6-thioguanine Nucleotide Levels Research Articles

Effects of allopurinol on 6-mercaptopurine metabolism in unselected patients with pediatric acute lymphoblastic leukemia: a prospective phase II study.

Allopurinol can be used in maintenance therapy (MT) for pediatric acute lymphoblastic leukemia (ALL) to mitigate hepatic toxicity in patients with skewed 6- mercaptopurine metabolism. Allopurinol increases the erythrocyte levels of thioguanine nucleotides (e-TGN), which is the proposed main mediator of the antileukemic effect and decreases methyl mercaptopurine (e-MeMP) levels, associated with hepatotoxicity. We investigated the effects of allopurinol in thiopurine methyltransferase (TPMT) wild-type patients without previous clinical signs of skewed 6MP metabolism. Fifty-one patients from Sweden and Finland were enrolled in this prospective beforeafter trial during ALL MT. Mean e-TGN increased from 280 nmol/mmol Hb after 12 weeks of standard MT to 440 after 12 weeks of MT with addition of allopurinol 50 mg/m2 (p.

P798 Study of the concordance between the enzymatic activity of Thiopurine S-methyltransferase and polymorphisms in the TPMT gene in patients with Inflammatory Bowel Disease

Abstract Background Thiopurine therapy is associated with an increased risk of serious adverse events in patients with Inflammatory Bowel Disease (IBD). Thiopurine S-methyltransferase (TPMT) is the enzyme responsible for metabolism, and its determination helps identify patients at risk of toxicity. This study aims to explore the level of correspondence between the TPMT genotype and enzymatic activity (phenotype) with the objective of preventing adverse events prior to the commencement of thiopurine therapy. Methods Consecutive patients with IBD, Crohn’s Disease (CD) or Ulcerative Colitis (UC), were included. Genotype was conducted for the most prevalent TPMT variants (TPMT*2, TPMT*3B, TPMT*3C), TPMT enzymatic activity assessment, and quantification of 6-thioguanine nucleotide (6-TGN) levels. Adverse effects were documented, along with the administration of biologics, mesalazine, ACE inhibitors or allopurinol. The primary objective was to evaluate the concordance between TPMT genotype and phenotype. Secondary objectives included determining the prevalence of genotype and phenotype within our patient cohort, 6-TGN serum levels, the prevalence of adverse events and identifying potential pharmacological inducers of the phenotype. Results A total of 218 patients were included (131 with CD and 81 with UC). Thirteen patients did not initiate thiopurines, and 34.63% (71/205) had discontinued treatment. The most observed adverse events were myelotoxicity (15.12%), gastrointestinal toxicity (12.20%), and hepatic toxicity (5.37%). TPMT activity exhibited a trimodal distribution, with 87.61% (191/218) having normal activity, 11.47% having intermediate activity, and 1% having low activity. Genotypically, 90.83% (198/218) had a wild-type genotype, 8.72% (18/218) had a heterozygous genotype, and 0.5% (1/218) had a homozygous genotype. The concordance index between phenotype and genotype was calculated at 0.57. Significantly, myelotoxicity was associated with the presence of genotypic variants (p=0.004). 6-TGN levels did not correlate with TPMT activity and concurrent medications did not affect TPMT activity . Conclusion The proactive assessment of TPMT (phenotype or genotype) before initiating thiopurine therapy effectively mitigates the occurrence of myelotoxicity in IBD patients. Importantly, the level of concordance between TPMT activity and genotype is moderate, with genotyping serving as the more reliable approach for preselecting patients susceptible to myelotoxic reactions.

The drug-survival of low-dose thioguanine in patients with inflammatory bowel disease: a retrospective observational study.

Thiopurines are commonly used to treat inflammatory bowel disease but withdrawal due to side effects are common. Thioguanine has been suggested to be better tolerated than conventional thiopurines. We studied drug-survival of low dose of thioguanine in real-life clinical practice in comparison to conventional thiopurines. Retrospective observational study. All patients born 1956 and later, and who at least once started thiopurine treatment between 2006 and 2022 were included. A medical chart review was performed that noted drug-survival for every thiopurine treatment attempt. The Mantel-Cox rank test was used to test differences in drug-survival for different thiopurines. Blood chemistry analysis and faecal calprotectin levels were registered for the first 5 years of treatment. In the study population, there was 379 initiated thiopurine treatments (210 for Crohn's disease and 169 for ulcerative colitis) in 307 patients with inflammatory bowel disease (IBD). Low-dose thioguanine (median dose 11 mg; 25-75th percentile 7-19 mg) had been initiated in 31 patients. Overall, when including all thiopurine attempts, thioguanine had the longest drug-survival [Mantel-Cox rank test: thioguanine versus azathioprine p = 0.014; thioguanine versus 6-mercaptopurine (6-MP) p < 0.001]. For second-line thiopurine treatment thioguanine had longer drug-survival than 6-MP (Mantel-Cox rank test: p = 0.006). At 60 months, 86% of the patients who started low-dose thioguanine were still on treatment compared to 42% of the patients who started 6-MP (p = 0.022). The median 6-thioguanine nucleotide levels in patients treated with thioguanine was 364 pmol/8 × 108. Patients on thioguanine treatment showed significantly lower values of median mean corpuscular volume at follow-up than patients treated with azathioprine and 6-MP. Patients treated with 6-MP showed significantly lower levels of FC in the third year of treatment compared to patient treated with azathioprine (59 versus 109 µg/g; p = 0.023), but there was no significant difference in FC levels for thioguanine compared to azathioprine (50 versus 109 µg/g; p = 0.33). Treatment with a low dose of thioguanine is well-tolerated in patients with IBD and had a significantly higher drug-survival than conventional thiopurines.

