[Progress of drug transport study based on absolute quantitative method for membrane transporter proteins].
Knowing the amount of membrane transporter expression in human tissue is one of the key issues in the rational and reliable prediction of pharmacokinetic profiles in humans. Recently, we have developed a simultaneous and highly sensitive method for the absolute quantification of multiple membrane transporter proteins in mammalian tissues. To develop quantitative analysis of high molecular-weight membrane proteins, we have solved problems using proteomics technology as follows: 1) The target proteins are detected via tryptic peptides that can be dissolved and analyzed with LC-MS/MS, while membrane protein is difficult to dissolve. 2) LC-MS/MS in multiple reaction monitoring (MRM) mode produce a highly sensitive and selective response for transporter proteins with low expression by separation from highly abundant molecules. 3) Analyte specificity for each peptide was demonstrated in amino acid sequences using multiple MRM detection. Selection of the peptide probe was very important for highly sensitive analysis with LC-MS/MS. We set criteria for peptide selection using an informatics approach. Peptides without unstable residue, double basic residues, and integral membrane domain were selected as useful probes. The developed method will provide an inclusive assay platform for all transporter proteins expressed in both animal/human tissues and will contribute to progress in drug discovery and development.
Kamiie
Junichi
J
Graduate School of Pharmaceutical Sciences, Tohoku University, SORST.JST, Aoba, Aramaki, Aoba-ku, Sendai, Japan. kamiie@azabu-u.ac.jp
D000818:Animals; D002853:Chromatography, Liquid; D015195:Drug Design; D006801:Humans; D013058:Mass Spectrometry; D026901:Membrane Transport Proteins / Q000032:analysis*; D010599:Pharmacokinetics; D040901:Proteomics / Q000379:methods*
D026901:Membrane Transport Proteins