2. dose-response Flashcards
Hazard
in North America, is the intrinsic toxic properties of a toxicant or toxicant mixture.
Risk
is the probability (so statistical…) of an adverse outcome based on the exposure and potency of the hazardous toxicant(s).
Risk assessment
characterization of the probability of potentially adverse health effects from human (or other animals!) exposures to hazardous agents.
Risk management
process by which policy actions are chosen to control hazards.
Hazard Identification
Do the chemicals/agents present cause — or have the potential to
cause — adverse health affects?
Epidemiology data (correlative)
studies the patterns, causes, and effects of health and disease conditions in defined populations.
In vitro tests
test how cell cultures respond to exposure)
Structure activity modeling
does the chemical look like a toxicant
bioassay experiments
One way to prove causation
Can be done on live animals (in vivo)
bioassay
quantitative estimation of the intensity or concentration of a biologically active chemical, measured via some biological response under standardized conditions.
Common animals used in bioassays
Test group
Mammals Rats, mice, guinea pigs, monkeys Fish Goldfish, fathead minnows, Japanese medaka, rainbow trout Invertebrates Cladocerans (water fleas), oysters
Goals of toxicity testing:
- Determine the range of doses over which the
toxic responses are produced - Identifythenatureoftheresponsestoa toxicant
- Extrapolation of these results for risk assessment analyses for human exposure
Uses of bioassays in environmental toxicology:
- Determine of the most sensitive species or life stage in an
assemblage of organisms. - Compare effects of different pollutants on a single organism.
- Compare effects of other environmental factors acting with the pollutant that modify its effects.
- Determine the maximum level of a pollutant that may occur in the environment without causing biological change (Maximum Allowable Toxicant Concentration: MATC).
Dose-Response assessments quantify the relationships between the exposure concentration (i.e., the dose) and a defined
endpoint (i.e., the response).
• Often represented graphically – a dose-response curve.
• Fundamental concept in toxicology.
• Usually the first step in toxicity testing
Endpoints can be
whatever we are most interested in measuring (e.g., molecular, enzymatic, organismal, etc.
• Recall adverse outcome pathways.
What to we mean by dose?
We tend to give the same amount of substance per unit of body weight in bioassays (and pharmaceutical drug tests)
• The same dose of a toxicant can have a greater effect on smaller individuals or organisms
• give80mgofatoxicanttoan80kgadult…adoseof1mg/kg
• give 80 mg to a 8 kg child… that child is dosed at 10 mg/kg
• This is why we standardize doses to body weight when studying organisms!
important for understanding the response in a test.
Duration and Frequency
Acute exposures
means different things to different people…
– Physiologists: acute is seconds to minutes
– Toxicologists: acute is hours to a day (i.e., 24 hrs)
– BUT, acute toxicity tests for aquatic organisms are often exposures for days
Chronic exposures
means different things to different people…
– Physiologists: chronic is weeks to months
– Toxicologists: chronic is months to years
This is why we must clearly state how long an individual was exposed to a toxicant
For example, we did an acute toxicity test and measured what concentration would kill (endpoint) 50% of the fish in a tank over 96 hrs.
• This is the definition of the lethal concentration (LC50) or lethal dose (LD50)
• Then we should report the concentration as the 96-hr LC50.
Graded dose-response relationships
use a continuous variable and occur in a single individual. They are characterized by a dose- related increase in the severity of the response.
• Examples:
– enzyme inhibition (pictured)
– changes in blood hormone levels
Quantal dose–response relationships occur in a
population
• At a given dose, an individual in the population is classified as either
a “responder” or a “non-responder.”
Often the endpoint is mortality (easy to quantify)
• Count how many individuals die (i.e., the responder…) in your experiment at a given concentration
• Example of a quantal dose-response relationship
-Often we want to know the dose when 50% of individuals die (i.e., the LD50 or LC50).
Can use dose-response curves to compare relative
toxicity of toxicants.
• We should use the same test conditions and test species.
toxicants can be ranked based upon their
LD50s into toxicity classes
Class 6
Relatively harmless (Acute toxicity of chemicals in rats*)
Class 5
Practically non-toxic (Acute toxicity of chemicals in rats*)
Class 4:
Slightly toxic
Acute toxicity of chemicals in rats*
Class 3
Moderately toxic (Acute toxicity of chemicals in rats*)
Class 2
Highly toxic
Acute toxicity of chemicals in rats*
Class 1
Super toxic
Acute toxicity of chemicals in rats*
Dose-Response curves will change based on
“route of exposure”
– Inhalation, Injection, Ingestion, Dermal (in order of effectiveness)
– The speed of the response is related to how quickly a toxicant can get into the bloodstream.
We can try to transform to linear functions if possible.
– Because it is hard to generate a nice sigmoidal curve without a lot of things dying
The response at 50% of the population is, by definition, the population mean.
Normally distributed curves have fixed standard deviations (SD) that always represent a percent of the population. For example 1 SD = 69.3% of the population.
