CPIC UPDATE
Clinical Pharmacogenetics Implementation Consortium Guideline for Thiopurine Dosing Based on TPMT and NUDT15 Genotypes: 2018 Update Mary V. Relling1, Matthias Schwab2,3,4 , Michelle Whirl-Carrillo5, Guilherme Suarez-Kurtz6, Ching-Hon Pui7, Charles M. Stein8, Ann M. Moyer9 , William E. Evans1, Teri E. Klein4, Federico Guillermo Antillon-Klussmann10,11, Kelly E. Caudle1, Motohiro Kato12, Allen E.J. Yeoh13,14, Kjeld Schmiegelow15,16 and Jun J. Yang1
Thiopurine methyltransferase (TPMT) activity exhibits a monogenic codominant inheritance and catabolizes thiopurines. TPMT variant alleles are associated with low enzyme activity and pronounced pharmacologic effects of thiopurines. Loss-of-function alleles in the NUDT15 gene are common in Asians and Hispanics and reduce the degradation of active thiopurine nucleotide metabolites, also predisposing to myelosuppression. We provide recommendations for adjusting starting doses of azathioprine, mercaptopurine, and thioguanine based on TPMT and NUDT15 genotypes (updates on www.cpicpgx.org).
This document is an update to the Clinical Implementation DRUGS: THIOPURINES Consortium (CPIC) Guidelines for Thiopurine Methyltransferase Background. Three thiopurines are used clinically: azathioprine Genotype and Thiopurine guideline updated last in April 2013. (a prodrug for mercaptopurine), mercaptopurine, and thioguanine. The guideline text, evidence table, and recommendations have been Although all three medications share many of the same pharmacologic updated to reflect new evidence. Specifically, this guideline adds a effects, mercaptopurine and azathioprine are generally used for nonma- recommendation for NUDT15 genotype with minor changes to lignant immunologic disorders, mercaptopurine for lymphoid malig- the TPMT recommendations. Although most of the dosing recom- nancies, and thioguanine for myeloid leukemias. Because azathioprine mendations have been generated from clinical studies in just a few is a prodrug for mercaptopurine, the two drugs can be considered to diseases, we have extrapolated recommended doses to all conditions, have identical interactions with thiopurine methyltransferase (TPMT) given the pharmacokinetic nature of the genotype/phenotype asso- and nudix (nucleoside diphosphate linked moiety X)-type motif 15 ciations. CPIC guidelines are published and periodically updated (NUDT15). Recommendations for individuals with variants in one or on www.cpicpgx.org. Detailed guidelines for use of phenotypic tests both of these genes will be addressed in detail in the following sections. (e.g., TPMT activity and thiopurine metabolite levels) as well as anal- yses of cost-effectiveness are beyond the scope of this document. GENES: TPMT AND NUDT15 Background FOCUSED LITERATURE REVIEW TPMT. TPMT activity is inherited as a monogenic, autosomal A systematic literature review focused on TPMT and NUDT15 codominant trait (Figure S1). Three TPMT single nucleotide genotypes and thiopurine use was conducted (details in polymorphisms (SNPs), which result in unstable proteins and Supplement). Definitive reviews1–4 were relied upon to summa- enhanced TPMT protein degradation,2,3 account for over 90% of rize much of the earlier literature. low activity phenotypes and are the most common inactivating
1Department of Pharmaceutical Sciences, St. Jude Children’s Research Hospital, Memphis, Tennessee, USA; 2Dr. Margarete Fischer-Bosch Institute of Clinical Pharmacology, Stuttgart, Germany; 3Department of Clinical Pharmacology, Institute of Experimental and Clinical Pharmacology and Toxicology, University Hospital, Tuebingen, Germany; 4Department of Pharmacy and Biochemistry, University of Tuebingen, Tuebingen, Germany; 5Department of Biomedical Data Science, Stanford University, Stanford, California, USA; 6Instituto Nacional de Câncer, Rio de Janeiro, Brazil Pharmacogenomics Network, Rio de Janeiro, Brazil; 7Department of Oncology, St. Jude Children’s Research Hospital, Memphis, Tennessee, USA; 8Division of Clinical Pharmacology, Vanderbilt University School of Medicine, Nashville, Tennessee, USA; 9Department of Laboratory Medicine and Pathology, Mayo Clinic, Rochester, Minnesota, USA; 10National Pediatric Oncology Unit, Guatemala City, Guatemala; 11School of Medicine, Universidad Francisco Marroquin, Guatemala City, Guatemala; 12Department of Pediatric Hematology and Oncology Research, National Center for Child Health and Development, Tokyo, Japan; 13National University Health System, National University Cancer Institute, Singapore; 14Viva University Children’s Cancer Centre, Yong Loo Lin School of Medicine, National University of Singapore, Singapore; 15Department of Paediatrics and Adolescent Medicine, Rigshospitalet University Hospital, Copenhagen, Denmark; 16Institute of Clinical Medicine, University of Copenhagen, Copenhagen, Denmark. Correspondence: Jun J. Yang ([email protected]; [email protected]) Received July 17, 2018; accepted October 24, 2018; advance online publication Month 00 2019. doi:10.1002/cpt.1304
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Azathioprine (AZA) 6-Thioguanine (6TG) GSTA1 XDH Thioxanthine GDA GSTA2 GSTM1 Thiouric acid
XDH Mercaptopurine (6MP) SLC29A2 SLC29A1 SLC28A3 SLC28A2 ABCC4 ABCC5
SLC29A2 SLC29A1 8-OHTG 6TG SLC28A3 AOX1 TITP ITPA HPRT1 rTG
SLC28A2 NT5C2 HPRT1 TIMP IMPDH1 GMPS TPMT TXMP TGMP 6MP PRPP meTGMP
NUDT15 RAC1 NT5C2 NME1 TPMT TGDP TGTP TPMT NME2 AdoMet RRM1
MPR RRM2 AdoHcy RNA damage TdGDP meMP TPMT Folate/ TdGMP Homocysteine Pathway apoptosis ADK NUDT15 meMPR meTIMP TdGTP
hepatotoxicity DNA damage
apoptosis PPAT de novo Purine Ribose 5P PRPP PRA Synthesis Pathway PRPS1
