LC-MS/MS monitoring of the colorectal carcinoma cellular uptake and entrapment of sorafenib and its N-oxide active metabolite

Sorafenib (SOR) is a multikinase inhibitor with a mild activity against colorectal cancer cells due to multi-drug resistance mechanisms. Potentiated SOR activity was expected upon combination with some ginger derived compounds due to their interference with intracellular drug metabolism. Studying such combination necessitates the development of a sensitive validated LC-MS/MS method for the determination of intra and extracellular concentration of SOR and its N-oxide metabolite (SNX) in colorectal cancer cells. SOR, SNX and the internal standard (diclofenac sodium) were efficiently separated on Eclipse plus C18 column (3.0 x150 mm, 5 mu m) using isocratic elution with acetonitrile and 0.01 M ammonium formate aqueous solution containing 0.1% formic acid (69:31, v/v). Sample pretreatment using solid phase extraction was optimized and the mean percent recoveries were more than 97.01% for both analytes. Detection was conducted at positive ion multiple reaction monitoring (MRM) mode and the monitored mass transitions were 465.2 -> 252.2 for SOR and 481.1 -> 286.0 for SNX. The method was linear over the range 0.25 - 200.00 ng/mL (r(2) >= 0.9992) for SOR and 0.10 - 125.00 ng/mL (r(2) >= 0.9990) for SNX in both intra and extracellular matrices. The lower limits of quantification (LLOQ) were 0.25 and 0.10 ng/mL for SOR and SNX, respectively. Accuracies were within 94.25 - 109.45% and precision CV values did not exceed 7.63%. The method was able to monitor the cellular uptake and entrapment of both analytes and to prove the positive effect of the ginger derived compounds on SOR activity.

Automated summary

A sensitive LC-MS/MS method was developed to determine the concentration of SOR and its metabolite SNX in colorectal cancer cells. The method was able to monitor the cellular uptake and entrapment of both compounds, and confirmed the positive effect of ginger derived compounds on SOR activity.