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Freidoun Albertioni - One of the best experts on this subject based on the ideXlab platform.
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Gene expression profiling of leukemia T-cells resistant to methotrexate and 7-Hydroxymethotrexate reveals alterations that preserve intracellular levels of folate and nucleotide biosynthesis
Biochemical Pharmacology, 2009Co-Authors: Alan Kambiz Fotoohi, Gerrit Jansen, Godefridus J Peters, Yehuda G. Assaraf, Ali Moshfegh, Jamileh Hashemi, Catharina Larsson, Freidoun AlbertioniAbstract:treatment of human T-cell leukemia cells with 7-Hydroxymethotrexate, the major metabolite of methotrexate resulted in acquired resistance as a result of the complete loss of folypolyglutamate synthetase (FPGS) activity. This was in contradistinction to the major modality of antifolate resistance of impaired drug transport in leukemia cells exposed to methotrexate.. To identify the genes associated with methotrexate and 7-Hydroxymethotrexate resistance, we herein explored the patterns of genome-wide expression profiles in these antifolte-resistant leukemia sublines. mRNA levels of the reduced folate carrier, the primary influx transporter of folates and antifolates, were down-regulated >2-fold in methotrexate -resistant cells. The dramatic loss of FPGS activity in 7-Hydroxymethotrexate - resistant cells was associated with alterations in the expression of various genes aimed at preserving reduced folates and/or enhancing purine nucleotide biosynthesis e.g. methylene tetrahydrofolate reductase, glycinamide ribonucleotide formyltransferase, adenosine deaminase, cystathionine β synthase, as well as the ATP-dependent folate exporters / and /. The observed changes in gene expression were generally not paralleled by acquired DNA copy numbers alterations, suggesting transcriptional regulatory mechanisms. Interestingly, gene expression of DNA/RNA metabolism and transport genes were more profoundly altered in methotrexate -resistant subline, whereas in 7-Hydroxymethotrexate -resistant cells, the most profoundly affected groups of genes were those encoding for proteins involved in metabolism and cellular proliferation. Thus, the present investigation provides evidence that 7-Hydroxymethotrexate induces gene expression alterations and an antifolate resistance modality that are distinct from its parent drug methotrexate.
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Gene expression profiling of leukemia T-cells resistant to methotrexate and 7-Hydroxymethotrexate reveals alterations that preserve intracellular levels of folate and nucleotide biosynthesis ☆
Biochemical Pharmacology, 2009Co-Authors: Alan Kambiz Fotoohi, Gerrit Jansen, Godefridus J Peters, Yehuda G. Assaraf, Ali Moshfegh, Jamileh Hashemi, Catharina Larsson, Freidoun AlbertioniAbstract:Abstract In vitro treatment of human T-cell leukemia cells with 7-Hydroxymethotrexate, the major metabolite of methotrexate resulted in acquired resistance as a result of the complete loss of folypolyglutamate synthetase (FPGS) activity. This was in contradistinction to the major modality of antifolate resistance of impaired drug transport in leukemia cells exposed to methotrexate. To identify the genes associated with methotrexate and 7-Hydroxymethotrexate resistance, we herein explored the patterns of genome-wide expression profiles in these antifolte-resistant leukemia sublines. mRNA levels of the reduced folate carrier, the primary influx transporter of folates and antifolates, were down-regulated more than two-fold in methotrexate-resistant cells. The dramatic loss of FPGS activity in 7-Hydroxymethotrexate-resistant cells was associated with alterations in the expression of various genes aimed at preserving reduced folates and/or enhancing purine nucleotide biosynthesis, e.g. methylene tetrahydrofolate reductase, glycinamide ribonucleotide formyltransferase, adenosine deaminase, cystathionine β synthase, as well as the ATP-dependent folate exporters BCRP / ABCG2 and MRP1 / ABCC1 . The observed changes in gene expression were generally not paralleled by acquired DNA copy numbers alterations, suggesting transcriptional regulatory mechanisms. Interestingly, gene expression of DNA/RNA metabolism and transport genes were more profoundly altered in methotrexate-resistant subline, whereas in 7-Hydroxymethotrexate-resistant cells, the most profoundly affected groups of genes were those encoding for proteins involved in metabolism and cellular proliferation. Thus, the present investigation provides evidence that 7-Hydroxymethotrexate induces gene expression alterations and an antifolate resistance modality that are distinct from its parent drug methotrexate.
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Disparate Mechanisms of Antifolate Resistance Provoked by Methotrexate and Its Metabolite 7-Hydroxymethotrexate in Leukemia Cells: Implications for Efficacy of Methotrexate Therapy.
