Lower doses of 6-mercaptopurine/azathioprine bring enough clinical efficacy and therapeutic concentration of erythrocyte 6-mercaptopurine metabolite in Japanese IBD patients

BackgroundAlthough 6-mercaptopurine (6-MP) and azathioprine (AZA) are prescribed at lower doses, their efficacy in patients with inflammatory bowel disease (IBD) in Japan is comparable to that in Europe/America. However, there has been no report concerning the measurement of erythrocyte 6-thioguanine nucleotides (6-TGN), which is an active metabolite of 6-MP or AZA, in Japanese IBD patients. This study was designed to elucidate the pharmacokinetic–pharmacodynamic properties of 6-MP and AZA in Japanese patients by measurement of erythrocyte 6-TGN level.Methods134 adult patients (99 males; 35 females) with IBD (75 ulcerative colitis; 59 Crohn's disease) who had been receiving a constant dose of 6-MP or AZA for three months or longer were enrolled. Erythrocyte 6-TGN levels were measured using the low-pressure gradient HPLC method, and correlated with treatment efficacy. The genetic polymorphism of thiopurine methyltransferase (TPMT) genotype was also assessed.ResultsThe mean erythrocyte 6-TGN level (mean ± SD) was 342.3 ± 220.9 pmol/8 × 10⁸RBC, which was supposed to be therapeutic concentration, although the mean daily doses of 6-MP and AZA were no more than 29.8 ± 9.9 mg/day of 6-MP equivalent. However, all patients were identified with the wild type of TPMT genotype. There was no significant difference in the mean 6-TGN levels between patients in remission and no-remission group. The mean 6-TGN level was significantly higher in the once-daily administration group than three times-daily group.ConclusionThirty mg/day of 6-MP or 50 mg/day of AZA, once-daily oral administration in Japanese IBD patients was sufficient to achieve the therapeutic target level of 6-TGN in Europeans/Americans.     

6-MP metabolite profiles provide a biochemical explanation for 6-MP resistance in patients with inflammatory bowel disease

BACKGROUND & AIMS: Approximately 40% of inflammatory bowel disease (IBD) patients fail to benefit from 6-mercaptopurine (6-MP)/azathioprine (AZA). Recent reports suggest 6-thioguanine nucleotide (6-TGN) levels (>235) independently correlate with remission. An inverse correlation between 6-TGN and thiopurine methyltransferase (TPMT) has been described. The objectives of this study were to determine whether dose escalation optimizes both 6-TGN levels and efficacy in patients failing therapy because of subtherapeutic 6-TGN levels and its effect on TPMT. METHODS: Therapeutic efficacy and adverse events were recorded at baseline and upon reevaluation after dose escalation in 51 IBD patients. 6-MP metabolite levels and TPMT activity were recorded blinded to clinical information. RESULTS: Fourteen of 51 failing 6-MP/AZA at baseline achieved remission upon dose escalation, which coincided with significant rises in 6-TGN levels. Despite increased 6-MP/AZA doses, 37 continued to fail therapy at follow-up. Dose escalation resulted in minor changes in 6-TGN, yet a significant increase in 6-methylmercaptopurine ribonucleotides (6-MMPR) (P < or = 0.01) and 6-MMPR:6-TGN ratio (P < 0.001). 6-MMPR rises were associated with dose-dependent hepatotoxicity in 12 patients (24%). TPMT was not influenced by dose escalation. CONCLUSIONS: Serial metabolite monitoring identifies a novel phenotype of IBD patients resistant to 6-MP/AZA therapy biochemically characterized by suboptimal 6-TGN and preferential 6-MMPR production upon dose escalation.     

