mercaptopurine and 6-thioguanylic-acid

Thiopurines in combination with glucocorticoids are used as first-line, second-line and maintenance therapies in autoimmune hepatitis and opportunities exist to improve and expand their use.. To describe the metabolic pathways and key factors implicated in the efficacy and toxicity of the thiopurine drugs and to indicate the opportunities to improve outcomes by monitoring and manipulating metabolic pathways, individualising dosage and strengthening the response.. English abstracts were identified in PubMed by multiple search terms. Full-length articles were selected for review, and secondary and tertiary bibliographies were developed.. Thiopurine methyltransferase activity and 6-tioguanine (6-thioguanine) nucleotide levels influence drug efficacy and safety, and they can be manipulated to improve treatment response and prevent myelosuppression. Methylated thiopurine metabolites are associated with hepatotoxicity, drug intolerance and nonresponse and their production can be reduced or bypassed. Universal pre-treatment assessment of thiopurine methyltransferase activity and individualisation of dosage to manipulate metabolite thresholds could improve outcomes. Early detection of thiopurine resistance by metabolite testing, accurate estimations of drug onset and strength by surrogate markers and adjunctive use of allopurinol could improve the management of refractory disease. Dose-restricted tioguanine (thioguanine) could expand treatment options by reducing methylated metabolites, increasing the bioavailability of 6-tioguanine nucleotides and ameliorating thiopurine intolerance or resistance.. The efficacy and safety of thiopurines in autoimmune hepatitis can be improved by investigational efforts that establish monitoring strategies that allow individualisation of dosage and prediction of outcome, increase bioavailability of the active metabolites and demonstrate superiority to alternative agents.

Topics: Allopurinol; Azathioprine; Drug-Related Side Effects and Adverse Reactions; Guanine Nucleotides; Hepatitis, Autoimmune; Humans; Inflammatory Bowel Diseases; Mercaptopurine; Purines; Quality Improvement; Thioguanine; Thionucleotides; Treatment Outcome

Thiopurine drugs, including azathioprine and 6-mercaptopurine, are used commonly in patients with inflammatory bowel disease for maintenance of remission. Although generally well tolerated, adverse effects lead to discontinuation in a significant minority of patients. Pharmacogenomic studies have suggested that metabolic breakdown of azathioprine in an individual is genetically determined. Coupled with the fact that certain thiopurine metabolites, notably 6-thioguanine nucleotide and 6-methylmercaptopurine, are associated with antiinflammatory effects and adverse effects, respectively, some investigators have examined intentionally shunting the metabolism of azathioprine toward increasing 6-thioguanine nucleotide levels by using low doses of the xanthine oxidoreductase inhibitor allopurinol to improve efficacy and decrease toxicity of azathioprine in patients with inflammatory bowel disease. We performed a search of the MEDLINE and Embase databases for basic and clinical research reports of this modality. Pertinent articles were retrieved, reviewed, and assessed by the authors. Case series, cohort studies, and one randomized trial have investigated adding allopurinol to azathioprine therapy in patients with inflammatory bowel disease. Most reports primarily examined metabolite levels in these patients. In general, the literature suggests that this modality was successful at significantly increasing 6-thioguanine nucleotide levels while decreasing 6-methylmercaptopurine levels. Several small reports have suggested that patients with increased 6-thioguanine nucleotide levels had improved symptoms or symptom remission. Adverse effects and discontinuation rates remained similar or were improved in patients who were taking a thiopurine and started allopurinol. In conclusion, the addition of allopurinol may be an option for optimizing thiopurine metabolite production in select patients with low 6-thioguanine nucleotide levels. Appropriate care and monitoring of these patients are mandatory to prevent neutropenia or other adverse effects.

Topics: Allopurinol; Drug Therapy, Combination; Enzyme Inhibitors; Guanine Nucleotides; Humans; Immunosuppressive Agents; Inflammatory Bowel Diseases; Mercaptopurine; Methyltransferases; Prospective Studies; Retrospective Studies; Thionucleotides

There is a growing interest in the use of thiopurines (azathioprine, 6-mercaptopurine and 6-thioguanine) for the management of inflammatory bowel disease. The genetically controlled thiopurine (S)-methyltransferase enzyme is involved in the metabolism of these agents and is hypothesised to determine the clinical response to thiopurines. Diminished activity of this enzyme decreases the methylation of thiopurines, theoretically resulting in potential overdosing, while high thiopurine (S)-methyltransferase status leads to overproduction of toxic metabolites and ineffectiveness of azathioprine and 6-mercaptopurine. In practice, controversies exist regarding the utility of standard thiopurine (S)-methyltransferase pheno- and genotyping. Current pharmacogenetic insights suggest that another enzyme system may participate in the efficacy and toxicity of thiopurines; inosine triphosphate pyrophosphatase. Other topics discussed in this review are the utilisation of therapeutic drug monitoring and the experimental use of 6-thioguanine in the treatment of inflammatory bowel disease. 6-Thioguanine has a less genetically controlled metabolism and skips genetically determined metabolic steps. On theoretical basis, 6-thioguanine might therefore have a more predictable profile than azathioprine and 6-mercaptopurine. However, the use of 6-thioguanine has been associated with an increased risk of nodular regenerative hyperplasia of the liver and veno-occlusive disease. Further research is warranted before 6-thioguanine can be considered as a treatment option for inflammatory bowel disease.

Topics: Antimetabolites; Azathioprine; Gene Frequency; Genotype; Guanine Nucleotides; Humans; Inflammatory Bowel Diseases; Inosine Triphosphatase; Mercaptopurine; Methyltransferases; Phenotype; Practice Guidelines as Topic; Pyrophosphatases; Racial Groups; Thionucleotides

Ulcerative colitis (UC) has been known as inflammatory bowel disease. Progress in UC management strategies has led to optimized approaches for achieving the two primary clinical goals of therapy: induction and maintenance of remission. We here review about immunosuppressants in management of UC; Cyclosporine A (CsA) has been used for the induction therapy in steroid resistant refractory UC. Although it has been reported that CsA has high response rate in severe UC, long-term efficacy (maintenance of remission) has not been proven. To improve maintenance therapy, immunosuppressant has been re-considered in management of UC. In recent years, it has been reported that efficacy of 6-mercaptopurine/azathioprine in maintenance of remission of UC is superior to 5-aminosalicylate (5-ASA). Pharmacological studies have indicated thiopurine methyltransferase (TPMT) activity is essential for maintenance of blood concentration of 6-thioguanine nucleotide (6-TG).

Topics: Azathioprine; Colitis, Ulcerative; Cyclosporine; Drug Therapy, Combination; Guanine Nucleotides; Humans; Immunosuppressive Agents; Mercaptopurine; Methyltransferases; Mycophenolic Acid; Tacrolimus; Thionucleotides

The thiopurine drugs azathioprine and 6-mercaptopurine (6-MP) are well-established in the treatment of inflammatory bowel disease (IBD). However, there is a wide inter- and intra-patient variation in the concentrations of active and toxic metabolites due to their complex metabolism and genetic polymorphisms in metabolizing enzymes. Serious drug toxicity leads to cessation of therapy in 9-25% of patients, and there is failure to achieve efficacy in approximately 15% of cases. Advances in the understanding of thiopurine drug metabolism have led to new genetic and metabolite tests to help clinicians optimize thiopurine use. Thiopurine methyltransferase (TPMT) enzyme activity can predict life-threatening myelotoxicity in the one in 300 patients who are TPMT-deficient. However, myelotoxicity can also occur in the presence of normal TPMT activity so blood count monitoring should remain standard practice. TPMT testing may also aid in dose individualization. 6-Thioguanine nucleotides (6-TGN) are thought to be the predominant active metabolites of the thiopurines. 6-thioguanine nucleotide concentration is correlated with bone marrow toxicity and may also correlate with efficacy in IBD. Measurement of 6-TGN and 6-methylmercaptopurine (6-MMP) concentration is most useful in determining why a patient is not responding to a standard dose of a thiopurine drug and may help in avoiding myelosuppression. The ratio of these metabolites can help distinguish non-compliance, under-dosing, thiopurine-resistant and thiopurine-refractory disease. Some of these investigations are entering routine clinical practice but more research is required to determine their optimal use in patients with IBD.

Topics: Antimetabolites, Antineoplastic; Azathioprine; Bone Marrow; Erythrocytes; Guanine Nucleotides; Humans; Inflammatory Bowel Diseases; Inosine Triphosphatase; Mercaptopurine; Methyltransferases; Polymorphism, Genetic; Pyrophosphatases; Thionucleotides

Topics: Bone Marrow Diseases; Child; Drug Resistance; Female; Guanine Nucleotides; Humans; Leukemia, Lymphoid; Male; Mercaptopurine; Methyltransferases; Thionucleotides

Patients' beliefs about medicine may either reflect the necessity for treatment or concerns regarding the treatment. We explored the extent to which these beliefs have an effect on thiopurine metabolite levels and premature discontinuation in patients with inflammatory bowel disease (IBD).. Patients enrolled in the 'Thiopurine response Optimization by Pharmacogenetic testing in Inflammatory Bowel Disease Clinics' (TOPIC) trial were asked to complete the Beliefs about Medicine Questionnaire (BMQ) 4 weeks after thiopurine initiation. The BMQ measures perceptions about treatment necessity and concerns. On the basis of the necessity and concern scores, patients can be categorized as accepting, ambivalent, indifferent, or skeptical. The thiopurine discontinuation rates for these belief subgroups were compared by Kaplan-Meier curves. Furthermore, clinical response and metabolite levels were compared between the belief subgroups.. A total of 767 patients with IBD started thiopurine treatment, of whom 576 (75%) completed the BMQ. Patients could be classified as accepting (34%), indifferent (17%), ambivalent (34%), or skeptical (15%). Compared with patients in the accepting group (discontinuation rate 22%), patients with an indifferent (35%; P=0.02), ambivalent (37%; P<0.01), or skeptical belief (54%; P<0.01) had higher thiopurine discontinuation rates. No differences were observed in the steady-state thiopurine metabolite levels between the different belief subgroups.. Patients with a low perceived treatment necessity or high concerns toward IBD treatment were more likely to discontinue thiopurine treatment prematurely. Extra attention toward these patients might prevent premature discontinuation.

Topics: Adult; Female; Guanine Nucleotides; Health Knowledge, Attitudes, Practice; Humans; Immunosuppressive Agents; Inflammatory Bowel Diseases; Male; Medication Adherence; Mercaptopurine; Middle Aged; Surveys and Questionnaires; Thioinosine; Thionucleotides; Treatment Outcome; Young Adult

Significance of monitoring adalimumab trough levels and anti-adalimumab antibodies (AAA) for disease outcome in Crohn's disease (CD) patients remained unclear.. To evaluate the association of adalimumab trough levels and AAA at week 26 with clinical remission at week 52, the effect of azathiopurine on AAA and factors influencing trough levels in CD patients in the DIAMOND trial.. We performed this study using adalimumab trough levels, AAA at week 26 and 6-thioguanine nucleotide (TGN) in red blood cells at week 12. A multiple regression model and receiver operating analysis was performed to identify factors influencing adalimumab trough levels and AAA, and adalimumab thresholds for predicting disease activity.. There was a significant difference of adalimumab trough level at week 26 between patients with disease remission and without at week 52 (7.7 ± 3.3 μg/mL vs 5.4 ± 4.3 μg/mL: P <.001). Adalimumab trough level of 5.0 μg/mL yielded optimal sensitivity and specificity for remission prediction (80.2% and 55.6%, respectively). AAA development at week 26 significantly affected remission at week 52 (P = .021), which was strongly associated with adalimumab trough levels. Female gender and increasing body weight were independently associated with low adalimumab trough levels, and female gender was associated with AAA development. A cut-off 6TGN level of >222.5 p mol/8 ×10. Adalimumab trough levels and AAA occurrence were significantly associated with clinical remission. Higher 6TGN affected AAA negativity. The combination therapy is beneficial in some relevant aspects for CD patients. (UMIN Registration No. 000005146).

Topics: Adalimumab; Anti-Inflammatory Agents; Antibodies; Crohn Disease; Drug Therapy, Combination; Female; Guanine Nucleotides; Humans; Male; Mercaptopurine; Sensitivity and Specificity; Thionucleotides; Treatment Outcome

Several studies have demonstrated a favourable safety profile for thiopurine use for inflammatory bowel disease (IBD) during pregnancy. We performed a study in pregnant patients with IBD who were using thiopurines, in order to determine the influence of pregnancy on thiopurine metabolism and to assess intrauterine exposure of the fetus to thiopurines.. Female patients with IBD receiving steady-state thiopurines and planning a pregnancy were prospectively enrolled. 6-Thioguanine nucleotide (6-TGN) and 6-methylmercaptopurine (6-MMP) concentrations were determined, combined with routine laboratory tests, before, during and after pregnancy. Thiopurine metabolites were measured in umbilical cord blood immediately after delivery.. Thirty patients who were using azathioprine (28 patients, median dose 1.93 mg/kg) or mercaptopurine (two patients, doses 1.32 and 0.94 mg/kg) were included. During pregnancy, median 6-TGN decreased over time (p=0.001). while 6-MMP increased, without causing myelotoxicity or hepatotoxicity. After delivery, both 6-TGN and 6-MMP levels returned to preconception baseline levels. Fetal 6-TGN concentrations correlated positively with maternal 6-TGN levels (p<0.0001). No 6-MMP was detected in the newborns, except one born with pancytopenia and high alkaline phosphatase activity; the mother of this infant had severe pre-eclampsia. All infants had normal Apgar scores, but 60% had anaemia at birth. No major congenital abnormalities were observed.. Pregnancy has a major effect on maternal thiopurine metabolism. In utero the unborn child is exposed to 6-TGN, but not to 6-MMP. Sixty per cent of the infants were born with anaemia, which raises the question whether infants should be tested for possible anaemia immediately after birth.

Topics: Abnormalities, Drug-Induced; Adolescent; Adult; Anemia, Neonatal; Azathioprine; Biomarkers; Female; Fetal Blood; Follow-Up Studies; Guanine Nucleotides; Humans; Immunosuppressive Agents; Infant, Newborn; Inflammatory Bowel Diseases; Mercaptopurine; Middle Aged; Pregnancy; Pregnancy Complications; Pregnancy Outcome; Prospective Studies; Thionucleotides; Treatment Outcome; Young Adult

Pediatric patients with acute lymphoblastic leukemia (ALL) are treated with oral 6-mercaptopurine (6MP) for nearly 2 years, but no pediatric formulation has been available. In this study, an oral 6MP liquid suitable for pediatric use was developed and tested in the target population.. A randomized cross-over study was performed in 20 pediatric ALL patients (age 1.9 - 14.6 years), comparing pharmacokinetics and pharmacodynamics of a newly developed 6MP liquid formulation to 6MP capsules, both taken orally for 4 weeks.. Based upon trough levels of the principal active metabolite,6-thioguanine nucleotides (6-TGN),a relative bioavailability of the liquid vs. capsules of 1.01 was found (90% CI 0.86 - 1.20), demonstrating bioequivalence. This was supported by the similarly observed 6MP dosages needed for leucocyte depletion, for both formulations (35 mg/day (range 10 - 115 mg)). 75% of the parents/patients (p = 0.005) preferred the oral liquid over the capsules because of the ease of administration.. We conclude that the novel 6MP liquid is a promising treatment for ALL.

Topics: Administration, Oral; Adolescent; Antimetabolites, Antineoplastic; Biological Availability; Child; Child, Preschool; Cross-Over Studies; Female; Guanine Nucleotides; Humans; Infant; Leukocytes; Male; Mercaptopurine; Precursor Cell Lymphoblastic Leukemia-Lymphoma; Therapeutic Equivalency; Thionucleotides; Young Adult

Topics: Antineoplastic Combined Chemotherapy Protocols; Child; DNA; Female; Genotype; Guanine Nucleotides; Humans; Male; Mercaptopurine; Methotrexate; Methyltransferases; Precursor Cell Lymphoblastic Leukemia-Lymphoma; Thionucleotides

Recent studies in patients with inflammatory bowel diseases (IBD) on thiopurine therapy suggest that too low 6-thioguanine nucleotide concentrations (6-TGN) and too high methylmercaptopurine nucleotide concentrations (MMPN) can be reversed by a combination therapy of allopurinol and low-dose thiopurines. To date, however, optimal dosing has not been established. The aim of this study was to evaluate the minimal allopurinol doses necessary to achieve adequate 6-TGN concentrations in combination with low-dose azathioprine.. A stepwise dose-escalation of allopurinol was performed in 11 azathioprine-pretreated IBD patients with inadequately low 6-TGN concentrations (<235 pmol/8 × 10(8) erythrocytes) and/or elevated MMPN concentrations (>5,000 pmol/8 × 10(8) erythrocytes) and/or elevated liver enzymes (alanine aminotransferase and/or aspartate aminotransferase levels one- to threefold the upper limit of normal). Six patients were recruited into an open study, and five were treated in the context of an individualized therapeutic approach. Adverse effects, azathioprine metabolites, liver enzymes and whole blood counts were monitored two to three times per month.. Adequate 6-TGN concentrations were achieved with a combination of 25 mg allopurinol and 50 mg azathioprine in one patient and with 50 mg allopurinol and 50 mg azathioprine in nine patients. Median 6-TGN concentrations (range) were 336 (290-488) pmol/8 × 10(8) erythrocytes after an 8-week-long intake of the final dose combination. One patient dropped out due to nausea after the first intake. MMPN concentrations and liver enzymes normalized immediately in all affected patients. All patients finishing the dose-escalation regimen tolerated the treatment without toxicity.. Combination therapy with only 50 mg allopurinol and 50 mg azathioprine daily is sufficient, efficacious and safe in most IBD patients with inadequate thiopurine metabolite concentrations to optimize azathioprine-based IBD therapy.

Topics: Adult; Aged; Allopurinol; Azathioprine; Female; Guanine Nucleotides; Humans; Inflammatory Bowel Diseases; Male; Mercaptopurine; Middle Aged; Thionucleotides

In children with acute lymphoblastic leukaemia (ALL) bone marrow activity can influence red blood cell (RBC) kinetics, the surrogate tissue for thiopurine methyltransferase (TPMT) measurements. The aim of this study was to investigate TPMT phenotype-genotype concordance in ALL, and the influence of TPMT on thiopurine metabolite formation.. We measured TPMT (activity, as units ml(-1) packed RBCs and genotype) at diagnosis (n = 1150) and TPMT and thioguanine nucleotide (TGN) and methylmercaptopurine nucleotide (MeMPN) metabolites (pmol/8 × 10(8) RBCs) during chemotherapy (n = 1131) in children randomized to thioguanine or mercaptopurine on the United Kingdom trial ALL97.. Median TPMT activity at diagnosis (8.5 units) was significantly lower than during chemotherapy (13.8 units, median difference 5.1 units, 95% confidence interval (CI) 4.8, 5.4, P < 0.0001). At diagnosis genotype-phenotype was discordant. During chemotherapy the overall concordance was 92%, but this fell to 55% in the intermediate activity cohort (45% had wild-type genotypes). For both thiopurines TGN concentrations differed by TPMT status. For mercaptopurine, median TGNs were higher in TPMT heterozygous genotype (754 pmol) than wild-type (360 pmol) patients (median difference 406 pmol, 95% CI 332, 478, P < 0.0001), whilst median MeMPNs, products of the TPMT reaction, were higher in wild-type (10 650 pmol) than heterozygous patients (3868 pmol) (P < 0.0001). In TPMT intermediate activity patients with a wild-type genotype, TGN (median 366 pmol) and MeMPN (median 8590 pmol) concentrations were similar to those in wild-type, high activity patients.. In childhood ALL, TPMT activity should not be used to predict heterozygosity particularly in blood samples obtained at disease diagnosis. Genotype is a better predictor of TGN accumulation during chemotherapy.