Blood, 2004Co-Authors: Freidoun Albertioni, Gerrit Jansen, Yehuda G. Assaraf, Kambiz Fotoohi, Lilah Rothem, Michal Stark, Letje Kathmann, Jacek Gregorczyk, Godefridus J PetersAbstract:Abstract Methotrexate (MTX) is one of the leading drugs in the treatment of leukemia, but extensive metabolism to 7-Hydroxymethotrexate (7-OHMTX) can limit its therapeutic efficacy. In this study we investigated whether 7-OHMTX itself can provoke antifolate resistance that may further disrupt MTX efficacy. For this purpose we developed resistance to 7-OHMTX as well as MTX in two human leukemia cell lines (CCRF-CEM and MOLT-4) by stepwise exposure to increasing concentrations of 7-OHMTX and MTX. Consequently, both leukemia cell lines displayed marked levels of resistance to 7-OHMTX (>10 fold) and MTX (>75 fold), respectively. The underlying mechanism of resistance in the MTX-exposed cells was a marked decrease (>10-fold) in reduced folate carrier (RFC)-mediated cellular uptake of MTX. This was associated with transcriptional silencing of the RFC gene in MTX-resistant CCRF-CEM cells. In contrast, the molecular basis for the resistance to 7-OHMTX was solely due to a marked decreased (> 95%) in folylpolyglutamate synthetase (FPGS) activity which conferred >100-fold MTX resistance upon a short term exposure to this drug. This is the first demonstration that 7-OHMTX can provoke distinct modalities of antifolate resistance as compared to the parent drug MTX.
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Disparate mechanisms of antifolate resistance provoked by methotrexate and its metabolite 7-Hydroxymethotrexate in leukemia cells: implications for efficacy of methotrexate therapy
Blood, 2004Co-Authors: Kambiz Fotoohi, Gerrit Jansen, Godefridus J Peters, Yehuda G. Assaraf, Lilah Rothem, Michal Stark, Jacek Gregorczyk, Ietje Kathmann, Freidoun AlbertioniAbstract:Methotrexate (MTX) is one of the leading drugs in the treatment of leukemia, but extensive metabolism to 7-Hydroxymethotrexate (7-OHMTX) can limit its therapeutic efficacy. In this study we investigated whether 7-OHMTX itself can provoke anti-folate resistance that may further disrupt MTX efficacy. For this purpose, we developed resistance to 7-OHMTX as well as MTX in 2 human leukemia cell lines (CCRF-CEM and MOLT-4) by stepwise exposure to increasing concentrations of 7-OHMTX and MTX. Consequently, both leukemia cell lines displayed marked levels of resistance to 7-OHMTX (> 10-fold) and MTX (> 75-fold). The underlying mechanism of resistance in the MTX-exposed cells was a marked decrease (> 10-fold) in reduced folate carrier (RFC)-mediated cellular uptake of MTX. This was associated with transcriptional silencing of the RFC gene in MTX-resistant CCRF-CEM cells. In contrast, the molecular basis for the resistance to 7-OHMTX was due solely to a marked decreased (> 95%) in folylpolyglutamate synthetase (FPGS) activity, which conferred more than 100-fold MTX resistance upon a short-term exposure to this drug. This is the first demonstration that 7-OHMTX can provoke distinct modalities of antifolate resistance compared with the parent drug MTX. The implications of this finding for MTX efficacy and strategies to circumvent MTX resistance are discussed.
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Monitoring of methotrexate and 7-Hydroxymethotrexate in saliva from children with acute lymphoblastic leukemia receiving high-dose consolidation treatment: relation to oral mucositis.
Anti-cancer drugs, 1997Co-Authors: Freidoun Albertioni, Charlotte Ulrikka Rask, Henrik Schroeder, Curt PetersonAbstract:The purpose of the study was to find out if saliva concentrations of methotrexate (MTX) and its main metabolite, 7-Hydroxymethotrexate (7-OHMTX), can predict oral mucositis in children with acute lymphoblastic leukemia (ALL) after treatment with high-dose consolidation therapy. We have also studied the relationship between the concentrations of MTX and 7-OHMTX in saliva and the unbound concentrations in plasma. Twelve patients (36 infusions) were studied during treatment with high-dose MTX as remission consolidation therapy (5-8 g/m2 by 24 h i.v. infusion followed by leucovorin rescue). Plasma and saliva concentrations of MTX and 7-OHMTX were determined concomitantly by HPLC at 20 h and at various times following infusion. Unbound plasma concentrations of MTX and 7-OHMTX were determined after ultrafiltration. Oral toxicity was graded according to the WHO criteria (grade 0-4). The concentrations of MTX and 7-OHMTX in saliva were not directly related to the development of mucositis. In patients with oral mucositis (WHO grade 1 or greater), the ratio to 7-OHMTX and MTX in saliva at 20 h was significantly lower than in patients without symptoms (p = 0.014, Mann Whitney rank sum test), but not at 42 and 66 h after starting the infusion. The salivary concentration of 7-OHMTX at 20 h ranged from undetectable (less than 1 nmol/l) to 1.6 micromol/l. No significant correlation was found between the unbound and total plasma concentrations of MTX and 7-OHMTX and the drug concentrations in saliva at different points in time. The concentrations of 7-OHMTX in saliva were 11, 23 and 13% of the unbound plasma concentrations at 20, 42 and 66 h, respectively, after starting the infusion. The respective median corresponding values for MTX were 1.6, 16.1 and 61.6%. The results suggest that determinations of saliva concentrations of MTX and 7-OHMTX may predict oral mucositis. This opens up the possibility of early identification of patients at high risk of developing oral mucositis in order to intensify topical or systemic treatment of these patients.
Alfred H. Schinkel - One of the best experts on this subject based on the ideXlab platform.
