A new, simple method for quantifying gemcitabine triphosphate in cancer cells using isocratic high-performance liquid...

Area of Science:

• Analytical Chemistry
• Cancer Pharmacology
• Biochemistry

Background:

• Gemcitabine (dFdC) is a vital chemotherapy drug for various cancers.
• Intracellular gemcitabine triphosphate (dFdCTP) concentration is key to dFdC's effectiveness.
• Monitoring dFdCTP levels can optimize dFdC treatment strategies.

Purpose of the Study:

• To develop a novel, sensitive, and simple method for quantifying dFdCTP in cancer cells.
• To establish an isocratic-elution high-performance liquid chromatography (HPLC) assay for dFdCTP measurement.
• To assess the method's applicability in biological samples and its potential for therapeutic drug monitoring.

Main Methods:

• Development of an isocratic-elution HPLC method using an anion-exchange column.
• Quantification of dFdCTP in cultured and primary leukemic cells.
• Validation of the method for sensitivity, linearity, and variability.
• Simultaneous determination of dFdCTP and cytarabine triphosphate (ara-CTP).

Main Results:

• The developed HPLC method demonstrated high sensitivity (20 pmol lower detection limit) and low variability.
• dFdCTP was effectively separated from other nucleoside triphosphates.
• The assay successfully measured dFdCTP in various cancer cell types.
• The method confirmed increased ara-CTP production following dFdC treatment.

Conclusions:

• The new isocratic HPLC method provides a sensitive and simple approach for quantifying dFdCTP in biological samples.
• This assay is valuable for research and potentially for therapeutic drug monitoring in cancer patients.
• The method's applicability extends to studying drug interactions, such as dFdC's effect on ara-CTP levels.