Heterogeneity of Toxicant Response: Sources of Human Variability

Justin E. Aldridge*,{dagger}, Jennifer A. Gibbons*,{dagger}, Meghan M. Flaherty*,{ddagger},§, Marisa L. Kreider*,{dagger}, Jocelyn A. Romano* and Edward D. Levin*,||,1

* Integrated Toxicology Program, {dagger} Departments of Pharmacology and Cancer Biology, {ddagger} Biological Chemistry Program, and || Department of Psychiatry, Duke University Medical Center, Durham, North Carolina 27710; § Department of Chemistry, Duke University, Durham, North Carolina 27708; and Nicholas School of the Environment, Duke University Marine Laboratory, Beaufort, North Carolina 28516

Received May 12, 2003; accepted July 17, 2003

ABSTRACT

While risk assessment models attempt to predict human risk to toxicant exposure, in many cases these models cannot account for the wide variety of human responses. This review addresses several primary sources of heterogeneity that may affect individual responses to drug or toxicant exposure. Consideration was given to genetic polymorphisms, age-related factors during development and senescence, gender differences associated with hormonal function, and preexisting diseases influenced by toxicant exposure. These selected examples demonstrate the need for additional steps in risk assessment that are needed to more accurately predict human responses to toxicants and drugs.

There is wide variation in individual human responses to environmental toxicants and drugs. Variations can be caused by seemingly unrelated differences such as genetic polymorphisms, age, and gender, as well as co-existing diseases or infections. A large diversity in responsiveness among individuals to environmental and industrial toxicants makes it difficult to determine actual risk, particularly at the low doses to which most people are exposed (Hanson and Solomon, 2002Go). The purpose of this review is to correlate human variability with risk assessment.

Opportunities now exist to study susceptibility for the development of cancer and other diseases in which an environmental component is present. Knowledge from such studies allows markers of susceptibility to be incorporated into epidemiologic studies. This is important because it is widely recognized that, the larger the body of knowledge regarding chemical exposure and human variability, the smaller the uncertainty of risk. A significant amount of research has gone into developing risk assessment strategies for better prediction of safe toxicant levels. By identifying those susceptibilities that affect individual toxic responses to environmental toxicants and drugs, researchers can better predict health risks and assist regulatory agencies in the development of health protection policies.

The rapid advances in molecular genetic technologies provide new opportunities to understand the genetic basis for individual differences in susceptibility to toxicant exposure. The link between genetic polymorphisms and disease is well documented; however, researchers are finding links with more subtle effects such as toxicant sensitivities. New polymorphisms are still being identified; provided in this review is information regarding commonly found polymorphisms as well as rarer genetic variants. The effects of these genetic variations must be understood before drug and toxicant sensitivities in an individual can be accounted for.

Response to environmental toxicants and the health problems associated with exposure often have an age bias. Developing individuals are particularly vulnerable to toxic insult because of the significant physiologic changes occurring over a short time span. Life-long exposure to toxicants increases the susceptibility of elderly individuals to new toxicants, especially when influenced by the overall aging process. Specific examples of toxicants with increased or decreased susceptibility related to age are provided in this paper.

Gender-specific toxicant response is intrinsically tied to age and genetics. The point in development at which exposure occurs often will be the deciding factor as to the extent of the toxicant effect on gender differentiation. Additionally, research has found that hormonal differences between the sexes influence the response an individual has to possible endocrine-disrupting toxicants. The wide range of possible toxicant effects associated with specific genders has made this a complex area of study; however, significant information can be obtained through consideration of known endocrine disruptors.

Superimposed on the heterogeneity of toxicant response caused by genetics, age, and gender is susceptibility conferred by preexisting diseases. Alterations in an individual’s immune system may result in increases in new infections as well as decreases in the body’s ability to fight existing pathogens. Toxicants that influence immune status may therefore increase the symptomatic progression of diseases or increase the likeliness of developing new diseases. It is therefore necessary, when determining an individual’s risk to toxicant exposure, to establish the preexisting disease state. This article discusses the need for risk assessment models in predicting risks to humans and the susceptibility of individuals to toxic exposures by numerous factors including genetics, age, gender, and disease state.

Uncertainty Factors

Due to large variations in human populations, risk assessment strategies have changed over the years to incorporate uncertainty factors that might aid in more accurate indications of acceptable risks. Genetics, age, gender, and disease state are all elements that must be addressed to further the advancement of predicting toxic exposure levels. Before we begin to investigate these various elements, it is critical to have a solid understanding of the current approach in risk assessment. The following is a brief introduction to the way chemicals are presently examined for toxicity.

Risk Assessments
The world’s production of chemicals has far outpaced our ability to evaluate their potential hazards to human health and the environment. Each year the United States adds between 1500 and 2000 new chemicals to an already staggering 60,000 chemicals currently used in industrial, pharmacological, and agricultural products (Harry, 2001Go). Adverse effects in humans occurring at sufficiently high exposure levels can range from systemic problems affecting the excretory, reproductive, nervous, respiratory, and circulatory systems, to a possible endpoint of death. Only a small fraction of chemicals has been evaluated for these adverse effects.

Due to the large uncertainty of human response to these unknown chemicals, risk assessment programs for chemicals have been established. Risk assessment refers to a complex action for predicting health risks in human populations caused by chemical exposures. The National Academy of Science (NAS) described four components in risk assessment: hazard identification, dose–response assessments, exposure assessment, and risk characterization. Hazard identification is the assessment of data that may suggest that a chemical produces adverse health effects. The data may come from laboratory studies, case studies, or epidemiological reports. Doseresponse assessment attempts to define the relationship between exposure level (dose) and the incidence of adverse effects in humans. Much of what is captured under the dose–response assessment deals with a variety of extrapolation issues. Since most of the available data are from laboratory animals only exposed for short periods of time to high concentration levels, extrapolations must be made to estimate the outcomes of chronic human cases of exposure. Exposure assessment focuses on the exposure levels actually encountered in the environment. Risk characterization puts together all the available information from the preceding components to arrive at estimates of human risk under varying exposure conditions.

Predictions of risk are often made despite incomplete data (Dourson et al., 1992Go). Approaches to risk assessment are therefore likely to be conservative, with the justification being that it is better to overestimate rather than underestimate risk to human health. A risk assessment typically begins by selecting an exposure level that is less than that which produces an adverse health effect and accounting for additional uncertainty to arrive at what is considered a safe or acceptable level for human exposure. No-observable-adverse effect levels (NOAELs) are used to establish human acceptable exposure levels for noncarcinogens. The reference dose (RfD) approach is widely used for setting environmental exposure limits for chemicals producing systemic toxicity. RfDs are derived from a NOAEL by applying uncertainty factors (UFs) and modifying factors (MFs) (Gaylor and Kodell, 2002Go):


(1)

Safety factors allow for interspecies and intraspecies variations. Default UFs of 10 are assigned when relevant research-based information is missing. However, a UF of 3 is commonly used in the presence of a plethora of data, as it represents the logarithmic mean of possible UF values (Dourson et al., 1996Go). MFs are used to adjust the UFs if data on pharmacokinetics, pharmacodynamics, or mechanisms of action are available to evaluate the relevance of animal information for human responses. The magnitude of the uncertainty factor depends on the nature of the data available, but generally consists of a multiple of 10 (Dourson et al., 1996Go).

Uncertainty in Results from Laboratory Animals
If adequate toxicity data on humans do not exist, animal models are used as the basis of the assessment, and an uncertainty factor of 10 is routinely applied to the NOAEL. The basic assumption for this uncertainty factor is that, although the results seen in experimental animals are relevant to humans, toxicokinetic and toxicodynamic differences exist among species, and humans are more sensitive than animals at a given mg/kg/day dosage (Jarabek, 1995Go).

Extrapolation of animal studies to humans depends on these factors: the critical site of toxicity within human cells or tissues, the presence of protective mechanisms, and the sensitivity of the site of toxicity to the active chemical entity (Dourson et al., 1996Go). The fate of the chemical in the body is influenced by species differences in the rate and extent of metabolism and by physiological differences, such as heart rate, cardiac output, and renal and hepatic blood flows. For example, hepatic and renal plasma flows are three to four times lower in humans than in rats (Renwick, 1993Go); therefore, humans are likely to have higher body loads for a comparable daily intake. When differences in liver weight and enzyme activity are taken into account, there may be larger differences in clearance, half-life, and body burden. These differences can be such that, during chronic intake, much or the entire default factor of 10 may be required for toxicokinetics.

Uncertainty in Human Variability
Whenever possible, data from humans are used to conduct noncancer risk assessment, thereby avoiding problems with interspecies extrapolation. If sufficient data on sensitive individuals exist, the subthreshold dose can be estimated directly without the need of an uncertainty factor. However, if there are inadequate data on sensitive humans, uncertainty is encountered and must be addressed. Most often this is covered by a ten-fold default factor. This uncertainty factor accounts for interhuman variability in responses that may not have been detected in the study. This factor may also assume that subpopulations of humans exist that are more sensitive to certain chemicals than the average population. An example of this is a genetic polymorphism found in over 90% of the Japanese population. This polymorphism results in a decrease in aldehyde dehydrogenase activity, resulting in an accumulation of acetaldehyde in the blood, causing various uncomfortable symptoms after alcohol consumption (Sun et al., 2002Go). In general, the default value of 10 for interhuman variability appears to be protective when starting from a median response or by assumptions made from a NOAEL assumed to be from an average group of humans. However, when no NOAELs are available for a known sensitive human subpopulation, or if human toxicokinetics or toxicodynamics are known with some certainty, this default value of 10 should be adjusted to a lower value (Gundert-Remy and Sonich-Mullin, 2002Go).

Uncertainty in Risk at Exposures below the Lowest-Observed Adverse Effect Level
A number of approaches can be taken in situations where a NOAEL cannot be identified but the data are of sufficient quality for use as the basis of the risk assessment (EPA, 1994Go). Uncertainty factors 1 through 10 can be applied to the lowest-observed-adverse effect level (LOAEL) depending on the endpoint and/or dose–response relationship. For example, in a study done by the EPA on the effects of di-(2-ethylhexyl)phthalate, no NOAEL was evident (EPA, 1996Go). A two-fold uncertainty factor was used for the extrapolation of NOAEL from LOAEL because the only effect observed in guinea pigs at the lowest dosage of 19 mg/kg/day was an increase in the relative kidney weight, which was not accompanied by any histopathological effects.

The data indicate that, when faced with a LOAEL and not a NOAEL, the choice of an uncertainty factor should generally depend on the severity of the effect at the LOAEL. More severe effects should be judged to need a larger uncertainty factor because the expected NOAEL is farther away from the LOAEL. Less severe effects would not require a large factor, presumably because the LOAEL is closer to the unknown NOAEL.

Uncertainty in Less-than-Lifetime Exposure
Lifetime or chronic studies may not be available, necessitating the use of data from a shorter subchronic study. Studies show that the ratio of the subchronic NOAEL or LOAEL to the corresponding chronic NOAEL or LOAEL was in most cases less than 10, and in 50% of cases was 2 or less, suggesting that a value less than 10 would be adequate (Dourson and Stara, 1983Go). If no adequately performed long-term studies are available, data on the extent of accumulation in the body and the rate of excretion may be relevant in selecting a database factor.

Uncertainty in Database
The information on a chemical may be considered incomplete because of incomplete information regarding particular routes of exposure, number of test subjects studied, or sensitivity in the subpopulation of test subjects. Major deficiencies in the adequacy of the database, which increase the uncertainty of the extrapolation process, can be accounted for by the use of an additional uncertainty factor (EPA, 1994Go). Since quality and completeness vary, the size of the uncertainty factor applied will also be variable.

It is generally assumed that a threshold exists for systemic toxicants and that toxicity results only after some minimum exposure concentration has been exceeded. While the RfD approach is generally very conservative, it may not account for all instances of susceptibility in human exposures (Hattis et al., 2002Go). Another approach that is commonly used in place of the NOAEL in RfD is the benchmark dose (BMD). This method tends to be less conservative and more predictive of threshold limits.