Thiopurine S-methyltransferase and inosine triphosphate pyrophosphohydrolase genes in Japanese patients with inflammatory bowel disease in whom adverse drug reactions were induced by azathioprine/6-mercaptopurine treatment

Background  The main cause of azathioprine (AZA)/6-mercaptopurine (6MP)-induced adverse reactions is a reduction in the activities of the metabolizing enzymes thiopurine S-methyltransferase (TPMT) and inosine triphosphate pyrophosphohydrolase (ITPA). Adverse reactions develop at a high frequency in Japanese patients at half the dose required for European and American patients; however, the association with TPMT and ITPA gene polymorphisms in Japanese has not been fully investigated. Methods  Gene mutations of TPMT and ITPA, the major AZA/6-MP -metabolizing enzymes, were investigated retrospectively in 16 Japanese patients with inflammatory bowel disease (IBD) in whom AZA/6MP treatment induced adverse reactions. Results  The TPMT gene was found to have a wild-type sequence in all patients, but in the ITPA gene a mutation, 94C>A, was detected at a rate of 50% (8/16), with 83.3% (5/6) occurring in patients with acute bone marrow suppression and 75% (3/4) in those with agranulocytosis. The 94C>A allele frequency was 10 of 32 (0.313; 95% CI, 0.180–0.486). Adverse reactions developed earlier in patients with the 94C>A mutation. However, in half the patients, no gene polymorphism was noted. Conclusions  It is suggested that the ITPA gene mutation is closely related to the adverse reactions of AZA/6-MP in Japanese patients, and screening for the mutant allele is useful for predicting the most serious adverse reactions, agranulocytosis and acute bone marrow suppression.     

5-氨基水杨酸对硫嘌呤类药物骨髓抑制的影响

目的 研究5-氨基水杨酸(5-ASA)对硫嘌呤类药物骨髓抑制的影响及其机制,探讨中国炎症性肠病(IBD)患者合用5-ASA时所需硫嘌呤类药物的剂量。方法 回顾性分析服用硫嘌呤类药物IBD患者的临床资料,检测硫嘌呤甲基转移酶(TPMT)活性和红细胞6-硫鸟嘌呤核苷酸(6-TGN)浓度。前瞻性研究中先予患者硫唑嘌呤(AZA) 50 mg/d治疗4周,继而加用5-ASA 3 g/d治疗4周,检测第4、8周末红细胞6-TGN浓度。结果 回顾性分析AZA/6-巯嘌呤(6-MP) +5-ASA组45例、AZA/6-MP组94例患者,两组骨髓抑制发生率分别为46.7％和16.0％,多因素回归分析显示合用5-ASA为增加骨髓抑制的惟一独立危险因素(OR= 3.45,95％CI：1.31～9.04)。TPMT活性在AZA/6-MP+ 5-ASA组和AZA/6-MP组之间差异无统计学意义(t=-0.351,P=0.734)。AZA/6-MP+5-ASA组6-TGN浓度显著高于AZA/6-MP组（中位浓度为384.9 pmol/8×108 RBC比286.4 pmol/8×108 RBC,F=29.15,P=0.00）。8例患者完成前瞻性研究,予AZA 50mg/d 4周后,7例患者6-TGN浓度＜230 pmol/8×108 RBC;加用5-ASA 4周后,7例患者6-TGN浓度≥230 pmol/8×108 RBC,其中3例6-TGN浓度≥420 pmol/8×108 RBC,2例发生骨髓抑制。结论 当中国IBD患者合用5-ASA治疗时,采用常规推荐剂量的AZA/6-MP时骨髓抑制的发生概率增加,其机制可能与红细胞内6-TGN浓度升高有关,降低AZA剂量有可能在保持疗效的同时降低骨髓抑制发生率。     

Steady-state of azathioprine during initiation treatment of pediatric inflammatory bowel disease

Background and AimAzathioprine (AZA) has a slow onset of action in treatment of pediatric inflammatory bowel disease (IBD). It is anticipated, that this delay correlates to the kinetics of 6-thioguanine nucletiodes (6-TGN) accumulation. The aim of this study was to evaluate the time to steady state of 6-TGN concentration in red blood cells.MethodsThe inclusion criteria were: a) age 0-19 years b) IBD diagnosis c) AZA treatment initiation. High performance liquid chromatography was used for the 6-TGN analysis. Concentrations of metabolites were studied in weeks 0, 1, 2, 5, and 8 after beginning of treatment.ResultsThe inclusion criteria were matched to 18 patients with IBD. The median time to steady state of 6-TGN was 55.3 days. The mean 6-TGN concentration at the steady state achieved 326 (SD 154) pmol/8.108 erythrocytes. High erythrocyte TPMT activity corresponds to the low steady state 6-TGN concentration and vice versa. This correlation reached statistical significance (p < 0.01) for the dose expressed in mg per square meter of body surface area.ConclusionThe time to steady state of 6-TGN erythrocyte concentration is significantly shorter than would expected according to clinical observation describe earlier.     

