Special feature of mixed phosphotriester derivatives of cytarabine.

Despite the unquestionable therapeutic interest of bis(SATE) pronucleotides, a presystemic metabolism preventing the delivery of the prodrugs in target cancer cells or tumours may constitute a limitation to the in vivo development of such derivatives. In order to overcome these drawbacks several strategies have been envisaged and we report herein the application of the S-acyl-2-thioethyl (SATE) phenyl pronucleotide approach to the well-known cytotoxic nucleoside cytosine-1-beta-D-arabinofuranoside (cytarabine, araC). We describe modifications of the SATE moieties with the introduction of polar groups on the acyl residue, in order to study how these changes affect antitumoral activity and metabolic stability.

F-ara-AMP is a substrate of cytoplasmic 5'-nucleotidase II (cN-II): HPLC and NMR studies of enzymatic dephosphorylation.

Intracellular accumulation of triphosphorylated derivatives is essential for the cytotoxic activity of nucleoside analogues. Different mechanisms opposing this accumulation have been described. We have investigated the dephosphorylation of monophosphorylated fludarabine (F-ara-AMP) by the purified cytoplasmic 5'-nucleotidase cN-II using HPLC and NMR.

The prognostic value of cN-II and cN-III enzymes in adult acute myeloid leukemia.

We analyzed the expression of deoxycytidine kinase (dCK), UMP/CMP-kinase (UMP/CMP-K), nucleotide diphosphokinase (NDPK-B) and 5'-nucleotidases cN-II, cN-III, cdN and mdN by quantitative polymerase chain reaction at diagnosis in leukemic blasts from 96 patients with acute myeloid leukemia (AML) treated with ara-C. Our results show that high mRNA levels of cN-II and low mRNA levels of cN-III are correlated with a worse clinical outcome and suggest that these enzymes may have a role in sensitivity to ara-C in AML patients.

Problems related to resistance to cytarabine in acute myeloid leukemia.

First-line chemotherapy treatment in acute-myeloid leukemia patients usually consists of a combination of cytarabine (ara-C) and an anthracycline. These regimens induce complete response (CR) rates in 65-80% of newly diagnosed AML patients. However, clinical outcome is unsatisfactory, as most of the patients who achieve a CR will relapse within 2 years from diagnosis, often with resistant disease and poor response to subsequent therapy.

Characterization of a gemcitabine-resistant murine leukemic cell line: reversion of in vitro resistance by a mononucleotide prodrug.

Resistance to cytotoxic nucleoside analogues is a major problem in cancer treatment. The cellular mechanisms involved in this phenomenon have been studied for several years, and some factors have been identified. Various strategies to overcome resistance have been suggested, but none has yet shown efficacy in vivo.

Resistance to gemcitabine in a human follicular lymphoma cell line is due to partial deletion of the deoxycytidine kinase gene.

Background: Gemcitabine is an analogue of deoxycytidine with activity against several solid tumors. In order to elucidate the mechanisms by which tumor cells become resistant to gemcitabine, we developed the resistant subline RL-G from the human follicular lymphoma cell line RL-7 by prolonged exposure of parental cells to increasing concentrations of gemcitabine.

Results: In vitro, the IC50 increased from 0.

5'-(3')-nucleotidase mRNA levels in blast cells are a prognostic factor in acute myeloid leukemia patients treated with cytarabine.

We analyzed cytosolic 5'-(3')-nucleotidase (dNT-1) mRNA expression by quantitative polymerase chain reaction at diagnosis in leukemic blasts from 114 patients with acute myeloid leukemia (AML) treated with ara-C. Our results show that low dNT-1 mRNA expression in leukemic blasts at diagnosis is correlated with a worse clinical outcome and suggest that this enzyme may have a role in sensitivity to ara-C in AML patients.

Frameshift mutation in the Dok1 gene in chronic lymphocytic leukemia.

