Evaluation of the application of national malaria treatment guidelines in private pharmacies in a rural area in the Democratic Republic of Congo.

In the Democratic Republic of the Congo, the first recourse in case of suspected malaria in the health system is the private pharmacy sector. This study was therefore designed to assess private provider adherence to national case management guidelines in Kimpese, a rural area of Central Kongo province. A descriptive cross-sectional survey of 103 pharmacies took place in March 2016. The study included 97 pharmacies. The artemether-lumefantrine combination recommended as the first-line treatment for uncomplicated P. falciparum malaria was available in 100% of pharmacies but only 3% stocked quality-assured medicines. The sulfadoxine-pyrimethamine recommended for intermittent preventive treatment of malaria in pregnant women and quinine, which is no longer part of national policy, were widely available (>97.0% of pharmacies). Among providers, fewer than 20% were aware of the national malaria treatment guidelines. The main reasons for non-adherence to national guidelines among private dispensers was the high cost (up to 10 times more expensive than sulfadoxine-pyrimethamine treatment) and adverse effects of artemisinin-based combination therapies. Governmental interventions to improve private sector engagement in implementation of the national guidelines and to prevent the spread of ineffective and non-quality assured antimalarial medicines must be intensified.

Access to artemisinin-based combination therapies and other anti-malarial drugs in Kinshasa.

OBJECTIVE: Artemisinin-based combination therapies have been available since 2005 in the Democratic Republic of the Congo to treat malaria and to overcome the challenge of anti-malarial drug resistance as well as to improve access to effective treatments. The private sector is the primary distribution source for anti-malarial drugs and thus, has a key position among the supply chain actors for a rational and proper use of anti-malarial drugs. We aimed to assess access to nationally recommended anti-malarial drugs in private sector pharmacies of the capital-city of Kinshasa. METHOD: We performed a cross-sectional survey of 404 pharmacies. RESULTS: Anti-malarial drugs were stocked in all surveyed pharmacies. Non-artemisinin-based anti-malarial therapies such as quinine or sulfadoxine-pyrimethamine, were the most frequently stocked drugs (93.8% of pharmacies). Artemisinin-based combination therapies were stocked in 88% of pharmacies. Artemether-lumefantrine combinations were the most frequently dispensed drugs (93% of pharmacies), but less than 3% were quality-assured products. Other non-officially recommended artemisinin-based therapies including oral monotherapies were widely available. CONCLUSION: Artemisinin-based combination therapies were widely available in the private pharmacies of Kinshasa. However, the private sector does not guarantee the use of nationally recommended anti-malarial drugs nor does it give priority to quality-assured anti-malarial drugs. These practices contribute to the risk of emergence and spread of resistance to anti-malarial drugs and to increasing treatment costs.

Pharmacokinetics and pharmacodynamics of a new ACT formulation: Artesunate/Amodiaquine (TRIMALACT) following oral administration in African malaria patients.

A new fixed-dose combination of artesunate (AS) plus amodiaquine (AQ) (TRIMALACT) was recently developed for the treatment of uncomplicated falciparum malaria. The originality of this combination lies in its galenic formulation which consists of a three-layer tablet with two layers containing each of the active ingredients, i.e. AS and AQ, and these are separated by a middle layer containing an antioxidant compound. To evaluate the efficacy and tolerability of this combination, adults with uncomplicated malaria received three administrations of two tablets (100:300 mg AS/AQ) in a 24-h interval, in Democratic Republic of Congo. Parasitemia and fever were measured and the plasma levels of parent compounds and metabolites [dihydroartemisinin (DHA) and monodesethylamodiaquine (MdAQ)] were determined by high-performance liquid chromatography. In addition, we determined the prevalence of molecular markers of resistance to chloroquine (CQ) and sulfadoxine/pyrimethamine (SP). The AS/AQ combination TRIMALACT demonstrated a good efficacy resulting in an excellent clinical and parasitological response rate of 100% after correction for PCR results. Treatment regimen was well tolerated. The main disposition parameters to AS+AQ were: for DHA, AUC = 632 +/- 475 ng h/ml and Cmax = 432 +/- 325 ng/ml, and for MdAQ = 14268 +/- 4114 ng h/ml and Cmax = 336 +/- 225 ng/ml (mean +/- standard deviation). Parasite genotyping show high frequencies of molecular SP- and CQ-resistance markers with more 80% of the samples showing more than three mutations linked to SP resistance and 93.48% carrying parasite with the CQ-resistant haplotype. This study shows that the AS/AQ combination TRIMALACT is safe and effective in the treatment of highly drug-resistant falciparum malaria.

