| 1. |
- Fotoohi, Alan Kambiz, et al.
(författare)
-
Impaired transport as a mechanism of resistance to thiopurines in human T-lymphoblastic leukemia cells
- 2006
-
Ingår i: Nucleosides, Nucleotides & Nucleic Acids. - : Informa UK Limited. - 1525-7770 .- 1532-2335. ; 25:9-11, s. 1039-1044
-
Tidskriftsartikel (refereegranskat)abstract
- In order to better understand the mechanisms of resistance to thiopurines, we studied two sublines of the MOLT4 T-lymphoblastic leukemia cell line, resistant to 6-mercaptopurine (6-MP) and 6-thioguanine (6-TG). We found that the underlying mechanism of resistance in both resistant cell lines was a markedly reduction in initial transport of 6-MP (3- and 5-fold, respectively, in 6-MP- and 6-TG-resistant cells). No significant alteration of activities of hypoxanthine-guanine phosphoribosyl transferase, thiopurine methyltransferase or inosine monophosphate dehydrogenase, the key enzymes involved in the metabolism of thiopurines was detected. We conclude that defected initial transport of thiopurines by cells may very well explain their resistance to these drugs. Copyright © Taylor & Francis Group, LLC.
|
|
| 2. |
- Fotoohi, Alan Kambiz, et al.
(författare)
-
Involvement of the concentrative nucleoside transporter 3 and equilibrative nucleoside transporter 2 in the resistance of T-lymphoblastic cell lines to thiopurines
- 2006
-
Ingår i: Biochemical and Biophysical Research Communications - BBRC. - : Elsevier BV. - 0006-291X .- 1090-2104. ; 343:1, s. 208-215
-
Tidskriftsartikel (refereegranskat)abstract
- Mechanisms of resistance to thiopurines, 6-mercaptopurine (6-MP) and 6-thioguanine (6-TG) were investigated in human leukemia cell lines. We developed two 6-MP- and 6-TG-resistant cell lines from the human T-lymphoblastic cell line (MOLT-4) by prolonged exposure to these drugs. The resistant cells were highly cross resistant to 6-MP and 6-TG, and exhibited marked reduction in cellular uptake of 6-MP (70% and 80%, respectively). No significant modification of the activities of hypoxanthine-guanine phosphoribosyl transferase, thiopurine methyltransferase or inosine monophosphate dehydrogenase was observed. Real-time PCR of concentrative nucleoside transporter 3 (CNT3) and equilibrative nucleoside transporter 2 (ENT2) of resistant cells showed substantial reductions in expression of messenger RNAs. Small interfering RNA designed to silence the CNT3 and ENT2 genes down-regulated the expression of these genes in leukemia cells. These decreases were accompanied by reduction of transport of 6-MP (47% and 21%, respectively) as well as its cytocidal effect (30% and 21%, respectively). Taken together these results show that CNT3 and ENT2 play a key role in the transport of 6-MP and 6-TG by leukemia cells. From a clinical point of view determination of CNT3 and ENT2 levels in leukemia cells may be useful in predicting the efficacy of thiopurine treatment. © 2006 Elsevier Inc. All rights reserved.
|
|
| 3. |
- Fotoohi, A K, et al.
(författare)
-
Molecular mechanisms underlying the enhanced sensitivity of thiopurine-resistant T-lymphoblastic cell lines to methyl mercaptopurineriboside
- 2006
-
Ingår i: Biochemical Pharmacology. - : Elsevier BV. - 0006-2952 .- 1356-1839. ; 72:7, s. 816-823
-
Tidskriftsartikel (refereegranskat)abstract
- Methylmercaptopurine riboside (meMPR), a cellular metabolite of 6-mercaptopurine (6-MP), is a potent inhibitor of de novo purine synthesis (DNPS). Human MOLT4 T-lymphoblastic leukaemia cells that have acquired resistance to 6-MP or 6-thioguanine (6-TG) as a consequence of defective transport exhibit enhanced sensitivity to meMPR. HPLC-based analysis of the transport of meMPR revealed normal uptake of this compound by our thiopurine-resistant cell sublines, suggesting a route of transport distinct from that for 6-MP and 6-TG. Studies on the wild-type parental leukemic cells showed that adenosine, dipyridamole and nitrobenzylthioinosine inhibit uptake of meMPR to a significant extent, whereas Na+ ions have no influence on this process. Transfection of these leukemic cells with small interference RNA molecules targeting the gene encoding the first member of the family of equiliberative nucleoside transporters (ENT1) strongly reduced the initial rate of meMPR transport. Our resistant cell lines exhibited 30-52% reductions (p < 0.005) in their levels of mRNA encoding several proteins involved in de novo purine synthesis, i.e., aminoimidazole carboxamide ribonucleotide formyltransferase, glycinamide ribonucleotide transformylase and guanine monophosphate synthetase. Consequently, the rate of de novo purine synthesis in these resistant sublines was decreased by 50%. Furthermore, the levels of ribonucleoside triphosphates in these cells were significantly lower than in the non-resistant parental cells. In combination, a reduced rate of de novo purine synthesis together with low levels of ribonucleoside triphosphates can explain the enhanced sensitivity of our thiopurine-resistant cell lines to meMPR. In this manner, meMPR bypasses the mechanisms of resistance to thiopurines and is even more cytotoxic towards resistant than towards wild-type cells. © 2006.