Association between 6-thioguanine nucleotide levels and preventing production of antibodies to infliximab in patients with inflammatory bowel disease

ObjectiveCombination therapy with infliximab and a thiopurine has been shown to be more effective than monotherapy in patients with inflammatory bowel disease (IBD). The therapeutic efficacy of thiopurines is correlated...

Humoral immune response to SARS-CoV-2 vaccination in patients with inflammatory bowel disease on immunosuppressive medication: association to serum drug levels and disease type

Objectives Immune responses following SARS-CoV-2 vaccination in patients with inflammatory bowel disease (IBD) are not well characterized. The aims of this study were to explore the serological response associated with IBD, and immunosuppressive medications including serum concentrations of biologics and thiopurine metabolites. Materials and methods This prospective, observational study included adult patients with ulcerative colitis (UC) and Crohn’s disease (CD), and healthy controls. Antibodies to the receptor-binding domain of SARS-CoV-2 spike proteins, and serum concentrations of ongoing biologic and immunomodulatory medications were assessed prior to, and 2-5 weeks after the second vaccine dose. Serologic response was defined as anti-Spike antibodies ≥70 AU/ml. Results In 958 IBD patients (380 UC, 578 CD) and 323 healthy controls, the median (Q1; Q3) anti-Spike antibody level (AU/ml) was lower in patients (618 (192; 4370)) compared to controls (3355 (896; 7849)) (p < 0.001). The antibody levels were lower in CD (439 (174; 3304)) compared to UC (1088 (251; 5975)) (p < 0.001). No associations were demonstrated between antibody levels and serum drug concentrations for TNF inhibitor (TNFi), vedolizumab and ustekinumab. Patients receiving TNFi + thiopurines with a subtherapeutic 6-thioguanine nucleotide (6-TGN) level had higher response rate (93%) compared to patients with 6-TGN within the therapeutic range (53%) (p = 0.003). A diagnosis of UC, mRNA-1273 vaccine, and other treatments than TNFi + thiopurines were associated with humoral response. Conclusions Patients with CD had an attenuated humoral response to SARS-COV-2 vaccination as compared to patients with UC. The lack of association between serum levels of biologics and serologic response indicates vaccination regardless of proximity to drug administration.

Comparison of the use of 6-thioguanine riboside versus 6-thioguanine as calibration standard to monitor 6-thioguanine nucleotides in red blood cells

The analytical methods reported to evaluate 6-Thioguanine nucleotides (6-TGNs) level in red blood cells (RBC) are based on the conversion of 6-TGNs to 6-Thioguanine (6-TG) using 6-thioguanine as calibration standard.Using the LC-DAD method previously reported by Dervieux and Boulieu in 1998 to determine 6-TGNs, we evaluated the use of 6-Thioguanine Riboside (6-TGR) as standard instead of the base 6-TG for the monitoring of 6-TGNs in RBC from patients with IBD.Our results show that 6-TGN values measured in RBC from 30 patients were significantly (p < 0,00001) higher when 6-TGR was used as calibration standard compared to 6-TG. The difference observed may be explained by the presence of ribose in the chemical structure of 6-TGR contrary to 6-TG. This difference in 6-TGN values was also observed in external quality control assay using 6-TGR versus 6-TG calibration standard.The use of the nucleoside 6-TGR as calibrator constitutes a better reflection of the chemical reaction which occurs in RBC compared to 6-TG. This preliminary observation suggests that the choice of calibration standard to monitor 6-TGNs may have a significant impact on the values measured in patients and laboratory should be aware of this potential pitfall.

Concomitant 5-aminosalicylic acid treatment does not affect 6-thioguanine nucleotide levels in patients with inflammatory bowel disease on thiopurines.

