Evaluation of the therapeutic efficacy of praziquantel against schistosomes in seven countries with ongoing large-scale deworming programs.

The World Health Organization (WHO) recommends periodic assessment of the therapeutic efficacy of praziquantel (PZQ) to detect reduced efficacy that may arise from drug resistance in schistosomes. In this multi-country study (2014), we assessed the therapeutic efficacy of a single oral dose of PZQ (40 mg/kg) against Schistosoma mansoni (Brazil, Cameroon, Ethiopia, Mali, Madagascar and Tanzania), S. haematobium (Cameroon, Ethiopia, Mali, Tanzania and Zanzibar) and S. japonicum (the Philippines) infections in school-aged children, across a total of 12 different trials. Each trial was performed according to the standardized methodology for evaluating PZQ efficacy as described by the WHO. Overall, therapeutic efficacy, measured as the reduction in arithmetic mean of schistosome egg counts following drug administration (egg reduction rate; ERR), was high for all three schistosome species (S. mansoni: 93.4% (95%CI: 88.8-96.8); S. haematobium: 97.7% (95%CI: 96.5-98.7) and S. japonicum: 90.0% (95%CI: 68.4-99.3). At the trial level, therapeutic efficacy was satisfactory (point estimate ERR ≥90%) for all three Schistosoma species with the exception of S. mansoni in Cameroon where the ERR was 88.5% (95%CI: 79.0-95.1). Furthermore, we observed that in some trials individual drug response could vary significantly (wide 95%CI) and that few non-responsive individuals could significantly impact ERR point estimates. In conclusion, these results do not suggest any established reduced efficacy of the standard PZQ treatment to any of the three schistosome species within these countries. Nevertheless, the substantial degree of variation in individual responses to treatment in some countries underpins the need for future monitoring. The reported ERR values serve as reference values to compare with outcomes of future PZQ efficacy studies to ensure early detection of reduced efficacies that could occur as drug pressure continues increase. Finally, this study highlights that 95%CI should be considered in WHO guidelines to classify the therapeutic efficacy of PZQ.

Building the capacity of public health programmes to become data rich, information rich and action rich.

Good quality, timely data are the cornerstone of health systems, but in many countries these data are not used for evidence-informed decision making and/or for improving public health. The SORT IT (Structured Operational Research and Training Initiative) model has, over 8 years, trained health workers in low- and middle-income countries to use data to answer important public health questions by taking research projects through to completion and publication in national or international journals. The D2P (data to policy) training initiative is relatively new, and it teaches health workers how to apply 'decision analysis' and develop policy briefs for policy makers: this includes description of a problem and the available evidence, quantitative comparisons of policy options that take into account predicted health and economic impacts, and political and feasibility assessments. Policies adopted from evidence-based information generated through the SORT IT and D2P approaches can be evaluated to assess their impact, and the cycle repeated to identify and resolve new public health problems. Ministries of Health could benefit from this twin-training approach to make themselves 'data rich, information rich and action rich', and thereby use routinely collected data in a synergistic manner to improve public health policy making and health care delivery.

Des données de bonne qualité et disponibles rapidement sont la pierre angulaire des systèmes de santé, mais dans de nombreux pays ces données ne sont pas utilisées pour les prises de décision fondées sur des preuves et/ou pour améliorer la santé publique. Le modèle SORT IT (Structured Operational Research and Training Initiative) a, en 8 années, formé le personnel de santé des pays à revenu faible et moyen à l'utilisation des données pour répondre à d'importantes questions de santé publique en amenant les projets de recherche jusqu'à leur achèvement et à la publication dans des revues nationales ou internationales. L'initiative de formation D2P (données pour la politique) est relativement nouvelle et forme le personnel de santé à la manière d'appliquer l'analyse de décision et à l'élaboration d'énoncés de politiques à l'intention des décideurs politiques : ceci inclut la description d'un problème et les preuves disponibles, une comparaison quantitative des options de politique qui tiennent compte des impacts prédits en matière de santé et d'économie, et une évaluation de politique et de faisabilité. Les politiques adoptées à partir d'informations basées sur des preuves générées grâce aux approches SORT IT et de D2P peuvent être évaluées en termes d'impact, et le cycle répété afin d'identifier et de résoudre de nouveaux problèmes de santé publique. Les ministres de la santé pourraient bénéficier de cette approche de formation double afin qu'ils soient « riches de données, riches d'information et riches d'action ¼, et donc utiliser les données recueillies en routine d'une manière synergique afin d'améliorer les choix en matière de politique de santé publique et de prestation des soins de santé.