So we can swap the y-axis from cumulative response to deviations away from the mean (Probit – Probability Units).
The y-axis gets converted to ‘probits’
The y-axis on a probit graph is always the same in probit plots (convenient for comparisons…)
1) the % response gets changed to the Normal Equivalent Deviation (NED
2) Add 5 to all to avoid negative numbers
3) The new probit Y-axis evenly spaces things to effectively turn the sigmoidal curve into a straight line
5 always corresponds to 50% of the population. 6 is one NED away from mean which represents cumulative 84% of the population.
The data points can be converted from % mortality to probits by looking up the values in a chart
- Convert%mortalityvaluestoprobitvalues 2. Any%mortalitybelow1or100isignored 3. Tolookupinthetable,the%moralityis
rounded to the nearest 0.5
These values will plot as a linear ‘probit plot’ when plotted vs
log dose
Can use this to calculate the LC50 when you don’t know it.
ED50
effective dose
where the drug is doing what we want it to do
in 50% of the populatio
TD50
toxic dose
where bad things start happening in 50% of the population
LD50:
lethal dose in 50% of the population
therapeutic index (TI)
is the ratio of TD50 and ED50
High values mean that the drug is relatively safe.
Low values usually lead to the rejection of the drug from further clinical testing.
TI = TD50 ED50
Potency
the amount required to produce an effect of given intensity A is more potent than B
Efficacy
refers to the maximum response achievable D has a greater efficacy than C
NOEC
No Observable Effect Concentration
LOEC:
Lowest Observable Effect Concentration
to distribute the contaminant to the animals.
Often use a “vehicle” or “carrier”
Chlorpyrifos
used on crops like strawberries, broccoli and citrus fruit as an insecticide.
– Residues can harm the developing brains of small children, even in the womb.
• Environmental Protection Agency (EPA) issued a proposal to ban chlorpyrifos 2 years ago.
• This was blocked by newly appointed EPA Administer Scott Pruitt (a lawyer)
– Has sued the EPA before, received money from the oil and gas industry, does not believe in climate change, no science background.
– Should NOT be in the position to interfere with the EPA trying to protect the health of citizens.
Dose-response curves can be
U-shaped or inversely U- shaped as well
– Some essential nutrients can have adverse effects at low and high doses (e.g., iron, chromium, vitamin A and D)
Hormesis
can refer to some types of dose-response curves
• Compounds that are beneficial/stimulatory at low doses but toxic at high doses
Nonmonotonic dose-response curves
Often observed with EDCs: toxicants that can have effects at very low levels
At low levels they interact with a hormonal receptor, leading to a response
At high levels, the receptor is ‘burned out’, thus the response decreases
Highlights the danger of extrapolating the effects of high doses to low dose responses
Basically curves that are not straight lines These indicate thresholds in the response
Sub-lethal
responses are very important
Many (most) toxic events occur before the animals died…
Can conduct a subchronic bioassay
at lower doses. Often with more than 1 species at different doses (i.e., 3 doses):
• High dose that causes toxicity but not more than 10% fatalities
• Low dose that should cause no toxic effects
• An intermediate dose
• Exposures for longer periods of time
• e.g., 90 days
Why do a subchronic test??
- To provide information on all types of subchronic
toxicity (other than carcinogenicity) that might occur – immunotoxicity, neurotoxicity - Establish dose regimens for chronic studies
- Provide data which will allow an estimate to be made of the MTD (maximum tolerable dose), where there is no significant impairment of growth.
- To develop biomarkers of exposure (e.g., expression of a gene, specific enzyme function).
Observed during the study:
– Altered growth rate
– Behavioural changes
After the exposure period, the animals are euthanized and sampled to see what deviates from ‘normal’:
– Mass – Blood chemistry (hormone levels) – Biochemical (glucose, ATP levels) – Cell membrane permeability – Enzyme activity – Organ and tissue damage (necrosis)
biomarker
can be described as a cellular or biochemical response to a xenobiotic that is measurable in a biological system or sample.
– There are biomarkers to other types of stressors, such as temperature in fish!!!
Biomarkers can include, but are not limited to:
i. Enzyme activity
• Certain enzymes in the blood can indicate liver toxicity
• Increase in cytochrome P450 activity for detoxification
• Metallothionein induction to bind heavy metals
ii. Changes in cellular receptor amount and activity
• Estrogen receptor in response to EDCs
iii. Changes in hormone levels
• Increase in cortisol (the ‘stress hormone’)
iv. Increased/decreased expression of genes that respond to toxicants
v. Changes in the histology of cells to detect cellular damage
chronic bioassay
Can conduct a chronic bioassay to look at longer-term endpoints like cancer development (months to years).
– Determine whether it is a carcinogenic compound.
• Cancer can take a long time to develop, and likely will not be identified in a subchronic study.
Carcinogens are classified based upon
what is known from humans (e.g., epidemiological studies) and animals (e.g., bioassays)
Rofecoxib (trade name Vioxx)
is a non-steroidal anti-inflammatory drug (NSAID)