|
|
| 4. |
|
|
| 5. |
- Hindorf, Ulf, et al.
(författare)
-
Pharmacogenetics during standardised initiation of thiopurine treatment in inflammatory bowel disease
- 2006
-
Ingår i: Gut. - : BMJ. - 0017-5749 .- 1468-3288. ; 55:10, s. 1423-1431
-
Tidskriftsartikel (refereegranskat)abstract
- Background: Firm recommendations about the way thiopurine drugs are introduced and the use of thiopurine methyltransferase (TPMT) and metabolite measurements during treatment in inflammatory bowel disease (IBD) are lacking. Aim: To evaluate pharmacokinetics and tolerance after initiation of thiopurine treatment with a fixed dosing schedule in patients with IBD. Patients: 60 consecutive patients with Crohn's disease (n = 33) or ulcerative colitis (n = 27) were included in a 20 week open, prospective study. Methods: Thiopurine treatment was introduced using a predefined dose escalation schedule, reaching a daily target dose at week 3 of 2.5 mg azathioprine or 1.25 mg 6-mercaptopurine per kg body weight. TPMT and ITPA genotypes, TPMT activity, TPMT gene expression, and thiopurine metabolites were determined. Clinical outcome and occurrence of adverse events were monitored. Results: 27 patients completed the study per protocol, while 33 were withdrawn (early protocol violation (n = 5), TPMT deficiency (n = 1), thiopurine related adverse events (n = 27)), 67% of patients with adverse events tolerated long term treatment on a lower dose (median 1.32 mg azathioprine/kg body weight). TPMT activity did not change during the 20 week course of the study but a significant decrease in TPMT gene expression was found (TPMT/huCYC ratio, p = 0.02). Patients with meTIMP concentrations > 11 450 pmol/8 × 108 red blood cells during steady state at week 5 had an increased risk of developing myelotoxicity (odds ratio = 45.0, p = 0.015). Conclusions: After initiation of thiopurine treatment using a fixed dosing schedule, no general induction of TPMT enzyme activity occurred, though TPMT gene expression decreased. The development of different types of toxicity was unpredictable, but we found that measurement of meTIMP early in the steady state phase helped to identify patients at risk of developing myelotoxicity.
|
|
| 6. |
- Lindqvist Appell, Malin, 1976-, et al.
(författare)
-
No induction of thiopurine methyltransferase during thiopurine treatment in inflammatory bowel disease
- 2006
-
Ingår i: Nucleosides, Nucleotides & Nucleic Acids. - : Informa UK Limited. - 1525-7770 .- 1532-2335. ; 25:9-11, s. 1033-1037
-
Tidskriftsartikel (refereegranskat)abstract
- The aim of this study was to follow, during standardized initiation of thiopurine treatment, thiopurine methyltransferase (TPMT) gene expression and enzyme activity and thiopurine metabolite concentrations, and to study the role of TPMT and ITPA 94C > A polymorphisms for the development of adverse drug reactions. Sixty patients with ulcerative colitis or Crohn's disease were included in this open and prospective multi-center study. Thiopurine naive patients were prescribed azathioprine (AZA), patients previously intolerant to AZA received 6-mercaptopurine (6-MP). The patients followed a predetermined dose escalation schedule, reaching target dose at Week 3; 2.5 and 1.25 mg/kg body weight for AZA and 6-MP, respectively. The patients were followed every week during Weeks 1-8 from baseline and then every 4 weeks until 20 weeks. TPMT activity and thiopurine metabolites were determined in erythrocytes, TPMT and ITPA genotypes, and TPMT gene expression were determined in whole blood. One homozygous TPMT-deficient patient was excluded. Five non compliant patients were withdrawn during the first weeks. Twenty-seven patients completed the study per protocol; 27 patients were withdrawn because of adverse events. Sixty-seven percent of the withdrawn patients tolerated thiopurines at a lower dose at Week 20. There was no difference in baseline TPMT enzyme activity between individuals completing the study and those withdrawn for adverse events (p = 0.45). A significant decrease in TPMT gene expression (TPMT/huCYC ratio, p = 0.02) was found, however TPMT enzyme activity did not change. TPMT heterozygous individuals had a lower probability of remaining in the study on the predetermined dose (p = 0.039). The ITPA 94C > A polymorphism was not predictive of adverse events (p = 0.35).
