P407 Thiopurines: use them or lose them? Results of an international survey

Background With the development of new drugs for inflammatory bowel disease (IBD), the role of thiopurines in the therapeutic algorithm of Crohn’s disease (CD) and Ulcerative Colitis (UC) is questioned. We aimed to investigate the current practice and future perspective of IBD physicians regarding the efficacy, safety, and role of precision medicine of thiopurines in IBD. Methods A 29-question web-based survey was developed and distributed to IBD physicians internationally, via national IBD associations and word of mouth. The final survey form was reviewed and approved by the clinical committee of ECCO. In the absence of a universal definition of an “IBD expert”, we defined them as senior physicians observing more than 20 patients per week. Results Complete answers from 408 physicians from 50 countries were obtained, the majority from European countries. Participants were mainly experienced physicians in IBD, with 26.0% meeting our definition of “IBD expert”. Only four physicians reported not using thiopurines in clinical practice. Azathioprine was the most used drug (97.1% of participants), while 61.3% of physicians reported using mercaptopurine; only 6.6% of participants used thioguanine. Most respondents used thiopurines in both monotherapy and in combination therapy, for both CD and UC, and consider them as effective treatments (Figures 1 and 2). Respondents tended to consider thiopurines as drugs with a good safety profile, with the agreement of 61.5% of the overall cohort; this was true especially in patients aged between 30 and 50 years old, with the agreement of 80.2% of the overall cohort (Figure 3). Access to TPMT activity assessment and metabolites measurement varied regionally. Physicians considered access to metabolites measurement as an important tool to improve thiopurines’ efficacy and safety, and to increase confidence when prescribing these drugs (Figure 4). Only a minority of the physicians (~6%) considered that thiopurines will not be used in the future in IBD, while 57.8% believed they will still be used both in mono and combination therapy in CD and UC. Conclusion Despite the many emerging treatments in IBD, thiopurines will still be part of the treatment algorithm of both CD and UC in the future. Access to tools for precision medicine varies around the globe, and wider implementation of these instruments, especially metabolite measurement, may improve confidence in thiopurines prescription, improving safety and efficacy.

Thiopurine Drugs in the Treatment of Ulcerative Colitis: Identification of a Novel Deleterious Mutation in TPMT

Chronic inflammatory bowel disease (IBD) includes Crohn’s disease and ulcerative colitis. Both are characterized by inflammation of part of the digestive tract lining. Azathioprine (AZA) is a well-known immunosuppressant that has been known for many years for its ability to provide long-term disease remission in IBDs, but has important side effects, most of which are related to a single nucleotide polymorphism in the gene for thiopurine methyltransferase (TPMT), which ensures the degradation and efficacy of AZA. Since a direct correlation between TPMT gene polymorphisms and the haematological toxicity of the AZA treatment has been widely demonstrated, TPMT genotyping has been made necessary prior to any introduction of AZA. The monitoring of thiopurine metabolites presents one of the factors that limit wide adaptation of these thiopurines in clinical practice. Thus, identifying patients with asymmetric metabolism could help clinicians provide an ideal treatment recommendation to improve response and reduce adverse effects. Here, we review the role of AZA in the treatment of IBD and discuss the usefulness of TPMT genotyping to guide clinical decision-making. In addition, we report the identification of a new molecular alteration, never described, TPMT mutation affecting the TPMT activity and responsible for deleterious side effects in a clinical case of a 20-year-old woman patient.

Normal Ranges of Thiopurine Methyltransferase Activity Do Not Affect Thioguanine Nucleotide Concentrations With Azathioprine Therapy in Inflammatory Bowel Disease