Figure 1 Metabolism of azathioprine, thioguanine, and mercaptopurine.41 Permission has been given by PharmGKB and Stanford to use figure (https://www.pharmgkb.org/pathway/PA2040). Pathway images and data are available under a Creative Commons BY-SA 4.0 license. alleles, and so genotyping tests including these three variants have wild-type patients, only about 30–60% of TPMT heterozygotes a high likelihood of being informative for TPMT phenotype.5,6 cannot tolerate full doses of mercaptopurine or azathioprine.8,10,11 Complementary phenotype laboratory tests can be helpful adjuncts Good thiopurine tolerance in some heterozygotes may be because, to genotyping tests (Supplement, Other Considerations).7 although they have higher TGNs than homozygous wild-type pa- TPMT catabolizes mercaptopurine to an inactive methylmercap- tients, they have lower concentrations (and, thus, fewer toxic effects) topurine base, leaving less parent drug available for eventual anab- of the MeMPNs than do normal metabolizers, which may offset the olism to active thioguanine nucleotides (TGNs; Figure 1). The toxic effects of having higher TGNs. Thus, there is less of a consensus secondary metabolite of mercaptopurine, thioinosine monophos- over how to dose azathioprine and mercaptopurine in patients who phate (TIMP), is also a substrate for TPMT, and methyl-TIMP are heterozygous for TPMT compared with those who are homozy- (and its further phosphorylated metabolites, methylmercaptopurine gous, although they are at a higher risk for toxicity compared with nucleotides (MeMPN)) have pharmacologic activity (mostly immu- patients carrying two normal function alleles.12 nosuppressive), inhibit de novo purine synthesis, and may contribute Although there is lower affinity between thioguanine and to some of the adverse effects of mercaptopurine, generally hepato- TPMT than between mercaptopurine and TPMT, TPMT sig- toxicity.2,8,9 Individuals who inherit two loss-of-function TPMT nificantly affects thioguanine pharmacokinetics and its cytotoxic alleles (homozygous or compound heterozygous TPMT deficient effects.12–16 Thioguanine is directly metabolized by TPMT to individuals) are at very high risk for life-threatening myelosuppres- inactive methylthioguanine base, leaving less drug available for sion, due to very high TGNs, if given conventional doses of mer- anabolism by HPRT and other enzymes to active TGN metabo- captopurine (or azathioprine). Despite having higher TGNs than lites. There is not a pharmacologically active secondary metabolite
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of thioguanine to undergo activation via TPMT (i.e., there are no based on the most common * allele diplotypes, and these assign- methyl-TIMP or methylmercaptopurine nucleotides). As a result, ments are used to link genotypes with thiopurine prescribing rec- patients receiving thioguanine are able to tolerate substantially ommendations. Of note, the phenotype of “possible intermediate higher TGN concentrations than do those receiving mercapto- metabolizer” has been introduced to this guideline to describe an purine or azathioprine.15 Within each TPMT phenotypic group, individual carrying one uncertain/unknown function allele plus the initial recommended relative dosage decreases are similar for one known no function allele, as this individual should be treated thioguanine, mercaptopurine, and azathioprine (Table 2). with “at least” the same precautions as would apply to an interme- diate metabolizer. Although inactivating TPMT and NUDT15 NUDT15. Through agnostic genomewide association studies, alleles have been identified in multiple populations (TPMT variants in NUDT15 have been identified that strongly influence Frequency Table21,22 and NUDT15 Frequency Table21,23), one thiopurine tolerance in patients with acute lymphoblastic leukemia of the limitations inherent in a commercial genotype-only test is (ALL)17 and those with inflammatory bowel diseases.18 As a that rare or previously undiscovered variants may not be included. nucleoside diphosphatase, NUDT15 catalyzes the conversion of cytotoxic thioguanine triphosphate (TGTP) metabolites to the Available genetic test options less toxic thioguanine monophosphate. Defects in NUDT15- See Supplement and the Genetic Testing Registry (https://www. mediated degradation of TGTP result in more TGTP available for ncbi.nlm.nih.gov/gtr/) for more information on commercially incorporation into DNA (DNA-TG; the primary antileukemic available clinical testing options. metabolite19), thus allowing for DNA damage and apoptosis. The SNP (rs116855232; c.415C>T) causing p.R139C was the first NUDT15 Incidental findings variant linked to thiopurine toxicity. It was shown that this There are no diseases or phenotypic traits that have been linked amino acid change results in a nearly complete loss of enzymatic to variation in TPMT or NUDT15 in the absence of thiopurine activity and protein stability in vitro. Patients carrying this allele treatment.2 showed excessive DNA-TG and severe myelosuppression.20 In children with ALL, patients homozygous for the p.R139C variant Linking genetic variability to variability in drug-related allele tolerated only 8% of the standard dose of mercaptopurine, phenotypes whereas tolerated dose intensity was 63% and 83.5% for There is substantial evidence linking TPMT and NUDT15 those heterozygous and wildtype for this SNP, respectively.17 genotype with phenotypic variability (see Tables S1 and S2). Although most clinical studies focused on mercaptopurine, Pre-emptive dose adjustments based on TPMT genotype have re- in vitro experiments using laboratory models indicated similar duced thiopurine-induced adverse effects without compromising influence of NUDT15 on the cytotoxicity