[Prevention and treatment of malaria: in vitro evaluation of new compounds]. / Prophylaxie et traitement du paludisme: évaluation in vitro de nouveaux composés.

One of the current options for reducing the morbidity and mortality of malaria are chemoprophylaxis and chemotherapy. For this reason, the increasing prevalence of strains of Plasmodium falciparum resistant to chloroquine and other antimalarial drugs poses a serious problem for control of malaria. There is an urgent need to find and develop novel compounds and to identify novel chemotherapeutic targets. Different approaches to discover new compounds are presented from examples of molecules studied in the Tropical Medicine Institute of the French Army Health Service (IMTSSA) evaluation against isolates of compounds in pharmaceutical development in collaboration with pharmaceuticals (pyronaridine, benflumetol, ferrochloroquine), screening of molecules which are still registered for other pathologies (antibiotics), screening of new synthesized compounds (artemisinin derivatives) and identification of parasitical targets and essential metabolic ways for parasite, and identification of molecules acting on these targets (reversal of resistance to chloroquine, iron chelators).

Involvement of calyculin A-sensitive phosphatase(s) in the differentiation of Trypanosoma cruzi trypomastigotes to amastigotes.

Differentiation of the non-dividing trypomastigote form of Trypanosoma cruzi, the causative agent of Chagas disease, to the dividing amastigote form normally occurs in cytoplasm of infected cells. Here we show that calyculin A. a potent inhibitor of protein phosphatases 1 and 2A, induces at pH 7.5 extracellular transformation of long slender trypomastigotes to round amastigote-like forms which acquire characteristic features observed after the normal differentiation process: repositioning and structural changes of the kinetoplast, release of surface neuraminidase, and expression of amastigote-specific epitopes. Calyculin A inhibits parasite phosphatases and changes in the phosphorylation of specific proteins occur during the transformation process. As an exposure of trypomastigotes to calyculin A concentrations as low as 1 nM and for only 1-2 h is sufficient to induce transformation, the inhibition of calyculin A-sensitive phosphatase(s) appears to play a major role in initiating the trypomastigote differentiation.

Selective and reversible effects of vinca alkaloids on Trypanosoma cruzi epimastigote forms: blockage of cytokinesis without inhibition of the organelle duplication.

Vinca alkaloids, vincristine and vinblastine, produce differential effects on the cell division of Trypanosoma cruzi epimastigote forms depending on drug concentrations. These effects are related to different microtubule-based mechanisms. For 15 microM vinblastine and 50 microM vincristine, the drugs inhibit both nuclear division and cytokinesis, and affect cell shape. At 3 microM vinblastine and 10 microM vincristine, however, cytokinesis is inhibited without major effect on the progression of the cell cycle; this yields giant cells having multiple nuclei, kinetoplasts and flagella. Cultures maintained over 1 week with daily drug replacement produced cells with more than 16 nuclei and 24 kinetoplasts, indicating that an equivalent of a fifth cell cycle was initiated. The ultrastructure of the multinucleate cells showed a basic organization closely similar to that of trypanosomes. Cytokinesis inhibition by vinca alkaloids seems to result from modulations of interactions between microtubules and associated proteins, rather than from an inhibition of microtubule dynamics as is usually proposed for vinca alkaloids. Cytokinesis inhibition is reversible: after removing the drug, epimastigotes emerge from the multinucleate cells. The emerging process follows a precise axis and polarity which are determined by the position of the flagellum/kinetoplast complex. This region could play an essential role in cell morphogenesis since zoids (cells without a nucleus) are frequently observed.

Host cell and malarial targets for docetaxel (Taxotere) during the erythrocytic development of Plasmodium falciparum.

