ReviewQuantitative Targeted Absolute Proteomics-Based Adme Research as A New Path to Drug Discovery and Development: Methodology, Advantages, Strategy, and Prospects
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INTRODCUTION
Transporters, enzymes, receptors, and channels are well known to play key roles in drug absorption, distribution, metabolism, elimination, and toxicity (ADMET) and therefore have an important influence on drug efficacy. Both pharmacogenomics and pharmacogenetics approaches have been used in drug discovery and development to elucidate the functions of these proteins in ADMET research,1,2 as well as to establish the extent of interindividual variability. Recent advances in mass spectrometry (MS)
Protein Quantification by MRM Using LC–MS/MS
Quantification by LC–MS/MS has been performed for small molecular compounds, such as drugs, and it was considered that a similar quantification strategy might be available for proteins. However, proteins are too large to separate by reversed-phase high-performance liquid chromatography (HPLC) and their mass lies above the range of the mass filter in MS for quantification [mass-to-charge ratio (m/z) <2800 in 4000QTRAP, <1250 in API5000 (ABSCIEX, Foster City, CA)]. Therefore, proteins must first
Sequence-based Target Peptide Selection for Multiplexed-MRM Analysis
The most important issue for high sensitivity and reliability in protein quantification by MRM analysis is the selection of the target peptide. The target peptide should have a unique amino acid sequence and give a high intensity in MS analysis. One selection strategy has been to identify the target peptide by global proteomics. Picotti et al.5 have developed a database of peptides based on previous proteomics experiments, called the MRM Atlas, and they reported an analysis of 1500 yeast
Comparison of LC–MS/MS-Based Quantification with Antibody-based Quantification
The principle of protein quantification using LC–MS/MS is to quantify specific peptides generated from target proteins by trypsin digestion. As MS selects the peptide based on molecular weight (m/z) with a mass filter, a major advantage of this quantification method is high selectivity, in contrast to the issue of cross-reactivity in antibody-based methods. A single amino acid difference in a peptide changes the m/z, and is able to be distinguished by MS (Table 1). An example of the superiority
Quantification of Membrane Transporter Protein
Membrane proteins are difficult to analyze quantitatively by western blotting or ELISA due to their poor solubility and high aggregability, although membrane proteins include important drug targets and play important roles in ADMET. By applying multiplexed-MRM analysis, we have quantified 43 transporters and Na+/K+ ATPase in mouse; 47 transporters, three junction proteins, two receptors, and Na+/K+ ATPase in monkey; and 106 transporters, 14 membrane proteins, and Na+/K+ ATPase in human
Reconstruction of in Vivo Transporter Protein Function
Variability, such as species difference, in vitro/in vivo difference, age difference, and normal/disease difference, in physiological function must be taken into account in drug discovery and development, as well as clinical application. Drug concentrations in the tissue and plasma, pharmacological and/or toxicological responses, and the pathophysiological state of the drug target organ depend upon the activities of functional proteins such as transporters, enzymes, channels, receptors, and so
QTAP-based Drug Discovery and Development
Great efforts have been directed to drug discovery and development in both the pharmaceutical industry and academia in the past decades. There have been remarkable developments in the methodology of life science, including whole-genome sequencing, genome-wide association study, and global proteomics, which have facilitated the discovery of disease-related proteins, but we are still far away from a “gold-rush era” of new drugs for these target proteins. The processes of drug ADMET/efficacy are
CONFLICT OF INTEREST
Tetsuya Terasaki and Sumio Ohtsuki are a full professor and an associate professor of Tohoku University (Sendai, Japan), respectively, and are also directors of Proteomedix Frontiers. This research was not supported by Proteomedix Frontiers and their positions at Proteomedix Frontiers do not present any financial conflicts. The other authors declared no conflict of interest.
Acknowledgements
The studies mentioned in this review were supported in part by a Grant for Development of Creative Technology Seeds Supporting Program for Creating University Ventures from Japan Science and Technology Agency (JST), and the Industrial Technology Research Grant Program from New Energy and the Industrial Technology Development Organization of Japan, as well as a Grant-in-Aid for Scientific Research (S) 18109002 from the JSPS, a Grant-in-Aid for Scientific Research on Priority Area 17081002 from
REFERENCES (46)
- et al.
Full dynamic range proteome analysis of S. cerevisiae by targeted proteomics
Cell
(2009) - et al.
Simultaneous absolute quantification of 11 cytochrome P450 isoforms in human liver microsomes by liquid chromatography tandem mass spectrometry with in silico target peptide selection
J Pharm Sci
(2011) - et al.
Quantitative, multiplexed assays for low abundance proteins in plasma by targeted mass spectrometry and stable isotope dilution
Mol Cell Proteomics
(2007) - et al.
High sensitivity detection of plasma proteins by multiple reaction monitoring of N-glycosides
Mol Cell Proteomics
(2007) - et al.
MALDI imaging mass spectrometry–Painting molecular pictures
Mol Oncol
(2010) - et al.
The 4F2hc/LAT1 complex transports L-DOPA across the blood–brain barrier
Brain Res
(2000) - et al.
Immunohistochemistry with an antibody to human liver carboxylesterase in human brain tissues
Brain Res
(1994) - et al.
Reduced brain 5-HT and elevated NE turnover and metabolites in bipolar affective disorder
Biol Psychiatry
(1994) - et al.
Effects of ketamine on dopamine metabolism during anesthesia in discrete brain regions in mice: Comparison with the effects during the recovery and subanesthetic phases
Brain Res
(1997) - et al.
New approaches to in vitro models of blood–brain barrier drug transport
Drug Discov Today
(2003)
Near completely humanized liver in mice shows human-type metabolic responses to drugs
Am J Pathol
A highly stable and nonintegrated human artificial chromosome (HAC) containing the 2.4 Mb entire human dystrophin gene
Mol Ther
Label-free quantitative proteomics using large peptide data sets generated by nanoflow liquid chromatography and mass spectrometry
Mol Cell Proteomics
Transporter-enzyme interactions: Implications for predicting drug–drug interactions from in vitro data
Curr Drug Metab
The role of transporters in the pharmacokinetics of orally administered drugs
Pharm Res
Analyzing antibody specificity with whole proteome microarrays
Nat Biotechnol
A preliminary study of serum concentrations of soluble epidermal growth factor receptor (sErbB1), gonadotropins, and steroid hormones in healthy men and women
Cancer Epidemiol Biomarkers Prev
Quantitative atlas of membrane transporter proteins: Development and application of a highly sensitive simultaneous LC/MS/MS method combined with novel in-silico peptide selection criteria
Pharm Res
A synthetic protein approach toward accurate mass spectrometric quantification of component stoichiometry of multiprotein complexes
J Proteome Res
Quantitative membrane protein expression at the blood-brain barrier of adult and younger cynomolgus monkeys
J Pharm Sci
Human platelets express organic anion-transporting peptide 2B1, an uptake transporter for atorvastatin
Drug Metab Dispos
Expression of ABC-type transport proteins in human platelets
Pharmacogenet Genomics
Quantitative targeted absolute proteomics of human blood–brain barrier transporters and receptors
J Neurochem
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