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Q: Toxicology ( Answered,   0 Comments )
Question  
Subject: Toxicology
Category: Science > Chemistry
Asked by: demi1981-ga
List Price: $30.00
Posted: 22 Jul 2003 08:28 PDT
Expires: 21 Aug 2003 08:28 PDT
Question ID: 233736
Giving named examples the role of enzymes which can either activate or
eliminate the offending toxins

Request for Question Clarification by tehuti-ga on 22 Jul 2003 09:30 PDT
Is any particular class of toxins being referred to in your question?

Clarification of Question by demi1981-ga on 22 Jul 2003 12:54 PDT
any type of toxins that harm the body
Answer  
Subject: Re: Toxicology
Answered By: tehuti-ga on 24 Jul 2003 19:31 PDT
 
Hello demi1981,

I want to start with a general introduction to set the background to
your question, and then move on to the specific named examples you
request.

The human organism has evolved a number of systems to deal with
xenobiotics (substances that enter into the body from outside),
whether these be toxins, nutrients or drugs, and also to deal with
some of the substances produced during its own physiological
processes.  One vital function of these systems is to deactivate
toxins and/or convert them into more soluble forms, so that they can
be more easily excreted by the body, rather than becoming accumulated
and remaining in the body for long enough to have a noticeable toxic
effect.  Thus, lipophilic, nonpolar molecules will tend to accumulate
within fat tissue, but if they are converted into polar, water-soluble
molecules, they will be readily excreted in urine (and also in faeces
and sweat).  However, there are so many different types of chemicals,
that in some cases some of these reactions, while normally making
substances less toxic, bring about metabolic activation of a substance
into a more reactive species, and at times may even convert a totally
non-toxic substance into a toxic intermediate. In some cases,
metabolic activation can be helpful, when it converts a nutrient into
a form more easily used by the body, or when it causes a drug to be
converted into its therapeutically active form.  There is a fine
balance between metabolic activation and detoxification and between
adsorption and excretion of a substance, which will decide whether or
how toxic a substance will be within the body.  This balance can be
affected by a number of factors, including genetic factors, diet,
health status and medication.

A very large part of this processing of xenobiotics takes place in the
liver. However, most cells have some capacity to metabolise
xenobiotics, and the kidneys, gastrointestinal tract and lungs can
account for quite significant metabolism.

These various reactions are classified into two major sets, which are
normally referred to as Phase I metabolism and Phase II metabolism.

Phase I metabolism is catalyzed by a group of enzyme collective known
as mixed function oxidases, or as the cytochrome P450 system;
individual enzymes are referred to by the prefix CYP followed by a
combination of letters and numbers, for example, CYP2C, CYP3A4,
CYP2E1.  These enzymes are located in the microsomes of the smooth
endoplasmic reticulum in liver cells, and also in the mucosa of the
gut, and, in lesser amounts in cells of the kidney, skin, brain and
airways. The reactions that are catalyzed by the cytochrome enzymes
are aromatic hydroxylation; aliphatic hydroxylation; N-, O-, and
S-dealkylation; N-hydroxylation; N-oxidation; sulfoxidation;
epoxidation; deamination; and dehalogenation.  There are also
non-cytochrome enzymes which catalyze some oxidation reactions, for
example, alcohol dehydrogenase. The cytochrome system also catalyzes
some reduction reactions such as hydrolysis, azo- and nitro-reduction,
hydration reactions.

These Phase I reactions serve to add or expose functional groups which
are then acted on by Phase II metabolism.  This involves conjugation
reactions which insert a hydrophilic species into the molecule, to
make it even more soluble and ready for excretion. Phase II reactions
include glucuronidation, sulfation, methylation, acetylation,
glutathione conjugation, fatty acid conjugation, amino acid
conjugation.  Glutathione conjugation by glutathione S-transferase is
a very important Phase II reaction that acts on a huge number of
substances. While Phase I reactions can decrease the activity of some
substances, they can also increase the activity of others.  It is the
Phase II reactions which are the true detoxification stage of the
whole process. They are mainly carried out by non-microsomal enzymes
in the cytoplasm of the liver and other tissues.

