Bioreductive activation of quinones: A mixed blessing

Quinones can be metabolized by various routes: substitution or reductive addition with nucleophilic compounds (mainly glutathione and protein thiol groups), one-electron reduction (mainly by NADPH: cytochrome P-450 reductase) and two-electron reduction (by D,T-diaphorase). During reduction semiquinone radicals and hydroquinones are formed, which can transfer electrons to molecular oxygen, resulting in the formation of reactive oxygen intermediates and back-formation of the parent quinone (redox cycling). Reaction of semiquinones and reactive oxygen intermediates with DNA and other macromolecules can lead to acute cytotoxicity and/or to mutagenicity and carcinogenicity. The enhanced DNA-alkylating properties of certain hydroquinones are exploited in the bioreductive alkylating quinones. Acute cytotoxicity of quinones appears to be related to glutathione depletion and to interaction with mitochondria and subsequent disturbance of cellular energy homoeostasis and calcium homoeostasis. These effects can to a certain extent be predicted from the electron-withdrawing and electron-donating effects of the substituents on the quinone nucleus of the molecule. Prediction of cytostatic potential remains much more complicated, because reduction of the quinones and the reactivity of the reduction products with DNA are modulated by the prevailing oxygen tension and by the prevalence of reducing enzymes in tumour cells.This article is based on a lecture given at the 16th LOF Symposium, 27 October 1989, Utrecht, the Netherlands.     

Fluconazole compared with ketoconazole for the treatment of Candida esophagitis in AIDS. A randomized trial.

OBJECTIVE: To determine the clinical and endoscopic response of candida esophagitis to antifungal therapy and to compare the two oral antifungal agents, fluconazole and ketoconazole. DESIGN: Multicenter, randomized, double-blind trial. SETTING: Fifteen U.S. centers including university, private practice, and county hospital settings. PATIENTS: A total of 169 patients with the acquired immunodeficiency syndrome (AIDS); odynophagia, dysphagia, or retrosternal pain; white esophageal plaques at endoscopy; and pseudohyphae on esophageal brushings or biopsies. INTERVENTION: Patients were randomly assigned to fluconazole (100 mg/d) or ketoconazole (200 mg/d). Doses were doubled at week 1 or 2 if no symptomatic improvement had occurred during the preceding week. Therapy was continued for 2 weeks after resolution of symptoms or for a maximum of 8 weeks. MEASUREMENTS: Patients were clinically evaluated weekly, and laboratory tests were done every 2 weeks. Endoscopy was repeated within 5 days after the end of therapy. RESULTS: A total of 143 patients were clinically evaluable (assessed within 7 days after therapy), and 129 patients were endoscopically evaluable (endoscopy repeated after therapy). Endoscopic cure occurred in 91% of patients treated with fluconazole and in 52% of those given ketoconazole for a difference of 39% (95% Cl, 24% to 52%; P less than 0.001). Esophageal symptoms resolved in 85% of fluconazole-treated patients and in 65% of ketoconazole-treated patients for a difference of 20% (Cl, 6% to 34%; P = 0.006). Intention-to-treat analyses also yielded statistically significant differences for the comparisons listed above. Side effects were minimal and comparable in the two groups; only one patient in each group had therapy discontinued for adverse effects that were possibly related to the study medications. CONCLUSIONS: Fluconazole is associated with significantly greater rates of endoscopic and clinical cure than ketoconazole in patients with AIDS and candida esophagitis. Both drugs appear to be safe and well tolerated.     

Decarboxylation of L-dopa in the rat isolated vascularly perfused small intestine: contribution to systemic elimination and dose-dependent first pass effect.

The contribution of the rat small intestine to systemic and presystemic elimination of L-dopa was studied. When L-dopa was administered into the vascular perfusate, a systemic extraction ratio of 0.38 was found, the major part being decarboxylated to dopamine. The intestinal L-dopa clearance was estimated to be 17.1 mL min-1 kg-1. Thus, L-dopa intestinal clearance in rat represents up to at least 20% of the total body clearance. After luminal administration of L-dopa 83-88% of the administered dose was absorbed within 60 min. The total amount of L-dopa appearing in the vascular perfusate increased more than proportionally to the increase in the dose. In contrast, the amount of dopamine increased less than proportionally to the dose. As a result, the intestinal first pass appeared to be strongly dose-dependent. Since the total percentage absorbed from the lumen was independent of the administered dose and the total amount that appeared in the vascular perfusate increased linearly with the dose, the dose dependency was probably due to saturation of intestinal L-dopa decarboxylation.     