硫嘌呤甲基转移酶基因型和酶活性检测在炎症性肠病治疗中的临床价值

目的 评价硫嘌呤甲基转移酶(TPMT)基因型和酶活性检测对炎症性肠病(IBD)患者服用硫唑嘌呤(AZA)发生不良反应的预测价值.方法 收集2004年4月-2009年12月有使用AZA指征的IBD确诊患者112例,其中溃疡性结肠炎(UC)26例,克罗恩病(CD)86例.患者每天均服用AZA 2 mg/kg.采用PCR技术检测患者TPMT基因型(*2、*3A、*3B*、3C),高效液相色谱法检测TPMT酶活性.分析达到随访终点(服药达半年或以上,或因不良反应停药)患者的TPMT基因多态性及酶活性与不良反应发生之间的关系及其影响因素.结果 112例患者中不良反应率为33.9%(38/112),以骨髓抑制最常见(20.5%).TPMT*3C杂合子突变率为0.9%(1/112).TPMT酶活性呈单峰正态分布,平均活性为(12.9士4.2)U/ml红细胞.1例TPMT*3C杂合子突变者于用药4周内发生骨髓抑制,TPMT基因型预测骨髓抑制发生的敏感度为4.4 0A(1/23),特异度为1/1.ROC曲线计算TPMT酶活性预测服药3个月内发生骨髓抑制的安全阈为≤4.5 U/ml红细胞,该安全阈预测3个月内骨髓抑制发生的敏感度为3/13,特异度为3/3.合用5-氨基水杨酸(5-ASA)制剂者骨髓抑制的发生率显著高于非合用者(44.4%比12.9%,P=0.000),但发生骨髓抑制者合用5-ASA前后TPMT酶活性差异无统计学意义(P>0.05).结论 TPMT基因突变和酶活性低下对预测发生骨髓抑制的特异性较高,但敏感性较差,合用5-ASA可增加AZA发生骨髓抑制的风险,但与TPMT酶活性无关.     

Thiopurine methyltransferase genotype and the toxicity of azathioprine in Japanese

OBJECTIVE: Thiopurine S-methyltransferase (TPMT) is a cytosolic enzyme that preferentially catalyzes the S-methylation of aromatic and heterocyclic sulfhydryl compounds, including Azathioprine (AZA). It has been reported that the level of AZA toxicity is dependent on the TPMT genotypes in Caucasian individuals; we thus investigated this relationship in Japanese. METHODS: The TPMT genotype was determined using peripheral blood cell DNA obtained from 36 Japanese patients with rheumatic diseases who were treated with AZA, by polymerase chain reaction (PCR) technique. Duration of AZA administration, white blood cell counts before and after AZA administration, and side effects were investigated in each subject, and were compared between the patients with or without TPMT mutation. RESULTS: The TPMT allelotype was TPMT*1/TPMT*1 in 33 (91.7%) and TPMT*1/TPMT*3C in 3 (8.3%) individuals. All 3 patients (100%) with the mutant TPMT allele (TPMT*3C) discontinued AZA treatment due to leucopenia while only 4 patients (12%) without mutant TPMT alleles showed leucopenia (p=0.0049, Fisher's exact test). However, leucopenia developed relatively late in patients with mutant TPMT. CONCLUSION: The TPMT mutant allele, TPMT*3C, also exists in Japanese individuals, and the bone marrow toxicity of AZA is likely stronger in patients with this mutant allele.     