There are conflicting data as to whether co-treatment with 5-aminosalicylic acid (5-ASA) in patients with inflammatory bowel disease (IBD) under azathioprine (AZA) or 6-mercaptopurine (6-MP) therapy may influence 6-thioguanine nucleotide (6-TGN) concentrations, and whether this combination puts patients at risk of side-effects. The aim of the study was to determine 6-TGN levels in patients treated with AZA/6-MP, either alone or in combination with 5-ASA. Available blood samples from patients treated with AZA or 6-MP were retrieved from the Swiss IBD Cohort Study (SIBDCS). The eligible individuals were divided into 2 groups: those with vs. without 5-ASA co-medication. Levels of 6-TGN and 6-methylmercaptopurine ribonucleotides (6-MMPR) were determined and compared. Potential confounders were compared between the groups, and also evaluated as potential predictors for a multivariate regression model. Of the 110 patients enrolled in this analysis, 40 received concomitant 5-ASA at the time of blood sampling. The median 6-TGN levels in patients with vs. those without 5-ASA co-treatment were 261 and 257 pmol/8×108 erythrocytes, respectively (P=0.97). Likewise, there were no significant differences in 6-MMPR levels (P=0.79). Through multivariate analysis, 6-TGN levels were found to be significantly higher in non-smokers, patients without prior surgery, and those without signs of stress-hyperarousal. Blood concentrations of 6-TGN and 6-MMPR did not differ between patients with vs. those without 5-ASA co-treatment. Our data warrant neither more frequent lab monitoring nor dose adaptation of AZA in patients receiving concomitant 5-ASA treatment.

Body mass index during maintenance therapy and relapse risk in children with acute lymphoblastic leukemia: AChildren's Oncology Group report.

Obesity at diagnosis of childhood acute lymphoblastic leukemia (ALL) is associated with greater risk of relapse; whether this association extends to obesity during maintenance is unstudied. This study used data from AALL03N1 to calculate median body mass index (BMI) for 676 children over 6 consecutive months during maintenance therapy; BMI percentile (BMI%ile) were operationalized as normal/underweight (<85%ile), overweight/obese (85%-98%ile), and extreme obesity (≥99%ile). Hazard of relapse was estimated using multivariable proportional subdistributional hazards regression after adjusting for all relevant demographic and clinical predictors. Median age at study enrollment was 6 years and median length of follow-up was 7.9years. Overall, 43.3% of the cohort was underweight/normal weight, 44.8% was overweight/obese, and 11.8% had extreme obesity. Cumulative incidence of relapse at 4 years from study enrollment was higher among those with extreme obesity (13.6%±4.5%) compared to those with underweight/normal weight (9.0%±2.1%). Multivariable analysis revealed that children with extreme obesity had a 2.4-fold (95% confidence interval [CI], 1.1-5.0; p=.01) greater hazard of relapse compared to those who were underweight/normal weight. Overweight/obese patients were at comparable risk to those who were underweight/normal weight (hazard ratio, 0.8; 95% CI, 0.4-1.6). Erythrocyte thioguanine nucleotide (TGN) levels were significantly lower among children with extreme obesity compared to those with underweight/normal weight (141.6 vs. 168.8 pmol/8×108 erythrocytes; p=.0002), however, the difference in TGN levels did not explain the greater hazard of relapse among those with extreme obesity. Extreme obesity during maintenance therapy is associated with greater hazard of relapse in children with ALL. Underlying mechanisms of this association needs further investigation. Findings from this study demonstrate that extreme obesity during maintenance therapy is associated with a greater hazard of relapse among children with acute lymphoblastic leukemia. We show that children with obesity have lower levels of erythrocyte thioguanine nucleotides even after adjusting for adherence to oral chemotherapy. However, these lower levels do not explain the greater hazard of relapse, paving the way for future studies to explore this association.

Combination Therapy With Immunomodulators Improves the Pharmacokinetics of Infliximab But Not Vedolizumab or Ustekinumab

Background and AimsThe aim of this study was to assess how 6-thioguanine nucleotide (6-TGN) levels and use of oral methotrexate relate to the pharmacokinetics of biologics. MethodsThis was a prospective cohort study including patients with inflammatory bowel diseases on maintenance doses of infliximab, vedolizumab, or ustekinumab on monotherapy or combination with a thiopurine or oral methotrexate. We collected 6-TGN concentrations, biomarker levels, and clinical and endoscopic disease activity. The primary outcomes were infliximab, vedolizumab, and ustekinumab concentrations as well as anti-drug antibodies (ADAs). ResultsA total of 369 patients were recruited (113 infliximab, 133 vedolizumab, and 123 ustekinumab). Patients with 6-TGN levels ≥146 pmol per 8 × 108 red blood cells (RBCs), and those receiving combination therapy with thiopurine or oral methotrexate had significantly higher infliximab concentrations when compared with monotherapy (median levels of 17.4 μg/mL on thiopurine with 6-TGN ≥146 pmol per 8 × 108 RBCs, 17.1 on methotrexate, and 3.9 on infliximab monotherapy; P = .001 for both comparisons). However, there was no association between the use of immunomodulators and 6-TGN concentrations with vedolizumab (median levels of 8.8 on thiopurine with 6-TGN ≥152 pmol per 8 × 108 RBCs, 6.8 on methotrexate, and 10.5 on vedolizumab monotherapy; P > .05 for both comparisons) or ustekinumab median concentrations (median levels of 5.0 on thiopurine with 6-TGN ≥154 pmol per 8 × 108 RBCs, 5.2 on methotrexate and 7.0 on ustekinumab monotherapy; P > .05 for both comparisons). Fourteen (12%) patients had anti-infliximab antibodies, while 1 patient had ADAs in each of the other drug cohorts. ConclusionsAchieving higher 6-TGN levels or the use of methotrexate improved the pharmacokinetics of infliximab. Conversely, these data do not support the use of combination therapy to augment pharmacokinetics with vedolizumab or ustekinumab.

