Effects of granulocyte-macrophage colony-stimulating factor on 3′-azido-3′-deoxythymidine uptake, phosphorylation and nucleotide retention in human U-937 cells

doi:10.1016/0006-2952(90)90589-D

Biochemical Pharmacology

Volume 40, Issue 12, 15 December 1990, Pages 2695-2700

https://doi.org/10.1016/0006-2952(90)90589-D Get rights and content

Abstract

Previous studies have demonstrated that granulocyte-macrophage colony-stimulating factor (GM-CSF) both increases and decreases levels of 3′-azido-3′-deoxythymidine (AZT) nucleotides in certain human myeloid cells. The present studies have examined the effects of GM-CSF on AZT metabolism in U-937 cells. The results demonstrate that GM-CSF stimulated AZT nucleotide formation in these cells. This stimulation was detectable during concurrent exposure to GM-CSF and AZT or as a result of pretreatment with GM-CSF. The GM-CSF-induced enhancement in AZT nucleotide formation was associated with a 4-fold increase in AZT uptake. The finding that uptake of AZT into U-937 cells was only partially sensitive to 6-[(4-nitrobenzyl)thio]-9-β-d-ribofuranosylpurine (NBMPR) suggested a process primarily involving nonfacilitated diffusion. The results also demonstrate that treatment of U-937 cells with GM-CSF was associated with nearly a 2-fold increase in thymidine kinase activity. Moreover, the findings indicate that retention of AZT-MP and AZP-TP was prolonged significantly (P < 0.05 and P < 0.01 respectively) in association with GM-CSF treatment. Taken together, these results suggest that GM-CSF enhances the formation of AZT nucleotides by increasing AZT uptake and phosphorylation, as well as increasing retention of phosphorylated derivatives.

References (23)

H Mitsuya et al.
3′-Azido-3′-deoxythymidine (BW A509U): An antiviral agent that inhibits the infectivity and cytopathic effect of human T-lymphotropic virus type III/lymphoadenopathy-associated virus in vitro
H Mitsuya et al.
Inhibition of the in vitro infectivity and cytopathic effect of human T-lymphotrophic virus type III/lymphoadenopathy-associated virus (HTLV-III/LAV), by 2',3'-dideoxy nucleosides
CF Perno et al.
Inhibition of human immunodeficiency virus (HIV/1/HTLV-III_Ba-L) replication in fresh and cultured human peripheral blood monocyte/macrophages by azidothymidine and related 2',3'-dideoxynucleosides
J Exp Med
(1988)
PA Furman et al.
Phosphorylation of 3'-azido-3'-deoxythymidine and selective interaction of the 5'-triphosphate with human immunodeficiency virus reverse transcriptase
K Ono et al.
Inhibition of reverse transcriptase activity by 2',3'-dideoxythymidine-5'-triphosphate and its derivatives modified on the 3'-position
Biochem Biophys Res Commun
(1986)
MH St. Clair et al.
3'-Azido-3'-deoxythymidine triphosphate as an inhibitor and substrate of purified human immunodeficiency virus reverse transcriptase
Antimicrob Agents Chemother
(1987)
Y-C Cheng et al.
Human immunodeficiency virus reverse transcriptase. General properties and its interactions with nucleoside triphosphate analogs
J Biol Chem
(1987)
CF Perno et al.
Replication of human immunodeficiency virus in monocytes. Granulocyte/macrophage colony-stimulating factor (GM-CSF) potentiates viral production yet enhances the antiviral effect mediated by 3'-azido-2',3'-dideoxythymdiine (AZT) and other dideoxynucleoside congeners of thymidine
J Exp Med
(1989)
K Bhalla et al.
The effect of recombinant human granulocyte-macrophage colony-stimulating factor (rGM-CSF) on 3'-azido-3'-deoxythymidine (AZT)-mediated biochemical and cytotoxic effects of normal human myeloid progenitor cells
Exp Hematol
(1989)
C Sundstrom et al.
Establishment and characterization of a human histiocytic lymphoma cell line (U-937)
Int J Cancer
(1976)

K Bhalla et al.

