Geoffrey Edwards

Thirty-month follow-up of sub-optimal responders to multiple treatments with ivermectin, in two onchocerciasis-endemic foci in Ghana.

Abstract The pathogenesis of the sub-optimal response of Onchocerca volvulus to ivermectin was investigated in a 30-month follow-up of 28 individuals who, in a previous study, had been found to show a sub-optimal (N = 15) or adequate response (N = 13) to multiple treatments with the drug. Verbal informed consent was obtained before each subject was given a general clinical and ocular examination. Skin snips were taken from both iliac crests and both calves. Seventeen nodule carriers were hospitalized for nodulectomy. Adult worms were harvested, embryogrammes were constructed and all developmental stages were counted; degenerate, stretched microfilariae were noted separately. All the subjects were in good general health and all except one had received at least one additional treatment with ivermectin since the earlier study. A large proportion of the adult female worms in 10 out of the 11 sub-optimal responders who were nodule carriers were in full embryonic production but most of the stretched microfilariae they carried were degenerate. This picture is similar to that found in adult worms exposed to the first dose of ivermectin. In one subject who had no viable worms in his nodules, the existence of occult but actively reproductive worms was inferred from the high level of microfilaridermia observed less than 12 months after treatment. These observations confirm the existence of populations of adult female O. volvulus that respond poorly to repeated doses of ivermectin. The use of suramin in the treatment of the sub-optimal responders is discussed.

Clinical Pharmacokinetics of Anthelmintic Drugs

SummaryA rational strategy for chemotherapy demands that dosage schedules be based on an adequate knowledge of clinical and biochemical pharmacology. Many anthelmintic drugs (e.g. suramin, diethylcarbamazine, hycanthone) were introduced before modern techniques for drug evaluation (controlled clinical trials) and before the development of specific and sensitive analytical methods for the assay of drugs and metabolites in biological fluids. Thus, many of the regimens used today for the treatment of parasitic diseases are largely empirically derived. By means of specific analytical methodology (high performance liquid chromatography, gas chromatography and mass-spectrometry) introduced in the 1960s, it is now possible to measure drugs and their metabolites with specificity and sensitivity.Much of this review deals with compounds which are active against the major systemic helminths, i.e., filariae (diethylcarbamazine, ivermectin and suramin) and schistosomes (niridazole, metrifonate, oxamniquine and praziquantel), but recent advances in the treatment of hydatid disease involving the benzimidazole carbamates albendazole and mebendazole are also discussed.Among the imidazole derivatives, mebendazole, a broad-spectrum anthelmintic, is poorly absorbed from the gastrointestinal tract after a therapeutic dose, but that fraction which is absorbed and escapes hepatic first-pass extraction is pharmacologically active against systemic helminths. Albendazole is more completely absorbed, but is almost undetectable in plasma due to its rapid conversion to an active sulphoxide metabolite. This compound may well become the drug of choice for the chemotherapy of echinococcosis. Levamisole, the 1-isomer of tetramisole, is rapidly and completely absorbed, but has not been widely used in systemic helminthiases because of severe side effects associated with prolonged dosage. Diethylcarbamazine is microfilaricidal against Onchocerca volvulus, but its use has been associated with major adverse effects resulting from its action on the microfilariae. These effects are related to the concentration of the drug in the plasma which, in turn, is influenced by urinary pH. The elimination half-life of diethylcarbamazine is prolonged and renal clearance reduced in alkaline urine. Under these conditions the microfilaricidal effect is enhanced, but the adverse reactions to treatment are more severe.Suramin is the only available antifilarial agent with macrofilaricidal activity. It has a long elimination half-life (36 to 54 days), and is highly (99.7%) bound to plasma protein which limits its removal from the blood. Ivermectin is a macrocyclic lactone with microfilarial activity which persists long after the drug has been eliminated from the plasma, suggesting that its mode of action is not related directly to its concentration in the systemic circulation. Metrifonate is an organophosphate, active against Schistosoma haematobium without major side effects. It is also microfilaricidal against Onchocerca volvulus but produces a greater degree of systemic side effects than diethylcarbamazine. The drug is well absorbed but is transformed into an active compound, dichlorvos, by a non-enzymic process. Both metrifonate and dichlorvos are eliminated rapidly by the kidneys.The schistosomicides niridazole, oxamniquine and praziquantel are all extensively metabolised. Niridazole and praziquantel are highly extracted on the first-pass through the liver. Oxamniquine is metabolised by the enzymes in the gut wall, but is relatively poorly cleared by the liver. All 3 compounds are rapidly eliminated. The presence of intra- and extrahepatic shunts in patients with hepatosplenic schistosomiasis might lead to reduced clearance and increased bioavailability of niridazole and praziquantel, necessitating dosage reductions in these subjects.