Low-dose thioguanine guided by therapeutic drug monitoring is a safe and effective alternative in inflammatory bowel disease patients intolerant to conventional thiopurines.

Thioguanine is an alternative thiopurine for inflammatory bowel disease (IBD) patients. To evaluate the short-term efficacy and safety of low-dose therapeutic drug-monitored (TDM) thioguanine. A retrospective evaluation of IBD patients intolerant to conventional thiopurines started on thioguanine from 2017 to 2019 with dosing guided by TDM was conducted. Clinical response was defined for ulcerative colitis (UC) as a reduction of partial Mayo score ≥3 with reduction in rectal bleeding score of at least 1 and a final rectal bleeding subscore of 0-1 at Week 12 of therapy. Crohn disease (CD) response was defined as a reduction of Harvey-Bradshaw index ≥3 (HBI) at Week 12 of therapy. Remission was defined in UC as partial Mayo score of <2 and in CD as HBI score of <5. Forty-six patients were included in the study. The median thioguanine dose was 20 mg/day (standard deviation 7.3; range: 10-40 mg/day) with a median 6-thioguanine nucleotide level of 577 pmol/8 × 108 (interquartile range (IQR) IQR 378.5-878.75) for CD and 677.5 pmol/8 × 108 (IQR 523.25-842.25) for UC. The overall clinical response rate was 62% (13/21), intention to treat (ITT). Maintenance of remission was 76% (19/25, ITT). Thirty-seven percent (17/46) of patients experienced an adverse effect. No early cases of nodular regenerative hyperplasia (NRH) were seen. Thioguanine was tolerated well in 63% of patients. A clinical response was seen in 62% of patients, and maintenance of remission was high at 76%. No cases of early NRH were seen. Longer-term follow up is required to ensure safety and to assess durability of response.

The Effect of Pregnancy and Inflammatory Bowel Disease on the Pharmacokinetics of Drugs Related to Inflammatory Bowel Disease-A Systematic Literature Review.

Due to ethical and practical reasons, a knowledge gap exists on the pharmacokinetics (PK) of inflammatory bowel disease (IBD)-related drugs in pregnant women with IBD. Before evidence-based dosing can be proposed, insight into the PK has to be gained to optimize drug therapy for both mother and fetus. This systematic review aimed to describe the effect of pregnancy and IBD on the PK of drugs used for IBD. One aminosalicylate study, two thiopurine studies and twelve studies with biologicals were included. Most drugs within these groups presented data over multiple moments before, during and after pregnancy, except for mesalazine, ustekinumab and golimumab. The studies for mesalazine, ustekinumab and golimumab did not provide enough data to demonstrate an effect of pregnancy on concentration and PK parameters. Therefore, no evidence-based dosing advice was given. The 6-thioguanine nucleotide levels decreased during pregnancy to 61% compared to pre-pregnancy levels. The potentially toxic metabolite 6-methylmercaptopurine (6-MMP) increased to maximal 209% of the pre-pregnancy levels. Although the PK of the thiopurines changed throughout pregnancy, no evidence-based dosing advice was provided. One study suggested that caution should be exercised when the thiopurine dose is adjusted, due to shunting 6-MMP levels. For the biologicals, infliximab levels increased, adalimumab stayed relatively stable and vedolizumab levels tended to decrease during pregnancy. Although the PK of the biologicals changed throughout pregnancy, no evidence-based dosing advice for biologicals was provided. Other drugs retrieved from the literature search were mesalazine, ustekinumab and golimumab. We conclude that limited studies have been performed on PK parameters during pregnancy for drugs used in IBD. Therefore, more extensive research to determine the values of PK parameters is warranted. After gathering the PK data, evidence-based dosing regimens can be developed.