Optimizing immunomodulator therapy for inflammatory bowel disease

6-Mercaptopurine (6-MP) and its prodrug azathioprine (AZA) remain the mainstay of immunomodulator therapy for the maintenance of steroid-free remission in patients with inflammatory bowel disease (IBD). Traditional dosing strategies for initiation of thiopurines are often based on weight or empirically chosen. Dosing based on an understanding of an inherited difference in drug disposition and metabolism may provide a safer alternative. The thiopurine methyltransferase (TPMT) enzyme plays a pivotal role in the metabolism of 6-MP and AZA and is critical to the determination of thiopurine toxicity. The therapeutic benefits of thiopurines correlate best with concentration of the active 6-thioguanine (6-TGN) metabolites. Reports suggest that therapeutic response can be maximized when patients achieve therapeutic 6-TGN levels. Pharmacogenetic dosing based on TPMT and pharmacokinetic dosing based on 6-TGN levels may offer a safety and efficacy advantage over traditional dosing strategies and provide a novel mechanism for optimizing immunomodulator therapy in IBD.     

Relationship between 6-mercaptopurine dose and 6-thioguanine nucleotide levels in patients with inflammatory bowel disease

BACKGROUND: 6-Thioguanine nucleotides (6-TGN) are active metabolites of azathioprine (AZA) and 6-mercaptopurine (6MP). Higher remission rates have been observed in patients with higher 6-TGN levels. However, many physicians prescribe AZA/6MP using milligrams per kilogram (mg/kg) dosing regimens without measuring 6-TGN levels. The aim of this study was to examine the association between 6MP dose and 6-TGN levels. METHODS: We conducted a cross-sectional study of patients treated for inflammatory bowel disease (IBD) with AZA or 6MP, whose 6-TGN levels were measured. Patients with low or intermediate thiopurine methyl transferase (TPMT) activity were excluded. AZA dose was converted to 6MP equivalents. The relationship between dose and 6-TGN levels was assessed with the Spearman correlation coefficient. We used logistic regression to assess the relationship between dose and 6-TGN levels of >230 pmol/8 x 10(8). RESULTS: In this study, 155 patients met our inclusion criteria (median dose, 1.01 +/- 0.40; range, 0.61-1.41 mg/kg). There was a weak correlation between 6-TGN levels and the absolute dose (rho = 0.18, P = 0.04) and the dose in mg/kg (rho = 0.19, P = 0.03). The correlation between mg/kg dosage and 6-TGN levels was slightly stronger in those using concomitant 5-aminosalicylate (5-ASA) medications (rho = 0.24, P = 0.02). Compared with <1.0 mg/kg per day, doses of > or =1.5 mg/kg per day were strongly associated with 6-TGN levels of >230 pmol/8 x 10(8) RBC (adjusted odds ratio [OR] = 3.7, 95% confidence interval [CI] = 1.1-11.8). However, only 37% of patients receiving > or =1.0 mg/kg per day had 6-TGN levels of >230 pmol/8 x 10(8) RBC. CONCLUSIONS: 6MP dose is weakly associated with 6-TGN levels. The use of standard mg/kg dosing regimens will result in low 6-TGN levels in most patients.     

Thiopurine S-methyltransferase polymorphisms and the relationship between the mutant alleles and the adverse effects in systemic lupus erythematosus patients taking azathioprine

OBJECTIVE: The present study sought to elucidate the genetic basis of thiopurine methyltransferase (TPMT) polymorphism and subsequently to investigate the relationship between mutant TPMT and an adverse response observed in Korean patients with systemic lupus erythematosus (SLE) taking azathioprine (AZA). METHODS: The TPMT genotype of 342 patients with SLE was determined by MALDI-TOF mass spectrometry and correlated with the effects of clinical exposure to AZA. RESULTS: TPMT polymorphism was detected in 17 of the 342 study subjects (5.0%), 12 heterozygous for the TPMT*3C allele and 5 heterozygous for the TPMT*6 allele. Numerous patients taking AZA demonstrated adverse drug responses. Severe nausea occurred in 4 patients with the TPMT*3C allele, while 1 patient with the TPMT*6 allele suffered severe bone marrow toxicity. Leucopenia (n = 17), nausea (n = 4), and abnormal liver function (n = 1) were suspected in 23 of the 94 lupus patients taking AZA. AZA was relatively well tolerated by the remainder of the patients. The heterozygous genotype for the TPMT*3C and *6 alleles was frequently detected in Korean SLE patients. CONCLUSION: Contrary to previous hypotheses, this study identified no statistical correlation between TPMT genotype and AZA toxicity. We thus conclude that TMPT genotyping cannot replace regular blood monitoring in SLE patients receiving AZA treatment.     

