Improved outcome for haemodialysis patients treated with high-flux membranes.

Improved survival for haemodialysis patients has been reported for synthetic, high-flux biocompatible membranes. The reported data fail to answer the question whether improved survival is related to an effect of enhanced biocompatibility or to increased clearance of larger molecular species of putative uraemic toxins. A retrospective analysis of 715 patients treated by continuous haemodialysis for up to 5 years was undertaken. Low-flux polysulfone dialysis was used exclusively for 252 patients and 463 patients were exposed for at least 3 months to high-flux polysulfone dialysis. Patients treated with high-flux dialysis had a lower mortality (21 vs 36 per 1000 years) and significantly lower standardized mortality ratio. For non-diabetic patients the 5-year probability of survival was significantly greater for high-flux patients (Kaplan-Meier: 92% vs 69%; P=0.036). High-flux dialysis significantly reduced the adverse effect of age on survival. In a Cox proportional hazard model membrane flux (high vs low) was one of the covariates with strong predictor value for reduction of death risk in non-diabetic patients. Although other variables may explain the better survival of patients exposed to high-flux dialysis the data reported here suggest that higher membrane flux, implying higher clearance of larger molecular species and independent of biocompatibility, is associated with improved survival for haemodialysis patients.

Meta-analysis of studies of the CYP2D6 polymorphism in relation to lung cancer and Parkinson's disease.

Studies of associations between the CYP2D6 polymorphism and susceptibility to specific diseases, particularly lung cancer and Parkinsonism, have produced conflicting results with respect to an under or overrepresentation of poor metabolizers. Accordingly, we have re-evaluated this primary research (18 studies on lung cancer and 18 on Parkinsonism) using meta-analysis. For lung cancer, the median odds ratio (OR) was 0.69 (95% confidence interval (CI) 0.52-0.90), which differed significantly from unity (P < 0.007). A trail comprising 3000 patient and an equal number of control individuals would be required to demonstrate that this observation had arisen purely by chance (i.e. OR = 1). For Parkinson's disease, the analysis gave an OR of 1.32 (95% CI 0.98-1.78), which was of borderline statistical significance (P < 0.074). If the only individual study that was statistically significant was excluded, the P-value increased greatly to 0.489. A study of at least 500 patients and an equal number of control individuals giving the same value as the current mean OR of 1.32 would be required to make the overall analysis statistically significant. In summary, poor metabolizers with respect to CYP2D6 show a small decrease in susceptibility to lung cancer compared with extensive metabolizers and its is hard to justify further studies. The relationship between the CYP2D6 polymorphism and lung cancer, as a determinant of individual susceptibility, is not appreciable (OR = 0.69) compared with that between smoking and lung cancer (OR > 11). Nevertheless, the epidemiological impact on the number of poor metabolizers who are protected from lung cancer may be considerable. With regard to Parkinson's disease, additional well designed studies may allow a definitive conclusion, although any risk for poor metabolizers is likely to be small and therefore of questionable clinical significance. An important lesson from the current review of studies is that much time, effort, expense and patient inconvenience might have been avoid if more attention had been paid to appropriate study design particularly in the selection of control groups.

Low seroconversion for hepatitis C virus (HCV) antibody achieved by universal precautions alone.

Since the cloning of Hepatitis C virus in 1988 positive serology for HCV antibody in haemodialysis populations has been reported at varying rates of 2 to 70%. To date there is no consensus regarding strategies which would curtail spread of HCV in the dialysis unit. In our satellite dialysis programme we implemented Universal Precautions as the sole strategy for containment and HCV monitored closely the outcome of this decisions.

Clinical manifestations of carcinoid disease.

Carcinoid tumors are relatively uncommon tumors and their presentation is varied. For these reasons, a high index of suspicion is necessary in order to consider the diagnosis. It is important to separate the "syndrome" from the primary tumor. It is obviously more effective to diagnose the tumor itself before the syndrome manifests itself, usually as a result of metastatic disease. Since the tumors are characteristically slow-growing, the physician may be misled into thinking the patient has functional problems rather than a tumor. Some data and guidelines are given for focusing on the signs and symptoms of carcinoid disease.

Octreotide therapy in carcinoid disease.

Octreotide therapy is expensive, but at present it and other somatostatin analogues appear to offer the best opportunity of controlling the symptoms of flushing and diarrhoea. It may also have other properties affecting general well-being. The question of whether it changes tumour growth remains unanswered and there is no convincing evidence that it alters survival. In all published studies the numbers of patients are small and there have been no control groups. However, since no other drug has yet proved effective against flushing, the somatostatin analogues, including octreotide, remain the treatment of choice for the symptomatic control of the carcinoid syndrome. Octreotide is of great therapeutic value pre-operatively and intra-operatively and it is essential that all operating theatres have this drug available for immediate use. Surgical debulking, if feasible, provides the best outcome potential in carcinoid disease. Present evidence suggests that the place of octreotide and other somatostatin analogues is in controlling the symptoms of the disease rather than its progress and in ensuring cardiovascular and respiratory stability during surgical procedures.