BMD to Estimate Reference Dose

Regulatory agencies mandated to protect public health are continually confronted with the task of estimating safe levels of toxic, noncarcinogenic chemicals. For more than 30 years such determinations have involved finding an experimental dose level that shows no observed effect and accounts for intraspecies and interspecies variability using uncertainty factors. In 1988, the EPA adopted the reference dose, a variant of this general procedure, which requires the identification of the NOAEL or LOAEL.

Many scientists have expressed concern that the use of the RfD approach has a number of limitations stemming from the use of NOAEL/LOAEL. Criticisms include (Crump, 1984Go):

NOAEL/LOAEL is limited to one of the doses in the study, and the ability to metabolize and eliminate is dependent on the study design.

NOAEL/LOAEL does not account for variability in the estimate of dose response.

NOAEL/LOAEL does not account for the slope of the dose–response curve.

NOAEL/LOAEL cannot be applied when there is no NOAEL, except through the application of an uncertainty factor.

The BMD was formulated to address these concerns by defining a starting point for the computation of an RfD or slope factor that is independent of the study design. BMD methods involve fitting mathematical models to dose–response data and using the different results to select a BMD that is associated with a predetermined benchmark response (BMR), such as a 10% increase in the incidence of a particular lesion or a 10% decrease in body weight gain. The benchmark dose is usually defined as the lower confidence limit of the dose that produces a specified magnitude of change in a specified adverse response (EPA, 1994Go). The BMD is determined by modeling the dose–response curve in the region of the dose–response relationship where biologically observable data are feasible. The benchmark dose method is not used for extrapolation to low doses where biological responses can only be estimated (Faustman et al., 1994Go)

Overall, the BMD approach to risk assessment has a number of advantages over the traditional use of NOAEL. The BMD provides estimates of a toxicological concentration that can replace, or refine, the NOAELs currently used in health risk assessment. In most cases, the BMD lies somewhere between the NOAEL and the LOAEL.

Genetic Susceptibility

Genetic susceptibility to the adverse effects of toxicants is a well-known phenomenon; this effect is accounted for by the uncertainty factor for human variability. Although some diseases have a well-defined etiology and therefore have many subjects available for genetic studies, it can be more difficult to discover the origin of more subtle genetic susceptibilities. In order to avoid drug toxicity as a result of genetic predisposition, genotyping in hospitals is necessary to tailor any drug combination to the genotype of the patient. Candidates for genotyping approaches include cytochrome P450 enzymes, DNA-associated enzymes, and receptors.

P450 Enzymes
The cytochrome P450 system (CYP450), which is required for the metabolism of toxicants and drugs in the human body, is of significant interest to scientists. Phase-I P450 enzymes add a functional group to increase polarity, and phase-II P450 enzymes add larger hydrophilic moieties to enhance excretion of the compound. There are many phase-I isozymes, each with a different specificity; examples of the wide variety of polymorphisms in the many phase-I enzymes are listed in Table 1Go.


View this table:
[in this window]
[in a new window]
 
TABLE 1 Information on Various P450 Enzymes
 
Many deleterious polymorphisms exist for phase-I enzymes. For example, CYP2D6, which metabolizes 25% of prescription drugs, has several clinically relevant polymorphisms (Pirmohamed and Park, 2001Go). The most common mutations for CYP2D6 are CYP2D6*4 and CYP2D6*5 (Ingelman-Sundberg et al., 1999Go); patients with these mutations are considered poor metabolizers (PMs). Since 6% of Caucasians are homozygous null for this P450, the mutation affects a large number of people and can cause serious side effects from overdosage, especially in drugs with a narrow therapeutic range. For example, an anti-psychotic drug such as zuclopenthixol requires precise dosages to be effective in treating schizophrenia. However, patients who cannot metabolize these drugs often have drug accumulation in their system that exacerbates side effects, such as parkinsonism and tardive dyskinesia (Jaanson et al., 2002Go). Similarly, if P450-mediated metabolism is required to activate the drug in vivo, a patient with the mutated allele may not experience a therapeutic effect. An excellent example of P450-mediated processing of a prodrug into a pharmacologically active form is the conversion by CYP2D6 of codeine into morphine, which causes the analgesic properties associated with codeine administration (Ingelman-Sundberg et al., 1999Go). Therefore, patients lacking CYP2D6 experience no analgesia from codeine, since it is never metabolized into morphine.

There exists a subpopulation of individuals that have an extra copy of the CYP2D6 gene (CYP2D6*2 x N), making them ultrafast metabolizers. The highest prevalence of this subpopulation has been found in Ethiopia and the surrounding countries (Ingelman-Sundberg, 2001bGo). Natural selection favors individuals with the CYP2D6*2 x N mutation. Due to the scarcity of food, those people who can supplement their diet with alkaloids without experiencing toxic effects have a greater chance of survival (Ingelman-Sundberg, 2001bGo). While this mutation can be favorable, in some situations, including pain management, this mutation can be deleterious. For example, patients treated with the alkaloid codeine would receive the expected analgesic effect; however, they are in jeopardy of morphine overdose (Ingelman-Sundberg et al., 1999Go). CYP2D6 metabolizes codeine faster than the P450 that metabolizes morphine, resulting in opioid side effects, including respiratory depression. Therefore, in a pharmacological context mutations in CYP2D6 can be deleterious, whether they enhance or decrease enzyme activity.

Sometimes polymorphisms in a P450 enzyme provide an advantage over the more common allele. Although CYP2A6 only metabolizes less than 1% of drugs (Anzenbacher and Anzenbacherova, 2001Go), it is medically important because of the toxicants it metabolizes. CYP2A6 is known to activate procarcinogens in tobacco smoke, such as 4-methylnitrosamino-1–3-pyridyl-1-butanone (NNK) (Raunio et al., 2001Go). Polymorphisms resulting in poor metabolism via CYP2A6 have a protective effect. This deficiency prevents the conversion of NNK, protecting the individual from carcinogenesis. Lung cancer patients have a lower percentage of CYP2A6 deficiency compared to the average population (Miyamoto et al., 1999Go), although more research is needed to support this conclusion.

N-acetyl transferase (NAT) is a phase-II enzyme that transfers an acetyl group to toxicants or drugs, enhancing their excretion. Patients are characterized as "slow" or "fast" acetylators, depending on the activity of their NAT2 (Table 2Go) (Miller et al., 2001Go). Slow acetylators will have more side effects from active drugs due to the accumulation of the drug in their system. However, if a drug needs to be metabolized for activity, fast acetylators will have increased side effects, as they will have higher levels of metabolites in their system.


View this table:
[in this window]
[in a new window]
 
TABLE 2 Main Mutations in NAT2
 
Slow acetylators are predisposed to bladder and prostate cancer caused by exposure to aromatic amine carcinogens found in tobacco smoke, overcooked meats, and the environment (Feng et al., 1999Go; Jiang et al., 1999Go). Increased susceptibility of slow acetylators to cancer is due to decreased NAT activity. Failure of NAT to N-acetylate carcinogens results in N-hydroxylation by CYP1A2 (Fig. 1aGo), which competes with NAT for the substrates. Fast acetylators do not have this cancer risk because their NAT, with higher activity, will outcompete CYP1A2 and N-acetylate the aromatic amines (Fig. 1bGo).



View larger version (16K):
[in this window]
[in a new window]
 
FIG. 1. (A) The effects of amines on slow acetylators. Slow acetylators have a greater carcinogenic effect with aromatic amines. NAT in slow acetylators has a lower activity for the substrates. In aromatic amines, this means that CYP1A2 N-hydroxylates the bulk of the aromatic amines, which will then be O-acetylated by NAT, activating the carcinogen. The pathway works the same for heterocyclic amines, but, since slow acetylators have less active NAT, less of the metabolite is O-acetylated compared to fast acetylators, activating the carcinogen. (B) The effects of amines on fast acetylators. Fast acetylators have a higher risk of colon cancer due to heterocyclic amines. Fast acetylators have very little N-hydroxylation by CYP1A2 on aromatic amines because their NAT enzyme has enough activity to outcompete CYP1A2 for the substrate. These acetylators have a higher risk with heterocyclic amines because these compounds are poor substrates for NAT. Therefore, they are N-hydroxylated by CYP1A2 and rapidly O-acetylated by NAT in fast acetylators because the O-acetylation makes the compound a good substrate for NAT.

 
Fast acetylators have an increased risk of colon cancer because they activate the procarcinogens rapidly (Ishibe et al., 2002Go). Heterocyclic amines, which are found in well-done meats (Hein, 2002Go), are poor substrates for NAT (Jaanson et al., 2002Go). Therefore, even if a person is a fast acetylator, CYP1A2 will outcompete NAT, and N-hydroxylate the heterocyclic amine (Fig. 1bGo).

DNA Repair and Metabolizing Enzymes
Not all polymorphisms associated with genetic susceptibility to pharmacological agents involve the P450 systems. Another system associated with susceptible polymorphisms involves DNA repair and nucleotide-metabolizing enzymes. DNA repair enzymes correct mutations in DNA, while nucleotide-metabolizing enzymes break down analogs of nucleotides. One cancer treatment protocol involves giving patients nucleotide analogs, which inhibit DNA synthesis by preventing the elongation of the nascent DNA strand. Therefore, these drugs are toxic to rapidly dividing cells, including tumor cells. Identification of polymorphisms in the enzymes required for metabolizing anti-cancer drugs are therefore of considerable interest.

Thiopurine methyltransferase (TMPT) is an enzyme that is important for nucleotide analog metabolism. TMPT metabolizes the active form of the anti-cancer drugs 6-mercaptopurine (6-MP) and thioguanine, both of which are used to treat acute lymphoblastic leukemia (ALL), and the immunosuppressant azathioprine (Pirmohamed and Park, 2001Go). Initially, these drugs are metabolized into thioguanine nucleotides, the active compounds that inhibit DNA synthesis. These active compounds are subsequently metabolized by TMPT, allowing for excretion by the kidney. Multiple mutations that reduce TMPT activity have been identified; many are amino acid substitutions that result in the rapid proteolysis of the enzyme (Krynetski and Evans, 1998Go). Ten percent of Caucasians and African-Americans are heterozygous for this enzyme, and 1 in 300 patients is homozygous null (Pirmohamed and Park, 2001Go). With low TMPT activity, patients only need 6–10% of the normal dose of the cancer treatment for efficacy (Relling et al., 1999Go). With a normal dose, these patients have an increased risk of potentially life-threatening hematopoietic toxicity. While low TMPT activity can cause serious side effects, studies of children with ALL have shown that higher levels of thioguanine in their system increase the chance of remission (Relling et al., 1999Go). Therefore, those with lower TMPT activity have a higher survival rate, if they can survive the initial treatment. The inverse is true for those having high TMPT activity, resulting in resistance to drug toxicity but a decreased incidence of remission. These effects can also be seen with azathioprine, used after tissue transplants to suppress the immune system. Those with higher TMPT levels have a higher chance of graft rejection, since they need a higher dose of azathioprine for equivalent efficacy (Krynetski and Evans, 1998Go). The role of TMPT in the metabolism of environmental and endogenous substrates is undefined and currently under investigation.

Transmembrane Receptors
Receptors, which are responsible for numerous cellular functions, are also susceptible to polymorphisms. Many receptors are gateways into a cell, allowing in necessary nutrients and components. Receptors are often the start of signaling pathways, activated by the binding of their ligands. Therefore, mutations affecting the activity of a receptor can be deleterious. For example, the ryanodine receptor is known to have many polymorphisms (McCarthy et al., 2000Go).