High prevalence of polymorphism and low activity of thiopurine methyltransferase in patients with inflammatory bowel disease

BackgroundGene polymorphism of thiopurine methyltransferase (TPMT) correlates with decreased enzyme activity which determines a significant risk of adverse effect reactions (ADR) in patients treated with thiopurines. The aim of this study was to investigate TPMT genotype and phenotype status in patients with inflammatory bowel diseases (IBD).MethodsFifty-one consecutive out-patients with IBD were genotyped for the following allelic variants: rs1800462 (referred as TPMT*2 allele), rs1800460 (referred as TPMT *3B allele), and 1142345 (referred as TPMT *3C allele). Red blood cell TPMT activity was measured using a competitive micro-well immunoassay for the semi-quantitative determination of TPMT activity in red blood cells (RBC) by means of a 6-MP substrate.ResultsPolymorphism of TPMT was found in 5 out of 51 patients (10%; 95% CI 2%–18%), three heterozygous and two homozygous carriers. Six patients (11.8%; 95% CI 2.4%–19.5%) displayed very low, 12 (23.5%; 95% CI 11.4%–34.5%) intermediate, and 33 (64.7%; 95% CI 52%–78%) normal/high TPMT activity. There were no differences between TPMT genotype and phenotype groups according to age, type of disease, smoking, and chronic medications. A 71% (95% CI 61%–81%; κ = 0.45) concordance rate was found between genotype and phenotype status. Six out of 27 (22%) current or past users of azathioprine developed ADR, with three (50%) displaying TPMT genotype and/or phenotype alterations.ConclusionCompared to the general population, IBD patients may have significantly higher prevalence of TPMT polymorphism and, even more, low activity. Phenotypic more than genotypic TPMT analysis could be useful to better manage IBD therapy with thiopurines.     

炎症性肠病患者巯基嘌啉甲基转移酶基因型检测与硫唑嘌啉安全性分析

目的初步探讨炎症性肠病(IBD)患者巯基嘌啉甲基转移酶(TPMT)基因突变情况,了解硫唑嘌啉的不良反应。方法对30例我院住院IBD患者TPMT*3C基因型通过基因PCR扩增、基因测序的方法进行检测,其中10例患者给予硫唑嘌啉1～2 mg/kg.d-1剂量口服。结果 30例患者TPMT*3C基因型均为野生型,未发现基因突变,2例发生不良反应。结论 TPMT*3C在IBD中突变率较低;应用硫唑嘌啉需严密观察其不良反应。     

Thiopurine methyltransferase enzyme activity determination before treatment of inflammatory bowel disease with azathioprine: effect on cost and adverse events.

BACKGROUND: Azathioprine (AZA), used to treat inflammatory bowel disease (IBD), is metabolized by thiopurine methyltransferase (TPMT). The accumulation of individual metabolites varies because humans display genetic polymorphism for TPMT expression. Deficiencies in TPMT result in accumulation of toxic metabolites, followed by neutropenia and hepatic inflammation. Concern over acute toxicity frequently leads to under dosing and frequent monitoring tests and visits. OBJECTIVE: To determine whether assessment of TPMT activity before the administration of AZA would predict acute toxicity and, thus, allow for reductions in health care costs related to biochemical screening for, and management of, AZA-induced adverse events. METHODS: Before AZA treatment, 29 patients with IBD were prospectively randomized to one of two groups: group 1, in which no TPMT assay was performed, was started on AZA at 1 mg/kg/day and then titrated every two weeks to a target dose of 2.5 mg/kg/day; and group 2, in which TPMT assays were performed, was started on AZA at the target dose of 2.5 mg/kg/day. For both groups, complete blood count and liver enzymes were monitored weekly for six weeks and at monthly intervals thereafter. Additional tests and health care interventions were undertaken at the discretion of the attending physicians. RESULTS: Of the 29 patients in the study, 15 were randomly assigned to group 1 and 14 to group 2. Demographics and disease activity were similar for both groups. Mean follow-up time was 7.1 months (range 3.5 to 10.7 months). Eight patients from group 1 and six patients from group 2 withdrew as a result of AZA-induced adverse events. There was no correlation between the TPMT activity and the development of AZA-induced adverse events. The direct health care costs for group 1 (300.11 dollars per patient) were lower than in group 2 (348.87 dollars per patient). CONCLUSION: The prospective assessment of TPMT enzyme activity before initiating AZA therapy in IBD patients incurred additional cost and did not predict AZA-induced toxicity.     

