Cytotoxic activity of 2′,2′-difluorodeoxycytidine, 5-aza-2′-deoxycytidine and cytosine arabinoside in cells transduced with deoxycytidine kinase gene

Beauséjour, Christian; Gagnon, Jacynthe; Primeau, Mélanie; Momparler, Richard L.

doi:10.1016/s0006-291x(02)00413-8

Cited by 28 publications

(22 citation statements)

References 23 publications

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“…Typically, phosphorylation of a nucleoside to its monophosphate is a rate-limiting step for the activation of many cytidine analogs (1,10). In order to understand the phosphorylation of PSI-6130 to PSI-6130-MP, human dCK and UCK-1 were cloned and purified.…”

Section: Discussionmentioning

confidence: 99%

Mechanism of Activation of β- d -2′-Deoxy-2′-Fluoro-2′- C -Methylcytidine and Inhibition of Hepatitis C Virus NS5B RNA Polymerase

Murakami¹,

Bao²,

Ramesh³

et al. 2007

Antimicrob Agents Chemother

101

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) is a potent specific inhibitor of hepatitis C virus (HCV) RNA synthesis in Huh-7 replicon cells. To inhibit the HCV NS5B RNA polymerase, PSI-6130 must be phosphorylated to the 5-triphosphate form. The phosphorylation of PSI-6130 and inhibition of HCV NS5B were investigated. The phosphorylation of PSI-6130 by recombinant human 2-deoxycytidine kinase (dCK) and uridine-cytidine kinase 1 (UCK-1) was measured by using a coupled spectrophotometric reaction. PSI-6130 was shown to be a substrate for purified dCK, with a K m of 81 M and a k cat of 0.007 s ؊1 , but was not a substrate for UCK-1. PSI-6130 monophosphate (PSI-6130-MP) was efficiently phosphorylated to the diphosphate and subsequently to the triphosphate by recombinant human UMP-CMP kinase and nucleoside diphosphate kinase, respectively. The inhibition of wild-type and mutated (S282T) HCV NS5B RNA polymerases was studied. The steady-state inhibition constant (K i ) for PSI-6130 triphosphate (PSI-6130-TP) with the wild-type enzyme was 4.3 M. Similar results were obtained with 2-C-methyladenosine triphosphate (K i ‫؍‬ 1.5 M) and 2-C-methylcytidine triphosphate (K i ‫؍‬ 1.6 M). NS5B with the S282T mutation, which is known to confer resistance to 2-C-methyladenosine, was inhibited by PSI-6130-TP as efficiently as the wild type. Incorporation of PSI-6130-MP into RNA catalyzed by purified NS5B RNA polymerase resulted in chain termination.Hepatitis C virus (HCV) is an RNA virus which possesses a single-stranded positive-sense RNA as the viral genome. This viral RNA plays important roles during viral replication, as it serves as an mRNA for viral protein synthesis, a template for RNA replication, and a nascent RNA genome for a newly formed virus (17). HCV NS5B RNA-dependent RNA polymerase is a key enzyme in viral RNA replication. This enzyme, which does not require a primer for initiation of RNA synthesis, catalyzes de novo RNA synthesis (8, 11). Nucleoside analogs have been used to treat viral infections, such as herpes simplex virus, human immunodeficiency virus, and hepatitis B virus infections (5,6,21). These drugs are designed to inhibit viral polymerases by a process called chain termination, in which DNA synthesis is quenched by incorporating the triphosphate forms of these drugs, which lack the 3Ј-hydroxyl group on the sugar moiety. In order for nucleoside analogs to inhibit a viral polymerase, they must be transported into the cell and converted to the active 5Ј-triphosphate form by cellular kinases. 2Ј-C-Methylnucleosides have been investigated as anti-HCV agents targeting HCV NS5B RNA polymerase (2, 20). 2Ј-C-Methyladenosine (2Ј-C-Me-A) and 2Ј-C-methylguanosine (2Ј-C-Me-G) showed potent anti-HCV activities in a cell-based replicon assay, and their triphosphate forms inhibited replicase and NS5B RNA polymerase in vitro (20). In addition, 2Ј-C-Me-A exhibited significant activity against HCV in a cell culture system which involves complete HCV replication and which produces infectious HCV (16). A resistant replicon has been selected by passage of HCV in the...

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Section: Discussionmentioning

confidence: 99%

Mechanism of Activation of β- d -2′-Deoxy-2′-Fluoro-2′- C -Methylcytidine and Inhibition of Hepatitis C Virus NS5B RNA Polymerase

Murakami¹,

Bao²,

Ramesh³

et al. 2007

Antimicrob Agents Chemother

101

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“…To test the activity of the drugs in vitro, we selected a non-small cell lung cancer cell line (A549) (29), a colon cancer cell line (WiDR) (30) and a variant of the WiDR cell line that has been cultured under low folate conditions (WiDR-LF). Normal cell culture medium contains supraphysiological folate levels, which reduce the effect of antifolates.…”

Section: Cell Linesmentioning

confidence: 99%

Cell cycle effects of fatty acid derivatives of cytarabine, CP-4055, and of gemcitabine, CP-4126, as basis for the interaction with oxaliplatin and docetaxel