Isolation and characterization of a deoxycytidine kinase deficient human promyelocytic leukemic cell line highly resistant to 1-β-d-arabinofuranosylcytosine

Cancer Res

(1984)

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Granulocyte macrophage colony-stimulating factor has come of age: From a vaccine adjuvant to antiviral immunotherapy
2021, Cytokine and Growth Factor Reviews
Citation Excerpt :
Nonetheless, a finding that is universal throughout the literature is that GM-CSF increases CD4 + T cell counts [97,101,104,105], which would be beneficial to the patient during the infection. GM-CSF has also been demonstrated to augment the effectiveness of the antiretroviral agent, zidovudine (AZT), in HIV infected macrophages in vitro [106,107] by upregulating thymidine kinase [107,108] to increase intracellular phosphorylation of AZT into its active triphosphate form [109]. In the context of HIV-1 infection of macrophages, other cellular markers in addition to CD4 have been investigated for their role in enabling HIV-1 infection of cells.
GM-CSF acts as a pro-inflammatory cytokine and a key growth factor produced by several immune cells such as macrophages and activated T cells. In this review, we discuss recent studies that point to the crucial role of GM-CSF in the immune response against infections. Upon induction, GM-CSF activates four main signalling networks including the JAK/STAT, PI3K, MAPK, and NFκB pathways. Many of these transduction pathways such as JAK/STAT signal via proteins commonly activated with other antiviral signalling cascades, such as those induced by IFNs.
GM-CSF also helps defend against respiratory infections by regulating alveolar macrophage differentiation and enhancing innate immunity in the lungs. Here, we also summarize the numerous clinical trials that have taken advantage of GM-CSF’s mechanistic attributes in immunotherapy. Moreover, we discuss how GM-CSF is used as an adjuvant in vaccines and how its activity is interfered with to reduce inflammation such as in the case of COVID-19. This review brings forth the current knowledge on the antiviral actions of GM-CSF, the associated signalling cascades, and its application in immunotherapy.
A physiologically based pharmacokinetic model of zidovudine (AZT) in the mouse: Model development and scale-up to humans
1997, Journal of Pharmaceutical Sciences
After having been used in treating HIV infection for a decade, zidovudine (AZT) continues to be an essential component of antiretroviral regimens. Because antiviral responses and toxicity of AZT seem to be related to cells in specific target tissues, being able to understand and predict the distribution of AZT into different pharmacologically and toxicologically relevant tissues is therefore critically important to improving the efficacy and minimizing the toxicity of AZT therapy. This study was designed to develop a physiologically based pharmacokinetic model to help describe and predict the time course of AZT levels in different tissues. The model was developed in the mouse and then scaled up to predict human situations.
Hematologic disorders and growth factor support in HIV infection
1996, Hematology/Oncology Clinics of North America
HIV disease highlights the continuum that links hematology to immunology. It is thought that the direct cellular interactions of HIV-1 are quite restricted. Yet a pancellular abnormality of the blood and immune systems develops that alters not just host defense but also the oxygen-carrying and coagulation function of these systems as well. Although the CD4+ T lymphocyte legitimately remains the focus of attention regarding the pathophysiology of AIDS, this article reviews the manifestations and possible mechanisms by which HIV-1 affects other dimensions of the lymphohematopoietic system.
Metabolism of zidovudine
1995, General Pharmacology
- 1.
  1. The anti-HIV drug zidovudine (3'-azido-2',3'-dideoxythymidine; ZDV) has three important pathways of metabolism. ZDV is a prodrug and must be phosphorylated in lymphocytes in order to exert its antiviral action. However, in quantitative terms this is a minor pathway probably accounting for less than 1% of the overall metabolic profile. The predominant pathway of metabolism is glucuronidation to GZDV and the metabolite is renally excreted. A further metabolite, derived by reduction of the azido moiety is 3'-amino-3'-deoxythymidine (AMT).