of azathioprine and desired antitumor and immunosuppressive therapeutic effects in thioguanine.20 Additional variant alleles have been identified with several clinical settings (Table S1). Similarly, retrospective studies varying prevalence among differing ancestral groups and varying strongly indicate that patients with loss-of-function NUDT15 al- degrees of functional effects (NUDT15 Allele Functionality leles are at excessive risk of thiopurine toxicity if the standard dose Table and Frequency Table)21. The variant p.R139C has been is administered. This body of evidence, rather than randomized studied most extensively in patients receiving thiopurine therapy, clinical trials, provides the basis for most of the dosing recommen- thus providing the strongest evidence for clinical implementation. dations in Tables 2 and 3. Subsequent studies reported additional variants, most of which are rare, and their associations with clinical thiopurine toxicity do not Therapeutic recommendations rise to clinical actionability at this point, even though some showed Thiopurines are used to treat malignant and nonmalignant condi- decreased NUDT15 activity in in vitro. For this reason, these tions, and, thus, the approach to dosing adjustments and the pro- variants (*4–*9) are designated as unclear function but may be pensity to initiate therapy at higher vs. lower starting doses based clarified as more data emerge. on TPMT/NUDT15 status may differ depending on the clinical Inherited TPMT deficiency is the primary genetic cause of thio- indication. Thiopurines have a unique role in the treatment of purine intolerance in Europeans and Africans, whereas risk alleles many malignancies. The “normal” starting doses of thiopurines in NUDT15 explain the majority of thiopurine-related myelosup- are generally “high” because they have been derived from trials pression in Asians and are also common in Hispanics. that have been heavily weighted by the ~ 90% of the population who are wildtype for TPMT and NUDT15 and receive maximal Genetic test interpretation tolerable doses by the standards of anticancer treatment (hence, Genetic testing analyzes the DNA sequence at specific SNP lo- full doses should be given to those who are normal metabolizers cations in the TPMT and NUDT15 genes (Supplement). Each for TPMT and NUDT15; Tables 2 and 3). Because the level of named star (*) allele is defined by the genotype at one or more thiopurine tolerance is highly correlated with genetic ancestry,17 specific loci (TPMT Allele Definition Table21,22 and NUDT15 the “normal” starting doses can also vary by geographic regions Allele Definition Table21,23) and is associated with a level of and clinical practice. enzyme activity (TPMT Allele Functionality Table21,22 and NUDT15 Allele Functionality Table21,23). Table 1 summarizes TPMT recommendation. If starting doses are already high (e.g., the assignment of the likely TPMT and NUDT15 phenotypes, 75 mg/m2 of mercaptopurine), as is true in some ALL treatment
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Table 1 Assignment of likely TPMT and NUDT15 phenotypes based on genotypes Likely phenotypea Genotypes Examples of diplotypes Assignment of likely TPMT phenotypes based on genotypes Normal metabolizer An individual carrying two normal function alleles *1/*1 Intermediate metabolizer An individual carrying one normal function allele *1/*2, *1/*3A, *1/*3B, *1/*3C, *1/*4 PLUS one no function allele Possible intermediate metabolizer An individual carrying one uncertain/unknown *2/*8, *3A/*7 function allele PLUS one no function allele Poor metabolizer An individual carrying two no function alleles *3A/*3A, *2/*3A, *3A/*3C, *3C/*4, *2/*3C, *3A/*4 Indeterminate An individual carrying two uncertain/unknown *6/*8 function alleles *1/*8 OR one normal function allele plus one uncertain allele function allele Assignment of likely NUDT15 phenotypes based on genotypes Normal metabolizer An individual carrying two normal function alleles *1/*1 Intermediate metabolizer An individual carrying one normal function allele *1/*2, *1/*3 PLUS one no function allele Possible intermediate metabolizer An individual carrying one uncertain function *2/*5, *3/*6 allele PLUS one no function allele Poor metabolizer An individual carrying two no function alleles *2/*2, *2/*3, *3/*3 Indeterminate An individual carrying two uncertain function *1/*4, *1/*5 alleles *4/*5, *5/*6 OR one normal function allele plus one uncertain function allele TPMT, thiopurine methyltransferase. NUDT15, Nudix (Nucleoside Diphosphate Linked Moiety X)-Type Motif 15 aSee TPMT and NUDT15 Frequency Table and Diplotype-Phenotype Table21–23 for estimates of phenotype frequencies among different ethnic/geographic groups and for a more comprehensive list of predicted metabolizer phenotypes. regimens, lower than normal starting doses should be considered carriers of TPMT vs. NUDT15 decreased function alleles,17 although in TPMT intermediate metabolizers11,15,24,25 and markedly there remains a paucity of multi-ethnic studies examining both TPMT reduced doses (10-fold reduction) should be used in TPMT poor and NUDT15 variants. Therefore, our NUDT15 recommendations metabolizers26 (Table 2). This approach has decreased the risk parallel those for TPMT. For NUDT15 normal metabolizers of acute toxicity without compromising relapse rate in ALL.27 (NUDT15*1/*1), starting doses do not need to be altered. For Even at these markedly reduced dosages, erythrocyte TGN NUDT15 intermediate metabolizers (e.g., NUDT15*1/*3; Table concentrations in TPMT poor metabolizers remain well above 2), reduced starting doses should be considered to minimize those tolerated and achieved by the majority