La buena calidad de los datos y su puntualidad constituyen los pilares de los sistemas de salud, pero en muchos países esta información no se utiliza con el fin de orientar la toma de decisiones basadas en la evidencia o mejorar la salud pública. El modelo de Investigación Operativa Estructurada e Iniciativa para la Formación (SORT IT) se ha aplicado durante más de 8 años en la capacitación de los profesionales de salud de países de ingresos bajos y medianos, en materia de aplicación de los datos para resolver importantes preguntas de salud pública, al acompañar los proyectos de investigación hasta su finalización y publicación en revistas de ámbito nacional o internacional. La iniciativa de formación D2P (de los datos a las políticas) es relativamente nueva e instruye a los trabajadores de salud sobre la forma de aplicar el 'análisis decisional' y formular documentos normativos destinados a las instancias decisorias; la iniciativa comprende la descripción de un problema y la evidencia disponible, una comparación cuantitativa de las opciones normativas que tiene en cuenta las repercusiones de salud y económicas previstas y una evaluación política y de factibilidad. Es posible evaluar las políticas adoptadas a partir de información basada en la evidencia científica generada por conducto de las estrategias SORT IT y D2P con el propósito de analizar su repercusión y se puede repetir el ciclo a fin de detectar y resolver nuevos problemas de salud pública. Los ministerios de salud pueden aprovechar este enfoque doble de capacitación con el objeto de alcanzar una situación de 'riqueza de datos, de información y de acción' y aplicar sinérgicamente los datos recogidos de manera sistemática con miras a optimizar la toma de decisiones de salud pública y la prestación de los servicios de salud.

Population pharmacokinetics of intravenous artesunate: a pooled analysis of individual data from patients with severe malaria.

There are ~660,000 deaths from severe malaria each year. Intravenous artesunate (i.v. ARS) is the first-line treatment in adults and children. To optimize the dosing regimen of i.v. ARS, the largest pooled population pharmacokinetic study to date of the active metabolite dihydroartemisinin (DHA) was performed. The pooled dataset consisted of 71 adults and 195 children with severe malaria, with a mixture of sparse and rich sampling within the first 12 h after drug administration. A one-compartment model described the population pharmacokinetics of DHA adequately. Body weight had the greatest impact on DHA pharmacokinetics, resulting in lower DHA exposure for smaller children (6-10 kg) than adults. Post hoc estimates of DHA exposure were not significantly associated with parasitological outcomes. Comparable DHA exposure in smaller children and adults after i.v. ARS was achieved under a dose modification for intramuscular ARS proposed in a separate analysis of children.

Leishmaniasis chemotherapy--challenges and opportunities.

Although there have been significant advances in the treatment of visceral leishmaniasis (VL), there remain challenges to ensure that treatments effective in India are also effective in other regions of the world and to identify treatment for post kala-azar dermal leishmaniasis as well as the opportunity to develop a safe oral short-course treatment. At the same time, there have been few advances for the treatment of simple or complex forms of cutaneous leishmaniasis (CL), other than topical paromomycin formulations. The main challenge for CL is to ensure that this disease is on the research and development agenda, so that new drugs are evaluated or compounds are screened in appropriate models, and that the standardization of quality of clinical trials is guaranteed. Problems also remain in the treatment of HIV/leishmaniasis co-infected patients. We are some way from having the ideal treatments for VL and CL and drug research and development for these diseases must remain focused.

Hypoxia modulates the effect of dihydroartemisinin on endothelial cells.

