Prospective Evaluation of the Pharmacogenetics of Azathioprine in the Treatment of Inflammatory Bowel Disease

A. Ansari; M. Arenas; S. M. Greenfield; D. Morris; J. Lindsay; K. Gilshenan; M. Smith; C. Lewis; A. Marinaki; J. Duley; J. Sanderson

Disclosures

Aliment Pharmacol Ther. 2008;28(8):973-983. 

In This Article

Discussion

This is the largest prospective evaluation of using full-dose AZA (2 mg/kg) and without dose adjustment in IBD patients. The study design minimized the effect of variable dosing on accurate evaluation of the pharmacokinetics of TGNs and side effects. Previous studies have demonstrated the increased risk of thiopurine toxicity in patients with TPMT deficiency in a variety of clinical settings, mainly haematological malignancy,[12] transplantation [30] and autoimmune disease.[31] In these studies, the chances of adverse effects varied from 20% to 35%.[32,33] However, the retrospective nature of most of these studies may not have provided a true measure of individual risk. Our study is one of the few comprehensive prospective studies and one could argue that the risk demonstrated (80% withdrawal) is a more accurate reflection of the actual risk of full-dose AZA therapy in those with intermediate TPMT activity.

The influence of genetic variation in TPMT activity on individual metabolism of thiopurines remains one of the classical applications of pharmacogenetics in modern medicine. Despite this, the uptake of TPMT testing remains variable, particularly in Europe.[34,35] Whilst pharmacoeconomic arguments for pretreatment TPMT testing are favourable[36,37,38,39,40,41,42] and numerous retrospective studies demonstrate the risks of exposure for TPMT deficiency individuals ('very low and intermediate') to standard doses of AZA or MP,[43,44,45,46,47] some uncertainty may stem from the lack of prospective studies of the influence of TPMT. Furthermore, emphasis on the fact that TPMT deficiency only explains a small proportion of overall thiopurine toxicity[19] has added to a misplaced belief that prior knowledge of TPMT activity adds little to conventional practice.

However, the results of the study confirm the major influence of TPMT carrier status on risk of AZA toxicity with most TPMT heterozygotes withdrawing due to early-onset adverse events. Only two TPMT heterozygotes continued AZA beyond the 6 month study period, one developed myelotoxicity at 9 months and the second was a self-confessed poor adherer. An important finding is that most patients with intermediate TPMT activity withdrew due to gastric intolerance within 6 weeks of treatment and this was particularly caused by nausea. This is likely to explain why myelotoxicity (which first occurred at 12 weeks) is not encountered more often amongst TPMT heterozygotes.

Equally, genetic variation in the activity of other enzymes may explain some of the toxicity to thiopurines that is not accounted for by variation in TPMT activity. In this study, we examined the influence of the two functional ITPase polymorphisms. We discovered a clear association between presence of the ITPA 94C>A mutation and drug withdrawal due to the flu-like illness. This relationship with adverse events,[21] withdrawal of treatment[23] and myelotoxicity[48] is supported by other studies. The mechanism of toxicity due to ITPase deficiency is uncertain, but toxicity could be predicted, perhaps as a result of interruption of a variety of nucleotide-dependent reactions from thio-ITP accumulation. As such, general toxicity symptoms such as flu-like illness might be expected rather than other more specific toxicities. In fact, the benefit of thioguanine in patients 'allergic' to AZA/6MP[49] could well be explained by the effective by-passing of ITPase with thioguanine. Whilst the influence of ITPase deficiency appears small, it is definite and may yet prove of sufficient value to merit pretreatment testing, if only to highlight a need for vigilance regarding toxicity and permit prompt an alternative therapeutic approach.

Most studies of the influence of TPMT have focused on the prediction of toxicity in those with genetic TPMT deficiency. However, high TPMT activity could also predict poor clinical response as a consequence of diversion to methylation of MP in preference to bioactivation to TGNs.[50] In this study, we have confirmed that prior knowledge of TPMT is an important predictor of clinical response. Patients with a high TPMT (>35 pmol/h/mg/Hb) have a significantly reduced chance of responding to AZA. This cut-off echoes earlier finding by our group and others of the influence of TPMT activity on AZA efficacy in a both a renal transplant setting[51] and IBD.[11,16] Prior knowledge of TPMT activity in the higher range indicates that empirical dosing (2-2.5 mg/kg) may be inadequate prompting earlier use of higher doses (i.e. 2.5-3 mg/kg) or alternative strategies to avoid lengthy periods of non-response. However, in a proportion of these patients, methylation may remain dominant making dose escalation futile or result in hepatotoxicity.[15,52] Hence, a very high TPMT might be a more reliable predictor of a need for co-therapy with allopurinol[52] or an alternative immunosuppressive.

We measured TGN concentrations throughout the study period and discovered a significant correlation between good clinical response and concentrations greater than 100 pmol/8 × 108 RBC. This level is considerably lower than that determined in other published series which have reported response cut-off values of about 250 pmol/8 × 108 RBC.[13] The difference is related to extraction and processing of TGNs (intact nucleotides by ion-pair HPLC) that was used in our study.[29] This method is considerably different from other reported TGN assays[53,54] and results in different reference ranges.[55] The optimal method for measurement of TGNs remains an unresolved question.

Our large prospective data set indicates that TGN level of above 100 pmol/8 × 108 RBC should predict a reasonable chance of successful clinical response. In practice, TGN concentrations are difficult to use as a guide to optimal dosing and uptake has therefore been poor in the UK. We would propose that the best use is still as an indicator of drug adherence and secondly, as a guide to the feasibility of dose escalation in the event of non-response (Data S3).

In our study there were several potential weaknesses. This includes the unexplained high incidence of adverse events (38%) compared with earlier studies.[6] However, recent large studies report a similar frequency of adverse events as ours,[17,56] and the reason for this remains uncertain (Data S4).

In conclusion, this large prospective study has demonstrated very clearly that patients with intermediate TPMT activity are highly likely to withdraw from standard-dose AZA treatment due to early side effects. This is frequently due to nausea and gastric intolerance prior to the onset of myelotoxicity (median time of onset 12 weeks). We strongly recommend that these individuals receive appropriately reduced AZA dose (1 mg/kg) to avoid toxicity. Furthermore, we have shown that, as one would expect, pretreatment TPMT activity is a strong predictor of clinical response. A significant association between adverse events, specifically flu-like symptoms, was found for the ITPA 94C>A polymorphism. Finally, the study also shows a strong correlation with successful outcome on AZA if TGN concentrations exceed 100 pmol/8 × 108 RBC. Pretreatment TPMT testing offers significant value to doctor and patient. The role of TGN monitoring is less certain but the results of this study add weight to the view that TGN monitoring is of clinical value.


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