Topics: Adolescent; Antimetabolites, Antineoplastic; Child; Child, Preschool; Female; Genotype; Guanine Nucleotides; Heterozygote; Humans; Infant; Male; Mercaptopurine; Methyltransferases; Phenotype; Precursor Cell Lymphoblastic Leukemia-Lymphoma; Thioguanine; Thionucleotides; United Kingdom

Therapeutic drug monitoring of active metabolites of thiopurines, azathioprine and 6-mercaptopurine, is relatively new. The proposed therapeutic threshold level of the active 6-thioguanine nucleotides (6-TGN) is ≥235 pmol/8×10(8) erythrocytes. The aim of this prospective cross-sectional study was to compare 6-TGN levels in adult thiopurine tolerant IBD patients with an exacerbation with those in remission, and to determine the therapeutic 6-TGN cut-off level.. Hundred IBD patients were included. Outcome measures were thiopurine metabolite levels, calculated therapeutic 6-TGN cut-off level, CDAI/CAI scores, thiopurine dose and TPMT enzyme activity.. Forty-one patients had an exacerbation, 59 patients were in remission. In 17% of all patients 6-TGN levels were compatible with non-compliance. The median 6-TGN levels were not significantly different between the exacerbation and remission group (227 versus 263 pmol/8×10(8) erythrocytes, p=0.29). The previous reported therapeutic 6-TGN cut-off level of 235 pmol/8×10(8) erythrocytes was confirmed in this study. Twenty-six of the 41 patients (63%) with active disease had 6-TGN levels below this threshold and 24 of 59 IBD patients (41%) in clinical remission (p=0.04).. Thiopurine non-compliance occurs frequently both in active and quiescent disease. 6-TGN levels below or above the therapeutic threshold are associated with a significant higher chance of IBD exacerbation and remission, respectively. These data support the role of therapeutic drug monitoring in thiopurine maintenance therapy in IBD to reveal non-compliance or underdosing, and can be used as a practical tool to optimize thiopurine therapy, especially in case of thiopurine non-response.

Topics: Adolescent; Adult; Aged; Azathioprine; Biomarkers, Pharmacological; Chromatography, High Pressure Liquid; Cross-Sectional Studies; Disease Progression; Dose-Response Relationship, Drug; Drug Monitoring; Female; Genetic Markers; Guanine Nucleotides; Humans; Immunosuppressive Agents; Inflammatory Bowel Diseases; Maintenance Chemotherapy; Male; Medication Adherence; Mercaptopurine; Methyltransferases; Middle Aged; Prospective Studies; Severity of Illness Index; Thioinosine; Thionucleotides; Young Adult

6-Mercaptopurine and its prodrug azathioprine are effective for the treatment of inflammatory bowel disease. Thiopurine methyltransferase is important for the metabolism of thiopurines. However, there is controversy as to the clinical utility of measuring thiopurine methyltransferase enzyme activity and 6-thioguanine nucleotide levels. Our aim was to determine if thiopurine methyltransferase enzyme activity and 6-thioguanine nucleotide level monitoring would predict response to therapy with thiopurines in patients with inflammatory bowel disease.. Baseline thiopurine methyltransferase enzyme activity prior to initiation of therapy with either 6-mercaptopurine or azathioprine was determined in 39 patients with inflammatory bowel disease. The association between clinical response and thiopurine methyltransferase activity and 6-thioguanine nucleotide levels singly or in combination were analysed.. Seventeen of 39 patients (44%) responded to 6-mercaptopurine or azathioprine therapy. Thiopurine methyltransferase enzyme activity below the mean of 30.5 U was significantly associated with clinical response. The thiopurine methyltransferase low phenotype was associated with response in 65% vs. 29% in individuals with thiopurine methyltransferase enzyme activity above 30.5 U (p = 0.05). There was no correlation between thiopurine methyltransferase activity and 6-thioguanine nucleotide levels. The maximal 6-thioguanine nucleotide levels did not predict clinical response. When combining thiopurine methyltransferase enzyme activity and 6-thioguanine nucleotide levels, the combination of thiopurine methyltransferase low/6-thioguanine nucleotide high was associated with response in 7/7 (100%) vs. only 2/8 (25%) with the combination of thiopurine methyltransferase high/6-thioguanine nucleotide low (p=0.01).. Thiopurine methyltransferase activity inversely correlated with clinical response to thiopurine treatment in inflammatory bowel disease. Thiopurine methyltransferase enzyme activity below 30.5 U combined with a post-treatment 6-thioguanine nucleotide level > 230 pmol/8 x 10(8) erythrocytes was the best predictor of response.

Topics: Adult; Antimetabolites, Antineoplastic; Azathioprine; Female; Forecasting; Guanine Nucleotides; Humans; Inflammatory Bowel Diseases; Male; Mercaptopurine; Methyltransferases; Thionucleotides; Treatment Outcome

Thiopurine methyltransferase (TPMT) activity determines biotransformation of azathioprine and, thereby, drug efficacy and safety. Evaluation of a possible long-term effect of mesalazine or azathioprine on TPMT activity is of particular clinical importance because both drugs can to be given for several years in inflammatory bowel disease. Monitoring of TPMT activity and three thiopurine metabolites was performed prospectively during a 1 year postoperative period in 21 patients with Crohn's disease randomly assigned to azathioprine (2.0-2.5 mg/kg per day) or mesalazine (4 g/day). TPMT activity did not change significantly within each treatment group during 52 weeks. At any study visit, TPMT activity was not different between 13 patients on azathioprine and eight patients on mesalazine. Concentrations of 6-thioguanine nucleotides (6-TGN, active moiety of azathioprine) and 6-methyl-mercaptopurine ribonucleotides (6-MMPR) did not alter significantly during the observation period, except for a slight decrease in 6-TGN levels when comparing the first with the last visit. In this first report of serial monitoring of 6-methyl-thioguanine nucleotides (6-MTGN) in patients with inflammatory bowel disease taking azathioprine, high levels of 6-TGN were correlated with high levels of 6-MTGN, with the mean 6-TGN:6-MTGN ratio being 2.4. In a well-standardized clinical setting of inflammatory bowel disease, neither mesalazine nor azathioprine significantly affected TPMT activity during a whole year of treatment.

Topics: Administration, Oral; Adult; Anti-Inflammatory Agents, Non-Steroidal; Azathioprine; Crohn Disease; Dose-Response Relationship, Drug; Double-Blind Method; Drug Administration Schedule; Drug Monitoring; Female; Gene Frequency; Genotype; Guanine Nucleotides; Humans; Immunosuppressive Agents; Male; Mercaptopurine; Mesalamine; Methyltransferases; Middle Aged; Phenotype; Postoperative Period; Prospective Studies; Thioguanine; Thionucleotides; Time Factors

A prospective randomized trial in patients with Crohn disease studied whether 6-thioguanine nucleotide (6-TGN) concentration-adapted azathioprine (AZA) therapy is clinically superior to a standard dose of 2.5 mg/kg/day AZA.. After 2 weeks of standard therapy, patients (n = 71) were randomized into standard (n = 32) or adapted-dose (n = 25) groups; 14 patients dropped out before randomization. In the adapted group, the AZA dose was adjusted to maintain 6-TGN concentrations between 250 and 400 pmol/8 x 10(8) erythrocytes (Ery). Response criteria were the number of patients in remission after 16 weeks without steroids (primary) and remission after 24 weeks, frequency of side effects, and quality of life (secondary).. After 16 weeks, 14 of 32 (43.8%) patients in the standard group vs 11 of 25 (44%) in the adapted group were in remission without steroids (intent-to-treat analysis). After 24 weeks, 43.8% vs 40% were in remission. No significant differences were found concerning quality of life, disease activity, 6-TGN concentrations, AZA dose, or dropouts due to side effects. Sixty-six patients had a wild-type thiopurine S-methyltransferase (TPMT) genotype, with TPMT activities of 8 to 20 nmol/(mL Ery x h). Five patients (dropouts after randomization) were heterozygous, with TPMT activities <8 nmol/(mL Ery x h). 6-Methyl mercaptopurine (6-MMP) concentrations >5700 pmol/8 x 10(8) Ery were not associated with hepatotoxicity.. Standard and adapted dosing with the provided dosing scheme led to identical 6-TGN concentrations and remission rates. Adapted dosing had no apparent clinical benefit for patients with TPMT activity between 8 and 20 nmol/(mL Ery x h). Additionally, 6-MMP monitoring had no predictive value for hepatotoxicity.

Topics: Adolescent; Adult; Aged; Chemical and Drug Induced Liver Injury; Crohn Disease; Dose-Response Relationship, Drug; Erythrocyte Indices; Female; Genotype; Guanine Nucleotides; Humans; Leukopenia; Male; Mercaptopurine; Methyltransferases; Middle Aged; Remission Induction; Thionucleotides; Thrombocytopenia

To elucidate interpatient variability in thioguanine nucleotide (TGN) concentrations in acute lymphoblastic leukemia (ALL) cells, we determined the TGN concentrations in leukemic blasts from 82 children with newly diagnosed ALL after intravenous administration of mercaptopurine (MP). Patients treated with MP alone achieved higher TGN concentrations than those treated with the combination of methotrexate plus mercaptopurine (MTX + MP). Analysis of the expression of approximately 9600 genes in ALL cells obtained at diagnosis identified 60 gene probes significantly associated with TGN accumulation in patients treated with MP alone and 75 gene probes in patients treated with MTX + MP, with no overlap between the 2 sets of genes. Genes significantly associated with intracellular TGN accumulation after MP alone included those encoding MP metabolic enzymes and transporters (eg, SLC29A1). Inhibition of SLC29A1 by nitrobenzylmercaptopurine ribonucleoside (NBMPR) caused a 33% to 45% reduction of TGN in ALL cells in vitro (P < .006), consistent with the gene expression findings. Genes associated with TGN concentration after combination therapy included those involved in protein and adenosine triphosphate (ATP)-biosynthesis. Together, these in vivo and in vitro data provide new insight into the genomic basis of interpatient differences in intracellular TGN accumulation and reveal significant differences between treatment with MP alone and treatment with MP and MTX.

Topics: Adolescent; Cell Line, Tumor; Cell Proliferation; Dose-Response Relationship, Drug; Gene Expression Profiling; Genotype; Guanine Nucleotides; Humans; Mercaptopurine; Phenotype; Phylogeny; Precursor Cell Lymphoblastic Leukemia-Lymphoma; Thionucleotides; Tissue Distribution

6-thioguanine (6-TG) has emerged as a promising therapeutic alternative in patients with Crohn's disease intolerant or resistant to azathioprine (AZA) and/or 6-mercaptopurine (6-MP). The aim of the present study was to evaluate the safety and efficacy of 6-TG in patients with ulcerative colitis (UC) or indeterminate colitis (IC) intolerant or resistant to AZA/6-MP.. Twenty patients with an acute flare, steroid-dependent or steroid-refractory disease attending our outpatient department were included in the study. Measurement of 6-TG nucleotide levels was done to check compliance. Complete, partial and non-response were defined by means of the clinical activity index and the daily steroid demand. Secondary outcome parameters included changes in cumulative steroid doses, C-reactive protein (CRP) levels, and an endoscopic score.. Out of 20 patients 4 were excluded owing to noncompliance; 2/16 compliant patients (13%) had to be prematurely withdrawn because of adverse events, which ceased upon drug discontinuation. By per-protocol analysis, 5/14 patients (36%) were complete, 6/14 (43%) partial and 3/14 (21%) non-responders. In addition to the reduction of the cumulative steroid dose over 3 months, CRP decreased in the study population and the endoscopic score decreased in treatment responders.. Treatment with 6-TG was effective in patients with UC or IC previously intolerant or resistant to AZA/6-MP. Future work is needed to define a subpopulation of patients at low risk for its potential hepatotoxicity, which we assume will benefit from 6-TG.

Topics: Adult; Aged; Anti-Inflammatory Agents; Azathioprine; Biomarkers; C-Reactive Protein; Colitis, Ulcerative; Dose-Response Relationship, Drug; Endoscopy, Gastrointestinal; Female; Genotype; Guanine Nucleotides; Humans; Male; Mercaptopurine; Methylthioinosine; Methyltransferases; Middle Aged; Orosomucoid; Patient Compliance; Prospective Studies; Quality of Life; Thioguanine; Thionucleotides; Treatment Outcome

Proper prospective pharmacokinetic studies of 6-mercaptopurine (6-MP) in inflammatory bowel disease (IBD) patients are lacking. As a result, conflicting recommendations have been made for metabolite monitoring in routine practice. The authors have evaluated 6-MP pharmacokinetics in IBD patients, including the genetic background for thiopurine methyltransferase (TPMT). Red blood cell (RBC) 6-thioguanine nucleotide (6-TGN) and 6-methylmercaptopurine ribonucleotide (6-MMPR) concentrations were measured in 30 IBD patients at 1, 2, 4, and 8 weeks after starting 6-MP, 50 mg once daily. Outcome measures included mean 6-TGN and 6-MMPR concentrations (+/- 95% confidence interval, CI95%) and their associations with TPMT genotype, 6-MP dose, and hematologic, hepatic, pancreatic, and efficacy parameters during the 8-week period. Steady-state concentrations were reached after 4 weeks, indicating a half-life of approximately 5 days for both 6-TGN and 6-MMPR; the concentrations were 368 (CI95% 284-452) and 2837 (CI95% 2101-3573) pmol/8 x 10 RBCs, respectively. Large interpatient variability occurred at all time points. TPMT genotype correlated with 6-TGN concentrations (0.576, P < 0.01), and patients with mutant alleles had a relative risk (RR) of 12.0 (CI95% 1.7-92.3) of developing leukopenia. A 6-MMPR/6-TGN ratio less than 11 was associated with therapeutic efficacy. Based on this pharmacokinetic analysis, therapeutic drug monitoring is essential for rational 6-MP dosing.

Topics: Adult; Aged; Drug Monitoring; Female; Genotype; Guanine Nucleotides; Half-Life; Humans; Immunosuppressive Agents; Inflammatory Bowel Diseases; Male; Mercaptopurine; Methyltransferases; Middle Aged; Prospective Studies; Purine Nucleotides; Thionucleotides

Mean corpuscular volume may correlate with erythrocyte 6-thioguanine nucleotide concentrations in patients treated with azathioprine and 6-mercaptourine. We conducted a study of 166 patients with inflammatory bowel disease treated with azathioprine or 6-mercaptopurine to determine the relationship between mean corpuscular volume and erythrocyte 6-thioguanine nucleotide concentrations, disease activity as measured by the Inflammatory Bowel Disease Questionnaire (active disease <170, remission >170), and leukopenia. Blood was submitted for mean corpuscular volume, whole blood 6-thioguanine nucleotide concentration, and leukocyte count. The mean +/- SD mean corpuscular volume during treatment was 94.7 +/- 6.6 fL and the mean +/- SD change in mean corpuscular volume was 7.5 +/- 6.3 fL. There were significant correlations between mean corpuscular volume and erythrocyte 6-thioguanine nucleotide concentration (r(s) = 0.33, p < 0.001) and between change from baseline in mean corpuscular volume and erythrocyte 6-thioguanine nucleotide concentration (r(s) = 0.26, p = 0.001). There was no correlation between Inflammatory Bowel Disease Questionnaire scores and mean corpuscular volume values (r(s) = 0.01, p = 0.94). The mean corpuscular volume values in 55 patients with active disease and 111 patients in remission were similar (95.1 vs. 94.5 fL, p = 0.57). There was a weak negative correlation between the mean corpuscular volume and the leukocyte count, (r(s) = -0.18, p = 0.022). In patients with inflammatory bowel disease treated with azathioprine or 6-mercaptopurine, mean corpuscular volume and change from baseline in mean corpuscular volume correlated with erythrocyte 6-thioguanine nucleotide concentrations and negatively with leukocyte counts, but did not correlate with disease activity as measured by the Inflammatory Bowel Disease Questionnaire. Measurement of mean corpuscular volume is a simple and inexpensive alternative to measurement of 6-thioguanine nucleotide concentrations in patients treated with azathioprine or 6-mercaptopurine.

Topics: Adolescent; Adult; Aged; Aged, 80 and over; Azathioprine; Biomarkers; Endpoint Determination; Erythrocyte Indices; Erythrocytes; Female; Guanine Nucleotides; Humans; Immunosuppressive Agents; Inflammatory Bowel Diseases; Leukocyte Count; Male; Mercaptopurine; Middle Aged; Severity of Illness Index; Surveys and Questionnaires; Thionucleotides; Treatment Outcome

6-Mercaptopurine (6MP) has been regarded as nonleukemogenic, even though the cytotoxicity of 6MP depends on the incorporation of 6-thioguanine nucleotides (6TGN) into DNA. In hematopoietic cells this pathway competes with S-methylation catalyzed by thiopurine methyltransferase (TPMT). However, methylated 6MP metabolites inhibit purine de novo synthesis and thus may enhance incorporation of 6TGN into DNA. Approximately 10% of white individuals have low TPMT activity as a result of polymorphisms in the TPMT gene. The authors attempted to test the hypothesis that the degree of DNA damage during 6MP therapy might reflect variations in 6MP metabolism and pharmacokinetics.. The authors measured TPMT activity as well as erythrocyte levels of 6TGN (E-6TGN) and methylated 6MP metabolites (E-MeMP) during 6MP therapy in 439 children with acute lymphoblastic leukemia, 5 of whom later developed secondary myelodysplasia or acute myeloid leukemia (sMDS/AML).. The patients who developed sMDS/AML had significantly lower TPMT activity compared with the remaining patients (P = 0.03). The 55 patients with TPMT activity <14 U/mL red blood cells (RBC) (antimode of the bimodal distribution) had a 5-year risk of sMDS/AML of 9 +/- 6% versus 1 +/- 1% for the remaining patients (P = 0.002). Cox regression analysis identified TPMT activity and E-MeMP level as the strongest predictors of risk for sMDS/AML (global P value = 0.02). Patients with low TPMT activity and high E-MeMP levels had the highest risk. All 5 patients with sMDS/AML had E-6TGN and/or E-MeMP levels > the 90% percentiles or had TPMT activity < 14 U/mL RBC.. These data demonstrate an increased leukemogenic risk when 6MP is administered with other cytotoxic agents in patients with low TPMT activity, and indicate that not only high 6TGN levels but also high levels of methylated metabolites may lead to DNA damage.

Topics: Adolescent; Antineoplastic Combined Chemotherapy Protocols; Bone Marrow Cells; Child; Child, Preschool; DNA Damage; Guanine Nucleotides; Humans; Immunophenotyping; Infant; Mercaptopurine; Methyltransferases; Myelodysplastic Syndromes; Precursor Cell Lymphoblastic Leukemia-Lymphoma; Risk; Thionucleotides

Mercaptopurine (6MP) has been the standard drug for maintenance therapy of acute lymphoblastic leukemia. In a multicenter study we investigated whether thioguanine (6TG), which is converted more directly to the cytotoxic thioguanine nucleotides (TGN), offers a therapeutic advantage over 6MP. In this study (COALL-92), 6TG was randomized versus 6MP in maintenance therapy, whereby the doses of both drugs had to be adjusted to a white blood cell (WBC) count of between 2 and 3/nl. In 19 children the plasma levels of both drugs and/or the accumulation of their metabolites in red blood cells (RBC) were measured during intensive treatment in two consecutive chemotherapy blocks, and in 54 children the metabolites in RBC were measured every 3 months during daily treatment in maintenance therapy. There was a marked interindividual difference in the plasma kinetics of the two drugs; after identical doses of 100 mg/m2 an about 4-fold higher peak concentration of the parent drug was reached with 6MP. The main metabolites of 6TG were thioguanine nucleotides (TGN), whereas during 6MP treatment, methylated thioinosine nucleotides (TIN) predominated in erythrocytes. In patients receiving 6TG during maintenance therapy (22 patients) the concentration of methylated TGN reached about 40% of that of unmethylated TGN; after 6MP administration (32 patients) the methylated TIN were concentrated about 26-fold higher in RBC than were TGN. In contrast to 6TG, for 6MP the pattern of metabolites shifted toward the methylated ones with increasing dose. The median TGN concentration was about 7-fold higher in the TG branch, although the median dose was only about 70% of that of 6MP. The WBC values were equivalent in the two treatment groups. Our results suggest that the cytotoxic effect of 6MP is not based solely on the formation of TGN.

Topics: Adolescent; Antimetabolites, Antineoplastic; Child; Child, Preschool; Erythrocytes; Female; Guanine Nucleotides; Humans; Infant; Leukocyte Count; Male; Mercaptopurine; Precursor Cell Lymphoblastic Leukemia-Lymphoma; Thioguanine; Thioinosine; Thionucleotides

Topics: Azathioprine; Erythrocytes; Graft Rejection; Guanine Nucleotides; Humans; Immunosuppressive Agents; Kidney Transplantation; Mercaptopurine; Methyltransferases; Prospective Studies; Thionucleotides

In the recent era of growing availability of biological agents, the role of thiopurines needs to be reassessed with the focus on toxicity.. We assessed the incidence and predictive factors of thiopurine-induced adverse events (AE) resulting in therapy cessation in pediatric inflammatory bowel disease (IBD), related to thiopurine metabolites and biochemical abnormalities, and determined overall drug survival.. We performed a retrospective, single-center study of children diagnosed with IBD between 2000 and 2019 and treated with thiopurine therapy. The incidence of AE and overall drug survival of thiopurines were evaluated using the Kaplan-Meier method. Correlations between thiopurine metabolites and biochemical tests were computed using Spearman's correlation coefficient.. Of 391 patients with IBD, 233 patients (162 Crohn's disease, 62 ulcerative colitis, and 9 IBD-unclassified) were prescribed thiopurines (230 azathioprine and 3 mercaptopurine), of whom 50 patients (22%) discontinued treatment, at least temporary, due to thiopurine-induced AE (median follow-up 20.7 months). Twenty-six patients (52%) were rechallenged and 18 of them (70%) tolerated this. Sixteen patients (6%) switched to a second thiopurine agent after azathioprine intolerance and 10 of them (63%) tolerated this. No predictive factors for development of AE could be identified. Concentrations of 6-thioguanine nucleotides (6-TGN) were significantly correlated with white blood cell and neutrophil count, 6-methylmercaptopurine (6-MMP) concentrations with alanine aminotransferase and gamma-glutamyltranspeptidase.. Approximately 20% of pediatric patients with IBD discontinued thiopurine treatment due to AE. A rechallenge or switch to mercaptopurine is an effective strategy after development of AE. Concentrations of 6-TGN and 6-MMP are associated with biochemical abnormalities.