Alcohol dehydrogenase, mentioned above, is a very good example of an
enzyme that can have a positive or negative effect on toxicity.  It
detoxifies ethanol (the alcohol in alcoholic drinks). However, the
first product of this oxidation reaction catalyzed by alcohol
dehydrogenase is acetaldehyde, which is actually even more toxic.
Fortunately, another enzyme, aldehyde dehydrogenase, then converts the
acetaldehyde into acetates and other molecules that can be used by the
body.  It is thought to be the build up of acetaldehyde levels which
is responsible for hangover symptoms. Some people of Asian descent
have a genetic mutation which makes their acetaldehyde dehydrogenase
less effective, and consequently they are less tolerant to alcohol,
because they rapidly develop high levels of acetaldehyde with all the
accompanying unpleasant symptoms even after drinking only a small
quantity. The drug disulfram (Antabuse) used for alcohol aversion
therapy in alcoholics works in the same way by blocking the activity
of acetaldehyde dehydrogenase, so that even a small drink can lead to
vomiting.  Alcohol dehydrogenase most probably evolved so that our
bodies could cope with the alcohol in naturally fermented fruits, or
produced by bacteria.  It is also involved in the metabolism of
retinol, steroids, and fatty acids. However, the very same oxidation
reaction that is an important step in the detoxification of ethanol is
also responsible for the highly toxic effects of methanol.  In this
case, the first reaction product is not acetaldehyde, but
formaldehyde. This is further metabolised by aldehyde dehydrogenase
into formate/formic acid. Formic acid inhibits mitochondrial
respiration.  It affects the optic nerve, causing blindness. In larger
doses it is lethal.  As little as 30 ml (just over one fluid ounce) of
40% methanol can be fatal to a human.

Acetaminophen toxicity is an example of the important balance between
Phase I and Phase II metabolism.  In normal circumstances, most of the
acetaminophen is acted on by the Phase II enzymes glucuronyl
transferase and sulfotransferase to form the glucuronide and sulfate,
which are easily excreted.  However, about 4% of a normal dose is
metabolised by the Phase I cytochrome CYP2E1 into a toxic metabolite,
N-acetyl-benzoquinoneimine (NAPQI). A phase II reaction mediated by
glutathione S-transferase detoxifies NAPQI by conjugating it to
glutathione. However, in the case of an overdose, the glucuronyl
transferase and sulfotransferase enzymes cannot metabolize sufficient
of the drug.  This results in more of it being acted on by CYP2E1, so
more NAPQI is formed. Once the body’s store of glutathione is
exhausted, the NAPQI can no longer be detoxified. Instead, it is able
to bind to elements within the liver cells causing their destruction,
and ultimately liver failure. Another factor can also be important in
acetaminophen toxicity. Some chemicals are able to induce the
production of higher levels of certain cytochrome enzymes. CYP2E1 is
induced by various drugs, including barbiturates, and also by alcohol.
An increased level of CYP2E1 means it is able to catalyze the
formation of more NAPQI. In this way, a dose of acetaminophen that is
normally safe can be toxic if taken in combination with a CYP2E1
inducer.

Such induction of enzymes, or their inhibition (as in the case of
disulfram mentioned above) is the reason why some drugs or even some
foods can interact with other drugs making a standard dose either
ineffective or toxic. An example of a food having an effect is the
finding that grapefruit juice can inhibit the activity of CYP3A4,
which is important in the metabolism of a number of drugs, including
cortisol, omeprazole and erythromycin.

