Evaluating whether a compound could make a good drug requires knowing its potency and selectivity, as well as how it is metabolized. The Drug Metabolism and Pharmacokinetics Core at UF Scripps has the expertise and equipment to perform such analysis and more.
In Vitro Services
Compound depletion in hepatic microsomes are commonly utilized to evaluate the innate stability of a compound with respect to hepatic metabolism. A variety of species are commonly evaluated. Stability in cytosol, S9, or hepatocytes are also available.
Plasma or Whole Blood Stability
Compound stability in plasma or whole blood is a significant concern for peptides and small molecule drugs containing ester or thioesters.
Plasma Protein Binding
Plasma protein binding influences the concentration of unbound compound available to diffuse into tissue. We typically evaluate plasma protein binding using equilibrium dialysis, but ultracentrifugation is also available.
Tissue non-specific binding
Total tissue concentration may not reflect cytosolic levels capable of interacting with your target. For lipophilic compounds the majority of the compound may be non-specifically bound to lipid. Tissue non-specific binding provides a scaling factor to estimate free drug in tissue.
Potent inhibition of cytochrome P450 is undesirable due to the potential to induce drug-drug interactions. We commonly evaluate the inhibition of common hepatic P450, specifically, CYP1A2, CYP2A6, CYP2B6, CYP2C8, CYP2C9, CYP2C19, CYP2D6, CYP2E1, and CYP3A4. Evaluation of time-dependent inhibition is an optional add-on. We also offer an economical cocktail inhibition assay where the inhibition of CYP1A2, CYP2C9, CYP2D6, and CYP3A4 is evaluated in the presence of 10 µM of your compound.
We use the PAMPA assay to evaluate the potential of a compound to cross a lipid membrane. Compounds are evaluated against high and low permeability reference compounds.
LogP and LogD
Calculated LogP and LogD values are based on structural similarity to a training set of compounds. With greater structural divergence, the calculated values incorporate additional error. LogD can be experimentally determined as user defined pH values using the octanol/water partitioning coefficient or with a higher throughput reverse phase HPLC method.
Kinetic solubility reflects most in vitro experiments where a stock solution prepared in an organic solvent is spiked into buffer and the solubility is determined at a predetermined time point. Equilibrium solubility mixes slid compound with buffer and determines soluble compound at a predetermined time point. Multiple pHs can be evaluated.
To design more stable compounds, it helps to know where your compound is metabolized. Studies can be customized to answer the specific needs of your individual project. Predicted metabolite structures are based on mass spectrometry experiments. Authentic standards may be required for definitive assignment.
In Vivo Services
Mouse or Rat Pharmacokinetic Study
PK studies quantitate plasma drug levels after intravenous, oral, intraperitoneal, or subcutaneous dosing. Study design usually uses three mice/rats and utilizes microsampling techniques to collect eight or more plasma samples per mouse/rat. Extended time points or multi-day designs are available.
Tissue Exposure Study
Tissue drug concentrations can be determined to verify tissue exposure and optimize dose levels or timing. Optional determination of free drug level is available using equilibrium dialysis.
We can match the conditions of your efficacy study to optimize the design of your experiment or to help interpret your results.
Discovery Toxicology/Tolerability Study
Single- or multi-dose studies are available. Behavior, general observation, body and organ weight, blood chemistry, and gross necropsy are available. Tissue can be collected and shipped to the sponsor or an external vendor if histology is desired.
LC-MS/MS quantitation of drug levels from user provided plasma, tissue, or tumor samples is available.
Frequently asked questions
1. How should we go about initiating a request for service?
Prior to sending any compounds, please send an email including your contact information, the compound name(s), and the specifics on which assays you are interested in. We will get back to you to work out the details.
Contact is Michael Cameron, PhD
Scientific Director, Sr. (561) 228-2223
2. Which species are available?
In vivo experiments are limited to mouse and rat. Most in vitro studies can be expanded to incorporate additional species.
3. What is the turnaround time?
Turnaround time varies with the experiment. In vitro studies are usually run when a sufficient number of compounds have been submitted for the assay. For example, if 20 compounds are submitted for mouse microsome stability, this request may have the data reported back within a week of compound delivery, but a single compound may take twice that long.
4. How much compound is required?
For in vitro studies, compounds can be submitted as a solid or in solution, preferably as a 10 mM DMSO stock solution. 30 µl is sufficient for most in vitro work. In vivo studies require solid compound. The amount differs with the species and dose. The following formula will help you predict the amount of compound required.
Compound (mg required) = 10 + 1.25 * (n * D * # * BW * SFC)
n=replicates, D = dose in mg/kg, # = number of doses, BW = body weight (use 0.025 for mouse and 0.45 for rat), SFC = salt form correction.
Example 1. A single rat PK study IV dosed at 1 mg/kg with n=3 rats. Compound is provided as a free base with a molecular mass of 400. Submitted compound should be:
10 + 1.25 * (3 * 1 * 1 * 0.45 * 400/400) = 12 mg
Example 2. A single rat PK study orally dosed at 30 mg/kg with n=3 rats. Compound has a molecular mass of 400 and TFA salt is submitted. This would require:
10 + 1.25 * (3 * 30 * 1 * 0.45 * 514/400) = 75 mg
When sending compound, make sure to include your contact information inside the package.
5. What other information do I need to provide?
We need the exact mass (molecular weight) of each compound. If the compound is a solid, we also need the batch molecular weight that accounts for the salt form. If there are numerous compounds, an electronic copy is preferred.
6. Will I get a final report?
Studies come with a datasheet and a brief description of the results. If you require a final report, an additional report fee will be charged.
7. Can we get a quote?
We do not require a formal quote for the initiation of studies for internal investigators, but can provide a quote if you desire one. Most outside institutions require a formal quote or an informal e-mail stating the service and cost.
8. How should we handle billing?
If a PO has been established prior to service, we will handle the invoicing with your purchasing department. If a PO has not been arranged, an invoice will be sent to you after the completion of the work. Prepayment of a portion of the study may be required if the materials cost of the study are high.
9. Can you help with grant proposals?
We have extensive experience working on NIH funded projects. We can provide descriptions of assays and compound progression plans for grant proposals and review the metabolism section of your proposal. Depending on the level of involvement we have served as a fee-for-service core, co-investigator, or co-PI.