Topics: Adolescent; Antimetabolites; Azathioprine; Biomarkers, Pharmacological; Child; Cohort Studies; Colitis, Ulcerative; Crohn Disease; Drug Substitution; Drug-Related Side Effects and Adverse Reactions; Female; Guanine Nucleotides; Humans; Male; Mercaptopurine; Netherlands; Retrospective Studies; Thionucleotides; Withholding Treatment

In inflammatory bowel disease (IBD), conventional thiopurine users cease treatment in 60% of cases within 5 years, mostly because of adverse events or nonresponse. In this study, the authors aimed to investigate the role of 6-thioguanine nucleotide (TGN) measurements, geno/phenotyping of thiopurine S-methyltransferase (TPMT), and their mutual relationship with TG therapy in IBD.. An international retrospective, multicenter cohort study was performed at 4 centers in the Netherlands (Máxima Medical Centre) and the United Kingdom (Guy's and St. Thomas' Hospital, Queen Elizabeth Hospital, and East Surrey Hospital).. Overall, 526 6-TGN measurements were performed in 316 patients with IBD. The median daily dosage of TG was 20 mg/d (range 10-40 mg/d), and the median duration of TG use was 21.1 months (SD, 28.0). In total, 129 patients (40.8%) had a known TPMT status. In the variant-type and wild-type TPMT genotype metabolism groups, median 6-TGN values were 1126 [interquartile range (IQR) 948-1562] and 467.5 pmol/8 × 10E8 red blood cells (RBCs) (IQR 334-593). A significant difference was observed between the 2 groups (P = 0.0001, t test). For TPMT phenotypes, in the slow, fast, and normal metabolism groups, the median 6-TGN values were 772.0 (IQR 459-1724), 296.0 (IQR 200-705), and 774.5 pmol/8 × 10E8 RBCs (IQR 500.5-981.5), with a significant difference observed between groups (P < 0.001, analysis of variance).. Our findings indicated that TPMT measurements at TG initiation can be useful but are not necessary for daily practice. TPMT genotypes and phenotypes are both associated with significant differences in 6-TGN levels between metabolic groups. However, the advantage of TG remains that RBC 6-TGN measurements are not crucial to monitor treatments in patients with IBD because these measurements did not correlate with laboratory result abnormalities. This presents as a major advantage in countries where patients cannot access these diagnostic tests.

Topics: Adult; Azathioprine; Female; Genotype; Guanine Nucleotides; Humans; Immunosuppressive Agents; Inflammatory Bowel Diseases; Male; Mercaptopurine; Methyltransferases; Middle Aged; Phenotype; Retrospective Studies; Thioguanine; Thionucleotides

Combining therapy with a thiopurine is favored when commencing infliximab in Crohn's disease; however, the optimal 6-thioguanine nucleotide (TGN) level and how long to continue thiopurines after induction are uncertain. We aimed to compare outcomes after induction and during maintenance in combination therapy versus infliximab monotherapy in Crohn's and to examine whether TGN levels were associated with outcomes.. Crohn's patients induced with infliximab with or without concomitant thiopurines were retrospectively identified. Response to induction and clinical outcomes in subsequent 6-month maintenance semesters were analyzed. A TGN level ≥235 pmol/8 × 10. In 89 patients, response to induction was higher in combination therapy than monotherapy (74 vs 47%, P = 0.04). This benefit was only seen in patients with a therapeutic TGN (odds ratio 3.72, confidence interval 1.07-13.0, P = 0.04). Combination therapy during induction yielded a three times longer time to subsequent need for treatment escalation or treatment failure compared with monotherapy (29 vs 9 months, P = 0.01), with both therapeutic and subtherapeutic TGNs independent predictors on multivariate analysis. Among 370 semesters, there was no difference in outcomes between combination therapy and monotherapy (P = 0.42), nor when combination semesters were stratified by therapeutic versus subtherapeutic TGN (P = 0.56). In semester 1 only, a significantly higher remission rate was observed with therapeutic compared with subtherapeutic TGN (76% vs 33%, P = 0.02).. Combination therapy dosed with an optimized thiopurine was superior to infliximab monotherapy for induction of response, durability of response, and clinical outcomes in the first 6 months following induction. Thereafter, combination therapy yielded no clinical advantage, supporting consideration of thiopurine withdrawal on a case-by-case basis.

Topics: Azathioprine; Biomarkers; Crohn Disease; Drug Therapy, Combination; Female; Guanine Nucleotides; Humans; Immunosuppressive Agents; Infliximab; Maintenance Chemotherapy; Male; Mercaptopurine; Remission Induction; Retrospective Studies; Thionucleotides; Treatment Outcome

Thiopurines are key drugs in maintenance therapy for treating inflammatory bowel disease (IBD). Time-dependent 5-aminosalicylates (5-ASA) releasing preparations (time-dependent 5-ASA) increase 6-thioguanine nucleotide (6-TGN), an active metabolite of thiopurines. However, the effects of pH-dependent 5-ASA releasing preparations (pH-dependent 5-ASA) on thiopurine metabolism were not reported.. We conducted a retrospective study of 134 IBD patients who received thiopurine treatment. The 6-methylmercaptopurine (6-MMP)/6-TGN values after taking the same dose of thiopurine preparations for at least 28 days were included.. There was a significant decrease in the 6-MMP/6-TGN ratio in time-dependent 5-ASA compared with group without 5-ASA preparations and the pH-dependent 5-ASA group (p = 0.008 and < 0.001 respectively). Spearman's rank correlation coefficient indicated a negative relationship between the daily oral dose of time-dependent 5-ASA and the 6-MMP/6-TGN ratio (r = -0.362, p = 0.003). Multivariate logistic regression analysis was performed in the groups with 6-MMP/6-TGN ratios of 1 or more and less than 1. The use of time-dependent 5-ASA and concomitant allopurinol negatively affected the independent 6-MMP/6-TGN ratio (p = 0.006 and 0.007 respectively).. Our study revealed that time-dependent but not pH-dependent 5-ASA decreases the 6-MMP/6-TGN ratio. We also confirmed that concomitant allopurinol results in a low 6-MMP/6TGN ratio.

Topics: Administration, Oral; Adolescent; Adult; Aged; Allopurinol; Anti-Inflammatory Agents, Non-Steroidal; Drug Interactions; Drug Liberation; Female; Guanine Nucleotides; Humans; Hydrogen-Ion Concentration; Immunosuppressive Agents; Inflammatory Bowel Diseases; Male; Mercaptopurine; Mesalamine; Middle Aged; Retrospective Studies; Thionucleotides; Time Factors; Young Adult

Thiopurines, commonly used to treat inflammatory bowel disease, cause lymphopenia and red blood cell macrocytosis, requiring therapeutic monitoring. Mean corpuscular volume/white blood cell (MCV/WBC) ratio has been proposed as a surrogate for therapeutic monitoring. Our aim was to investigate MCV/WBC ratio for assessing clinical response to thiopurines among pediatric patients with inflammatory bowel disease.. We performed a retrospective cross-sectional study at a tertiary care center using laboratory results and standardized physician global assessments (PGA) among pediatric patients taking thiopurines. Erythrocyte sedimentation rate (ESR), C-reactive protein (CRP), fecal calprotectin, and 6-thioguanine nucleotides were assessed when available. The primary outcome was association between MCV/WBC ratio and clinical remission assessed by ESR, CRP, calprotectin, or PGA. We also used a composite outcome requiring all available data to be normal. Analyses were limited to 1 occurrence per patient, >60 days after starting thiopurine, and comparators were required to be within 14 days of one another.. A total of 471 patients met inclusion criteria. MCV/WBC ratio poorly predicted quiescent disease as defined by PGA (area under receiver operating characteristic curve [AuROC] 0.55, 95% confidence interval [CI] 0.43-0.66). MCV/WBC ratio better predicted quiescent disease defined as normal CRP (AuROC 0.64, 95% CI 0.58-0.70) or normal ESR (AuROC 0.59, 95% CI 0.52-0.66). When the composite outcome measure was used, MCV/WBC ratio had an AuROC of 0.65 (95% CI 0.59-0.70), indicating it is reasonably accurate in discriminating between clinical remission and active disease.. MCV/WBC ratio is a noninferior, easy, and low-cost alternative to thiopurine metabolite monitoring.

Topics: Adolescent; Area Under Curve; Azathioprine; C-Reactive Protein; Child; Colitis, Ulcerative; Crohn Disease; Cross-Sectional Studies; Erythrocyte Indices; Feces; Female; Guanine Nucleotides; Humans; Immunosuppressive Agents; Leukocyte Count; Leukocyte L1 Antigen Complex; Male; Mercaptopurine; Predictive Value of Tests; Retrospective Studies; ROC Curve; Severity of Illness Index; Thionucleotides; Young Adult

The aim of this study was to assess whether sustained 6-thioguaninenucleotide (6-TGN) levels were associated with improved long-term outcomes in patients with inflammatory bowel diseases (IBD).. Cross-sectional data have shown that thiopurine metabolites are correlated with clinical efficacy in patient receiving thiopurines for IBD but the role for serial measurements through treatment course is unclear.. We conducted a retrospective cohort study including patients with IBD on thiopurine monotherapy and had serial 6-TGN levels measured. Predictive variables included demographics, disease phenotype, 6-TGN levels (nadir, median, and peak levels). The primary outcome was the development of a disease relapse. The secondary outcome was the need for IBD surgery.. Two hundred eighteen 6-TGN samples from 87 patients were analyzed. Nadir and median 6-TGN levels were significantly higher in patients who did not relapse [185 and 233 pmol per 8×10 red blood cells (RBCs)] as compared with levels in patients who did relapse (150 and 167 pmol per 8×10 RBCs, P=0.025) but there was no significant difference in peak 6-TGN level. When adjusted for confounding factors, a nadir 6-TGN level ≥161 and a median 6-TGN level ≥264 were associated with a significant decrease in the rate of disease exacerbation (hazard ratio: 0.5; 95% confidence interval, 0.26-0.87; P=0.016 and hazard ratio: 0.4; 95% confidence interval, 0.2-0.82; P=0.14).. Serial thiopurine metabolite level assessments and dose adjustment aiming to maintain higher 6-TGN levels may be helpful to improve long-term outcomes in patients with IBD.

Topics: Adult; Anti-Inflammatory Agents; Azathioprine; Biomarkers; Colitis, Ulcerative; Crohn Disease; Digestive System Surgical Procedures; Drug Monitoring; Female; Gastrointestinal Agents; Guanine Nucleotides; Humans; Longitudinal Studies; Male; Mercaptopurine; Middle Aged; Predictive Value of Tests; Recurrence; Remission Induction; Retrospective Studies; Thionucleotides; Time Factors; Treatment Outcome; Up-Regulation; Young Adult

We aimed to identify the impact of NUDT15 variants on thiopurine intolerance and 6-thioguanine nucleotide (6-TGN) levels in Korean children with acute lymphoblastic leukemia (ALL).. Genotyping of NUDT15 was tested in 258 patients with ALL registered at Samsung Medical Center. Patients were classified into normal-activity (wild-type), intermediate-activity (heterozygous variant), and low-activity groups (homozygous or compound heterozygous variant). Clinical and laboratory features during the first year of maintenance therapy were investigated.. A total of 182 patients were included in the final analysis. There were five (2.7%), 46 (25.3%), and 131 (72.0%) patients in low-, intermediate-, and normal-activity groups, respectively. The lowest 6-mercaptopurine (6-MP) dose (mg/m2/day) was administered to the low-activity group (low-activity group 7.5 vs. intermediate-activity group 24.4 vs. normalactivity group 31.1, p < 0.01) from three months to a year after beginning maintenance therapy. The low-activity group experienced the longest duration of therapy interruption during the first year (low-activity group 169 days vs. intermediate-activity group 30 days vs. normal-activity group 16 days, p < 0.01). They also showed the lowest blood cell counts and had a longer duration of leukopenia (low-activity group 131 days vs. intermediate-activity group 92 days vs. normal-activity group 59 days, p < 0.01). 6-TGN level and its ratio to 6-MP dose were lowest in the low-activity group.. NUDT15 variants cause hematopoietic toxicity with low 6-TGN levels. NUDT15 genotyping should be conducted before administering thiopurine, and dose adjustments require caution regardless of 6-TGN levels.

Topics: Adolescent; Child; Child, Preschool; Drug Administration Schedule; Female; Guanine Nucleotides; Humans; Infant; Male; Mercaptopurine; Pharmacogenomic Variants; Polymorphism, Single Nucleotide; Precursor Cell Lymphoblastic Leukemia-Lymphoma; Pyrophosphatases; Republic of Korea; Retrospective Studies; Sequence Analysis, DNA; Thionucleotides; Young Adult

Azathioprine is frequently used in severe eczema. It is converted in the liver into active metabolites, including 6-thioguanine nucleotide (6-TGN) and methylated 6-methylmercaptopurine (6-MMP). In the past, the therapeutic potential of azathioprine may have not been fully utilized. Recent investigations on inflammatory bowel disease have led to a better understanding of azathioprine metabolism and optimizing treatment.. To investigate whether measuring thiopurine metabolites in circulation can improve the effectiveness and safety of azathioprine treatment in patients with atopic dermatitis and/or chronic hand/foot eczema.. Azathioprine metabolite levels were measured in eczema patients during maintenance treatment (Part I) and dose escalation (Part II). Clinical effectiveness, hepatotoxicity, and bone marrow suppression were analyzed and TPMT genotype was assessed.. A wide variation in metabolite levels in all dose groups was observed. In Part I (32 patients), there were no significant differences in 6-TGN levels between clinical responders and non-responders (p = .806). No hepatoxicity or myelotoxicity was observed. In Part II, all 6-TGN and 6-MMP levels increased during dose escalation. Hypermethylation was observed in 2/8 patients.. For individual eczema patients treated with azathioprine, routinely measuring 6-TGN and 6-MMP can be helpful in optimizing azathioprine dose, improving clinical effectiveness, and preventing side effects.

Topics: Adult; Azathioprine; Chromatography, High Pressure Liquid; Dermatitis, Atopic; Eczema; Female; Guanine Nucleotides; Humans; Immunosuppressive Agents; Male; Mercaptopurine; Middle Aged; Thionucleotides; Treatment Outcome

Topics: Adult; Allopurinol; Azathioprine; Chronic Disease; Dermatitis, Atopic; Drug Therapy, Combination; Eczema; Female; Foot Dermatoses; Guanine Nucleotides; Hand Dermatoses; Humans; Immunosuppressive Agents; Male; Mercaptopurine; Middle Aged; Prospective Studies; Thionucleotides; Time Factors; Treatment Outcome

Interactions between principal cytotoxic thiopurine metabolites, that is 6-thioguanine nucleotides [6-TGN], and infliximab [IFX] and anti-IFX antibodies [Abs] may contribute to higher effectiveness of IFX-thiopurine combination therapy than monotherapies in inflammatory bowel disease.. To examine if thiopurine metabolites influenced trough IFX and anti-IFX Abs, 89 patients previously assessed for anti-IFX Abs were included. To assess if IFX influenced thiopurine metabolites, eight patients who had responded to 12 weeks of intensified IFX at a constant thiopurine dosing were included.. In the first cohort, IFX-thiopurine combination therapy reduced anti-IFX Ab detection [8/40; 20%] as compared with IFX monotherapy [22/49; 45%], odds ratio [OR] 0.31 [0.12-0.80], p < 0.05. 6-TGN was significantly lower in anti-IFX Ab-positive patients (50 pmol/8 × 108 red blood cells [RBC] vs 105, p < 0.01). All anti-IFX Ab-positive patients had 6-TGN < 117 pmol/8 × 108 RBC (sensitivity 100% [63-100], specificity 47% [29-65], area under the curveROC = 0.82, p < 0.01). Trough IFX was similar between anti-IFX Ab-negative patients in IFX monotherapy and IFX-thiopurine combination therapy [5.1 μg/mL vs 4.9, p = 0.76]. 6-TGN and IFX did not correlate [rP = 0.04, p = 0.83; rS = 0.02, p = 0.89, respectively]. In the second cohort, trough IFX increased during IFX intensification [ΔIFX median 6.5 μg/mL, p = 0.02], but 6-TGN was stable [6-TGN at Weeks 0, 4, 8, 12: 90 pmol/8 × 108 RBC, 93, 101, 90; p > 0.05]. Methylated mercaptopurine metabolite associations were consistently negative.. Superior effect of IFX-thiopurine combination therapy over monotherapies partly relates to decrease in anti-IFX Abs, which associates with 6-TGN levels and has a lower therapeutic threshold than during thiopurine monotherapy. Additional benefit likely ascribes to synergy between different anti-inflammatory modes of action rather than direct drug interactions.

Topics: Adult; Antibodies; Drug Synergism; Drug Therapy, Combination; Erythrocytes; Female; Gastrointestinal Agents; Guanine Nucleotides; Humans; Immunosuppressive Agents; Inflammatory Bowel Diseases; Infliximab; Male; Mercaptopurine; Methyltransferases; Middle Aged; Retrospective Studies; Thionucleotides

Up to 20% of patients with inflammatory bowel disease (IBD) who are refractory to thiopurine therapy preferentially produce 6-methylmercaptopurine (6-MMP) at the expense of 6-thioguanine nucleotides (6-TGN), resulting in a high 6-MMP:6-TGN ratio (>20). The objective of this study was to evaluate whether genetic variability in guanine monophosphate synthetase (GMPS) contributes to preferential 6-MMP metabolizer phenotype.. Exome sequencing was performed in a cohort of IBD patients with 6-MMP:6-TGN ratios of >100 to identify nonsynonymous single nucleotide polymorphisms (nsSNPs). In vitro assays were performed to measure GMPS activity associated with these nsSNPs. Frequency of the nsSNPs was measured in a cohort of 530 Caucasian IBD patients.. Two nsSNPs in GMPS (rs747629729, rs61750370) were detected in 11 patients with very high 6-MMP:6-TGN ratios. The 2 nsSNPs were predicted to be damaging by in silico analysis. In vitro assays demonstrated that both nsSNPs resulted in a significant reduction in GMPS activity (P < 0.05). The SNP rs61750370 was significantly associated with 6-MMP:6-TGN ratios ≥100 (odds ratio, 5.64; 95% confidence interval, 1.01-25.12; P < 0.031) in a subset of 264 Caucasian IBD patients.. The GMPS SNP rs61750370 may be a reliable risk factor for extreme 6MMP preferential metabolism.

Topics: Adult; Azathioprine; Carbon-Nitrogen Ligases with Glutamine as Amide-N-Donor; Cohort Studies; Female; Guanine Nucleotides; Humans; Inflammatory Bowel Diseases; Male; Mercaptopurine; Middle Aged; Polymorphism, Single Nucleotide; Risk Factors; Thionucleotides; Young Adult

Discontinuation of thiopurine analogues is common prior to live vaccines, during infection or when de-escalating therapy. Data regarding clearance of active metabolites and immune re-constitution is scant. We aimed to determine drug elimination and immune re-constitution following thiopurine cessation.. The elimination kinetics of 6-thioguanine nucleotides (6-TGN) were determined in nine inflammatory bowel disease [IBD] patients discontinuing thiopurines. Immune reconstitution was evaluated by toxic shock syndrome toxin 1 [TSST1] or anti-CD3 [OKT3]-induced CD4+ T-cell proliferation, following an initial exposure to TSST1 and 6-mercaptopurine [6MP], separately or combined.. All patients discontinuing thiopurines displayed first-order elimination kinetics of 6-TGN, with a median elimination half-life of 6.8 days [interquartile range 5.9-8.4]. Resting CD4+ T-cells exposed to 6MP preserved their response to subsequent polyclonal or Vβ2+-preferential stimulation. By contrast, exposure of TSST1-activated CD4+ T-cells to 6MP inhibited their subsequent Vβ2+clonal response to further stimulation [p = 0.008], whereas overall response to further non-Vβ2-selective stimulation with OKT3 was unaltered [p = 0.9].. Upon 6MP/azathioprine discontinuation, a 6-TGN elimination half-life of less than 10 days is expected in most patients. Immune reconstitution, however, may take longer for T-cell clones exposed to stimulation during thiopurine treatment. These findings may be useful when considering thiopurine cessation.

