UMLS:C0023418 - FACTA Search

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Query: UMLS:C0023418 (leukemia)
93,477 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

In the past decade, fludarabine has had a major impact in increasing the effectiveness of treatment of patients with indolent B-cell malignancies. This has come about in a variety of clinical circumstances, including use of fludarabine alone as well as in combinations with DNA-damaging agents or membrane-targeted antibodies. Other strategies have used fludarabine to reduce immunological function, thus facilitating non-myeloablative stem cell transplants. Fludarabine is a prodrug that is converted to the free nucleoside 9-beta-D-arabinosyl-2-fluoroadenine (F-ara-A) which enters cells and accumulates mainly as the 5'-triphosphate, F-ara-ATP. The rate-limiting step in the formation of triphosphate is conversion of F-ara-A to its monophosphate, which is catalyzed by deoxycytidine kinase. Although F-ara-A is not a good substrate for this enzyme, the high specific activity of this protein results in efficient phosphorylation of F-ara-A in certain tissues. F-ara-ATP has multiple mechanisms of action, which are mostly directed toward DNA. These include inhibition of ribonucleotide reductase, incorporation into DNA resulting in repression of further DNA polymerisation, and inhibition of DNA ligase and DNA primase. Collectively these actions affect DNA synthesis, which is the major mechanism of F-ara-A-induced cytotoxicity. Secondarily, incorporation into RNA and inhibition of transcription has been shown in cell lines. With the standard dose of fludarabine (25 to 30 mg/m(2)/day given over 30 minutes for 5 days), plasma concentrations of about 3 micromol/L F-ara-A are achieved at the end of each infusion. Serial sampling of leukaemia cells from patients receiving these standard doses of fludarabine has demonstrated that the peak concentrations of F-ara-ATP are achieved 4 hours after start of fludarabine infusion. Although there is heterogeneity among individuals with respect to rate of F-ara-ATP accumulation, the peak concentrations are generally proportional to the dose of the drug. Knowledge of the plasma pharmacokinetics of its principal nucleoside metabolite F-ara-A, and the cellular pharmacology of the proximal active metabolite, F-ara-ATP, has provided some understanding of the activity of fludarabine when used as a single agent. Preclinical studies directed toward learning the mechanisms of action of this agent have formed the basis for several mechanism-based strategies for its combination and scheduling with other agents. As a single agent fludarabine has been effective for the indolent leukaemias. Biochemical modulation strategies resulted in enhanced accumulation of cytarabine triphosphate and led to the use of fludarabine for the treatment of acute leukaemias. Combination of fludarabine with DNA damaging agents to inhibit DNA repair processes has been highly effective for indolent leukaemias and lymphomas. The current review brings together knowledge of the mechanisms of fludarabine, the state of understanding of the plasma pharmacokinetics, and cellular pharmacodynamics of fludarabine nucleotides. This may be useful in the design of future therapeutic approaches.
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PMID:Cellular and clinical pharmacology of fludarabine. 1188 30

The relative levels of the deoxycytidine kinase (dCK), deoxyguanosine kinase (dGK), and the 5'-nucleotidase (5'-NT) are of importance for the effect of many nucleoside analogues used in the treatment of hematological malignancies. To elucidate dCK, dGK and 5'-NT gene expressions in cell lines and in samples from patients with leukemia, we have established a real-time quantitative PCR (RQ-PCR) method. From the available dCK, dGK and 5'-NT cDNA sequences we designed specific primers and fluorogenic probes for the respective genes. The mRNA of dCK, dGK and 5'-NT was also measured by semi-quantitative RT-PCR, the enzyme activities by a radioactive substrate-based technique and Western blot was used to measure the amount of dCK and dGK protein. A MOLT-4 wild-type and its 9-beta-D-arabinofuranosylguanine (Ara-G)-resistant subline was used for the methods comparisons and the RQ-PCR assay was used in 35 samples from pediatric patients with ALL and AML. The results from RQ-PCR for the cell lines were in agreement with the semi-quantitative RT-PCR. The mRNA expression for dCK, dGK and 5'-NT (expressed as the ratio of the respective gene and the reference gene) in pediatric ALL and AML patients showed a large interindividual variability from 0.06 to 2.34, non-detectable to 0.06 and 0.04 to 0.30, respectively. These results show that the quantitative evaluation by RQ-PCR is a valuable tool in the determination of dCK, dGK and 5'-NT mRNA levels in cell lines and in clinical samples which were expressed at various levels. This rapid, convenient and specific method is suitable for further studies of these genes in clinical samples.
Leukemia 2002 Mar
PMID:Real-time quantitative PCR assays for deoxycytidine kinase, deoxyguanosine kinase and 5'-nucleotidase mRNA measurement in cell lines and in patients with leukemia. 1189 43