- 2.
  2. Zidovudine glucuronidation has been characterised in human liver microsomes. A number of drugs (e.g., naproxen, indomethacin and probenecid) have been shown to inhibit the in vitro conjugation of ZDV. Some of these drugs have also been co-administered with ZDV in HIV-positive patients. Significant pharmacokinetic interactions have been demonstrated with probenecid, naproxen and fluconazole.
- 3.
  3. 3'-amino-3'-deoxythymidine formation is probably mediated by both cytochrome P450 isozymes and NADPH-cytochrome P450 reductase. Peak plasma concentrations of AMT are approximately 10–15% of ZDV in patients. This is a potentially important metabolite because of its alleged cytotoxicity.
- 4.
  4. Measurement of intracellular ZDV phosphates in patients provides the key to our understanding of both the efficacy and toxicity of ZDV. Important recent work has demonstrated that as patients deteriorate (i.e., CD4 counts decrease below 100 × 10⁶/L), there is a corresponding increase in intracellular ZDV-monophosphate. This could have toxicological implications.
Simultaneous analysis of azidothymidine and its mono-, di- and triphosphate derivatives in biological fluids, tissue and cultured cells by a rapid high-performance liquid chromatographic method
1992, Journal of Chromatography B: Biomedical Sciences and Applications
A rapid high-performance liquid chromatographic (HPLC) method for the simultaneous analysis of the antiviral drug azidothymidine (AZT), AZT monophosphate, AZT diphosphate and AZT triphosphate, with ultraviolet detection in the nanomolar range, is described. Determination of these compounds in vitro in the human MT-4 lymphocyte cell line did not require a prior extraction, and AZT and its phosphorylated derivatives could be accurately analysed in one HPLC run. However, plasma, bile, liver homogenate and urine samples could not be injected directly into the chromatograph. Therefore, a solid-phase extraction procedure was developed, using azidodideoxyinosine as internal standard. The extractions of the compounds of interest from all but urine samples were reproducible, with recoveries between 65% (AZT triphosphate from plasma) and 100% (AZT from plasma).
Different pattern of activity of inhibitors of the human immunodeficiency virus in lymphocytes and monocyte/macrophages
1992, Antiviral Research
Monocyte/macrophages (M/M) are important targets for HIV in the body, and represent the majority of cells infected by the virus in some body compartments such as the central nervous system (CNS). M/M can be different from T-lymphocytes in terms of surface antigens, cell replication and drug metabolism. Thus, we evaluated, in M/M and in T-lymphocytes, the pattern of viral inhibition induced by various anti-HIV drugs, and assessed some of the mechanisms of action related to such antiviral activity.
Inhibitors of HIV binding on CD4 receptors have similar activity in M/M and T-lymphocytes, while AZT and other dideoxynucleosides (ddN) are in general more active against HIV in M/M than in T-lymphocytes. This phenomenon can be related to the increased ratio in M/M of ddN-triphosphate/deoxynucleoside-triphosphate, and can at least in part explain the ability of zidovudine and didanosine in improving neurological dysfunctions in AIDS patients. Moreover, the antiviral activity of AZT (but not of other ddN- or HIV-binding inhibitors) is potently enhanced by cytokines like granulocyte-macrophage colony stimulating factor (GM-CSF) in M/M, while anti-HIV activity of TIBO compounds in M/M is not down-modulated by GM-CSF and other cytokines. Finally, non-toxic concentrations of adriamycin, an anticancer drug reported to be active against DNA viruses, can inhibit HIV replication in M/M (but not in T-lymphocytes).
Taken together, these results suggest that M/M are selective targets for HIV with peculiarities different from those of T-lymphocytes. Thus, promising anti-HIV compounds should be evaluated both in T-cells and in M/M before reaching clinical trials. This may help in selecting drugs with good chances of being effective in patients with HIV-related disease.