Topics: Adult; Azathioprine; Bacterial Toxins; Cell Proliferation; Enterotoxins; Female; Guanine Nucleotides; Humans; Immune Reconstitution; Immunosuppressive Agents; Inflammatory Bowel Diseases; Male; Mercaptopurine; Metabolic Clearance Rate; Muromonab-CD3; Superantigens; T-Lymphocytes; Thionucleotides; Withholding Treatment

Azathioprine (AZA) is the mainstay of maintenance therapy in pediatric autoimmune hepatitis (AIH). The use of thiopurines metabolites to individualize therapy and avoid toxicity has not, however, been clearly defined.. Retrospective analysis of children ≤18 years diagnosed with AIH between January 2001 and 2016. Standard definitions were used for treatment response and disease flare. Thiopurine metabolite levels were correlated with the corresponding liver function test.. A total of 56 children (32 girls) were diagnosed with AIH at a median age of 11 years (interquartile range [IQR] 9). No difference in 6-thioguanine-nucleotide (6-TG) levels (271[IQR 251] pmol/8 × 10 red blood cell vs 224 [IQR 147] pmol/8 × 10 red blood cell, P = 0.06) was observed in children in remission when compared with those who were not in remission. No correlation was observed between the 6-TG and alanine aminotransferase levels (r = -0.179, P = 0.109) or between 6-methyl-mercaptopurine (6-MMP) and alanine aminotransferase levels (r = 0.139, P = 0.213). The 6-MMP/6-TG ratio was significantly lower in patients who were in remission (2[7] vs 5 (10), P = 0.04). Using a quartile analysis, we found that having a ratio of <4 was significantly associated with being in remission with OR 2.50 (95% confidence interval 1.02-6.10), P = 0.047. Use of allopurinol with low-dose AZA in 6 children with preferential 6-MMP production brought about remission in 5/6 (83.3%).. Thiopurine metabolite levels should be measured in patients with AIH who have experienced a loss of remission. A 6-MMP/6-TG ratio of <4 with the addition of allopurinol could be considered in these patients.

Topics: Allopurinol; Antimetabolites; Azathioprine; Child; Female; Guanine Nucleotides; Hepatitis, Autoimmune; Humans; Liver Function Tests; Maintenance Chemotherapy; Male; Mercaptopurine; Retrospective Studies; Thionucleotides; Treatment Outcome

Thiopurines are the prerequisite for immunomodulation in inflammatory bowel disease (IBD) therapy. When administered in high (oncological) dose, thiopurine metabolites act as purine antagonists, causing DNA-strand breakage and myelotoxicity. In lower IBD dosages, the mode of action is primarily restricted to anti-inflammatory effects. Then, myelosuppression and hepatotoxicity are the most common adverse events of thiopurines. The aim of this study was to assess the effect of thiopurine metabolites on hematologic and hepatic parameters and to determine which patient characteristics are related to generation of thiopurine metabolites.. The authors scrutinized the therapeutic drug monitoring database of the VU University medical center and subsequently merged this database with the Clinical Laboratory database of our hospital covering the same time period (2010-2015).. The authors included 940 laboratory findings of 424 unique patients in this study. Concentrations of 6-thioguanine nucleotides (6-TGN) correlated negatively with red blood cell count, white blood cell count, and neutrophil count in both azathioprine (AZA) and mercaptopurine users. There was a positive correlation with mean corpuscular volume. In patients using 6-thioguanine, 6-TGN concentrations correlated positively with white blood cell count. Furthermore, there was an inverse correlation between patient's age and 6-TGN concentrations in patients using AZA or 6-thioguanine, and we observed an inverse correlation between body mass index and 6-TGN concentrations in patients using AZA or mercaptopurine. No relations were observed with liver test abnormalities.. Thiopurine derivative therapy influenced bone marrow production and the size of red blood cells. Age and body mass index were important pharmacokinetic factors in the generation of 6-TGN.

Topics: Adult; Blood Cell Count; Databases, Factual; Drug Monitoring; Female; Guanine Nucleotides; Humans; Inflammatory Bowel Diseases; Male; Mercaptopurine; Middle Aged; Thionucleotides; Time Factors; Treatment Outcome

NUDT15 R139C (rs116855232) is a recently identified genetic factor responsible for thiopurine-induced leukocytopenia and hair loss. In this study, we investigated the association of NUDT15 R139C with 6-thioguanine nucleotide (6-TGN) levels and thiopurine-induced leukocytopenia in Japanese patients with inflammatory bowel disease (IBD).. Two hundred and sixty-four subjects (103 healthy volunteers and 161 IBD patients treated with thiopurines) were enrolled. Genotyping for NUDT15 R139C was performed using Custom TaqMan® SNP genotyping assays.. The NUDT15 C/C, C/T, and T/T genotypes were 80.7, 18.2, and 1.1 %, respectively. The allelic frequency was 10.2 %. Among 161 IBD patients, there was no significant difference in 6-TGN levels among the NUDT15 genotypes. Forty-five patients (27.9 %) developed leukocytopenia (WBC <3000/μl), and the C/T and T/T genotypes were significantly associated with the development of leukocytopenia (P = 1.7 × 10(-5)). In these patients, 6-TGN levels were not significantly different between NUDT15 genotypes. NUDT15 R139C was significantly associated with early (<8 weeks) (P = 1.03 × 10(-4)) and late (>8 weeks) leukocytopenia (P = 4.3 × 10(-4)). The decrease in WBC count at 2 and 4 weeks was significantly higher in patients with the C/T or T/T genotypes as compared to the patients with the C/C genotype. All patients with the T/T genotype (n = 2) developed early severe hair loss and severe leukocytopenia (<1000/μl). The logistic regression analysis revealed that NUDT15 R139C was the sole genetic factor responsible for the thiopurine-induced leukocytopenia (P = 0.001).. These results suggest that NUDT15 R139C-related thiopurine-induced leukocytopenia is mediated by a 6-TGN-independent mechanism.

Topics: Adult; Alopecia; Case-Control Studies; Female; Gene Frequency; Genetic Predisposition to Disease; Guanine Nucleotides; Humans; Inflammatory Bowel Diseases; Leukocyte Count; Leukopenia; Male; Mercaptopurine; Middle Aged; Pyrophosphatases; Thionucleotides

6-Mercaptopurine (6-MP) and its prodrug azathioprine (AZA) are used in many autoimmune diseases and after solid-organ transplantation. Their properties are mediated by active metabolites, 6-thioguanine nucleotides (6-TGN), and 6-methylmercaptopurine (6-MMP). The most common adverse effects are myelo- and hepato-toxicity. The aim of the study was quantification of 6-TG and 6-MMP, with the use of liquid chromatography combined with tandem mass spectrometry (LC/MS/MS) in solid-organ transplant recipients.. In 33 patients, kidney transplant recipient (n = 25) and liver transplant recipient (n = 8) intra-erythrocyte concentrations of 6-TG and 6-MMP were measured with the use of LC/MS/MS.. The mean concentration of 6-TG was 205.35 ± 157.62 pmol/8 × 10(8) red blood cells (RBC); median concentration of 6-MMP was 1064.1 (35.78-11,552.9) pmol/8 × 10(8) RBC. There were no correlations between 6-TG levels and peripheral blood parameters (white blood cell count, WBC; hemoglobin, Hb concentration; PLT, blood platelet count) or alanine aminotransferase activity (AlAT) activity. Relationships between 6-MMP concentrations and peripheral blood parameters (WBC, Hb, PLT) or AlAT activity have not been found. Subgroups with leukopenia, anemia, thrombocytopenia, and liver dysfunction did not differ in concentrations of 6-TG or 6-MMP. We have observed a negative correlation between daily azathioprine dose and WBC count (r = -0.37, P = .04).. Relationships between concentrations of azathioprine metabolites and myelotoxicity or hepatotoxicity have not been confirmed. Further studies on larger groups of patients would be helpful in a more accurate understanding of the impact of azathioprine metabolites on parameters of bone marrow and liver function.

Topics: Adult; Azathioprine; Chromatography, Liquid; Erythrocytes; Female; Guanine Nucleotides; Humans; Immunosuppressive Agents; Leukocyte Count; Male; Mercaptopurine; Organ Transplantation; Tandem Mass Spectrometry; Thioguanine; Thionucleotides

Clinical efficacy and risk of complications are associated with intracellular levels of thiopurine metabolites 6-thioguanine nucleotides (6-TGN) and 6-methylmercaptopurines (6-MMP) in patients with Crohn's disease. Therapeutic monitoring of thiopurine metabolites is not widely available. Surrogate markers such as hematologic indices (MCV, leukopenia) have been proposed.. To evaluate accuracy of hematologic indices for prediction of therapeutic levels of thiopurine metabolites.. A retrospective cross-sectional study. We included patients treated with thiopurines for IBD between February 2008 and November 2013. Hematologic indices were correlated with thiopurine metabolites and compared to pre-treatment levels.. A total of 168 patients with 608 measurements were included. Macrocytosis was observed in 4.5 % of the patients. On multivariate analysis, macrocytosis was associated with 6-TGN levels >235 pmol/8 × 10(8) erythrocytes and 6-mmp levels >5,700 pmol/8 × 10(8) erythrocytes. Therapeutic 6-TGN levels were associated with MCV, ΔMCV, macrocytosis and lymphocyte count. Sensitivity and Spearman's r correlation for prediction of therapeutic metabolite levels were poor for all hematologic indices.. Although macrocytosis and an elevated MCV are associated with therapeutic 6-TGN levels, the correlation is weak. None of the evaluated hematologic indices is a reliable surrogate marker for thiopurine metabolite levels.

Topics: Anti-Inflammatory Agents; Azathioprine; Biomarkers; Colitis, Ulcerative; Crohn Disease; Cross-Sectional Studies; Drug Monitoring; Erythrocyte Indices; Female; Guanine Nucleotides; Hematologic Tests; Hemoglobins; Humans; Lymphocyte Count; Male; Mercaptopurine; Platelet Count; Predictive Value of Tests; Retrospective Studies; Thionucleotides; Treatment Outcome

The conventional thiopurines azathioprine and mercaptopurine are considered maintenance immunosuppressive drugs of choice in the treatment of inflammatory bowel disease (IBD). Unfortunately, treatment is often discontinued because of adverse events (AEs) or refractoriness, retrospectively associated with the high levels of the thiopurine metabolites 6-methylmercaptopurine ribonucleotides (6-MMPR). Patients with a clinically "skewed" thiopurine metabolism may be particularly at risk for therapy failure. We determined the predictive value of this pharmacological phenomenon in patients with IBD during regular thiopurine therapy.. Clinical effectiveness and tolerability of weight-based thiopurine therapy were determined in all patients with IBD displaying a skewed metabolism [ratio 6-MMPR/6-thioguanine nucleotide (6-TGN) >20]. All samples were routinely assessed between 2008 and 2012, as part of standard clinical follow-up after initiation of conventional thiopurine therapy.. Forty-one (84%) of 49 included patients with IBD discontinued thiopurines (55% female, 53% with Crohn disease) with a median duration of 14 weeks (range, 7-155). The majority of patients with a skewed metabolism discontinued thiopurines because of adverse events (55%) or refractoriness (12%). The most commonly observed adverse event was hepatotoxicity (18 patients, 37%). Median 6-TGN level was 159 pmol/8 × 10 RBC (range, 46-419), median 6-MMPR level was 11,020 pmol/8 × 10 RBC (range, 3610-43,670), and the median 6-MMPR/6-TGN ratio was 72 (range, 29-367). Thiopurine therapy failure was associated with a ratio above 50 (P < 0.03). Hepatotoxicity occurred more frequently in patients with an extremely skewed metabolism (6-MMPR/6-TGN ratio >100) (P < 0.01).. This study demonstrates that a routinely established skewed metabolism is a major risk factor for future thiopurine failure in patients with IBD. These observations imply that routine thiopurine metabolite measurements may be used as a prognostic tool to identify those patients with an aberrant-skewed metabolism at an early stage, possibly benefitting from therapy adjustments.

Topics: Adolescent; Adult; Aged; Azathioprine; Cohort Studies; Colitis, Ulcerative; Crohn Disease; Female; Follow-Up Studies; Guanine Nucleotides; Humans; Immunosuppressive Agents; Male; Mercaptopurine; Middle Aged; Retrospective Studies; Risk Factors; Thioinosine; Thionucleotides; Treatment Failure; Young Adult

Optimal levels of the thiopurine metabolite, 6-thioguanine nucleotides [6-TGN] correlate with remission of inflammatory bowel disease [IBD]. Apart from variations in the thiopurine methyl transferase [TPMT] gene, little is known about other predictors of 6-TGN levels. Obesity adversely affects response to infliximab and adalimumab and clinical course in IBD, but little is known about the interaction of thiopurines and obesity. We investigated the relationship between body mass index [BMI] and 6-TGN levels and sought to examine other predictors of 6-TGN levels.. This retrospective cohort study included patients with concurrent measurements of 6-TGN and BMI. The association between 6-TGN and clinical variables including BMI was estimated using a multivariable linear regression model.. Of 132 observations, 77 [58%] had Crohn's disease and 55 [42%] ulcerative colitis. BMI, smoking, and TPMT levels were associated with 6-TGN levels in multivariable analysis. Every 5kg/m(2) increase in BMI was associated with an 8% decrease in 6-TGN (0.92; 95% confidence interval [CI] 0.87-0.98; p = 0.009). Smokers had higher 6-TGN levels in comparison with non-/ex-smokers [1.43; 95% CI 1.02-2.02; p = 0.041]. Patients with intermediate TPMT had higher 6-TGN compared to those with normal levels [2.13; 95% CI 1.62-2.80; p < 0.001]. Obese patients were more likely to have sub-therapeutic 6-TGN levels and a higher methyl mercaptopurine nucleotide [MMPN/TGN] ratio despite a similar dose of thiopurines.. Active smoking and intermediate TPMT values were associated with higher 6-TGN levels but increasing BMI resulted in lower 6-TGN and higher MMPN levels. This may explain the worse outcome that has been reported previously in obese IBD subjects.

Topics: Adult; Azathioprine; Biomarkers; Body Mass Index; Colitis, Ulcerative; Crohn Disease; Drug Administration Schedule; Female; Guanine Nucleotides; Humans; Immunosuppressive Agents; Linear Models; Male; Mercaptopurine; Middle Aged; Obesity; Retrospective Studies; Smoking; Thionucleotides; Treatment Outcome

6-Mercaptopurine (6-MP) is one of the key drugs in the treatment of many pediatric cancers, auto immune diseases and inflammatory bowel disease. 6-MP is a prodrug, converted to an active metabolite 6-thioguanine nucleotide (6-TGN) through enzymatic reaction involving thiopurine methyltransferase (TPMT). Pharmacogenomic variation observed in the TPMT enzyme produces a significant variation in drug response among the patient population. Despite 6-MP's widespread use and observed variation in treatment response, efforts at quantitative optimization of dose regimens for individual patients are limited. In addition, research efforts devoted on pharmacogenomics to predict clinical responses are proving far from ideal. In this work, we present a Bayesian population modeling approach to develop a pharmacological model for 6-MP metabolism in humans. In the face of scarcity of data in clinical settings, a global sensitivity analysis based model reduction approach is used to minimize the parameter space. For accurate estimation of sensitive parameters, robust optimal experimental design based on D-optimality criteria was exploited. With the patient-specific model, a model predictive control algorithm is used to optimize the dose scheduling with the objective of maintaining the 6-TGN concentration within its therapeutic window. More importantly, for the first time, we show how the incorporation of information from different levels of biological chain-of response (i.e. gene expression-enzyme phenotype-drug phenotype) plays a critical role in determining the uncertainty in predicting therapeutic target. The model and the control approach can be utilized in the clinical setting to individualize 6-MP dosing based on the patient's ability to metabolize the drug instead of the traditional standard-dose-for-all approach.

Topics: Antineoplastic Agents; Bayes Theorem; Genetic Association Studies; Guanine Nucleotides; Humans; Immunosuppressive Agents; Mercaptopurine; Models, Biological; Neoplasms; Precision Medicine; Prodrugs; Thionucleotides

Mercaptopurine (6-MP), a critical component of acute lymphoblastic leukemia (ALL) therapy, is metabolized to 6-thioguanine (6-TGN) which is responsible for its anti-leukemic effect, and to 6-methylmercaptopurine nucleotides (6-MMPN/6-MMP) which can be hepatotoxic. Some patients preferentially metabolize 6-MP to 6-MMPN which may increase the risk of liver injury, reduce serum levels of 6-TGN and potentially increase the risk of relapse. The addition of allopurinol to oral 6-MP has been shown to optimize metabolism towards 6-TGN in patients with inflammatory bowel disease (IBD); however, this use has not been reported in patients undergoing treatment for ALL.

Topics: Adolescent; Allopurinol; Antimetabolites, Antineoplastic; Antineoplastic Combined Chemotherapy Protocols; Biotransformation; Chemical and Drug Induced Liver Injury; Child, Preschool; Drug Evaluation; Female; Guanine Nucleotides; Humans; Hyperbilirubinemia; Hypoxanthine Phosphoribosyltransferase; Maintenance Chemotherapy; Male; Mercaptopurine; Methotrexate; Methyltransferases; Neutropenia; Precursor Cell Lymphoblastic Leukemia-Lymphoma; Thionucleotides; Xanthine Oxidase

Symptomatic fasting hypoglycemia has been reported as an unusual side effect in patients with acute lymphoblastic leukemia (ALL) on maintenance therapy. We evaluated the relation of the red cell 6-mercaptopurine (6-MP) metabolite 6-methyl-mercaptopurine (6MMP) with hypoglycemia.. We retrospectively reviewed charts of three patients with ALL and symptomatic hypoglycemia while fasting who were noted to have high levels of 6MMP. All patients had an empiric trial of switching from evening to morning 6-MP administration, and two patients were subsequently switched to twice daily dosing. Patients also received complex carbohydrates at bedtime.. Switching 6-MP from evening to morning administration reduced 6MMP levels yet preserved adequate levels of the active metabolite red cell 6-thioguanine nucleotide (6TGN). All patients had decreased hypoglycemic events when changed from evening to morning dosing. Two patients showed a rebound in 6MMP levels with return of hypoglycemic symptoms. Both were then switched to twice daily 6-MP dosing with one having a decrease in 6MMP and hypoglycemic symptoms.. High levels of 6MMP are associated with symptomatic hypoglycemia which may be mitigated by switching to morning or twice daily 6-MP dose administration.

Topics: Antimetabolites, Antineoplastic; Antineoplastic Combined Chemotherapy Protocols; Biotransformation; Blood Glucose; Child; Child, Preschool; Dexamethasone; Dietary Carbohydrates; Drug Administration Schedule; Fasting; Female; Guanine Nucleotides; Humans; Hyperglycemia; Hypoglycemia; Hypoxanthine Phosphoribosyltransferase; Infant; Liver Glycogen; Maintenance Chemotherapy; Male; Mercaptopurine; Methotrexate; Methylation; Methyltransferases; Polymorphism, Genetic; Precursor B-Cell Lymphoblastic Leukemia-Lymphoma; Prodrugs; Retrospective Studies; Thionucleotides

To report the use of febuxostat in order to potentiate thiopurines' metabolism in a patient on azathioprine (AZA) therapy with low metabolite 6-thioguanine nucleotides (6-TGN) levels and elevated metabolite 6-methylmercaptopurine (6-MMP) levels.. A 44-year-old woman with a history of anti-signal recognition particle necrotizing myopathy was treated with AZA-allopurinol combination therapy. When she developed an atypical drug-induced hypersensitivity syndrome, allopurinol was replaced by the new xanthine oxidase (XO) inhibitor febuxostat, at a daily dose of 40 mg. Febuxostat-AZA combination was successful with 6-TGN reaching therapeutic levels while 6-MMP levels remained low. After 5 months, she developed similar manifestations that she had presented on AZA-allopurinol combination. Febuxostat and AZA were then stopped.. AZA and 6-MP are both inactive pro-drugs that undergo a complex metabolic transformation leading to active 6-TGN and potentially hepatotoxic 6-MMP. Some patients with unfavorable thiopurine metabolism might benefit from addition of XO inhibitor allopurinol in order to potentiate 6-TGN and reduce 6-MMP levels. It is likely that febuxostat, via its XO inhibition, would exhibit the same effect on thiopurines' metabolism.. It has been shown that low dose of febuxostat was able to prevent hypermethylation and to potentiate 6-TGN levels in an AZA-treated patient. Thus, febuxostat could be useful in optimizing thiopurines' metabolism, but more data are needed before this practice can be recommended. The mechanisms by which febuxostat optimizes thiopurines' metabolism remain to be confirmed. Also, the optimal dose of febuxostat for this use remains to be determined.