of patients (who are toxicity, particularly if the starting doses are high (e.g., 75 mg/m2/ TPMT normal metabolizers).4,26 day for mercaptopurine). For NUDT15 poor metabolizers (e.g., In some nonmalignant conditions, alternative agents may be NUDT15*3/*3), substantially reduced doses (e.g., 10 mg/m2/ chosen for TPMT intermediate or poor metabolizers rather than day of mercaptopurine) or the use of an alternative agent should be reduced doses of thiopurines; if thiopurines are used, full starting considered (Table 2).20 doses are recommended for TPMT normal metabolizers, reduced As for TPMT, there is substantial variability in the tolerated doses (30–80% of target dose) in TPMT intermediate metaboliz- thiopurine dosages within NUDT15 intermediate metabolizers, ers,28,29 and substantially reduced doses (or use of an alternative with a minority of individuals who do not seem to require signifi- agent) in TPMT poor metabolizers (Table 2).4,30 cant dose reduction.17,20 Therefore, genotype-guided prescribing Some of the clinical data upon which dosing recommendations recommendations apply primarily to starting doses; subsequent are based (Table 2) rely on measures of TPMT phenotype rather dosing adjustments should be made based on close monitoring than genotype; however, because TPMT genotype is strongly of clinical myelosuppression (or disease-specific guidelines). In linked to TPMT phenotype,5–7,31 these recommendations apply contrast, a full dose of mercaptopurine poses a severe risk of pro- regardless of the method used to assess TPMT status. longed hematopoietic toxicity in NUDT15 poor metabolizers and pre-emptive dose reductions are strongly recommended.32,33 NUDT15 recommendation. Similar to TPMT, tolerated merca The NUDT15 poor metabolizer phenotype is observed at a ptopurine dosage is also correlated with the number of nonfunctional frequency of about 1 in every 50 patients of East Asian descent, alleles of the NUDT15 gene.17,18 In fact, the degree of thiopurine which is more common than the TPMT poor metabolizer pheno- intolerance (e.g., for mercaptopurine) is largely comparable between type in Europeans, and, thus, genotyping NUDT15 in the Asian
4 VOLUME 0 NUMBER 0 | Month 2019 | www.cpt-journal.com CPIC UPDATE b (Continues) Strong Moderate Classification of recommendations - /day 2 /day) (e.g., 2 a specific 4,16 4,16 state after state after mg/m is a Thioguanine /day) and adjust/day) 2 40–60 and adjust doses of and thioguanine of other myelosuppressive therapy without any special emphasis on thioguanine. Allow 2 weeks to reach steady- each dose adjustment. (e.g., 20–48(e.g., mg/ m doses of thioguanine based on degree of myelosuppression and disease- Allow guidelines. 2–4 weeks to reach steady- each dose adjustment. If myelosuppression occurs, and depending on other therapy, emphasis should be on thioguanine reducing over other agents. Start with normal starting dose Start with reduced doses (50–80% of normal dose) if normal starting dose ≥ 40–60 mg/m Dosing recommendaDosing thioguanine for tions
related related leukopenia, leukopenia, neutropenia, myelosuppression. leukopenia, neutropenia, myelosuppression. Lower Lower concentrations of TGN metabolites, but note that TGN after thioguanine are 5–10 × higher than TGN after mercaptopurine azathioprine. or Normal risk of thiopurine- Moderate to high concentrations of TGN metabolites; but note that TGN after thioguanine are 5–10 × higher than TGN after mercaptopurine azathioprine. or Increased risk of thiopurine- Implications for for Implications thioguanine phenotypic measures Strong Strong Classification of recommendations - 4,30,37 Azathioprine (e.g., a 4,30,37,38 specific specific state after each state after is 2–3 mg/kg/ a 2–3 mg/kg/day) and adjust doses of azathioprine based on disease- Allow guidelines. 2 weeks to reach steady- dose adjustment. Dosing recommendaDosing azathioprine for tions day 0.6–2.4 (e.g., mg/ kg/day), and adjust doses of azathioprine based on degree of myelosuppression and disease- Allow guidelines. 2–4 weeks to reach steady- each dose adjustment. Start with normal starting dose Start with reduced starting doses (30–80% of normal dose) if normal starting dose - Classification of recommenda tions Strong Strong / 2 - /day /day 2 /day /day 2 4,27,30 (e.g., a /day or /day 2 state after each state after each 4,11,15,24,25,27,30,38,39 is ≥ 75 mg/m a Mercaptopurine mg/m mg/kg/day (e.g.,or ≥ 1.5 mg/kg/day start at 22.5–60 mg/m 75 Dosing recommendaDosing mercaptopurine for tions If normal starting dose is already < 75 mg/m 1.5 mg/kg/day) and adjust doses of mercaptopurine of (and any other myelosuppressive therapy) without any special emphasis on mercaptopurine compared with other agents. Allow at least 2 weeks to reach steady- dose adjustment. mg/kg/day, dose or < 1.5 mg/kg/day, not be reduction may recommended. day or 0.45–1.2 mg/kg/ day and adjust doses of day) mercaptopurine based on degree of and myelosuppression disease- specific guidelines. Allow to reach 2–4 weeks steady- dose adjustment. If occurs,myelosuppression and depending on other emphasis should therapy, be on reducing othermercaptopurine over agents. Start with normal starting dose Start with reduced starting doses (30–80% of normal dose) if normal starting dose
related related
Implications for Implications for mercaptopurine and azathioprine phenotypic measures leukopenia, leukopenia, neutropenia, myelosuppression. Normal risk of thiopurine- Lower Lower concentrations of TGN metabolites, thishigher MeTIMP, is the “normal” pattern. Moderate to high concentrations of TGN metabolites; low concentrations of MeTIMP. Increased risk of thiopurine- leukopenia, neutropenia, myelosuppression.
