Alexander Ure MD, FRCS (1808-1866), and the beginning of drug metabolism studies.

Although many fields of endeavour emerge owing to the coalescence of the work and observations of numerous individuals, there is usually one seminal event that unites and acts as a catalyst to stimulate and advance the process. Such was the case with Alexander Ure. Up to this point it had been speculated that chemicals taken into the body may undergo bio-transformation, akin to the digestion of nutrients, but no unequivocal and quantitative experiments had been performed before those of Ure. Following his observations the subject began to flourish; to him may be attributed the beginnings of xenobiochemistry and the field now known as drug metabolism.

Plastic contamination of the food chain: A threat to human health?

Macro-plastic pollution is found in terrestrial and marine environments and is degraded to micro-particles (MP) and nano-particles (NP) of plastic. These can enter the human food chain either by inhalation or by ingestion, particularly of shellfish and crustaceans. Absorption across the gastrointestinal tract is relatively low, especially for MPs, which appear to have little toxicity. However, NPs are more readily absorbed and may accumulate in the brain, liver and other tissues in aquatic species and other animals. Studies using nanoparticles of other materials suggest that toxicity could potentially affect the central nervous system and the reproductive system, although this would be unlikely unless exposure levels were very high and absorption was increased by physiological factors.

Drug metabolism in the elderly: A multifactorial problem?

Whether or not an individual's drug metabolising capacity declines with advancing age is a vexing question. There is no clear evidence that drug metabolism itself ('the biologically-assisted chemical alteration of the administered parent molecule') is less efficient in healthy old age than at younger ages, whereas a decreased capacity may be associated with ill-health and frailty. However, elderly individuals do show a reduced enzyme induction capability and are less able to tolerate overdoses. It appears that the majority of deleterious clinical outcomes related to drug therapy in an elderly (usually ill or frail) population may be ascribed to various anatomical and physiological age-related changes. These may affect both pharmacodynamics and pharmacokinetics, but not necessarily drug metabolism. Information gleaned from animal studies undertaken mainly in rodents does not seem to be of relevance to humans and studies in healthy aged human populations may not highlight possible problems. However, certain circumstances may influence metabolic competence, and phenotyping rather than genotyping is of more value in identifying those susceptible to adverse drug reactions. This short review discusses the potential contributions of four factors (inflammation, circadian rhythm, gut microbes, epigenetic aspects) which may lead to alterations in drug metabolism with increasing age.

Enteral feeding reduces metabolic activity of the intestinal microbiome in Crohn's disease: an observational study.

BACKGROUND/OBJECTIVES: Enteral feeding will induce remission in as many as 80-90% of compliant patients with active Crohn's disease (CD), but its method of action remains uncertain. This study was designed to examine its effects on the colonic microbiome. METHODS/SUBJECTS: Healthy volunteers and patients with CD followed a regimen confined to enteral feeds alone for 1 or 2 weeks, respectively. Chemicals excreted on breath or in faeces were characterised at the start and at the end of the feeding period by gas chromatography/mass spectrometry. RESULTS: One week of feeding in healthy volunteers caused significant changes in stool colour and deterioration in breath odour, together with increased excretion of phenol and indoles on the breath. Feeding for 2 weeks in patients with CD produced significant improvements in symptoms and a decrease in the concentration of C-reactive protein. The faecal concentrations of microbial products, including short-chain fatty acids (SCFAs), and potentially toxic substances, including 1-propanol, 1-butanol and the methyl and ethyl esters of SCFAs, showed significant falls. CONCLUSIONS: A significant change occurs in the production of microbial metabolites after enteral feeding in both healthy volunteers and patients with CD. Many of those detected in CD are toxic and may feasibly lead to the immunological attack on the gut microbiota, which is characteristic of inflammatory bowel disease. The reduction in the production of such metabolites after enteral feeding may be the reason for its effectiveness in CD.

In utero exposure to carcinogens: Epigenetics, developmental disruption and consequences in later life.

The uterine environment is often viewed as a relatively safe haven, being guarded by the placenta which acts as a filter, permitting required materials to enter and unwanted products to be removed. However, this defensive barrier is sometimes breached by potential chemical hazards to which the mother may be subjected. Many of these toxins have immediate and recognisable deleterious effects on the embryo, foetus or neonate, but a few are insidious and leave a legacy of health issues that may emerge in later life. Several substances, falling into the categories of metals and metalloids, endocrine disruptors, solvents and other industrial chemicals, have been implicated in the development of long-term health problems in the offspring following maternal and subsequent in utero exposure. The mechanisms involved are complex but often involve epigenetic changes which disrupt normal cell processes leading to the development of cancers and also dysregulation of biochemical pathways.

Xenobiotic sulphation and its variability during inflammation: a factor in adverse drug reactions?