|
|
| 7. |
- Lindqvist Appell, Malin, 1976-, et al.
(författare)
-
Thiopurines in inflammatory bowel disease - The role of pharmacogenetics and therapeutic drug monitoring
- 2006
-
Ingår i: Current Pharmacogenomics. - : Bentham Science Publishers Ltd.. - 1570-1603. ; 4:4, s. 285-300
-
Tidskriftsartikel (refereegranskat)abstract
- Pharmacogenetics represents the study of variability in drug response due to genetic variations. Inflammatory bowel disease (IBD, i.e. primarily Crohn's disease and ulcerative colitis) is characterized by a chronic or relapsing inflammation of the digestive tract. The thiopurines 6-mercaptopurine (6-MP) and azathioprine (AZA), an imidazol derivative and pro-drug of 6-MP, are widely used in IBD, particularly in Crohn's disease. The metabolism of thiopurines is complex and individually variable. Thiopurine methyltransferase (TPMT) is a key enzyme in this metabolism and exhibits a genetic variability due to a number of variant alleles coding for a defective enzyme. The formation of biologically active thioguanine nucleotides (TGN) and methylated metabolites may vary considerably due to the TPMT activity. Patients with decreased TPMT activity are at increased risk of developing severe side effects if treated with conventional thiopurine doses, due to the accumulation of toxic metabolites. Determination of the TPMT phenotype or genotype is often used to identify individuals with increased risk for adverse events. Twenty-one variant TPMT alleles have been described, of which three are more common than the others. An association between inosine triphosphate pyrophosphatase polymorphisms and adverse events during thiopurine treatment has also been proposed. In this review, the clinical value of TPMT status determination and pharmacological monitoring of thiopurine metabolites are discussed as well as the increased interest in the use of 6-thioguanine, a thiopurine with a less complex metabolism, as an alternative for patients who do not tolerate AZA or 6-MP. It can be concluded that TPMT determination before start of thiopurine therapy is of value to identify individuals with increased risk for adverse reactions due to genetic enzyme deficiency. However, large prospective studies are still needed to evaluate the true benefit of monitoring thiopurine metabolites during thiopurine treatment. © 2006 Bentham Science Publishers Ltd.
|
|
| 8. |
- Lotfi, Kourosh, et al.