Recommended dosing of thiopurines TPMT by phenotype
Phenotype TPMT normal metabolizer TPMT intermediate metabolizer OR TPMT possible intermediate metabolizer 2 Table
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a
16 4 fold and dose Thioguanine (reduce daily dose by 10- by instead thrice weekly of daily) and adjust doses of thioguanine based on degree of myelosuppression and disease- specific guidelines. Allow to reach 4–6 weeks steady- state after each dose adjustment. If myelosuppression emphasis occurs, should be on reducing thioguanine other agents. over nonmalignant For consider conditions, alternative nonthiopurine immunosuppressant therapy. Start with drastically reduced doses Dosing recommendaDosing thioguanine for tions
Extremely high Extremely high concentrations of TGN metabolites; fatal toxicity possible without decrease. dose increased Greatly risk of thiopurine- related leukopenia, neutropenia, myelosuppression. Implications for for Implications thioguanine phenotypic measures Strong Classification of recommendations - Azathioprine fold and 28,30,37,38,40 specific
state after by 10- a dose thrice weekly instead of daily) and adjust doses of azathioprine based on degree of myelosuppression and disease- Allow guidelines. 4–6 weeks to reach steady- each dose adjustment. Dosing recommendaDosing azathioprine for tions For nonmalignant nonmalignant For consider conditions, alternative nonthiopurine immunosuppressant therapy. For malignancy, start reduced drastically with doses (reduce daily dose - Classification of recommenda tions Strong / - 2 fold and . specific 4,26,30,38 state after each by 10- a Mercaptopurine reduce frequency to to frequency reduce thrice weekly instead of daily mg/m 10 (e.g., Dosing recommendaDosing mercaptopurine for tions day given just 3 days/ week) and adjust doses of mercaptopurine based on degree of myelosuppression and disease- Allow guidelines. 4–6 weeks to reach steady- dose adjustment. If myelosuppression occurs, emphasis should be on reducing mercaptopurine over other agents. For conditions, nonmalignant alternativeconsider nonthiopurine immunosuppressant therapy. For malignancy, start reduced drastically with doses (reduce daily dose
Implications for Implications for mercaptopurine and azathioprine phenotypic measures Extremely high Extremely high concentrations of TGN metabolites; fatal toxicity possible without dose decrease; no MeTIMP metabolites. increased Greatly risk of thiopurine- related leukopenia, neutropenia, myelosuppression. (Continued)
2 Rating scheme described Material in Supplemental Normal starting doses vary by race/ethnicity and treatment regimens. If standard dose is below normal recommended dose, dose reduction might not be recommended for intermediate metabolizers. Phenotype TPMT poor metabolizer MeTIMP, metabolitesMeTIMP, of thiopurine methyltransferase; TGN, thioguanine nucleotides;a TPMT, thiopurine methyltransferase. b Table
6 VOLUME 0 NUMBER 0 | Month 2019 | www.cpt-journal.com CPIC UPDATE b (Continues) Classification of recommendations Strong Moderate - / 2 4,16 Thioguanine 4,16 /day 2 (40– is a a /day). Adjust/day). 2 specific state after each state after each mg/m Start with normal starting dose Start with reduced doses (50–80% of normal dose) if normal starting dose 60 ≥ 40–60 mg/m (e.g., 20–48(e.g., mg/m Dosing recommendaDosing thioguanine for tions doses of thioguanine and of other myelosuppressive therapy without any special emphasis on thioguanine. Allow 2 weeks to reach steady- dose adjustment. day) and adjust doses of based thioguanine on degree of myelosuppression and disease- Allow guidelines. 2–4 weeks to reach steady- dose adjustment. If myelosuppression occurs, and depending on other therapy, emphasis should be on thioguanine reducing over other agents. Strong Strong Classification of recommendations is a - 4,30,37 Azathioprine (e.g., a 4,30,37,38 specific specific state after each state after each disease-
2–3 mg/kg/day) and adjust doses of azathioprine based on guidelines. Allow Allow guidelines. 2 weeks to reach steady- dose adjustment. Dosing recommendaDosing azathioprine for tions Start with reduced starting doses (30–80% of normal dose) if normal starting dose 2–3 mg/kg/day (e.g., 0.6–2.4 mg/kg/day), and adjust doses of azathioprine based on degree of myelosuppression and disease- Allow guidelines. 2–4 weeks to reach steady- dose adjustment. Start with normal starting dose Classification of recommendations Strong Strong
/ 2 /day 2 /day 2 4,27,30 specific 4,11,15,24,25,27,30,38,39 state after each state after each Mercaptopurine mg/m 75 (e.g., mg/m is ≥ 75 a a Dosing recommendations recommendations Dosing mercaptopurine for day or 0.45–1.2 mg/kg/ day) and adjust doses of mercaptopurine based on degree of myelosuppression and disease- Allow guidelines. 2–4 weeks to reach steady- dose adjustment. If myelosuppression occurs, and depending on other therapy, emphasis should be on reducing mercaptopurine other over agents. Start with reduced starting doses (30–80% of normal dose) if normal starting dose If normal starting dose is already mg/m2/day < 75 or < 1.5 mg/kg/day, dose reduction may not be recommended. Start with normal starting dose or ≥ 1.5 mg/kg/day (e.g., start at 22.5–60 mg/m or 1.5 mg/kg/day) and adjust doses of mercaptopurine of (and any other myelosuppressive therapy) without any special emphasis on mercaptopurine compared with other agents. Allow at least 2 weeks to reach steady- dose adjustment. related related Implications for for Implications thiopurine phenotypic measures leukopenia, leukopenia, neutropenia, myelosuppression Increased risk of thiopurine- Normal risk of thiopurine- leukopenia, neutropenia, myelosuppression