The prodrug of 9-beta-D-arabinosylguanine (ara-G), nelarabine, demonstrated efficacy against T-cell acute lymphoblastic leukemia, and its effectiveness correlated with the accumulation of the triphosphate form (ara-GTP). Although in vitro investigations using purified deoxycytidine kinase (dCK) or deoxyguanosine kinase (dGK) suggested that ara-G is a substrate for both enzymes, controversy exists in regard to the role of these enzymes in whole cells. In this work, we used a CEM mutant cell line containing low endogenous levels of dGK and deficient in dCK (dCK-) to assess the role of these kinases in ara-G phosphorylation. Using a retroviral vector system, we infected the dCK- mutant cell line to obtain cell lines with overexpression of dCK (dCK+) or dGK (dGK+). Only the dCK+ cell line phosphorylated 1-beta-D-arabinofuranosylcytosine (used as a substrate for dCK) in a cell-free system; phosphorylation of this compound by dGK+ was below the limit of detection. Again in in vitro assays, the dCK-and dCK+ cell lines phosphorylated dGuo to similar levels (0.91 +/- 0.15 and 0.93 +/- 0.19 pmol/mg/min, respectively), whereas dGK+ phosphorylated dGuo more efficiently (150 pmol at 60 min). When ara-G was used as a substrate in a cell-free system, the maximum accumulation of phosphorylated product was observed in dGK+ extracts at low ara-G levels (10 microM) and in dCK+ extracts at high concentrations of ara-G (100 microM). Thus, both dCK and dGK can phosphorylate ara-G, but at low ara-G concentrations, dGK seems to predominate, whereas at higher ara-G concentrations, dCK seems to be the preferred enzyme. In whole-cell systems after a 3-h incubation with 10 microM ara-G, both dCK+ and dGK+ cells accumulated ara-GTP; however, the levels were significantly (P = 0.0008) higher in dGK+ cells. In contrast, at 100 microM ara-G, intracellular ara-GTP accumulated to similar levels (P = 0.5529) in these cell types; 25 +/- 3.7 microM in dCK+, and 27.8 +/- 2.7 microM in the dGK+ cells. These results from whole-cell experiments are consistent with those from the cell-free system and strongly suggest that ara-G is phosphorylated by both kinases, and at low substrate concentrations, dGK is preferred enzyme. Evaluation of the expression of each of these kinases in primary leukemia cells may reveal a biochemical basis for the pharmacological differences in the accumulation of ara-GTP.
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PMID:Arabinosylguanine is phosphorylated by both cytoplasmic deoxycytidine kinase and mitochondrial deoxyguanosine kinase. 1203 20

