Abstract：OBJECTIVE: Thymidylate synthase (TS) catalyses the conversion of deoxy-uridylate to deoxy-thymidylate and is a key enzyme for DNA synthesis. TS is the target enzyme of 5-fluorouracil (5-FU) and involved in folate metabolism. TS gene polymorphisms play an important role in the efficiency of fluorouracil activity in vivo. This study investigated the allelic frequencies and distribution characters of single-nucleotide polymorphisms within the coding region (cSNPs) of TS gene in Chinese children with acute leukemia (AL) and normal control children in order to explore the possible relationship between the cSNP in human TS gene and chemotherapeutic effects of 5-fluorouracils. METHODS: Bone marrow samples from 53 children with AL and peripheral blood samples from 115 normal children were obtained to prepare complementary DNAs (cDNAs). The cDNAs were analyzed for the polymorphisms in TS gene by reverse transcriptase (RT)-polymerase chain reaction (PCR)-denaturing gradient gel electrophoresis (DGGE) and direct sequencing. The distributive difference of each genotype between AL children and control children was evaluated. RESULTS: A polymorphism 381 A＞G (E127E) in the coding region of TS gene was firstly identified in the Chinese population. The 381 A＞G allelic frequency in AL children and control children was 12.3% and 13.5% respectively (P>0.05), which were similar to that in the International SNP Bank (12.3%). The allelic frequency of cSNPs was not associated with the susceptibility to AL. CONCLUSIONS: A polymorphism 381 A＞G (E127E) in TS gene was successfully identified in children using RT-PCR-DGGE combined with DNA sequencing. There was no significant difference in the allelic frequency of cSNPs in AL children and normal children.［Chin J Contemp Pediatr, 2009, 11 (4):251-254］
CHEN Xiao-Wen,YUE Li-Jie,LI Chang-Gang et al. Polymorphisms of thymidylate synthase gene detected by RT-PCR-denaturing gradient gel electrophoresis in children with acute leukemia[J]. CJCP, 2009, 11(04): 251-254.
［2］Pullarkat ST, Stoehlmacher J, Ghaderi V, Xiong YP, Ingles SA, Sherrod A, et al. Thymidylate synthase gene polymorphism determines response and toxicity of 5-FU chemotherapy［J］. Pharmacogenomics J, 2001, 1 (1): 65-70.
［3］Nief N, Le Morvan V, Robert J. Involvement of gene polymorphisms of thymidylate synthase in gene expression, protein activity and anticancer drug cytotoxicity using the NCI-60 panel［J］. Eur J Cancer, 2007, 43 (5): 955-962.
［4］Kuramochi H, Tanaka K, Oh D, Lehman BJ, Dunst CM, Yang DY, et al. Thymidylate synthase polymorphisms and mRNA expression are independent chemotherapy predictive markers in esophageal adenocarcinoma patients［J］. Int J Oncol, 2008, 32(1): 201-208.
［5］Matsui T, Omura K, Kawakami K, Morita S, Sakamoto J. Genotype of thymidylate synthase likely to affect efficacy of adjuvant 5-FU based chemotherapy in colon cancer［J］. Oncol Rep, 2006, 16(5):1111-1115.
［6］Marsh S. Thymidylate synthase pharmacogenetics［J］. Invest New Drugs, 2005, 23(6):533-537.
［7］Krajinovic M, Costea I, Primeau M, Dulucq S, Moghrabi A. Combining several polymorphisms of thymidylate synthase gene for pharmacogenetic analysis［J］. Pharmacogenomics J, 2005, 5(6): 374-380.
［8］Danenberg PV. Pharmacogenomics of thymidylate synthase in cancer treatment［J］. Front Biosci, 2004, 9:2484-2494.
［14］van der Hout AH, van den Ouweland AM, van der Luijt RB, Gille HJ, Bodmer D, Brüggenwirth H, et al. A DGGE System for comprehentive Mutation Screening of BRCA1 and BRCA2: Application in a Dutch Cancer Clinic Setting［J］. Hum Mutat, 2006, 27 (7): 654-666.
［15］Rendtorff ND, Bjerregaard B, Frodin M, Kjaergaard S, Hove H, Skovby F, et al. Analysis of 65 tuberous sclerosis complex (TSC) patients by TSC2 DGGE, TSC1/TSC2 MLPA, and TSC1 long-range PCR sequencing, and report of 28 novel mutations［J］. Hum Mutat, 2005, 26(4): 374-383.
［16］Lacerra G, Fiorito M, Musollino G, Di Noce F, Esposito M, Nigro V, et al. Sequence variations of the alpha-globin genes: scanning of high CG content genes with DHPLC and DG-DGGE［J］. Hum Mutat, 2004, 24(4): 338-349.