(författare)
-
The pattern of deoxycytidine- and deoxyguanosine kinase activity in relation to messenger RNA expression in blood cells from untreated patients with B-cell chronic lymphocytic leukemia
- 2006
-
Ingår i: Biochemical Pharmacology. - : Elsevier BV. - 0006-2952 .- 1356-1839. ; 71:6, s. 882-890
-
Tidskriftsartikel (refereegranskat)abstract
- Deoxycytidine kinase (dCK) and deoxyguanosine kinase (dGK) catalyze the first step in the intracellular cascade of fludarabine (2-fluoroadenine-beta-D-arabinofuranoside) and cladribine (2-chlorodeoxyadenosine) phosphorylation, which leads to activation of these prodrugs, commonly used for treatment of chronic lymphocytic leukemia (CLL). Thus, resistance to nucleoside analogues may primarily be due to low levels of deoxynucleoside kinase activity. The purpose of this study was to investigate the activity profiles of dCK and dGK and characterize the possible relationship between the levels of dCK enzymatic activities and mRNA levels in B-CLL cells from untreated patient samples in an attempt to determine the best approach for predicting sensitivity to nucleoside analogues and thereby optimizing treatment of CLL. For this purpose, dCK and dGK analyses were done in blood cells from 59 untreated symptomatic patients with CLL. The dGK activity towards 2-chlorodeoxyadenosine was significantly lower than of dCK (median 73 pmol/mg protein/min (85-121, 95% CI) versus 353 pmol/mg protein/min (331-421)). The median dCK mRNA level was 0.107 (0.096-0.120, 95% CI). There was a lack of correlation between the activities of dCK and dGK, which indicates that these proteins are regulated independently. We also found that the dCK and dGK activity measurement towards their endogenous substrates were comparable to the nucleoside analogues tested. Such variations in enzyme activities and mRNA levels may well explain differences in clinical responses to treatment. There was no correlation between the levels of dCK mRNAs and enzymatic activities using a quantitative real-time PCR procedure. Sequencing of dCK mRNA did not reveal alternate splicing or mutations in the coding region. The relation between activity and mRNA levels was studied by short interfering RNA (siRNA) method, which showed that in the siRNA treated cells the down-regulation of dCK expression, and activity followed each other. However, in control cells the mRNA levels remained stable but the protein activity markedly decreased. These data demonstrate that the dCK activity is not reflected by dCK mRNA expression that indicates a post-translational mechanism(s). (c) 2005 Elsevier Inc. All rights reserved.
|
|
| 9. |
- Masquelier, Michéle, et al.
(författare)
-
Cytotoxic effect of a lipophilic alkylating agent after incorporation into low density lipoprotein or emulsions : Studies in human leukemic cells
- 2006
-
Ingår i: Leukemia Research. - : Elsevier BV. - 0145-2126 .- 1873-5835. ; 30:2, s. 136-144
-
Tidskriftsartikel (refereegranskat)abstract
- The use of low density lipoprotein (LDL) as drug carrier in acute myeloblastic leukemia chemotherapy is attractive due to high LDL uptake by leukemic cells. Lipid-based formulations, such as liposomes or microemulsions are promising alternatives. In the current study, we incorporated N-trifluoroacetyl-adriamycin-14-valerate (AD32), a lipophilic derivative of daunorubicin (DNR), and WB4291, a lipophilic alkylating agent, into LDL or lipid microemulsions and evaluated their cytotoxic activities towards leukemic cell lines using as references DNR and melphalan. The incorporation of AD32 into LDL or emulsion resulted in complexes with poor cytotoxicity. WB4291-LDL and WB4291-emulsion exerted, on the other hand, promising cytotoxic effects towards parental and resistant K562 and HL60 cell lines. © 2005 Elsevier Ltd. All rights reserved.
|
|
| 10. |
- Palle, Josefine, et al.
(författare)
-
Doxorubicin pharmacokinetics is correlated to the effect of induction therapy in children with acute myeloid leukemia
- 2006
-
Ingår i: Anti-Cancer Drugs. - : Ovid Technologies (Wolters Kluwer Health). - 0959-4973 .- 1473-5741. ; 17:4, s. 385-392
-
Tidskriftsartikel (refereegranskat)abstract
- We studied the pharmacokinetics of doxorubicin in 41 children treated for newly diagnosed acute myeloid leukemia. Doxorubicin, 75 mg/m(2) body surface area, was administered by constant i.v. infusion over 8 h. Four children with Down's syndrome (DS), 1.2-2.3 years old, had a median total body clearance of 523 ml/min/m(2). The median clearance in non-DS children, 0.6-1.8 years old (n=4) and 2.5-17.7 years old (n=33), was 446 and 538 ml/min/m(2), respectively. Patients who went into complete remission (CR) after induction therapy had a significantly higher median plasma concentration of doxorubicin than those who did not, 249 compared with 180 ng/ml, respectively (P=0.036; analysis restricted to non-DS patients). Doxorubicin plasma concentration was an independent factor for CR, both in univariate (P=0.031) and multivariate analysis including sex, age and white blood cell count at diagnosis (P=0.021). Patients who reached CR had a significantly lower doxorubicin clearance than those who did not, 513 and 657 ml/min/m(2), respectively (P=0.017). In conclusion, doxorubicin plasma concentration and total body clearance during up-front treatment were correlated to the effect of induction therapy. Prospective studies should be performed to confirm the concentration-effect relationship and explore the possibility of therapeutic monitoring.
|
|
