Toxicity Testing

Question 1

toxicity is an extension of a drug’s pharmacological action and therefore all toxic effects are predictable

true or false

Answer 1

true

Question 2

Acute toxicity of a drug occurs after

Answer 2

a single dose

Question 3

subacute or subchronic tox is after exposure to the drug for

Answer 3

days or weeks

Question 4

chronic toxicity of drugs is observed after exposure for

Answer 4

months to years

Question 5

what are the goals (5) of non-clinical safety evaluations?

Answer 5

1. characterization of toxic effects with respect to target organs
2. dose dependence of toxic effects, i.e. dose response relationship
3. relationship to exposure
4. reversal potential
5. dose determination for human clinical trials

Question 6

animal tox studies can be seen as risk reduction measures for clinical trials

True/false

Answer 6

True

Question 7

How are tox studies performed, 6 key attributes of a well designed study.

Answer 7

1. according to a protocol
2. use toxic doses
3. record functional, structural or biochemical abnormalities compared with a control group
4. determine the No Adverse Effect Level
5. determine Maximum Tolerated Dose
6. Evaluate safety margin over the therapeutic dose range in man

Question 8

what are the four types of tox studies often performed?

Answer 8

1. Acute Tox - single dose
2. Short term chronic - up to 3 months (Repeat Dose)
3. Long term chronic - 1-2 years (Repeat Dose)
4. Special: Mutagenicity, teratogenicity, carcinogenicity

Question 9

what is a key pharmaceutical consideration for conducting tox studies?

Answer 9

you want your molecule or test article to be representative in terms of purity and formulation to that used in man or long term tox studies to ensure you get consistent and good results

Question 10

what other factors about species choice and dosage schedule are important to understand when designing tox studies?

Answer 10

• species have different metabolic rates and toxic effects
• compare metabolic profile in chosen species with man
• justify choice
• understand the plasma concentration of drug in the animal model and compare with man

Question 11

When conducting acute tox studies the following must be performed prior to starting the study?

Answer 11

dose ranging study to determine doses that will demonstrate toxic effects

Question 12

acute tox studies (single dose studies) should include which routes of administration

Answer 12

parenteral, that planned to be used in man, and any other appropriate routes

Question 13

acute tox studies must include: (4 things)

Answer 13

1. 2 species
2. have equal male to female ratio
3. include observation of physical and behavioural changes
4. performed for 7 days

Question 14

Lethal dose studies determine

Answer 14

the LD50, or the dose at which 50% of the population dies

Question 15

what are some criticisms of the LD50 Methodology (7 things)

Answer 15

1. kill animals unnecessarily
2. require a large number of animals to complete
3. can achieve the same if not better results with less animals using geometric methods
4. varies between species and within strains of the same species
5. values can differ by 500x or more with age
6. nutritional status influences results
7. environmental factors such as acclimatization, light, humidity, temperature, noise, socialization can infuence results

Question 16

Outline requirements for each of the following for Chronic toxicity studies (Repeat Dose):

• Duration
• route of admin
• Dose
• Species
• Observation
• QC

Answer 16

• Duration
  
  • 3 months - 1 year
  • ICHM3 6 months rodent, 9 months non-rodent for exposure > 6 months
• route of admin
  
  • that used in man
  • for inhalation or topical include parenteral
• Dose
  
  • use dose that gives toxic effects but not lethal dose
  • 5x the human dose
  • a dose in between
• Species
  
  • 2 species, equal male/female
  • metabolism resembles man
  • usually rats or dogs
• Observation
  
  • body weight, food/water intake, behavioural
  • biological such as urinalysis, CBC
• QC
  
  • done under GLP
  • test facility management
  • procedures
  • study director
  • quality assurance and quality control
  • raw data available, reviewed and retained

Question 17

Teratogens are

Answer 17

compounds which impair fertility and pre or post natal development

Question 18

teratogens have their highest effect during blastula stage or organogenesis stage

Answer 18

organogenesis

Question 19

the impact of teratogens from conception to blastula stage is

Answer 19

is either lethal or not lethal

Question 20

When are repro tox studies required

Answer 20

Reproduction development tox studies are required for any drug indicated for women of child bearing

Question 21

this type of repro tox study involves the exposure of germ cells of males and females to mutagenic agents that may impair fertility

Answer 21

Fertility study

Question 22

Fertility studies require

Answer 22

• pretreatment of animals prior to mating
• males 70 days
• females 14 days

Question 23

the teratogenic effect of fertility studies is determined based upon

Answer 23

the number of non-viable implants and # surviving fetuses

Question 24

describe elements of an effect on organogenesis repro tox study

Answer 24

• females exposed d6-16
• some sacrificed during exposure and observed for resorption and dead fetuses
  
  • fetus are studied for abnormalities
• some litter
  
  • litter observed for behavioral and developmental effect

Question 25

repro tox studies for drugs for long term use must be studied over

Answer 25

2 generations

Question 26

repro tox studies examing the effect on organogenesis must include this number and level of dose

Answer 26

* 3 dose levels * maternal toxic dose * pharmacological effect * geometric mean in between

Question 27

repro tox studies are commonly performed in these species

Answer 27

rat and rabbit

Question 28

mutagenic studies are required to be performed under these circumstances

Answer 28

* if the drug reacts with nuclear DNA or if it can be transformed by metabolism to a reactive species

Question 29

a mutagen is

Answer 29

capable of inducing stable heritable changes in DNA sequence of a cell or organism

Question 30

Mutagenicity occurs in three ways

Answer 30

* point mutations - change of a base sequences * structure breakage in chromosome - clastogenic activity * chromosomal breakage - formation of additional nuclei

Question 31

chemical mutations that occur in germ cells are\_\_\_\_and in somatic cells are\_\_\_\_

Answer 31

* teratogenic * carcinogenic

Question 32

what is the Ames test

Answer 32

* an in vitro assay used to determine if a compound is a potential carcinogen by assessing its mutagenic effect on bacteria

Question 33

what is the basis of the the Ames test?

Answer 33

* uses a strain of bacteria which has a mutation that requires histadine to grow * use media with minimal amount of histadine. When histadine is depleted, only those cells which are able to revert or mutate are able to grow * if a mutagen is present the cells will grow

Question 34

what are some criticisms of the ames test

Answer 34

* it is prokaryotic and doesn't reflect the mammalian metabolic process * does not work with compounds that are metabolicly processed into mutagens

Question 35

what does the current ames test require?

Answer 35

* use of S9 rat mitochondria fraction and NADPH obtained from glc -6 phosphate and NADP * multiple bacterial strains demonstrating different mutation types * control with similar structure to the test article * negative control to account for solvent effect * 5 concentrations with 3 fold difference between, upper concentration limit 5 mg/plate

Question 36

what are four other mutagenicity tests that can be conducted in vitro?

Answer 36

* metaphase chromosome abberation assay * micronucleus test * unschdeuled DNA synthesis * sister chromatid exchange

Question 37

when do carcinogenicity studies need to be conducted (5)

Answer 37

1. for novel drugs 2. long term use 3. long half life 4. persistent metabolites 5. if any mutagenicity test is positive

Question 38

long term invivo carcinogenicity tests usually include these species and are of this duration

Answer 38

* rats or mice * 2 years of dosing and 130 weeks of observation