Determination of ddATP Levels in Human Immunodeficiency Virus-Infected Patients Treated with Dideoxyinosine

Saint, Cécile Le; Terreux, Raphaël; Duval, Danièle; Durant, J.; Ettesse, Helene; Dellamonica, P.; Guedj, R.; Vincent, Jean Pierre; Cupo, A.

doi:10.1128/aac.48.2.589-595.2004

Cited by 8 publications

(3 citation statements)

References 43 publications

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“…5). Immunoassays have been used without any sample separation (Goujon et al, 1998;Le Saint et al, 2004), but most methods require chromatographic separation before detection.…”

Section: Introductionmentioning

confidence: 99%

Mass spectrometry in the quantitative analysis of therapeutic intracellular nucleotide analogs

Jansen

Rosing

Schellens

et al. 2011

Mass Spectrometry Reviews

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Nucleoside analogs are widely used in anti-cancer, anti-(retro)viral, and immunosuppressive therapy. Nucleosides are prodrugs that require intracellular activation to mono-, di-, and finally triphosphates. Monitoring of these intracellular nucleotides is important to understand their pharmacology. The relatively involatile salts and ion-pairing agents traditionally used for the separation of these ionic analytes limit the applicability of mass spectrometry (MS) for detection. Both indirect and direct methods have been developed to circumvent this apparent incompatibility. Indirect methods consist of de-phosphorylation of the nucleotides into nucleosides before the actual analysis. Various direct approaches have been developed, ranging from the use of relatively volatile or very low levels of regular ion-pairing agents, hydrophilic interaction chromatography (HILIC), weak anion-exchange, or porous graphitic carbon columns to capillary electrophoresis and matrix-assisted light desorption--time of flight (MALDI-TOF) MS. In this review we present an overview of the publications describing the quantitative analysis of therapeutic intracellular nucleotide analogs using MS. The focus is on the different approaches for their direct analysis. We conclude that despite the technical hurdles, several useful MS-compatible chromatographic approaches have been developed, enabling the use of the excellent selectivity and sensitivity of MS for the quantitative analysis of intracellular nucleotides.

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“…5). Immunoassays have been used without any sample separation (Goujon et al, 1998;Le Saint et al, 2004), but most methods require chromatographic separation before detection.…”

Section: Introductionmentioning

confidence: 99%

Mass spectrometry in the quantitative analysis of therapeutic intracellular nucleotide analogs

Jansen

Rosing

Schellens

et al. 2011

Mass Spectrometry Reviews

View full text Add to dashboard Cite

show abstract

“…5). Immunoassays have been used without any sample separation (Goujon et al, 1998; Le Saint et al, 2004), but most methods require chromatographic separation before detection.…”

Section: Introductionmentioning

confidence: 99%

Integrin repertoire on myogenic cells changes during the course of primary myogenesis in the mouse

Cachaço

Pereira

Pardal

et al. 2005

Developmental Dynamics

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Here, we determined ␣ subunit expression patterns at embryonic day (E) 11.5-E14.5. Differentiated myotomal myocytes express all ␣ subunits studied. As the muscle masses form both in trunk (E12.5) and limbs (E11.5-E12.5), laminin receptors ␣6␤1 and ␣7␤1 are undetectable, and an assembled laminin matrix is absent. Instead ␣1␤1, ␣4␤1, ␣5␤1, and an ␣v-containing integrin are expressed and unassembled laminin and fibronectin are abundant around myogenic cells. At E13.5-E14.5, ␣6␤1 and ␣7␤1 are expressed, and a laminin matrix forms around individual myotubes. Thus, myogenic cells change their integrin expression pattern during the course of primary myogenesis in the mouse, suggesting different roles for fibronectin-and laminin-containing matrices in this process.

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“…RIA was used by raising antibodies against a stable isostere of ddATP. The three phosphorylated metabolites of ddI (ddAMP, ddADP and ddATP) had to be purified by anion exchange chromatography and the amount of each metabolite was determined with or without prior phosphatase treatment [7]. LC-MS has been applied to quantitate 2 0 ,3 0 -dideoxyadenosine (ddA) which was obtained from dephosphorylation of ddATP present in cellular extracts of cells treated with ddI.…”

Section: Introductionmentioning

confidence: 99%

Analysis of dideoxyadenosine triphosphate by CE with fluorescence detection. I. Derivatization through the phosphate group

2007

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A CZE method was developed, which separates 2',3'-dideoxyadenosine-5'-triphosphate (ddATP) from other metabolites and endogenous nucleotides at high concentrations (20-200 microg/mL) to allow UV detection. To enhance sensitivity, fluorescence detection which requires prior derivatization of compounds was examined. Precapillary derivatization of ddATP in the presence of N-(3-dimethylaminopropyl)-N'-ethylcarbodiimide hydrochloride (EDAC) with dansyl ethylenediamine (dansyl EDA) was faster and stable compared to that of 4,4-difluoro-5,7-dimethyl-4-bora-3a,4a-diaza-s-indacene-3-propionyl ethylenediamine (BODIPY FL EDA). Reaction conditions, reagent concentrations and detection parameters were optimized and highest derivatization efficiency was achieved in 0.1 M 1-methylimidazole buffer (pH 8.0) with 140 mM EDAC in 1-methylimidazole buffer and 30 mM dansyl EDA in DMF for 90 min at 60 degrees C. Dansyl EDA derivatives of ddATP, 2'-deoxyadenosine-5'-triphosphate (dATP) and ATP were comigrating with the CZE method; therefore, a MEKC method was developed and optimized for repeatable separations. Upon dansylation, sensitivity of ddATP with fluorescence detection (LOQ = 12 ng/mL) was 160 times higher than UV detection (LOQ = 1.9 microg/mL).

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Determination of ddATP Levels in Human Immunodeficiency Virus-Infected Patients Treated with Dideoxyinosine

Cited by 8 publications

References 43 publications

Mass spectrometry in the quantitative analysis of therapeutic intracellular nucleotide analogs

Mass spectrometry in the quantitative analysis of therapeutic intracellular nucleotide analogs

Integrin repertoire on myogenic cells changes during the course of primary myogenesis in the mouse

Analysis of dideoxyadenosine triphosphate by CE with fluorescence detection. I. Derivatization through the phosphate group

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