The interactions between disease processes and the metabolism of therapeutic drugs have not been systematically investigated. Inflammation, with the presence of pro-inflammatory cytokines, affects Phase 1 metabolism, particularly the activity of the CYP isoforms. Inflammatory factors also alter the activity of some Phase 2 enzymes, particularly the sulphotransferases (SULT isoforms) responsible for drug sulphonation and the enzyme pathway involved in the supply of sulphate for this reaction. Being ill may, therefore, in itself make drug metabolism unpredictable.

Diethylstilboestrol--a long-term legacy.

Diethylstilboestrol (DES) is an endocrine disrupter which causes cancer in rodents. It was prescribed in large amounts to treat women with gynaecological problems; some of the daughters of these women subsequently developed a rare cancer (vaginal clear cell adenocarcinoma) while genital abnormalities were found in some of the sons. It was used for decades in livestock feed and this may have contaminated the food chain leading to the exposure of the more general population. DES appears to cause epigenetic effects in animals and there is some evidence that this also occurs in man. The mechanisms of carcinogenesis are complex and the effects are difficult to prove due to the background of dietary and environmental phyto- and xenooestrogens. It has been suggested that, like other endocrine disrupters, DES may have acted as an obesogen in the human population.

Detection of endocrine disruptors - from simple assays to whole genome scanning.

Endocrine disruptors frequently bear little structural relationship to the hormone whose actions they disrupt. Consequently, the threat of an uninvestigated chemical cannot easily be assessed. Here three different approaches to assessment are discussed. The first presumes an endocrine-disrupting property, following which a cell model capable of responding to such a hormone is used. Although simple and cheap, it provides limited data. A second approach involves multiple assays to detect multiple hormones. Increasing the amount of data increased the difficulty in assessing the significance of results. To meet this problem, cluster analysis based on a simple mathematical matrix was adopted. The matrix was used to determine (i) a limited number of assays to identify a maximum number of endocrine disruptors and (ii) the chemicals with the most wide-ranging effects. A third approach was a whole genome expression analysis based on expression of mRNAs in human TE671 medulloblastoma cells. Expression of individual mRNAs was assessed using the Affymetrix GeneChip(®) Human Genome U133 Plus 2.0 chip. The significance of differential expressed genes was assessed based on gene ontology and pathways analyses using DAVID and GenMaPP programs. The results illustrated the very wide-ranging effects of these chemicals across the genome.

Biomarkers of endocrine disruption: cluster analysis of effects of plasticisers on Phase 1 and Phase 2 metabolism of steroids.

Although some endocrine disruptors (EDs) act at steroid receptors, it is now apparent that compounds may have ED potential if they alter steroid synthesis or metabolism, particularly if they affect Phase 1 or Phase 2 pathways. In the ENDOMET project (EU-funded 5th Framework programme), 23 different assays were used on a wide range of EDs. Cluster analysis of the matrix results enabled identification of four integrated test systems that can be used to pinpoint compounds that are able to alter steroid metabolism or function. Critical pathways were shown to include oestrogen synthesis and sulphonation, synthesis of sulphate/PAPS and thyroid hormone regulation so that the activity profiles of some Phase 1 and Phase 2 reactions can be used as biomarkers for detection of compounds with ED potential.

Reactive oxygen species and oxidative DNA damage mediate the cytotoxicity of tungsten-nickel-cobalt alloys in vitro.

Tungsten alloys (WA) have been introduced in an attempt to find safer alternatives to depleted uranium and lead munitions. However, it is known that at least one alloy, 91% tungsten-6% nickel-3% cobalt (WNC-91-6-3), causes rhabdomyosarcomas when fragments are implanted in rat muscle. This raises concerns that shrapnel, if not surgically removable, may result in similar tumours in humans. There is therefore a clear need to develop rapid and robust in vitro methods to characterise the toxicity of different WAs in order to identify those that are most likely to be harmful to human health and to guide development of new materials in the future. In the current study we have developed a rapid visual in vitro assay to detect toxicity mediated by individual WA particles in cultured L6-C11 rat muscle cells. Using a variety of techniques (histology, comet assay, caspase-3 activity, oxidation of 2'7'-dichlorofluorescin to measure the production of reactive oxygen species and whole-genome microarrays) we show that, in agreement with the in vivo rat carcinogenicity studies, WNC-91-6-3 was the most toxic of the alloys tested. On dissolution, it produces large amounts of reactive oxygen species, causes significant amounts of DNA damage, inhibits caspase-3, triggers a severe hypoxic response and kills the cells in the immediate vicinity of the alloy particles within 24h. By combining these in vitro data we offer a mechanistic explanation of the effect of this alloy in vivo and show that in vitro tests are a viable alternative for assessing new alloys in the future.