Ivermectin: does P-glycoprotein play a role in neurotoxicity?

The macrocyclic lactone ivermectin (Mectizan®) is widely used for the control of human filarial infections, particularly as a donated product for onchocerciasis and lymphatic filariasis. In the case of control of lymphatic filariasis in Africa, it is used in combination with donated albendazole. In areas co-endemic for Onchocerciasis and Loa loa, serious adverse reactions have been observed in patients with apparently high microfilaria counts of Loa loa. Recent findings suggest that the severe central nervous system side effects seen in various vertebrates following ivermectin treatment may be due to an absence of, or functional deficiency in P-glycoprotein. P-glycoprotein is expressed in the apical membrane of brain capillary epithelial cells and is responsible for limiting the brain penetration of a range of compounds. Toxicity of ivermectin in some collie dogs may be explained by a 4-bp deletion mutation of the mdr1 gene resulting in a frame shift, generating stop codons that prematurely terminate synthesis of P-glycoprotein. Additionally, sub-populations of CF-1 identified as expressing reduced levels of P-glycoprotein exhibit increased toxicity to substrates of this transporter. Furthermore, while the traditional view of drug-drug interactions is alteration in drug clearance mediated through a change in hepatic drug metabolism, some of these changes may arise through competition for binding sites on P-glycoprotein in the blood-brain barrier, resulting in reduced extracellular efflux and enhanced CNS toxicity. In conclusion, P-glycoprotein is an integral component of the human blood brain barrier and plays a central role in limiting drug uptake into the brain. Altered expression or function of p-glycoprotein could conceivably allow elevation of brain concentrations of ivermectin and produce severe neurotoxicity. This might arise through a genetic polymorphism in p-glycoprotein or co-administration of ivermectin with a drug or foodstuff that might inhibit this efflux transporter.

The toxicity of artemisinin and related compounds on neuronal and glial cells in culture

The antimalarial drug artemisinin and a number of its derivatives were tested for their effects on proliferation of undifferentiated neuroblastoma Nb2a cells and glioma C6 cells in culture as well as their ability to inhibit neurite outgrowth from Nb2a cells differentiated by removal of serum and addition of dibutyryl cyclic AMP. In the Nb2a and C6 cell cultures, all drugs except desoxyartemisinin significantly inhibited cell proliferation in a dose-related manner with the lowest effective concentration being that of artemisinin at 0.1 microM. Artemether, arteether, artemisinin and dihydroartemisinin also produced a dose-related decrease in the number of neurites/extensions formed by differentiating Nb2a cells, with an effect of dihydroartemisinin at a concentration as low as 1 nM. Desoxyartemisinin had no effect on extension/neurite formation. We propose a potential mechanism for neurotoxicity of artemisinin and its derivatives that involves the endoperoxide bridge which is also known to be necessary for their antimalarial action.

The relative systemic availability of ivermectin after administration as capsule, tablet, and oral solution

SummaryAdministration of 12-mg doses of ivermectin (H2B1a) to 12 healthy volunteers in the form of tablets, capsules, and alcoholic oral solution showed the solution to have approximately twice the systemic availability as either of the solid forms, as evidence both by the maximum concentrations of drug attained in plasma and by the corresponding areas under the plasma concentration vs time curves. However, the two solid formulations showed similar systemic availability.

Onchocerciasis Control: Vision for the Future from a Ghanian perspective

Since 1987 onchocerciasis control has relied on the donation of ivermectin (Mectizan®, Merck & Co., Inc.) through the Mectizan Donation Programme. Recently, concern has been raised over the appearance of suboptimal responses to ivermectin in Ghana – highlighting the potential threat of the development of resistance to ivermectin. This report summarises a meeting held in Ghana to set the research agenda for future onchocerciasis control. The aim of this workshop was to define the research priorities for alternative drug and treatment regimes and control strategies to treat populations with existing evidence of suboptimal responsiveness and define research priorities for future control strategies in the event of the development of widespread ivermectin resistance.

Selective determination, in plasma, of artemether and its major metabolite, dihydroartemisinin, by high-performance liquid chromatography with ultraviolet detection

A sensitive and selective reversed-phase high-performance liquid chromatographic method for the determination of artemether and its major metabolite dihydroartemisinin in plasma has been developed. It involves extraction of plasma with dichloromethane, solid-phase separation of the two analytes and acid decomposition prior to chromatography on a C18 Spherisorb column with a mobile phase of acetonitrile-water (50:50, v/v). Run time is 30 min. The assay satisfies all of the criteria required for use in clinical pharmacokinetic studies.