The microtubular stabilizing agent docetaxel (Taxotere) is known to inhibit the intraerythrocytic development of Plasmodium falciparum. To investigate the mechanism(s) of inhibition, we analyzed the structural organization of the mitotic spindle by immunofluorescence and electron microscopy. When 30 microM docetaxel was applied for five hours on ring forms, alterations in the mitotic spindles leading to abnormal nuclear divisions were observed. At the trophozoite- and schizont-stage, docetaxel pulses prevent mitosis by stabilizing microtubular structures associated with the mitotic apparatus, giving abnormal spindles. However, this inhibition did not interfere with parasite DNA synthesis indicating the absence of a checkpoint that couples exit from mitosis with proper spindle assembly as observed in higher eukaryotic cells. In parallel, intraerythrocytic concentration of docetaxel was measured in parasitized erythrocytes, after incubation of cells with 3H-docetaxel for five hours. It was found to be 14-fold increased at the ring-stage of infected erythrocytes compared to normal ones, 170-fold increased at the trophozoite-stage and 1,500-fold increased at the schizont-stage. Our data show that, even though the overall intracellular concentration of docetaxel is low in docetaxel-pulsed rings, the agent might be sufficient to disturb the spindle organization. However, the existence of targets for docetaxel other than mitotic spindle microtubules, i.e. erythrocyte membrane components could interfere with mitotic spindle formation.

In vitro and in vivo inhibition of erythrocytic development of malarial parasites by docetaxel.

The stage-dependent susceptibility of Plasmodium falciparum to a short exposure to docetaxel (Taxotere) was evaluated by subjecting ring-infected, trophozoite-infected, and schizont-infected erythrocytes to a 5-h exposure to various concentrations of the drug. The schizont stage was shown to be the most sensitive stage; an inhibition of more than 60% of parasite development was observed at 10 nM. At this drug concentration, the development of the younger ring and trophozoite forms was unaffected. The in vivo antimalarial activity of docetaxel against the development in blood of old trophozoites of a species that causes malaria in rodents, Plasmodium vinckei petteri, was evaluated in IOPS-OF1 mice. Two tests were performed: the 4-day suppressive test, as described by Peters (W. Peters, p. 145-273, in Chemotherapy, and Drug Resistance in Malaria, vol. 1, 1987), and the effects of a single injection of docetaxel after inoculation of the parasites. A single injection of docetaxel at 40 mg/kg of body weight was sufficient to reduce drastically the level of parasitemia; 90% inhibition of the development of parasites in blood was observed 5 days after drug injection. This program avoided the toxicity observed when mice were treated with four injections of docetaxel. The possibility of using a single bolus of taxoids to treat malaria infections is discussed.

New trends in chemotherapy on human and animal blood parasites.

Blood-parasite protozoa are causative agents of some of the major tropical or infectious diseases for humans and animals, such as Plasmodium for malaria (about 270 million infected people), Trypanosoma cruzi for Chagas' disease (about 18-20 million individuals), African trypanosomes for human and bovine trypanosomiasis, and Babesia for cattle and dogs. The absence of efficient vaccines against these diseases, the absence or the high toxicity of the few drugs against American and African trypanosomiasis, and the emergence of chemoresistance against Plasmodium falciparum emphasize the necessity to propose new antiparasitic strategies. Among these strategies, the biological strategy is based on the identification of key molecules for parasite development such that structural analogs can be designed that are parasite-specific or sufficiently inactive for the host. This requires a careful biochemical analysis of each step of the parasite life cycle. For blood-parasite protozoa, the lipid metabolism required for membrane biogenesis, antimicrotubular drugs or inhibitors of the mitotic spindle, and drug targeting offer new trends in chemotherapy against Plasmodium, Babesia, and trypanosomes.

Interactions between docetaxel (Taxotere) and Plasmodium falciparum-infected erythrocytes.

Taxotere (docetaxel) inhibits Plasmodium falciparum erythrocytic development in vitro at nanomolar concentrations, both in chloroquine-sensitive (F32/Tanzania) and chloroquine-resistant (FcB1/Colombia, FcR3/Gambia) strains. The dose-response assays performed on asynchronous cultures during 42 hr showed clear biphasic curves with a plateau from 50 microM to 10 nM and a single sigmoid curve with a concentration inhibiting 50% of growth (IC50) of 3-6 nM observed after a 72-hr incubation. Addition of Taxotere to different stages of FcB1 revealed two types of targets: one type on ring/trophozoite-infected erythrocytes (RBCs), at the micromolar level, and another type on schizont-infected RBCs with Taxotere at micromolar concentrations inhibited the merozoite invasion of erythrocytes and parasite growth. These Taxotere-RBC interactions were stable, at least for 1 day. Pulse experiments of 5 hr with Taxotere efficiently inhibit parasite development regardless of the period of the parasite's erythrocytic life cycle. However, different cellular effects were obtained depending upon periods of drug incubations. The inhibition of P. falciparum development by Taxotere should provide additional strategies to block parasite development.