Mutations in the ryanodine receptor are responsible for approximately 50% of the incidences of malignant hyperthermia (MH) (Pirmohamed and Park, 2001Go). Patients with such a mutation can develop MH when given halogenated anesthetics or non-depolarizing muscle relaxants to prepare for surgery. MH, caused by a dysregulation of calcium in the skeletal muscle, results in increased body temperature, tachycardia, lactic acidosis, and respiratory acidosis, which can be fatal. In skeletal muscle, the ryanodine receptor causes calcium release from the sarcoplasmic reticulum into the myoplasm, initiating muscle contractions. More than 16 mutations have been found in this receptor (McCarthy et al., 2000Go). The best-characterized mutation, R614C, causes abnormal calcium inactivation, hypersensitivity to activating ligands (halothane), hyposensitivity to inactivating ligands (calmodulin), and a three-fold higher maximum calcium peak on release. Therefore, patients with this mutation have a hypersensitive ryanodine receptor, causing excessive calcium release and muscle contractions when activated by halothane. These mutations are rare, with familial genetic predisposition. Once discovered, these mutations do not cause many problems, since other anesthetics can be used. There is no known phenotype for this receptor other than MH; therefore no physiological problems are known except by man-made agents (McCarthy et al., 2000Go).

There is a wide variety of polymorphisms that can be implicated in genetic susceptibility. The difficulty in discovering these susceptibilities lies in the wide variability of patient response. Humans, unlike laboratory animals, are very heterogeneous, and most effects are probably not due to one simple polymorphism, but the combined effects of many. Still, much has been discovered, and this summary only covers a tiny fraction of that knowledge. With what is known, hospitals could soon start genotyping patients, if not to tailor their prescriptions, to at least check for dangerous mutations that are common in the patient’s ethnicity.

Age-Related Susceptibility

Age is an important factor that can influence an individual’s metabolic processes and their ability to eliminate a toxicant. There are many factors that differ between individuals of different ages and periods of development and influence susceptibility to toxic insult. There are many examples in toxicology that show either increased or decreased susceptibility related to age, demonstrating that generalizations cannot be made. The toxicity depends on the chemical under consideration, the period of life of the individual, as well as many other factors.

Developmental and Childhood Susceptibility
The fetus, neonate, and child are often more sensitive than adults to environmental toxicants. The nature of the fetus as a target for toxicity is difficult to pinpoint because of rapidly occuring developmental changes. Although the development of the fetus is a continuum, critical developmental windows can be identified. Fetal tissues are specific targets for toxicity because the cells are differentiating, proliferating, and migrating (Rodier, 1994Go). Rapidly dividing cells confer increased susceptibility due to the fact that there are more opportunities for the cell to inaccurately copy DNA, which can have deleterious effects on the individual. Also, mRNA and proteins are being rapidly synthesized, leading to increased opportunities for error in the transcriptional and translational processes, which can result in mutated or nonfunctional proteins. During fetal development, cell migration and differentiation are vital for establishing correct cell function, and toxic agents are capable of interfering in this process. Migration and differentiation either do not occur or occur at remarkably lower rates in adults. Therefore, a mature adult may be resistant to certain toxic insults while the fetus is highly susceptible.

The toxicokinetics and toxicodynamics of the developing individual can increase the susceptibility to toxic insult. During the fetal period, the major pathway of chemical absorption is through the placenta. It was once believed that the placenta served to protect the fetus from maternal exposure, but the thalidomide catastrophe significantly changed this way of thinking (McBride, 1961Go). Thalidomide was introduced in 1956 to aid sleep and relieve nausea and vomiting in pregnancy. After maternal use of thalidomide, there was a large increase in newborns with rare limb malformations including phocomelia (reduction of the long bones of the limbs) or amelia (absence of limbs). Over 12,000 children were born with skeletal abnormalities. The exact mechanism underlying the formation of these abnormalities is unknown, although thalidomide has been found to have a broad range of effects on angiogenesis, immune function, cytokine secretion, and cell proliferation (Parman et al., 1999Go).

Following this incident, researchers became aware that numerous compounds readily cross the placenta and may affect the fetus. This demonstrated that the placenta was a potential site of toxicity and has an important role in influencing the exposure of the fetus to toxic chemicals. The placenta does this by regulating blood flow, offering a semi-permeable barrier to some chemicals, and by metabolizing chemicals. The placenta functions as a lipid membrane that allows the transfer of chemicals from the mother to the fetus. The passage of most chemicals across the placenta occurs by passive diffusion and depends on the characteristics of the specific chemical, such as lipid solubility, size of the chemical, plasma protein binding, and the ionization state. For example, the ionization state of weak acids allows them to easily cross the placental barrier. Numerous chemicals have been shown to exert toxicity by crossing the placenta, making the fetus especially susceptible to toxic insult.

Lead poisoning is a well-researched example of how exposure to a toxicant during a critical period of development can affect an individual (Goyer, 1996Go). The primary route of exposure in the general population is through food. However, for children it has been found that a major source of exposure was the hand-to-mouth behavior used as a way of exploring an environment. These toxic effects, ranging from subtle and undiagnosed to overt, involve many organ systems and biochemical activities. The effects depend on the developmental stage of the child at the time of exposure. A correlation between lead exposure and problematic behaviors has been identified in children as young as 2 years old. In older children, associations have been made between lead exposure and negative behaviors including aggression, delinquency, and social problems.

Many mechanisms have been proposed to explain the increased susceptibility of young children to lead toxicity. Lead may disrupt interneuronal programming, thus modifying neuronal circuitry; it may also influence glial differentiation and migration, affecting the overall structure of the brain. Lead has also been found to influence the concentrations of dopamine and norepinephrine in neurons and synapses, thereby affecting their signaling properties.

Another mechanism by which lead causes neurotoxicity is by interfering with the role of calcium, which is important for proper brain function. Lead affects normal calcium homeostasis and uptake by calcium membrane channels and acts as a substitute in calcium-sodium ATP pumps, blocks the entry of calcium into nerve terminals, and inhibits calcium uptake in mitochondria in the brain. All of these effects lead to a decrease in energy production and therefore decrease the ability of the brain to perform vital functions. The most important action of lead involving calcium is the interruption of second messenger signaling. Calcium signals are received by calcium receptor proteins including calmodulin and protein kinase C (PKC). Calmodulin is a sensor for the intracellular concentration of calcium. Lead disrupts calmodulin by displacing bound calcium ions. PKC signaling influences cell division and proliferation, cell–cell communication, and organization of the cytoskeleton. All of these processes are critical to the developing brain. Disruption of calmodulin and PKC by lead affects the development of the blood brain barrier, increasing susceptibility to lead toxicity (Needleman, 1994Go).

Another example of a developmental toxicant is the organophosphate pesticide chlorpyrifos (CPF). CPF is used extensively in agricultural and domestic settings and has replaced many other organophosphate pesticides because of its chemical stability and reported lower toxicity. The systemic toxicity and neurotoxicity of CPF has been widely studied in adults; however, the amount of research on the effects of CPF in the developing organism is far less (Slotkin, 1999Go). This is of particular concern because children come into contact with contaminated surfaces more than adults, due to repeated hand-to-mouth contact at a young age, thus increasing their susceptibility to CPF toxicity (Fenske et al., 1990Go). One of the more desirable characteristics of CPF as a pesticide is its persistence, which reduces the need for repeated applications. However, the increased persistence of CPF could cause increased duration of exposure, leading to increased toxicity.

CPF, through its active metabolite CPF oxon, inhibits the enzyme cholinesterase and prevents the breakdown of acetylcholine, leading to cholinergic hyperstimulation. Acetylcholine plays an important role in the development of the central nervous system by regulating the replication, differentiation, growth, death, and migration of target cells (Hohmann et al., 1988Go). Therefore, measurements of the activity of cholinesterase in exposed fetal or neonate models have been determined to be the standard index of developmental toxicity. CPF has been found to be approximately 100 times more toxic to neonatal rats than to adult rats. Some of this increased toxicity can be explained by the differential rates of development of cholinesterase when compared to the development of detoxifying enzymes, and also by an increased accumulation of CPF in the developing brain when compared with adult accumulation. However, the inhibition of cholinesterase does not explain all of the neurotoxic effects seen with CPF exposure. CPF has direct effects on cholinergic receptors, causing the desensitization of nicotinic receptors (Huff and Abou-Donia, 1995Go) and the inhibition of muscarinic actions (Katz et al., 1997Go). CPF can also interact with downstream signaling cascades such as the adenylyl cyclase pathway (Huff et al., 1994Go). Adenylyl cyclase and cyclic AMP help to control cell replication and differentiation. Therefore, a disturbance of this pathway by CPF would be expected to have deleterious effects on brain development. Other neurotoxic actions of CPF include the inhibition of DNA synthesis (Dam et al., 1999bGo), interference with DNA binding of transcription factors involved in cell differentiation (Crumpton et al., 1999Go), a reduction in RNA synthesis during differentiation that impairs neurite outgrowth (Song et al., 1998Go), and the disturbance of neurotransmitter systems during critical developmental periods (Dam et al., 1999aGo; Qiao et al., 2003Go; Slotkin et al., 2001Go, 2002Go). Many of these neurotoxic effects during development occur at doses that lie well below the threshold for systemic toxicity, and these effects are not seen in adults. Therefore, it is clear that the fetus, newborn, and child are more susceptible to the neurotoxic effects of CPF, and measures should be taken to monitor exposure to CPF closely.

Elderly Susceptibility
Similar to developing children, elderly individuals possess increased susceptibility to toxic insult. In the elderly, the consequences of neuronal damage caused by toxic insult may be more severe because a substantial loss of neurons may have already occurred. In this situation, a further loss of neurons caused by a chemical may be critical in loss of function (Lotti, 2002Go; Troen, 2003Go). Enhanced effects caused by a toxic insult in the elderly may be due to changes in renal function, hepatic extraction, plasma protein binding, and volume of distribution for hydrophilic compounds. Phase-I enzyme activity is usually decreased in elderly patients, affecting the rate of elimination, causing certain xenobiotics to be present in the body for a longer period of time, and possibly increasing the toxic effect. Part of the aging process is a decrease in cardiac output (Dybing and Soderlund, 1999Go), which affects the distribution and rate of blood flow throughout the body. Therefore, delivery of chemicals to the liver and kidney is slower, and the inactivation of the toxicants is delayed. When chemicals arrive at the kidney for excretion, the process may also be slowed due to the fact that a large percentage of the elderly have decreased renal function, resulting in xenobiotics persisting longer in the elderly population.

The toxic effects of lead have already been discussed for the developing individual. However, elderly people also have an increased susceptibility to lead toxicity due to many of the factors described above (Vig and Hu, 2000Go). Elderly adults may have sustained exposure to sources of lead from both occupational and nonoccupational sources. Most people are exposed to lead through respiratory and gastrointestinal routes. Once in the body, lead is carried in the blood, where it can remain for days to weeks. Lead can stay in soft tissues for months, while lead incorporated into bone has a half-life of over 20 years. Lead deposited in bone may not be directly harmful, but as it reenters the bloodstream through bone resorption, which is often associated with osteoporosis, harmful effects can result (Tsaih et al., 2001Go). Even low blood lead levels can lead to renal impairment, hypertension, and cognitive disorders. Therefore, elderly patients need to be monitored for lead toxicity, especially individuals who have been exposed to high levels of lead throughout their lives.

When considering the susceptibility of individuals to toxic exposure, the stage of life needs to be an important consideration. Age may be a determining factor for the risk of toxicity and how the toxicity may affect life later. Lead poisoning is a well-researched example of how environmental toxicant exposure during development or aging influences the risk of toxicity. Behavioral, physiologic, biochemical, social, and many other factors interact with an individual to result in age-related toxicological susceptibility.

Gender-Related Susceptibility

A genetic polymorphism is not the only means by which an individual’s genetic makeup influences their response to toxicants. Indeed, our gender, which is genetically predetermined, sets the stage for individual differences in toxicant susceptibility (see Table 3Go). Although significant physiological changes occur during development, thus making age an important factor when considering gender-specific susceptibilities, hormones regulate biological function throughout the course of our life.