Artemisinin derivatives, the current cornerstone of malaria treatment, possess also anti-angiogenic and anti-tumor activity. Hypoxia plays a crucial role both in severe malaria (as a consequence of the cytoadherence of infected erythrocytes to the microvasculature) and in cancer (due to the restricted blood supply in the growing tumor mass). However, the consequences of hypoxia onto the effects of artemisinins is under-researched. This study aimed at assessing how the inhibition of microvascular endothelial cell (HMEC-1) growth induced by dihydroartemisinin (DHA, an antimalarial drug and the active metabolite of currently in-use artemisinins) is affected by oxygen tension. Low doses of DHA (achieved in the patients' plasma when treating malaria) were more inhibitory in hypoxia, whereas high doses (required for anti-angiogenic or anti-tumor activity) were more effective in normoxia. The peroxide bridge is essential for cellular toxicity (deoxyDHA was inactive). High doses of DHA caused HMEC-1 apoptosis and G2 cell cycle arrest. Effects were mediated by the generation of oxidative stress as demonstrated by DCF-DA fluorescence and membrane lipid peroxidation analysis. Overall, these results suggest that DHA inhibition of endothelial cell growth is related to the level of tissue oxygenation and drug concentration. This should be considered when studying both the effects of artemisinin derivatives as antimalarials and the potential therapeutic applications of these drugs as anti-tumor agents.

Effect of artesunate and mefloquine in combination on the Fridericia corrected QT intervals in Plasmodium falciparum infected adults from Thailand.

OBJECTIVE: To ascertain whether mefloquine (MQ) produces electrocardiogram (ECG) changes that could be a risk for Torsades de Pointe (TdP), a potentially malignant, ventricular tachyarrhythmia. METHODS: We measured the Fridericia corrected QT (QTcF) intervals on 12 lead ECGs on days (D) 0, 3, 7 in Plasmodium falciparum infected adults, treated with oral artesunate (AS) and MQ as a new fixed dose (n = 25) combination or loose tablets (n = 25) over 3 days. Target total doses were 12 mg/kg of AS and 24-25 mg/kg of MQ. MQ concentrations ([MQ]) were measured by HPLC. RESULTS: All ECG intervals were similar between drug arms and were combined for analysis. Mean QTcF values were 389 (D0), 407 (D3) and 399 (D7) ms (Ps < 0.003 vs. D0); corresponding heart rates and [MQ]s were 83, 67 and 73 beats/minute (Ps ≤ 0.0003 vs. D0) and 0, 3095 and 1721 ng/ml. One male patient (loose arm) had a D3 QTcF 504 ms (D0 406 ms, D7 433 ms). In the modelling of QTcF and JTcF from D0 to D7, significant effects were observed individually for [MQ], temperature and heart rate (HR). The MQ AUC(0-∞) was not a significant factor. Using a manual descending, model building approach to select variables, the HR was the only significant variable (P = 0.001) over time in the model that best explained the changes in the QTcF and JTcF intervals. CONCLUSIONS: In this small group of patients, slowing heart rates due to malaria resolution best explained the observed increases in the QTcF intervals.

Changing species distribution and antimicrobial susceptibility pattern of Shigella over a 29-year period (1980-2008).

We studied changes in species distribution and antimicrobial resistance patterns of Shigella during 1980-2008, using the Diarrhoeal Diseases Surveillance system of Dhaka Hospital of ICDDR,B. In hospitalized patients Shigella prevalence decreased steadily from 8-12% in the 1980s to 3% in 2008. Endemic S. flexneri was the most commonly isolated species (54%). Epidemic S. dysenteriae type 1 had two peaks in 1984 and 1993, but was not found after 2000, except for one case in 2004. The therapeutic options are now limited: in 2008 a total of 33% of S. flexneri were resistant to ciprofloxacin and 57% to mecillinam. In the <5 years age group, severely underweight, wasted and stunted children were more at risk of shigellosis compared to well-nourished children (P<0·001). Although hospitalization for Shigella diarrhoea is decreasing, the high levels of antimicrobial resistance and increased susceptibility of malnourished children continue to pose an ongoing risk.