Another example of genetic effects on toxicity, in addition to that of
alcohol dehydrogenase, is that of isoniazid, which is the first line
drug used in the treatment of tuberculosis. Isoniazid is detoxified by
an acetylation reaction mediated by N-acetyl-transferase. Most people
are so-called “fast acetylators”, however, some people have a mutation
which results in a less effective enzyme, and these are known as “slow
acetylators”.  A fast acetylator needs 70 minutes to eliminate half of
a dose of isoniazid, while a slow acetylator needs 180 minutes.
Therefore, unless slow acetylators are given lower doses of isoniazid,
they are likely to build up toxic levels of the drug. This is
reflected in the fact that slow acetylators given isoniazid are more
likely to develop peripheral neuropathy (a neurological disorder
causing pain, numbness, weakness or tingling in the arms, hands, feet
and legs.

N-acetyl transferase is also important in detoxifying aromatic amines,
which are a class of mutagens found in tobacco smoke and other
environmental pollutants. A mutagen can affect the DNA of a cell and
cause it to become abnormal, resulting in cancer. A study which was
the subject of an NIH news release (ref. 9 below)  looked at its
effect in post-menopausal women: “Neither slow acetylators nor heavy
smokers by themselves had an increased risk of breast cancer before or
after menopause. But post-menopausal women who were both slow
acetylators and heavy smokers had an increased incidence of breast
cancer. The number of cigarettes smoked daily and smoking at young age
rather than number of years smoked, appeared to confer the most risk
in this latter group. For example, post-menopausal women who were slow
acetylators and who smoked more than a pack a day for 20 years, had
3.9 times greater risk than the non-smoking slow acetylators.”

A number of different enzymes can together play a part in determining
toxicity.  For example, here is a small extract taken from an
invitation for grant applications to the NIH (ref 10) which summarizes
some of the factors that can determine the susceptibility to
tobacco-induced cancer:
“Many of these enzymes involved in tobacco carcinogen metabolism are
also induced by environmental factors such as alcohol usage, dietary
constituents, pesticide and xenobiotic exposure, hormonal status and
occupation.  Induction of activation (P450s) stimulates
carcinogenesis, while induction of detoxification enzymes (epoxide
hydrolase, glutathione transferase, N-acetyl  transferase,
sulfotransferase, UDPG transferase) decreases tumorigenic response. 
Polymorphism [existence of more than one form / mutation ] in genes
that code for the synthesis of metabolizing enzymes may result in
lower affinity or differential specificity.  Polymorphism in DNA
repair genes and the influence of fragile sites or mutational hot
spots in the genome may also be involved.”

The heterocyclic amines (HCAs) found in cooked meat need metabolic
activation in order to be converted into reactive metabolites which
cause DNA damage which can lead to cancer:
“Cooked meat contains a variety of mutagens, including HCAs formed
from the cooking of proteinaceous foodstuffs (particularly meat and
fish) at high temperatures. These compounds are carcinogens in the rat
mammary and prostate glands following high-dose oral administration,
and are suspected of being mammary carcinogens in women and prostate
carcinogens in men. HCAs require metabolic activation to form
DNA-damaging metabolites. This can occur via a two-step pathway
involving first a CYP [cytochrome P450] catalyzed N-hydroxylation
followed by O-esterification catalyzed by NAT [N-acetyltransferase]
and/or SULT [sulfotransferase] enzymes. The acetoxy or sulfoxy esters
formed are unstable and generate a nitrenium ion as the ultimate
DNA-reactive intermediate, and quantitatively,
N-(deoxyguanosin-8-yl)–HCAs are the major adducts formed. Parent HCAs
can also be activated by peroxidases to generate nitrenium ions. (ref
11)

Ref 11 also mentions that peroxidases can be implicated in breast
cancer through the activation of mutagens:
“Two interesting epidemiological studies have shown that women with
allergies/atopic diseases (i.e. those associated with inflammation and
elevated peroxidase activity), compared with women without atopic
diseases, have an elevated risk of developing cancer of the breast
….  Levels of MPO [myeloperoxidase]-containing neutrophils are
elevated in inflamed mammary tissues compared with non-inflamed
tissues, and are present in breast milk and breast secretions. The
gene encoding the human form of another peroxidase enzyme,
lactoperoxidase, is arranged `tail-to-tail' with the MPO gene. The
enzyme encoded by this gene also has bacteriocidal functions, and is
secreted into the milk ducts. The catalytic activity of the human form
of this enzyme has not been well studied, but the bovine form of this
enzyme can activate 17-estradiol, aromatic amines and HCAs to
mutagenic metabolites.”