HRM技术检测炎症性肠病患者TPMT的基因多态性

目的应用HRM技术检测炎症性肠病(IBD)患者TPMT基因多态性,进而探索IBD患者TPMT基因型与硫唑嘌呤(AZA)致骨髓抑制的关系。方法采用聚合酶链反应-高分辨熔解曲线(PCR-HRM)与Sanger法序列测定相结合的方法,对82例IBD患者和53名健康志愿者TPMT基因第7、10外显子进行检测。结果 IBD患者中TPMT*1/*3C杂合子4例,健康对照者中TPMT*1/*3C杂合子2例。未检测出TPMT*3A和TPMT*3B型突变。IBD患者4例出现骨髓抑制的患者中,1例是TPMT*1/*3C杂合子;其余3例为TPMT野生型。结论 TPMT*3C(A719G)基因突变在中国的IBD患者中较TPMT*3A(G460A/A719G)、TPMT*3B(G460A)发生率高,而存在此基因突变的IBD患者对硫唑嘌呤不能耐受,导致骨髓抑制。然而TPMT基因突变只能解释部分AZA治疗IBD导致骨髓抑制的病例。     

Genotyping should be considered the primary choice for pre-treatment evaluation of thiopurine methyltransferase function

Background and aimsA pre-treatment determination of the thiopurine S-methyltransferase (TPMT) genotype or phenotype can identify patients at risk of developing severe adverse reactions from thiopurine treatment. The risk of misclassifying a patient might be dependent on the method used. The aim of this study was to investigate the concordance between TPMT genotyping and phenotyping.MethodsThe data consist of 7195 unselected and consecutive TPMT genotype and phenotype determinations sent to the division of Clinical Pharmacology, Linköping, Sweden. TPMT activity was measured in red blood cells (RBC) and the genotype determined by pyrosequencing for the three most common TPMT variants (TPMT *2, *3A, *3C).ResultsTPMT genotyping identified 89% as TPMT wild type (*1/*1), 10% as TPMT heterozygous and 0.5% as TMPT defective. The overall concordance between genotyping and phenotyping was 95%, while it was 96% among IBD patients (n = 4024). Genotyping would have misclassified 8% of the TPMT defectives as heterozygous as compared to 11% if only TPMT activity had been measured. 11% of the heterozygous patients had a normal TPMT activity (> 8.9 U/ml RBC) and 3% of the TPMT wild-type patients had an intermediate TPMT activity (2.5–8.9 U/ml RBC).ConclusionsThere is a risk for TPMT misclassification when only genotyping or phenotyping is used, but it is not reasonable to check both in all patients. Since TPMT genotyping is the more reliable test, especially in TPMT heterozygotes, we suggest that genotyping should be considered the primary choice for the pre-treatment evaluation of TPMT function before initiation of thiopurine therapy.     

Comparison of TPMT and NUDT15 polymorphisms in Chinese patients with inflammatory bowel disease