Inhalation of the Phosphodiesterase-3 Inhibitor Milrinone Attenuates Pulmonary Hypertension in a Rat Model of Congestive Heart Failure

Background: Most patients with congestive heart failure (CHF) develop pulmonary venous hypertension, but right ventricular afterload is frequently further elevated by increased pulmonary vascular resistance. To investigate whether inhalation of a vasodilatory phosphodiesterase-3 inhibitor may reverse this potentially detrimental process, the authors studied the effects of inhaled or intravenous milrinone on pulmonary and systemic hemodynamics in a rat model of CHF.

Methods: In male Sprague-Dawley rats, CHF was induced by supracoronary aortic banding, whereas sham-operated rats served as controls. Milrinone was administered as an intravenous infusion (0.2-1 [mu]g [middle dot] kg body weight-1 [middle dot] min-1) or by inhalation (0.2-5 mg/ml), and effects on pulmonary and systemic hemodynamics and lung water content were measured.

Results: In CHF rats, intravenous infusion of milrinone reduced both pulmonary and systemic arterial blood pressure. In contrast, inhalation of milrinone predominantly dilated pulmonary blood vessels, resulting in a reduced pulmonary-to-systemic vascular resistance ratio. Repeated milrinone inhalations in 20-min intervals caused a stable reduction of pulmonary artery pressure. No hemodynamic effects were detected when 0.9% NaCl was administered instead of milrinone or when milrinone was inhaled in sham-operated rats. No indications of potentially adverse effects of milrinone inhalation in CHF, such as left ventricular volume overload, were detected. Moreover, lung edema was significantly reduced by repeated milrinone inhalation.     

Hepatic,intestinal and renal transport of 1-naphthol-β-d-glucuronide in mutant rats with hereditary-conjugated hyperbilirubinemia

Summary Recently, a mutant rat strain was described with a genetic defect for the biliary excretion of organic anions (TR^– rats). To determine the possible heterogeneity of the transport systems in liver, intestine and kidney we investigated the transport of the anion 1-naphthol--d-glucuronide (1-NG) in isolated vascularly perfused organ preparations of the rat liver, intestine and kidney of both Wistar rats and TR^– rats. 1-NG was administered as such (liver and kidney experiments) or formed intracellularly from 1-naphthol (1-N) (liver and gut experiments). Independent of the type of exposure to 1-NG, the biliary excretion was considerably impaired in TR^– rats. In the intestine the total appearance and the vascular/luminal distribution pattern of 1-NG were not significantly different from the values in control rats. Furthermore, no significant disturbance was found with respect to the renal clearance of 1-NG in the TR^– rat when compared with the Wistar rat. Thus, the genetic defect in the TR^– rat is restricted to an impaired hepatobiliary excretion of 1-NG and does not affect the excretory systems of the intestine and kidney. These results suggest that the excretion of 1-NG by the liver, intestine and kidney involves distinct organ-specific transport systems.     

Absorption and presystemic glucuronidation of 1-naphthol in the vascularly fluorocarbon emulsion perfused rat small intestine

Summary Using the isolated vascularly fluorocarbon emulsion perfused rat small intestine some factors which determine the extent of the intestinal glucuronidation of 1-naphthol to 1-naphthol--d-glucuronide were studied. Increasing the luminal 1-naphthol concentration resulted in a concomitant increase in the 1-naphthol appearance in the vascular perfusate. In contrast, the total appearance of 1-naphthol--d-glucuronide increased less than proportional to the increase in the luminal 1-naphthol concentration. About 88% of the total amount of 1-naphthol--d-glucuronide excreted was released into the vascular perfusate. The capacity-limited intestinal glucuronide efflux is most likely due to saturation of the excretory mechanism for 1-naphthol--d-glucuronide. Decreasing the vascular flow rate influenced both the appearance of 1-naphthol and 1-naphtol--d-glucuronide in the vascular perfusate, whereas the appearance of 1-naphthol--d-glucuronide in the luminal perfusate was essentially flow-independent. A noradrenaline-induced change in the haemodynamic state of the vascular bed (with the total flow kept constant) resulted in a marked decrease in the 1-naphthol vascular concentration. The vascular 1-naphthol--d-glucuronide concentration was only slightly affected. These results indicate that changes in blood flow and blood flow distribution within the intestinal wall can affect the extent of presystemic intestinal metabolism by interfering with the absorption of the parent compound and the efflux of formed conjugates. These parameters can be of paramount importance for causing variable intestinal first-pass effects of drugs in vivo. Send offprint requests to M. H. de Vries at the above address     

Distinct pattern of antibody reactivity with oligomeric or polymeric forms of the capsular polysaccharide of Haemophilus influenzae type b.