Topics: Adult; Azathioprine; Drug Interactions; Enzyme Inhibitors; Febuxostat; Female; Guanine Nucleotides; Humans; Immunosuppressive Agents; Mercaptopurine; Muscular Diseases; Thiazoles; Thionucleotides; Xanthine Oxidase

Thiopurines are used as a maintenance therapy in patients with ulcerative colitis (UC). For some patients the metabolism of thiopurines is unfavorable, leading to increased adverse effects, including hepatotoxicity. There are many reports in the adult literature concerning the manipulation of thiopurine metabolism with allopurinol; however, there is only 1 publication in this respect for pediatric UC. We present 3 pediatric cases of UC wherein the combination of allopurinol and low-dose 6-mercaptopurine allowed for shunting of thiopurine metabolites to a more favorable pattern. This intervention supported clinical remission in all, including one case poorly responsive to infliximab.

Topics: Adolescent; Allopurinol; Antimetabolites; Child; Colitis, Ulcerative; Drug Therapy, Combination; Female; Guanine Nucleotides; Humans; Immunosuppressive Agents; Male; Mercaptopurine; Thionucleotides

Thiopurines (azathioprine, 6-mercaptopurine) are a mainstay of treatment in Crohn disease (CD). Monitoring intracellular metabolite (6-thioguanine nucleotides [6-TGN] and 6-methylmercaptopurine [6-MMP]) levels can help optimize therapeutic efficacy and minimize potential toxicity. Determination of 6-MMP/6-TGN ratios may provide additional useful information, such as the identification of individuals with excessive thiopurine methyltransferase activity and disadvantageous 6-MMP overproduction. These patients are at increased risk of therapeutic failure and hepatotoxicity. The aim of the study was to evaluate the correlation of 6-MMP/6-TGN ratios with therapeutic efficacy and risk of hepatotoxicity in CD.. The present study was a single-center cross-sectional study including pediatric patients with CD studied prospectively with clinical and laboratory assessments along with serial measurements of 6-MMP and 6-TGN. Clinical response was determined using established clinical indices.. The study included 238 pediatric patients with CD with a total of 1648 evaluation points. The patients were in steroid-free remission at 59.1% of the evaluation points. 6-MMP/6-TGN ratios of 4 to 24 were protective against relapse (odds ratio [OR] 0.52, 95% confidence interval [CI] -0.39 to 0.69, P = 0.001). Hepatotoxicity was associated with high 6-MMP levels (>3919  pmol/8 × 10 red blood cell count: OR 7.65, 95% CI 3.7-15.9, P = 0.001) and high 6-MMP/6-TGN ratios (>24: OR 5.35, 95% CI -3.43 to 8.43, P = 0.001).. We observed significant associations between 6-MMP/6-TGN ratios and clinical response, and risk of hepatotoxicity. Our results suggest that determination of thiopurine metabolite ratios is a valuable tool for identification of patients at increased risk of therapeutic failure and hepatotoxicity.

Topics: Adolescent; Azathioprine; Chemical and Drug Induced Liver Injury; Child; Crohn Disease; Female; Guanine Nucleotides; Humans; Immunosuppressive Agents; Male; Mercaptopurine; Methyltransferases; Odds Ratio; Thionucleotides; Treatment Failure

Previous studies have reported no or only a very poor correlation between 6-methylmercaptopurine/6-thioguanine nucleotide (6-MeMPN/6-TGN) and azathioprine (AZA) dose in the treatment of inflammatory bowel disease (IBD). However, metabolite levels are often repeatedly measured yielding a hierarchical data structure that requires more appropriate data analysis.. This study explored the relationship between the weight-based dosage of AZA and metabolites levels in 86 pediatric IBD patients using multilevel analysis. Other covariates related to patient characteristics and treatment were evaluated.. This is the first study to demonstrate positive correlations between AZA dose and 6-TGN and 6-MeMPN levels and 6-MeMPN/6-TGN ratio (P < 0.001) in IBD children. Other novel predictors of metabolites were reported. Younger children exhibited lower 6-TGN and 6-MeMPN levels, probably suggesting age-related differences in metabolism and/or absorption of thiopurines. Coadministration of infliximab resulted in a significant increase in 6-TGN levels (P = 0.023). Moreover, alanine aminotransferase values positively correlated with 6-MeMPN levels (P = 0.032). The duration of AZA therapy, gender, and thiopurine methyltransferase activity were associated with metabolite levels. The wide interindividual variability in metabolite levels that accounted for 67.7%, 48.6%, and 49.4% of variance in the 6-TGN and 6-MeMPN levels and the ratio, respectively, were confirmed.. The reliable AZA dose-metabolites relationship is useful for clinicians to guide the dosing regimen to maximize clinical response and minimize side effects or to consider alternative therapies when patients have preferential production of the toxic 6-MeMPN. These results may be of potential interest for optimizing thiopurine therapy to achieve safe and efficacious AZA use in pediatric IBD patients.

Topics: Adolescent; Age Factors; Azathioprine; Child; Dose-Response Relationship, Drug; Female; Guanine Nucleotides; Humans; Immunosuppressive Agents; Inflammatory Bowel Diseases; Male; Mercaptopurine; Multilevel Analysis; Retrospective Studies; Thionucleotides

Thiopurine drugs (azathioprine, 6-mercaptopurine) show wide interindividual variability and a narrow therapeutic range thus making therapeutic monitoring of their active metabolite 6-thioguanine nucleotides (6-TGN) desirable. We improved the currently available laborious and complex methodology of therapeutic drug monitoring of 6-TGN and the metabolite 6-methylmercaptopurine (6-MMP) in washed erythrocytes (ery) based on a whole-blood method.. The analytes were hydrolyzed and extracted from 25-µL ethylenediaminetetraacetic acid-anticoagulated whole-blood spiked with isotope labeled 6-TG-C2N and 6-MMP-d3 internal standards. Chromatography was performed in 5.1 minutes on a C18 reverse phase column followed by detection via electrospray interface-coupled API 4000 mass spectrometer set up in the positive multiple reaction monitoring mode. The hemoglobin concentration was measured in 20 µL of the original sample (AHD575 method), and the results were standardized to 120 g/L of hemoglobin.. Calibration curves were linear with r > 0.999 (6-TGN and 6-MMP up to 10,000 pmol/0.2 mL). The limit of quantification was 30 pmol/0.2 mL for 6-TGN and 6-MMP. Intraassay and interassay imprecision was <7.5% at 3 tested levels for 6-TGN and 6-MMP, respectively. Method comparisons were as follows: Ery 6-TGN: y = 1.3x - 11 and ery 6-MMP y = 1.1x - 124.. The new method compares favorably with established ones, allowing for rapid single run determination of 6-TGN and 6-MMP from <50 µL of fresh or frozen whole blood. Linearity and limits of quantification cover the clinically relevant range. Variability during sample preparation and matrix effects are compensated by the use of isotope-labeled internal standards. The whole-blood method is hemoglobin standardized to avoid falsely low results in the case of anemia. The method correlates well with 6-TGN measured in washed erythrocytes, but it requires significantly less hands-on time. Preliminary therapeutic ranges for the most common indications of azathioprine and 6-MP are provided.

Topics: Calibration; Chromatography, High Pressure Liquid; Drug Monitoring; Erythrocytes; Guanine Nucleotides; Humans; Limit of Detection; Mercaptopurine; Tandem Mass Spectrometry; Thionucleotides; Time Factors

Azathioprine is prescribed as a corticosteroid-sparing agent for many inflammatory conditions, including refractory atopic dermatitis (AD). There are limited prospective data on its appropriate use and monitoring for children with AD.. This study was designed to assess clinical response to azathioprine, determine the necessity for repeated measurement of thiopurine methyltransferase (TPMT) activity during treatment, and test the utility of measuring levels of the metabolites 6-thioguanine nucleotide and 6-methylmercaptopurine.. Twelve children with severe, recalcitrant AD were treated with oral azathioprine and followed prospectively. Disease severity was determined by the SCORing AD index. Baseline TPMT activity was measured and this was repeated along with 6-thioguanine nucleotide and 6-methylmercaptopurine measurement at times of stable improvement, inadequate response, or change in response.. Azathioprine therapy was associated with clinical improvement in all but 1 patient. There were few adverse effects. Three patients showed a significant change in TPMT activity during treatment: 2 had a mild decrease and 1 demonstrated enzyme inducibility with an increase from the intermediate to the normal activity range. These changes, but not 6-thioguanine nucleotide or 6-methylmercaptopurine levels, inversely correlated with the clinical response to therapy.. Small sample size is a limitation.. Azathioprine can be of benefit in the treatment of recalcitrant pediatric AD. Repeat assessment of TPMT activity may be helpful for evaluation of nonresponse or change in response and warrants further study. In contrast, measurement of thiopurine metabolites during treatment was not clinically useful.

Topics: Administration, Oral; Adolescent; Azathioprine; Child; Child, Preschool; Dermatitis, Atopic; Female; Guanine Nucleotides; Humans; Immunosuppressive Agents; Male; Mercaptopurine; Methyltransferases; Prospective Studies; Severity of Illness Index; Thionucleotides

The thiopurines azathioprine and 6-mercaptopurine are recommended for maintenance of remission in inflammatory bowel disease (IBD). Measurement of concentrations of the metabolites 6-thioguanine nucleotide and 6-methylmercaptopurine helps delineate interindividual variation in metabolism that may underlie variability in efficacy and toxicity.. We aimed to perform a retrospective observational study to determine the utility of thiopurine metabolite testing following its introduction into South Australia.. All patients having thiopurine metabolite tests done at Flinders Medical Centre between November 2008 and January 2010 were identified. Case notes of patients with testing done in the context of treatment for IBD were interrogated to determine the reason for testing, clinical context and outcome.. One hundred and fifty-one patients were identified with thiopurine metabolite testing for IBD with 157 testing episodes. Eighty (51.0%) had testing done for flare or inefficacy, 18 (11.5%) for adverse effects, 5 (3.2%) for a combination of inefficacy and adverse effects, and 54 (34.4%) for routine or other reasons. Testing was followed by improved outcomes of increased efficacy, reduced toxicity or change to alternative therapy in 55.0% of the inefficacy/flare group, 27.8% of the suspected adverse reaction group, 60.0% of the combination group, and 13.0% of the routine/other group. Allopurinol was used as cotherapy in 16 patients and led to marked improvements in metabolite concentrations.. Thiopurine metabolite testing has quickly become established in South Australia. When used for inefficacy or adverse effects, it often leads to improved outcomes. Prospective studies are needed to determine whether routine testing to guide dosing is of benefit.

Topics: Adult; Biomarkers; Disease Management; Female; Guanine Nucleotides; Humans; Inflammatory Bowel Diseases; Male; Mercaptopurine; Middle Aged; Retrospective Studies; Thionucleotides

Thiopurine therapy is often discontinued in inflammatory bowel disease (IBD) patients. The xanthine oxidase (XO) inhibitor allopurinol has previously shown to enhance thiopurine efficacy and to prevent adverse reactions, the mechanism of this beneficial interaction is not completely clarified. The aim of this study is to observe possible effects of allopurinol and low-dose thiopurine combination therapy on the activity of three pivotal thiopurine metabolizing enzymes.. A prospective study of IBD patients failing thiopurine therapy due to a skewed thiopurine metabolism was performed. Patients were treated with allopurinol and azathioprine or mercaptopurine. Xanthine oxidase, hypoxanthine-guanine phosphoribosyl transferase (HGPRT) and thiopurine S-methyl transferase (TPMT) activities, and thiopurine metabolites concentrations were measured during thiopurine monotherapy, and after 4 and 12 weeks of combination therapy.. Of fifteen IBD patients, XO activity decreased from 0.18 (IQR 0.08-0.3) during thiopurine monotherapy to 0.14 (IQR 0.06-0.2) and 0.11 (IQR 0.06-0.2; p=0.008) mU/hour/ml at 4 and 12 weeks, respectively. HGPRT activity increased from 150 (IQR 114-176) to 180 (IQR 135-213) and 204 nmol/(h×mg protein) (IQR 173-213; p=0.013). TPMT activity seemed not to be affected. 6-Thioguanine nucleotide concentrations increased from 138 (IQR 119-188) to 235 (223-304) and to 265 pmol/8×10^8 (IQR 188-344), whereas 6-methyl mercaptopurine ribonucleotides concentrations decreased from 13230 (IQR 7130-17420) to 690 (IQR 378-1325) and 540 (IQR 240-790) pmol/8×10^8 at 4 and 12 weeks of combination therapy (both p<0.001).. Allopurinol and thiopurine combination-therapy seems to increase HGPRT and decrease XO activity in IBD patients, which at least in part may explain the observed changes in thiopurine metabolite concentrations.

Topics: Adult; Allopurinol; Azathioprine; Drug Therapy, Combination; Enzyme Activation; Enzyme Inhibitors; Fatigue; Female; Guanine Nucleotides; Humans; Hypoxanthine Phosphoribosyltransferase; Immunosuppressive Agents; Inflammatory Bowel Diseases; Male; Mercaptopurine; Methyltransferases; Middle Aged; Nausea; Prospective Studies; Thioinosine; Thionucleotides; Xanthine Oxidase

Thiopurine therapy effectively maintains remission in inflammatory bowel disease. However, many patients are unable to achieve optimum benefits from azathioprine or 6-mercaptopurine because of undesirable metabolism related to high thiopurine methyltransferase (TPMT) activity characterized by hepatic transaminitis secondary to increased 6-methylmercaptopurine (6-MMP) production and reduced levels of therapeutic 6-thioguanine nucleotide (6-TGN). Allopurinol can optimize this skewed metabolism. We discuss two brothers who were both diagnosed with ulcerative colitis (UC). Their disease remained active despite oral and topical mesalamines. Steroids followed by 6-mercaptopurine (MP) were unsuccessfully introduced for both patients and both were found to have high 6-MMP and low 6-TGN levels, despite normal TMPT enzyme activity, accompanied by transaminitis. Allopurinol was introduced in combination with MP dose reduction. For both brothers addition of allopurinol was associated with successful remission and optimized MP metabolites. These siblings with active UC illustrate that skewed thiopurine metabolism may occur despite normal TPMT enzyme activity and can lead to adverse events in the absence of disease control. We confirm previous data showing that addition of allopurinol can reverse this skewed metabolism, and reduce both hepatotoxicity and disease activity, but we now also introduce the concept of a family history of preferential MP metabolism as a clue to effective management for other family members.

Topics: Allopurinol; Antimetabolites; Colitis, Ulcerative; Dose-Response Relationship, Drug; Drug Therapy, Combination; Guanine Nucleotides; Humans; Liver; Male; Mercaptopurine; Methyltransferases; Steroids; Thionucleotides; Treatment Outcome; Young Adult

The prodrugs azathioprine and 6-mercaptopurine, which are well-established anticancer and immunosuppressive agents, are extensively metabolized by activating and inactivating enzymes. Whereas the 6-thioguanine nucleotides (TGN) are currently being considered as major active metabolites, methylthioinosine nucleotides seem to contribute to the cytotoxic effect as well. Thiopurine-related adverse drug reactions and thiopurine failure are frequent. Thus, therapeutic monitoring of TGN and methylthioinosine derivatives has been suggested to improve thiopurine therapy, however with limited success. To elucidate systematically underlying molecular mechanisms as potential explanation for interindividual variability of thiopurine response, we developed a novel highly specific and sensitive liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for simultaneous quantitation of eleven mono-, di-, and triphosphates of thioguanosine, methylthioinosine, methylthioguanosine, and thioinosine. Using stable isotope-labeled analogues as internal standards obtained by chemical synthesis, an intra- and interassay variability below 8% and an accuracy of 92% to 107% were achieved in spiked quality control samples with known standards. All eleven metabolites could be determined in red blood cells from patients with inflammatory bowel diseases and long-term azathioprine therapy. Thus, our novel method opens a new avenue for the understanding of the thiopurine metabolism by quantitation of all important thiopurine nucleotide metabolites in one run.

Topics: Azathioprine; Chromatography, High Pressure Liquid; Crohn Disease; Erythrocytes; Guanine Nucleotides; Humans; Immunosuppressive Agents; Isotope Labeling; Mercaptopurine; Methylthioinosine; Tandem Mass Spectrometry; Thionucleotides

Up to 20% of patients on thiopurine therapy fail to achieve adequate drug response. Many of these patients preferentially produce the toxic 6-methylmercaptopurine metabolites (6-MMP) rather than the active 6-thioguanine nucleotides (6-TGN) resulting in a high 6-MMP/6-TGN ratio (>20) and increased risk of hepatotoxicity.. To determine the prevalence of preferential 6-MMP producers and define the relationships between 6-TGN, 6-MMP and thiopurine methyltransferase (TPMT).. The database of 6-TGN, 6-MMP and TPMT measurements from patients throughout New Zealand was used to calculate patients' 6-MMP/6-TGN ratios and identify those with high (>20) or normal ratio (≤20).The TPMT enzyme activity was compared amongst the groups.. Of 1879 patients with TPMT, 6-TGN and 6-MMP results, 349 (19%) had a 6-MMP/6-TGN ratio >20. The mean TPMT enzyme activity was slightly lower for those with a 6-MMP/6-TGN ratio ≤20 vs. >20, which achieved statistical significance (12.2 vs. 13.2; P < 0.001). However, the distributions of TPMT enzyme activity were similar, with 97% of TPMT results falling between 5.0 and 17.6 IU/mL for both groups. In all, 17% of those with 6-MMP/6-TGN ratio ≤20 were intermediate TPMT metabolisers (TPMT 5.0-9.2 IU/mL) vs. 7% in those with a ratio >20.. In this patient population with measured 6-MMP/6-TGN ratios, 19% of patients were preferential 6-MMP producers. The results show that high TPMT enzyme activity is not the major reason for preferential 6-MMP production in most patients with a high metabolite ratio. This suggests that there are one or more important alternative mechanisms for preferentially producing 6-MMP.

Topics: Antimetabolites, Antineoplastic; Azathioprine; Chromatography, High Pressure Liquid; Drug Resistance; Erythrocyte Membrane; Female; Genotype; Guanine Nucleotides; Humans; Inflammatory Bowel Diseases; Male; Mercaptopurine; Methyltransferases; Statistics as Topic; Thioguanine; Thionucleotides

To investigate the potential influence of 5-aminosalicylic acid (5-ASA) on the induction of myelotoxicity during thiopurine therapy in inflammatory bowel disease patients.. (a) The retrospective study included inflammatory bowel disease patients treated with azathioprine (AZA)/6-mercaptopurine (6-MP). Thiopurine methyltransferase (TPMT) activity and 6-thioguanine nucleotide (6-TGN) levels were detected at stable medication points. (b) The prospective study was performed in active disease patients: 4 weeks of AZA 50 mg/day followed by concomitant 5-ASA 3 g/day for another 4 weeks. 6-TGN was analyzed at weeks 4 and 8.. (a) Of the 139 retrospective study patients, 45 were on AZA/6-MP+5-ASA and 94 on AZA/6-MP alone. The myelotoxicity rates were 47 and 16%, respectively. Multivariates regression analysis indicated that the administration of concomitant 5-ASA was the only risk factor associated with myelotoxicity (odds ratio=3.45, 95% confidence interval 1.31-9.04, P=0.01). (b) Thiopurine methyltransferase activity was not significantly different between patients on AZA/6-MP+5-ASA and patients on AZA/6-MP alone (P=0.78). (c) 6-TGN levels were significantly higher in samples on AZA/6-MP+5-ASA than those on AZA/6-MP (P=0.003) alone. (d) Sixteen patients completed the prospective study. After 4 weeks on AZA 50 mg/day, 6-TGN levels of 13 patients were less than 230 pmol/8×10 RBC. After another 4 weeks' cotreatment with mesalazine 3 g/day, 12 patients had 6-TGN levels at least 230 pmol/8×10 RBC, five patients had 6-TGN levels at least 420 pmol/8×10 RBC, and two of these five patients developed myelotoxicity.. The risk of thiopurine-induced myelotoxicity markedly increases in patients treated with combined 5-ASA and 2 mg/kg/day AZA therapy, which may be correlated to the increase in 6-TGN. 50 mg daily AZA when concomitant 5-ASA might help maintain an effective 6-TGN level without increasing the risk of myelotoxicity.