P592 Effectiveness and safety of thioguanine in thiopurine-naïve Inflammatory Bowel Disease patients

Abstract Background Currently thioguanine is considered as off-label rescue therapy for Inflammatory Bowel Disease (IBD) after conventional thiopurine failure. This study aimed to determine the safety, effectiveness and 12-month drug tolerability of thioguanine in thiopurine-naïve IBD patients. Methods We performed an analysis of our multicenter, retrospective cohort study including thiopurine-naïve IBD patients treated with thioguanine as first-line maintenance therapy without concomitant biological therapy. Clinical effectiveness was defined as a sustained clinical response (based on physician’s global assessment) without the (re-)initiation of concurrent biological therapy, corticosteroids or a IBD-related surgical intervention. All adverse events that occurred during follow-up were categorized by the Common Terminology Criteria for Adverse Events. Elevation of two concurrent liver tests were categorized as drug-induced liver injury. Results A total of 103 IBD patients (female 61%, Crohn’s disease 52%) were included with a median daily thioguanine dose of 20mg and median 6-thioguanine nucleotide (6-TGN) levels of 635 pmol/8x108 RBC (IQR 425–1100). Clinical effectiveness at 12 months was observed in 60 out of 99 patients (61%) and 80% of patients were still using thioguanine 12 months after initiation. Four patients did not reach the 12-month follow-up period but were in remission at time of data collection. Of the responding patients at 12 months 88% (N=53) remained responsive until the end of follow-up (median follow-up period 28 months, IQR 17–40 months). Forty-nine patients (48%) developed adverse events (grade 1 or 2), of which 24% graded as moderate (grade 2) and none as severe. Seven patients ceased therapy due to the occurrence of adverse events. Adverse events consisted mainly of elevated liver tests (26%) and gastrointestinal complaints (17%). An infection was documented in three patients, none of them requiring hospitalization. Pancytopenia occurred in two other patients with 6-TGN levels of respectively 2900 and 140 pmol/8x108 RBC. None of the included patients had signs of (noncirrhotic) portal hypertension or underwent a liver biopsy during follow-up. Conclusion This is the first cohort study that reports on the safety and effectiveness of first-line thioguanine maintenance therapy in IBD. Thioguanine therapy was, 12 months after initiation, still used by 80% and clinically effective in 61% of thiopurine-naive patients. Adverse events were relatively common but mainly mild (grade 1) and the discontinuation rate related to adverse events was lower than observed during conventional thiopurine therapy. No signs of (noncirrhotic) portal hypertension were reported

P320 6-Thioguanine nucleotide levels are associated with trough levels of infliximab and adalimumab in Inflammatory Bowel Disease patients on combination therapy: A retrospective multicentre study

Abstract Background Thiopurine co-therapy with anti-tumour necrosis factor alpha (anti-TNFα) agents is associated with higher anti-TNFα drug levels and reduced immunogenicity in patients with Inflammatory Bowel Disease (IBD). However, the, 6-thioguanine nucleotide (6-TGN) threshold level required to confer this benefit remains unclear. We aimed to evaluate the relationship between serum, 6-TGN levels and trough levels of infliximab (IFX) and adalimumab (ADA) in IBD patients on combination therapy. Methods We performed a retrospective multicentre study of IBD patients receiving combination anti-TNFα and thiopurine maintenance therapy, for at least, 3 months duration with stable doses for at least, 2 months. All, 6-TGN levels measured within, 3 months prior to or on the same date as IFX or ADA trough levels, between January, 2015 and August, 2021, were included. Undetectable 6-TGN levels (&amp;lt;5pmol/8x108 RBC) were excluded. Patient demographics, disease characteristics, treatment details, biochemical results, and endoscopy results were collected. Primary outcomes were IFX and ADA levels. Secondary outcomes were antibodies to IFX (ATI) or ADA (ATA). Spearman’s rho was used to correlate, 6-TGN and anti-TNFα levels. The Mann-Whitney U test was used to compare anti-TNFα levels between, 6-TGN quartiles, and, 6-TGN levels between patients with and without detectable ATI. Results 483 paired, 6-TGN and anti-TNFα levels (426 IFX, 57 ADA) from, 266 patients (217 IFX, 49 ADA) were included. The median, 6-TGN level was, 240pmol/8x108 RBC, median IFX level was, 6.55mcg/ml and median ADA level was, 7.60mcg/ml. There were significant positive correlations between, 6-TGN and IFX levels (rho, 0.147, p=0.003), and, 6-TGN and ADA levels (rho, 0.266, p=0.047). Figure, 1 shows median IFX levels stratified by, 6-TGN quartiles (Q1:, 0–156, Q2:, 157–237, Q3:, 238–331 and Q4: &amp;gt;331pmol/8x108 RBC). There was a significant difference in IFX levels between Q2 and Q3 (p=0.017), but not between Q1 and Q2 (p=0.232), or Q3 and Q4 (p=0.544). Figure, 2 shows median ADA levels stratified by, 6-TGN quartiles (Q1:, 0–183, Q2:, 184–252, Q3:, 253–352, Q4: &amp;gt;352pmol/8x108 RBC). ADA levels did not differ significantly between, 6-TGN quartiles., 6 patients had detectable ATI. Of these, 5 patients had, 6-TGN levels in Q1 and, 1 patient had a, 6-TGN level in Q2. Patients with ATI had significantly lower, 6-TGN levels compared to those without ATI (median, 6-TGN, 147 vs, 239.5pmol/8x108 RBC, p=0.017). No patients had detectable ATA. Conclusion 6-TGN levels are associated with IFX and ADA trough levels. Target, 6-TGN levels in the lower end of the therapeutic range may be adequate to maintain anti-TNFα levels and prevent anti-drug antibodies in combination therapy.