The influence of human variability upon food safety legislation.

This paper considers the influence of human variations in the kinetics and dynamics of chemicals in food upon the formulation of food safety legislation. Legislation is designed to protect and benefit all and not solely specific groups or individuals within the population. A consideration of the adequacy of the safety factors used to calculate Acceptable Daily Intake (ADI) leads on to the discussion of those circumstances in which a part of the population may exceed the ADI for a particular chemical. The conclusion is that the ADI does take human variability into consideration and that current legislation is satisfactory and made more so through the surveillance of dietary additive and contaminant intakes in man.

The demethylenation of methylenedioxymethamphetamine ("ecstasy") by debrisoquine hydroxylase (CYP2D6).

The metabolism of methylenedioxymethamphetamine (MDMA, "ecstasy") was examined in a microsomal preparation of the yeast Saccharomyces cerevisiae expressing human debrisoquine hydroxylase, CYP2D6. Only one product, dihydroxymethylamphetamine (DHMA), was detected in the incubation mixture, and this product accounted for all of the substrate consumption at low concentration (10 microM). Mean +/- SD values of apparent Km(microM) and Vmax (nmol/min per nmol P450) for the demethylenation of (+) and (-)-MDMA at low concentrations (1-100 microM) were 1.72, 0.12 and 6.45, 0.10 and 2.90, 0.10 and 7.61, 0.06, respectively. At high concentrations (> 1000 microM) substrate inhibition was noted, with Ki values of 14.2 and 28.2 mM, respectively, for the (+) and (-) enantiomers. Incubation of MDMA isomers with human liver microsomes indicated that their demethylenation is deficient in the poor metabolizer phenotype. Thus, MDMA is converted to the catecholamine DHMA by CYP2D6, and this may give rise to genetically-determined differences in toxicity.

Beeturia and the biological fate of beetroot pigments.

Beeturia, the passage of pink or red urine after the ingestion of beetroot, is said to occur in 10-14% of the population, and is more common in iron deficiency and malabsorption. A specific HPLC assay for betacyanins, the red beetroot pigments, in biological fluids was developed to study the prevalence of this apparent polymorphism in humans, and to investigate its basis in rats. Two major peaks were observed in chromatograms of extracts of unpickled beetroot. They had identical UV absorption spectra (lambda max = 535 nm) by diode array analysis, and mass spectrometry indicated that one (betacyanin 1) was betanin or its epimer and the other (betacyanin 2) a disaccharide of betacyanin 1. In a population of 100 normal subjects the 0-8 h urinary recoveries after an oral dose of 60 mg beetroot extract were 0.06-0.54% for betacyanin 1 and 0.01-0.6% for betacyanin 2. The distributions of these data were skewed but not clearly bimodal by visual inspection or by kernel density analysis. Four subjects produced visibly red urine and had betacyanin recoveries at the upper end of the population range. Studies using in situ isolated perfused rat jejunum and liver preparations indicated a negligible absorption of the pigments after 1 h and no detectable metabolism or biliary secretion. Intact anaesthetized rats given i.v. bolus doses of beetroot extract cleared both betacyanins from plasma at the rate of 3.3 +/- 0.9 (SD) ml min-1 (n = 5). The total urinary recovery of both pigments amounted to 80% of the dose, and their renal clearances approached their plasma clearances. These data suggest that beeturia does not arise from deficiencies in hepatic metabolism or renal excretion of betacyanins. After oral administration of beetroot extract to rats the betacyanin content of the stomach decreased rapidly with time but neither the intestines nor the bile duct were stained visibly red. These findings together with those showing instability of the betacyanins in acid conditions suggest that variability in the biological fate of beetroot pigments may be determined largely by gastric pH and emptying rate.

Catalytic activities of human debrisoquine 4-hydroxylase cytochrome P450 (CYP2D6) expressed in yeast.