Endocrine disrupters--a threat to women's health?

Endocrine disruption has been a topic of public concern for many years and its study remains high on the scientific agenda. Endocrine disrupters (EDs) are compounds which may be of industrial or natural origin and which act to dysregulate steroid function and metabolism. As well as their actions on nuclear steroid receptors, EDs can inhibit the pathways of steroid synthesis and degradation. They not only affect reproductive function but also affect a range of tissues which are steroid sensitive such as the central nervous system and thyroid. Results from the latest studies suggest that EDs may also affect the immune system, glucose homeostasis and can act as epigenetic modulators resulting in transgenerational effects. Research in this area has led to the development of drugs used in the treatment of several types of hormone-sensitive cancer. However, despite many years of effort, the effects on human health of long-term environmental exposure to EDs, whether singly or as mixtures, remain unknown.

The menstrual cycle and drug metabolism.

Within the field of drug metabolism, when addressing quantitative aspects, an average value is traditionally quoted, commonly the arithmetic mean with perhaps an indication of spread. Better still a range of values may be given, thereby acknowledging that various factors may precipitate differences between individuals. A single subject, however, usually only merits a single value. Nevertheless, events such as an acute illness or concurrent drug therapy serve to alert that this value may change substantially over a relatively short time-period, although any potential effects of naturally occurring phenomena, such as the female menstrual cycle, are often overlooked or disregarded. Are the biochemical and physiological changes that occur during the menstrual cycle able to influence xenobiotic metabolism? Is the idea of a stable and unwavering baseline within a single healthy individual flawed? Is it time to reassess our thinking with regards to such aspects? This brief review explores these issues and examines information available within the literature for evidence of potential influences of menstrual cycle events upon drug metabolism, defined as the actual chemical alteration of the parent molecule into another chemical species.

Allometric relationships predicting foliar biomass and leaf area:sapwood area ratio from tree height in five Costa Rican rain forest species.

We developed allometric equations to predict whole-tree leaf area (A(l)), leaf biomass (M(l)) and leaf area to sapwood area ratio (A(l):A(s)) in five rain forest tree species of Costa Rica: Pentaclethra macroloba (Willd.) Kuntze (Fabaceae/Mim), Carapa guianensis Aubl. (Meliaceae), Vochysia ferru-gi-nea Mart. (Vochysiaceae), Virola koshnii Warb. (Myristicaceae) and Tetragastris panamensis (Engl.) Kuntze (Burseraceae). By destructive analyses (n = 11-14 trees per species), we observed strong nonlinear allometric relationships (r(2) > or = 0.9) for predicting A(l) or M(l) from stem diameters or A(s) measured at breast height. Linear relationships were less accurate. In general, A(l):A(s) at breast height increased linearly with tree height except for Penta-clethra, which showed a negative trend. All species, however, showed increased total A(l) with height. The observation that four of the five species increased in A(l):A(s) with height is consistent with hypotheses about trade--offs between morphological and anatomical adaptations that favor efficient water flow through variation in the amount of leaf area supported by sapwood and those imposed by the need to respond quickly to light gaps in the canopy.

Sulfotransferase inhibition: potential impact of diet and environmental chemicals on steroid metabolism and drug detoxification.

The cytosolic sulfotransferase enzymes (SULT isoforms) utilise PAPS (3'-phosphoadenosine-5'-phosphosulfate) as co-factor to transfer sulfonate groups onto a wide range of substrates. SULT1A3 has catecholamines such as dopamine as substrates while SULT 1E1 sulfonates oestrogens. SULT 1A1 sulfonates phenols and also oestrogens at a higher K(m) than SULT 1E1. SULT 2A1 mainly sulfonates DHEA and some steroids, with hydroxy derivatives of polycyclic aromatic hydrocarbons. Studies on these isoforms with a range of environmental chemicals and dietary components have shown that SULT 1A1 is significantly inhibited by flavonoids; all flavones and flavonols with a 3',4'-dihydroxy motif had an IC(50) of < 100 nm against 3 microM 4-nitrophenol as the standard substrate. SULTs 1A3 and 2A1 were less strongly inhibited by flavonoids or isoflavonoids although tricin (3',5'-dimethoxy-4',5,7-trihydroxyflavone is a competitive inhibitor of SULT 1E1 with an inhibition constant of approximately 1 nM. Fruit and vegetable cytosols also inhibit SULT isoforms, as do long-chain alkylphenols and chlorinated phenols. Phthalates (used as plasticisers) inhibited SULTs 1E1 and 2A1. As these environmental contaminants and dietary components all act at the same site, their effects would be expected to be additive and could potentially therefore reduce sulfonation of drugs and lead to altered pharmacological responses.

