Susan M. Pond

Conversion of irinotecan (CPT-11) to its active metabolite, 7-ethyl-10-hydroxycamptothecin (SN-38), by human liver carboxylesterase

We have investigated the conversion of the novel anti-topoisomerase I agent CPT-11 (irinotecan; 7-ethyl-10[4-(1-piperidino)-1-piperidno]carbonyloxycamptothecin ) to its active metabolite, SN-38 (7-ethyl-10-hydroxycamptothecin), by human liver carboxylesterase (HLC). Production of SN-38 was relatively inefficient and was enzyme deacylation rate-limited with a steady-state phase occurring after 15-20 min of incubation. This later phase followed Michaelis-Menten kinetics with an apparent Km of 52.9 +/- 5.9 microM and a specific activity of 200 +/- 10 mumol/sec/mol. However, the total enzyme concentration estimated from the intercept concentrations of SN-38 was much lower than that estimated directly from the titration of active sites with paraoxon (0.65 vs. 2.0 microM, respectively). Because deacylation rate-limiting kinetics result in the accumulation of inactive acyl-enzyme complex, we postulated that incubation of CPT-11 with HLC would result in an inhibition of the HLC-catalysed hydrolysis of p-nitrophenylacetate (p-NPA), an excellent substrate for this enzyme. Indeed, this was found to be the case although complete inhibition could not be attained. Analysis of possible kinetic schemes revealed that the most likely explanation for the disparity in estimated enzyme concentrations and the incomplete inhibition of p-NPA hydrolysis is that CPT-11 also interacts at a modulator site on the enzyme, which profoundly reduces substrate hydrolysis. Furthermore, loperamide, a drug often used for the treatment of CPT-11-associated diarrhea, was found to inhibit both CPT-11 and p-NPA HLC-catalysed hydrolysis, most likely by a similar interaction. These observations have direct implications for the clinical use of CPT-11.

The association between polymorphisms in the cytochrome P-450 2D6 gene and Parkinson's disease: a case-control study and meta-analysis

We performed a case-control study to investigate the association of the poor metaboliser genotype of the cytochrome P450 2D6 gene with Parkinsons disease (PD). Genotyping was determined by the polymerase chain reaction followed by digestion with restriction enzymes. The poor metaboliser genotype was more frequent in 112 patients with PD than in 206 matched controls (odds ratio 1.7, 95% CI: 0.94-2.45). A meta-analysis of these results together with ten other published studies gave a pooled odds ratio for the poor metaboliser genotype of 1.47 (95% CI: 1.18-1.96, P=0.01). Thus, the poor metaboliser genotype has a small but highly significant association with PD which would be easily missed in small studies. Research now should focus on the mechanism of this association.

THE EPIDEMIOLOGY OF PARKINSON'S DISEASE IN AN AUSTRALIAN POPULATION

A prevalence study of Parkinson’s disease (PD) was conducted in the rural town of Nambour, Australia. There were 5 cases of PD in a study population of 1207, yielding a crude prevalence ratio of 414 per 100,000 (95% confidence interval; 53–775). We performed a separate case-control study involving 224 patients with PD and 310 controls from South East Queensland and Central West New South Wales, to determine which factors increase the risk for PD in Australia. A positive family history of PD was the strongest risk factor for the development of the disease (odds ratio = 3.4; p < 0.001). In addition, rural residency was a significant risk factor for PD (odds ratio = 1.8, p < 0.001). Hypertension, stroke and well water ingestion were inversely correlated with the development of PD. There was no significant difference between patients and controls for exposure to herbicides and pesticides, head injury, smoking or depression. The high prevalence of PD in Nambour may be explained by rural residency. However, the most significant risk factor for PD was a positive family hisotry. This demonstrates the need for improved understanding of the genetic nature of the disease.

Phenytoin-Induced Methadone Withdrawal

Methadone-maintained volunteers experienced moderately severe opiate withdrawal symptoms within 3 or 4 days of beginning phenytoin in therapeutic doses. The area under the methadone plasma concentration-time curve decreased while the ratio of the pyrrolidine-to-metabolite excretion in urine to this area increased significantly. This suggests that phenytoin accelerates methadone metabolism. Methadone dosing adjustments should be anticipated when phenytoin is initiated or discontinued in methadone-maintained patients.

Serum Formate Concentrations in Methanol Intoxication as a Criterion for Hemodialysis

To evaluate the utility of serum formate concentrations, four patients were studied after ingestion of a methanolic copying fluid. All patients were initially intoxicated. Twelve to twenty-four hours later, signs and symptoms included nausea, abdominal pain, hypokalemia, acidosis (three patients), and pathologic ocular findings (two patients). All patients were treated with ethanol and folate. The two patients with ocular signs and acidosis had high serum formate concentrations (75 and 55 mg/dL, respectively). One of the two patients had a high methanol concentration (222 mg/dL) and required hemodialysis; the other patient did not (methanol concentration, 24 mg/dL). In the other two patients without ocular signs, initial formate concentrations were undetectable (limit of detection, 0.5 mg/dL); however, one patient required hemodialysis because the methanol concentration was 72 mg/dL. Formate is the mediator of ocular injury and acidosis. In these patients formate concentrations correlated with the clinical condition but methanol concentrations did not.