Ribonucleotide reductase is the rate-limiting enzyme for the de novo synthesis of deoxynucleoside triphosphates and therefore represents a good target for cancer chemotherapy. Trimidox (3,4,5-trihydroxybenzamidoxime) was identified as a potent inhibitor of this enzyme and was shown to significantly decrease deoxycytidine triphosphate (dCTP) pools in HL-60 leukemia cells. We now investigated the ability of trimidox to increase the antitumor effect of 1-beta-D-arabinofuranosyl cytosine (Ara-C). Ara-C is phosphorylated by deoxycytidine kinase, which is subject to negative allosteric regulation by dCTP. Therefore, a decrease of dCTP may cause increased Ara-C phosphorylation and enhanced incorporation of Ara-C into DNA. Ara-C incorporation indeed increased 1.51- and 1.89-fold after preincubation with 75 and 100 microM trimidox, respectively. This was due to the significantly increased 1-beta-D-arabinofuranosyl cytosine triphosphate pools (1.9- and 2.5-fold) after preincubation with trimidox. We also investigated the effects of a combination of trimidox and Ara-C on the colony formation of HL-60 cells. A synergistic potentiation of the effect of Ara-C could be observed, when trimidox was added. Trimidox, which decreases intracellular deoxynucleoside triphosphate concentrations thus leading to apoptosis, enhanced the induction of apoptosis caused by Ara-C. We conclude, that trimidox is capable of synergistically enhancing the effects of Ara-C and therefore this drug combination might be further tested in animals.
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PMID:Trimidox, an inhibitor of ribonucleotide reductase, synergistically enhances the inhibition of colony formation by Ara-C in HL-60 human promyelocytic leukemia cells. 1214

Infant acute lymphoblastic leukemia (ALL) is characterized by a high incidence of mixed lineage leukemia (MLL) gene rearrangements, a poor outcome, and resistance to chemotherapeutic drugs. One exception is cytosine arabinoside (Ara-C), to which infant ALL cells are highly sensitive. To investigate the mechanism underlying Ara-C sensitivity in infants with ALL, mRNA levels of Ara-C-metabolizing enzymes were measured in infants (n = 18) and older children (noninfants) with ALL (n = 24). In the present study, infant ALL cells were 3.3-fold more sensitive to Ara-C (P =.007) and accumulated 2.3-fold more Ara-CTP (P =.011) upon exposure to Ara-C, compared with older children with ALL. Real-time quantitative reverse trancriptase-polymerase chain reaction (RT-PCR) (TaqMan) revealed that infants express 2-fold less of the Ara-C phosphorylating enzyme deoxycytidine kinase (dCK) mRNA (P =.026) but 2.5-fold more mRNA of the equilibrative nucleoside transporter 1 (hENT1), responsible for Ara-C membrane transport (P =.001). The mRNA expression of pyrimidine nucleotidase I (PN-I), cytidine deaminase (CDA), and deoxycytidylate deaminase (dCMPD) did not differ significantly between both groups. hENT1 mRNA expression inversely correlated with in vitro resistance to Ara-C (r(s) = -0.58, P =.006). The same differences concerning dCK and hENT1 mRNA expression were observed between MLL gene-rearranged (n = 14) and germ line MLL cases (n = 25). An oligonucleotide microarray screen (Affymetrix) comparing patients with MLL gene-rearranged ALL with those with nonrearranged ALL also showed a 1.9-fold lower dCK (P =.001) and a 2.7-fold higher hENT1 (P =.046) mRNA expression in patients with MLL gene-rearranged ALL. We conclude that an elevated expression of hENT1, which transports Ara-C across the cell membrane, contributes to Ara-C sensitivity in MLL gene-rearranged infant ALL.
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PMID:Differential mRNA expression of Ara-C-metabolizing enzymes explains Ara-C sensitivity in MLL gene-rearranged infant acute lymphoblastic leukemia. 1240 12