View this table:
[in this window]
[in a new window]
 
TABLE 3 Examples of Endocrine Disrupting Chemicals
 
At first glance, the essential bodily functions of males and females are the same; however, increasing amounts of research are pointing toward crucial developmental, hormonal, and metabolic differences. Proper development and function of the reproductive system is essential for homeostatic maintenance of a species. The basic reproductive unit is composed of the mother, father, and offspring (Hoyer, 2001Go). It is therefore necessary, when considering toxic effects, to consider all three aspects of this unit. Typically, toxicity to the reproductive system in males is accompanied by reduced sperm production, impotence, and/or infertility. Reproductive toxic response among females often coincides with acyclicity, delayed conception, early menopause, or troubled pregnancies. The effects of chemicals on the reproductive unit are often difficult to define by long-term health consequences (Melnick et al., 2002Go). Therefore, many studies choose to look at biological changes in reproductive endpoints versus adverse effects (Daston et al., 1997Go).

The mechanism of action of a toxicant often indicates if there will be a greater toxic susceptibility for one developmental stage or gender. Research has found that a large number of toxicants have the ability to interact with hormone transport proteins, disrupt hormone metabolism, or bind to the hormone receptor. Through these actions the toxicant either mimics endogenous hormones or blocks their action (Hoyer, 2001Go). These chemicals, many of which have similar chemical structures to endogenous hormones, disrupt both the endocrine systems as well as alter the cognitive functions of animals. Agents that act on the endocrine system can be divided into a number of categories including, but not limited to: estrogenic, anti-estrogenic, androgenic, and anti-androgenic. Some common endocrine disrupters include phthalates; alkylphenolic compounds; polychlorinated biphenyls (PCBs); polychlorinated dibenzodioxins; bisphenol A; and heavy metals, including lead, mercury, and cadmium.

Estrogenic/Anti-Estrogenic Compounds
Estrogenic compounds, which have been the most widely studied of the endocrine disruptors, are defined as those chemicals that produce responses qualitatively similar to those produced by 17ß–estradiol, an endogenous estrogen (Daston et al., 1997Go). Estrogens play a role in the expression of specific genes and production of several vital hormones. Through pleiotropic actions, estrogens influence cell proliferation, cell differentiation, and tissue organization. In humans, estrogen in the bloodstream binds to sex hormone binding globulin (SHBG) or remains in the unbound state. Free estrogen binds to estrogen receptors (ER) in various tissues. ERs are nuclear receptor proteins and are located in a target cell within the cytosol. When estradiol, or another estrogen-like ligand, encounters a potential target cell, the chemical diffuses through the cell membrane and binds to the ER. Upon binding, the ER is activated, and the subsequent hormone–receptor complex is translocated into the nucleus. Once in the nucleus, a dimer of the ligand-bound ER binds to the estrogen response element (ERE) on the DNA and recruits the binding of additional co-regulatory proteins. These co-regulatory proteins have the potential to either stimulate or inhibit transcription (Fang et al., 2000Go; Witorsch, 2002Go).

Due to the complex role that estrogen plays in the body, it has been extremely difficult for researchers to elucidate the site and mechanism of environmental estrogenic compounds. Exogenous estrogenic compounds may elicit biological effects through the activation of intracellular estrogen receptors and changes in gene expression characteristics of endogenous estrogen (Degen et al., 2002Go). Toxicants may also indirectly produce an estrogenic response by increasing estrogen synthesis, facilitating the binding of estrogen to the receptor, or changing the rate of estrogen degradation. Anti-estrogenic compounds often act by preventing exogenous estrogens from interacting with their receptors. Numerous studies investigated the impact of low-level exposure to various estrogenic and anti-estrogenic compounds. For many compounds, the information is inconclusive, with different labs giving different results. The discrepancies between positive and negative experiments can be attributed to numerous factors, including differing observed endpoints; different data analysis; and, in animals studies using different diets with differing background levels of phytoestrogens, differences in strains of animals used, dosing regimens, and animal housings (Melnick et al., 2002Go). There have been a few compounds where definite statements can be made regarding the toxic effects of low-level exposure. The next few paragraphs will include information on some of the well-characterized compounds as well as those whose effects are still rather debatable.

Diethylstilbestrol (DES), an estrogen agonist with a higher affinity for the ER than endogenous estradiol, is partially responsible for calling attention to the potential hazards that some chemicals may have on human reproductive health. In the mid–twentieth century, pregnant women with a history of miscarriages were told to take DES to stabilize their pregnancy. Severe reproductive effects were observed in the offspring of these women. Estrogens play a significant role in the development of the male reproductive system; however, the mechanism by which this action occurs is still unknown. Researchers have found that estrogen receptors are widely expressed in the male reproductive tract and that overexposure of the fetal/neonatal male to exogenous estrogens, such as DES, results in major abnormalities and life-long dysfunction (Williams et al., 2000Go). Studies have shown that estrogen exposure causes various reproductive tract abnormalities through the reorganization of tissue composition, an effect that can be observed in the prostate gland (Pylkkanen et al., 1991Go, 1992Go). Another mechanism of estrogen toxic action involves alterations in, or the induction of, abnormal gene expression such as was observed with c-myc in the prostate (Pylkkanen et al., 1993Go).

Researchers have looked at the abnormal induction of progesterone receptor in the reproductive tract of male rats neonatally exposed to the estrogenic compound DES. Overexposure of neonatal rats to DES produced a marked, dose-dependent expression of the progesterone receptor in the stromal tissue of the male reproductive tract. This is not an effect that is usually observed in male rats. The authors admit that this information does not constitute direct proof of male feminization; however, it does suggest that inappropriate exposure to estrogenic compounds alters the structure of the male reproductive system (Williams et al., 2000Go).

Toxicology studies on DES have also been performed in humans. In one such study, pregnant women were dosed with DES throughout the second and third trimesters. There was no postnatal exposure to DES. The male offspring of women exposed to DES had increased incidences of genital malformations, including testicular abnormalities and hypoplastic testes. There was no decrease in fertility observed in these men, whose ages ranged from 38 to 41(Wilcox et al., 1995Go).

In recent years, numerous studies have been published regarding the disparities between genders with regard to susceptibility to a variety of neurological disorders. Parkinson’s disease, ischaemia, neurotrauma, and drug-induced injuries all seem to impact females less than males. These differences are significantly reduced when comparisons are made with post-menopausal women who are not receiving hormone replacement therapy. Additionally, studies have found that parkinsonian symptoms are more severe in women during times of low estrogen in the menstrual cycle. This information suggests that estrogen and estrogenic compounds may act as a preventative barrier for neurodegenerative damage. To study this effect and determine if estrogen would have a similar effect on men as it does on women, a series of rats were treated with a neurodegenerative toxin, 6-hydroxydopamine (6-OHDA). There was a significant difference in the magnitude of toxic effects seen in the females versus the males. Overall, gonadal hormones in the males exacerbated the effects of 6-OHDA, while it was decreased in the females. Males undergoing a gonadectomy had reduced toxic effects, whereas females undergoing the same procedure experienced increased neuronal degeneration. Treatment of male rats with 6-OHDA and estrogen increased the degeneration of dopamine-producing cells. Numerous studies on other toxins have looked at this effect, and the results are inconclusive. This research concludes that not only does estrogen have a protective effect against some neurological diseases, but also sex hormones in both males and females can influence neurotoxicity (Murray et al., 2003Go).

Androgenic/Anti-Androgenic Compounds
Researchers are finding increasing numbers of developmental tract abnormalities in a variety of organisms, including humans. Some studies have suggested that there is an overall decrease in male sperm production in various regions around the world (Daston et al., 1997Go). Although this is a much-debated issue, the discussion has led to an increased interest in the androgenic/anti-androgenic properties of environmental chemicals. The anti-hormone action of most anti-androgenic chemicals is classified as either type I antagonists, which bind androgen receptors (AR) and prevent DNA binding, or type II antagonists, which induce DNA binding but do not initiate transcription. ARs, found in high concentrations in the male reproductive tract, are responsible for a multitude of functions, including sexual differentiation in the developing fetus and androgen action in post-embryonic life. A majority of anti-androgenic chemicals have a low affinity for the AR receptor and primarily act as type I antagonists. Researchers have found that these antagonists are responsible for the formation of heterodimers with the AR. Inhibition of AR–DNA binding occurs for a number of reasons, including increased AR degradation due to improper receptor conformation or increased susceptibility of receptor toward proteases, inhibition of proper AR dimerization, and failure of mixed-ligand AR dimers to bind DNA.

Studies have found that the administration of androgenic compounds has a significant impact on rodent sexual development. Females adopt masculine features; have an increased anogenital distance (AGD); and exhibit abnormalities of the uterus, vagina, and mammary glands. Males have small hypospermatogenic testes and reduced epididymal weights (Gray et al., 1999Go). Vinclozolin, a dicarboximide fungicide, has been widely used in Europe and the United States as a means of protecting herbs, fruits, and ornamental plants from fungi damage (Wolf et al., 2000Go). Not only is vinclozolin a known antiandrogenic compound, but it has provided researchers with significant insight into the roles that metabolites and dosing regimens can have on toxicity (Kelce et al., 1994Go; Wolf et al., 2000Go).

The phthalates represent another class of antiandrogenic compounds, and, although they have been extensively studied, little is known about their mechanism of toxic action. Di(n-butyl) phthalate (DBP) is one of the primary components of adhesives, printing inks, and aerosols (Mylchreest et al., 2000Go). Numerous studies have indicated that DBP exposure resulted in alterations in male reproductive development and function. In order to establish a NOAEL, studies were undertaken in which gestating rats were incrementally dosed with DBP. At dosages of 100 mg/kg/day, male offspring exhibited feminization, including decreased AGD and retained areolas. At 500 mg/kg/day, absent or partially developed epididymis, vas deferens, seminal vesicles, and ventral prostate were observed. In 110-day-old males, the weights of the testis, epididymis, dorsolateral and ventral prostates, seminal vesicles, and levator anibulbocavernosus muscle were decreased at 500 mg/kg/day. Overall, the NOAEL for DBP was determined to be 50–100 mg/kg/day. No analogous effects were observed in female offspring. This is likely explained by the anti-androgenic activity of DBP. During fetal development, androgenic influences are significantly more vital for the sexual differentiation of males than females (Mylchreest et al., 2000Go).

Other Endocrine Disruptors
The last class of endocrine disruptors discussed in this review will be those that influence the thyroid. Thyroid hormones regulate growth and development, cellular metabolism, and the rate of basal metabolism. Key hormones involved in thyroid function are thyroxine (T4), 3,5,3'-triiodothyronine (T3), and the thyroid stimulating hormone (TSH). TSH regulates the synthesis and stimulates the release of T3 and T4 into circulation. In peripheral tissues, T4 is converted into the active T3, which is then capable of stimulating gene transcription. Numerous studies have been performed outlining the significance of T3, T4, and TSH on various developmental stages in animals and humans (EPA, 1997Go).

Given the broad influence of the thyroid, researchers have naturally been interested in the effects that various chemicals could have on its function. One study involved the exposure of immature female rats to the environmental toxicant 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). The rats exposed to single doses of TCDD showed increased concentrations of luteinizing hormone (LH) and a decrease in ovulation. Interestingly, the expected decrease in estradiol that usually accompanies an LH increase was absent. This suggests that direct effects on the ovary caused alterations in the hypothalamic-pituitary regulation of ovulation (EPA, 1997Go).

It is clear that exposure to a variety of chemicals, both at low and high doses, can influence the complex endocrine system. However, significant work is still necessary. Researchers need to carefully elucidate the mechanisms by which these chemicals work. Therefore, it is important to determine the cellular target and the intracellular pathways affected. Development of testing schemes sensitive to estrogenic and androgenic effects is necessary to clearly determine the deleterious effects of endocrine disruption.

Disease-Related Susceptibility

The previous examples have clearly indicated the existence of subpopulations that are vulnerable to toxicant exposure due to their genetics, developmental status, and gender. However, a commonality in all of these subpopulations is the existence of disease. Diseases themselves have the ability to influence toxicity and therefore warrant consideration in this review. The following will address a few examples in which diseases cause increases in toxicant susceptibility and therefore identify the need for proper identification of the disease state before designating an individual’s risk to toxicant exposure.