New fixed-dose artesunate-mefloquine formulation against multidrug-resistant Plasmodium falciparum in adults: a comparative phase IIb safety and pharmacokinetic study with standard-dose nonfixed artesunate plus mefloquine.

A new fixed-dose artesunate (AS)-mefloquine (MQ) was assessed in adults hospitalized for 28 days with uncomplicated drug-resistant falciparum malaria. The patients (n = 25/arm) were treated with (i) two fixed-dose tablets (AS-MQ arm; 100 mg AS-200 mg MQ/tablet) daily for 3 days (days 0, 1, and 2) or (ii) nonfixed AS (AS-plus-MQ arm; 4 mg/kg of body weight/day for 3 days) plus MQ (15 mg/kg on day 1 and 10 mg/kg on day 2), dosed by weight. Clinical laboratory electrocardiogram (ECG), adverse events (AEs), efficacy, and pharmacokinetic parameters were assessed over 28 days. Both regimens were well tolerated. No AEs were drug related. Two serious AEs of malaria-induced hypotension occurring in the AS-MQ arm necessitated rescue treatment. There were no significant changes in hematology, biochemistry, or PR and QRS intervals. For all patients, mean Fridericia-corrected QT intervals were significantly (P < or = 0.0027) prolonged on day 3 (407 ms) and day 7 (399 ms) versus day 0 (389 ms), in parallel with significant (P < or = 0.0003) falls in heart rates (67 [day 3], 73 [day 7], and 83 [day 0] beats/minute). Fixed-nonfixed formulations were bioequivalent for MQ, but not for AS and dihydroartemisinin (DHA). One AS-MQ patient developed a new infection on day 28; his day 28 plasma MQ concentration was 503.8 ng/ml. Fixed-dose AS-MQ was well tolerated, had pharmacokinetic (PK) profiles broadly similar to those of nonfixed AS plus MQ, and is a suitable replacement.

Treatment of urinary schistosomiasis: methodological issues and research needs identified through a Cochrane systematic review.

Guidelines recommend praziquantel (PZQ) for the treatment and control of schistosomiasis, with no real alternative. Metrifonate was still widely used against Schistosoma haematobium in the 1990s, and then withdrawn. Experimental studies and clinical trials suggest that artemisinin compounds are active against S. haematobium. In a Cochrane systematic review assessing the efficacy and safety of drugs for treating urinary schistosomiasis, 24 randomized controlled trials (n=6315 individuals) met our inclusion criteria. These trials compared a variety of single agent and combination regimens with PZQ, metrifonate or artemisinin derivatives. The review confirmed that both the standard recommended doses of PZQ (single 40 mg/kg oral dose) and metrifonate (3x7.5-10 mg/kg oral doses administered fortnightly) are efficacious and safe in treating urinary schistosomiasis, but there is no study comparing these two regimens head-to-head. There is currently not enough evidence to evaluate artemisinin compounds. Most of the studies included in the Cochrane systematic review were insufficiently powered, lacked standardization in assessing and reporting outcomes, and had a number of methodological limitations. In this paper we discuss the implications of these findings with respect to public health and research methodology and propose priority research needs.

The global portfolio of new antimalarial medicines under development.

With the elimination of malaria now considered a realistic goal, it would be useful for scientists and policy makers to have an inventory of the arsenal of antimalarials, current and prospective, that could help make this goal a reality. In order to provide an overview of antimalarial projects in recent clinical development, we review here the global portfolio of antimalarial drugs in clinical phases of development complemented by projects in the preclinical and early discovery phases. The portfolio is discussed in terms of the novelty of the new molecules and their potential health impact in terms of addressing the requirements for the control and eventual eradication of malaria.

A single LC-tandem mass spectrometry method for the simultaneous determination of 14 antimalarial drugs and their metabolites in human plasma.

