Analysis Of Polymorphisms Of TPMT Gene And Its Relevance To Drug Adverse Reactions Of6-MP In Child Patients With Acute Lymphoblastic Leukemia

Objective: Thiopurine S-methyltransferase (TPMT) plays one of the important roles in thecatabolic metabolism of thiopurine drugs, and there may be correlations betweencoding-region single nucleotide polymorphisms (cSNPs) of TPMT gene and adversereactions of thiopurine drugs. This study was aimed to detect mutations existed in codingsequence region (CDS) of TPMT gene, based on which to investigate the distributions ofgenotypes and alleles frequencies in Han nationality children with acute leukemia (AL),and to look forward to finding new mutations. Moreover, a retrospective study wasperformed by analysing each different clinical data of child patient with acutelymphoblastic leukemia (ALL) during maintenance treatment period, and to assess theassociations between the polymorphisms of TPMT gene and these6-mercaptopurine(6-MP) related toxicities. Providing the basis for clinical developments of individualmedicine programs.Methods: Total RNA was extracted from bone marrow samples of103child patients withAL and111peripheral blood samples of the control group after informing to theirguardians, cDNA was got by reverse transcriptase-ploymerase chain reaction (RT-PCR).Gene mutations of single DNA base of all samples were detected by denaturing gradientgel electrophoresis (DGGE) combining DNA sequencing technology. Drug toxicities ofchemotherapy group were classified according to national cancer institute-commontoxicity criteria, version3.0(NCI CTC v3.0), meanwhile, the relationships between thepolymorphisms of TPMT gene and the adverse reactions of6-MP were analyzed. Theincidences of adverse reactions of6-MP in TPMT*3C GG or TPMT*1S CC/TT wereanalysed by Fisher probabilities in2*2table. Sex, age, the frequencies of genotypes andalleles, and the incidence of adverse reactions of genotypes in two groups were made bychi-square test.Results:1. There were no statistical difference between patients and controls in age and sex(P>0.05). Only two known mutations,210C>T (70Cys>Cys，rs1862148744) and TPMT*26(622T>C，208Phe>Leu，rs72556347) were screened in the CDS of TPMT gene,the former was synonymous mutation and the latter was missense mutation. Likewise, onlytwo cSNPs, TPMT*3C (474T>C,158Ile>Ile, rs2842934) and*1S (719A>G,240Try>Cys,rs1142345) were found.2. The genotypes and alleles frequencies of TPMT*1S were adherence toHardy-Weinberg equilibrium in controls. The C allele frequency of TPMT*1S in this studyof the Han nationality was20.3%. There were no significant differences in genotype andallele frequencies of TPMT*1S between patients and cotrols, ALL and AML, respectively(P>0.05).3. The genotypes and alleles frequencies of TPMT*3C were adherence toHardy-Weinberg equilibrium in controls. The G allele frequency of TPMT*3C in this studyof the Han nationality was2.1%. There were no significant differences in genotype andallele frequencies of TPMT*3C between patients and cotrols, ALL and AML, respectively(P>0.05).4. The cumulative time of each child in the hemotherapy group was742.2d (163~985d),10cases (21.7%) were discontinued because of serious adverse reactions of6-MP,and the total interrupt day was7.9d (2~22d), the drug withdrawal was5.3d (2~14d)at every turn.5. Neutropenia (93.5%)，lower hemoglobin (69.9%)，thrombocytopenia (50.0%) andhigher aminotransferase (82.6%and80.4%) were the most common adverse reaction of6-MP. The serious adverse reaction was appeared frequently in neutropenia (60.9%),however, others were mild in contrast.6. To compare with standard-risk, there were no significant differences in high-riskand medium-risk of TPMT*1S in hemotherapy group. There were no significantdifferences between TPMT*1S genotypes and severe myelosuppression or hepatotoxicitycaused by6-MP (P>0.05). However, Both of two children with TPMT*3C genotypes hadserious adverse reactions. Among them, one with homozygote showed6-MP dose-relatedintolerance.Conclusion:1. The genotypes of TPMT*26、C210T were screened in the Han nationality children.2. There may be no correlations between the risk of childhood AL and TPMT*1S or*3C that was found in the Han nationality children. 3. Neutropenia was the most common reverse adverse reaction of6-MP duringmaintenance treatment period of child patients with ALL.4. There was no relationship between TPMT*1S genotypes and a variety of adversereactions.5. TPMT*3C may correlate with sever adverse reactions caused by6-MP.