Background Thiopurine methyltransferase (TPMT) activity influences azathioprine conversion into active metabolite 6-thioguanine nucleotide (6-TGN). Low TPMT activity correlates with high 6-TGN and risk for myelosuppression. Conversely, normal-to-high TPMT activity may be associated with low 6-TGN and drug resistance, the so-called hypermetabolizers. Our aim was to identify the effect of normal-to-high TPMT activity on 6-TGN concentrations in an inflammatory bowel disease population. Methods A retrospective chart review of patients aged ≥18 with inflammatory bowel disease, on azathioprine, with documented TPMT activity and 6-TGN concentration was performed. Correlations were evaluated via the Spearman rho correlation coefficient. Linear regression was used to determine the effect of TPMT activity on 6-TGN accounting for confounders. Relationships between TPMT activity, drug dose, and 6-TGN levels were defined via average causal mediation effects. Results One hundred patients were included. No correlation was observed between TPMT activity, azathioprine dosing, and metabolite concentrations. Overall, 39% of the cohort had a therapeutic 6-TGN level of >230 pmol/8 × 108 red blood cells (RBCs). No patient under 1 mg/kg achieved a therapeutic 6-TGN level, whereas 42% of patients taking 2.5 mg/kg did. The median 6-TGN concentration was higher for those in remission (254 pmol/8 × 108 RBCs, interquartile range: 174, 309) versus those not in remission (177 pmol/8 × 108 RBCs, interquartile range: 94.3, 287.8), though not significantly (P = 0.08). Smoking was the only clinical factor associated with 6-TGN level. On multivariate linear regression, only age, azathioprine dose, and obese body mass index were predictive of metabolite concentration. Conclusions Variations within the normal range of TPMT activity do not affect 6-TGN concentration.

P709 Is azathioprine safe as long-term treatment in paediatric patients with inflammatory bowel disease?

Background The toxicity of azathioprine (AZA) includes myelosuppression, infections, pancreatitis, photosensitivity, and hepatotoxicity. The aim of this study was to describe the adverse effects profile of azathioprine as long-term treatment in paediatric inflammatory bowel disease (IBD). Methods An observational, descriptive and retrospective study was performed in the paediatric IBD Unit of a tertiary care hospital from September 2008 to December 2018. It was included patients under 18 diagnosed with IBD who were treated with AZA during their follow-up. We recorded epidemiological data, thiopurine methyltransferase (TPMT) enzyme activity, AZA side effects, and the dosage the patients were receiving when these effects took place. Bone marrow suppression (BMS) was defined as leukopenia, thrombocytopenia and/or anaemia. Acute pancreatitis (AP) induced by azathioprine was considered when two of these criteria (Atlanta 2012) were met: lipase increase (> 3 times normal value), congruent signs and symptoms and/or echographic findings, without other possible aetiology and with complete recovery after AZA withdrawal. Results We included 52 patients, being 31 men (59.6%). They were diagnosed with Crohn′s disease (CD) (73%), ulcerative colitis (UC) (21%) and IBD-unclassified (6%). The median TPMT activity was 17 U/ml (14.2–19.2). Up to 63.5% developed adverse effects by AZA with a median time from the beginning of treatment of 11.4 months (2.6–26.4) and a median dosage of 2 mg/kg/day (1.7–2.3). The most frequent side effect was BMS (52%). These patients had a median TPMT activity of 16.9 U/ml (14.2–18.9), the median duration of treatment was 14 months (3.9–27.7), and the median dosage was 2 mg/kg/d (1.8–2.5). BMS was more frequent in patients with UC (p 0.04) and longer treatment (p 0.08). No differences were found according to age, sex or TPMT activity. Up to 11.5% developed AP, the median duration of treatment until its appearance was 1.5 months (0.7–43.3) and the median dosage was 2 mg/kg/d (1.5–2.5). No differences were found related to age, sex, diagnosis or dosage. Other side effects were: 3 flu-like symptoms, 3 opportunistic infections, 2 hypertransaminasemia, and 1 patient with elevated pancreatic enzymes and hyperbilirubinemia. AZA was discontinued in 14 patients (43.8%): in 6 due to AP, in 4 due to severe lymphopenia, in 2 because of Epstein-Barr virus infection, in 1 due to flu-like symptoms and in 1 with several adverse effects. Conclusion More than half of the patients treated with AZA presented side effects, mainly BMS, although most of them were mild and temporary, and the withdrawal of the drug was not necessary. It seems that TPMT activity is not useful to predict BMS, but this adverse effect could be related to a longer treatment.