Among the various determinants of treatment response, the achievement of sufficient blood levels is essential for curing malaria. For helping us at improving our current understanding of antimalarial drugs pharmacokinetics, efficacy and toxicity, we have developed a liquid chromatography-tandem mass spectrometry method (LC-MS/MS) requiring 200mul of plasma for the simultaneous determination of 14 antimalarial drugs and their metabolites which are the components of the current first-line combination treatments for malaria (artemether, artesunate, dihydroartemisinin, amodiaquine, N-desethyl-amodiaquine, lumefantrine, desbutyl-lumefantrine, piperaquine, pyronaridine, mefloquine, chloroquine, quinine, pyrimethamine and sulfadoxine). Plasma is purified by a combination of protein precipitation, evaporation and reconstitution in methanol/ammonium formate 20mM (pH 4.0) 1:1. Reverse-phase chromatographic separation of antimalarial drugs is obtained using a gradient elution of 20mM ammonium formate and acetonitrile both containing 0.5% formic acid, followed by rinsing and re-equilibration to the initial solvent composition up to 21min. Analyte quantification, using matrix-matched calibration samples, is performed by electro-spray ionization-triple quadrupole mass spectrometry by selected reaction monitoring detection in the positive mode. The method was validated according to FDA recommendations, including assessment of extraction yield, matrix effect variability, overall process efficiency, standard addition experiments as well as antimalarials short- and long-term stability in plasma. The reactivity of endoperoxide-containing antimalarials in the presence of hemolysis was tested both in vitro and on malaria patients samples. With this method, signal intensity of artemisinin decreased by about 20% in the presence of 0.2% hemolysed red-blood cells in plasma, whereas its derivatives were essentially not affected. The method is precise (inter-day CV%: 3.1-12.6%) and sensitive (lower limits of quantification 0.15-3.0 and 0.75-5ng/ml for basic/neutral antimalarials and artemisinin derivatives, respectively). This is the first broad-range LC-MS/MS assay covering the currently in-use antimalarials. It is an improvement over previous methods in terms of convenience (a single extraction procedure for 14 major antimalarials and metabolites reducing significantly the analytical time), sensitivity, selectivity and throughput. While its main limitation is investment costs for the equipment, plasma samples can be collected in the field and kept at 4 degrees C for up to 48h before storage at -80 degrees C. It is suited to detecting the presence of drug in subjects for screening purposes and quantifying drug exposure after treatment. It may contribute to filling the current knowledge gaps in the pharmacokinetics/pharmacodynamics relationships of antimalarials and better define the therapeutic dose ranges in different patient populations.

Validation of high performance liquid chromatography-electrochemical detection methods with simultaneous extraction procedure for the determination of artesunate, dihydroartemisinin, amodiaquine and desethylamodiaquine in human plasma for application in clinical pharmacological studies of artesunate-amodiaquine drug combination.

With the expanded use of the combination of artesunate (AS) and amodiaquine (AQ) for the treatment of falciparum malaria and the abundance of products on the market, comes the need for rapid and reliable bioanalytical methods for the determination of the parent compounds and their metabolites. While the existing methods were developed for the determination of either AS or AQ in biological fluids, the current validated method allows simultaneous extraction and determination of AS and AQ in human plasma. Extraction is carried out on Supelclean LC-18 extraction cartridges where AS, its metabolite dihydroartemisinin (DHA) and the internal standard artemisinin (QHS) are separated from AQ, its metabolite desethylamodiaquine (DeAQ) and the internal standard, an isobutyl analogue of desethylamodiaquine (IB-DeAQ). AS, DHA and QHS are then analysed using Hypersil C4 column with acetonitrile-acetic acid (0.05M adjusted to pH 5.2 with 1.00M NaOH) (42:58, v/v) as mobile phase at flow rate 1.50ml/min. The analytes are detected with an electrochemical detector operating in the reductive mode. Chromatography of AQ, DeAQ and IB-DeAQ is carried out on an Inertsil C4 column with acetonitrile-KH(2)PO(4) (pH 4.0, 0.05M) (11:89, v/v) as mobile phase at flow rate 1.00ml/min. The analytes are detected by an electrochemical detector operating in the oxidative mode. The recoveries of AS, DHA, AQ and DeAQ vary between 79.1% and 104.0% over the concentration range of 50-1400ng/ml plasma. The accuracies of the determination of all the analytes are 96.8-103.9%, while the variation for within-day and day-to-day analysis are <15%. The lower limit of quantification for all the analytes is 20ng/ml and limit of detection is 8ng/ml. The method is sensitive, selective, accurate, reproducible and suited particularly for pharmacokinetic study of AS-AQ drug combination and can also be used to compare the bioavailability of different formulations, including a fixed-dose AS-AQ co-formulation.

