Comparison of UPLC-MS/MS-based targeted quantitation and conventional quantitative methods for the analysis of MRP1...

Multidrug resistance (MDR), which is mainly caused by the overexpression of ATP-binding cassette (ABC) transporters, limits the effectiveness of clinical chemotherapy. Although multidrug resistance-associated protein 1 (MRP1) is an important ABC transporter, the relationship between MRP1 expression and drug-resistance has rarely been studied. The results of previous studies either cannot be compared or are contradictory, mainly due to limitations of conventional methods, such as semiquantification and low sensitivity. In addition, few reports have studied low resistance index (RI) levels, which are similar to clinical conditions. In this study, a novel UPLC-MS/MS method was developed and validated using the surrogate peptide (EDAQVDLFR) for the accurate and sensitive quantification of MRP1 expression in drug-resistance at low RI levels. The method was proven to be accurate (±20%, RE%), sensitive (0.09 pM, LLOQ), precise (±15%, RSD%), and stable under different conditions within 0.09-91.56 pM. In addition, the precision and stability of real samples further confirmed that the UPLC-MS/MS method was reliable. Quantification of MRP1 expression showed that there were significant differences (P < 0.05) between parental cell lines and drug-resistant cell lines, as well as between various drug-resistance cell lines, which could not be accurately monitored by qRT-PCR and Western blot. Therefore, the UPLC-MS/MS method is more accurate and sensitive when detecting slight changes in MRP1 expression at low RI levels and is an effective choice for related studies. Additionally, MRP1 expression was positively correlated with the RI level based on the Pearson correlation coefficient, which was not discussed in previous studies.