A 1.57kb BamH1 fragment containing a full-length human debrisoquine 4-hydroxylase cytochrome P450 (CYP2D6) cDNA was inserted into the BglII site of the yeast expression plasmid pMA91 and the resulting recombinant plasmid, PELT1, introduced into Saccharomyces cerevisiae strain AH22. Microsomes prepared from AH22/pELT1 cells gave an absorption maximum at 448 nm and a P450 content of 67 +/- 31 pmol/mg of microsomal protein. No P450 was detectable in microsomes prepared from AH22/pMA91 control cells. A western blot of microsomes prepared from yeast transformed with pELT1 were probed with a monoclonal antibody to CYP2D6 and revealed a strong band with a molecular mass consistent with that of CYP2D6 from human liver microsomes. No corresponding band was observed with microsomes from control yeast transformed with pMA91 alone. Microsomes from AH22/pELT cells showed catalytic activity towards metoprolol (alpha-hydroxylation and O-demethylation, 0.17 and 0.78 nmol/mg protein/h, respectively); and towards sparteine (2- and 5-dehydrogenation, 1.82 and 0.59 nmol/mg protein/h, respectively). The inhibition of metoprolol metabolism by quinidine (Qd) was 200 times more potent than that of quinine (Qn), both for alpha-hydroxylation (Qd IC50 = 0.05 microM; Qn IC50 = 4 microM) and O-demethylation (Qd IC50 = 0.05 microM; Qn IC50 = 4 microM). Negligible metabolism of tolbutamide and S-mephenytoin, substrates of the 2C sub-family, and of p-nitrophenol, a substrate of CYP2E1, was detected, although a trace of the N-deethylated metabolite of lignocaine, thought to be metabolised by CYP3A4, was detected with microsomes from CYP2D6-expressing yeast cells. The results indicate that yeast cells containing human CYP2D6 cDNA express a functionally active form of the enzyme, the immunochemical and catalytic properties of which are consistent with those of human liver.

Evidence for a dissociation in the control of sparteine, debrisoquine and metoprolol metabolism in Nigerians.

The 0-8 hour urinary distributions of the metabolic ratios of sparteine (100 mg), debrisoquine (10 mg) and metoprolol (100 mg) were measured in 165 healthy, unrelated, black Nigerian medical students. There was a weak correlation (rs = 0.51, p < 0.001; n = 82) between the metoprolol/alpha-hydroxymetoprolol (M/HM) and the sparteine/total (2- + 5-) dehydrosparteine (S/DHS) ratios. No significant correlations were found between the debrisoquine/4-hydroxydebrisoquine (D/HD) and M/HM ratios (rs = 0.16, n = 33) and between the D/HD and S/DHS ratios (rs = 0.31, n = 38). Both visual inspection and kernel density analysis of the data suggested the presence of two phenotypic groups for sparteine oxidation, with 4% of the population studied being putative poor metabolizers. In contrast biomodality was not apparent in the distribution of the log10M/HM and log10D/HD ratios. These findings provide evidence for a dissociation in the control of metoprolol, sparteine and debrisoquine oxidation in Nigerians and highlight the difficulties in the interpretation of data from pharmacogenetic studies in different ethnic groups.

The expression of human cytochrome P450IA1 in the yeast Saccharomyces cerevisiae.

Data from animal studies suggest that cytochrome P450IA1 catalyses the metabolic activation of several procarcinogenic compounds. In the present study, we have expressed human cytochrome P450IA1 in yeast cells. A 1.70 kb BclI/BamHI fragment containing a full-length human cytochrome P450IA1 cDNA was inserted into the BglII expression site of the yeast expression plasmid pMA91 thereby allowing the ATG initiation codon to be located adjacent to the PGK (phosphoglycerate kinase) promoter. The resulting recombinant plasmid, pCK-1, was introduced into Saccharomyces cerevisiae strains ATCC 44773 and AH22. Microsomes prepared from yeast transformatants of strain ATCC 44773 contained undetectable levels of cytochrome P450. In contrast, microsomes from strain AH22 contained cytochrome P450 with a specific content of 33.3 +/- 10.8 pmol/mg of microsomal protein and showed a reduced carbon monoxide difference spectrum with a peak at 448 nm. Control yeast cells transformed with pMA91 showed no cytochrome P450. Western blots were carried out using an antibody that reacts against rat cytochrome P450IA1 and an antibody that reacts against a synthetic peptide representing a short sequence of human cytochrome P450IA1. A band with a molecular weight of 54 kD was observed in microsomes of yeast transformed with pCK-1, but not with pMA91. When microsomes from yeast transformed with pCK-1 were incubated with benzo(a)pyrene (10 min, 10-160 microM), an estimated Km value of 7 microM was obtained. The availability of yeast cells with functionally active human cytochrome P450IA1 will facilitate molecular structure-activity studies of procarcinogen and drug metabolism by this enzyme in man.

Backtracking booze with Bayes--the retrospective interpretation of blood alcohol data.

1. A Bayesian method is described which allows the explicit estimation of errors produced in estimating drug concentrations at times for which samples are not available for analysis. 2. This method was applied to the problem of 'backtracking' alcohol concentrations for medico-legal purposes. 3. Computer simulation allowed the effect of continuing alcohol absorption on the position and range of estimates of alcohol concentrations to be studied.