There were no significant differences in thiopurine dose between patients who did and did not carry an ABCC4 2269A allele and all patients had normal TPMT activity.42 However, white blood cell counts were significantly lower and red blood cell 6-TGN concentrations were significantly higher in ABCC4 2269A carriers compared to wild type homozygotes.

Furthermore, the incidence of leucopenia tended to be higher in patients with the variant allele (20.6% vs 8.3% of 2269G homozygotes, P = 0.053).42 These findings suggest that some cases of 6-TGN toxicity, in the absence of TPMT deficiency, may be explained by impaired efflux of this metabolite out of cells. Prospective testing of IBD patients for ABCC4 2269G>A may be useful in Asian populations where this SNP occurs at appreciable frequencies. Methotrexate, CHIR-99021 in vitro an analog of dihydrofolic acid (DHF), enters cells primarily through the reduced folate carrier

1 (RFC1)43 (Fig. 2). Once within the cell, methotrexate (also known as MTX-Glu1 because it is a monoglutamate) is quickly converted to its active polyglutamated forms (MTXGlu2-5) by the enzyme folylpolyglutamate synthase (FPGS).43 This sequential addition of glutamate residues prevents methotrexate efflux selleck kinase inhibitor from the cell via a range of multi-drug resistance proteins.43 Glutamation can be reversed by γ-glutamyl hydrolase (GGH), and the balance between these two enzymes determines how long methotrexate is retained within a cell and its efficacy.43 MTXGlu2-5 inhibit the folate pathway by displacing DHF as the preferred substrate of the folate-dependent enzymes DHF reductase (DHFR),44 thymidylate synthase (TYMS)45 and 5-aminoimidazole-4-carboxamide 4-Aminobutyrate aminotransferase ribonucleotide formyltransferase/IMP cyclohydrolase (ATIC).46 At high dose, MTX has an anti-proliferative effect and this has been attributed to MTXGlu2-5

blocking DHFR and TYMS-mediated nucleic acid methylation and pyrimidine synthesis.46 However, inhibition of these enzymes does not explain the anti-inflammatory effect of low dose MTX. Instead this has been attributed to the inhibition of ATIC which, in turn, leads to the accumulation and eventual release of adenosine from the cell.47 The subsequent binding of extracellular adenosine to adenosine receptors on target cells prevents the production of the pro-inflammatory cytokines while promoting transcription of the anti-inflammatory agents.47 Many genetic polymorphisms have been documented in the folate pathway and an ever-increasing number of studies on patients with rheumatoid arthritis48,49 and patients with hematological malignancies50 have reported association of folate pathway polymorphisms with methotrexate response and toxicity. However, discordance among studies is common, and associations observed in one cohort seldom replicate in another.