We have previously reported that in a MOLT-4 leukemia cell line the acquired resistance to 9-beta-D-arabinofuranosylguanine (Ara-G) is due to deficiency of the activating enzymes deoxyguanosine kinase and deoxycytidine kinase [Biochem. Biophys. Res. Commun. 293 (5) (2002) 1489]. In this study we investigated whether apoptotic pathways are affected in two human T-cell lymphoblastic MOLT-4 cell lines with acquired resistance to Ara-G. In contrast to the MOLT-4 wild type cells, Ara-G resistant cells displayed no increase in caspase-3 or caspase-9 activity, DNA fragmentation, cytochrome c release or a drop in the mitochondrial membrane potential (DeltaPsi(mito)) upon Ara-G treatment. A drop in the DeltaPsi(mito) was induced in wild type cells after treatment with tributyltin, an inducer of mitochondrial permeability transition, and with carbonyl cyanide m-chlorophenylhydrazone, an uncoupling agent that reduces the DeltaPsi(mito), although not in Ara-G resistant cells. Ara-G resistant cells displayed higher levels of the anti-apoptotic protein Bcl-xL in immunoblots. A recent study indicates that Ara-G-induced apoptosis is mediated in part via the Fas pathway [Cancer Res. 43 (2047) (2002) 411]. When cells were treated with anti-Fas antibody, the wild type cell line exhibited increased caspase-3-like activity but the Ara-G resistant cells did not. Using FACS analysis and semi-quantitative PCR, 3-6-fold decreased protein levels and almost no detectable mRNA levels of Fas in the resistant cells were recorded. These data indicate that the inability to induce apoptosis via both the apoptosome pathway and the Fas pathway, due to increased levels of Bcl-xL and a lack of Fas, contributes to Ara-G resistance. This resistance to apoptosis in Ara-G resistant cells may serve to explain the overall resistance to a variety of anti-neoplastic drugs.
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PMID:Resistance to mitochondrial- and Fas-mediated apoptosis in human leukemic cells with acquired resistance to 9-beta-D-arabinofuranosylguanosine. 1241 45

1-beta-D-arabinofuranosylcytosine (ara-C) is a deoxycytidine analog with activity in leukemia, which requires phosphorylation by deoxycytidine kinase (dCK) to allow formation of its active phosphate 1-beta-D-arabinofuranosylcytosine triphosphate, but can be deaminated by deoxycytidine deaminase. Altered membrane transport is also a mechanism of drug resistance. In order to facilitate ara-C uptake and prolong retention in the cell, lipophilic prodrugs were synthesized. Fatty acid groups with a varying acyl chain length and number of double bonds were esterified at the 5' position on the sugar moiety of ara-C. The compounds were tested in two pairs of ara-C resistant leukemic cell lines (murine L1210 and rat BCLO and their resistant variants L4A6 and Bara-C, respectively) and two pairs of cell lines with a resistance to gemcitabine, another deoxycytidine analog (human ovarian cancer A2780 and murine colon cancer C26-A and their resistant variants AG6000 and C26-G, respectively). L4A6, Bara-C and AG6000 have varying degrees of decreased dCK activity, while the mechanism for C26-G is not yet clear. In the parent cell lines, ara-C was more active, but in the resistant variants several of the analogs were more active, while the degree of cross-resistance varied. In AG6000 with a total dCK deficiency, all compounds were inactive. Structure-activity relation analysis showed that ara-C derivatives with shorter acyl chains and more double bonds were more active in the parental and drug resistant cells. Further mechanistic studies were performed with the elaidic acid derivative of ara-C (CP-4055). CP-4055 inhibited deamination of dCyd partly and induced DNA synthesis inhibition effectively in C26-A and C26-G cells, but the retention of inhibition was much longer for CP-4055 than for ara-C. In contrast to ara-C, CP-4055 inhibited RNA synthesis for 60% after drug exposure. In conclusion, CP-4055 seems to be a promising prodrug, whose effects were different and longer lasting than for the parent drug.
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PMID:Antiproliferative activity and mechanism of action of fatty acid derivatives of arabinofuranosylcytosine in leukemia and solid tumor cell lines. 1503 2