Retrospective Data Analysis of the Influence of Age and Sex on TPMT Activity and Its Phenotype–Genotype Correlation

Background Therapeutic efficacy and toxicity of thiopurine drugs (used as anticancer and immunosuppressant agents) are affected by thiopurine S-methyltransferase (TPMT) enzyme activity. TPMT genotype and/or phenotype is used to predict the risk for adverse effects before drug administration. Inosine triphosphate pyrophosphatase (ITPA) is another enzyme involved in thiopurine metabolism. In this study, we aimed to evaluate (a) frequency of various TPMT phenotypes and genotypes, (b) correlations between them, (c) influence of age and sex on TPMT activity, and (d) distribution of ITPA variants among various TPMT subgroups. Methods TPMT enzyme activity was determined by LC-MS/MS. TPMT (*2,*3A–C) and ITPA (rs1127354, rs7270101) genotypes were determined using a customized TaqMan® OpenArray®. Results TPMT enzyme activity varied largely (6.3–90 U/mL). The frequency of low, intermediate, normal, and high activity was 0.5% (n = 230), 13.1% (n = 5998), 86.1% (n = 39448), and 0.28% (n = 126), respectively. No significant difference in TPMT activity in relation to age and sex was found. Genotype analysis revealed the frequency of variant TPMT alleles was 6.73% (*3A, n = 344), 0.05% (*3B, n = 2), 2.22% (*3C, n = 95), and 0.42% (*2, n = 19). Analysis of paired phenotype and genotype showed that TPMT activity in samples with variant allele(s) was significantly lower than those without variant alleles. Lastly, an equal distribution of ITPA variants was found among normal and abnormal TPMT activity. Conclusions This retrospective data analysis demonstrated a clustering of variant TPMT genotypes with phenotypes, no significant influence of age and sex on TPMT activity, and an equal distribution of ITPA variants among various TPMT subgroups.

Validation of a high-performance liquid chromatography method for thiopurine S-methyltransferase activity in whole blood using 6-mercaptopurine as substrate

Background:Variation in metabolism, toxicity and therapeutic efficacy of thiopurine drugs is largely influenced by genetic polymorphisms in the thiopurine S-methyltransferase (TPMT) gene. Determination of TPMT activity is routinely performed in patients to adjust drug therapy.Methods:We further optimized a previously established high-performance liquid chromatography (HPLC) method by measuring TPMT activity in whole blood instead of isolated erythrocytes, which is based on conversion of 6-mercaptopurine to 6-methylmercaptopurine using S-adenosyl-methionine as methyl donor.Results:The simplified TPMT whole-blood method showed similar or better analytical and diagnostic performance compared with the former erythrocyte assay. The whole-blood method was linear for TPMT activities between 0 and 40 nmol/(mL·h) with a quantification limit of 0.1 nmol/(mL·h). Within-day imprecision and between-day imprecision were ≤5.1% and ≤8.5%, respectively. The optimized method determining TPMT activity in whole blood (y) showed agreement with the former method determining TPMT activity in erythrocytes (x) (n=45, y=1.218+0.882x; p>0.05). Phenotype-genotype concordance (n=300) of the whole-blood method was better when TPMT activity was expressed per volume of whole blood (specificity 92.2%), whereas correction for hematocrit resulted in lower genotype concordance (specificity 86.9%). A new cutoff for the whole-blood method to distinguish normal from reduced TPMT activity was determined at ≤6.7 nmol/(mL·h).Conclusions:This optimized TPMT phenotyping assay from whole blood using 6-MP as substrate is suitable for research and routine clinical analysis.

AZATHIOPRINE INDUCED PANCYTOPENIA-A CASE REPORT

<p>Azathioprine (AZA), a prodrug of 6 mercaptopurine, is an immunosuppressant that can be used as adjunctive therapy with corticosteroids in the treatment of arteritic form of ischemic optic neuropathy. Although myelotoxicity is known to occur while using azathioprine, severe pancytopenia is rarely seen. Patients with thiopurine methyltransferase (TPMT) deficiency are at high risk of developing severe myelosuppression. A 63 year* old female with ischemic optic neuropathy was initiated treatment with oral methylprednisone. As two courses of oral steroids showed no significant improvement, oral azathioprine 25 mg twice daily was added and gradually increased to 50 mg twice daily with relief of symptoms. 6 mo later, she was admitted with acute stroke and lab reports showed low levels of total blood counts or pancytopenia. The patient was put on broad spectrum antibiotics; given injection of granulocyte colony stimulating factor 300 mcg subcutaneously and blood transfusion as correction till counts normalised. She improved over 14 d and on next follow-up counts were in normal range. Causality was assessed by Naranjo causality assessment scale and a probable relationship was obtained between azathioprine and pancytopenia with a score of 6. Variations in TPMT activity occurs due to genetic polymorphism. Physicians should be aware of the possibility of myelosuppression while prescribing azathioprine. Frequent blood count monitoring is the most convenient way to avoid this problem where testing for TPMT deficiency is not possible.</p>