Topics: Adolescent; Adult; Aged; Anti-Inflammatory Agents, Non-Steroidal; Asian People; Azathioprine; Bone Marrow; Child; Child, Preschool; Cohort Studies; Drug Therapy, Combination; Female; Guanine Nucleotides; Humans; Immunosuppressive Agents; Infant; Inflammatory Bowel Diseases; Male; Mercaptopurine; Mesalamine; Methyltransferases; Middle Aged; Prospective Studies; Retrospective Studies; Thionucleotides; Young Adult

Thiopurine S-methyltransferase (TPMT) is an excellent example of an enzyme whose pharmacogenetic polymorphisms affect efficacy and toxicity of a drug. The association between TPMT activity and thiopurine-related myelosuppression is well recognized. To study the significance of TPMT deficiency in thiopurine metabolism and immunosuppressive activity in vitro, we established RNA interference-based TPMT knockdown (kd) in a Jurkat cell line.. In Jurkat TPMT kd cells, TPMT expression was reduced to 73% at the RNA level and 83% at the protein level. TPMT kd cells were more sensitive to 6-mercaptopurine (6-MP) (10 μmol/L) and 6-thioguanine (6-TG) (8 μmol/L) than wild-type (wt) cells, (32% versus 20%) and (18% versus 9%), respectively. Both Jurkat wt and kd cells were more sensitive to 6-TG-induced apoptosis than to 6-MP. 6-TG activity was also more affected by TPMT levels than was 6-MP as reflected by IC60, concentrations that is, 6-MP [4.6 μmol/L (wt) and 4.7 μmol/L (kd)], 6-TG [2.7 μmol/L (wt) and 0.8 μmol/L (kd)]. IC60 concentrations induced significant apoptosis in both Jurkat wt and kd cells (257%, versus 314%) with 6-MP and (323% versus 306%) with 6-TG, respectively. At IC60 (6-MP) 6-thioguanine nucleotides (6-TGN) accumulation in cells was 518 versus 447 pmol/million cells in wt and kd cells, respectively. On the other hand 6-TGN accumulation at IC60 (6-TG) was 477 versus 570 pmol/million cells in wt and kd cells, respectively. 6-Methylated mercaptopurine (6-MeMP) concentrations were more affected than 6-TGN by TPMT kd (194 versus 10 pmol/million cells) in wt and kd cells, respectively.. We conclude that TPMT kd cells are an appropriate in vitro model to investigate the significance of TPMT deficiency with thiopurine therapy and could be helpful in understanding possible clinical consequences of TPMT polymorphism.

Topics: Apoptosis; Cell Line, Tumor; Drug Hypersensitivity; Gene Knockdown Techniques; Guanine Nucleotides; Humans; Immune Tolerance; Jurkat Cells; Mercaptopurine; Methyltransferases; Polymorphism, Genetic; Purine-Pyrimidine Metabolism, Inborn Errors; T-Lymphocytes; Thioguanine; Thionucleotides

Allopurinol potentiates azathioprine and 6-mercaptopurine (6-MP) by increasing 6-thioguanine nucleotide (6-TGN) metabolite concentrations. The outcome might also be improved by adding allopurinol in individuals who preferentially produce 6-methylmercaptopurine nucleotides (6-MMPN), rather than 6-TGN. The aim of the present study was to investigate the effect of allopurinol on concentrations of 6-MMPN and 6-TGN in individuals with a high ratio of these metabolites (>20), which is indicative of a poor thiopurine response.. Sixteen individuals were identified who were taking azathioprine or 6-MP, and were commenced on allopurinol to improve a high 6-MMPN:TGN ratio. Metabolite concentrations were compared before and after commencing allopurinol, and markers of disease control were compared.. The addition of 100-300 mg allopurinol daily and thiopurine dose reduction (17-50% of the original dose) resulted in a reduction of the median (and range) 6-MMPN concentration, from 11,643 (3,365-27,832) to 221 (55-844) pmol/8×10(8) red blood cells (RBC; P=0.0005), increased 6-TGN from 162 (125-300) to 332 (135-923) pmol/8×10(8) RBC (P=0.0005), and reduced the 6-MMPN:6-TGN ratio from 63 (12-199) to 1 (0.1-4.5) (P=0.0005). There was a significant reduction in steroid dose requirements at 12 months (P=0.04) and trends for improvement in other markers of disease control. One patient developed red cell aplasia that resolved upon stopping azathioprine and allopurinol.. In those with a high 6-MMPN:6-TGN ratio (>20), response to thiopurine treatment might be improved by the addition of allopurinol, together with a reduced thiopurine dose and close hematological monitoring.

Topics: Adult; Allopurinol; Anti-Inflammatory Agents; Azathioprine; Biotransformation; Drug Therapy, Combination; Enzyme Inhibitors; Erythrocyte Count; Female; Gastrointestinal Agents; Guanine Nucleotides; Humans; Inflammatory Bowel Diseases; Male; Mercaptopurine; Middle Aged; New Zealand; Red-Cell Aplasia, Pure; Retrospective Studies; Steroids; Thionucleotides; Time Factors; Treatment Outcome; Xanthine Oxidase

Our study showed that a combination of 6-thiopurine (6-TP) drugs and a redox agent effectively inhibits the motility of SUM cells derived from human inflammatory breast cancer (IBC) cells and RhoC-overexpressed mammary epithelium cells. This 6-TP-mediated inhibition of cell motility occurs because the treated 6-TPs target and inactivate RhoC. A molecular mechanism for inactivation by the 6-TP-mediated RhoC is proposed by which treated TPs are converted in cells into 6-thioguanosine phosphate (6-TGNP). This 6-TGNP in turn reacts with the Cys(20) side chain of the redox-sensitive GXXXCGK(S/T)C motif of RhoC to produce a 6-TGNP-RhoC disulfide adduct. A redox agent synergistically enhances the formation process of this disulfide. The adduct that is formed impedes RhoC guanine nucleotide exchange, which populates an inactive RhoC. Our results suggest that 6-TGNP can also react with the redox-sensitive GXXXCGK(S/T)C and GXXXXGK(S/T)C motif of RhoA and Rac, respectively, to produce a 6-TGNP-RhoA and 6-TGNP-Rac disulfide adduct. However, given that RhoC has been shown to be overexpressed in ∼90% of IBC lesions, the populated RhoC but not other Rho proteins is likely to be a primary target for 6-TPs and a redox agent to terminate the metastasis of IBC.

Topics: Breast Neoplasms; Cell Line, Tumor; Cell Migration Inhibition; Crystallography, X-Ray; Drug Delivery Systems; Drug Synergism; Female; Gene Silencing; Guanine Nucleotides; Humans; Inflammation Mediators; Mercaptopurine; Neoplasm Invasiveness; Oxidation-Reduction; Prodrugs; rho GTP-Binding Proteins; rhoC GTP-Binding Protein; Thionucleotides

Low thiopurine-methyl-transferase (TPMT) activity and high 6-thioguanine-nucleotide (6TGN) concentrations have been linked to therapeutic success in inflammatory bowel disease patients treated with thiopurines; however, this has not been implemented in clinical practice.. To identify a therapeutic threshold value for TPMT or 6TGN concentrations, and their capability to predict treatment safety and efficacy.. Prospective multicentre study including steroid-resistant/dependent patients starting thiopurines. The TPMT activity was determined at inclusion (>5 U/mL required). Azathioprine metabolites [6TGN, 6-methyl-mercaptopurine ribonucleotides (6MMP), and 6TGN/6MMP and 6TGN/TPMT ratios] were periodically monitored during steroid tapering and after withdrawal for 6 months or until a new flare occurred.. A total of 113 patients were analysed (62% clinical response). Areas under the receiver operating characteristic (ROC) curve (AUC) relating clinical response and metabolite levels at 2, 4 and 6 months after steroid withdrawal were less than 0.7. The AUCs relating final response and initial TPMT activity or metabolite concentrations at 2, 4, 8 and 16 weeks after starting thiopurines were less than 0.7. No cut-off point with worthwhile sensitivity/specificity was found. Eight (7%) patients developed thiopurine-related toxicity that could not be linked to TPMT activity or 6TGN levels.. Our results do not support determination of TPMT activity or 6TGN concentrations to predict treatment outcome, and no useful serum metabolites threshold value to adjust the drug's dose was identified.

Topics: Adolescent; Adult; Aged; Area Under Curve; Azathioprine; Biomarkers; Dose-Response Relationship, Drug; Female; Guanine Nucleotides; Humans; Immunosuppressive Agents; Inflammatory Bowel Diseases; Male; Mercaptopurine; Methyltransferases; Middle Aged; Prospective Studies; ROC Curve; Thionucleotides; Treatment Outcome; Young Adult

Clinical response to thiopurine medication is related to the concentration of its metabolites. Proxy measures are traditionally used to assess dose adequacy. We present our experience of using tioguanine (previously known/formerly referred to as thioguanine) metabolite measurements in paediatric patients and evaluate their effect on clinical practice.. To report our experience of using tioguanine metabolite measurements in paediatric patients and to evaluate their effects on clinical practice.. The 6-tioguanine nucleotide (6-TGN) and 6-methylmercaptopurine (6-MMP) were measured in children prescribed thiopurine medication for at least 3 months. Data were collected on thiopurine methyl transferase (TPMT) genotype, drug dose, laboratory indices and management changes. Therapeutic 6-TGN levels were defined as 235-400 pmol/8 × 10(8) RBCs. Seventy individuals (30 males) with a median age of 15 years. Underlying diagnoses were 'IBD' (68/70) and two cases of eosinophilic colitis. Sixty-three were treated with azathioprine and seven with mercaptopurine. A total of 103 separate measurements were made.. On initial measurement, 68% of patients had 6-TGN levels outside therapeutic levels despite standard thiopurine dosing. Initial 6-TGN levels were significantly higher in patients with TPMT mutations. Toxicity occurred in seven cases. The 6-TGN levels were significantly higher in those with signs of marrow toxicity. The 6-TGN level correlated with WBC, leukocyte count, mean corpuscular volume (MCV) and ΔMCV; however, the ability of each of these to predict therapeutic 6-TGN levels was poor. After initial measurement, management was changed in 25/70 cases (36%).. 6-TGN levels were therapeutic in a minority of those patients who were tested. Proxy measures perform poorly in predicting therapeutic 6-TGN levels. Measuring thiopurine metabolites is useful for dosage adjustment in children, and for the detection of potential toxicity.

Topics: Adolescent; Azathioprine; Biomarkers; Chromatography, High Pressure Liquid; Dose-Response Relationship, Drug; Female; Guanine Nucleotides; Humans; Immunosuppressive Agents; Inflammatory Bowel Diseases; Male; Mercaptopurine; Methyltransferases; Retrospective Studies; Thionucleotides; Time Factors; Treatment Outcome

Levels of the thiopurine metabolites 6-thioguanine nucleotide (6-TGN) and 6-methylmercaptopurine commonly are monitored during thiopurine therapy for inflammatory bowel disease despite this test's high cost and poor prediction of clinical response (sensitivity, 62%; specificity, 72%). We investigated whether patterns in common laboratory parameters might be used to identify appropriate immunologic responses to thiopurine and whether they are more accurate than measurements of thiopurine metabolites in identifying patients who respond to therapy.. We identified 774 patients with inflammatory bowel disease on thiopurine therapy using metabolite and standard laboratory tests over a 24-hour time period. Machine learning algorithms were developed using laboratory values and age in a random training set of 70% of the cases; these algorithms were tested in the remaining 30% of the cases.. A random forest algorithm was developed based on laboratory and age data; it differentiated clinical responders from nonresponders in the test set with an area under the receiver operating characteristic (AUROC) curve of 0.856. In contrast, 6-TGN levels differentiated clinical responders from nonresponders with an AUROC of 0.594 (P < .001). Algorithms developed to identify thiopurine nonadherence (AUROC, 0.813) and thiopurine shunters (AUROC, 0.797) were accurate.. Algorithms that use age and laboratory values can differentiate clinical response, nonadherence, and shunting of thiopurine metabolism among patients who take thiopurines. This approach was less costly and more accurate than 6-TGN metabolite measurements in predicting clinical response. If validated, this approach would provide a low-cost, rapid alternative to metabolite measurements for monitoring thiopurine use.

Topics: Adolescent; Adult; Aged; Aged, 80 and over; Algorithms; Artificial Intelligence; Child; Child, Preschool; Drug Monitoring; Female; Guanine Nucleotides; Humans; Immunologic Factors; Inflammatory Bowel Diseases; Male; Mercaptopurine; Middle Aged; Thionucleotides; Young Adult

Topics: DNA; Guanine Nucleotides; Humans; Mercaptopurine; Prodrugs; Signal Transduction; Thionucleotides

To explore the DNA incorporation of 6-thioguanine nucleotide levels (DNA-6TGN) during 6-mercaptopurine (6MP) therapy of childhood acute lymphoblastic leukaemia (ALL) and non-Hodgkin lymphoma (NHL) and its relation to erythrocyte levels of their metabolites: 6-thioguanine-nucleotides (E-6TGN), methylated metabolites (E-MeMP), Methotrexate polyglutamates (E-MTX), and to thiopurine methyltransferase activity (TPMT).. We studied these metabolites in 229 blood samples from 18 children with ALL (N = 16) or NHL (N = 2) on 6MP/Methotrexate maintenance therapy.. DNA-6TGN levels were significantly correlated to E-6TGN (r (p) = 0.66, p = 0.003) with a trend to reach a plateau at high E-6TGN levels. To explore the relative DNA incorporation of 6TGN in relation to cytosol 6TGN levels, a DNA-6TGN index was calculated as DNA-6TGN/E-6TGN. The DNA-6TGN index was inversely correlated to E-6TGN (r (p) = -0.58, p = 0.012), which implies that with increasing levels of E-6TGN relatively less 6TGN are incorporated into DNA. E-MeMP levels were correlated to the DNA-TGN index (r (p) = 0.60, p = 0.008), indicating that high levels of MeMP result in enhanced DNA-6TGN incorporation, possibly due to inhibition of purine de novo synthesis, mediated by some of the methylated 6MP metabolites.. DNA-6TGN may prove to be a more relevant pharmacokinetic parameter for monitoring 6MP treatment intensity than the previously used erythrocyte 6MP metabolites levels. Prospective clinical trials are needed to evaluate the usefulness of DNA-6TGN for individual dose adjustments.

Topics: Antineoplastic Combined Chemotherapy Protocols; Child; Child, Preschool; Cytosol; DNA; Drug Monitoring; Erythrocytes; Female; Guanine Nucleotides; Humans; Infant; Lymphoma, Non-Hodgkin; Male; Mercaptopurine; Methotrexate; Precursor Cell Lymphoblastic Leukemia-Lymphoma; Purines; Thionucleotides

6-Mercaptopurine (6-MP) and azathioprine (AZA) are widely used as maintenance therapy in children with inflammatory bowel disease (IBD). However, proper 6-thioguanine nucleotide (6-TGN) concentrations in Japanese children with IBD have not been reported.. This retrospective review examines 32 ulcerative colitis (UC) patients and 19 Crohn's disease (CD) patients (12.87 ± 3.56 years) who required 6-MP or AZA to maintain disease remission. All patients were treated with 6-MP or AZA for at least 3 weeks prior to this study in addition to previous treatment. 6-MP dose, 6-TGN levels, assayed by high-performance liquid chromatography, as well as laboratory data were evaluated.. Thirty-five children were successfully kept in remission with 6-MP and AZA therapy after weaning off corticosteroids. Overall, 123 measurements (59 active disease, 64 in remission) were analyzed. The mean 6-TGN concentration of the entire study population was 499.61 ± 249.35 pmol/8 × 10(8) red blood cell. The mean 6-MP dose in patients with active disease (0.910 ± 0.326 mg/kg per day) was significantly higher than for patients in remission (0.749 ± 0.225) (P = 0.0016). A significant inverse correlation was found between white blood cell counts and 6-TGN concentrations (r = 0.275, P < 0.002). Two patients experienced leukopenia with alopecia, and four transiently experienced increased serum levels of pancreatic enzymes, although no thiopurine S-methyl transferase mutations were confirmed.. The doses of 6-MP or AZA needed to maintain remission in Japanese children with IBD are lower than those reported in Western countries. However, 6-TGN concentrations in this population are higher than previously reported.

Topics: Adolescent; Azathioprine; Biomarkers, Pharmacological; Child; Chromatography, High Pressure Liquid; Colitis, Ulcerative; Crohn Disease; Dose-Response Relationship, Drug; Drug Monitoring; Female; Follow-Up Studies; Guanine Nucleotides; Humans; Immunosuppressive Agents; Inflammatory Bowel Diseases; Japan; Male; Mercaptopurine; Remission Induction; Retrospective Studies; Thionucleotides; Treatment Outcome

6-mercaptopurine (6-MP) is used for the induction and maintenance of remission of inflammatory bowel disease (IBD). 6-MP is converted into 6-methylmercaptopurine (6-MMP) or 6-thioguanine nucleotides (6-TGN) intracellularly. Treatment response in IBD patients correlates with 6-TGN levels. This study prospectively evaluated the effect of allopurinol on 6-MP metabolites in adult and pediatric IBD patients. Additionally, we quantified the prevalence of preferential metabolism towards 6-MMP through a retrospective analysis of IBD patients.. Twenty patients (10 adult; 10 pediatric) with evidence of preferential metabolism towards 6-MMP, (6-TGN<250 pmol/8×10⁸ RBCs and 6-MMP>5000 pmol/8×10⁸ RBCs) were prospectively treated with allopurinol 100 mg daily and up to 100 mg of 6-MP. 6-MP dose was adjusted after a 3-week metabolite measurement.. The median dose of 6-MP for adults decreased from 100mg daily (range: 37.5-150 mg) to 25mg daily (range: 12.5-50 mg). The median dose of 6-MP for pediatric patients decreased from 50 mg (range: 25-50 mg) to 10.7 mg (range: 10.7 to 21.4 mg). Mean 6-TGN levels in all subjects increased from 197.4 (± 59) to 284.8 (± 107) pmol/8×10⁸ RBCs (p=0.0005). Mean 6-MMP levels in all subjects decreased from a mean of 7719.8 (± 4716) to 404.8 (± 332) pmol/8×10⁸ RBCs (p=0.0004). There were no complications associated with allopurinol therapy. Eighty-eight (30.9%) of 285 IBD patients had evidence of preferential metabolism towards 6-MMP. The proportion of preferential metabolism was equal in adults and pediatric patients.. Our results indicate that the addition of allopurinol safely shifts metabolite production in both adult and pediatric IBD patients and that there is a high prevalence of preferential metabolism towards 6-MMP among IBD patients.

Topics: Adolescent; Adult; Aged; Aged, 80 and over; Allopurinol; Child; Child, Preschool; Drug Administration Schedule; Enzyme Inhibitors; Female; Guanine Nucleotides; Humans; Immunosuppressive Agents; Inflammatory Bowel Diseases; Male; Mercaptopurine; Methyltransferases; Middle Aged; Phenotype; Prevalence; Prospective Studies; Thionucleotides; Young Adult

Azathioprine is a thiopurine immunosuppressive antimetabolite used to chronically treat inflammatory bowel disease and autoimmune hepatitis. Azathioprine treatment is a long-term therapy and therefore it is at risk for non-adherence, which is considered an important determinant of treatment inefficacy. Measurement of 6-thioguanine and 6-methylmercaptopurine nucleotides has been recently suggested as a screener for non-adherence detection. We describe four young patients in which non-adherence to azathioprine therapy was detected only through the measurement of drug metabolite concentrations, and the criterion for non-adherence was undetectable metabolite levels. After the identification of non-adherence, patients and their families were approached and the importance of a correct drug administration was thoroughly enlightened and discussed; this allowed obtaining a full remission in all subjects. Our observations support the use of undetectable metabolite levels as indicators of non-adherence to therapy in azathioprine treated patients. The additional level of medical supervision given by this assay allows getting a better adherence to medical treatment, which results in an improvement in the response to therapy; these benefits may justify the costs associated with the assay.

Topics: Adolescent; Azathioprine; Child; Child, Preschool; Female; Guanine Nucleotides; Hepatitis, Autoimmune; Humans; Immunosuppressive Agents; Inflammatory Bowel Diseases; Male; Medication Adherence; Mercaptopurine; Thionucleotides; Young Adult

Azathioprine (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 nucleotides (6-TGN) accumulation. The aim of this study was to evaluate the time to steady state of 6-TGN concentration in red blood cells.. The 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.. The 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.. The time to steady state of 6-TGN erythrocyte concentration is significantly shorter than would expected according to clinical observation describe earlier.