Of the many factors affecting an individual’s health, the immune system status is of great importance. Alterations in an individual’s immune system result in an increased susceptibility to new infections as well as a decreased ability to combat existing pathogens. In addition, the immune system has been implicated in defending against carcinogenesis by monitoring the formation of abnormal cells and subsequently eliminating them. Because of this, both preexisting diseases and exposure to toxicants that result in immune suppression are important considerations when assessing an individual’s risk.

Human Immunodeficiency Virus (HIV)
Human immunodeficiency virus is one of the most well-studied diseases in which infection results in decreased immune function. Consequently, when coupled with HIV, several toxicants result in more extreme conditions than HIV alone. Several studies have recently shown that opioids, including heroin, have the potential to advance HIV symptoms, including a more rapid progression toward HIV-associated dementia (Nath et al., 2002Go). Methadone, an opioid used to treat heroin and morphine addiction, induces the expression of CCR5, the receptor responsible for HIV entry into lymphocytes, resulting in promotion of HIV infection (Li et al., 2002Go; Miyagi et al., 2000Go). In addition, methadone was shown to enhance viral replication (Li et al., 2002Go) as well as inhibit DNA damage repair (Madden et al., 2002Go). As a result, mutations within the rapidly replicating virus will be maintained, conferring resistance to drugs used to treat the disease. Clearly, opioid use has the ability to increase HIV viral load, resulting in a more rapid progression of the disease.

Opioids have also been shown to increase HIV-associated dementia through a mechanism independent from the increase in viral load. Chronic opiate use has been known to alter dopaminergic signaling. As with Parkinson’s disease, these alterations lead to an increase in the formation of reactive oxygen species and may sensitize neurons to the effects of HIV, synergizing to result in neuronal damage. Studies indicate these synergistic effects are mediated through the opioid receptor and may result in mitochondrial toxicity through Akt, PI-3 kinase, and caspases (Nath et al., 2002Go). Because needle sharing during intravenous heroin use is a risk factor for contracting HIV, the effects of opioid use on the progression of HIV and HIV-associated dementia are of particular concern.

Exposure to carcinogens may also be of concern to those suffering from HIV infection. A reduction in cell-mediated immunity has been seen in several malignant diseases, including melanoma, colorectal cancer, and prostate cancer, implying that compromised immune function may result in carcinogenesis (Dalgleish and O’Byrne, 2002Go). While many of the tumors associated with HIV are likely a result of reduction in the ability of the immune system to monitor tumor formation, other factors have been implicated in oncogenesis in the presence of HIV. The HIV-1 tat protein, which is involved in growth promotion, angiogenesis, and cell survival, has been proposed to contribute to tumor progression (Albini et al., 1995Go; Ensoli et al., 1994Go; Zauli et al., 1993Go). Given their predisposition to the development of malignant conditions and the tumor promoting properties of HIV-1 tat, patients suffering from HIV may be more susceptible to exposure to carcinogens. Diethylnitrosamine, a hepatic carcinogen, has been shown to increase both liver tumors and preneoplastic lesions in mice that express the HIV-1 tat gene, indicating susceptibility to chemical carcinogenesis with HIV infection (Altavilla et al., 2000Go).

Hepatitis C Virus (HCV)
Hepatitis C virus has also been associated with carcinogenesis, a consequence that is exacerbated by concurrent ethanol exposure. While HCV generally runs a fairly benign course in most people, the development of severe liver damage, cirrhosis, or hepatocellular carcinoma (HCC) occurs in some patients. Several studies indicate that alcohol has the ability to enhance the progression of HCV toward liver damage (Danta et al., 2002Go; Westin et al., 2002Go). Studies show that the relative risk for development of cirrhosis, a precursor to HCC, was more than additive in patients with HCV who drink alcohol compared to those with just HCV or ethanol exposure (Donato et al., 2002Go; Vento and Cainelli, 2002Go). In fact, there is a significant increase in the incidence of HCC in alcohol-consuming populations, likely a result of an increase in incidence and severity of liver fibrosis and cirrhosis (Danta et al., 2002Go; Westin et al., 2002Go).

Immune suppression is an important consequence of chronic ethanol use. Reports show that chronic ethanol use inhibits T-helper and T-lymphocyte responses to HCV (Geissler et al., 1997Go), as well as T-cell proliferation (Szabo et al., 2001Go), resulting in an increased viral load. Ethanol has also been shown to increase the quasispecies complexity of HCV (Takahashi et al., 2001Go). Therefore, the immune system has a more difficult time combating the increased number of mutated viruses. When an HCV-infected patient consumes alcohol, the ability of the body to defend against the infection is reduced. Consequently, the infection is able to progress at a more rapid pace toward liver damage and HCC.

In addition to the properties of ethanol alone impacting the disease state, common signaling pathways between HCV and ethanol may be influenced, resulting in synergism. HCV is an RNA virus encoding several proteins, including the HCV core protein. The HCV core protein signals through several intracellular cascades, including activator protein-1 (AP-1) and nuclear-factor kappa B (NF-{kappa}B) (Kato et al., 2000Go). It is the activation of these pathways that is thought to lead to liver injury, cirrhosis, and HCC. Together, both NF-{kappa}B and AP-1 are able to promote cell proliferation in response to the HCV core protein, and both have been shown to be activated in HCC (Liu et al., 2002Go). From human in vivo studies, it is known that AP-1 and NF-{kappa}B activation precedes the development of HCC, indicating that activation may be a cause of HCC and not a consequence (Liu et al., 2002Go). Alone, both ethanol and the HCV core protein have been shown to activate NF-{kappa}B. When combined, the metabolism of ethanol potentiates the activation of NF-{kappa}B by the HCV core protein (Kim et al., 2001Go), indicating a possible mechanism of synergism between ethanol and HCV in the formation of HCC.

It is clear that a preexisting disease increases susceptibility to the effects of a toxicant. However, what is often overlooked is the impact that a toxicant can have on an individual bordering on disease state. Toxicant exposure may result in the manifestation of the disease in a patient who may not otherwise have ever reached the disease state. Parkinson’s disease, a disease with an unknown etiology, may be resultant of such a phenomenon.

Parkinson’s Disease (PD)
In most cases of Parkinson’s disease, a 25% reduction in mitochondrial complex I activity has been noted (Greenamyre et al., 2001Go). As complex I is the target of several mitochondrial toxins, including the pesticide rotenone, exposure to such toxins results in the PD phenotype (Betarbet et al., 2000Go). Should an individual have a lower activity of complex I and be on the verge of PD, exposure to chemicals such as rotenone may cause the manifestation of PD symptoms.

Parkinson’s disease is thought to result from oxidative damage to dopaminergic neurons in the substantia nigra. Deficiency in mitochondrial complex I, which is responsible for transferring electrons and creating the electrochemical gradient that drives ATP synthesis, is associated with an increase in the formation of reactive oxygen species, suggesting a mechanism by which complex I deficiency leads to PD (Sipos et al., 2003Go). In the mitochondria, when the electrochemical gradient is compromised, the cell is unable to synthesize ATP, ultimately resulting in cell death. Hydrogen peroxide, which is abundantly produced in dopamine neurons as a by-product of dopamine metabolism, is unable to alter the electrochemical gradient. However, when coupled with the partial inhibition of complex I, hydrogen peroxide causes a gradual decrease in the electrochemical gradient and ultimately leads to the death of the cell (Chinopoulos and Adam-Vizi, 2001Go). Therefore, a dopamine neuron with a deficiency in complex I activity may suffer cell death upon the formation of hydrogen peroxide. Pesticides such as rotenone inhibit complex I and may therefore cause a more rapid development of PD in an individual with compromised mitochondrial function.

Rotenone, a common pesticide, is a known inhibitor of complex I. An individual with a deficiency in complex I, and therefore on the verge of PD, may develop PD as a consequence of rotenone exposure. In addition to rotenone, chronic exposure to the pesticide manganese ethylene-bis-dithiocarbamate (maneb) has been shown to cause Parkinsonism in humans. Recent work indicates that maneb, like rotenone, may alter mitochondrial respiration via the inhibition of complex III in the electron transport chain in vitro (Zhang et al., 2003Go). Humans with polymorphisms in complex III may be susceptible to the development of PD as a result of chronic maneb exposure.

In addition to such polymorphisms, exposure to both maneb and the pesticide paraquat results in increased susceptibility to the development of PD. Combined exposure to these two pesticides results in increased dopaminergic cell loss within the striatum, as well as increases in dopamine turnover and release (Thiruchelvam et al., 2000Go). Little is known about the mechanism by which these two pesticides synergize, although one may speculate that oxidative stress and the inhibition of mitochondrial respiration may be involved. The interaction of these toxicants, which already synergize to form the PD phenotype, may be exacerbated by a preexisting disease condition or polymorphism resulting in the manifestation of PD in cases in which it would not occur under normal conditions.

Preexisting diseases, when coupled with exposure to a toxicant, may make an individual more susceptible to the effects of the toxicant. These few examples represent only a subset of the diseases that result in a predisposition to toxicity. Both immune status–dependent and immune status–independent mechanisms are involved in this predisposition. In determining an individual’s risk, it is of particular importance to define the disease status of the individual and take the proper precautions to reduce the occurrence of resultant adverse effects.

Conclusions

Human variability to toxicant exposure remains a concern both in identifying hazardous chemicals and assessing their risk to humans. Although genetics, age, gender, and disease state clearly influence toxic responses, these factors and their combinations have essentially been ignored in clinical trials and toxicity determinations. Currently, the use of uncertainty factors is intended to account for human variability. However, these uncertainty factors may not be adequate in the estimation of tolerable exposures.

Genetic polymorphisms, which are critical in drug and toxicant metabolism, have often been overlooked because testing is not cost-effective due to low occurrences of polymorphisms. This neglect has carried over to basic research, as cost and volume of animals are minimized and thereby decrease the ability to account for less recognized susceptibilities. Age has received considerably more attention than genetic polymorphisms with regard to both drug testing and toxicant evaluations. However, subtle effects associated with specific periods of development often go unstudied, perhaps due to the difficulty in identifying effects, as manifestations may occur many years after exposure. Toxicant exposure studies have also been skewed when considering gender-specific effects. Female animals, unless used in developmental or maternal studies, are often omitted from tests due to the inconsistencies associated with hormone cycling. This biases toxicant exposure levels toward one gender or the other. Disease state is especially difficult to address in toxicant and drug exposure studies, as the sheer volume of diseases is overwhelming. This is further complicated by the differences in disease etiology and progression between animals and humans. Because risk assessment models currently only evaluate healthy individual responses, those with chronic diseases must be given separate consideration.

While it is impossible to individually assess susceptibilities within a population, it is necessary for researchers, clinicians, and policy makers to address these issues. Possible actions include the improvement of basic research tools, recognition of the need for personalized treatments in clinical settings, and modification of risk assessment procedures to include the aforementioned factors, thereby allowing for more accurate toxicant evaluations.

These factors must be considered in defining uncertainty factors and identifying risk in conferring vulnerability to exposure on human populations. However, what is often neglected is the possibility that a given individual may exhibit multiple susceptibilities. Combinations of susceptibilities may underlie the manifestation of toxicity in cases in which human variability was accounted for during the risk assessment process. While it is impossible to individually assess susceptibility within a population and tailor policies to encompass the entire range of human responses, policy makers must consider the potential for a broad range of responses during risk assessment. Currently, the use of uncertainty factors is intended to account for human variability. However, given the incidents of toxicity demonstrated in this review, these uncertainty factors may not be adequate in the estimation of tolerable exposures. For future risk assessment, it may be necessary to assign more accurate uncertainty factors to account for such examples of human variability.

ACKNOWLEDGMENTS

The authors thank Dr. Richard DiGuilio and Dr. Theodore Slotkin for their helpful comments on the manuscript. The NIEHS Toxicology Training Grant T32 ES07031 provided support.

NOTES

All authors contributed equally to this work.