Discontinuation of Azathioprine could be considered in pediatric patients with Crohn\u2019s disease who have sustained clinical and deep remission

Few studies have demonstrated treatment strategies about the duration and cessation of medications in patients with Crohn’s disease (CD). We investigated factors affecting clinical relapse after infliximab (IFX) or azathioprine (AZA) withdrawal in pediatric patients with CD on combination therapy. Pediatric patients with moderate-to-severe CD receiving combination therapy were analyzed retrospectively and factors associated with clinical relapse were investigated. Discontinuation of IFX or AZA was performed in patients who sustained clinical remission (CR) for at least two years and achieved deep remission. A total of 75 patients were included. Forty-four patients (58.7%) continued with combination therapy and 31 patients (41.3%) discontinued AZA or IFX (AZA withdrawal 10, IFX withdrawal 15, both withdrawal 6). Cox proportional-hazards regression and statistical internal validation identified three factors associated with clinical relapse: IFX cessation (hazard ratio; HR 2.982, P = 0.0081), IFX TLs during maintenance therapy (HR 0.581, P = 0.003), 6-thioguanine nucleotide (6-TGN) level (HR 0.978, P < 0.001). However, AZA cessation was not associated with clinical relapse (P = 0.9021). Even when applied in pediatric patients who met stringent criteria, IFX cessation increased the relapse risk. However, withdrawal of AZA could be contemplated in pediatric patients with CD who have sustained CR for at least 2 years and achieved deep remission.

Pharmacogenetics-based personalized treatment in patients with inflammatory bowel disease: A review

The development of treatment options has revolutionized the prognosis of inflammatory bowel disease (IBD). However, a particular group of patients still experience therapeutic failure or drug side effects. Although the high inter-patient variability in therapy is associated with clinical factors, including age, disease behavior, and disease duration, they attribute only a small proportion of inter-individual variability. Thus, pharmacogenetics evaluating associations between specific genetic variations and drug responses or side effects have focused on optimizing therapeutic efficacy and minimizing toxicity in IBD treatment. Thiopurine S-methyltransferase (TPMT) and nudix hydrolase 15 (NUDT15) are well-established predictive markers of thiopurine-induced myelosuppression. Low TPMT activity is related to increased 6-thioguanine nucleotide levels, subsequently leading to myelotoxicity. NUDT15 variants are strongly associated with thiopurine-induced early leukopenia in Asians, with a lower incidence of TPMT-deficient allele. The Korean Association for the Study of Intestinal Diseases guidelines recommend pretreatment determination of NUDT15 genotypes, especially in East Asians, and NUDT15 R139C measurement has been approved for clinical use since 2019. Several studies have attempted to identify powerful genetic markers for personalized medicine. In this article, we review the identified pharmacogenetics of currently available drugs, focusing on 5-aminosalicylic acid, glucocorticosteroids, thiopurines, and anti-tumor necrosis factor-alpha agents.

No Association between Relapse Risk and Thiopurine Methyltransferase Activity By Either Geno- or Phenotype in the NOPHO ALL-2008 Protocol