Anthropometrically derived dosing and drug costing calculations for treating visceral leishmaniasis in Bihar, India.

OBJECTIVE AND METHOD: To estimate drug costs of treating visceral leishmaniasis (VL) based on data on the VL population structure from the high-burden, antimony-resistant area of Northern Bihar, India. RESULTS: Paromomycin is the cheapest option ($7450 to treat 1000 patients). Treating 1000 patients with oral miltefosine would cost $119,250 at the current private market price or $64,383-$75,129 at preferential public sector price depending on the size of the order. With AmBisome it would be $163,600 or $229,500 depending on the dose (10 or 15 mg/kg total). These costs are without considering other direct costs (daily intramuscular injections for 3 weeks for paromomycin; intravenous devices and hospitalization for AmBisome; directly observed treatment if applied for miltefosine) and indirect costs. CONCLUSION: These calculations provide useful basic information for projections.

Dosing accuracy of artesunate and amodiaquine as treatment for falciparum malaria in Casamance, Senegal.

OBJECTIVES: Several products of artesunate plus amodiaquine (AS + AQ) are being deployed in malaria-endemic countries for treating uncomplicated falciparum malaria but dosing accuracy and consequential effects on efficacy and tolerability have not been examined. METHODS: Patients with parasitologically confirmed, uncomplicated falciparum malaria were treated and followed by research teams or local health centre staff in Casamance, Senegal. AS + AQ was given as: (i) loose combination (AS 50 mg, AQ 200 mg), dosed on body weight, or (ii) co-blistered product (AS 50 mg, AQ 153 mg) dosed by weight or age. Target doses were: (i) AS 4 (2-10) mg/kg/day and (ii) AQ 10 (7.5-15) mg/kg/day. Patients receiving therapeutic doses defined dosing accuracy. Treatment-emergent signs and symptoms (TESS) were recorded. RESULTS: A total of 3277 patients were treated with loose (n = 1972, weight-dosed) or co-blistered (n = 1305, 962 age-dosed, 343 weight-dosed) AS + AQ by the research team (n = 966) or clinic staff (n = 2311). AS was dosed correctly in >99% with all regimens. Loose AQ by weight was 98% correct. The co-blister AQ overdosed 18% of patients when dosed by age and underdosed 13% by weight. Low weight was an independent risk factor for overdosing. The co-blister had significantly more TESS than the loose product [117/1305 (9%) vs. 41/1972 (2%), relative risk = 4.3 (95% CI: 3.0-6.1, P < 0.0001)]. Age-based dosing accounted for the difference. TESS occurred mostly within one day (72%) and were mild or moderate (75%). CONCLUSION: Artesunate is easier to dose than AQ. Currently available age-dosed, co-blistered AS + AQ tends to overdose AQ and is less well tolerated than loose tablets. It is not the optimal presentation of AS + AQ.

Pre-referral rectal artesunate to prevent death and disability in severe malaria: a placebo-controlled trial.