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Impact of Abcc2 [Multidrug Resistance-Associated Protein (Mrp) 2], Abcc3 (Mrp3), and Abcg2 (Breast Cancer Resistance Protein) on the Oral Pharmacokinetics of Methotrexate and Its Main Metabolite 7-Hydroxymethotrexate
Drug metabolism and disposition: the biological fate of chemicals, 2011Co-Authors: Maria L. H. Vlaming, Els Wagenaar, Zeliha Pala, Anita Van Esch, Olaf Van Tellingen, Evita Van De Steeg, Alfred H. SchinkelAbstract:The ATP-binding cassette (ABC) transporters ABCC2 [multidrug resistance-associated protein (MRP) 2], ABCC3 (MRP3), and ABCG2 (breast cancer resistance protein) are involved in the efflux of potentially toxic compounds from the body. We have shown before that ABCC2, ABCC3, and ABCG2 together influence the pharmacokinetics of the anticancer and antirheumatic drug methotrexate (MTX) and its toxic metabolite 7-Hydroxymethotrexate (7OH-MTX) after intravenous MTX administration. We now have used Abcc2;Abcc3;Abcg2(-/-) and corresponding single and double knockout mice to investigate the relative impact of these transporters on MTX and 7OH-MTX pharmacokinetics after oral MTX administration (50 mg/kg). The plasma areas under the curve (AUC plasma) in Abcg2(-/-) and Abcc2;Abcg2(-/-) mice were 1.7- and 3.0-fold higher than those in wild-type mice, respectively, suggesting additive effects of Abcc2 and Abcg2 on oral MTX pharmacokinetics. However, the AUC plasma in Abcc2;Abcc3; Abcg2(-/-) mice was not different from that in wild-type mice, indicating that Abcc3 protein is necessary for increased MTX plasma concentrations in the absence of Abcc2 and/or Abcg2. Furthermore, 2 h after administration, MTX liver levels were increased in Abcg2-deficient strains and MTX kidney levels were 2.2-fold increased in Abcc2;Abcg2(-/-) mice compared with those in wild-type mice. The absence of Abcc2 and/or Abcg2 also led to significantly increased liver and kidney levels of 7OH-MTX. Our results suggest that inhibition of ABCG2 and/or ABCC2, genetic polymorphisms or mutations reducing expression or activity of these proteins may increase the oral availability of MTX. Such conditions may also present risk factors for increased MTX-related toxicity in patients treated with oral MTX. Copyright © 2011 by The American Society for Pharmacology and Experimental Therapeutics.
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Abcc2 (Mrp2), Abcc3 (Mrp3), and Abcg2 (Bcrp1) are the main determinants for rapid elimination of methotrexate and its toxic metabolite 7-Hydroxymethotrexate in vivo
Molecular cancer therapeutics, 2009Co-Authors: Maria L. H. Vlaming, Koen Van De Wetering, Els Wagenaar, Zeliha Pala, Anita Van Esch, Olaf Van Tellingen, Alfred H. SchinkelAbstract:The multidrug transporters ABCC2, ABCC3, and ABCG2 can eliminate potentially toxic compounds from the body and have overlapping substrate specificities. To investigate the overlapping functions of Abcc2, Abcc3, and Abcg2 in vivo, we generated and characterized Abcc3;Abcg2-/- and Abcc2;Abcc3;Abcg2-/- mice. We subsequently analyzed the relative impact of these transport proteins on the pharmacokinetics of the anticancer drug methotrexate (MTX) and its main, toxic, metabolite 7-Hydroxymethotrexate (7OH-MTX) after i.v. administration of MTX (50 mg/kg). Whereas in single and double knockout mice, the plasma and liver concentrations of MTX and 7OH-MTX decreased rapidly after MTX administration, in the Abcc2;Abcc3;Abcg2-/- mice, they remained very high. One hour after administration, 67% of the MTX dose was still present in livers of Abcc2;Abcc3;Abcg2-/- mice as MTX or 7OH-MTX versus 7% in wild-type, showing dramatic liver accumulation of these toxic compounds when Abcc2, Abcc3, and Abcg2 were all absent. Furthermore, the urinary and fecal excretion of the nephrotoxic metabolite 7OH-MTX were increased 27- and 7-fold, respectively, in Abcc2;Abcc3;Abcg2-/- mice. Thus, Abcc2, Abcc3, and Abcg2 together mediate the rapid elimination of MTX and 7OH-MTX after i.v. administration and can to a large extent compensate for each other's absence. This may explain why it is still comparatively safe to use a toxic drug such as MTX in the clinic, as the risk of highly increased toxicity due to dysfunctioning of ABCC2, ABCC3, or ABCG2 alone is limited. Nevertheless, cotreatment with possible inhibitors of ABCC2, ABCC3, and ABCG2 should be done with utmost caution when treating patients with methotrexate.
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Functionally overlapping roles of Abcg2 (Bcrp1) and Abcc2 (Mrp2) in the elimination of methotrexate and its main toxic metabolite 7-Hydroxymethotrexate in vivo.