Wide-ranging genomic effects of plasticisers and related compounds.

UNLABELLED: The effects of four compounds, bis(2-ethylhexyl)phthalate (BEHP); diisodecylphthalate (DIP); 4-n-octylphenol (OP); 4-chloro-3-methylphenol (CMP), on gene expression (steady-state mRNA levels) across the whole human genome were studied in human TE671 cells. Effects were studied using the Affymetrics GeneChip Human Genome U133 Plus 2.0, HG-U133 Plus 2.0 arrays, The array analyses the expression of 47,000 transcripts and variants, including approximately 38,500 well characterised. All four compounds exerted statistically significant actions, affecting between 4 and 6.5% of all genes. Each compound had its own expression signature. In most instances where there was an effect, steady-state mRNA levels were decreased, although not always. CMP treatment caused most increases in mRNA levels. A mixture of DIP and CMP caused fewer changes in mRNA levels than either of the individual compounds. CONCLUSIONS: These plasticisers affected the steady-state mRNA levels of many human genes. Exposure to these compounds over many years has the potential to influence human health.

Phytoestrogens and xenoestrogens: the contribution of diet and environment to endocrine disruption.

Some endocrine disrupting compounds such as phthalates and phenols act non-genomically by inhibiting the sulfotransferase (SULT 1E1 and SULT 1A1) isoforms which inactivate estrogens by sulfonation. A range of environmental phenolic contaminants and dietary flavonoids was tested for inhibition of the human SULT 1A1, 1E1 and 2A1 isoforms. In particular, the plasticisers 4-n-octyl- and 4-n-nonyl-phenol inhibit SULT 1E1 with IC(50) values of 0.16 microM vs. 10nM estradiol while the 2-substituted chlorophenols show similar values. Flavonoids are also SULT inhibitors; tricin is a competitive inhibitor of SULT 1E1 with a K(i) of 1.5+/-0.8 nM. In a small pilot study to determine whether ingestion of soy flavonoids would affect SULT1A1 activity in vivo as well as in vitro, sulfonation of daidzein was reduced in a group of women 'at risk' of breast cancer, as compared with controls, although the SULT 1A1*1/SULT 1A1*2 allele ratio was not different. Endocrine disrupting effects in man may be multifactorial when components from both the diet and the environment act at the same point in steroid metabolism.

Induction of cysteine dioxygenase activity by oral administration of cysteine analogues to the rat: implications for drug efficacy and safety.

One of the major steps in the oxidation of the sulphur-containing amino acid, L-cysteine, is the production of cysteine sulphinic acid, catalysed by the enzyme cysteine dioxygenase. This enzyme plays a key role in the intermediary metabolism of sulphur-containing compounds. The activity of this crucial enzyme is known to be influenced by sulphur-compound intake, being increased in animals fed an excess of L-cysteine or methionine. However, the affects on this enzyme of the chronic administration of drugs similar in structure to cysteine are unknown. This has now been investigated using the anti-rheumatic agent, D-penicillamine, and the mucoactive compound, S-carboxymethyl-L-cysteine. Repeated oral administration of these sulphur-containing drugs to male Wistar rats for five consecutive days led to a significant increase in hepatic cysteine dioxygenase activity. This increase in the production rate of cysteine sulphinic acid remained evident until returning to control levels four days after cessation of drug administration. These observations provide evidence that these two drugs interact with the intermediary biochemistry of sulphur compounds and may provide hitherto unappreciated insights into mechanisms by which therapeutic effects and adverse reactions may occur.

Endocrine disrupters: a human risk?

Endocrine disrupters (EDs) alter normal hormonal regulation and may be naturally occurring or environmental contaminants. Classically, EDs act genomically, with agonistic or antagonistic effects on steroid receptors and may alter reproductive function and/or cause feminisation by binding to oestrogen or androgen receptors; their binding to the thyroid receptor may dysregulate the neuroendocrine system. Recently, it has been shown that EDs can also act by non-genomic mechanisms, altering steroid synthesis (inhibition of cytochrome P450 isoforms) or steroid metabolism. The alkylphenol and phthalate plasticisers inhibit the inactivation of oestrogens by sulphation (via SULT 1A1 and 1E1 isoforms) and so cause a rise in levels of the free active endogenous oestrogens. A range of ED effects have been shown in mammals, fish, birds, reptiles, amphibia and aquatic invertebrates but it is not yet clear whether these processes also occur in human beings. It is evident that EDs, as well as altering reproduction, can cause changes in neurosteroid levels and so have the potential to affect immune function, behaviour and memory. This may be of long-term concern since traces of EDs such as plasticisers, brominated fire retardants, sunscreen agents and cosmetic ingredients are widely distributed in the environment and in human biofluids.