tou J, Wang L, Liu L, Wang Y, et al (2011) Genetic variants in RET and risk of Hirschsprung's disease in Southeastern Chinese: a haplotype-based analysis. BMC Med Genet 12:32-37.

Gray VE, Kukurba KR, Kumar S (2012) Performance of computational tools in evaluating the functional impact of laboratory-induced amino acid mutations. Bioinformatics 28(16):2093-2096.

Borrello MG, Aiello A, Peissel B, et al (2011) Functional characterization of the MTC-associated germline RET-K666E mutation: evidence of oncogenic potential enhanced by the G691S polymorphism. Endocr Relat Cancer 18(4):519-527.

Fitze G, Cramer J, Serra A, et al (2003) Within-gene interaction between c.135 G/A genotypes and RET proto-oncogene germline mutations in HSCR families. Eur J Pediatr Surg 13(3):152-157.

Gath R, Goessling A, Keller KM, et al (2001) Analysis of the RET, GDNF, EDN3, and EDNRB genes in patients with intestinal neuronal dysplasia and Hirschsprung disease. Gut 48(5):671-675.

Luzon-Toro B, Torroglosa A, Nunez-Torres R, et al (2012) Comprehensive analysis of NRG1 common and rare variants in Hirschsprung patients. PLoS One 7(5):e36524.

Wallace AS, Anderson RB (2011) Genetic interactions and modifier genes in Hirschsprung's disease. World J Gastroenterol 17(45):4937-4944.


  • There are currently no refbacks.
Asia-Pacific Journal of Molecular Medicine (APJMM),
C/o: Level 8, UKM Medical Molecular Biology Institute (UMBI), UKM Medical Centre,
Jalan Ya’acob Latiff, Bandar Tun Razak,
56000 Cheras, Kuala Lumpur, MALAYSIA
Tel: +6 03 9145 6321
Fax: +6 03 9171 7185