Clinical cancer research : an official journal of the American Association for Cancer Research, 2009Co-Authors: Maria L. H. Vlaming, Koen Van De Wetering, Dirk R. De Waart, Ronald P.j. Oude Elferink, Els Wagenaar, Zeliha Pala, Anita Van Esch, Olaf Van Tellingen, Cornelia M.m. Van Der Kruijssen, Alfred H. SchinkelAbstract:Purpose: ABCC2 (MRP2) and ABCG2 (BCRP) transport various endogenous and exogenous compounds, including many anticancer drugs, into bile, feces, and urine. We investigated the possibly overlapping roles of Abcg2 and Abcc2 in the elimination of the anticancer drug methotrexate (MTX) and its toxic metabolite 7-Hydroxymethotrexate (7OH-MTX). Experimental Design: We generated and characterized Abcc2;Abcg2 -/- mice, and used these to determine the overlapping roles of Abcc2 and Abcg2 in the elimination of MTX and 7OH-MTX after i.v. administration of 50 mg/kg MTX. Results: Compared with wild-type, the plasma areas under the curve (AUC) for MTX were 1.6-fold and 2.0-fold higher in Abcg2 -/- and Abcc2 -/- mice, respectively, and 3.3-fold increased in Abcc2;Abcg2 -/- mice. The biliary excretion of MTX was 23-fold reduced in Abcc2;Abcg2 -/- mice, and the MTX levels in the small intestine were dramatically decreased. Plasma levels of 7OH-MTX were not significantly altered in Abcg2 -/- mice, but the areas under the curve were 6.2-fold and even 12.4-fold increased in Abcc2 -/- and Abcc2;Abcg2 -/- mice, respectively. This indicates that Abcc2 compensates for Abcg2 deficiency but that Abcg2 can only partly compensate for Abcc2 absence. Furthermore, 21-fold decreased biliary 7OH-MTX excretion in Abcc2;Abcg2 -/- mice and substantial 7OH-MTX accumulation in the liver and kidney were seen. We additionally found that in the absence of Abcc2, Abcg2 mediated substantial urinary excretion of MTX and 7OH-MTX. Conclusions: Abcc2 and Abcg2 together are major determinants of MTX and 7OH-MTX pharmacokinetics. Variations in ABCC2 and/or ABCG2 activity due to polymorphisms or coadministered inhibitors may therefore substantially affect the therapeutic efficacy and toxicity in patients treated with MTX.
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Impact of Abcc2 (Mrp2) and Abcc3 (Mrp3) on the in vivo elimination of methotrexate and its main toxic metabolite 7-Hydroxymethotrexate.
Clinical cancer research : an official journal of the American Association for Cancer Research, 2008Co-Authors: Maria L. H. Vlaming, Koen Van De Wetering, Dirk R. De Waart, Ronald P.j. Oude Elferink, Els Wagenaar, Zeliha Pala, Anita Van Esch, Olaf Van Tellingen, Alfred H. SchinkelAbstract:Purpose: ATP-binding cassette sub-family C member 2 [ABCC2; multidrug resistance–associated protein 2 (MRP2)] and ABCC3 (MRP3) mediate the elimination of toxic compounds, such as drugs and carcinogens, and have a large overlap in substrate specificity. We investigated the roles of Abcc2 and Abcc3 in the elimination of the anticancer drug methotrexate (MTX) and its toxic metabolite 7-Hydroxymethotrexate (7OH-MTX) in vivo . Experimental Design: Abcc2;Abcc3 −/− mice were generated, characterized, and used to investigate possibly overlapping or complementary roles of Abcc2 and Abcc3 in the elimination of MTX and 7OH-MTX after i.v. administration of 50 mg/kg MTX. Results: Abcc2;Abcc3 −/− mice were viable and fertile. In Abcc2 −/− mice, the plasma area under the curve (AUC i.v. ) for MTX was 2.0-fold increased compared with wild type, leading to 1.6-fold increased urinary excretion, which was not seen in Abcc2;Abcc3 −/− mice. Biliary excretion of MTX was 3.7-fold reduced in Abcc2 −/− but unchanged in Abcc2;Abcc3 −/− mice. The plasma AUC i.v. s of 7OH-MTX were 6.0-fold and 4.3-fold increased in Abcc2 −/− and Abcc2;Abcc3 −/− mice, respectively, leading to increased urinary excretion. The biliary excretion of 7OH-MTX was 5.8-fold reduced in Abcc2 −/− but unchanged in Abcc2;Abcc3 −/− mice. 7OH-MTX accumulated substantially in the liver of Abcc2 −/− and especially Abcc2;Abcc3 −/− mice. Conclusions: Abcc2 is important for (biliary) excretion of MTX and its toxic metabolite 7OH-MTX. When Abcc2 is absent, Abcc3 transports MTX and 7OH-MTX back from the liver into the circulation, leading to increased plasma levels and urinary excretion. Variation in ABCC2 and/or ABCC3 activity may therefore have profound effects on the elimination and severity of toxicity of MTX and 7OH-MTX after MTX treatment of patients.