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Biochemical Pharmacology

Abstract

References (23)

3′-Azido-3′-deoxythymidine (BW A509U): An antiviral agent that inhibits the infectivity and cytopathic effect of human T-lymphotropic virus type III/lymphoadenopathy-associated virus in vitro

Inhibition of the in vitro infectivity and cytopathic effect of human T-lymphotrophic virus type III/lymphoadenopathy-associated virus (HTLV-III/LAV), by 2',3'-dideoxy nucleosides

Inhibition of human immunodeficiency virus (HIV/1/HTLV-III_Ba-L) replication in fresh and cultured human peripheral blood monocyte/macrophages by azidothymidine and related 2',3'-dideoxynucleosides

J Exp Med

Phosphorylation of 3'-azido-3'-deoxythymidine and selective interaction of the 5'-triphosphate with human immunodeficiency virus reverse transcriptase

Inhibition of reverse transcriptase activity by 2',3'-dideoxythymidine-5'-triphosphate and its derivatives modified on the 3'-position

Biochem Biophys Res Commun

3'-Azido-3'-deoxythymidine triphosphate as an inhibitor and substrate of purified human immunodeficiency virus reverse transcriptase

Antimicrob Agents Chemother

Human immunodeficiency virus reverse transcriptase. General properties and its interactions with nucleoside triphosphate analogs

J Biol Chem

Replication of human immunodeficiency virus in monocytes. Granulocyte/macrophage colony-stimulating factor (GM-CSF) potentiates viral production yet enhances the antiviral effect mediated by 3'-azido-2',3'-dideoxythymdiine (AZT) and other dideoxynucleoside congeners of thymidine

J Exp Med

The effect of recombinant human granulocyte-macrophage colony-stimulating factor (rGM-CSF) on 3'-azido-3'-deoxythymidine (AZT)-mediated biochemical and cytotoxic effects of normal human myeloid progenitor cells

Exp Hematol

Establishment and characterization of a human histiocytic lymphoma cell line (U-937)

Int J Cancer

Isolation and characterization of a deoxycytidine kinase deficient human promyelocytic leukemic cell line highly resistant to 1-β-d-arabinofuranosylcytosine

Cancer Res

Cited by (15)

Granulocyte macrophage colony-stimulating factor has come of age: From a vaccine adjuvant to antiviral immunotherapy

A physiologically based pharmacokinetic model of zidovudine (AZT) in the mouse: Model development and scale-up to humans

Hematologic disorders and growth factor support in HIV infection

Metabolism of zidovudine

Simultaneous analysis of azidothymidine and its mono-, di- and triphosphate derivatives in biological fluids, tissue and cultured cells by a rapid high-performance liquid chromatographic method

Different pattern of activity of inhibitors of the human immunodeficiency virus in lymphocytes and monocyte/macrophages

Effects of granulocyte-macrophage colony-stimulating factor on 3′-azido-3′-deoxythymidine uptake, phosphorylation and nucleotide retention in human U-937 cells

Abstract

3′-Azido-3′-deoxythymidine (BW A509U): An antiviral agent that inhibits the infectivity and cytopathic effect of human T-lymphotropic virus type III/lymphoadenopathy-associated virus in vitro

Inhibition of the in vitro infectivity and cytopathic effect of human T-lymphotrophic virus type III/lymphoadenopathy-associated virus (HTLV-III/LAV), by 2',3'-dideoxy nucleosides

Inhibition of human immunodeficiency virus (HIV/1/HTLV-IIIBa-L) replication in fresh and cultured human peripheral blood monocyte/macrophages by azidothymidine and related 2',3'-dideoxynucleosides

J Exp Med

Phosphorylation of 3'-azido-3'-deoxythymidine and selective interaction of the 5'-triphosphate with human immunodeficiency virus reverse transcriptase

Inhibition of reverse transcriptase activity by 2',3'-dideoxythymidine-5'-triphosphate and its derivatives modified on the 3'-position

Biochem Biophys Res Commun

3'-Azido-3'-deoxythymidine triphosphate as an inhibitor and substrate of purified human immunodeficiency virus reverse transcriptase

Antimicrob Agents Chemother

Human immunodeficiency virus reverse transcriptase. General properties and its interactions with nucleoside triphosphate analogs

J Biol Chem

Replication of human immunodeficiency virus in monocytes. Granulocyte/macrophage colony-stimulating factor (GM-CSF) potentiates viral production yet enhances the antiviral effect mediated by 3'-azido-2',3'-dideoxythymdiine (AZT) and other dideoxynucleoside congeners of thymidine

J Exp Med

The effect of recombinant human granulocyte-macrophage colony-stimulating factor (rGM-CSF) on 3'-azido-3'-deoxythymidine (AZT)-mediated biochemical and cytotoxic effects of normal human myeloid progenitor cells

Exp Hematol

Establishment and characterization of a human histiocytic lymphoma cell line (U-937)

Int J Cancer

Isolation and characterization of a deoxycytidine kinase deficient human promyelocytic leukemic cell line highly resistant to 1-β-d-arabinofuranosylcytosine

Cancer Res

Inhibition of human immunodeficiency virus (HIV/1/HTLV-III_Ba-L) replication in fresh and cultured human peripheral blood monocyte/macrophages by azidothymidine and related 2',3'-dideoxynucleosides