SWATI SARIN

Benefit –risk ratio can be calculated

Prediction of therapeutic index

Therapeutic index= Maximum tolerated dose Minimum curative dose

Smaller ratio, better safety of the drug

Pharmacological effects are same in man as in animals

Toxic effect in species will predict adverse effects in man

Giving high doses in animals improves predictability to man

Risk assessment can be made by comparison of toxic doses in test species with predicted therapeutic dose in man

PHASES OF DRUG DEVELOPMENT

(ANIMAL MAN)

PHASE III PHASE IVPHASE IPHASE IPHASE IPRECLINICALPRECLINICAL PHASE II

Product Approval (NDA/MAA)

Patient studies

Entry to man(IND / CTA)

NoneNone

Healthy subjects

Safety andtolerability

Healthy subjects

Safety andtolerability

Genetic toxicity(in vivo)

Repeat dose toxicity testing

+Bioanalysis /

Toxicokinetics

Drug Metabolism

Reproductive Toxicity Testing(teratogenicity)

Genetic toxicity(in vivo)

Repeat dose toxicity testing

+Bioanalysis /

Toxicokinetics

Drug Metabolism

Reproductive Toxicity Testing(teratogenicity)

Patients

Small scale efficacy studies

Patients

Small scale efficacy studies

Patients

Large scalemulticentre

studies

Patients

Large scalemulticentre

studies

Chronic (long term) toxicity testing+

Bioanalysis / Toxicokinetics

Reproductive Toxicity Testing (fertility and pre/post natal)

Carcinogenicity studies

Drug Metabolism

Chronic (long term) toxicity testing+

Bioanalysis / Toxicokinetics

Reproductive Toxicity Testing (fertility and pre/post natal)

Carcinogenicity studies

Drug Metabolism

Patients

Large scalepost-marketing

studies

Patients

Large scalepost-marketing

studies

As requiredAs required

Genetic toxicity(in vitro)

Single / repeat dose

toxicity studies+

Bioanalysis / Toxicokinetics

Safety Pharmacology

Drug Metabolism

Lead candidateIdentified

Clin

ical

Non

-clin

ical

MOLECULE

Studies should comply with GLPPerformed by trained and qualified staffUse of standardized and calibrated equipmentSOP’s followed in laboratory tasksAll documents should be preserved for minimum 5 years after marketing of the drug

TOXICOKINETIC STUDIES

Generation of Pharmacokinetic data to access systemic exposure achieved in animals

Relation to dose level and the time course of toxicity study

To support choice of species & Treatment regimen

Design on clinical studies accordingly

Pharmacodynamic responses

Pharmacokinetic profile Species, sex, age of experimental animals

Susceptibility, sensitivity and reproducibility of test system

In vitro: Isolated organs, tissues cell-cultures

Mechanism of effect in vivo

Systemic toxicology studies

Single dose studies Repeated dose studies

Reproductive toxicology studies

Male fertility Female reproduction & Developmental studiesLocal toxicity studies

Hypersensitivity studies

Genotoxicity studies

Carcinogenicity studies

Preliminary Definitive

• Maximum Non Lethal dose(MNLD) determined

• MTD and MLD determined• Evaluate effects • Target organ of toxicity may be determined

a) SINGLE DOSE STUDIES/ ACUTE TOXICITY

METHOD

Single dose tested in 2 rodent species 2 routes of administration Oral dosing of 2g/kg or 10 times of normal

human dose Observation for 14 days after dosing MNLD established Symptoms , signs reported Microscopic and Macroscopic evaluation

METHOD

Group of 20 animals of either sex dosed at MNLD 5 animals of each sex are observed for 48 hr and

conduct autopsy for early pathological changes Remaining 5 of each sex are observed for 14 days MTD and MLD established Signs of intoxication or recovery, changes in body

weight, pathological changes Complete macroscopic and microscopic examination Target organs can be identified

Two mammalian species(one should be non-rodent)

Long duration studies (30-180 days) Dose is dependent on dose-escalating studies Drug administered by clinical route Parameters monitored and recorded are:

Behavioral Physiological Biochemical Microscopic observations

b) REPEATED DOSE STUDIES/SUB-ACUTE OR CHRONIC TOXICITY

a) MALE FERTILITY

METHODOne rodent species(rat)

3 dose groups taken (each with 6 adult males),

1 control

Drug treatment by clinical route for 28-72 days

Mated with females in 1:2 ratio

Females getting pregnant should be examined

After 13 days of gestation

All male animals sacrificed

•Weights of testis, epididymus recorded & examined for their histology

•Sperms examined for motility & morphology

Segment I

15

Fertility and general reproductive performance study

Segment II Teratogenicity

Segment III Peri and post-natal study

Fertility and early embryonic development (rat) Embryo- foetal development (rat & rabbit)

Post natal development (rat) (post natal survival of offspring), growth parameters, vital senses, behavioral effects

b) FEMALE FETILITY

Drug administered to both males (28days) and females (14 days) before mating

Implantation Embryogenesis

Required when drug is administered by special route (other than oral) in humans

Study design: 2 species along with control used Dose dependent on dose escalating studies 3 dose levels

Dermal toxicity studies

Dermal photo-toxicity studies

Vaginal toxicity studies

Rectal tolerance studies

Rats & RabbitLocal signs (erythema, oedema), histological examination

Guinea pigUsed in treatment of leucodermaExamination of erythema & oedema formation

Rabbit or DogObservation of swelling, histopathology of vaginal wall

Rabbit or DogSigns of pain, blood or mucous, histology examination of rectal mucosa

Ocular toxicity studies

Parenteral drugs

Inhalation toxicity studies

Albino RabbitChanges in cornea ,Iris & aqueous humor, histological examination of eye

For intravenous/ intramuscular/ subcutaneous/ intra-dermal injectionSites of injection examined grossly and microscopically

One rodent and non rodent speciesAcute , sub-acute and chronic studies performedObservation of respiratory rateHistological examination of respiratory passages, lung tissue

Guinea Pig Maximization test

Local lymph node assay

Determination of Maximum non irritant or minimum irritant doseEvaluation of Erythema and oedema

Mice of one sex(either male or female)Drug treatment given on ear skinAuricular lymph node dissection after 5 daysIncrease in 3h-thymidine used for evaluation

To detect early tumorigenic effects in cases of chronic illness

In vitro tests:Test for gene mutation in BacteriaCytogenetic evaluation of chromosomal damage in mammalian cells

E.g.; Ames’s Salmonella Assay detects increased number of aberrations in metaphase chromosomesDNA strand breaks, DNA repair or recombination, Measurements of DNA adducts

In vivo tests:Chromosome damage in rodent hematopoietic cells

E.g.; Micronucleus Assay

Life-time Bioassays

Carcinogenicity studies are performed on:

Drug used for >6 months or frequent intermittent use for chronic diseases

Chemical structure of drug indicates carcinogenic potential

Therapeutic class of drugs which have produced positive carcinogenicity

Group sizes of 50 animals/sex at each of 3 dose levels

Control group is of double size

Record for onset of tumor development

Usually carried out for 24 months in rats and 18 months in mice (life span studies)

CONDUCT OF STUDY

EVALUATION OF RESULT

Incidence of cancers in control and test

Trend towards increasing incidence with increasing doses

Number of animals with single/multiple tumors

Macroscopic changes observed by autopsy

Histopathology of organs and tissues