Practice Essentials
Azathioprine is a purine analogue that interferes with DNA synthesis and inhibits the proliferation of quickly growing cells, especially cells of the immune system. It is used as an immunosuppressant in patients undergoing organ transplantation, and its metabolite 6-mercaptopurine is used in the treatment of autoimmune diseases and acute lymphoblastic leukemia. During metabolism, hypoxanthine-guanine phosphoribosyltransferase (HGPRT) converts 6-mercaptopurine to cytotoxic 6-thioguanine nucleotide analogues, while thiopurine methyltransferase (TPMT) inactivates 6-mercaptopurine through methylation to form 6-methylmercaptopurine. In patients with TPMT deficiency, active 6-mercaptopurine accumulates, and a larger proportion of 6-mercaptopurine is converted to the cytotoxic 6-thioguanine nucleotide analogues, which can lead to bone marrow toxicity and myelosuppression. [1, 2, 3, 4, 5, 6, 7]
Most of the variant alleles of TPMT that have been identified are associated with decreased activity in vitro, but only a subset have known clinical effects. [8] TPMT*3A, the most common variant (5% frequency in Caucasians), includes 2 nonsynonymous coding changes: an exon 7 A154T variant and an exon 10 Y240C variant. The second most frequent variant, TPMT*3C, contains only the exon 10 variant and is more common in Asian populations (2% frequency). TPMT*3B occurs rarely and contains only the exon 7 variant. [9] TPMT*2 is a rare nonsynonymous variant (A80P) with reduced catalytic activity. [10] Other variants, such as TPMT*8, occur more commonly in African populations (2% frequency). [11, 4, 5]
Azathioprine is also commonly used in the management of inflammatory bowel disease (IBD). Thiopurine-induced myelosuppression (TIM) has a cumulative incidence of 7% and usually occurs within a few weeks of starting the drug. TPMT variants are only found in 25% of patients of European ancestry affected by TIM, suggesting the presence of other genetic and environmental determinants. Studies in patients of other populations identified variants in nudix hydrolase 15 (NUDT15) as risk factors for TIM. [12, 13] The Clinical Pharmacogenetics Implementation Consortium Guidelines updated in November 2018 suggest that both TPMT and NUDT15 genotypes should be evaluated for pharmacogenetic dosing of thiopurine. [24]
Enzymatic activity of TPMT in blood can be determined by a variety of methods, including HPLC (high-performance liquid chromatography), and radiochemical assays. Care must be taken to ensure assays measure activity, not concentration, since the common genetic polymorphisms effect enzymatic activity but not necessarily enzymatic levels. Note that TPMT testing cannot substitute for complete blood count monitoring.
Clinical Implications of the Genetic Mutation
Patients with TPMT deficiency treated with standard doses of azathioprine or 6-mercaptopurine are at significantly increased risk of myelosuppression, bleeding, infection, and death associated with increased levels of cytotoxic 6-thioguanine nucleotide levels in the red blood cells. [15, 16] Therefore, it is not recommended that patients with known TPMT deficiency be treated with azathioprine or 6-mercaptopurine; if treatment with these agents is deemed necessary, a low-dosage regimen should be used along with extremely careful hematologic monitoring.
Intermediate TPMT activity is associated with an increased risk of developing leukopenia, but there is significant clinical variability due to other modifying factors. [17] Nevertheless, lower azathioprine dosages have been suggested in some cases for patients with intermediate TPMT activity. [18] At a minimum, the drug should be administered with caution in these patients.
The US Food and Drug Administration (FDA) recommends but does not require genetic testing for TPMT. As noted on the product label for azathioprine and 6-mercaptopurine, TPMT genotype testing is recommended for the most common nonfunctional alleles, TPMT*2, *3A, and *3C, which account for the vast majority of patients with low or intermediate enzyme activity. For patients heterozygous for a nonfunctional TPMT allele, a reduced dose of azathioprine is recommended. Individuals with an inherited deficiency of the TPMT enzyme may be unusually sensitive to the myelosuppressive effects of thioguanine and prone to developing rapid bone marrow suppression following treatment initiation. Substantial dosage reductions or alternative nonthiopurine immunosuppressant therapy may be required to avoid the development of life-threatening bone marrow suppression in these patients. [5]
Phenotyping quantitates TPMT enzyme activity in erythrocytes, and based on the result, patients are classified as having normal, intermediate, or low TPMT activity. [1]
According to one study, more than 20% of patients with inflammatory bowel disease (IBD) discontinue thiopurine therapy because of severe adverse drug reactions (ADRs), with leukopenia being one of the most serious ADRs. This prospective study was performed to determine whether genotype analysis of TPMT before thiopurine treatment (and dose selection based on the results) would affect the outcomes of patients with IBD. According to the authors, screening for variants in TPMT did not reduce the proportions of patients with hematologic ADRs during thiopurine treatment for IBD, but there was a 10-fold reduction in hematologic ADRs in variant carriers who were identified and received a dose reduction, compared with variant carriers who did not, without differences in treatment efficacy. [19, 20]
In a randomized, controlled study of patients undergoing thiopurine therapy for IBD, selection of therapy based on genetic variants associated with thiopurine-induced leukopenia significantly reduced the proportion of patients with myelosuppression during treatment. Pretreatment genotype analysis also reduced numbers of outpatient clinic visit and numbers of patients with drug discontinuation or dose reductions. [21]
Updated guidelines suggest NUDT15 genotypes should also be evaluated for pharmacogenetic dosing of thiopurine. [24]