Recommended dosing of thiopurines NUDT15 by phenotype
Phenotype NUDT15 intermediate metabolizer OR NUDT15 possible intermediate metabolizer 3 Table NUDT15 normal metabolizer
CLINICAL PHARMACOLOGY & THERAPEUTICS | VOLUME 0 NUMBER 0 | Month 2019 7 CPIC UPDATE b Classification of recommendations Strong - Thioguanine and adjust a specific 4 state after each doses of thioguanine based on degree of myelosuppression and disease- Allow guidelines. 4–6 weeks to reach steady- dose adjustment. In setting of myelosuppression, emphasis should be on thioguanine reducing over other agents. For nonmalignant consider conditions, nonthiopurine alternative immunosuppressant therapy. Reduce doses of to 25% normal dose Dosing recommendaDosing thioguanine for tions Strong Classification of recommendations -
a Azathioprine 28,30,37,38,40 state after each fold) and adjust Dosing recommendaDosing azathioprine for tions (reduce daily dose by (reduce daily dose by 10- doses of azathioprine based on degree of and myelosuppression disease- specific guidelines. Allow to reach 4–6 weeks steady- dose adjustment. For nonmalignant For consider conditions, alternative nonthiopurine immunosuppressant therapy. malignant For start with conditions, drastically reduced normal daily doses Classification of recommendations Strong /day and /day 2 specific . 4,26,30,38 Mercaptopurine Dosing recommendations recommendations Dosing mercaptopurine for For malignancy, initiate dose mg/m at 10 adjust dose based on myelosuppression and disease- Allow guidelines. 4–6 weeks to reach steady state after each dose adjustment. If myelosuppression occurs, emphasis should be on reducing mercaptopurine over other agents. nonmalignant For consider conditions, nonthiopurine alternative immunosuppressant therapy. Implications for for Implications thiopurine phenotypic measures Greatly increased increased Greatly risk of thiopurine- related leukopenia, neutropenia, myelosuppression (Continued)
3 Rating scheme described Material in Supplemental Normal starting doses vary by race/ethnicity and treatment regimens. If standard dose is below normal recommended dose, dose reduction might not be recommended for intermediate metabolizers. Table Phenotype NUDT15 poor metabolizer a b
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Figure 2 Recommended starting doses of thiopurines by thiopurine methyltransferase (TPMT) and NUDT15 phenotype. aWhether a dose reduction is recommended from the starting dose depends on the level of the standard starting dose; for example, if the standard starting dose of mercaptopurine is 75 mg/m2/day or higher, then a lower starting dose may be considered in intermediate metabolizers and would be recommended in poor metabolizers, whereas if the starting dose is 50 mg/m2/day or lower, a reduced starting dose may not be necessary in intermediate metabolizers. bSee Table 2 for recommendation. cFor patients who are intermediate metabolizers for both TPMT and NUDT15, further dose reduction might be needed compared with those who are only intermediate metabolizers with respect to one gene (TPMT or NUDT15). populations may be of particular clinical importance. NUDT15 be considered. In addition, there may be other reasons underlying deficiency is also more prevalent in individuals of Hispanic ethnic- poor tolerance to thiopurines that are not related to TPMT or to ity, particularly those with high levels of Native American genetic NUDT15 genetic variation. ancestry.17 Complementary clinical laboratory tests are available to measure thiopurine metabolites in erythrocytes: TGNs (for mercaptopu- rine, azathioprine, and thioguanine) and MeMPNs (or MeTIMP) TPMT and NUDT15 recommendation. Figure 2 outlines the recom- for those on mercaptopurine or azathioprine (see Supplement mended course of action if both TPMT and NUDT15 genotypes are known. There have been reports of patients with intermediate for details on associations with TPMT). Erythrocyte TGNs or 34–36, but there metabolizer status for both TPMT and NUDT15 (i.e., compound MeMPNs are not related to NUDT15 genotypes is evidence that intermediate and poor metabolizers for NUDT15 intermediate metabolizers), and there was a trend for a lower thio- accumulate higher levels of DNA-TG than normal metabolizers purine tolerance in these individuals compared with intermediate given the same mercaptopurine dosage.20 Thus, currently available metabolizers for only TPMT or NUDT15. The two genes are inde- erythrocyte therapeutic drug monitoring tests do not distinguish pendent: the likelihood of an individual being an intermediate me- NUDT15 metabolizer phenotypes. tabolizer for both genes depends upon the population frequencies for variant alleles. For example, given an estimate of no function alleles Implementation of this guideline for NUDT15 of 11% and of no function alleles for TPMT of 2%, the frequency of the compound intermediate phenotype would be The guideline supplement contains resources that can be used estimated to be 0.2%. However, the evidence for a different starting within electronic health records to assist clinicians in applying ge- dosage recommendation for the compound intermediate metaboliz- netic information to patient care for the purpose of drug therapy ers remains limited. optimization (see Resources to incorporate pharmacogenetics into an electronic health record with clinical