Preclinical Testing Flashcards
Data from lead optimisation
Preliminary CMC (Chemistry, Manufacture and Control)
*Formulation
*Stability of active ingredient
*Detailed physicochemical characterization
*Impurity analysis
Benchmark in vivo models
*In vivo models
*Validated disease models
*Models in other disease areas
ADME Profiling
*Optimized analytical method development
*Pharmaco-kinetic(PK) profile
*Oral bioavalibility
*Determine metabolism of drug
Preliminary Toxicology (non-GLP)
*Maximum tolerated dose(MTD)
*Repeat Dose toxicity (non-GLP)
*Preliminary Cardiovascular Safety
GMP
– Good Manufacturing Practice: is a quality system covering the
manufacture and testing of active pharmaceutical ingredients,
diagnostics, foods, pharmaceutical products, and medical
devices. Guidelines that outline the aspects of production and
testing can impact on the quality of a product
GLP
– Good laboratory practice: a system of management controls for
laboratories and research organizations to ensure the
consistency and reliability of results
GCLP
– Good clinical laboratory practice: a framework for a quality
system in analysis of clinical trial samples, ensuring GCLP
compliance overall of processes and results
Data from initial preclinical stage
Detail Preliminary CMC
ICH Stability
*ICH Impurity Analysis
*Develop prototype clinical formulation (pill, liquid, etc.)
Comprehensive ADME
*GLP PK profile
*GLP toxicokinetics (TK) profile
*Comprehensive determination of metabolites
GLP Toxicology Package
*Acute Tox study
*Subchronic repeat dose study
*Genotoxicity Battery
*Safety Pharmacology Core Battery
Chemical development
Chemical Development
* Improvement of the synthesis
– To reduce cost,
– To increase output, safety and quality (purity and consistency)
* Chemical development is non-GMP
* Chemical manufacturing (supplies for clinical trials) must be GMP
Salts
Salt and Formulation
* Find best salt for
– Stability
– Ease of formulation
* Find best formulation for chosen route of administration
– Tablet, capsule, solution, controlled release, protection in GI tract, etc.
Optimized analytical method development
Optimized analytical method development
Crucial to demonstrate exposure levels in toxicology studies – human starting doses are based on this data!
*Small molecules - HPLC/MS
-Can definitively show the molecular structure
*Biologics (proteins, antibodies, etc.) - ELISA
-Does not show structure
-Uses binding as an endpoint
-Does not demonstrate activity
Assay Validation
Assay needs to be validated for use in GLP studies and be performed GLP
-Extraction technique recovery
-Linearity of standard curve
-Intra- and inter-assay precision
-Bench top and freeze/thaw stability
-Sensitivity (lower limit of quantitation; LOQ)
-Establish Quality Control (QC) standards
Bioavailability
Bioavailability and PK (BAPK)
* Bioanalytical method development and validation
– use to quantify drug and/or metabolites usually in plasma
* Bioavailability – single dose, iv and intended route
* PK – rodent and non-rodent, drug availability by intended route, mean residence time, half-life
* Blood-brain barrier bioavailability – measure drug accumulation in brain, brain vs plasma levels
Interspecies scaling improves human PK predictions = better clinical trial dosage design
Metabolic Profile
- Comparative metabolism
– Important when interspecies differences are observed
– Hepatic microsomes and cytosolic fractions from different species : human, mouse, rat, rabbit, dog, non-human primate, guinea pig, etc
– Time course of loss of parent compound
– Metabolite identification - Metabolic inhibition
– Potential drug-drug interaction
Toxicology Evaluation
*Appropriate species – one rodent, one second species (dog, pig or monkey generally)
-Good exposure
-Metabolism similar to human – must cover all human metabolites
-Same pharmacologic activity as humans (same target binding, effect in disease models, pharmacologic effects)
*Exposures achieved in test species should be sufficient to cover multiples of the intended human dose/exposure in order to establish a safety margin
*Higher doses to evaluate possible toxicities that could occur
-FDA guidance to dose up to 1 g/kg, if possible
*Administer compound long enough to support intended clinical study
*Example Endpoints: body weight, clinical observations, serum chemistry, hematology, organ weights, histology, drug exposure (toxicokinetics)
Single-Dose (Acute) Toxicity (Non-GLP)
Single-Dose (Acute) Toxicity (Non-GLP)
* Determination of adverse effects within short time frame of single dose administration
* Animals observed for 14 days after dosing
* Minimal endpoints focused on clinical observation and may be non-terminal (e.g. dogs)
* Identifies single dose MTD
Range-finding
– Longer schedule of repeated dosing
– Establish dosage levels for subsequent toxicity studies
Acute and Repeated-Dose Toxicity (GLP)
- Acute and repeated dose toxicity studies in 2 species (rodent and non-rodent) selected from non-GLP range-finding studies.
- More comprehensive: (greater number/gender/species
- More complete toxicology study.
- Determination of adverse effects resulting from daily dosing to identify MTD and NOAEL (No Observed Adverse Effect Level)
Clinical Endpoints
-Survival
-Body weight
-Clinical signs
-Behavior (Eating, movement)
Genotoxicity/Mutagenicity Testing (GLP)
- in vitro non-mammalian cell system – e.g. Ames Test – Salmonella typhimurium
- in vitro mammalian cell system – e.g. CHO (Chinese Hamster Ovarian) cells -> Determine % chromosomal aberration across a range of drug concentrations
- in vivo mammalian system –e.g. mouse micronucleus assay ->
– Immature mice treated with mutagen for period of 2-4 weeks
– RBCs observed under microscope for increased % of micronuclei
