Thiopurine metabolites variations during co-treatment with aminosalicylates for inflammatory bowel disease:Effect of N-acetyl transferase polymorphisms

AIM: To evaluate variation of the concentration of thiopurine metabolites after 5-aminosalicylate(5-ASA) interruption and the role of genetic polymorphisms of N-acetyl transferase(NAT) 1 and 2. METHODS: Concentrations of thioguanine nucleotides(TGN) and methymercaptopurine nucleotides(MMPN), metabolites of thiopurines, were measured by high performance liquid chromatography in 12 young patients(3 females and 9 males, median age 16 years) with inflammatory bowel disease(6 Crohn's disease and 6 ulcerative colitis) treated with thiopurines(7 mercaptopurine and 5 azathioprine) and 5-ASA. Blood samples were collected one month before and one month after the interruption of 5-ASA. DNA was extracted and genotyping of NAT1, NAT2, inosine triphosphate pyrophosphatase(ITPA) and thiopurine methyl transferase(TPMT) genes was performed using PCR assays. RESULTS: Median TGN concentration before 5-ASA interruption was 270 pmol/8 x 108 erythrocytes(range: 145-750); after the interruption of the aminosalicylate, a 35% reduction in TGN mean concentrations(absolutemean reduction 109 pmol/8 × 108 erythrocytes) was observed(median 221 pmol/8 × 108 erythrocytes, range: 96-427, P value linear mixed effects model 0.0011). Demographic and clinical covariates were not related to thiopurine metabolites concentrations. All patients were wild-type for the most relevant ITPA and TPMT variants. For NAT1 genotyping, 7 subjects presented an allele combination corresponding to fast enzymatic activity and 5 to slow activity. NAT1 genotypes corresponding to fast enzymatic activity were associated with reduced TGN concentration(P value linear mixed effects model 0.033), putatively because of increased 5-ASA inactivation and consequent reduced inhibition of thiopurine metabolism. The effect of NAT1 status on TGN seems to be persistent even after one month since the interruption of the aminosalicylate. No effect of NAT1 genotypes was shown on MMPN concentrations. NAT2 genotyping revealed that 6 patients presented a genotype corresponding to fast enzymatic activity and 6 to slow activity; NAT2 genotypes were not related to thiopurine metabolites concentration in this study. CONCLUSION: NAT1 genotype affects TGN levels in patients treated with thiopurines and aminosalicylates and could therefore influence the toxicity and efficacy of these drugs; however the number of patients evaluated is limited and this has to be considered a pilot study.

Failure of interferon-γ pre-treated mesenchymal stem cell treatment in a patient with crohn's disease

Mesenchymal stem cells(MSC) are cells of stromal origin which exhibit unlimited self-renewal capacity and pluripotency in vitro.It has recently been observed that MSC may also exert a profound immunosuppressive and anti-inflammatory effect both in vitro and in vivo with consequent potential use in autoimmune disorders.We present the case of a patient suffering from childhood-onset, multidrug resistant and steroiddependent Crohn's disease who underwent systemic infusions of MSC, which led to a temporary reduction in CCR4, CCR7 and CXCR4 expression by T-cells, and a temporary decrease in switched memory B-cells, In addition, following MSC infusion, lower doses of steroids were needed to inhibit proliferation of the patient's peripheral blood mononuclear cells.Despite these changes, no significant clinical benefit was observed, and the patient required rescue therapy with infliximab and subsequent autologous hematopoietic stem cell transplantation.The results of biological and in vitro observations after MSC use and the clinical effects of infusion are discussed, and a brief description is provided of previous data on MSC-based therapy in autoimmune disorders.

Pharmacogenetics of azathioprine in inflammatory bowel disease: A role for glutathione-S-transferase?

Azathioprine is a purine antimetabolite drug commonly used to treat inflammatory bowel disease (IBD). In vivo it is active after reaction with reduced glutathione (GSH) and conversion to mercaptopurine. Although this reaction may occur spontaneously, the presence of isoforms M and A of the enzyme glutathione-S-transferase (GST) may increase its speed. Indeed, in pediatric patients with IBD, deletion of GST-M1, which determines reduced enzymatic activity, was recently associated with reduced sensitivity to azathioprine and reduced production of azathioprine active metabolites. In addition to increase the activation of azathioprine to mercaptopurine, GSTs may contribute to azathioprine effects even by modulating GSH consumption, oxidative stress and apoptosis. Therefore, genetic polymorphisms in genes for GSTs may be useful to predict response to azathioprine even if more in vitro and clinical validation studies are needed.