decision support sections of the Recommendations for incidental findings Supplement). Not applicable. POTENTIAL BENEFITS AND RISKS FOR THE PATIENT Other considerations The benefits of pre-emptive TPMT testing are that doses that are If test results are available for only one gene (TPMT or NUDT15, customized based on TPMT status reduce the likelihood of acute but not both), prescribing recommendations based on that gene’s myelosuppression without compromising disease control.4,8,24,25 results may be implemented with the caveat that the other gene’s The risks would be that a proportion of TPMT intermediate results are missing and may have important implications. The metabolizers may spend a period of time at lower thiopurine higher frequency of decreased function NUDT15 variants among doses than they can eventually tolerate, because only ~ 30–60% individuals of Asian and Hispanic backgrounds and of TPMT of TPMT heterozygous patients receiving conventional thiopu- variants in those with European and African backgrounds should rine doses experience severe myelosuppression.4,8,11 However,
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because steady state is reached in 2–4 weeks, any period of “un- CONFLICT OF INTEREST derdosing” should be short, and, using this approach, at least in The authors declared no competing interests for this work. ALL and in inflammatory bowel disease, outcomes were not 4,8,24,25,28 DISCLAIMER compromised. Clinical Pharmacogenetics Implementation Consortium (CPIC) Similar benefits are expected with pre-emptive NUDT15 geno- guidelines reflect expert consensus based on clinical evidence and typing, especially for Asian patients, given that these variants have peer-reviewed literature available at the time they are written and are intended only to assist clinicians in decision making, as well as comparable effects as risk alleles in TPMT. At least in ALL, leuke- to identify questions for further research. New evidence may have mia cells with no function NUDT15 alleles are also more sensitive to emerged since the time a guideline was submitted for publication. mercaptopurine20 and, thus, in theory, NUDT15 genotype guided Guidelines are updated periodically on https://cpicpgx.org/guidelines/ dosing would not compromise antileukemic efficacy of this drug. and it is the responsibility of the guideline user to consult this website 4 for updates. Guidelines are limited in scope and are not applicable A possible risk to the patient is an error in genotyping. As shown to interventions or diseases not specifically identified. Guidelines do in preclinical models, some TPMT and/or NUDT15 variants may not account for all individual variation among patients and cannot be not be included in the genotype test used, and patients with these vari- considered inclusive of all proper methods of care or exclusive of other treatments. It remains the responsibility of the healthcare provider to ants may be assigned a “wildtype” (*1) genotype by default. Thus, an determine the best course of treatment for the patient. Adherence to assigned “wildtype” allele could potentially harbor a no or decreased any guideline is voluntary, with the ultimate determination regarding function variant. Because genotypes are life-long test results, any such its application to be solely made by the clinician and the patient. CPIC assumes no responsibility for any injury to persons or damage error could stay in the medical record for the life of the patient. to property related to any use of CPIC’s guidelines, or for any errors or omissions. CAVEATS: APPROPRIATE USE AND/OR POTENTIAL MISUSE OF GENETIC TESTS © 2019 American Society for Clinical Pharmacology and Therapeutics Most of the time, thiopurines are given orally daily for a period of at least several months. Genotype-based starting doses are just that— starting doses, and, in most diseases, titration to the desired degree (or 1. Sandborn, W.J. Pharmacogenomics and IBD: TPMT and thiopu- lack thereof ) of myelosuppression is required. Thus, clinicians must rines. Inflamm. Bowel Dis. 10(suppl. 1), S35–S37 (2004). continue to evaluate markers of disease progression and/or of myelo- 2. Evans, W.E. Pharmacogenetics of thiopurine S-methyltransferase suppression to adjust thiopurine doses up or down from the genotype- and thiopurine therapy. Ther. Drug Monit. 26, 186–191 (2004). 3. Weinshilboum, R. Inheritance and drug response. N. Engl. J. Med. directed starting doses. One caveat is that some serious long-term 348, 529–537 (2003). adverse effects (secondary tumors) have been associated with defective 4. Ford, L.T. & Berg, J.D. Thiopurine S-methyltransferase (TPMT) TPMT activity without necessarily causing serious acute myelosup- assessment prior to starting thiopurine drug treatment: a pharmacogenomic test whose time has come. J. Clin. Pathol. 63, pression; whether capping doses of thiopurines in those with a TPMT 288–295 (2010). defect will decrease the risk of the late effect of secondary cancer is not 5. Schaeffeler, E. et al. Comprehensive analysis of thiopurine known (see Supplement for additional information). Some adverse re- S-methyltransferase phenotype-genotype correlation in a large population of German-Caucasians and identification of novel actions to thiopurines, such as pancreatitis and hepatotoxicity, are not TPMT variants. Pharmacogenetics 14, 407–417 (2004). related to low TPMT activity. 