Topics: Adolescent; Azathioprine; Child; Child, Preschool; Female; Genotype; Guanine Nucleotides; Humans; Immunosuppressive Agents; Inflammatory Bowel Diseases; Male; Mercaptopurine; Methyltransferases; Prospective Studies; Severity of Illness Index; Thionucleotides; Treatment Outcome

To examine the allelic variation of three enzymes involved in 6-mercaptopurine/azathioprine (6-MP/AZA) metabolism and evaluate the influence of these polymorphisms on toxicity, haematological parameters and metabolite levels in patients with acute lymphoblastic leukaemia (ALL) or inflammatory bowel disease (IBD).. Clinical data and blood samples were collected from 19 ALL paediatric patients and 35 IBD patients who were receiving 6-MP/AZA therapy. All patients were screened for seven genetic polymorphisms in three enzymes involved in mercaptopurine metabolism [xanthine oxidase, inosine triphosphatase (C94-->A and IVS2+21A-->C) and thiopurine methyltransferase]. Erythrocyte and plasma metabolite concentrations were also determined. The associations between the various genotypes and myelotoxicity, haematological parameters and metabolite concentrations were determined.. Thiopurine methyltransferase variant alleles were associated with a preferential metabolism away from 6-methylmercaptopurine nucleotides (P = 0.008 in ALL patients, P = 0.038 in IBD patients) favouring 6-thioguanine nucleotides (6-TGNs) (P = 0.021 in ALL patients). Interestingly, carriers of inosine triphosphatase IVS2+21A-->C variants among ALL and IBD patients had significantly higher concentrations of the active cytotoxic metabolites, 6-TGNs (P = 0.008 in ALL patients, P = 0.047 in IBD patients). The study confirmed the association of thiopurine methyltransferase heterozygosity with leucopenia and neutropenia in ALL patients and reported a significant association between inosine triphosphatase IVS2+21A-->C variants with thrombocytopenia (P = 0.012). CONCLUSIONS; Pharmacogenetic polymorphisms in the 6-MP pathway may help identify patients at risk for associated toxicities and may serve as a guide for dose individualization.

Topics: Adolescent; Antimetabolites, Antineoplastic; Azathioprine; Child; Child, Preschool; Female; Gene Frequency; Genotype; Guanine Nucleotides; Humans; Inflammatory Bowel Diseases; Male; Mercaptopurine; Methyltransferases; Pharmacogenetics; Polymorphism, Genetic; Precursor Cell Lymphoblastic Leukemia-Lymphoma; Thionucleotides; Treatment Outcome

Azathioprine and its initial metabolite, 6-mercaptopurine (6-MP), are associated with high rates of treatment cessation due to toxicity or inadequate response. Individualization of thiopurine dose based on concentrations of the active 6-thioguanine nucleotide (6-TGN) metabolites can help improve outcomes with this class. Some individuals, however, preferentially metabolize thiopurine drugs to the potentially hepatotoxic 6-methylmercaptopurine nucleotide (6-MMPN) metabolites rather than the 6-TGNs. For these patients, escalation in thiopurine dose is not likely to increase 6-TGN concentrations sufficiently but may lead to a disproportionate increase in exposure to the 6-MMPNs. We present three cases in whom thiopurine dose escalation based on clinical status and low 6-TGN concentrations (100-262 pmol/8 x 10 RBC) resulted in severe hepatotoxicity (liver failure in two cases) associated with unrecognized extremely high 6-MMPN concentrations of 26,000-40,000 pmol/8 x 10 RBC. These cases illustrate a risk with thiopurine dose adjustment based on monitoring of 6-TGN metabolites without also monitoring 6-MMPN.

Topics: Adult; Azathioprine; Chemical and Drug Induced Liver Injury; Drug Administration Schedule; Drug Monitoring; Female; Guanine Nucleotides; Humans; Immunosuppressive Agents; Mercaptopurine; Middle Aged; Thionucleotides

Topics: Bias; Guanine Nucleotides; Humans; Inflammatory Bowel Diseases; Mercaptopurine; Prospective Studies; Thionucleotides

To examine the relationship between medication adherence and quality of life (QOL) in adolescent patients with inflammatory bowel disease (IBD) utilizing a multimethod adherence assessment approach.. Medication adherence in 36 adolescents with IBD was assessed via interviews, pill counts, and biological assays. QOL was assessed via patient and parent report. Pediatric gastroenterologists provided disease severity assessments.. Hierarchical multiple regression analyses revealed that adherence contributed significant variance to patient-reported QOL but not parent-reported QOL. Nonadherence to 6-MP/azathioprine was related to poorer patient-reported physical health QOL. Greater self-reported 5-ASA adherence was related to poorer overall psychological health QOL, and particularly social functioning QOL.. Results provide preliminary support for the negative effects of 6-MP/azathioprine nonadherence on QOL and an inverse relationship between 5-ASA adherence and QOL in this population. Adherence burden in patients and the utility of multimethod adherence assessment in research are discussed.

Topics: Adolescent; Anti-Inflammatory Agents; Azathioprine; Colitis, Ulcerative; Crohn Disease; Depression; Dose-Response Relationship, Drug; Drug Administration Schedule; Female; Guanine Nucleotides; Humans; Illness Behavior; Male; Medication Adherence; Mercaptopurine; Mesalamine; Quality of Life; Thionucleotides

We evaluated how our use of thiopurines was altered by determination of thiopurine methyltransferase (TPMT) level and drug dose adjustment guided by a 6-mercaptopurine metabolite assay. We further examined whether these resulted in better selection of the drug dose, improved control of disease, and decreased corticosteroid use in pediatric inflammatory bowel disease (IBD).. This is a retrospective review of 101 pediatric patients with IBD receiving a stable dose of azathioprine (AZA) for 4 months or longer. The study group (n = 64) consisted of patients who received AZA and had metabolite levels measured. The comparison group (n = 37) consisted of patients who were receiving AZA before the availability of metabolite measurement. The TPMT level was measured in study group patients before starting AZA.. Patients with normal TPMT level received a higher starting dose of AZA than in patients who were heterozygous for TPMT deficiency (1.7 vs 0.9 mg/[kg x d], P < 0.0001). Study group patients received a higher starting dose (1.6 vs 1.2 mg/[kg x d], P = 0.001) and a higher final dose of AZA (2.4 vs 1.7 mg/[kg x d], P < 0.0001) compared with patients in the comparison group. These patients also had more dose adjustments (0.8 vs 0.4 mg/kg, P < 0.002). The number of disease exacerbations per patient per year was 55% less in the study group (95% CI, 17%-76%, P < 0.0001). The study group patients received less prednisone (P < 0.0001) and had lower disease activity scores (P < 0.05). There was no difference between groups in infliximab use or surgery rate.. Azathioprine dose adjustment using a 6-mercaptopurine metabolite assay was associated with use of higher doses, improved control of disease and decreased corticosteroid use in pediatric patients with IBD.

Topics: Adolescent; Adult; Azathioprine; Child; Child, Preschool; Colitis, Ulcerative; Crohn Disease; Erythrocytes; Female; Guanine Nucleotides; Heterozygote; Humans; Infant; Inflammatory Bowel Diseases; Male; Mercaptopurine; Methyltransferases; Prednisone; Remission Induction; Retrospective Studies; Thionucleotides

Polymorphic thiopurine S-methyltransferase (TPMT) is a major determinant of thiopurine toxicity.. We extracted 6-thioguanine nucleotides (6-TGNs) and 6-methylmercaptopurine nucleotides (6-MMPNs) from erythrocytes with perchloric acid and converted them to 6-thioguanine (6-TG) and a 6-methylmercaptopurine (6-MMP) derivative during a 60-min acid hydrolysis step. The liquid chromatography system consisted of a C(18) column with an ammonium acetate-formic acid-acetonitrile buffer. 8-Bromoadenine was the internal standard. Analytes were measured with positive ionization and multiple reaction monitoring mode. With PCR-restriction fragment length polymorphism analysis and TaqMan allelic discrimination, common TPMT alleles (*1, *2, *3A, *3B, *3C) were determined in 31 792 individuals. We used perchloric acid extraction, acid hydrolysis, and HPLC with ultraviolet detection to measure erythrocyte 6-TG and 6-MMP nucleotide concentrations in 6189 patients with inflammatory bowel disease receiving azathioprine/6-mercaptopurine therapy.. Intra- and interday imprecision were <10% at low and high analyte concentrations. The conversion of 6-TG and 6-MMP nucleoside mono-, di-, and triphosphates was complete after hydrolysis. Allelic frequency for TPMT variant alleles ranged from 0.0063% (*3B) to 3.61% (*3A). Compared with wild types, TPMT heterozygotes had an 8.3-fold higher risk for 6-TGNs >450 pmol/8 x 10(8) erythrocytes (concentration associated with increased risk for leukopenia), but an 8.2-fold lower risk for 6-MMPNs >5700 pmol/8 x 10(8) erythrocytes (concentration associated with increased risk for hepatotoxicity).. The liquid chromatography-tandem mass spectrometry method can be applied to the routine monitoring of thiopurine therapy. The association between TPMT genotype and metabolite concentrations illustrates the utility of pharmacogenetics in the management of patients undergoing treatment with thiopurines.

Topics: Adult; Azathioprine; Chromatography, Liquid; Cohort Studies; Drug Therapy, Combination; Erythrocytes; Female; Genetic Variation; Guanine Nucleotides; Humans; Hydrolysis; Inflammatory Bowel Diseases; Male; Mass Spectrometry; Mercaptopurine; Methyltransferases; Purine Nucleotides; Thionucleotides

Azathioprine and 6-mercaptopurine (6-MP) are well established for the treatment of inflammatory bowel disease (IBD). Assessing thiopurine methyltransferase (TPMT) status has been recommended to reduce the risk of serious toxicity. Measuring red blood cell (RBC) 6-thioguanine nucleotide (6-TGN) concentrations has been recommended for dose adjustment.. To describe the results of measuring TPMT activity and genotype, and 6-TGN concentration in New Zealand.. Canterbury Health Laboratories provided these analyses for New Zealand. Those with low TPMT activity also underwent genotyping. All results were collated and analysed descriptively. 6-TGN concentrations were correlated with the dose of thiopurine when known.. TPMT enzyme activity (range 1-22 U/mL) from 574 patients showed a trimodal distribution. Genotyping results matched this distribution with only mild overlap between (*1/*1) homozygote and (*1/*3) heterozygote groups. One patient without TPMT measurement before therapy had life-threatening neutropenia and was later found to have (*3/*3) genotype. TPMT analysis probably prevented two further such cases. Of 884 6-TGN concentrations (range 0-1434 pmol/10(8) RBC), 41, 39 and 20% were within, below, and above the therapeutic range of 235-450 pmol/10(8) RBC, respectively. Leucopenia was seen in some patients with high 6-TGN. 6-MMP concentrations in 177 patients with low 6-TGN suggested non-compliance in 31, underdosing in 130, and preferential metabolism of 6-MP to 6-methylmercaptopurine in 16. There was poor correlation between azathioprine dose and 6-TGN concentration (r(2) = 0.002), supporting 6-TGN monitoring.. Measurement of TPMT enzyme activity and 6-TGN concentration has been well-integrated into clinical practice. These tests should reduce the risk of toxicity and improve efficacy with thiopurines in patients with IBD.

Topics: Azathioprine; Biomarkers; Erythrocytes; Genotype; Guanine Nucleotides; Humans; Immunosuppressive Agents; Inflammatory Bowel Diseases; Mercaptopurine; Methyltransferases; Thionucleotides

To study the distribution of the thiopurine methyltransferase (TPMT) genotype among azathioprine (Aza)-tolerant and -intolerant patients with various disorders, and to investigate a possible relationship with the Aza metabolite levels.. Forty-six Aza-tolerant and six Aza-intolerant patients had the TPMT genotype distribution determined using a polymerase chain reaction (PCR) assay and the forty-six Aza-tolerant patients had the Aza metabolite levels determined using a high-pressure liquid chromatography (HPLC) analysis.. One non-functional TPMT mutant allele was demonstrated in 2 of the 46 Aza-tolerant patients (4.4%) and one or two non-functional mutant alleles in 2 of the 6 Aza-intolerant patients (33.3%). Of the 4 patients, with one or two non-functional mutant alleles 2 (50%) were intolerant to Aza compared with 4 of the 48 patients (8.3%) with no mutations detected. The time to hepatotoxicity did not differ significantly between the 2 patients with one or two non-functional mutant alleles and the remaining 3 patients ( P=0.5). The TPMT genotype distribution differed slightly in the three different categories of disorders ( P=0.05). The median E-6-TGN level among the 2 TPMT heterozygous patients was 275 pmol/8x10(8) RBC (range 240-310), whereas the remaining 44 patients had a median E-6-TGN level of 110 pmol/8x10(8) RBC (range 0-440) ( P=0.07).. Although TPMT genotyping cannot be recommended on behalf of the present study, it is to be expected that half of the patients with one or two non-functional TPMT mutant alleles will develop Aza intolerance leading to withdrawal of therapy. Thus, clinicians may anticipate about 5% of the patients to develop intolerance to Aza therapy solely for that reason.

Topics: Adjuvants, Immunologic; Adult; Aged; Aged, 80 and over; Anti-Inflammatory Agents, Non-Steroidal; Azathioprine; Female; Genotype; Guanine Nucleotides; Humans; Male; Mercaptopurine; Methyltransferases; Middle Aged; Mutation; Polymerase Chain Reaction; Thionucleotides

Genetic polymorphism in thiopurine methyltransferase (TPMT) activity may influence clinical responsiveness to azathioprine (AZA) therapy. Our aim was to determine if the measurement of erythrocyte TPMT enzyme activity could be used to optimize clinical responsiveness to AZA therapy in patients with inflammatory bowel disease (IBD).. A total of 142 consecutive patients were studied. Forty-one patients (32 with Crohn's disease [CD] and 9 with ulcerative colitis [UC]) were enrolled in a 4-month prospective nonrandomized study with AZA, and 101 (65 with CD and 36 with UC) were on either maintenance AZA or 6-mercaptopurine (6-MP). Erythrocyte TPMT activity and AZA metabolite levels were measured blinded to the clinical response.. The response rate after 4 months of continuous AZA therapy was 69% (9/13) in those patients with below-average (12 U/mL blood (P < 0.001). Patients with TPMT activity 12 (218 +/- 28), despite similar mean (1.6 mg/kg/day) dosages of AZA (P < 0.001). By multivariate logistic regression analysis, patients with a TPMT level <15.3 U/mL blood were 6.2 times more likely to respond to AZA therapy. A 6-TGn level of >292 pmol/8 x 10(8) RBCs was associated with a positive predictive value of clinical response of 85.7%.. Patients with higher than average TPMT activity (>12) may remain refractory to conventional dosages of AZA, and may require high (>292) 6-TGn levels. Prospective, randomized, controlled trials are needed to determine whether prior TPMT phenotype testing can be used to adjust the dose of AZA effectively to improve clinical response time and rate.

Topics: Adult; Azathioprine; Colitis, Ulcerative; Crohn Disease; Erythrocytes; Female; Guanine Nucleotides; Humans; Immunosuppressive Agents; Logistic Models; Male; Mercaptopurine; Methyltransferases; Prospective Studies; Remission Induction; Thionucleotides; Treatment Outcome

The immunosuppressive effects of thiopurine drugs are mainly mediated through their intracellular metabolism into active 6-thioguanine nucleotide (6-TGN) metabolites, which are incorporated into DNA. Erythrocyte 6-TGN (E-6TGN) levels have been proposed as an instrument for monitoring treatment. The aim of the study was to use erythrocyte E-6TGN, methylated mercaptopurine (MeMP) metabolites, and thiopurine methyltransferase (TPMT) measurements in a clinical setting to determine the clinical outcome in relation to thiopurine metabolism.. Fifty-five adult patients with inflammatory bowel disease were included in a prospective study and followed for 6 months. Metabolite levels were measured and correlated to outcome and AZA/6-MP dose.. The E-6TGN level was significantly related to the TPMT genotype (P = 0.008). Patients in disease remission had higher E-6TGN levels than patients with disease activity both at baseline (P < 0.05) and after 6 months (P = 0.02). Active disease was more frequent among subjects with E-6TGN < or = 125 nmol/mmol Hb at baseline (P = 0.04), but not at 6 months. AZA/6-MP drug dose was positively correlated to E-MeMP levels (r = 0.48; P < 0.001) and E-MeMP/E-6TGN ratio (r = 0.41; P = 0.002). Dose changes were positively correlated with the changes in E-MeMP levels (P = 0.01) and E-MeMP/E-6TGN ratio (P = 0.03).. E-6TGN level was the only factor in this study related to disease activity, while there was no relationship between AZA/6-MP dose and E-6TGN levels. This finding illustrates the clinical usefulness of E-6TGN monitoring in the evaluation of treatment intensity.

Topics: Adolescent; Adult; Azathioprine; Drug Monitoring; Erythrocytes; Female; Genotype; Guanine Nucleotides; Humans; Immunosuppressive Agents; Inflammatory Bowel Diseases; Male; Mercaptopurine; Methyltransferases; Middle Aged; Sulfasalazine; Thionucleotides

Thiopurine drugs are commonly used immunosuppressants in the treatment of inflammatory bowel disease (IBD) as well as in autoimmune hepatitis (AIH), rheumatic diseases and in transplantation medicine. The relatively narrow therapeutic range requires useful therapy control. Therefore, the purpose of this study was to further investigate the rationale and usefulness of therapeutic drug monitoring in the surveillance of thiopurine drug therapy in Crohn's disease, ulcerative colitis and autoimmune hepatitis.. 6-Thioguanine nucleotide (TGN) and 6-methylmercaptopurine nucleotide (MMPN) levels were measured in 182 IBD patients and 18 AIH patients using HPLC-UV.. In our cohort of IBD patients, 18% had TGN levels < 235 pmol/8 x 10 red blood cells (RBC) (recommended range, 235-450 pmol/8 x 10 RBC), 41% of these patients were sent for drug failure. Twenty-four per cent of the IBD patients had TGN levels > 450 pmol/8 x 10 RBC, but only 27% of these experienced adverse effects. Fifty-nine per cent of the patients having drug failure had TGN levels in the recommended range and could therefore be classified as non-responders. In the AIH cohort 33% of the patients had TGN levels below the recommended range but showed clinical response to therapy. MMPN levels increased with the duration of treatment and could be useful for controlling compliance. There was 8.8% of IBD patients who were heterozygous for non-functional TPMT alleles.. TGN monitoring did not identify significant differences between patient groups but allowed the identification of non-responders from non-compliant patients and allowed the differentiation of mild side effects, such as malaise, from genuine toxicity caused by highly increased TGN levels.

Topics: Adolescent; Adult; Aged; Child; Child, Preschool; Chromatography, High Pressure Liquid; Cohort Studies; Colitis, Ulcerative; Crohn Disease; Drug Monitoring; Female; Guanine Nucleotides; Hepatitis, Autoimmune; Heterozygote; Humans; Immunosuppressive Agents; Inflammatory Bowel Diseases; Male; Mercaptopurine; Middle Aged; Purines; Thioguanine; Thionucleotides

Topics: Azathioprine; Child; Drug Monitoring; Guanine Nucleotides; Humans; Immunosuppressive Agents; Inflammatory Bowel Diseases; Mercaptopurine; Methylthioinosine; Methyltransferases; Polymorphism, Genetic; Thionucleotides

Topics: Azathioprine; Child; Drug Monitoring; Guanine Nucleotides; Humans; Immunosuppressive Agents; Inflammatory Bowel Diseases; Mercaptopurine; Methylthioinosine; Methyltransferases; Polymorphism, Genetic; Thionucleotides

Nineteen pediatric patients affected by acute lymphoblastic leukemia (ALL) were examined weekly with respect to 6-mercaptopurine nucleotide (6-MPN) and 6-thioguanine nucleotide (6-TGN) levels in erythrocytes during the course of maintenance treatment with 6-MP 50 mg/m2 per d and results were related to various parameters of bone marrow function to assess, in the same individual, the level of reliability of 6-MP metabolites in predicting a later change in peripheral blood cell counts. Median values for 6-MPN and 6-TGN were 57 and 200 pmol/8 x 10(8) erythrocytes, respectively, as measured by reversed-phase high-performance liquid chromatography (HPLC). 6-TGN levels in erythrocytes were inversely related with white blood cell count (r = -0.463, p < 0.0001, n = 361), absolute neutrophil count (r = -0.386, p < 0.0001, n = 347), erythrocyte (r = -0.354, p < 0.0001, n = 287), and platelet counts (r = -0.24, p < 0.0001, n = 319) in the majority of patients (n = 10-12), while no correlation was found for 6-MPN. In the remaining children, no evidence of correlation was demonstrated between 6-TGN levels and myelotoxicity. The results confirm the role of 6-TGN as the reference cytotoxic metabolite for evaluating the exposure to 6-MP and identifying treatment compliance in ALL children but indicate the limits of a follow-up based solely on metabolite levels and suggest that a more correct approach remains the double monitoring of 6-TGN and blood cell count with differential.