Background6-mercaptopurine/methotrexate (6MP/MTX) maintenance therapy is essential for high cure rates of childhood acute lymphoblastic leukemia (ALL). 6MP exerts its cytotoxicity through conversion into thioguanine nucleotides (TGN) that are substrates for DNA polymerase and compete with normal guanine for DNA incorporation. DNA-TGN occasionally mismatches to thymidine causing cell death due to futile attempts of repetitive mismatch repair. We recently showed that higher levels of DNA-incorporated TGN were associated with a reduced relapse risk in non-high risk ALL (Nielsen, Lancet Oncol 2017). Common germline variants in TPMT that codes for thiopurine methyltransferase (TPMT) lead to reduced TPMT activity and higher levels of cytosol TGN when measured in erythrocytes. Thus, low TPMT activity could theoretically lead to higher DNA-TGN and a lower relapse rate. Low TPMT activity (phenotype) has been associated with a lower relapse risk in the NOPHO ALL92 protocol (Schmiegelow, Leukemia 2009). We explored if TPMT status evaluated by either geno- or phenotype was related to DNA-TGN levels and relapse rate in the NOPHO ALL2008 protocol.MethodTPMTgenotype and repeated phenotyping (2141 measurements in total), and levels of DNA-TGN (10830 measurements in total) during maintenance therapy were determined for 918 children treated for non-high risk ALL (89% of all eligible patients). On a subset (N=204) TPMT phenotype was available both at diagnosis and during maintenance therapy. 6MP was started at 50 and 75 mg/m2 for TPMT heterozygous and wildtype patients, respectively, and subsequently adjusted to a target WBC of 1.5-3.0 x109/L. Cox analyses were adjusted by age, sex and WBC at diagnosis.Results78 of the 918 non-high risk patients were TPMT heterozygous (none homozygous deficient). TPMT activities were significantly higher during maintenance therapy than at diagnosis for both TPMT wildtype and heterozygous children (N=204; wildtype: 16.6 vs. 11.3 U/mL ery., p <0.001; heterozygous: 9.9 vs. 6.8 U/mL ery., p=0.047). Mean DNA-TGN levels were higher for TPMT heterozygous than wildtype children (N=523, children in remission at end of therapy with Maintenance II measurements, median 760.9 fmol/µg, range 125.7−1255.0 vs. 494.9, range 44.0−1559.0, p <0.001; Figure). The 5-year cumulative incidence of relapse was 4.2% (95% CI: 0-8.7%) for TPMT heterozygous and 5.7% (95% CI: 3.8-7.5%) forwildtype children (hazard ratio =0.87, 95% CI: 0.27−2.83, p=0.81). No association between mean TPMT activity during maintenance therapy (as a time-dependent covariate) and relapse-specific hazard was seen (N=812; hazard ratio =0.98 per one unit increase in TPMT activity (U/mL ery.), 95% CI: 0.89−1.08, p=0.63). In patients with positive MRD at end of induction (d29) results were similar (N=476; hazard ratio =1.04 per one unit increase in TPMT activity (U/mL ery.), 95% CI: 0.93−1.16, p=0.50).ConclusionAlthough mean DNA-TGN levels were higher in TPMT heterozygous than wildtype children, variability was high in both groups and DNA-TGN levels were largely overlapping, and no association with relapse hazard was seen for neither TPMT geno- nor phenotype. In accordance with previously published data TPMT phenotype determinations done in erythrocytes present at ALL diagnosis can lead to TPMT misclassification.FIGUREa) Mean DNA-TGN levels (N=523) during Maintenance II of children in clinical remission with completed therapy and at least one measurement.b) Cumulative incidence curves of relapse of children carrying TPMT wildtype vs. heterozygous. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Increments in DNA-thioguanine level during thiopurine-enhanced maintenance therapy of acute lymphoblastic leukemia.

Maintenance therapy containing methotrexate and 6-mercapto - purine is essential to cure acute lymphoblastic leukemia (ALL). Cytotoxicity is elicited by incorporation of thioguanine nucleotides into DNA (DNA-TG), and higher leukocyte DNA-TG is associated with increased relapse-free survival. As 6-thioguanine provides 6- fold higher cytosolic levels of thioguanine nucleotides than does 6- mercapto purine, we added low-dose 6-thioguanine to methotrexate/6- mercapto purine maintenance therapy to explore if this combination results in significantly higher DNA-TG. The target population of the “Thiopurine Enhanced ALL Maintenance therapy” (TEAM) study was 30 patients with non-high-risk ALL, aged 1-45 years on methotrexate/6-mercaptopurine maintenance therapy receiving no other systemic chemotherapy. Incremental doses of 6-thioguanine were added to methotrexate/6-mercaptopurine maintenance therapy (starting 6-thioguanine dose: 2.5 mg/m2/day, maximum: 12.5 mg/m2/day). The primary endpoint was DNA-TG increments. Thirty-four patients were included, and 30 patients completed maintenance therapy according to the TEAM strategy. Of these 30 patients, 26 (87%) tolerated 10.0-12.5 mg/m2/day as the maximum 6-thioguanine dose. TEAM resulted in significantly higher DNA-TG levels compared to those in both TEAM patients before their inclusion in TEAM (on average 251 fmol/mg DNA higher [95% confidence interval: 160-341; P<0.0001]), and with historical patients receiving standard methotrexate/6-mercapto - purine maintenance therapy (on average 272 fmol/mg DNA higher [95% confidence interval: 147-398; P<0.0001]). TEAM did not increase myelotoxicity or hepatotoxicity. In conclusion, TEAM is an innovative and feasible approach to improve maintenance therapy and results in higher DNA-TG levels without inducing additional toxicity. It may therefore be an effective strategy to reduce the risk of ALL relapse through increased DNA-TG. This will be tested in a randomized ALLTogether-1 substudy.

