16 Safety Testing New Medicines Flashcards
ICH approved non-clinical
safety testing
• ICH: International conference on harmonization (US, EU, Japan)
• Toxicokinetics
– PK studies associated with toxicology studies
• Doses
– Single dose - “acute toxicity”
– Repeat dose (between 2 weeks and 6-9 months) –“chronic toxicity”
– NOEL & NOAEL (No Observed Effect Level; No Observed Adverse Effect Level)
• Studies to assess:
– Safety Pharmacology
– Reproductive toxicology
– Genotoxicity (Mutagenicity)
– Carcinogenicity
– Local tolerance (for route of administration)
– Any special considerations relating to the drug
Toxicokinetics
The study of how a chemical or toxin gets into the body and what the body does to the chemical
By obtaining toxicokinetic data we can:
- Understand relationship between dose and systemic exposure
- Understand relevance to human exposure at high dose
- Relate observed toxicity to exposure (including tissue distribution)
- Show the toxicity measured in animals is a reasonable model of what might happen in clinical trials
- Level of exposure relevant
- Metabolites relevant
- Pharmacology of toxicology
Single-dose toxicity studies
- Two mammalian species (often rodent)
- Two routes of administration
- Route to be used in man
- Route giving F=1 (eg i.v.)
- Measure exposure
- Range of drug doses to observe Therapeutic exposure & NOEL → NOAEL →LD (Lethal Dose)
- Observe for ≥14 days for evidence of toxicity (eg body weight, behaviour)
- Autopsy: organs with macroscopic abnormality undergo histopathological examination
Repeat-dose toxicity studies
• Rodent (mouse, rat) and non-rodent (dog, monkey)
• Exposure (dose/PK, duration, route) to reflect expected clinical use
– 28 day (Normally non-regulatory – justifies investment in longer studies)
– 6-12 months carried out to GLP (For very short clinical trials may be shorter)
• Toxicokinetics
• Monitor
– Weight, behaviour, hematology, urine analysis, food consumption, cardiovascular (rate, bp, ECG)
– Organ weights, gross pathology, histology
• Mutagenicity
• Indication of reproductive toxicology (fertility, pre & post
natal development)
• Indication of carcinogenicity
Repeat-dose drug distribution
studies
• Single-dose drug distribution studies performed routinely as part of toxicokinetics package
• Repeated dose distribution studies performed when
– Plasma levels higher after repeated dose than single dose
– Slow elimination from tissues seen in single dose studies
– Unexpected histopathological (tissues become diseased) effect observed compared to results from single dose distribution study
GLP
Good Laboratory Practice
• 1970s - FDA investigation identified poor laboratory practice in US toxicology labs
-Examples of some of these poor
lab practices found were
1. Equipment not been calibrated to standard form , therefore giving wrong measurements.
2. Incorrect/inaccurate accounts of the actual lab study
3. Inadequate test systems
-Industrial Bio Test – excluded mice suffering severe adverse events from safety-testing studies for cosmetics
Safety Pharmacology
• Investigation of undesirable pharmaco-dynamic effects (eg receptor antagonism)
– Test: drug and major metabolite
• In particular look for effects on – cardiovascular system,
– respiration,
– CNS.
• Supplementary tests
– Renal/urinary system; autonomic nervous system; GI function
• Also any effects anticipated from:
– Target class; Drug class; Target physiological function
• Use: animals, organs, tissue, cells in vitro, cell fragments, enzyme assay, receptor binding assays
– Relatively few models have regulatory recognition
– Used by Pharma companies for internal decision-making
– Some may be incorporated into make-test drug discovery cycle
Reproductive toxicity
• To asses:
– Fertility (♀ & ♂),Embryonic, fetal and post-natal development
• Species
– Same as for other toxicity studies (often rat)
– Teratogenicity – include second non rodent species (often rabbit)
• Study for full life cycle:
– Pre-mating→Conception→implantation →embryo →fetus →birth →weaning →sexual maturity
• Phase 1&2 trials can be carried out without Reprotox studies
– Only in Men & Women Not of Childbearing Potential
Preclinical tests for Genotoxicity
In vitro • Ames bacterial reverse mutation • Mammalian cells – Metaphase chromosome aberration – Micronucleus – Mouse Lymphoma TK assay
In vivo
• Can be part of repeated dose study
• Chromosome aberration assay or micronucleus assay
What ames test
The Ames test is a widely employed method that uses bacteria to test whether a given chemical can cause mutations in the DNA of the test organism.
Ames test for mutation
- Salmonella strain requires Histidine for growth on agar due to DNA mutation
- Test for reversal of mutation – point mutation and frameshift
- Optional: Liver homogenate to generate & test drug metabolites – biotransformation
Tests for chromosomal aberration & aneuploidy
Cells treated with drug (in vitro or in vivo) –>
Giemsa Stain
-Metaphase chromosome aberration assay • Light microscopy • Metaphase cells • Count chromosomes • Morphological assessment
-Micronucleus Assay
•Light microscopy •Chromosome fragment not incorporated into daughter cell’s nucleus
-Mouse Lymphoma TK assay
•If drug induces mutation in Thymidine kinase gene– cells survive
•Detects broad spectrum of DNA alterations – anything that interferes with function of TK
•Point mutations, frameshifts, translocations, deletions, aneuploidy
Define aneuploidy
presence of an abnormal number of chromosomes in a cell
Carcinogenicity
• Most human carcinogens are also carcinogens in animals
• Study design:
– 2 year or animal lifetime study
– Drug dose MTD & 0.5 x MTD (maximum tolerated dose) – Rat or mouse
– Monitor appearance of tumours
• Only performed if
– Drug will be used continually > 6 months (eg anti depresants) – OR if there is cause for concern
• Compound class/SAR
• Evidence of carcinogenicity from repeat dose study • Slow elimination
• Genotoxic – presumed to be carcinogenic
• Sometimes delayed until/after MAA for serious disease and not required if life expectancy short – i.e may not be done before First in Human (FIH) trial!