Theophylline overdose: Acute single ingestion versus chronic repeated overmedication

Currently available guidelines for managing theophylline intoxication do not distinguish between acute single ingestion and chronic repeated overmedication and do not reliably predict which patients should undergo hemoperfusion. Although hemoperfusion is widely recommended when serum concentrations exceed 40-60 mg/l, many patients with acute overdose tolerate much higher levels without serious toxicity. Because manifestations of toxicity might be dependent on the chronicity of the overdose, the authors retrospectively compared the clinical features of 15 patients with chronic repeated overmedication with those of 27 patients suffering acute single overdose. Patients suffering chronic repeated overmedication developed seizures (7/15) and serious arrhythmias (4/15) with serum levels of 28-70 mg/l. By contrast, only one of 19 patients suffering acute single overdose with peak levels less than 100 mg/l had seizures, and only two of 19 with levels less than 100 mg/l had serious arrhythmias. However, of the eight single-overdose patients with levels over 100 mg/l, seven had seizures and three had serious arrhythmias. Single-overdose patients were easily recognized by the presence of hypotension, hypokalemia, and low serum bicarbonate, features not present in chronic-type patients. Thus, while patients with theophylline overdose caused by chronic repeated overmedication frequently develop seizures and arrhythmias with serum levels of 40-70 mg/l, those with acute single ingestion are highly unlikely to suffer serious complications unless serum levels exceed 100 mg/l. Management of the intoxication, especially selection of patients for hemoperfusion, should be based on whether the overdose is caused by an acute single ingestion or chronic repeated overmedication.

Identification of potentially neurotoxic pyridinium metabolite in the urine of schizophrenic patients treated with haloperidol

Evidence that the parkinsonian inducing agent MPTP is biotransformed to a pyridinium species that selectively destroys nigrostriatal neurons in humans and subhuman primates has prompted studies to evaluate the metabolic fate of the structurally related neuroleptic agent haloperidol. With the aid of a highly sophisticated atmospheric pressure ionspray HPLC/MS/MS assay, unambiguous evidence has been obtained for the presence of the haloperidol pyridinium species in extracts of urine obtained from haloperidol-treated patients and in extracts of NADPH-supplemented human liver microsomal incubation mixtures containing haloperidol. The potential significance of the formation of this putative neurotoxic pyridinium species is considered.

Evaluation of equilibrium constants for antigen-antibody interactions by solid-phase immunoassay: the binding of paraquat to its elicited mouse monoclonal antibody

A procedure is devised for simple graphical evaluation of the association constant for antibody-antigen interactions from data obtained by conventional solid-phase immunoassay for antigen. Application of the method to situations involving a univalent antigen is illustrated by means of ELISA data for the interactions of paraquat with its elicited murine monoclonal antibody, and the Fab fragment derived therefrom. Although the completely general situation with both antibody and antigen multivalent is not amenable to study by the present procedure, the quantitative expression is readily modified to accommodate antigen multivalency provided that the univalent Fab fragment may be substituted for the multivalent antibody (IgG or IgM) as partitioning species in the solid-phase immunoassay.

Failure of haemoperfusion and haemodialysis to prevent death in paraquat poisoning. A retrospective review of 42 patients.

SummaryIn this review the efficacy of haemoperfusion in the treatment of paraquat poisoning is addressed. 42 reports containing sufficient information of paraquat-poisoned patients were evaluated. These reports, from 35 patients reported in the literature and 7 new cases, were chosen for the following reasons: the timed plasma paraquat concentrations were known, patient outcome was known, and details of haemoperfusion were available. In some cases, haemodialysis was also performed. The plasma paraquat concentrations and the specific times post-ingestion were plotted on a contour graph that predicts the probability of survival. Comparison of the predicted probability of survival versus the actual outcome showed that haemoperfusion, single or repeated, did not affect patient survival. None of the patients whose initial plasma concentrations were greater than 3 mg/L paraquat survived, regardless of the time after ingestion that the concentrations were measured, and despite haemoperfusion. Therefore, such patients might not be considered for haemoperfusion because of their uniformly bad prognosis, despite the procedure being used, and because of the morbidity, discomfort and costs associated with it. Clearly, the need for better techniques to remove paraquat and to prevent the consequences of the metabolic effects of the compound are required urgently before the treatment of the paraquat-poisoned patient will be successful.

The Puzzle of Rates of Cellular Uptake of Protein-Bound Ligands

The rates of hepatic and cerebral uptake of physiologically important ligands, such as fatty acids, drugs and dyes, have long been known to be surprisingly high when most ligand is bound to plasma proteins (Baker & Bradley, 1966, Pardridge & Landaw, 1984). The magnitudes of these rates, as well as the unexpected forms of their dependence on protein concentration, have motivated the hypothesis of specific albumin receptors on the hepatocyte (Weisiger, Gollan & Ockner, 1981) helping to translocate the ligand from albumin into the cell; and the hypothesis of a catalytic mechanism of dissociation of ligand from protein at the cellular surface (for example, Baker & Bradley, 1966; Forker & Luxon, 1981). We shall refer to both proposals as facilitation hypotheses. We do not include in this term facilitation of cellular uptake of unbound ligands (see for example Stremmel, Strohmeyer & Berk, 1986).