Cytarabine (araC) is a highly active antimetabolite against hematological malignancy while the agent shows limited activity against carcinomas. In this study, we focused on cellular transport and catalysis of the nucleoside in order to elucidate the mechanism of intrinsic resistance to araC in carcinomas. Activities of two metabolizing enzymes for araC, deoxycytidine kinase (DCK) and cytidine deaminase (CDA), and cellular transport of the agent were examined in 9 carcinoma cell lines. These variables in carcinoma lines were compared with those in 14 araC-sensitive leukemia lines and one leukemia line with acquired resistance. The mean IC50 in 9 carcinoma lines was 3 x 10(3)-fold higher than that in 14 leukemia lines (4.6 x 10(3) vs. 1.3 microM, p<0.01). A cell line with acquired resistance (U937R), which was established from U937 monocytoid leukemia cells, showed more than 10(3)-fold higher IC50 than the parent cells (1.6 x 10(3) vs. 1.3 microM). The resistance in carcinomas was associated with higher CDA activity and lower influx when compared to araC sensitive leukemias. Especially, these two types of malignant cell lines were clearly distinguished by CDA activity. The acquired resistance in U937R cells was followed by increase in cytidine deaminase (CDA) activity, decrease in DCK activity and decrease in influx of the drug. In conclusion, carcinomas are intrinsically resistant to cytarabine through high CDA activity and low cellular transport, but not low DCK activity. This finding suggests that treatment of carcinoma with deoxycytidine analogues should conquer the high CDA activity.
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PMID:Impact of cytidine deaminase activity on intrinsic resistance to cytarabine in carcinoma cells. 1549 2

To explain why 2-chloro-2'-deoxyadenosine (CdA) is unable to block DNA synthesis and cell cycle progression, and paradoxically enhances progression from G1 into S phase in the CdA-resistant leukemia EHEB cell line, we studied its metabolism and effects on proteins regulating the transition from G1 to S phase. A low deoxycytidine kinase activity and CdATP accumulation, and a lack of p21 induction despite p53 phosphorylation and accumulation may account for the inability of CdA to block the cell cycle. An alternative pathway involving pRb phosphorylation seems implicated in the CdA-induced increase in G1 to S phase progression.
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PMID:Effects of 2-chloro-2'-deoxyadenosine on the cell cycle in the human leukemia EHEB cell line. 1557 Dec 71

Recently, we reported a novel approach for the intracellular delivery of the anti-cancer nucleotide 5-fluoro-2'-deoxyuridine 5'-monophosphate (FdUMP) using phosphoramidate-based prodrugs. These phosphoramidate prodrugs contain an ester group that undergoes intracellular activation, liberating phosphoramidate anion, which in turn undergoes spontaneous cyclization and P-N bond cleavage to yield the nucleoside monophosphate quantitatively. This approach has now been extended to cytarabine [1-beta-D-arabinofuranosylcytosine (Ara-C)], an anti-cancer nucleoside that is limited in its utility because of poor intracellular transport characteristics and weak activity as a substrate for tumor cell kinases. The cytarabine phosphoramidate prodrug 1 has been synthesized and evaluated in comparison with cytarabine for growth inhibitory activity against wild-type, nucleoside transport-deficient, and nucleoside kinase-deficient CEM leukemia cell lines. The prodrug was comparable in growth inhibitory activity (IC50 = 32 nM) to cytarabine (IC50 = 16 nM) in wild-type CCRF-CEM cells following drug treatment for 72 h. The nucleoside transport-deficient CEM/AraC8C exhibited a high level of resistance (6400-fold) to cytarabine but was more sensitive (210-fold resistant vs CCRF-CEM cells) to prodrug 1. Similarly, the deoxycytidine kinase-deficient cell line (CEM/dCK-) was highly resistant to cytarabine (13900-fold) but more sensitive (106-fold resistant vs CCRF-CEM cells) to prodrug 1. These results indicate that prodrug 1 is significantly more potent than cytarabine against transport- and kinase-deficient cell lines and are consistent with a mechanism involving intracellular delivery of cytarabine 5'-monophosphate.
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PMID:Synthesis and biological evaluation of a cytarabine phosphoramidate prodrug. 1583 7

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