The chain length of oligosaccharides required for antibody binding has been studied by using the capsular polysaccharide from Haemophilus influenzae type b or oligosaccharides derived from it. The concentration of competing antigens required to achieve a 50% inhibition of antibody binding by human polyclonal antisera in an in vitro competition enzyme-linked immunosorbent assay decreased progressively from greater than 10(-3) to 5 x 10(-7) M as the inhibiting saccharide chain length increased from 1 to 262 repeat units. Even small oligosaccharides (one or two repeat units) are potentially capable of competing to a significant level if a high enough concentration of saccharides is used. A similar pattern of reactivity was seen with a monoclonal anti-polyribosyl ribitol phosphate antibody, suggesting that the differences in the avidity of the antibody subpopulations in the polyclonal antisera do not contribute to the binding patterns observed. The binding reaction was specific as evaluated with pneumococcal saccharides. Furthermore, an oligosaccharide-protein conjugate binds antibody better than the free oligosaccharides do. Such a difference in binding was not observed between the polysaccharide and a polysaccharide-protein conjugate. Overall, the data suggest that identical epitopes are expressed by oligomeric and polymeric forms of the antigen and that a particularly more stable conformation in polysaccharides is preferred by antibodies. Covalent coupling of oligomers to protein increases the expression of stable conformation of epitopes. The data further suggest that this kind of antigenic analysis may be important for the design and synthesis of glycoconjugate vaccines.     

Oligonucleotide typing is a perfect tool to identify antigens stimulatory in the mixed lymphocyte culture

An important criterion for the selection of donors for bone marrow transplantation is the grade of matching for HLA between donor and recipient. For patients that lack an HLA-identical sibling, an extending pool of unrelated volunteers for bone marrow donation is available. From these donors the best matched candidate can be selected by serological typing, followed by a mixed lymphocyte culture (MLC).Oligonucleotide genotyping for HLA class II antigens is considered to be valuable for the prediction of MLC reactivity. We investigated whether this typing method, in combination with serological typing, would cover the recognition of all MLC stimulatory determinants. One hundred thirty-six combinations of HLA-A, -B, and -DR serologically identical individuals were tested in the MLC. Additional typing for HLA-DRB and HLA-DPB by oligonucleotide genotyping made it possible to evaluate the influence of these genes on MLC reactivity. Combinations that were matched for HLA-DRB gave significantly lower responses than those that were mismatched. Nevertheless, in the matched combinations responses were observed to 94% relative response index. These responses could all be attributed to HLA-DP, since all combinations that were identical by HLA-DPB genotyping were negative in the MLC. In conclusion, with the combined use of serology and oligonucleotide genotyping, responder-stimulator combinations can be selected that are identical for all MLC stimulatory determinats. 245 (1991)     

Activation of neutrophil migration by dioctanoyl-sn-glycerol and fMet-Leu-Phe is controlled by different pathways

Migration activated by fMet-Leu-Phe is inhibited by GTP[S] and is little affected by protein kinase C inhibitors. We investigated the effects of GTP[S] and the protein kinase C inhibitor AMG-C₁₆ on dioctanoyl-sn-glycerol (DiC8)-activated migration of rabbit neutrophils and compared them with the effects on fMet-Leu-Phe-activated migration and random migration. GTP[S] did not inhibit DiC8-activated migration or random migration but inhibited fMet-Leu-Phe-activated migration. AMG-C₁₆ gave a strong inhibition of DiC8-activated migration but had only a small effect on fMet-Leu-Phe-activated migration and random migration. When fMet-Leu-Phe and DiC8 were added together in suboptimal concentrations an additive effect was found. Pretreatment with the diacylglycerol kinase inhibitor R59022 enhanced random migration. The enhancement was completely inhibited by AMG-C₁₆ and was unaffected by GTP[S]. These findings suggest that DiC8-activated migration and fMet-Leu-Phe-activated migration are controlled by different pathways.