The thiopurine drugs azathioprine (AZA) and 6-mercaptopurine (6-MP) are effective for the treatment of inflammatory bowel disease (IBD) and their prescription is increasing. Haematotoxicity, which can lead to potentially life threatening bone marrow suppression, represents the most serious side effect of thiopurine therapy. It has been attributed to the accumulation of active cytotoxic metabolites of AZA/6-MP, collectively called 6-thioguanine nucleotides, resulting from a deficiency in thiopurine catabolism specifically catalysed by the thiopurine S-methyltransferase (TPMT) enzyme. Genotyping tests are now available to identify deficient and intermediate methylators who are, respectively, homozygous and heterozygous for non-functional alleles of the TPMT gene. As pointed out by Lennard in the leading article (Gut 2002;51:143–6), it is clear that myelosuppression may be caused by other factors in addition to variable TPMT.
Since the identification of the molecular basis of inosine triphosphate pyrophosphatase (ITPAse) deficiency,1 a clinically benign condition characterised by abnormal accumulation of inosine triphosphate in erythrocytes, the possibility of a correlation between thiopurine toxicity and ITPAse deficiency has been raised. Complete ITPase deficiency was found to be associated with a homozygous missense 94C>A mutation that encodes a Pro32Thr exchange, whereas an intronic IVS2+21A>C polymorphism was shown to have a less severe effect, homozygotes retaining 60% ITPAse activity. It was then postulated that in ITPAse deficient patients treated with thiopurine drugs, a 6-thio-ITP metabolite could accumulate resulting in toxicity.1 A recent study in 62 patients with inflammatory bowel disease reported a significant association between the ITPA 94C>A polymorphism and AZA related adverse effects, specifically flu-like symptoms, rash, and pancreatitis.2 No correlation was observed with occurrence of neutropenia but only 11 patients were studied. We previously reported TPMT genotype analysis in 41 Crohn’s disease (CD) patients who had experienced leucopenia during AZA/6-MP therapy.3 Even though this study confirmed the efficiency of TPMT genotyping in identifying patients at risk of developing myelosuppression, it also highlighted its limitations, as only 27% of patients carried mutant alleles of the TPMT gene that were associated with enzyme deficiency. This prompted us to investigate the occurrence of ITPA mutations in this series of patients in order to evaluate whether genotyping of the ITPAse gene could improve the detection rate of patients at risk of thiopurine myelotoxicity.
Our population comprising 41 patients with CD has been described in detail previously.3 Briefly, all patients had either leucopenia (white blood cell count <3000/mm3; n = 24) or thrombocytopenia (platelets <100 000/mm3; n = 30), or both (n = 14), leading either to discontinuation of treatment or reduction of dose by 50% or more during AZA (n = 33) or 6-MP (n = 8) treatment. Patients were genotyped for the ITPA 94C>A and IVS2+21A>C mutations according to a previously described procedure based on endonuclease digestion of polymerase chain reaction products.1 Distribution of the 41 patients according to their ITPA genotype is presented in table 1 ▶ and compared with that of a previously published control population of 100 healthy Caucasians.1 Allele frequencies in the CD population were 0.085 for the 94C>A mutation and 0.12 for the IVS2+21A>C mutation, similar to frequencies observed in the control population (0.06 and 0.13, respectively). There was no significant difference in the genotypes distribution between the two populations, which confirmed the lack of association between ITPAse deficiency and myelosuppression during thiopurine therapy. Due to the retrospective nature of the study, no correlation with other side effects could be investigated.
Table 1.
Distribution of ITPA genotypes in 41 Crohn’s disease (CD) patients and 100 healthy Caucasians
| ITPA genotype | CD patients (n = 41) | Control population† (n = 100) |
| Wt/Wt | 26 (0.63)* | 64 (0.64) |
| Wt/94C>A | 6 (0.15) | 10 (0.10) |
| Wt/IVS2+21A>C | 7 (0.17) | 24 (0.24) |
| 94C>A/94C>A | 0 (0.00) | 0 (0.00) |
| IVS2+21A>C/IVS2+21A>C | 1 (0.02) | 0 (0.00) |
| 94C>A/IVS2+21A>C | 1 (0.02) | 2 (0.02) |
*Values in parentheses represent genotype frequencies.
†The control population comprised 100 healthy Caucasians who were genotyped in a previous study.1
In conclusion, application of ITPA genotyping tests does not seem to improve the identification of patients at risk of myelosuppression with AZA/6-MP therapy. Although we believe that conventional TPMT genotyping tests should still be applied before the initiation of thiopurine treatment, further work is needed on the role of other candidate genes that may be involved in thiopurine haematotoxicity.
Acknowledgements
We thank N Ferrari and A Vincent for their assistance in performing the study and the members of the GETAID for recruiting patients in the study.
Competing Interests: None declared.
References
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