BACKGROUND: Most malaria deaths occur in rural areas. Rapid progression from illness to death can be interrupted by prompt, effective medication. Antimalarial treatment cannot rescue terminally ill patients but could be effective if given earlier. If patients who cannot be treated orally are several hours from facilities for injections, rectal artesunate can be given before referral and acts rapidly on parasites. We investigated whether this intervention reduced mortality and permanent disability. METHODS: In Bangladesh, Ghana, and Tanzania, patients with suspected severe malaria who could not be treated orally were allocated randomly to a single artesunate (n=8954) or placebo (n=8872) suppository by taking the next numbered box, then referred to clinics at which injections could be given. Those with antimalarial injections or negative blood smears before randomisation were excluded, leaving 12 068 patients (6072 artesunate, 5996 placebo) for analysis. Primary endpoints were mortality, assessed 7-30 days later, and permanent disability, reassessed periodically. All investigators were masked to group assignment. Analysis was by intention to treat. This study is registered in all three countries, numbers ISRCTN83979018, 46343627, and 76987662. RESULTS: Mortality was 154 of 6072 artesunate versus 177 of 5996 placebo (2.5%vs 3.0%, p=0.1). Two versus 13 (0.03%vs 0.22%, p=0.0020) were permanently disabled; total dead or disabled: 156 versus 190 (2.6%vs 3.2%, p=0.0484). There was no reduction in early mortality (56 vs 51 deaths within 6 h; median 2 h). In patients reaching clinic within 6 h (median 3 h), pre-referral artesunate had no significant effect on death after 6 h or permanent disability (71/4450 [1.6%] vs 82/4426 [1.9%], risk ratio 0.86 [95% CI 0.63-1.18], p=0.35). In patients still not in clinic after more than 6 h, however, half were still not there after more than 15 h, and pre-referral rectal artesunate significantly reduced death or permanent disability (29/1566 [1.9%] vs 57/1519 [3.8%], risk ratio 0.49 [95% CI 0.32-0.77], p=0.0013). INTERPRETATION: If patients with severe malaria cannot be treated orally and access to injections will take several hours, a single inexpensive artesunate suppository at the time of referral substantially reduces the risk of death or permanent disability. FUNDING: UNICEF/UNDP/World Bank Special Programme for Research and Training in Tropical Diseases (WHO/TDR); WHO Global Malaria Programme (WHO/GMP); Sall Family Foundation; the European Union (QLRT-2000-01430); the UK Medical Research Council; USAID; Irish Aid; the Karolinska Institute; and the University of Oxford Clinical Trial Service Unit (CTSU).

Old and new targets for innovative antimalarial compounds: the different strategies of the Italian Malaria Network.

Clinical treatment-failures to affordable drugs encouraged new investigation for discovery and development of new prophylactic and therapeutic interventions against malaria. The Drug Discovery Cluster (DDcl) of the Italian Malaria Network gathers several highly integrated and complementary laboratories from different Italian Institutions to identify, synthesise, screen in vitro and in vivo new antimalarial molecules directed against the intraerythrocytic stage of P. falciparum parasites and/or with transmission blocking activity to select lead compounds for further development. Complementary research activities, both in vitro and in the clinics, aim at investigating the pathogenetic mechanisms of severe malaria anaemia and the different manifestations of the disease in malaria-HIV co-infected patients to identify new therapies and improve survival.

[Visceral leishmaniasis: clinical sensitivity and resistance to various therapeutic agents]. / Leishmaniose viscérale: efficacité clinique et résistances aux différentes molécules.

Visceral leishmaniasis is present in 61 countries but 90% of the 500,000 new cases that arise annually occur in five countries, i.e., India, Bangladesh, Nepal, Sudan, and Brazil. Annual mortality is approximately 59000 cases. Agents based on pentavalent antimony have been the mainstay of treatment for the last 60 years. In recent years, however, clinical resistance to these agents has been reported especially in the state of Bihar in India. Pentamidine and amphotericin B were introduced in the 1950s and 1960s. More recent additions to the therapeutic arsenal include liposomal amphotericin B, miltefosine, and paromomycin. Among these recent molecules, miltefosine, i.e., the only oral agent, appears most vulnerable because it involves long-term treatment and has a long half-life. The main therapeutic problems now being encountered are the emergence of acquired resistance to antimonials, the high cost of treatment, and failure of therapy in immunocompromised patients mainly due to concurrent human immunodeficiency virus (HIV) infection. For eradication initiatives such as the one aimed at eliminating leishmaniasis on the Indian subcontinent, the appearance of drug resistance increases the risk associated to parasite infection and, as for malaria, tuberculosis and HIV infection, raises fears that the problems in the implementation of public health policies will lead to highly refractory forms.

An analytical method with a single extraction procedure and two separate high performance liquid chromatographic systems for the determination of artesunate, dihydroartemisinin and mefloquine in human plasma for application in clinical pharmacological studies of the drug combination.