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Mechanism of the Pharmacokinetic Interaction between Methotrexate and Benzimidazoles: Potential Role for Breast Cancer Resistance Protein in Clinical Drug-Drug Interactions
Cancer research, 2004Co-Authors: Pauline Breedveld, Alfred H. Schinkel, Olaf Van Tellingen, Jos H. Beijnen, Noam Zelcer, Dick Pluim, Özgür Sönmezer, M. M. Tibben, Piet Borst, Jan H.m. SchellensAbstract:The antifolate drug methotrexate (MTX) is transported by breast cancer resistance protein (BCRP; ABCG2) and multidrug resistance-associated protein1-4 (MRP1-4; ABCC1-4). In cancer patients, coadministration of benzimidazoles and MTX can result in profound MTX-induced toxicity coinciding with an increase in the serum concentrations of MTX and its main metabolite 7-Hydroxymethotrexate. We hypothesized that benzimidazoles interfere with the clearance of MTX and/or 7-Hydroxymethotrexate by inhibition of the ATP-binding cassette drug transporters BCRP and/or MRP2, two transporters known to transport MTX and located in apical membranes of epithelia involved in drug disposition. First, we investigated the mechanism of interaction between benzimidazoles (pantoprazole and omeprazole) and MTX in vitro in membrane vesicles from Sf9 cells infected with a baculovirus containing human BCRP or human MRP2 cDNA. In Sf9-BCRP vesicles, pantoprazole and omeprazole inhibited MTX transport (IC50 13 microm and 36 microm, respectively). In Sf9-MRP2 vesicles, pantoprazole did not inhibit MTX transport and at high concentrations (1 mm), it even stimulated MTX transport 1.6-fold. Secondly, we studied the transport of pantoprazole in MDCKII monolayers transfected with mouse Bcrp1 or human MRP2. Pantoprazole was actively transported by Bcrp1 but not by MRP2. Finally, the mechanism of the interaction was studied in vivo using Bcrp1-/- mice and wild-type mice. Both in wild-type mice pretreated with pantoprazole to inhibit Bcrp1 and in Bcrp1-/- mice that lack Bcrp1, the clearance of i.v. MTX was decreased significantly 1.8- to 1.9-fold compared with the clearance of i.v. MTX in wild-type mice. The conclusion is as follows: benzimidazoles differentially affect transport of MTX mediated by BCRP and MRP2. Competition for BCRP may explain the clinical interaction between MTX and benzimidazoles.
Curt Peterson - One of the best experts on this subject based on the ideXlab platform.
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Monitoring of methotrexate and 7-Hydroxymethotrexate in saliva from children with acute lymphoblastic leukemia receiving high-dose consolidation treatment: relation to oral mucositis.
Anti-cancer drugs, 1997Co-Authors: Freidoun Albertioni, Charlotte Ulrikka Rask, Henrik Schroeder, Curt PetersonAbstract:The purpose of the study was to find out if saliva concentrations of methotrexate (MTX) and its main metabolite, 7-Hydroxymethotrexate (7-OHMTX), can predict oral mucositis in children with acute lymphoblastic leukemia (ALL) after treatment with high-dose consolidation therapy. We have also studied the relationship between the concentrations of MTX and 7-OHMTX in saliva and the unbound concentrations in plasma. Twelve patients (36 infusions) were studied during treatment with high-dose MTX as remission consolidation therapy (5-8 g/m2 by 24 h i.v. infusion followed by leucovorin rescue). Plasma and saliva concentrations of MTX and 7-OHMTX were determined concomitantly by HPLC at 20 h and at various times following infusion. Unbound plasma concentrations of MTX and 7-OHMTX were determined after ultrafiltration. Oral toxicity was graded according to the WHO criteria (grade 0-4). The concentrations of MTX and 7-OHMTX in saliva were not directly related to the development of mucositis. In patients with oral mucositis (WHO grade 1 or greater), the ratio to 7-OHMTX and MTX in saliva at 20 h was significantly lower than in patients without symptoms (p = 0.014, Mann Whitney rank sum test), but not at 42 and 66 h after starting the infusion. The salivary concentration of 7-OHMTX at 20 h ranged from undetectable (less than 1 nmol/l) to 1.6 micromol/l. No significant correlation was found between the unbound and total plasma concentrations of MTX and 7-OHMTX and the drug concentrations in saliva at different points in time. The concentrations of 7-OHMTX in saliva were 11, 23 and 13% of the unbound plasma concentrations at 20, 42 and 66 h, respectively, after starting the infusion. The respective median corresponding values for MTX were 1.6, 16.1 and 61.6%. The results suggest that determinations of saliva concentrations of MTX and 7-OHMTX may predict oral mucositis. This opens up the possibility of early identification of patients at high risk of developing oral mucositis in order to intensify topical or systemic treatment of these patients.
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Methotrexate in juvenile rheumatoid arthritis. Evidence of age dependent pharmacokinetics.
European journal of clinical pharmacology, 1995Co-Authors: Albertioni F, Olof Beck, Peter Seideman, Curt Peterson, Flatø B, Vinje O, Staffan EksborgAbstract:Children with juvenile rheumatoid arthritis (JRA) have been reported to require higher doses (per kg body weight) of methotrexate (MTX) than adults with rheumatoid arthritis to control their disease. The purpose of the present study was to characterise the plasma pharmacokinetics of MTX and its major metabolite, 7-Hydroxymethotrexate (7-OHMTX) in children, and to compare the results with those previously obtained in adults. Thirteen patients (age 5–16 y) with JRA (median disease duration 5.5 y) were studied after once weekly oral administration of MTX (median 0.21 mg·kg−1).
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Plasma and urine levels of methotrexate and 7-Hydroxymethotrexate in children with ALL during maintenance therapy with weekly oral methotrexate.