6. Yates, C.R. et al. Molecular diagnosis of thiopurine S-methyltrans The discovery and clinical implementation of NUDT15 vari- ferase deficiency: genetic basis for azathioprine and mercaptopu- ants in thiopurine dosing is relatively recent, and the exact im- rine intolerance. Ann. Intern. Med. 126, 608–614 (1997). 7. Liu, C. et al. Genomewide approach validates thiopurine methyl- pact of NUDT15 genotype-guided dose adjustments on toxicity transferase activity is a monogenic pharmacogenomic trait. Clin. and efficacy are less clear compared with TPMT. Pharmacol. Ther. 101, 373–381 (2017). 8. Relling, M.V. et al. Mercaptopurine therapy intolerance and hetero- SUPPORTING INFORMATION zygosity at the thiopurine S-methyltransferase gene locus. J. Natl. Supplementary information accompanies this paper on the Clinical Cancer Inst. 91, 2001–2008 (1999). Pharmacology & Therapeutics website (www.cpt-journal.com) and 9. Nygaard, U., Toft, N. & Schmiegelow, K. Methylated metabolites https://cpicpgx.org/guidelines/guideline-for-thiopurines-and-tpmt/. of 6-mercaptopurine are associated with hepatotoxicity. Clin. Pharmacol. Ther. 75, 274–281 (2004). Supplement. Clinical Pharmacogenetics Implementation Consortium 10. Evans, W.E. et al. Preponderance of thiopurine S- (CPIC) guideline for thiopurine dosing based on TPMT and NUDT15 gen- methyltransferase deficiency and heterozygosity among patients otypes: 2018 update. intolerant to mercaptopurine or azathioprine. J. Clin. Oncol. 19, 2293–2301 (2001). 11. Stocco, G. et al. Genetic polymorphism of inosine triphosphate ACKNOWLEDGMENTS pyrophosphatase is a determinant of mercaptopurine metabolism We acknowledge the critical input of members of the Clinical Pharmaco and toxicity during treatment for acute lymphoblastic leukemia. genetics Implementation Consortium (CPIC) of the Pharmacogenomics Clin. Pharmacol. Ther. 85, 164–172 (2009). Research Network (PGRN), and U24HG010135, R01GM118578, and 12. Higgs, J.E., Payne, K., Roberts, C. & Newman, W.G. Are patients P50GM115279. and the PharmVar TPMT and NUDT15 expert panels with intermediate TPMT activity at increased risk of myelosup- especially Andrea Gaedigk, PhD. This work is also supported by the pression when taking thiopurine medications? Pharmacogenomics Robert Bosch Foundation, Stuttgart, Germany. 11, 177–188 (2010). 13. Hartford, C. et al. Differential effects of targeted disruption of FUNDING thiopurine methyltransferase on mercaptopurine and thioguanine This guideline is funded by the National Institutes of Health (CPIC pharmacodynamics. Cancer Res. 67, 4965–4972 (2007). (R24GM115264 and U24HG010135) and PharmGKB (R24GM61374, 14. Hosni-Ahmed, A., Barnes, J.D., Wan, J. & Jones, T.S. Thiopurine R01GM118578, and P50GM115279)). methyltransferase predicts the extent of cytotoxicity and DNA
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damage in astroglial cells after thioguanine exposure. PLoS One atopic eczema: a double-blind, randomised controlled trial. Lancet 6, e29163 (2011). 367, 839–846 (2006). 15. Lennard, L. & Lilleyman, J.S. Individualizing therapy with 29. Coenen, M.J. et al. Identification of patients with variants in TPMT 6-mercaptopurine and 6-thioguanine related to the thiopurine and dose reduction reduces hematologic events during thiopurine methyltransferase genetic polymorphism. Ther. Drug Monit. 18, treatment of inflammatory bowel disease. Gastroenterology 149, 328–334 (1996). 907–917 e7 (2015). 16. McBride, K.L., Gilchrist, G.S., Smithson, W.A., Weinshilboum, 30. Sandborn, W.J. Rational dosing of azathioprine and R.M. & Szumlanski, C.L. Severe 6-thioguanine-induced marrow 6-mercaptopurine. Gut 48, 591–592 (2001). aplasia in a child with acute lymphoblastic leukemia and inhibited 31. Tamm, R. et al. Polymorphic variation in TPMT is the principal deter- thiopurine methyltransferase deficiency. J. Pediatr. Hematol. minant of TPMT phenotype: a meta-analysis of three genome-wide Oncol. 22, 441–445 (2000). association studies. Clin. Pharmacol. Ther. 101, 684–695 (2017). 17. Yang, J.J. et al. Inherited NUDT15 variant is a genetic determinant 32. Zhu, Y. et al. Combination of common and novel rare NUDT15 vari- of mercaptopurine intolerance in children with acute lymphoblas- ants improves predictive sensitivity of thiopurine-induced leukope- tic leukemia. J. Clin. Oncol. 33, 1235–1242 (2015). nia in children with acute lymphoblastic leukemia. Haematologica 18. Yang, S.K. et al. A common missense variant in NUDT15 confers 103, e293–e295 (2018). susceptibility to thiopurine-induced leukopenia. Nat. Genet. 46, 33. Ailing, Z., Jing, Y., Jingli, L., Yun, X. & Xiaojian, Z. Further evidence 1017–1020 (2014). that a variant of the gene NUDT15 may be an important predic- 19. Nielsen, S.N. et al. DNA-thioguanine nucleotide concentration and tor of azathioprine-induced toxicity in Chinese subjects: a case relapse-free survival during maintenance therapy of childhood report. J. Clin. Pharm. Ther. 41, 572–574 (2016). acute lymphoblastic leukaemia (NOPHO ALL2008): a prospective 34. Moriyama, T. et al. The effects of inherited NUDT15 polymor- substudy of a phase 3 trial. Lancet Oncol. 18, 515–524 (2017). phisms on thiopurine active metabolites in Japanese children 20. Moriyama, T. et al. NUDT15 polymorphisms alter thiopurine metabo- with acute lymphoblastic leukemia. Pharmacogenet. Genomics 27, lism and hematopoietic toxicity. Nat. Genet. 48, 367–373 (2016). 236–239 (2017). 21. Clinical Pharmacogenetics Implementation Consortium (CPIC). 35. Lee, J.H. et al. Measurements of 6-thioguanine nucleotide levels CPIC Guideline for Thiopurines and TPMT and NUDT15.
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