B-cell chronic lymphocytic leukemia (B-CLL) is a malignant disease characterized by an accumulation of monoclonal CD5+ mature B cells, with a high percentage of cells arrested in the G0/G1 phase of the cell cycle, and a particular resistance toward apoptosis-inducing agents. Dok1 (downstream of tyrosine kinases) is an abundant Ras-GTPase-activating protein (Ras-GAP)-associated tyrosine kinase substrate, which negatively regulates cell proliferation, downregulates MAP kinase activation and promotes cell migration. The gene encoding Dok1 maps to human chromosome 2p13, a region previously found to be rearranged in B-CLL.

Sensitization of ara-C-resistant lymphoma cells by a pronucleotide analogue.

Adequate intracellular concentrations of ara-CMP, the monophosphorylated derivative of ara-C, are essential for its cytotoxicity. The critical step for ara-CMP formation is intracellular phosphorylation of ara-C by deoxycytidine kinase (dCK). A common nucleoside resistance mechanism is mutation affecting the expression or the specificity of dCK.

Deoxycytidine kinase and cN-II nucleotidase expression in blast cells predict survival in acute myeloid leukaemia patients treated with cytarabine.

The cytotoxic activity of cytarabine (ara-C) in leukaemic blasts depends on activating enzymes such as deoxycytidine kinase (dCK) and inactivating enzymes such as the 5'-nucleotidases. We have analysed dCK and 'high-Km' 5'-nucleotidase (cN-II) mRNA expression by the quantitative real-time polymerase chain reaction at diagnosis in leukaemic blasts from 115 acute myeloid leukaemia (AML) patients treated with ara-C. The prognostic value of these parameters as well as that of the cN-II/dCK ratio was determined.

Influence of p53 and p21(WAF1) expression on sensitivity of cancer cells to cladribine.

The present study was performed to gain insight into the role of p53 and p21(WAF1) on the cytotoxicity of the purine analogue cladribine (2-CdA) on cancer cells. Drug sensitivity, cell cycle distribution and drug-induced cell death were compared in three lines derived from the colorectal carcinoma HCT116: the p53+/+ cell line containing wild-type p53 and the p53-/- and p21(WAF1)-/- lines, in which both alleles of p53 or p21(WAF1) were deleted by homologous recombination, respectively. p53-/- and p21(WAF1)-/- cells were significantly more resistant to the cytotoxic effects of 2-CdA than the p53+/+ cells.

In vivo mechanisms of resistance to cytarabine in acute myeloid leukaemia.

Factors that reduce the intracellular concentration of triphosphorylated cytarabine (ara-CTP), the active form of cytarabine (ara-C), may induce chemoresistance in acute myeloid leukaemia (AML) patients. These factors include reduced influx of ara-C by the hENT1 transporter, reduced phosphorylation by deoxycytidine kinase (dCK), and increased degradation by high Km cytoplasmic 5'-nucleotidase (5NT) and/or cytidine deaminase (CDD). Increased levels of DNA polymerase alpha (DNA POL) and reduced levels of topoisomerase I (TOPO I) and topoisomerase II (TOPO II) have also been detected in ara-C-resistant cell lines.

Potential mechanisms of resistance to cytarabine in AML patients.

To determine whether the human equilibrative nucleoside transporter 1 (hENT1), deoxycytidine kinase (dCK), cytoplasmic 5'-nucleotidase (5NT), cytidine deaminase (CDD), topoisomerase I (TOPO I) and topoisomerase II alpha (TOPO II) are involved in clinical resistance to cytarabine (ara-C), we analyzed the level of expression of these parameters by reverse transcriptase polymerase chain reaction (rt-PCR), at diagnosis in the blast cells of 77 acute myeloid leukemia (AML) patients treated with ara-C, including 31 for whom samples were collected at first relapse. By univariate and/or multivariate analyses, patients with expression of 5NT or hENT1 deficiency at diagnosis had significantly shorter disease-free survival (DFS) and overall survival (OS). These results suggest that expression of 5NT and reduced hENT1 in leukemic blasts at diagnosis are correlated with clinical outcome and may play a role in resistance mechanisms to ara-C in patients with AML.