Medical and pediatric oncology, 1994Co-Authors: Kathleen A. Skoglund, Olof Beck, Mona Wennberg, Curt Peterson, Stefan Söderhäll, Salah Hayder, Olle BjörkAbstract:A new high-performance liquid chromatographic assay was used to determine methotrexate (MTX) and its main metabolite, 7-Hydroxymethotrexate (7-OH-MTX), in the plasma (n = 17) and urine (n = 14) of children (age 3-12 years) on maintenance therapy for acute lymphocytic leukemia (n = 14) or non-Hodgkin's lymphoma (n = 3). Each child received oral doses of weekly MTX (4.0-29 mg/m2) and daily 6-mercaptopurine (40-111 mg/m2). Plasma samples were collected daily from two children during the 1-week dose interval. A limited sampling strategy was designed, whereby 2 days of blood sampling were used in the other 15 patients. Morning urine samples were collected daily for 1 week following MTX intake from 14 of the children. MTX was detectable in all plasma and urine samples for the entire dose interval. The main metabolite, 7-OH-MTX, could be detected in plasma and urine from all patients on the first day after dose intake but only in a few patients during the whole dose interval. Interpatient variability of MTX and 7-OH-MTX levels was high at all points during the week. Significant correlation were found between the urinary MTX levels on days 2 and 7 and plasma MTX levels on day 2 after intake. No significant correlation was found between drug levels in plasma or urine and liver function tests in the children showing signs of mild liver injury. This assay provides a tool for further studies on the role of pharmacokinetics for the clinical effects of weekly oral low-dose MTX given alone or in combination with 6-mercaptopurine.
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Trace analysis of methotrexate and 7-Hydroxymethotrexate in human plasma and urine by a novel high-performance liquid chromatographic method.
Therapeutic drug monitoring, 1991Co-Authors: Olof Beck, Peter Seideman, Mona Wennberg, Curt PetersonAbstract:A new high-performance liquid chromatographic method for the quantitative determination of methotrexate (MTX) and its metabolite 7-Hydroxymethotrexate (7-OHMTX) in blood plasma and urine was developed. The method utilized a solid-phase extraction procedure (Certify II cartridges) for the simultaneou
Jarle Aarbakke - One of the best experts on this subject based on the ideXlab platform.
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Microvascular perturbations in rats receiving the maximum tolerated dose of methotrexate or its major metabolite 7-Hydroxymethotrexate.
Ultrastructural pathology, 2000Co-Authors: Ole Martin Fuskevåg, Jarle Aarbakke, Randi Olsen, Christel Kristiansen, Sigurd LindalAbstract:Methotrexate (MTX) is a clinically important cytostatic antifolate. The study describes the acute effects of maximum tolerated doses of MTX or its major metabolite 7-Hydroxymethotrexate (7-OH-MTX) on the ultrastructure of rat liver and kidneys. The ultrastructural changes in rats receiving MTX or 7-OH-MTX were, in principle, indistinguishable and their severity and extension increased with time of survival or doses of medication. All lesions were focal, microvascular, or parenchymal. Microvascular changes were more severe in nature when blood cells were present. The endothelial cells were swollen with loss of pinocytotic vesicles, their luminal plasma membrane formed blebs or were disrupted. Partly detached endothelial cells or deendothelialized areas, various types of white blood cells, in particular, neutrophil granulocytes, were observed in the microcirculation. Single platelets or small platelet aggregates were found either in the lumen or adhering to deendothelialized areas of injured endothelial cells. Hepatocytes exhibited steatosis, edema, and manifest single cell necrosis. There were also nuclear changes, marked proliferation of smooth endoplasmatic reticulum, increased amounts of intracellular lipid vacuoles, and a decrease in glycogen particles in hepatocytes. The kidney presented the major changes in the tubules and in the interstitial part. MTX and 7-OH-MTX acute toxicity may primarily be related to microvascular perturbation.
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High-dose 7-Hydroxymethotrexate: Acute toxicity and lethality in a rat model
Cancer Chemotherapy and Pharmacology, 1996Co-Authors: Eivind Smeland, Roy M. Bremnes, Ole Martin Fuskevåg, Kirsten Nymann, John Sigurd Svendsen, Randi Olsen, Sigurd Lindal, Jarle AarbakkeAbstract:To elucidate mechanisms for methotrexate (MTX)-induced renal and hepatic toxicity, we investigated the acute effects of bolus plus continuous infusion of up to 0.4 g/kg 7-Hydroxymethotrexate (7-OH-MTX) in the rat. We demonstrate for the first time in any species the occurrence of acute lethal toxicity within a few hours after 7-OH-MTX administration. Serum concentrations of 7-OH-MTX measured at the time of death were 1.4 mM (mean), about one-half of those achieved in some patients after infusion of high-dose MTX (HD-MTX) in the clinic. The data suggest an approximate LD50 (the dose lethal to 50% of the study population) of 0.3 g/kg and a steep dose/ lethality curve for 7-OH-MTX. Moreover, acute renal and hepatic toxicity occurred as evidenced by severe morphological findings and increased serum levels of creatinine and liver transaminases. In all rats subjected to continuous infusion of 7-OH-MTX, yellow microscopic precipitations were apparent in the kidney tubules. Crystallization was also seen in bile ducts of the liver in some of the rats. These results further support that the formation of 7-OH-MTX is disadvantageous and that reported attempts to prevent its formation during MTX treatment are warranted.