1 To whom correspondence should be addressed at Box 3412, Department of Psychiatry, Duke University Medical Center, Durham, NC 27710. Fax: (919) 681-3416. E-mail: edlevin{at}duke.edu. Back

REFERENCES

Aklillu, E., Persson, I., Bertilsson, L., Johansson, I., Rodrigues, F., and Ingelman-Sundberg, M. (1996). Frequent distribution of ultrarapid metabolizers of debrisoquine in an ethiopian population carrying duplicated and multiduplicated functional CYP2D6 alleles. J. Pharmacol. Exp. Ther. 278, 441–446.[Abstract]

Albini, A., Barillari, G., Benelli, R., Gallo, R., and Ensoli, B. (1995). Tat, the human immunodeficiency virus type regulatory protein, has angiogenic properties. Proc. Natl. Acad. Sci. 92, 4838–4842.[Abstract]

Altavilla, G., Caputo, A., Lanfredi, M., Piola, C., Barbanti-Brodano, G., and Corallini, A. (2000). Enhancement of chemical hepatocarcinogenesis by the HIV-1 tat gene. Am. J. Pathol. 157, 1081–1089.[Abstract/Free Full Text]

Anzenbacher, P., and Anzenbacherova, E. (2001). Cytochromes P450 and metabolism of xenobiotics. Cell Mol. Life Sci. 58(5–6), 737–747.[ISI][Medline]

Betarbet, R., Sherer, T. B., MacKenzie, G., Garcia-Osuna, M., Panov, A. V., and Greenamyre, J. T. (2000). Chronic systemic pesticide exposure reproduces features of Parkinson’s Disease. Nat. Neurosci. 3, 1301–1306.[CrossRef][ISI][Medline]

Burroughs, C. D., Bern, H. A., and Stokstad, E. L. R. (1985). Prolonged vaginal cornification and other changes in mice treated neonatally with coumestrol, a plant estrogen. J. Toxicol. Environ. Health 15, 51–61.[ISI][Medline]

Burroughs, C. D., Mills, K. T., and Bern, H. A. (1990). Reproductive abnormalities in female mice exposed neonatally to various doses of coumestrol. J. Toxicol. Environ. Health 30, 105–122.[ISI][Medline]

Chinopoulos, C., and Adam-Vizi, V. (2001). Mitochondria deficient in complex I activity are depolarized by hydrogen peroxide in nerve terminals: Relevance to Parkinson’s disease. J. Neurochem. 76, 302–306.[CrossRef][ISI][Medline]

Crump, K. S. (1984). A new method for determining allowable daily intakes. Fundam. Appl. Toxicol. 4, 854–871.[ISI][Medline]

Crumpton, T., Seidler, F., and Slotkin, T. (1999). Developmental neurotoxicity of chlorpyrifos in vivo and in vitro: Effects on nuclear transcription factor involved in cell replication and differentiation. Brain Res. 857, 87–98.[CrossRef][ISI]

Csanady, G. A., Oberste-Frielinghaus, H. R., Semder, B., Baur, C., Schneider, K. T., and Filser, J. G. (2002). Distribution and unspecific protein binding of the xenoestrogens bisphenol A and daidzein. Arch. Toxicol. 76(5–6), 299–305.[CrossRef][ISI][Medline]

Dalgleish, A., and O’Byrne, K. (2002). Chronic immune activation and inflammation in the pathogenesis of AIDS and cancer. Adv. Cancer Res. 84, 231–276.[ISI][Medline]

Dam, K., Garcia, S., Seidler, F., and Slotkin, T. (1999a). Neonatal chlorpyrifos exposure alters synaptic development and neuronal activity in cholinergic and catecholaminergic pathways. Dev. Brain Res. 116, 9–20.[ISI][Medline]

Dam, K., Seidler, F., and Slotkin, T. (1999b). Developmental neurotoxicity of chlorpyrifos: Delayed targeting of DNA synthesis after repeated administration. Dev. Brain Res. 108, 39–45.[ISI]

Danta, M., Dore, G. J., Hennessy, L., Li, Y., Vickers, C. R., Harley, H., Ngu, M., Reed, W., Desmond, P. V., Sievert, et al. (2002). Factors associated with severity of hepatic fibrosis in people with chronic hepatitis C infection. MJA 177, 240–245.[Medline]

Daston, G. P., Gooch, J. W., Breslin, W. J., Shuey, D. L., Nikiforov, A. I., Fico, T. A., and Gorsuch, J. W. (1997). Environmental estrogens and reproductive health: a discussion of the human and environmental data. Reprod. Toxicol. 11, 465–481.[CrossRef][ISI][Medline]

De Waard, M. A., and Van Nistelrooy, J. G. M. (1982). Antagonistic and synergistic activities of various chemicals on the toxicity of fenarimol to Aspergillus nidulans. Pestic. Sci. 13, 279–286.[ISI]

Degen, G. H., Janning, P., Wittsiepe, J., Upmeier, A., and Bolt, H. M. (2002). Integration of mechanistic data in the toxicological evaluation of endocrine modulators. Toxicol. Lett. 127(1–3), 225–237.[CrossRef][ISI][Medline]

Donato, F., Tagger, A., Gelatti, U., Parrinello, G., Boffetta, P., Albertini, A., Decarli, A., Trevisi, P., Ribero, M. L., Martelli, C., et al. (2002). Alcohol and hepatocellular carcinoma: the effect of lifetim intake and hepatitis virus infections in men and women. Am. J. Epidemiol. 155, 323–331.[Abstract/Free Full Text]

Dourson, M. L., Felter, S. P., and Robinson, D. (1996). Evolution of science-based uncertainty factors in noncancer risk assessment. Regul. Toxicol. Pharmacol. 24(2, Pt. 1), 108–120.[CrossRef][ISI][Medline]

Dourson, M. L., Knauf, L. A., and Swartout, J. C. (1992). On reference dose (RfD) and its underlying toxicity data base. Toxicol. Ind. Health 8, 171–189.[ISI][Medline]

Dourson, M. L., and Stara, J. F. (1983). Regulatory history and experimental support of uncertainty (safety) factors. Regul. Toxicol. Pharmacol. 3, 224–238.[Medline]

Dybing, E., and Soderlund, E. (1999). Situations with enhanced chemical risks due to toxicokinetic and toxicodynamic factors. Regul. Toxicol. Pharmacol. 30, 27–30.[CrossRef]

Ensoli, B., Gendelman, R., Markham, P., Fiorelli, V., Colombini, S., Raffeld, M., Cafaro, A., Chang, H., Brady, J., and Gallo, R. (1994). Synergy between basic fibroblast growth factor and HIV-1 Tat protein in induction of Kaposi’s sarcoma. Nature 371, 674–680.[CrossRef][ISI][Medline]

Environmental Protection Agency (EPA). (1994). Methods for derivation of inhalation reference concentrations and application of inhalation dosimetry. Dept. Health and Environmental Assessment, Washington, DC.

Environmental Protection Agency (EPA). (1996). The integrated risk information system. Dept. Environmental Assessment, Washington, DC.

Environmental Protection Agency (EPA). (1997). Special report on environmental endocrine disruption: An effects assessment and analysis. Washington, DC.

Fairbrother, K. S., Grove, J., de Waziers, I., Steimel, D. T., Day, C. P., Crespi, C. L., and Daly, A. K. (1998). Detection and characterization of novel polymorphisms in the CYP2E1 gene. Pharmacogenetics 8, 543–552.[ISI][Medline]

Fang, H., Tong, W., Perkins, R., Soto, A. M., Prechtl, N. V., and Sheehan, D. M. (2000). Quantitative comparisons of in vitro assays for estrogenic activities. Environ. Health Perspect. 108, 723–729.[ISI][Medline]

Faustman, E. M., Allen, B. C., Kavlock, R. J., and Kimmel, C. A. (1994). Dose-response assessment for developmental toxicity. I. Characterization of database and determination of no observed adverse effect levels. Fund. Appl. Toxicol. 23(4), 478–486.[CrossRef][ISI][Medline]

Feng, Y., Finley, J. W., Davis, C. D., Becker, W. K., Fretland, A. J., and Hein, D. W. (1999). Dietary selenium reduces the formation of aberrant crypts in rats administered 3,2'-dimethyl-4-aminobiphenyl. Toxicol. Appl. Pharmacol. 157, 36–42.[CrossRef][ISI][Medline]

Fenske, R., Black, K., Elkner, K., Lee, C., Methner, M., and Soto, R. (1990). Potential exposure and health risks of infants following indoor residential pesticide applications. Am. J. Public Health 80, 689–693.[Abstract]

Fredricks, G. R., Kincaid, R. L., Bondioli, K. R., and Wright, R. W., Jr. (1981). Ovulation rates and embryo degeneracy in female mice fed the phytoestrogen, coumestrol. Proc. Soc. Exp. Biol. Med. 167, 237–241.

Gaylor, D. W., and Kodell, R. L. (2002). A procedure for developing risk-based reference doses. Regul. Toxicol. Pharmacol. 35(2, Pt. 1), 137–141.[CrossRef][ISI][Medline]

Geissler, M., Gesien, A., and Wands, J. (1997). Inhibitory effects of chronic ethanol consumption on cellular immune responses to hepatitix C virus core protein are reversed by genetic immunizations augmented with cytokine-expressing plasmids. J. Immunol. 159, 5107–5113.[Abstract]

Goldstein, J. A. (2001). Clinical relevance of genetic polymorphisms in the human CYP2C subfamily. Br. J. Clin. Pharmacol. 52, 349–355.[CrossRef][ISI][Medline]

Goldstein, L. A., and Sengelaub, D. R. (1994). Differential effects of dihydrotestosterone and estrogen on the development of motoneuron morphology in a sexually dimorphic rat spinal nucleus. J. Neurobiol. 25, 878–892.[ISI][Medline]

Goyer, R. (1996). Results of lead research: Prenatal exposure and neurological consequences. Environ. Health Perspect. 104, 1050–1054.[ISI][Medline]

Gray, L. E., Jr., Wolf, C., Lambright, C., Mann, P., Price, M., Cooper, R. L., and Ostby, J. (1999). Administration of potentially antiandrogenic pesticides (procymidone, linuron, iprodione, chlozolinate, p,p'-DDE, and ketoconazole) and toxic substances (dibutyl- and diethylhexyl phthalate, PCB 169, and ethane dimethane sulphonate) during sexual differentiation produces diverse profiles of reproductive malformations in the male rat. Toxicol. Ind. Health 15(1–2), 94–118.[ISI][Medline]

Greenamyre, J. T., Sherer, T. B., Betarbet, R., and Panov, A. V. (2001). Complex I and Parkinson’s Disease. IUBMB Life 52, 135–141.[ISI][Medline]

Gundert-Remy, U., and Sonich-Mullin, C. (2002). The use of toxicokinetic and toxicodynamic data in risk assessment: An international perspective. Sci. Total Environ. 288(1–2), 3–11.[CrossRef][ISI][Medline]

Hamdy, S. I., Hiratsuka, M., Narahara, K., El-Enany, M., Moursi, N., Ahmed, M. S., and Mizugaki, M. (2002). Allele and genotype frequencies of polymorphic cytochromes P450 (CYP2C9, CYP2C19, CYP2E1) and dihydropyrimidine dehydrogenase (DPYD) in the Egyptian population. Br. J. Clin. Pharmacol. 53, 596–603.[CrossRef][ISI][Medline]

Hanson, M. L., and Solomon, K. R. (2002). New technique for estimating thresholds of toxicity in ecological risk assessment. Environ. Sci. Technol. 36, 3257–3264.[CrossRef][ISI][Medline]

Harry, J. (2001). Neurotoxicity Risk Assessment for Human Health: Principles and Approaches. World Health Organization, Geneva, Switzerland.

