Split-dose administration of thiopurine drugs: a novel and effective strategy for managing preferential 6-MMP metabolism.

BACKGROUND: Mercaptopurine and azathioprine (AZA) are efficacious in treating IBD. 6-tioguanine (6-TGN) levels correlate with therapeutic efficacy, whereas high 6-methylmercaptopurine (6-MMP) levels are associated with hepatotoxicity and myelotoxicity. Some IBD patients exhibit dose-limiting preferential 6-MMP production, which may lead to undesired side effects and impact efficacy. AIM: To review the outcomes of thiopurine split-dosing in patients with preferential 6-MMP metabolism. METHODS: A retrospective chart review of 179 IBD patients treated at the Cedars-Sinai IBD Center with AZA or mercaptopurine was performed. Preferential 6-MMP metabolisers with 6-MMP levels greater than 7000 pmol/8 × 10(8) erythrocytes who underwent split-dosing were identified and assessed for biochemical and clinical responses to these dose modifications. RESULTS: A total of 20 of 179 patients met the criteria for preferential 6-MMP metabolism and underwent thiopurine split-dosing. Dividing the total daily thiopurine dose led to a reduction in 6-MMP levels (11785 vs. 5324 pmol/8 × 10(8) erythrocytes; P < 0.0001) without negatively affecting clinical disease activity or 6-TGN levels (239 vs. 216 pmol/8 × 10(8) erythrocytes; P = N.S.) and led to resolution of 6-MMP associated side effects (elevated transaminases, leucopenia and flu-like symptoms) in all but two patients. After mean follow-up of 36 months, 12 patients remained in clinical remission on split-dose mercaptopurine. Five of the remaining eight patients escalated to anti-TNF therapy, two progressed to surgery, and one switched to tioguanine therapy. CONCLUSION: Split-dose administration of mercaptopurine/AZA represents an alternative option in IBD patients with preferential 6-MMP metabolism who might otherwise require steroid exposure or escalation of therapy.

Thiopurine methyltransferase activity combined with 6-thioguanine metabolite levels predicts clinical response to thiopurines in patients with inflammatory bowel disease.

BACKGROUND/AIMS: 6-Mercaptopurine and its prodrug azathioprine are effective for the treatment of inflammatory bowel disease. Thiopurine methyltransferase is important for the metabolism of thiopurines. However, there is controversy as to the clinical utility of measuring thiopurine methyltransferase enzyme activity and 6-thioguanine nucleotide levels. Our aim was to determine if thiopurine methyltransferase enzyme activity and 6-thioguanine nucleotide level monitoring would predict response to therapy with thiopurines in patients with inflammatory bowel disease. METHODS: Baseline thiopurine methyltransferase enzyme activity prior to initiation of therapy with either 6-mercaptopurine or azathioprine was determined in 39 patients with inflammatory bowel disease. The association between clinical response and thiopurine methyltransferase activity and 6-thioguanine nucleotide levels singly or in combination were analysed. RESULTS: Seventeen of 39 patients (44%) responded to 6-mercaptopurine or azathioprine therapy. Thiopurine methyltransferase enzyme activity below the mean of 30.5 U was significantly associated with clinical response. The thiopurine methyltransferase low phenotype was associated with response in 65% vs. 29% in individuals with thiopurine methyltransferase enzyme activity above 30.5 U (p = 0.05). There was no correlation between thiopurine methyltransferase activity and 6-thioguanine nucleotide levels. The maximal 6-thioguanine nucleotide levels did not predict clinical response. When combining thiopurine methyltransferase enzyme activity and 6-thioguanine nucleotide levels, the combination of thiopurine methyltransferase low/6-thioguanine nucleotide high was associated with response in 7/7 (100%) vs. only 2/8 (25%) with the combination of thiopurine methyltransferase high/6-thioguanine nucleotide low (p=0.01). CONCLUSIONS: Thiopurine methyltransferase activity inversely correlated with clinical response to thiopurine treatment in inflammatory bowel disease. Thiopurine methyltransferase enzyme activity below 30.5 U combined with a post-treatment 6-thioguanine nucleotide level > 230 pmol/8 x 10(8) erythrocytes was the best predictor of response.

Identification and characterization of a phase-variable nonfimbrial Salmonella typhimurium gene that alters O-antigen production.

Salmonella typhimurium 798, which was isolated from a pig, is known to phase vary from a nonadhesive to an adhesive phenotype. Cells of the adhesive phenotype adhere to porcine enterocytes, are more readily phagocytized by porcine neutrophils and macrophages, and once phagocytized can survive intracellularly, while cells of the nonadhesive phenotype die rapidly. The effect of phenotypic switching also can be visualized by changes in colony morphologies and the presence of between 10 and 15 proteins in the envelopes of cells in the adhesive phenotype. Mutants previously constructed with cells in the adhesive phenotype and the transposon TnphoA were screened to identify mutants lacking one or more of the unique proteins. One mutation was cloned and sequenced, and the mutation was shown to be in rfaL (O-antigen ligase). Expression of O antigen was shown to be phase variable. The adhesive strain expressed an O antigen that was at least eightfold longer than that for the nonadhesive strain and by virtue of O-antigen production was resistant to porcine complement. The mutant survived intracellularly in phagocytic cells as well as its wild-type parent.

Beta-MHC and SMLC1 transgene induction in overloaded skeletal muscle of transgenic mice.

The hypertrophic responses of white fast-twitch muscle to mechanical overload has been investigated using transgenic mice. After 7 wk of overload, endogenous beta-myosin heavy chain (MHC) and slow myosin light chain 1 and 2 (SMLC1, SMLC2) protein were increased in the overloaded plantaris (OP) muscle compared with sham-operated control plantaris (CP)muscle. Concurrently, the levels of endogenous beta-MHC, SMLC1, SMLC2, and cardiac/slow troponin C (CTnC) mRNA transcripts were significantly increased in OP muscles, whereas skeletal troponin C (sTnC) mRNA transcript levels decreased. As an initial attempt to locate DNA sequence(s) that governs beta-MHC induction in response to mechanical overload, multiple independent transgenic lines harboring four different human beta-MHC transgenes (beta 1286, beta 988, beta 450, beta 141) were generated. Except for transgene beta 141, muscle-specific expression and induction (3- to 22-fold) in OP muscles were observed by measuring chloramphenicol acetyltransferase activity (CAT assay). Induction of a SMLC1 transgene (3920SMLC1) in OP muscles was also observed. Collectively, these in vivo data provide evidence that 1) a mechanical overload inducible element(s) is located between nucleotides -450 and +120 of the human beta-MHC transgene, 2) 3,900 bp of 5' sequence is sufficient to confer mechanical overload induction of a SMLC1 transgene, and 3) the increased expression of slow/type I isomyosin (beta-MHC, SMLC1, SMLC2) in response to mechanical overload is regulated, in part, transcriptionally.