The combination of two sensitive, selective and reproducible reversed phase liquid chromatographic (RP-HPLC) methods was developed for the determination of artesunate (AS), its active metabolite dihydroartemisinin (DHA) and mefloquine (MQ) in human plasma. Solid phase extraction (SPE) of the plasma samples was carried out on Supelclean LC-18 extraction cartridges. Chromatographic separation of AS, DHA and the internal standard, artemisinin (QHS) was obtained on a Hypersil C4 column with mobile phase consisting of acetonitrile-0.05 M acetic acid adjusted to pH 5.2 with 1.0M NaOH (42:58, v/v) at the flow rate of 1.50 ml/min. The analytes were detected using an electrochemical detector operating in the reductive mode. Chromatography of MQ and the internal standard, chlorpromazine hydrochloride (CPM) was carried out on an Inertsil C8-3 column using methanol-acetonitrile-0.05 M potassium dihydrogen phosphate adjusted to pH 3.9 with 0.5% orthophosphoric acid (50:8:42, v/v/v) at a flow rate of 1.00 ml/min with ultraviolet detection at 284 nm. The mean recoveries of AS and DHA over a concentration range of 30-750 ng/0.5 ml plasma and MQ over a concentration of 75-1500 ng/0.5 ml plasma were above 80% and the accuracy ranged from 91.1 to 103.5%. The within-day coefficients of variation were 1.0-1.4% for AS, 0.4-3.4% for DHA and 0.7-1.5% for MQ. The day-to-day coefficients of variation were 1.3-7.6%, 1.8-7.8% and 2.0-3.4%, respectively. Both the lower limit of quantifications for AS and DHA were at 10 ng/0.5 ml and the lower limit of quantification for MQ was at 25 ng/0.5 ml. The limit of detections were 4 ng/0.5 ml for AS and DHA and 15 ng/0.5 ml for MQ. The method was found to be suitable for use in clinical pharmacological studies.

Effect of artesunate plus sulfadoxine-pyrimethamine on haematological recovery and anaemia, in Kenyan children with uncomplicated, Plasmodium falciparum malaria.

Malaria-associated anaemia is a major public-health problem. Although the treatment of uncomplicated, Plasmodium falciparum malaria aims to clear the parasites, relieve the symptoms and permit haematological recovery, data on the impact of antimalarial treatment on haematological recovery are few. Haematological recovery and the prevalence of anaemia were therefore evaluated in 600 Kenyan children with uncomplicated malaria who were randomly assigned to one of three treatment groups. The children were given sulfadoxine-pyrimethamine (SP) on day 0, SP plus artesunate on day 0 (AS1), or SP on day 0 and artesunate on each of days 0-2 (AS3). Haemoglobin (Hb) concentrations were measured on days 0, 7, 14, 21 and 28, with haematological recovery defined as a day-28 Hb concentration of at least 11 g/dl. Only 96 (18%) of the 543 children who were anaemic (i.e. with <11.0 g Hb/dl) at enrolment achieved haematological recovery. The prevalence of anaemia fell from 91% on day 0 to 74% (252/340) by day 28 (P=0.065). Compared with SP alone, neither artesunate regimen resulted in higher Hb concentrations on day 28 (with means of 10.2, 9.9 and 10.2 g/dl for AS3, AS1 and SP, respectively; P=0.254), a higher frequency of haematological recovery (19%, 14% and 20% for AS3, AS1 and SP, respectively; P=0.301) or a greater reduction in the prevalence of anaemia (prevalences in the AS3, AS1 and SP arms falling from 90%, 89% and 93%, respectively, on day 0, to corresponding values of 71%, 82% and 69% on day 28; P=0.40). In fact, between days 0 and 7, the children in the AS3 arm showed a larger drop in mean Hb than the children in the other two treatment arms. In general, haematological recovery was most likely in older children who had mild anaemia at presentation and were parasitologically cured. Overall, the frequencies of haematological recovery were modest and not influenced by the artesunate treatments. Other factors contributing to anaemia need to be explored more fully.