Pharmacologically relevant measurement of high potency drug candidates with improved sensitivity in in-vitro transporter assays 

Using the SCIEX Triple Quad™ 7500 LC-MS/MS System - QTRAP® Ready

Brendon Kapinos1, Ian Moore2, Rolf Kern3
1
Pfizer, Groton, CT, USA; 2SCIEX, Canada, 3SCIEX, USA  

Abstract

The enhanced sensitivity of the SCIEX 7500 System will allow lower compound concentrations to be used in transporter assays, more closely align with the lower predicted systemic exposures for more potent drug candidates. Higher quality data will better guide compound rank ordering and selection in early stage drug development.

12617 7500

Introduction

In-vitro transporter assays are routinely run in mid-stage drug discovery to assess the impact of active transport activity on drug candidates’ ADME (absorption distribution metabolism excretion) behavior. Evaluation of transporter activity requires the analysis of many individual samples for a single compound, so to improve the efficiency of these assays, they are generally performed in a microtiter plate format. This dramatically improves sample handling but also serves to limit the sample size available for LC-MS/MS analysis of the collected samples. Because of the small available sample size and the need to be able to measure small percentages of the analyte at its initial concentration, high concentrations of analyte are typically tested. For potent compounds, these levels can be significantly higher than efficacious in-vivo concentrations, which makes prediction of in-vivo transporter activity from the in-vitro assays much more prone to error. In effect, the analytical technique can guide the assay concentration, as opposed to tailoring the assay around the more relevant predicted exposures.

Here, the use of the SCIEX Triple Quad 7500 LC-MS/MS System–QTRAP Ready is explored for the analysis of known compounds in OATP1B1 and OAT2 transporter assays, highlighting the sensitivity gains of this quantitative platform.

Figure 1. Valsartan chromatograms for wild type cell line. Comparing the QTRAP 6500+ System MRM signal (blue) with the SCIEX 7500 System MRM signal (pink) shows an approximately 10x signal improvement. 

Key features for transporter activity analysis

  • The combination of the OptiFlow® Pro Ion source, E Lens™ Technology and the D Jet™ Ion Guide on the SCIEX 7500 System provides increased sensitivity through the generation and transfer more ions into the mass spectrometer2
  • Enhanced sensitivity of the SCIEX 7500 System enables lower level analyte detection providing greater flexibility in experimental design
  • Ability to perform transporter assays with lower concentrations of analyte to better mimic biology
  • Reduced failures, reduced repeat injections, improved efficiency in early stage compound selection