Peters¹

2009

Int J Oncol

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Abstract. To bypass resistance due to limited entry into the cell derivatives of cytarabine (CP-4055, elacytarabine) and gemcitabine (CP-4126) containing a fatty acid chain at the 5' position of the nucleoside were developed. CP-4055 showed an increased retention of the active metabolite, the triphosphate. This characteristic was supposed to favor combinations, such as with the tubulin antagonist docetaxel, the platinum oxaliplatin and the antifolate pemetrexed. The role of the cell cycle effects of CP-4055 and CP-4126 on the efficacy of the combination with docetaxel or pemetrexed was determined. The combination of CP-4055 with oxaliplatin and docetaxel was also evaluated in a mouse xenograft model. CP-4055 induced a G2/M and S phase accumulation and CP-4126 an S phase accumulation. Both analogs induced a dose-dependent cell kill (apoptosis and necrosis). None of the docetaxel combinations induced a synergistic effect. The combination of docetaxel with CP-4055 or CP-4126 induced a G2/M accumulation in the A549 (lung cancer) cell line, but a G0/G1 accumulation in the WiDR (colon cancer) cell line. Preincubation with docetaxel induced an increased cell kill in both cell lines. The combination with oxaliplatin showed a synergistic effect in both cell lines. Combinations with pemetrexed were antagonistic in both cell lines. In the A549 cell line pemetrexed with CP-4055 led to an increase of the G0/G1 phase and the S phase. In WiDR the combination of pemetrexed with CP-4055 increased the G0/G1 phase and increased the cell kill. Pemetrexed with CP-4126 induced an increase in the G0/G1 phase and the S phase in the A549 cell line. In the xenograft study, on a colon cancer and a lung metastasis model, the combination of CP-4055 with docetaxel showed the best results. Treatment with CP-4055 followed by docetaxel after 4 h resulted in a reduction in metastasis in a lung metastasis model, and a favorable toxicity profile was observed. In conclusion, the combinations with oxaliplatin showed a synergistic effect in the combination studies. Although the combinations with docetaxel did not show an enhanced effect in the in vitro studies, this combination revealed an increased effect in the xenograft model.

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“…Blackstock et al 34 used the same retroviral construct in HT-29 cells. Beause´jour et al 35 utilized another retroviral construct to transfect the human dCK gene into human A549 lung carcinoma and murine 36 transduced the Colon 26 mouse colon carcinoma and 9L rat gliosarcoma cell lines with another human dCK encoding retroviral vector. None of these authors mentioned signs of increased toxicity in vitro, but one should take into account that much higher dCK levels can be achieved with adenoviral vectors than with retroviruses.…”

Section: -31mentioning

confidence: 99%

“…They showed a strong correlation between dCK activity and 1-b-D-arabinofuranosylcytosine (AraC), 2-chloro-2 0 -deoxyadenosine (CdA) or 2-fluoro-9-b-D-arabinofuranosyladenine (FAraA) toxicity, but a lesser level of correlation existed between dCK activity and gemcitabine toxicity. Beause´jour and co-workers 35 reported that dCK overexpression could increase AraC and 5-aza-2 0 -deoxycytidine (5-aza-CdR) toxicity in human A-549 and murine NIH 3T3 cell lines, but led to an unexpected drug resistance against gemcitabine. Our results show that increased dCK activity upon adenoviral transfer of the gene could sensitize cells to gemcitabine.…”

Section: -31mentioning

confidence: 99%

Adenoviral vector transduction of the human deoxycytidine kinase gene enhances the cytotoxic and radiosensitizing effect of gemcitabine on experimental gliomas

et al. 2008

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The aim of this work was to improve the cytotoxic and radiosensitizing effects of gemcitabine using a gene-directed enzyme prodrug therapy approach. Murine Gl261, rat C6 and human U373 glioma cell lines were transduced with an adenoviral vector encoding the human deoxycytidine kinase gene (Ad-HudCK). Intracranial tumors were established in C57BL/6 mice and Wistar rats using either wild-type or Ad-HudCK-transduced Gl261 and C6 glioma cells. In vitro growing cells and established tumors were treated with gemcitabine and irradiation either alone or in combination. Deoxycytidine kinase overexpression substantially increased both the toxic and radiosensitizing effects of gemcitabine in each cell line, but the enhancement rate varied: it was mild in the Gl261 cells and much stronger in the C6 and U373 cells. In vivo experiments showed a mild radiosensitizing effect of dCK overexpression both in the Gl261 and C6 models. The combination of dCK overexpression, gemcitabine treatment and irradiation improved the survival rate of C6 bearing rats significantly. In conclusion, overexpression of the dCK gene can improve the cytotoxic and radiosensitizing effect of gemcitabine both in vitro and in vivo in a tumor-specific manner.

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Cytotoxic activity of 2′,2′-difluorodeoxycytidine, 5-aza-2′-deoxycytidine and cytosine arabinoside in cells transduced with deoxycytidine kinase gene

Cited by 28 publications

References 23 publications

Mechanism of Activation of β- d -2′-Deoxy-2′-Fluoro-2′- C -Methylcytidine and Inhibition of Hepatitis C Virus NS5B RNA Polymerase

Mechanism of Activation of β- d -2′-Deoxy-2′-Fluoro-2′- C -Methylcytidine and Inhibition of Hepatitis C Virus NS5B RNA Polymerase

Cell cycle effects of fatty acid derivatives of cytarabine, CP-4055, and of gemcitabine, CP-4126, as basis for the interaction with oxaliplatin and docetaxel

Adenoviral vector transduction of the human deoxycytidine kinase gene enhances the cytotoxic and radiosensitizing effect of gemcitabine on experimental gliomas

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