Hattis, D., Baird, S., and Goble, R. (2002). A straw man proposal for a quantitative definition of the RfD. Drug Chem. Toxicol. 25, 403–436.[CrossRef][ISI][Medline]

Hein, D. W. (2002). Molecular genetics and function of NAT1 and NAT2: Role in aromatic amine metabolism and carcinogenesis. Mutat. Res. 506–507, 65–77.[ISI]

Hohmann, C., Brooks, A., and Coyle, J. (1988). Neonatal lesions of the basal forebrain cholinergic neurons result in abnormal development. Dev. Brain Res. 42, 253–264.[ISI]

Hosokawa, S., Murakami, M., Ineyama, M., Yamada, T., Koyama, Y., Okuno, Y., Yoshitake, A., Yamada, H., and Miyamoto, J. (1993). Effects of procymidone on reproductive organs and serum gonadotropins in male rats. J. Toxicol. Sci. 18, 111–124.[Medline]

Hoyer, P. B. (2001). Reproductive toxicology: current and future directions. Biochem. Pharmacol. 62, 1557–1564.[CrossRef][ISI][Medline]

Hsieh, K. P., Lin, Y. Y., Cheng, C. L., Lai, M. L., Lin, M. S., Siest, J. P., and Huang, J. D. (2001). Novel mutations of CYP3A4 in Chinese. Drug Metab. Dispos. 29, 268–273.[Abstract/Free Full Text]

Hu, J.-y., Aizawa, T., and Ookubo, S. (2002). Products of aqueous chlorination of bisphenol A and their estrogenic activity. Environ. Sci. Technol. 36, 1980–1987.[CrossRef][ISI][Medline]

Hu, Y., Oscarson, M., Johansson, I., Yue, Q. Y., Dahl, M. L., Tabone, M., Arinco, S., Albano, E., and Ingelman-Sundberg, M. (1997). Genetic polymorphism of human CYP2E1: Characterization of two variant alleles. Mol. Pharmacol. 51, 370–376.[Abstract/Free Full Text]

Huff, R., and Abou-Donia, M. (1995). In vitro effect of chlorpyrifos oxon on muscarinic receptors and adenylate cyclase. Neurotoxicology 16, 281–290.[ISI][Medline]

Huff, R., Corcoran, J., Anderson, J., and Abou-Donia, M. (1994). Chlorpyrifos oxon binds directly to muscarinic receptors and inhibits cAMP accumulation in rat striatum. J. Pharmacol. Exp. Ther. 269, 329–335.[Abstract]

Ingelman-Sundberg, M. (2001a). Genetic susceptibility to adverse effects of drugs and environmental toxicants. The role of the CYP family of enzymes. Mutat. Res. 482(1–2), 11–19.[ISI][Medline]

Ingelman-Sundberg, M. (2001b). Genetic variability in susceptibility and response to toxicants. Toxicol. Lett. 120(1–3), 259–268.[CrossRef][ISI][Medline]

Ingelman-Sundberg, M., Oscarson, M., and McLellan, R. A. (1999). Polymorphic human cytochrome P450 enzymes: An opportunity for individualized drug treatment. Trends Pharmacol. Sci. 20, 342–349.[CrossRef][ISI][Medline]

Ishibe, N., Sinha, R., Hein, D. W., Kulldorff, M., Strickland, P., Fretland, A. J., Chow, W. H., Kadlubar, F. F., Lang, N. P., and Rothman, N. (2002). Genetic polymorphisms in heterocyclic amine metabolism and risk of colorectal adenomas. Pharmacogenetics 12, 145–150.[CrossRef][ISI][Medline]

Jaanson, P., Marandi, T., Kiivet, R. A., Vasar, V., Vaan, S., Svensson, J. O., and Dahl, M. L. (2002). Maintenance therapy with zuclopenthixol decanoate: Associations between plasma concentrations, neurological side effects and CYP2D6 genotype. Psychopharmacology (Berl) 162, 67–73.[CrossRef][ISI][Medline]

Jarabek, A. M. (1995). Interspecies extrapolation based on mechanistic determinants of chemical disposition. J. Ecol. Risk Assess. 15, 641–652.

Jiang, W., Feng, Y., and Hein, D. W. (1999). Higher DNA adduct levels in urinary bladder and prostate of slow acetylator inbred rats administered3,2'-dimethyl-4-aminobiphenyl. Toxicol. Appl. Pharmacol. 156, 187–194.[CrossRef][ISI][Medline]

Kato, N., Yoshida, H., Ono-Nita, S. K., Kato, J., Goto, T., Otsuka, M., Lan, K.-H., Matsushima, K., Shiratori, Y., and Omata, M. (2000). Activation of intracellular signaling by hepatitis B and C viruses: C-Viral core is most potent signal inducer. Hepatology 32, 405–412.[ISI][Medline]

Katz, E., Cortes, V., Eldefrawi, M., and Eldefrawi, A. (1997). Chlorpyrifos, parathion, and their oxons bind to and desensitize a nicotinic acetylcholine receptor: Relevance to their toxicities. Toxicol. Appl. Pharmacol. 146, 227–236.[CrossRef][ISI][Medline]

Kelce, W. R., Monosson, E., Gamcsik, M. P., Laws, S. C., and Gray, L. E., Jr. (1994). Environmental hormone disruptors: Evidence that vinclozolin developmental toxicity is mediated by antiandrogenic metabolites. Toxicol. Appl. Pharmacol. 126, 276–285.[CrossRef][ISI][Medline]

Kim, W.-H., Hong, F., Jaruga, B., Hu, Z., Fan, S., Liang, T. J., and Gao, B. (2001). Additive activation of hepatic NF-kB by ethanol and hepatitis B proteinxor HCV core protein: Involvement of TNF-a receptor 1-independent and dependent mechanisms. FASEB 15, 2551–2553.[Abstract/Free Full Text]

Krynetski, E. Y., and Evans, W. E. (1998). Pharmacogenetics of cancer therapy: Getting personal. Am. J. Hum. Genet. 63, 11–16.[CrossRef][ISI][Medline]

Li, Y., Wang, X., Tian, S., Guo, C.-J., Douglas, S. D., and Ho, W.-Z. (2002). Methadone enhances human immunodeficiency virus infection of human immune cells. J. Infect. Dis. 185, 118–122.[CrossRef][ISI][Medline]

Liu, P., Kimmoun, E., Legrand, A., Sauvanet, A., Degott, C., Lardeux, B., and Bernuau, D. (2002). Activation of NF-kappaB, AP-1, and STAT transcription factors is a frequent and early event in human hepatocellular carcinoma. J. Hepatol. 37, 63–71.[CrossRef][ISI][Medline]

Lotti, M. (2002). Age-related sensitivity of the nervous system to neurotoxic insults. Toxicol. Lett. 127, 183–187.[CrossRef][ISI][Medline]

Ma, M. K., Woo, M. H., and McLeod, H. L. (2002). Genetic basis of drug metabolism. Am. J. Health Syst. Pharm. 59, 2061–2069.[ISI][Medline]

Madden, J. J., Wang, Y., Lankford-Turner, P., and Donahoe, R. M. (2002). Does reduced DNA repair capacity play a role in HIV infection and progression in the lymphocytes of opiate addicts? JAIDS 31, S78–S83,[ISI][Medline]

Masimirembwa, C., Persson, I., Bertilsson, L., Hasler, J., and Ingelman-Sundberg, M. (1996). A novel mutant variant of the CYP2D6 gene (CYP2D6*17) common in a black African population: Association with diminished debrisoquine hydroxylase activity. Br. J. Clin. Pharmacol. 42, 713–719.[ISI][Medline]

McBride, W. (1961). Thalidomide and congenital anomalies. Lancet 2, 1358.[ISI]

McCarthy, T. V., Quane, K. A., and Lynch, P. J. (2000). Ryanodine receptor mutations in malignant hyperthermia and central core disease. Hum. Mutat. 15, 410–417.[CrossRef][ISI][Medline]

Melnick, R., Lucier, G., Wolfe, M., Hall, R., Stancel, G., Prins, G., Gallo, M., Reuhl, K., Ho, S.-M., Brown, T., et al. (2002). Summary of the national toxicology program’s report of the endocrine disruptors low-dose peer review. Environ. Health Perspect. 110, 427–431.[Medline]

Miller, M. C., III, Mohrenweiser, H. W., and Bell, D. A. (2001). Genetic variability in susceptibility and response to toxicants. Toxicol. Lett. 120(1–3), 269–280.[CrossRef][ISI][Medline]

Milman, H. A., Bosland, M. C., Walden, P. D., and Heinze, J. E. (2002). Evaluation of the adequacy of published studies of low-dose effects of bisphenol A on the rodent prostate for use in human risk assessment. Regul. Toxicol. Pharmacol. 35, 338–346.[CrossRef][ISI][Medline]

Miyagi, T., Chuang, L. F., Doi, R. H., Carlos, M. P., Torres, J. V., and Chuang, R. Y. (2000). Morphine induces gene expression of CCR5 in human CEM x174 lymphocytes. J. Biol. Chem. 275, 31305–31310.[Abstract/Free Full Text]

Miyamoto, M., Umetsu, Y., Dosaka-Akita, H., Sawamura, Y., Yokota, J., Kunitoh, H., Nemoto, N., Sato, K., Ariyoshi, N., and Kamataki, T. (1999). CYP2A6 gene deletion reduces susceptibility to lung cancer. Biochem. Biophys. Res. Commun. 261, 658–660.[CrossRef][ISI][Medline]

Murray, H. E., Pillai, A. V., McArthur, S. R., Razvi, N., Datla, K. P., Dexter, D. T., and Gillies, G. E. (2003). Dose- and sex-dependent effects of the neurotoxin 6-hydroxydopamine on the nigrostriatal dopaminergic pathway of adult rats: Differential actions of estrogen in males and females. Neuroscience 116, 213–222.[CrossRef][ISI][Medline]

Mylchreest, E., Wallace, D. G., Cattley, R. C., and Foster, P. M. D. (2000). Dose-dependent alterations in androgen-regulated male reproductive development in rats exposed to di(n-butyl) phthalate during late gestation. Toxicol. Sci. 55, 143–151.[Abstract/Free Full Text]

Nath, A., Hauser, K. F., Wojna, V., Booze, R. M., Maragos, W., Prendergast, M., Cass, W., and Turchan, J. T. (2002). Molecular basis for interactions of HIV and drugs of abuse. JAIDS 31, S62–S69.[ISI][Medline]

Needleman, H. (1994). Preventing childhood lead poisoning. Prev. Med. 23, 634–637.[CrossRef][ISI][Medline]

Oscarson, M., Gullsten, H., Rautio, A., Bernal, M. L., Sinues, B., Dahl, M. L., Stengard, J. H., Pelkonen, O., Raunio, H., and Ingelman-Sundberg, M. (1998). Genotyping of human cytochrome P450 2A6 (CYP2A6), a nicotine C-oxidase. FEBS Lett. 438, 201–205.[CrossRef][ISI][Medline]

Oscarson, M., McLellan, R. A., Gullsten, H., Agundez, J. A., Benitez, J., Rautio, A., Raunio, H., Pelkonen, O., and Ingelman-Sundberg, M. (1999a). Identification and characterisation of novel polymorphisms in the CYP2A locus: Implications for nicotine metabolism. FEBS Lett. 460, 321–327.[CrossRef][ISI][Medline]

Oscarson, M., McLellan, R. A., Gullsten, H., Yue, Q. Y., Lang, M. A., Bernal, M. L., Sinues, B., Hirvonen, A., Raunio, H., Pelkonen, O., et al. (1999b). Characterisation and PCR-based detection of a CYP2A6 gene deletion found at a high frequency in a Chinese population. FEBS Lett. 448, 105–110.[CrossRef][ISI][Medline]

Paolini, M., Mesirca, R., Pozzetti, L., Maffei, F., Vigagni, F., Hrelia, P., and Cantelli-Forti, G. (1996). Genetic and non-genetic biomarkers related to carcinogenesis in evaluating toxicological risk from Fenarimol. Mutat. Res. 368, 27–39.[ISI][Medline]

Parman, T., Wiley, M., and Wells, P. (1999). Free radical mediated oxidative DNA damage in the mechanism of thalidomide teratogenicity. Nature Med. 5, 582–585.[CrossRef][ISI][Medline]

Paschke, T., Riefler, M., Schuler-Metz, A., Wolz, L., Scherer, G., McBride, C. M., and Bepler, G. (2001). Comparison of cytochrome P450 2A6 polymorphism frequencies in Caucasians and African-Americans using a new one-step PCR-RFLP genotyping method. Toxicology 168, 259–268.[CrossRef][ISI][Medline]

Pirmohamed, M., and Park, B. K. (2001). Genetic susceptibility to adverse drug reactions. Trends Pharmacol. Sci. 22, 298–305.[CrossRef][ISI][Medline]

Pruett, S. B., Myers, L. P., and Keil, D. E. (2001). Toxicology of metam sodium. J. Toxicol. Environ. Health 4, 207–222.[ISI]

Pylkkanen, L., Makela, S., Valve, E., Harkonen, P., Toikkanen, S., and Santti, R. (1993). Prostatic dysplasia associated with increased expression of c-myc in neonatally estrogenized mice. J. Urol. 149, 1593–1601.[ISI][Medline]

Pylkkanen, L., Santti, R., Maentausta, O., and Vihko, R. (1992). Distribution of estradiol-17 beta hydroxysteroid oxidoreductase in the urogenital tract of control and neonatally estrogenized male mice: Immunohistochemical, enzymehistochemical, and biochemical study. Prostrate 20, 59–72.

