Development and validation of a useful UPLC–MS/MS method for quantification of total and phosphorylated-ribavirin in peripheral blood mononuclear cells of HCV+ patients

Agnesod, Danilo; Nicolò, Amedeo De; Simiele, Marco; Abdi, Adnan Mohamed; Boglione, Lucio; Perri, Giovanni Di; D’Avolio, Antonio

doi:10.1016/j.jpba.2013.11.027

Cited by 21 publications

(14 citation statements)

References 47 publications

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“…TAC and EVE amounts of all patients' samples were in the calibration curve range. Concentrations from each patient were calculated using the personalized MCV . It was observed that intra‐PBMCs concentration was meanly 19.23 and 218.61 times higher than the hematic one for TAC and EVE, respectively.…”

Section: Resultssupporting

confidence: 81%

First UHPLC-MS/MS method coupled with automated online SPE for quantification both of tacrolimus and everolimus in peripheral blood mononuclear cells and its application on samples from co-treated pediatric patients.

Pensi

Nicolò²,

Pinon

et al. 2017

J. Mass Spectrom.

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Tacrolimus (TAC, FK-506) and everolimus (EVE, RAD001) are immunosuppressors used to treat pediatric patients undergoing liver transplantation. Their hematic TDM by liquid chromatography became standard practice. However, it does not always reflect concentrations at their active site. Our aim was to develop and validate a new method for the simultaneous TAC and EVE quantification into target cells: peripheral blood mononuclear cells (PBMCs). Peripheral blood mononuclear cells were collected using cell preparation tubes; cells number and mean cell volume were evaluated by an automatic cell counter. TAC and EVE were quantified using UHPLC-MS/MS coupled with an automated online solid-phase extraction platform. Chromatographic run was performed on an Acquity UPLC® BEH C18 1.7 μm (2.1 × 50 mm) column at 45 °C, for 6 min at 0.5 ml/min. Mobile phases were water and methanol, both with 2 mm ammonium acetate and 1 ml/l formic acid). XBridge® C8 10 μm (1 × 10 mm) SPE cartridges were used, and the internal standard was ascomycin. Following Food and Drug Administration guidelines, method validation resulted in high sensitivity and specificity. Calibration curves were linear (r = 0.998) and intra-day and inter-day imprecision and inaccuracy were <15%. A reproducible matrix effect was observed, with a good recovery for all compounds. Drug amounts in 15 'real' PBMCs samples from five pediatric patients in co-treatment resulted within the calibration range (0.039-5 ng). Concentrations from each patient were standardized using their evaluated mean cell volume: intra-PBMCs concentration was meanly 19.23 and 218.61 times higher than the hematic one for TAC and EVE, respectively. This method might be useful in clinical routine, giving reliable data on drugs concentration at the active site. Copyright © 2017 John Wiley & Sons, Ltd.

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Section: Resultssupporting

confidence: 81%

First UHPLC-MS/MS method coupled with automated online SPE for quantification both of tacrolimus and everolimus in peripheral blood mononuclear cells and its application on samples from co-treated pediatric patients.

Pensi

Nicolò²,

Pinon

et al. 2017

J. Mass Spectrom.

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“…Consequently, the development of a parallel method for the intracellular quantification of remdesivir and of the triphosphate active form in the near future is guaranteed, following an already tested protocol. [26][27][28]…”

Section: Discussionmentioning

confidence: 99%

Development and validation of a UHPLC-MS/MS method for quantification of the prodrug remdesivir and its metabolite GS-441524: a tool for clinical pharmacokinetics of SARS-CoV-2/COVID-19 and Ebola virus disease

Avataneo

Nicolò

Cusato

et al. 2020

Journal of Antimicrobial Chemotherapy

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Background: Remdesivir has received significant attention for its potential application in the treatment of COVID-19, caused by SARS-CoV-2. Remdesivir has already been tested for Ebola virus disease treatment and found to have activity against SARS and MERS coronaviruses. The remdesivir core contains GS-441524, which interferes with RNA-dependent RNA polymerases alone. In non-human primates, following IV administration, remdesivir is rapidly distributed into PBMCs and converted within 2 h to the active nucleoside triphosphate form, while GS-441524 is detectable in plasma for up to 24 h. Nevertheless, remdesivir pharmacokinetics and pharmacodynamics in humans are still unexplored, highlighting the need for a precise analytical method for remdesivir and GS-441524 quantification.Objectives: The validation of a reliable UHPLC-MS/MS method for remdesivir and GS-441524 quantification in human plasma.Methods: Remdesivir and GS-441524 standards and quality controls were prepared in plasma from healthy donors. Sample preparation consisted of protein precipitation, followed by dilution and injection into the QSight 220 UHPLC-MS/MS system. Chromatographic separation was obtained through an Acquity HSS T3 1.8 lm, 2.1%50 mm column, with a gradient of water and acetonitrile with 0.05% formic acid. The method was validated using EMA and FDA guidelines.Results: Analyte stability has been evaluated and described in detail. The method successfully fulfilled the validation process and it was demonstrated that, when possible, sample thermal inactivation could be a good choice in order to improve biosafety.Conclusions: This method represents a useful tool for studying remdesivir and GS-441524 clinical pharmacokinetics, particularly during the current COVID-19 outbreak.

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“…After separation, PBMC samples were stored at −80 • C until analysis. After each analysis, drug amounts quantified in PBMC samples were normalized by cell number and MCV in each single sample, as described by Simiele, according to the formula: [TAC] PBMC = TAC AMOUNT /(Cell N • × MCV) [21][22][23][24][25][26][27][28].…”

Section: Clinical Applicationmentioning

confidence: 99%

An UPLC–MS/MS method coupled with automated on-line SPE for quantification of tacrolimus in peripheral blood mononuclear cells

Pensi

Nicolò

Pinon

et al. 2015

Journal of Pharmaceutical and Biomedical Analysis

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Development and validation of a useful UPLC–MS/MS method for quantification of total and phosphorylated-ribavirin in peripheral blood mononuclear cells of HCV+ patients

Cited by 21 publications

References 47 publications

First UHPLC-MS/MS method coupled with automated online SPE for quantification both of tacrolimus and everolimus in peripheral blood mononuclear cells and its application on samples from co-treated pediatric patients.

First UHPLC-MS/MS method coupled with automated online SPE for quantification both of tacrolimus and everolimus in peripheral blood mononuclear cells and its application on samples from co-treated pediatric patients.

Development and validation of a UHPLC-MS/MS method for quantification of the prodrug remdesivir and its metabolite GS-441524: a tool for clinical pharmacokinetics of SARS-CoV-2/COVID-19 and Ebola virus disease

An UPLC–MS/MS method coupled with automated on-line SPE for quantification of tacrolimus in peripheral blood mononuclear cells

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