AIM To observe gene polymorphisms of TPMT and NUDT15, and compare their predictive value for azathioprine(AZA)-induced leukopenia in inflammatory bowel disease(IBD).METHODS This study enrolled 219 patients diagnosed with IBD in Xiangya Hospital, Central South University, Changsha, China from February 2016 to November 2017. Peripheral blood of all patients was collected to detect their genotypes of TPMT and NUDT15 by pyrosequencing at the Department of Clinical Pharmacology, Hunan Key Laboratory of Pharmacogenetics, Xiangya Hospital. Eighty patients were treated with AZA according to the disease condition. During the first month, patients who received AZA underwent routine blood tests and liver function tests once a week. The endpoint of the study was leukopenia induced by AZA. By analyzing patient characteristics, genotypes and leukopenia induced by drug use, we found the risk factors associated with AZA-induced leukopenia.RESULTS There were 219 patients with IBD(160 men and 59 women), including 39 who were confirmed with ulcerative colitis(UC), 176 with Crohn's disease(CD) and 4 with undetermined IBD(UIBD). There were 44 patients(20.1%) with mutant genotype of NUDT15(C/T); among them, 16 received AZA, and 8(50%) developed leukopenia. There were 175 patients(79.7%) with wild genotype of NUDT15(C/C); among them, 64 received AZA, and 11(17.2%) developed leukopenia. A significant difference was found between NUDT15 C/T and its wild-type C/C(P = 0.004). There were only 3 patients with TPMT mutant genotype of A/G(1.4%) who participated in the research, and 1 of them was treated with AZA and developed leukopenia. The remaining 216 patients(98.6%) were found to bear the wild genotype of TPMT(A/A); among them, 79 patients received AZA, and 18(22.8%) developed leukopenia, and there was no significant difference from those with A/G(P = 0.071). The frequency of TPMT mutation was 1.4%, and NUDT15 mutation rate was significantly higher and reached 20.1%(P = 0.000). Therefore, NUDT15 gene polymorphism was obviously a better biomarker than TPMT gene polymorphism in the prediction of AZA-induced leukopenia.CONCLUSION Mutation rate of NUDT15 in Chinese IBD patients is higher than that of TPMT. NUDT15 polymorphism is a better predictor for AZA-induced leukopenia than TPMT polymorphism.     

巯嘌呤甲基转移酶检测在炎症性肠病治疗中的作用

巯嘌呤甲基转移酶（TPMT）是硫唑嘌呤（AZA）／6-巯基嘌呤（6-MP）代谢的关键酶之一,而AZA／6-MP广泛用于难治性炎症性肠病（IBD）的治疗。TPMT的活性／基因型与AZA／6-MP的疗效和不良反应相关,进行TPMT活性／基因型检测有可能预测骨髓抑制等不良反应的发生,并能指导个体化治疗。     

Azathioprine-induced severe pancytopenia due to a homozygous two-point mutation of the thiopurine methyltransferase gene in a patient with juvenile HLA-B27-associated spondylarthritis

Severe pancytopenia due to azathioprine (AZA) toxicity in patients with autoimmune diseases is not uncommon. We describe a 14-year-old girl with HLA-B27+ spondylarthritis who was treated with AZA 3 mg/kg/day and who suddenly developed severe pancytopenia in the seventh week of treatment. Analysis of the catabolic pathway of AZA revealed a homozygous deficiency of thiopurine methyltransferase (TPMT) on the basis of a combined 2-point mutation at nucleotide positions 460 and 719 in the gene for TPMT, causing a toxic level of the metabolic active 6-thioguanine nucleotides (6-TGN) (2,394 pmoles/8 × 10⁸ red blood cells). The patient was transfusion dependent and finally recovered 8 weeks after the development of the pancytopenia. At that time, 6-TGN had already returned to normal therapeutic levels. Family studies revealed another homozygous deficiency in the mother, while the other family members were heterozygous.     

Azathioprine-induced fatal myelosuppression in systemic lupus erythematosus patient carrying TPMT*3C polymorphism

Azathioprine (AZA) is a commonly used immunosuppressant for systemic lupus erythematosus (SLE). Myelosuppression is a serious adverse reaction due to AZA and its metabolites. Thiopurine S-methyltransferase (TPMT) is the rate-limiting enzyme. Variations of TPMT enzyme activity may be responsible for myelosuppression. However, a correlation between certain mutant alleles of low TPMT enzyme activity and myelotoxicity has also been suggested as a factor. We describe herein a case of AZA-induced severe myelosuppression associated with TPMT*3C heterozygous mutant allele in a SLE patient. The patient presented with pancytopenia, sepsis, typhlitis and disseminated intravascular coagulopathy after a short period of AZA therapy. The patient had low TPMT activity and TPMT*3C genotype. Measurement of TPMT activity and determination of TPMT variant allele may identify patients at risk for AZA-induced myelosuppression.