Pylkkanen, L., Santti, R., Newbold, R., and McLachlan, J. A. (1991). Regional differences in the prostate of the neonatally estrogenized mouse. Prostrate 18, 117–129.

Qiao, D., Seidler, F. J., Abreu-Villaça, Y., Tate, C. A., Cousins, M. M., Thillai, I., and Slotkin, T. A. (2003). Chlorpyrifos exposure during neurulation: Cholinergic synaptic dysfunction and cellular alterations in brain regions at adolescence and adulthood. Brain Res. (under review).

Quesada, I., Fuentes, E., Viso-Leon, M. C., Soria, B., Ripoll, C., and Nadal, A. (2002). Low doses of the endocrine disruptor bisphenol-A and the native hormone 17.beta-estradiol rapidly activate the transcription factor CREB. FASEB J. 16, 1671–1673.[Abstract/Free Full Text]

Radice, S., Ferraris, M., Marabini, L., and Chiesara, E. (2002). Estrogenic activity of procymidone in primary cultured rainbow trout hepatocytes (Oncorhynchus mykiss). Toxicol. in vitro 16, 475–80.[CrossRef][ISI][Medline]

Rao, S. B., and Mehendale, H. M. (1993). Halomethane-chlordecone (CD) interactive hepatotoxicity—Current concepts on the mechanism. Ind. J. Biochem. Biophys. 30, 191–198.[Medline]

Raunio, H., Rautio, A., Gullsten, H., and Pelkonen, O. (2001). Polymorphisms of CYP2A6 and its practical consequences. Br. J. Clin. Pharmacol. 52, 357–363.[CrossRef][ISI][Medline]

Relling, M. V., Hancock, M. L., Boyett, J. M., Pui, C. H., and Evans, W. E. (1999). Prognostic importance of 6-mercaptopurine dose intensity in acute lymphoblastic leukemia. Blood 93, 2817–2823.[Abstract/Free Full Text]

Renwick, A. G. (1993). Data-derived safety factors for the evaluation of food additives and environmental contaminants. Food Addit. Contam. 10, 275–305.[ISI][Medline]

Rodier, P. M. (1994). Vulnerable periods and processes during central nervous system development. Environ. Health Perspect. 102, 121–124.

Sanderson, J. T., Letcher, R. J., Heneweer, M., Giesy, J. P., and Van den Berg, M. (2001). Effects of chloro-s-triazine herbicides and metabolites on aromatase activity in various human cell lines and on vitellogenin production in male carp hepatocytes. Environ. Health Perspect. 109, 1027–1031.[ISI][Medline]

Sata, F., Sapone, A., Elizondo, G., Stocker, P., Miller, V. P., Zheng, W., Raunio, H., Crespi, C. L., and Gonzalez, F. J. (2000). CYP3A4 allelic variants with amino acid substitutions in exons 7 and 12: Evidence for an allelic variant with altered catalytic activity. Clin. Pharmacol. Ther. 67, 48–56.[ISI][Medline]

Scordo, M. G., Aklillu, E., Yasar, U., Dahl, M. L., Spina, E., and Ingelman-Sundberg, M. (2001). Genetic polymorphism of cytochrome P450 2C9 in a Caucasian and a black African population. Br. J. Clin. Pharmacol. 52, 447–450.[CrossRef][ISI][Medline]

Sipos, I., Tretter, L., and Adam-Vizi, V. (2003). Quantitative relationship between inhibition of respiratory complexes and formation of reactive oxygen species in isolated nerve terminals. J. Neurochem. 84, 112–118.[CrossRef][ISI][Medline]

Slotkin, T. (1999). Developmental cholinotoxicants: Nicotine and chlorpyrifos. Environ. Health Persp. 107(Suppl. 1), 71–80.[ISI][Medline]

Slotkin, T. A., Cousins, M. L., Tate, C. A., and Seidler, F. J. (2001). Persistent cholinergic presynaptic deficits after neonatal chlorpyrifos exposure. Brain Res. 902, 229–243.[CrossRef][ISI][Medline]

Slotkin, T. A., Tate, C. A., Cousins, M. M., and Seidler, F. J. (2002). Functional alterations in CNS catecholamine systems in adolescence and adulthood after neonatal chlorpyrifos exposure. Dev. Brain Res. 133, 163–173.[ISI][Medline]

Song, X., Violin, J. D., Seidler, F. J., and Slotkin, T. A. (1998). Modeling the developmental neurotoxicity of chlorpyrifos in vitro: Macromolecule synthesis in PC12 cells. Toxicol. Appl. Pharmacol. 151, 182–191.[CrossRef][ISI][Medline]

Sun, F., Tsuritani, I., and Yamada, Y. (2002). Contribution of genetic polymorphisms in ethanol-metabolizing enzymes to problem drinking behavior in middle-aged Japanese men. Beh. Genetics 32, 229–236.[CrossRef][ISI]

Szabo, G., Mandrekar, P., Dolganuic, A., Catalano, D., and Kodys, K. (2001). Reduced alloreactive T-cell activation after alcohol intake is due to impaired monocyte accessory cell function and correlates with elevated IL-10, IL-13, and decreased IFN gamma levels. Alcohol Clin. Exp. Res. 25, 1766–1772.[ISI][Medline]

Takahashi, K., Takahashi, T., Takahashi, S., Watanabe, K., Boku, S., Matsui, S., Arai, F., and Asakura, H. (2001). Difference in quasispecies of the hypervariable region 1 of hepatitis C virus between alcoholic and non-alcoholic patients. J. Gastroenterol. Hepatol. 16, 416–423.[CrossRef][ISI][Medline]

Takiguchi, M., Cherrington, N. J., Hartley, D. P., Klaassen, C. D., and Waalkes, M. P. (2001). Cyproterone acetate induces a cellular tolerance to cadmium in rat liver epithelial cells involving reduced cadmium accumulation. Toxicology 165, 13–25.[CrossRef][ISI][Medline]

Thiruchelvam, M., Brockel, B., Richfield, E., Baggs, R., and Cory-Slechta, D. (2000). Potentiated and preferential effects of combined paraquat and maneb on nigrostriatal dopamine systems: Environmental risk factors for Parkinson’s disease? Brain Res. 873, 225–234.[CrossRef][ISI][Medline]

Troen, B. R. (2003). The biology of aging. Mount Sinai J. Med. 70, 3–22.[ISI][Medline]

Tsaih, S., Korrick, S., Schwartz, J., Lee, M., Amarasiriwardena, C., Aro, A., Sparrow, D., and Hu, H. (2001). Influence of bone resorption on the mobilization of lead from bone among middle-aged and elderly men: The normative aging study. Environ. Health Perspect. 109, 995–999.[ISI][Medline]

Turner, R. T., Wakley, G. K., Hannon, K. S., and Bell, N. H. (1988). Tamoxifen inhibits osteoclast-mediated resorption of trabecular bone in ovarian hormone-deficient rats. Endocrinology 122, 1146–1150.[Abstract]

Vento, S., and Cainelli, F. (2002). Does hepatitis C virus cause severe liver disease only in people who drink alcohol? Lancet Inf. Dis. 2, 303–309.[CrossRef][ISI][Medline]

Vermeulen, A., and Comhaire, F. (1978). Hormonal effects of an antiestrogen, tamoxifen, in normal and oligospermic men. Fert. Steril. 29, 320–327.[ISI][Medline]

Vig, E., and Hu, H. (2000). Lead toxicity in older adults. J. Am. Geriatr. Soc. 48, 1501–1506.[ISI][Medline]

Westin, J., Lagging, L. M., Spak, F., Aires, N., Svensson, E., Lindh, M., Dhillon, A. P., Norkrans, G., and Wejstal, R. (2002). Moderate alcohol intake increases fibrosis progression in untreated patients with hepatitis C virus infection. J. Viral Hep. 9, 235–241.[CrossRef][ISI]

Wilcox, A. J., Baird, D. D., Weinberg, C. R., Hornsby, P. P., and Herbst, A. L. (1995). Fertility in men exposed prenatally to diethylstilbestrol. N. Engl. J. Med. 332, 1411–1416.[Abstract/Free Full Text]

Williams, K., Saunders, P. T. K., Atanassova, N., Fisher, J. S., Turner, K. J., Millar, M. R., McKinnell, C., and Sharpe, R. M. (2000). Induction of progesterone receptor immunoexpression in stromal tissue throughout the male reproductive tract after neonatal oestrogen treatment of rats. Mol. Cell. Endocrinol. 164(1–2), 117–131.[CrossRef][ISI][Medline]

Witorsch, R. J. (2002). Endocrine disruptors: Can biological effects and environmental risks be predicted? Regul. Toxicol. Pharmacol. 36, 118–130.[CrossRef][ISI][Medline]

Wolf, C. J., LeBlanc, G. A., Ostby, J. S., and Gray, L. E., Jr. (2000). Characterization of the period of sensitivity of fetal male sexual development to vinclozolin. Toxicol. Sci. 55, 152–161.[Abstract/Free Full Text]

You, L., Casanova, M., Bartolucci, E. J., Fryczynski, M. W., Dorman, D. C., Everitt, J. I., Gaido, K. W., Ross, S. M., and Heck, H. D. A. (2002). Combined effects of dietary phytoestrogen and synthetic endocrine-active compound on reproductive development in Sprague-Dawley rats: Genistein and methoxychlor. Toxicol. Sci. 66, 91–104.[Abstract/Free Full Text]

Zauli, G., Gibellini, D., MIlani, D., Mazzoni, M., Borgatti, P., Placa, M. L., and Capitani, S. (1993). Human immunodeficiency virus type 1 tat protein protects lymphoid, epithelial, and neuronal cell lines from death by apoptosis. Cancer Res. 53, 4481–4485.[Abstract]

Zhang, J., Fitsanakis, V. A., Gu, G., Jing, D., Ao, M., Amarnath, V., and Montine, T. J. (2003). Manganese ethylene-bis-dithiocarbamate and selective dopaminergic neurodegeneration in rat: A link through mitochondrial dysfunction. J. Neurochem. 84, 336–346.[CrossRef][ISI][Medline]





This Article
Abstract
FREE Full Text (PDF)
Supplementary tables
All Versions of this Article:
76/1/3    most recent
kfg204v1
Alert me when this article is cited
Alert me if a correction is posted
Services
Email this article to a friend
Similar articles in this journal
Similar articles in PubMed
Alert me to new issues of the journal
Add to My Personal Archive
Download to citation manager
Search for citing articles in:
ISI Web of Science (2)
Disclaimer
Request Permissions
Google Scholar
Articles by Aldridge, J. E.
Articles by Levin, E. D.
PubMed
PubMed Citation
Articles by Aldridge, J. E.
Articles by Levin, E. D.