The co-administration of ivermectin and albendazole - safety, pharmacokinetics and efficacy against Onchocerca volvulus

Abstract A randomized, double-blind, placebo-controlled trial was conducted, to determine whether the co-administration of ivermectin with albendazole is safe and more effective against Onchocerca volvulus than ivermectin alone, and whether a significant pharmacokinetic interaction occurs. Forty-two male onchocerciasis patients received ivermectin (200 mug/kg) alone, albendazole (400 mg) alone or the combination. Safety was determined from the results of detailed clinical and laboratory examinations before treatment, during hospitalization and on day 30. Microfilaricidal efficacy was estimated from the reductions in skin counts between day 0 (pretreatment) and day 30. To determine efficacy against the adult worms, two independent observers examined histology slides prepared from nodules excised on day 180; changes in the skin counts of skin microfilariae between days 30 and 365 provided additional indicators of the level of adulticidal activity. Pharmacokinetic parameters for ivermectin and albendazole sulphoxide were defined over 72 h post-treatment. The co-administration of ivermectin with albendazole did not produce more severe adverse effects than ivermectin alone. Both nodule examiners found that the combination was not macrofilaricidal and that it was not clearly superior to ivermectin alone in the effects on reproductive activity; this was supported by the similar efficacy of the two regimens in the suppression of skin microfilariae. There was no significant pharmacokinetic interaction. Although the co-administration of ivermectin with albendazole appears safe, it offers no advantage over ivermectin alone in the control of onchocerciasis. The combination does not require an alteration in the dosage of either component.

Simultaneous determination of artemether and its major metabolite dihydroartemisinin in plasma by gas chromatography–mass spectrometry-selected ion monitoring

A sensitive, selective, and reproducible GC-MS-SIM method was developed for determination of artemether (ARM) and dihydroartemisinin (DHA) in plasma using artemisinin (ART) as internal standard. Solid phase extraction was performed using C18 Bond Elut cartridges. The analysis was carried out using a HP-5MS 5% phenylmethylsiloxane capillary column. The recoveries of ARM, DHA and ART were 94.9 +/- 1.6%, 92.2 +/- 4.1% and 81.3 +/- 1.2%, respectively. The limit of quantification in plasma was 5 ng/ml (C.V. < or = 17.4% for ARM and 15.2% for DHA). Calibration curves were linear with R2 > or = 0.988. Within day coefficients of variation were 3-10.4% for ARM and 7.7-14.5% for DHA. Between day coefficients of variations were 6.5-15.4% and 7.6-14.1% for ARM and DHA. The method is currently being used for pharmacokinetic studies. Preliminary data on pharmacokinetics showed Cmax of 245.2 and 35.6 ng/ml reached at 2 and 3 h and AUC0-8 h of 2463.6 and 111.8 ngh/ml for ARM and DHA, respectively.

The role of glutathione in the neurotoxicity of artemisinin derivatives in vitro

The role of antioxidants in the neurotoxicity of the antimalarial endoperoxides artemether and dihydroartemisinin was studied in vitro by quantitative image analysis of neurite outgrowth in the neuroblastoma cell line NB2a. Intracellular glutathione concentrations were measured by high performance liquid chromatography with fluorescence detection. Both dihydroartemisinin (1 microM) and a combination of artemether (0.3 microM) plus haemin (2 microM) significantly inhibited neurite outgrowth from differentiating NB2a cells to 11.5 +/- 11.0% (SD) and 19.6 +/- 15.2% of controls, respectively. The inhibition by artemether/haemin was prevented by the antioxidants superoxide dismutase (109.7 +/- 47.8% of control), catalase (107.0 +/- 29.3%) glutathione (123.8 +/- 12.4%), L-cysteine (88.0 +/- 6.3%), N-acetyl-L-cysteine (107.8 +/- 14.9%), and ascorbic acid (104.3 +/- 12.7%). Dihydroartemisinin-induced neurotoxicity was completely or partially prevented by L-cysteine (99.5 +/- 17.7% of control), glutathione (57.9 +/- 23.4% of control), and N-acetyl-L-cysteine (57.3 +/- 9.5%), but was not prevented by superoxide dismutase, catalase, or ascorbic acid. Buthionine sulphoximine, an inhibitor of gamma-glutamylcysteine synthetase, significantly increased the neurotoxic effect of non-toxic concentrations of artemether/haemin (0.1 microM/2 microM) and dihydroartemisinin (0.2 microM), suggesting that endogenous glutathione participates in the prevention of the neurotoxicity of artemether/haemin and dihydroartemisinin. Artemether/haemin completely depleted intracellular glutathione levels, whereas dihydroartemisinin had no effect. We conclude that although glutathione status is an important determinant in the neurotoxicity of endoperoxides, depletion of glutathione is not a prerequisite for their toxicity. This is consistent with their mechanisms of toxicity being free radical-mediated damage to redox-sensitive proteins essential for neurite outgrowth, or alteration of a redox-sensitive signalling system which regulates neurite outgrowth.