The author of ref 11 also makes a very pertinent concluding statement,
which has wider validity than just to the area of environmental
carcinogenesis to which he refers:
“Information on all the competing pathways of metabolic activation and
detoxication of suspected human carcinogens should also be taken into
account. Peroxidases and SULT enzymes have wider substrate
specificities than NAT enzymes. Thus, while NAT enzymes are limited to
activation of N-hydroxylated heterocyclic and aromatic amines,
peroxidases can metabolically activate both parent and hydroxylated
heterocyclic and aromatic amines and hydroxylated PAHs, and SULTs can
activate the hydroxylated promutagenic derivatives of all three
classes of mutagens. Such mixtures are more representative of the
mixtures present in cooked meat and tobacco smoke. Investigation of
polymorphisms in enzymes with wide substrate specificity, with regard
to carcinogen activation therefore looks promising for the detection
of links between environmental or lifestyle factors and cancer risk.”

Sources consulted:

1. Personal knowledge of toxicology picked up as part of my
professional activities.
2. Small Molecule Drug Metabolism by Walter Yu,  Celera Corporation
(Ionsource.com monograph)
http://www.ionsource.com/tutorial/metabolism/drug_metabolism.htm 
3. Phase I Metabolism: Reactions of the Cytochrome P450 Mixed Function
Oxidase System
http://www.pharmacy.wsu.edu/courses/PharS541/riley/Phase%20I%20Metabolism_files/frame.htm#slide0001.htm
4. Phase II Metabolism
http://www.pharmacy.wsu.edu/courses/PharS541/riley/Phase%20II%20Metabolism_files/frame.htm
Both the above are slide show presentations for the course
Pharmacological Basis of Therapeutics by John Riley, College of
Pharmacy, Washington State University
5. PDB (Protein Databank) Molecule of the Month: Alcohol
Dehydrogenase, by David S. Goodsell
http://www.rcsb.org/pdb/molecules/pdb13_1.html 
6. Wikipedia entry for Acetaldehyde 
http://www.wikipedia.org/wiki/Acetaldehyde
7. Minnesota Poison Control System: Methanol
http://www.mnpoison.org/index.asp?pageID=145
8. Emedicine.com article on acetaminophen toxicity by Jeffrey Tucker
http://www.emedicine.com/ped/topic7.htm 
9. Smoking May Be Risk Factor for Breast Cancer in Post-Menopausal
Women With Mutations in Detoxifying Enzyme (NIH News Release)
http://www.nih.gov/news/pr/nov96/nci-12.htm 
10. Genetic Regulation Of Susceptibility To Tobacco-Related
Carcinogenesis
http://grants1.nih.gov/grants/guide/pa-files/PA-98-095.html 
11. “Single nucleotide polymorphisms, metabolic activation and
environmental carcinogenesis: why molecular epidemiologists should
think about enzyme expression”  by J.A. Williams
Published in: Carcinogenesis, Vol. 22, No. 2, 209-214, February 2001
Full text: http://carcin.oupjournals.org/cgi/content/full/22/2/209 

Search strategy:  I started by searching on the terms “Phase I
metabolism” “Phase II metabolism”, which I knew from previous
experience to be the key to finding this information.  I then carried
out a number of searches on individual drug and enzyme names, as well
as on: carcinogenesis, enzymes.
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