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Renal and hepatic toxicity after high-dose 7-Hydroxymethotrexate in the rat
Cancer Chemotherapy and Pharmacology, 1994Co-Authors: Eivind Smeland, Roy M. Bremnes, Anders Andersen, Ragnhild Jæger, Tor J. Eide, Nils-e Huseby, Jarle AarbakkeAbstract:To examine directly the hepatic and renal toxicity of 7-Hydroxymethotrexate (7-OH-MTX) without interference of the parent compound methotrexate (MTX), we purified and gave 100 mg/kg 7-OH-MTX to rats, a dose resulting in serum levels of 7-OH-MTX comparable with those achieved in the clinic after the administration of high-dose MTX (HD-MTX). After only 5 h, the 7-OH-MTX-treated rats demonstrated 2.6-fold increases in serum creatinine values and 2-fold elevations in serum aspartate aminotransferase (ASAT) levels as compared with the controls. Morphologic evidence of toxicity, however, was apparent only in the kidneys. Intraluminal cellular debris containing membranous material and deteriorated organelles was seen, but no precipitate of the delivered drug. The peak serum concentration of 7-OH was up to 939 μ M , and concentrations of 7-OH-MTX declined triphasically, showing a t 1/2α value of 2.45 min, a t 1/2β value of 30.5 min, and a terminal half-life ( t 1/2γ) of 240 min. The total clearance value was 14.5 ml min^−1 kg, and the postdistributional volume of distribution (Vβ) was 5070 ml/kg. Our results may indicate a direct toxic effect of 7-OH-MTX on kidney and liver cells.
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Inhibition of 7-Hydroxymethotrexate formation by amsacrine
Cancer Chemotherapy and Pharmacology, 1991Co-Authors: Roy M. Bremnes, Eivind Smeland, Nils P. Willassen, Erik Wist, Jarle AarbakkeAbstract:The inhibition of methotrexate (MTX) biotransformation to 7-Hydroxymethotrexate (7-OH-MTX) by 4′-(9-acridinylamino)-methanesulfon- m -anisidide (mAMSA) was studied in bile-drained rats in vivo and in incubates of isolated rat hepatocytes and rat-liver homogenate in vitro. In vivo, i.v. administration of 10 mg/kg mAMSA prior to [^3H]-MTX infusion (50 mg/kg) led to a significant alteration in 7-OH-MTX kinetics. 7-OH-MTX peak concentrations and AUC in bile and serum were reduced by 75% and the recovery of MTX as 7-OH-MTX in bile and urine decreased by 70%, whereas MTX pharmacokinetics remained unaltered. In suspensions of isolated hepatocytes. 10 μ m mAMSA led to a 54% decrease in 7-OH-MTX formation. However, the hepatocellular influx and efflux of MTX was not perturbed by mAMSA. Preincubation of rat-liver homogenates with 1.25–10 μ m mAMSA reduced the formation of 7-OH-MTX by up to 73%. mAMSA appeared to inhibit MTX hydroxylation competitively, exhibiting a K _iof 3 μ m . Due to its inhibition of the MTX-oxidizing system, mAMSA may be beneficial in combination chemotherapy with MTX by reducing 7-OH-MTX-associated toxicity and, possibly, enhancing the cytotoxic effects of MTX.
Godefridus J Peters - One of the best experts on this subject based on the ideXlab platform.
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Gene expression profiling of leukemia T-cells resistant to methotrexate and 7-Hydroxymethotrexate reveals alterations that preserve intracellular levels of folate and nucleotide biosynthesis
Biochemical Pharmacology, 2009Co-Authors: Alan Kambiz Fotoohi, Gerrit Jansen, Godefridus J Peters, Yehuda G. Assaraf, Ali Moshfegh, Jamileh Hashemi, Catharina Larsson, Freidoun AlbertioniAbstract:treatment of human T-cell leukemia cells with 7-Hydroxymethotrexate, the major metabolite of methotrexate resulted in acquired resistance as a result of the complete loss of folypolyglutamate synthetase (FPGS) activity. This was in contradistinction to the major modality of antifolate resistance of impaired drug transport in leukemia cells exposed to methotrexate.. To identify the genes associated with methotrexate and 7-Hydroxymethotrexate resistance, we herein explored the patterns of genome-wide expression profiles in these antifolte-resistant leukemia sublines. mRNA levels of the reduced folate carrier, the primary influx transporter of folates and antifolates, were down-regulated >2-fold in methotrexate -resistant cells. The dramatic loss of FPGS activity in 7-Hydroxymethotrexate - resistant cells was associated with alterations in the expression of various genes aimed at preserving reduced folates and/or enhancing purine nucleotide biosynthesis e.g. methylene tetrahydrofolate reductase, glycinamide ribonucleotide formyltransferase, adenosine deaminase, cystathionine β synthase, as well as the ATP-dependent folate exporters / and /. The observed changes in gene expression were generally not paralleled by acquired DNA copy numbers alterations, suggesting transcriptional regulatory mechanisms. Interestingly, gene expression of DNA/RNA metabolism and transport genes were more profoundly altered in methotrexate -resistant subline, whereas in 7-Hydroxymethotrexate -resistant cells, the most profoundly affected groups of genes were those encoding for proteins involved in metabolism and cellular proliferation. Thus, the present investigation provides evidence that 7-Hydroxymethotrexate induces gene expression alterations and an antifolate resistance modality that are distinct from its parent drug methotrexate.