Towards personalised medicine in autoimmune hepatitis: Measurement of thiopurine metabolites results in higher biochemical response rates

Patients with autoimmune hepatitis (AIH) usually receive maintenance therapy with thiopurines, such as azathioprine (AZA) or mercaptopurine. Genetic polymorphisms in AZA metabolism can lead to variations in thioguanine nucleotide (TGN) and 6-methylmercaptopurine, both of which can cause adverse drug reactions (ADRs). In inflammatory bowel disease, a therapeutic TGN range (225-450 pmol/8x108 erythrocytes) has been identified to optimise effectiveness. We evaluated the benefits of a personalised medicine approach to thiopurine dosing, in comparison to standard weight-based dosing. A retrospective matched cohort study of 214 patients with AIH who were seen at King's College between 1999-2019 was performed. Metabolite levels were measured in 109 patients. The control group included 105 patients on weight-based thiopurine dosing with no metabolite monitoring. Biochemical response (BR) occurred more frequently at 6-month follow-up in patients with metabolite monitoring compared to those on a weight-based regimen (77% vs. 60%, p= 0.008). This remained true with data analysis based on clinicians who measure metabolites and those who do not (BR at 6 months: 84% vs. 64%, p= 0.016). Patients with BR had TGN levels within the therapeutic range of 225-450 pmol/8x108 erythrocytes significantly more often than those who failed to achieve or lost BR (40% vs. 13%, p <0.0001). Moreover, TGN levels within the pre-defined therapeutic range predicted more stable disease within 6 months of testing compared to levels outside the range (p <0.0001). A high proportion of patients with sub-therapeutic TGN levels (75-225 pmol/8x108 erythrocytes) remained in BR (75% vs. 81%, p= 0.589) with fewer ADRs (44% vs. 86%, p= 0.0002) when compared to patients with therapeutic TGN levels. A strategy of personalised medicine using metabolite levels can optimise treatment regimens in AIH, resulting in fewer ADRs whilst maintaining BR. This study looked to see if measuring the breakdown products of a medication used in autoimmune hepatitis increases the chances of gaining good control of the disease, when compared to a group of patients who were on a dose of this medication based on their weight. A group of 214 patients with autoimmune hepatitis were split into 2 groups: roughly half had their medication dose adjusted according to measurements of breakdown products in the blood, whilst the other half received their weight-based dose as normal. The results confirmed that using a personalised approach and checking drug breakdown products resulted in fewer side effects and potentially improved control of disease.

Dynamics of leucocyte DNA thioguanine nucleotide levels during maintenance therapy of childhood acute lymphoblastic leukemia.

Methotrexate (MTX)/6-Mercaptopurine (6MP)-based maintenance therapy is crucial to cure childhood acute lymphoblastic leukemia (ALL). Cytotoxicity is mediated by incorporation of thioguanine nucleotides (TGN) into DNA (DNA-TG) with higher levels in leucocytes being associated with reduced relapse risk. To further understand the dynamics of DNA-TG formation, we measured DNA-TG levels in leucocyte subsets during maintenance therapy and in the months following its discontinuation. DNA-TG levels were measured in leucocytes (DNA-TGTotal), polymorph nucleated granulocytes (neutrophils, eosinophils, basophils [DNA-TGPMN]) and mononucleated cells (lymphocytes, monocytes [DNA-TGMNC]) in 1013 samples from 52 patients on ALL maintenance therapy (951 samples during therapy and 62 samples after therapy discontinuation, respectively). Median DNA-TGTotal, DNA-TGPMN and DNA-TGMNC during maintenance therapy were 539, 563 and 384 fmol/µg DNA, respectively. DNA-TGPMN displayed more pronounced fluctuation than DNA-TGMNC (range 0-3084 [interquartile range IQR 271-881] versus 30-1411 [IQR 270-509] fmol/µg DNA). DNA-TGTotal was more strongly correlated with DNA-TGPMN (rS = 0.95, p < 0.0001) than DNA-TGMNC (rS = 0.73, p < 0.0001). DNA-TGPMN correlated less with DNA-TGMNC (rS = 0.64, p < 0.0001) and to a much lesser extent with absolute neutrophil count (rS = 0.35, p < 0.0001). Following discontinuation of therapy, DNA-TGPMN was rapidly eliminated, and not measurable beyond day 22 after discontinuation, whereas DNA-TGMNC was slowly eliminated, and five patients demonstrated a measurable DNA-TGMNC more than 365days after therapy discontinuation. Fluctuations in DNA-TGTotal are predominantly caused by corresponding fluctuations in DNA-TGPMN, thus DNA-TGTotal measures recent TGN incorporation in these short-lived cells. Measurement of DNA-TGTotal at 2-4weeks intervals provides a reliable profile of DNA-TG levels.

Relationship Between Thiopurine S-Methyltransferase Genotype/Phenotype and 6-Thioguanine Nucleotide Levels in 316 Patients With Inflammatory Bowel Disease on 6-Thioguanine.

Background: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.Methods: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).Results: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).Conclusions: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.