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The Pharmacoanalytic Shared Resource (PhASR) supports pre-clinical and clinical drug development at Ohio State by providing high-quality and cost-effective bioanalytical method development, quantitative sample analysis and pharmacokinetic/pharmacodynamic experimental design, data analysis and modeling.
Analytical Method Development and Validation
PhASR staff offers expertise in developing and validating quantitative assays optimally suited for small molecules and peptides (LC-MS/MS), protein therapeutics (ELISA), and oligonucleotide therapeutics (ELISA and qRT-PCR). Proper analytical method development ensures appropriate understanding of factors that could influence results obtained with bioanalytical measurements or with measures of drug activity. For example, lack of understanding about compound solubility and stability may lead to incorrect conclusions from in vitro or in vivo pharmacokinetic or pharmacodynamic studies. Full validation of a newly developed method, based on the Guidance for Industry, Bioanalytical Method Validation published by the U.S. Food and Drug Administration, is completed for all work involving clinical trials, and it is recommended for all methods producing results intended for publication in non-clinical studies.
Accurate and sensitive quantification of drugs, metabolites and endogenous biomolecules requires developing and validating analytical methods optimized for precision and accuracy. For LC-MS methods, this involves appropriate selection of internal standards, solvents or solid-phase media for efficient sample extraction, LC mobile phases, analytical columns, liquid flow rates, gradients, ion polarity, daughter ions, gas flow rates and other general MS parameters. For some compounds, LC-MS may not be the most appropriate method of quantitation and an alternative detection method should be considered.
The outcome of the Analytical Method Development and Validation service is a validated method for accurate and precise quantitation of a specific compound within a given matrix (e.g. blood, plasma, tissue, urine, etc.).
Sample Analysis for General Pharmaceutics/Pharmacology Experimentation
Once an analytical method is developed, compound quantitation can be achieved. Using validated method procedures, PhASR processes and analyzes samples containing unknown concentrations of the compounds of interest. Samples may arise from biological matrices, such as plasma, urine or feces, or they may come from in vitro systems. The validity of results is ensured by the method validation process, processing quality control samples and standard samples—all processed and analyzed in parallel to the unknown samples.
Once a run is complete, PhASR reviews the data to validate the analytical run and ensure that quality control sample measurements fall within the allowed accuracy range and that the measured standard sample concentrations produce a linear range for quantitation.
PhASR provides expertise in metabolite and product identification. Knowledge of the metabolic pathways of a drug or drug class is critical in the drug development process, and mass spectrometry provides a means to specifically identify metabolites formed in small quantities in in vitro or in vivo systems.
Both LC and MS conditions are developed to separate the parent compound and metabolites adequately. For cases in which the metabolic pathways are known, specific metabolites may be anticipated, and the mass spectrometer can be tuned to look for these specific compounds. For cases in which the metabolic pathways are not known, searches for all possible metabolites can be conducted.
Experimental Design and Data Analysis
Experimental Design and Data Analysis offers users full access to PhASR’s expertise in characterizing relationships between drug pharmacokinetics and pharmacodynamics, including specialized needs for sample collection, specimen preparation and storage, data analysis and statistical modeling. The service provides clinical and pre-clinical research expertise in understanding the expected kinetics and fate of a drug and/or its metabolites in a biological system. Through proper design of a pharmacokinetic sa mpling strategy, representative biological specimens are obtained before, during and after drug administration. Samples are processed and stored appropriately to ensure they remain representative of the biological samples originally obtained.
For clinical studies, PhASR works closely with the Clinical Treatment Unit to advise on sampling and storage strategies that will provide the highest quality data while considering patient needs and clinical procedures.
After samples are analyzed, PhASR completes appropriate analysis to ensure the quality of the raw data. The outcome of this service is a carefully designed PK/PD study, which yields data and results that achieve the investigator’s project objectives
Pharmacokinetic/Pharmacodynamic Data Analysis and Modeling
After samples have been analyzed, PhASR can complete analyses of the data generated. Relevant PK/PD parameters and statistical considerations are defined during the experimental design process, and the data analysis follows the outlined plan.
PhASR staff is highly trained and experienced in PK/PD data analysis and modeling, which ranges from basic statistical comparisons to linear and non-linear, mixed-effects data modeling. The quantity and level of data analysis will differ for each project. Analyses could range from a simple comparison of drug concentrations between experimental and control samples sets to a more complex analysis requiring a population PK/PD model to be constructed, qualified and validated for predictive capabilities.
PhASR also offers additional services based on the methodological needs of OSUCCC – James members.
- Global DNA Methylation (GDM)
- Regional DNA Methylation (RDM)
- Protein/Peptide Quantification
- Quantitation of dNTP/NTP Pools
- Oligonucleotide Therapeutic Quantification using qRT-PCR or Hybridization-based ELISA
- Metabolite/Product Identification
- Drug Binding and Transport
Instrumentation and Equipment
The Pharmacoanalytical Shared Resource supports pre-clinical and clinical drug development at OSUCCC – James by providing high-quality, cost-effective and reliable bioanalytical method development, quantitative sample analysis and pharmacokinetic/pharmacodynamic experimental design and data analysis.
Liquid Chromatography/Mass Spectrometry (LC-MS) System Details: Primary PhASR equipment comprises three liquid chromatography/mass spectrometry (LC-MS) systems.
ThermoFinnigan TSQ Quantiva: a triple-quadrupole mass spectrometer with ultra-high-sensitivity coupled with a Dionex RSLCnano system (with ternary loading pumps, quaternary nano-flow pumps, degasser, temperature-controlled autosampler and column compartment). This system also incorporates an Easy-Spray nano-flow electrospray ionization source to achieve the maximal sensitivity possible for accurate quantification of small molecules, peptides or other biomolecules in complex biological matrices or in small sample quantities.
ThermoFinnigan TSQ Quantum Ultra: a triple-quadrupole mass spectrometer with extended mass range and heated electrospray ionization probe connected to a Dionex RSLCnano system (with ternary loading pumps, quaternary nano-flow pumps, degasser, temperature-controlled autosampler and column compartment).
ThermoFinnigan TSQ Quantum Discovery Max: a triple-quadrupole mass spectrometer with ESI and APCI ionization probes and a ThermoFisher Acella ultra-high-performance LC system (degasser, quaternary pumps, and photo-diode array detector), which includes an autosampler that is expandable for multiplexing, online extraction and high-throughput analysis.
Additional LC-MS systems (ion trap, triple-quadrupole and time-of-flight mass spectrometers) available for PhASR use are housed in PhASR or in Ohio State’s College of Pharmacy.
PhASR also maintains full access to other resources necessary to develop and run biological correlative assays, including UV-Vis and fluorescence spectrometers, plate-readers, thermal cyclers, centrifuges, cell culture and animal facilities, and freezer and refrigerator storage.
WinNonlin and NONMEM Pharmacokinetic Analysis Software Packages are also available for data analysis.
All PhASR fees are based on hourly rates for PhASR personnel time ($75/hour) or LC-MS instrumentation time ($95/hour).