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Gene expression profiling of leukemia T-cells resistant to methotrexate and 7-Hydroxymethotrexate reveals alterations that preserve intracellular levels of folate and nucleotide biosynthesis ☆
Biochemical Pharmacology, 2009Co-Authors: Alan Kambiz Fotoohi, Gerrit Jansen, Godefridus J Peters, Yehuda G. Assaraf, Ali Moshfegh, Jamileh Hashemi, Catharina Larsson, Freidoun AlbertioniAbstract:Abstract In vitro treatment of human T-cell leukemia cells with 7-Hydroxymethotrexate, the major metabolite of methotrexate resulted in acquired resistance as a result of the complete loss of folypolyglutamate synthetase (FPGS) activity. This was in contradistinction to the major modality of antifolate resistance of impaired drug transport in leukemia cells exposed to methotrexate. To identify the genes associated with methotrexate and 7-Hydroxymethotrexate resistance, we herein explored the patterns of genome-wide expression profiles in these antifolte-resistant leukemia sublines. mRNA levels of the reduced folate carrier, the primary influx transporter of folates and antifolates, were down-regulated more than two-fold in methotrexate-resistant cells. The dramatic loss of FPGS activity in 7-Hydroxymethotrexate-resistant cells was associated with alterations in the expression of various genes aimed at preserving reduced folates and/or enhancing purine nucleotide biosynthesis, e.g. methylene tetrahydrofolate reductase, glycinamide ribonucleotide formyltransferase, adenosine deaminase, cystathionine β synthase, as well as the ATP-dependent folate exporters BCRP / ABCG2 and MRP1 / ABCC1 . The observed changes in gene expression were generally not paralleled by acquired DNA copy numbers alterations, suggesting transcriptional regulatory mechanisms. Interestingly, gene expression of DNA/RNA metabolism and transport genes were more profoundly altered in methotrexate-resistant subline, whereas in 7-Hydroxymethotrexate-resistant cells, the most profoundly affected groups of genes were those encoding for proteins involved in metabolism and cellular proliferation. Thus, the present investigation provides evidence that 7-Hydroxymethotrexate induces gene expression alterations and an antifolate resistance modality that are distinct from its parent drug methotrexate.
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Disparate Mechanisms of Antifolate Resistance Provoked by Methotrexate and Its Metabolite 7-Hydroxymethotrexate in Leukemia Cells: Implications for Efficacy of Methotrexate Therapy.
Blood, 2004Co-Authors: Freidoun Albertioni, Gerrit Jansen, Yehuda G. Assaraf, Kambiz Fotoohi, Lilah Rothem, Michal Stark, Letje Kathmann, Jacek Gregorczyk, Godefridus J PetersAbstract:Abstract Methotrexate (MTX) is one of the leading drugs in the treatment of leukemia, but extensive metabolism to 7-Hydroxymethotrexate (7-OHMTX) can limit its therapeutic efficacy. In this study we investigated whether 7-OHMTX itself can provoke antifolate resistance that may further disrupt MTX efficacy. For this purpose we developed resistance to 7-OHMTX as well as MTX in two human leukemia cell lines (CCRF-CEM and MOLT-4) by stepwise exposure to increasing concentrations of 7-OHMTX and MTX. Consequently, both leukemia cell lines displayed marked levels of resistance to 7-OHMTX (>10 fold) and MTX (>75 fold), respectively. The underlying mechanism of resistance in the MTX-exposed cells was a marked decrease (>10-fold) in reduced folate carrier (RFC)-mediated cellular uptake of MTX. This was associated with transcriptional silencing of the RFC gene in MTX-resistant CCRF-CEM cells. In contrast, the molecular basis for the resistance to 7-OHMTX was solely due to a marked decreased (> 95%) in folylpolyglutamate synthetase (FPGS) activity which conferred >100-fold MTX resistance upon a short term exposure to this drug. This is the first demonstration that 7-OHMTX can provoke distinct modalities of antifolate resistance as compared to the parent drug MTX.
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Disparate mechanisms of antifolate resistance provoked by methotrexate and its metabolite 7-Hydroxymethotrexate in leukemia cells: implications for efficacy of methotrexate therapy
Blood, 2004Co-Authors: Kambiz Fotoohi, Gerrit Jansen, Godefridus J Peters, Yehuda G. Assaraf, Lilah Rothem, Michal Stark, Jacek Gregorczyk, Ietje Kathmann, Freidoun AlbertioniAbstract:Methotrexate (MTX) is one of the leading drugs in the treatment of leukemia, but extensive metabolism to 7-Hydroxymethotrexate (7-OHMTX) can limit its therapeutic efficacy. In this study we investigated whether 7-OHMTX itself can provoke anti-folate resistance that may further disrupt MTX efficacy. For this purpose, we developed resistance to 7-OHMTX as well as MTX in 2 human leukemia cell lines (CCRF-CEM and MOLT-4) by stepwise exposure to increasing concentrations of 7-OHMTX and MTX. Consequently, both leukemia cell lines displayed marked levels of resistance to 7-OHMTX (> 10-fold) and MTX (> 75-fold). The underlying mechanism of resistance in the MTX-exposed cells was a marked decrease (> 10-fold) in reduced folate carrier (RFC)-mediated cellular uptake of MTX. This was associated with transcriptional silencing of the RFC gene in MTX-resistant CCRF-CEM cells. In contrast, the molecular basis for the resistance to 7-OHMTX was due solely to a marked decreased (> 95%) in folylpolyglutamate synthetase (FPGS) activity, which conferred more than 100-fold MTX resistance upon a short-term exposure to this drug. This is the first demonstration that 7-OHMTX can provoke distinct modalities of antifolate resistance compared with the parent drug MTX. The implications of this finding for MTX efficacy and strategies to circumvent MTX resistance are discussed.
