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| Quantitative Membrane Proteomics and its Application in Translational Pharmacology |
| Yurong Lai* |
| Pfizer Global Research and Development, Groton, CT, USA |
| *Corresponding author: |
Yurong Lai
Department of Pharmacokinetics, Dynamics,
and Metabolism
Pfizer Global Research and Development
Pfizer Inc., Groton, CT
06340, USA
E-mail: yurong.lai@pfizer.com |
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| Received August 25, 2012; Accepted August 25, 2012; Published August 27,
2012 |
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| Citation: Lai Y (2012) Quantitative Membrane Proteomics and its Application
in Translational Pharmacology. J Proteomics Bioinform 5: vii-viii. doi:10.4172/jpb.10000e13 |
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| Copyright: © 2012 Lai Y, et al. This is an open-access article distributed under
the terms of the Creative Commons Attribution License, which permits unrestricted
use, distribution, and reproduction in any medium, provided the original author and
source are credited. |
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| The drug development process is expensive, time-consuming
and of high attrition. Often in spite of the efforts on improving drug
discovery ability and the excellent preclinical results, a significant
number of candidate drugs failed during clinical trial because of the
unfavorable efficacy and/or safety properties. To aim at improving
success rates in the crucial preclinical stage of development, scientists
in the pharmaceutical industry identify a number of fundamental
elements to guide decision making in drug discovery and development.
The elements are referred as three pillars of Phase II survival [1], which
include: 1) a drug is present at the target site of action. 2) a drug binds
to the pharmacological target. 3) pharmacological activity is expressed
associated with the shown target exposure and target binding. |
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| Membrane proteins including transporters, receptors and channels
are expressed on cellular membrane and play essential roles in the
transport of nutrients, ions and physiological compounds to sustain
cell survival. A subset of membrane proteins called drug transporters
also transport therapeutical xenobiotics across cellular barrier to
regulate the exposure on the site responsible for their effect and/
or toxicity. Accordingly membrane transporters become potential
pharmacological targets, biomarkers, carriers for drug delivery and
regulators of drug absorption, disposition, metabolism and elimination
(ADME) that may be involved in clinical drug-drug interactions and
adverse effects. |
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| Since the predictive information allows extrapolation to human
from in vitro or preclinical results, model based predictions, e.g.
physiologically-based pharmacokinetics (PBPK) model, become
emerging approaches used to predict complex drug disposition in a
way of holistic perspectives and are essential tools in the translational
research for the systemic and quantitative integration of diverse
preclinical information for the sake of rational drug design [2,3]. Rapid
advance in this regard requires analytical tools that can quantitatively
determine the components associated with biological processes
and measure the differences between two or more physiological
states of a biological system. Toward this end, technologies for
characterizing membrane proteins at different molecular levels such as
transcriptomics, proteomics, and metabolomics have been a growing
field, and proteomics in particular becomes a key enabling technology
and is continuing to evolve rapidly. While tissues selective expressions
at mRNA level are well-addressed in the literatures [4-6], quantitative
expressions of membrane proteins in human organs at protein level
are still missing. |
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| Liquid chromatography tandem mass spectrometry (LC-MS/
MS)-based proteomics address two shortcomings from classical
proteomics quantifications technologies---resolution of separation
provided by gels and identity of the underlying protein, and offer
considerable opportunities for biological understanding and
translational pharmacology in drug discovery and development. While
global proteomics can be used to indentify thousands of proteins in
cells or other biological samples, targeted proteomics quantifications
use peptides unique to the protein of interest, which can be readily
obtained from commercial sources and serve as surrogate standards to overcome the absence of protein standards, to assess quantitative
protein expressions in various biological matrices by way of a sensitive
and selective method that is amenable to high-throughput formats
[7]. However, despite of advances in protein analytical technologies
have been offering broad applications in drug discovery programs,
significant analytical challenges in quantitative membrane proteomics
remain, as membrane proteins are expressed at relatively low levels
and often comprise multiple hydrophobic domains that resist exposure
to aqueous environments leading to solubilization and denaturation
limitations with respect to facilitating protease access and digestion
efficiency. To overcome aforementioned hurdles, optimizations
tailored to a specific protein often include membrane solubilization
strategies, in which the organic solvents, detergents, and chaotropic
agents are examined to be compatible with the route of digestion and
subsequent MS analysis. The released proteolytic peptides are also
monitored over the course of the digestion to attain the optimized
digestion condition. The combination of proteases, example Lys-C and
trypsin, and the use of isotope labeled internal standards at different
level, e.g. stable isotope label by amino acid in cell culture (SILAC), are
commonly applied for addressing the incomplete digestion [8,9]. New
tools are continually being explored. For example, recently lipid-based
protein immobilization that offer immobilization and digestion of
bilayer-embedded native membrane proteins is used to rapidly probe
the solvent exposed domains in a flow cell format [10]. |
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| Quantitative membrane proteomics is now beginning to learn
for which type of study these methods can be meaningfully applied;
however, significant further improvements to experimental strategies
are required particularly for the quantitative analysis of posttranslational
modifications. Improvement in sample preparation is
equally important in order to differentiate the sub-cellular components
for investigating proteomes of intracellular membranes. As such,
significant technological advances and method optimizations that
affect quantification in bottom-up proteomic workflows for membrane
proteomics are further required to improve detection and accuracy
through addressing the sample handling, digestion efficiency, and
separation challenges. |
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| References |
| |
- Morgan P, Van Der Graaf PH, Arrowsmith J, Feltner DE, Drummond KS, et al. (2012) Can the flow of medicines be improved? Fundamental pharmacokinetic and pharmacological principles toward improving phase II survival. Drug Discov Today 17: 419-424.
- Atkinson AJ Jr, Smith BP (2012) Models of physiology and physiologically based models in clinical pharmacology. Clin Pharmacol Ther 92: 3-6.
- Smith BJ (2012) An industrial perspective on contemporary applications of PBPK models in drug discovery and development. Biopharm Drug Dispos 33: 53-54.
- Nishimura M, Naito S (2005) Tissue-specific mRNA expression profiles of human ATP-binding cassette and solute carrier transporter superfamilies. Drug Metab Pharmacokinet 20: 452-477.
- Nishimura M, Naito S (2008) Tissue-specific mRNA expression profiles of human solute carrier transporter superfamilies. Drug Metab Pharmacokinet 23: 22-44.
- Nishimura M, Suzuki S, Satoh T, Naito S (2009) Tissue-specific mRNA expression profiles of human solute carrier 35 transporters. Drug Metab Pharmacokinet 24: 91-99.
- Kamiie J, Ohtsuki S, Iwase R, Ohmine K, Katsukura Y, et al. (2008) 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 25: 1469-1483.
- Balogh LM, Kimoto E, Chupka J, Zhang H, Lai Y (2012) Membrane protein quantification by peptide-based mass spectrometry approaches: Studies on the organic anion-transporting polypeptide family. J Proteomics and Bioinformatics S4: 003.
- Wu CC, Yates JR 3rd (2003) The application of mass spectrometry to membrane proteomics. Nat Biotechnol 21: 262-267.
- Sui P, Miliotis T, Davidson M, Karlsson R, Karlsson A (2011) Membrane protein digestion - comparison of LPI hexalane with traditional techniques. Methods Mol Biol 753: 129-142.
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