Preanalytical Stability of Gemcitabine and its Metabolite 2′, 2′-Difluoro-2′-Deoxyuridine in Whole Blood—Assessed by Liquid Chromatography Tandem Mass Spectrometry

Tormod Bjånes,Tina Kamčeva,Torunn Eide,Bettina Riedel,Jan Schjøtt,Asbjørn Svardal

doi:10.1002/jps.24638

Abstract

Gemcitabine (2',2'-difluoro-2'-deoxycytidine, dFdC) and metabolite (2',2'-difluoro-2'-deoxyuridine, dFdU) quantification is warranted for individualized treatment strategies. Analyte stability is crucial for the validity of such quantification. We therefore studied the impact of the time interval from blood sampling to separation of plasma on gemcitabine stability. Blood from gemcitabine-treated patients was drawn into tetrahydrouridine (THU)-spiked heparin and ethylenediaminetetraacetic acid tubes and kept on ice until separation. Plasma was separated sequentially up to 24 h after sampling and dFdC and dFdU were quantified by liquid chromatography tandem mass spectrometry (LC-MS/MS). The change in plasma concentrations over time was compared with the highest imprecision for concentrations above the lower limit of quantification of the LC-MS/MS method. Analyte concentrations decreased slightly over time, but for samples stored for 4 h on ice, the decline was smaller than the expected analytical imprecision. After 24 h, the maximum decline was 14.0%, which exceeded the expected analytical imprecision. dFdC and dFdU stabilities were acceptable for at least 4 h when THU-spiked whole blood samples were kept on ice. This is within the scope of routine sampling procedures. Further, variations in separation time intervals within this time frame are negligible when interpreting drug concentrations.

Highlights

The following three main sections cover background topics related to the current research field, focused on pancreatic cancer and chemotherapeutic drugdelivery by sonoporation, with special emphasis on the nucleoside analogue gemcitabine and quantitative aspects of cellular uptake and metabolism. ௺௾1.1
In paper IV, we showed that intracellular cytidine deaminase (CDA) could inactivate gemcitabine extensively and hereby regulate intracellular difluoro-2′-deoxycytidine triphosphate (dFdCTP) accumulation
In paper V, we demonstrated that sonoporation contributed to only a minor extent of gemcitabine uptake compared to physiological membrane transporters

Summary

Introduction

The following three main sections cover background topics related to the current research field, focused on pancreatic cancer and chemotherapeutic drugdelivery by sonoporation, with special emphasis on the nucleoside analogue gemcitabine and quantitative aspects of cellular uptake and metabolism. ௺௾1.1. Gemcitabine (2′,2′-difluoro-2′-deoxycytidine, dFdC) is a nucleoside analog used in chemotherapeutic drug regimens against several human cancers, including pancreatic ductal adenocarcinoma (PDAC).. In vitro deamination of dFdC to dFdU by cytidine deaminase (CDA) is a major issue and can be inhibited by tetrahydrouridine (THU).11 Assessment of both analytes and of their ratio contributes as a control of the sample quality in this setting. Gemcitabine (2’,2’-difluoro-2’-deoxycytidine, dFdC) is a nucleoside analogue used either alone or in combination with other cytostatic agents for treatment of inoperable pancreatic ductal adenocarcinoma (PDAC), and several other human cancers (Norwegian Medicines Agency, 2018). Conclusions: It is possible to combine ultrasound, microbubbles, and chemotherapy in a clinical setting using commercially available equipment with no additional toxicities This combined treatment may improve the clinical efficacy of gemcitabine, prolong the quality of life, and extend survival in patients with pancreatic ductal adenocarcinoma

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