Topics: Adolescent; Antimetabolites, Antineoplastic; Bone Marrow; Child; Child, Preschool; Drug Monitoring; Erythrocytes; Female; Guanine Nucleotides; Humans; Male; Mercaptopurine; Precursor Cell Lymphoblastic Leukemia-Lymphoma; Thionucleotides

Topics: Adult; Azathioprine; DNA Methylation; Dose-Response Relationship, Drug; Erythrocytes; Female; Guanine Nucleotides; Heart-Lung Transplantation; Humans; Immunosuppressive Agents; Male; Mercaptopurine; Methyltransferases; Middle Aged; Monitoring, Physiologic; Thionucleotides

To elucidate the effect of methotrexate (MTX) on 6-mercaptopurine (6-MP) metabolism in rats.. Fourteen rats were given 6-MP 20 mg/kg daily for 7 days. Seven of the rats were also given MTX 20 mg/kg on days 5 and 7. Blood samples were obtained from all rats on days 0.5 and 8, and red blood cell (RBC) lysates were analysed for thiopurine methyltransferase (TPMT) activity and the concentration of methylated 6-MP metabolites [methyl mercaptopurine ribonucleotides (MMPRP)] and 6-thioguanine nucleotides (6-TGN).. The concentration of MMPRP increased 2.4 times from day 5 to day 8 in RBCs from rats given MTX in addition to 6-MP, as against 1.2 times in rats given 6-MP alone (P = 0.003). 6-TGN levels increased and TPMT activity decreased from day 5 to day 8, with no difference between the 6-MP and the 6-MP plus MTX groups.. Single bolus doses of MTX increase the concentration of MMPRP in rats given daily s.c. doses of 6-MP, with no effect on 6-TGN concentration or TPMT activity.

Topics: Animals; Erythrocytes; Guanine Nucleotides; Male; Mercaptopurine; Methotrexate; Methylthioinosine; Methyltransferases; Nucleic Acid Synthesis Inhibitors; Rats; Rats, Wistar; Thionucleotides

Daily 6-mercaptopurine (6MP) forms the backbone of continuing chemotherapy for childhood lymphoblastic leukaemia (ALL). A major metabolic route is catalysed by thiopurine methyltransferase (TPMT). TPMT deficiency occurs in 1 in 300 individuals and results in high concentrations of thioguanine nucleotides (TGNs), cytotoxic 6MP metabolites. A leukaemic child taking 6MP repeatedly developed profound pancytopenias. TPMT deficiency was confirmed. TGN formation was then studied on attenuated 6MP dosages. Four weekly oral doses of 75 mg/m2 6MP produced TGNs of 2348 pmol/8 x 10(8) red cells, nearly double the maximum TGNs recorded in ALL children with TPMT activity taking long term daily 75 mg/m2 6MP. Grossly elevated TGN concentrations were also produced at 10% standard 6MP dosage (7.5 mg/m2 daily), accompanied by unacceptable 6MP toxicity (neutropenia, diarrhoea, vomiting). The child was eventually stabilised on 10% alternate day therapy and after 15 weeks TGNs were 1670 pmol, just above the upper end of the TGN range for ALL children with TPMT activity.

Topics: Antimetabolites, Antineoplastic; Child; Drug Administration Schedule; Erythrocytes; Female; Guanine Nucleotides; Humans; Mercaptopurine; Methyltransferases; Neutropenia; Precursor Cell Lymphoblastic Leukemia-Lymphoma; Thionucleotides; Thrombocytopenia

Measurement of red cell 6-mercaptopurine (MP) derived 6-thioguanine nucleotide (TGN) and methylmercaptopurine metabolites (MeMPs) can be used to monitor therapy in children who are administered MP for childhood lymphoblastic leukemia. Red cell TGNs are not influenced by the time of blood sampling in relation to the last MP dose. The purpose of this study was to find out whether the same is true for the MeMPs. Plasma MP and red cell MP metabolite pharmacokinetics were studied in seven children immediately before and for 4 hours after a protocol standardized dose of MP. Duplicate blood samples were taken, one was processed immediately whereas one was left at an ambient temperature for 24 hours. The variation in TGN and MeMP metabolites over the 0- to 4-hour period (10 time points per child) was within the error of the assays used. The coefficients of variation for the TGNs ranged from 2.7% to 7% and for the MeMPs, 4% to 10.7%. There was no difference in the TGN and MeMP concentrations measured when the blood samples were left for 24 hours. If a child takes a MP tablet immediately before a clinic appointment, it has no major influence on MeMP measurements.

Topics: Administration, Oral; Antimetabolites, Antineoplastic; Child; Child, Preschool; Erythrocytes; Female; Guanine Nucleotides; Humans; Male; Mercaptopurine; Precursor Cell Lymphoblastic Leukemia-Lymphoma; Thionucleotides; Time Factors

During maintenance chemotherapy for childhood acute lymphoblastic leukemia (ALL), the cytotoxic metabolites of methotrexate (MTX polyglutamates) and mercaptopurine (6MP) (thioguanine nucleotides [6TGN]) accumulate intracellularly, including in erythrocytes (E-MTX and E-6TGN) with large interindividual variations. In the present Nordic Society for Pediatric Hematology and Oncology (NOPHO) study, the relation of E-MTX and E-6TGN to relapse risk was explored.. Two hundred ninety-seven patients with non-B-cell ALL, aged 1 to 14 years, on oral MTX and 6MP had E-MTX and E-6TGN levels measured three to 35 (median, eight) and three to 75 (median, nine) times, respectively. For each patient, a mean of all E-MTX (mE-MTX) and E-6TGN (mE-6TGN) measurements was calculated, as well as the product of mE-MTX and mE-6TGN (mE-MTX-6TGN), since MTX and 6MP may have synergistic action.. For patients in remission, the median mE-MTX and mE-6TGN values were 4.7 nmol/mmol hemoglobin (Hgb) (range, 0.4 to 10.3) and 173 nmol/mmol Hgb (range, 58 to 846). With a median follow-up duration of 66 months for patients in remission, 64 patients relapsed. Cox regression analysis identified mE-MTX-6TGN and sex to be the most significant parameters to predict relapse (global P = .001). Factors that predicted a better prognosis were high mE-MTX 6TGN and female sex. Patients who had a mE-MTX-6TGN less than the product of the median mE-MTX and median mE-6TGN (813 [nmol/mmol Hgb]2) had a significantly poorer event-free survival (EFS) than did patients with higher values (5-year probability of EFS [pEFS5y], 0.70 v 0.86; P = .001).. The pharmacokinetics of MTX and 6MP may have significant influence on the risk of relapse. The value of dose adjustments by E-MTX and E-6TGN remains to be determined.

Topics: Administration, Oral; Adolescent; Child; Child, Preschool; Erythrocytes; Female; Follow-Up Studies; Guanine Nucleotides; Humans; Infant; Male; Mercaptopurine; Methotrexate; Polyglutamic Acid; Precursor Cell Lymphoblastic Leukemia-Lymphoma; Prognosis; Recurrence; Regression Analysis; Remission Induction; Risk; Thionucleotides; Time Factors

Capillary electrophoresis proved to be a useful technique for the analysis of intracellular levels of 6-thioguanosine mono-, di-, and triphosphate with analysis times of 20 min. Conditions required for baseline separation of the thioguanine nucleotides consisted of a 25 mM KH2PO4 (pH 8.0) buffer and a separation voltage of +28 kV. Laser-induced fluorescence detection (lambda ex = 325 nm, lambda em = 410 nm) of the thioguanine nucleotide metabolites of 6-mercaptopurine (6-MP) was possible following oxidation of the thiol functionality. Tedious extraction procedures involving mercury cellulose resins or phenyl mercury adduct formation, which had been required previously for the selective extraction of thiopurines from erythrocytes, were unnecessary due to the overall specificity of the approach. However, the inclusion of 50 mM EDTA in the sample preparation was required to inhibit the anabolic/catabolic enzymatic activity, which was responsible for the degradation of the analytes. The method demonstrated linearity from 5 to 1700 pmol/100 microliters red blood cells for the three analytes (RSDs < or = 8%). The feasibility of the method was demonstrated for the quantitation of 6-thioguanine nucleotides in patients receiving either oral or intravenous 6-MP therapy.

Topics: Electrophoresis; Erythrocytes; Fluorescence; Guanine Nucleotides; Guanosine Diphosphate; Guanosine Triphosphate; Humans; Mercaptopurine; Thionucleotides

Two children with acute lymphoblastic leukaemia (ALL) taking daily 6-mercaptopurine as part of a national UK therapeutic trial repeatedly developed profound myelosuppression on 25% of the standard protocol dose. Both were found to have undetectable intracellular activity of thiopurine methyltransferase (TPMT), an enzyme controlling one of the major alternative catabolic pathways of 6-mercaptopurine, and both produced higher concentrations of cytotoxic drug metabolites at 10-25% of the protocol dose than other patients taking 100%. It is supposed that these patients represent the 0.33% of the normal population constitutionally lacking TPMT. It is important to recognise such individuals both to avoid fatal bone marrow failure through inadvertent overdosage, and to be reassured that an adequate drug effect can be achieved at around 10% of the standard dose.

Topics: Bone Marrow; Child; Child, Preschool; Female; Guanine Nucleotides; Humans; Male; Mercaptopurine; Methotrexate; Methyltransferases; Precursor Cell Lymphoblastic Leukemia-Lymphoma; Reference Values; Thionucleotides; Vincristine

Topics: Adolescent; Child; Female; Guanine Nucleotides; Humans; Male; Mercaptopurine; Patient Compliance; Precursor Cell Lymphoblastic Leukemia-Lymphoma; Thionucleotides

6-mercaptopurine (6-MP) can be inactivated by S-methylation, which is catalysed by thiopurine methyltransferase (TPMT). An alternative metabolic route leads to the formation of cytotoxic 6-thioguanine nucleotides (6-TGN). To investigate whether these two pathways compete with each other to affect the therapeutic response to 6-MP, 6-TGN concentrations and TPMT enzymatic activity were measured in erythrocytes (RBC) from 95 children on long-term 6-MP therapy for lymphoblastic leukaemia (ALL). RBC TPMT activities were also measured in 130 control children and 104 long-term survivors of ALL no longer on treatment. The 95 children on 6-MP showed wide interindividual differences in RBC 6-TGN concentrations at the full protocol dose of 75 mg/m2, and RBC 6-TGN concentrations correlated negatively with RBC TPMT activity. Children with 6-TGN concentrations below the group median had higher TPMT activities and a higher subsequent relapse rate. 50 of the 104 long-term survivors had been treated with "gentle" low-dose protocols, and this subgroup contained an excess of children with lower TPMT activities compared with normal controls. These results indicate that genetically determined TPMT activity may be a substantial regulator of the cytotoxic effect of 6-MP, an effect which in turn could be important in influencing the outcome of therapy for childhood ALL.

Topics: Actuarial Analysis; Administration, Oral; Adolescent; Adult; Analysis of Variance; Child; Child, Preschool; Drug Administration Schedule; Drug Evaluation; Erythrocytes; Female; Follow-Up Studies; Guanine Nucleotides; Homozygote; Humans; Male; Mercaptopurine; Methyltransferases; Precursor Cell Lymphoblastic Leukemia-Lymphoma; Prognosis; Remission Induction; Thionucleotides; Time Factors

A fast and reliable two-step method has been established for the chemical synthesis of 6-thioguanosine 5'-monophosphate, 6-thioguanosine 5'-diphosphate and 6-thioguanosine 5'-triphosphate starting from the ribonucleoside. In the first step, 6-thioguanosine dissolved in triethyl phosphate, at high yield reacts with phosphorus oxide trichloride to 6-thioguanosine 5'-monophosphate which is purified by anion-exchange chromatography on DEAE-Sephadex using a step gradient of hydrochloric acid. In the second step, 6-thioguanosine 5'-monophosphate dissolved in water, reacts with phosphoric acid in the presence of pyridine/dicyclohexyl carbodiimide and is converted to 6-thioguanosine 5'-diphosphate and 6-thioguanosine 5'-triphosphate which are separated from each other and from the 6-thioguanosine 5'-monophosphate by anion-exchange chromatography on DEAE-Sephadex using a gradient of ammonium bicarbonate. Material from each step of the preparation procedure is separated by reversed-phase HPLC chromatography and analyzed for its free ribonucleoside content, 5'-monophosphate, 5'-diphosphate, 5'-triphosphate and small amounts of unidentified phosphorylated compounds. The purity of the final preparations and the identity of each 6-thioguanosine 5'-phosphate are proven by highly specific enzymatic peak-shifting/HPLC analyses using alkaline phosphatase, 5'-nucleotidase, pyruvate kinase, nucleoside diphosphate kinase and combined hexokinase/glucose 6-phosphate dehydrogenase.

Topics: Animals; Chromatography, High Pressure Liquid; Enzymes; Guanine Nucleotides; Guanosine Diphosphate; Guanosine Triphosphate; Kinetics; Mercaptopurine; Phosphorylation; Rabbits; Thionucleotides

In the present study of 12 boys and 19 girls 2-16 years of age (median, 7 years) on oral 6-mercaptopurine (6MP) and methotrexate (MTX) maintenance therapy (MT) for non-B-cell acute lymphoblastic leukemia (ALL), we found that (a) during MT, 6-thioguanine nucleotides (6TGN) (the major cytotoxic metabolite of 6MP) accumulate in the erythrocytes (E-6TGN); (b) for patients receiving an unchanged dose of 6MP, no significant correlation could be demonstrated between the mean E-6TGN (mE-6TGN) and the dose of 6MP (r = -0.11, P = 0.28) (31 patients); (c) among 21 patients receiving 50-75 mg/m2 6MP, a variation of up to 3 orders of magnitude in mE-6TGN could be demonstrated, with the interindividual coefficient of variation (CV) in mE-6TGN for these patients being 0.31; (d) the median intraindividual CV in E-6TGN at an unchanged dose of 6MP was 0.11 (range, 0.04-0.18); and (e) the degree of myelodepression as measured by the mean white cell count was related to mE-6TGN (r = -0.55, P = 0.0006). These results indicate that E-6TGN could be a useful parameter for monitoring 6MP maintenance chemotherapy, although this needs to be explored in prospective studies.

Topics: Adolescent; Antineoplastic Combined Chemotherapy Protocols; Child; Child, Preschool; Dose-Response Relationship, Drug; Erythrocytes; Female; Guanine Nucleotides; Humans; Leukopenia; Male; Mercaptopurine; Methotrexate; Monitoring, Physiologic; Precursor Cell Lymphoblastic Leukemia-Lymphoma; Thionucleotides

A sensitive and rapid assay for the quantitation of thioinosine monophosphate and thioguanosine monophosphate, the major intracellular active metabolites of mercaptopurine and thioguanine, respectively, has been developed. Neoplastic cells are extracted with trichloracetic acid, and the neutralized acid extracts are analyzed by ion-pairing high-performance liquid chromatography with dual-channel uv-wavelength detection. This technique provides a lower limit of sensitivity of 30 pmol of thioinosine monophosphate and 10 pmol of thioguanosine monophosphate. The number of cells assayed per sample was 2 X 10(7). This assay makes it possible to detect and quantitate low levels of thioinosine monophosphate and thioguanosine monophosphate present in neoplastic cells obtained directly from patients receiving mercaptopurine or thioguanine chemotherapy.

Topics: Bone Marrow; Cell Line; Chromatography, High Pressure Liquid; Guanine Nucleotides; Humans; Inosine Monophosphate; Inosine Nucleotides; Leukemia, Myeloid, Acute; Lymphoma; Mercaptopurine; Thioguanine; Thionucleotides

The metabolism of 6-mercaptopurine (6-MP) in L-1210 mouse leukemia cells and human chronic myelocytic leukemia cells (CML cells) was examined. The acid-soluble fractions obtained from cells incubated with [8-14C]6-MP were chromatographed on a Dowex-1 formate resin column using a formic acid linear gradient elution system. Chromatography of the extract of L-1210 cells revealed four principal radioactive peaks. The fraction containing the third peak was hydrolyzed by snake venom 5'-nucleotidase (Crotalus adamanteus). Cellulose thin layer chromatography revealed that the radioactive peak of the hydrolysate corresponded to 6-thioguanosine. The results showed that 6-MP was converted to 6-thioinosinic acid (6-TIMP) and 6-thioguanylic acid (6-TGMP) in L-1210 cells. In order to elucidate the pathway of 6-MP conversion to 6-TGMP, we examined the interaction of [8-14C]6-TIMP and purified IMP dehydrogenase. It was found by DEAE-cellulose thin layer chromatography that the IMP dehydrogenase converted 6-TIMP to 6-thioxanthylic acid (6-TXMP). Dowex-1 chromatography of the acid-soluble extract of human CML cells incubated with [8-14C]-6-MP also revealed a radioactive peak corresponding to 6-TGMP. These results suggest that 6-MP is metabolized to 6-TGMP by serial conversion to 6-TIMP and 6-TXMP through the de novo GMP synthetic pathway in L-1210 cells and human CML cells.

Topics: Animals; Cells, Cultured; Chromatography; Chromatography, DEAE-Cellulose; Guanine Nucleotides; Guanosine; Humans; IMP Dehydrogenase; Kinetics; Leukemia L1210; Leukemia, Myeloid; Mercaptopurine; Mice; Thionucleosides; Thionucleotides

Twelve boys and 10 girls on similar long term remission maintenance treatment for lymphoblastic leukaemia had 79 random assays of their red cell 6 thioguanine nucleotide ( 6TGN ) concentrations performed as an index of cytotoxic activity generated by oral 6-mercaptopurine ( 6MP ). Correlation between the dose of 6MP and 6TGN was statistically significant in the girls (r = 0.58, P less than 0.001) but not in the boys (r = 0.15). Additionally, as a group the boys tolerated more 6MP (P less than 0.05), despite similar prescribing criteria, but this did not result in a higher mean 6TGN concentration or increased myelotoxicity. It appears that girls develop 6MP cytotoxicity at lower doses and more predictably than boys. If so, this may be relevant to the as yet unexplained but marked sex difference in prognosis apparent in some studies.

Topics: Adolescent; Child; Child, Preschool; Erythrocytes; Female; Guanine Nucleotides; Humans; Leukemia, Lymphoid; Leukocyte Count; Male; Mercaptopurine; Neutrophils; Sex Factors; Thionucleotides

The effect of cotrimoxazole on the utilization of 6-mercaptopurine (6MP) was studied in a group of children receiving remission maintenance treatment for lymphoblastic leukaemia (ALL). This was done by measuring the level of an active metabolite of 6MP, 6-thioguanine nucleotide (6TGN), and comparing it both with the drug dose and with subsequent neutropenia in the presence or absence of concurrent cotrimoxazole. In children who were not taken cotrimoxazole, the concentration of 6TGN showed a significant positive correlation with the dose and a significant negative correlation with the absolute neutrophil count 2 weeks later. In those who were taking the antibiotic both these relationships were lost. This suggests that cotrimoxazole can interfere with both the absorption and the cytotoxicity of 6MP and may, in turn, alter its antileukaemic effect.

Topics: Absorption; Adolescent; Child; Child, Preschool; Drug Combinations; Drug Interactions; Female; Guanine Nucleotides; Humans; Leukemia, Lymphoid; Male; Mercaptopurine; Sulfamethoxazole; Thionucleotides; Trimethoprim; Trimethoprim, Sulfamethoxazole Drug Combination

An assay for 6-thioguanine (6-TG) nucleotide, a major metabolite of the cytotoxic drugs azathioprine, 6-mercaptopurine and 6-thioguanine in human red blood cells (RBCs) has been developed. The metabolite was not detected in RBCs when azathioprine or 6-mercaptopurine was incubated with whole blood in-vitro. The assay for intracellular 6-TG nucleotide is specific and requires 8 X 10(8) RBCs (100 microliters packed cells) for which the limit of sensitivity is 30 pmole 6-TG nucleotide. Pre-dose blood samples were obtained, 12 h after the last azathioprine dose, from 10 renal transplant recipients with stable functioning cadaver grafts on a total daily dose of 150 mg azathioprine. The mean 6-TG nucleotide concentration was 171 pmole/8 X 10(8) RBCs (s.d. = 84). The assay is also suitable for use in measuring 6-TG nucleotide in the RBCs of leukaemic children undergoing 6-mercaptopurine treatment.

Topics: Azathioprine; Erythrocytes; Guanine Nucleotides; Humans; In Vitro Techniques; Leukemia, Lymphoid; Mercaptopurine; Thioguanine; Thionucleotides

Topics: Adolescent; Biological Availability; Child; Child, Preschool; Dose-Response Relationship, Drug; Erythrocytes; Female; Guanine Nucleotides; Humans; Leukemia, Lymphoid; Leukocyte Count; Male; Mercaptopurine; Neutrophils; Thionucleotides