肾移植受者TPMT和ITPA基因多态性与硫唑嘌呤所致不良反应关系的研究

目的探讨硫嘌呤甲基转移酶（TPMT）和三磷酸肌苷焦磷酸酶（ITPA）基因多态性与硫唑嘌呤（AZA）所致各种不良反应的关系。方法应用高效液相色谱法（HPLC）Ng定155例肾移植受者红细胞TPMT和ITPA活性，采用等位基因特异性的PCR（ASPCR）和限制性片断长度多态性（PCR-RFLP）的方法检测TPMT＊2、＊3A、＊3B和＊3C四种基因型，采用实时荧光定量PCR和PCR-RFLP的方法分别检测ITPA 94C〉A和IVS2＋21A〉C两种基因型，分析TPMT和ITPA活性和基因多态性与AZA所致不良反应的关系。结果155例肾移植受者中，120例患者未发生不良反应，将其作为对照组，平均TPMT活性为（37．26±11．60）U，平均ITPA活性为（97．9±34．7）U。另外35例患者由于发生了不良反应而停用AZA或减少了AZA的剂量。其中12例患者产生了血液毒性，平均TPMT活性为（22．92±10．67）U，明显低于对照组（P〈0．05），其中仅2例患者为TPMT＊3C突变基因杂合子，AZA所致的血液毒性主要与患者红细胞TPMT活性低下有关。18例患者产生了肝脏毒性，其TPMT和ITPA活性离散度较大，其均值分别与对照组比较均无统计学差异（P〉0．05）。其中仅发现2例TPMT＊3C和4例ITPA94C〉A突变基因杂合子，TPMT和ITPA活性和基因多态性与AZA所致的肝脏毒性无明显相关性。5例患者产生了胃肠紊乱症状（包括1例患者同时产生了肝脏毒性），其中有2例患者ITPA活性缺乏，为ITPA94C〉A突变基因纯合子，另外3例患者ITPA活性较低，为ITPA94C〉A突变基因杂合子。剩余1例患者出现了流感样症状，ITPA活性缺乏，为ITPA94C〉A突变基因纯合子。ITPA活性缺乏或ITPA94C〉A基因突变纯合子患者极易发生AZA所致胃肠紊乱症状和流感样症状的不良反应。结论一在服用AZA之前，对患者TPMT活性进行测定，有利于避免AZA所致血液毒性的发生。而对患者进行ITPA活性和基因多态性检测，有利于避免AZA所致的胃肠紊乱和流感样症状的产生，使AZA的使用更加安全、合理。

Associations of Thiopurine S-methyltransferase and Inosine Triphosphate Pyrohoshatase Genetic Polymorphisms with Azathioprine-related Adverse Drug Reactions in Renal Transplant Recipients

Objective To systematically investigate the relationships between thiopurine S-methyltransferase （TPMT） and inosine triphosphate pyrophosphatase （ITPA） polymorphisms and azathioprine-related adverse drug reactions in patients with kidney transplantation. Methods Erythrocyte TPMT and ITPA activity of 155 patients with kidney transplantation under AZA therapy was determined by high performance liquid chromatography （HPLC）. The frequency of 4 common TPMT mutant alleles, TPMT ＊ 2, ＊ 3A, ＊ 3B and ＊ 3C, was determined by allele-specific PCR and PCR- restriction fragment length polymorphism （PCR-PFLP） analysis. Meanwhile, the frequency of ITPA 94C〉A and IVS2 ＋ 21A〉C was determined by real-time fluorescence PCR and PCR-PFLP. Results Among 155 patients in this study, 120 did not experience any adverse reactions under standard dosage of AZA and they were classified as control group. The mean TPMT activity of control group was （37. 26 ± 11.60） U and the mean ITPA activity was （97. 9 ± 34. 7） U. Thirty- five patients stopped azathioprine medication or were on reduced dose due to azathioprine-related side effects, of whom twelve developed hematotoxicity with mean TPMT activity of （22. 92 ± 10. 67） U, mueh lower than control group （P〈0. 05）. Only two patients with TPMT＊3C heterozygous alleles were found in these twelve patients. AZA induced hematotoxieity was related to the lower TPMT activity. Eighteen patients developed hepatotoxicity and there was no statistical difference between the mean TPMT or ITPA activity and the control mean （P〉0. 05）. Only two cases with TPMT ＊ 3C and four cases with ITPA 94C〉A heterozygous alleles were found. No significant asso- ciations between TPMT or ITPA genetic polymorphisms and AZA-related hepatotoxicity could be detected. In five patients with gastrointestinal disturbance （including one patient who developed hepatotoxicity simultaneously）, two patients with deficient ITPA activity were homozygote for 94C〉A and the other three patients with low ITPA activity had 94C〉 A heterozygous alleles. The patient who experienced flu-like symptoms showed deficient ITPA activity and he was the remaining homozygote for 94C〉A. Patients with ITPA 94C〉A homozygous alleles are at high risk to develop AZA-related gastrointestinal disturbance and flu-like symptoms. Conclusion Before commencing AZA treatment, it is important to measure erythrocyte TPMT activity in renal transplant recipients in order to prevent AZA-related hematotoxicity. Pretherapeutic screening of patients for erythrocyte ITPA activity and genetic polymorphisms should be useful to avoid AZA- related gastrointestinal disturbance and flu-like symptoms for safer and more tolerable AZA treatment.