Pascal Millet

Manslaughter by fake artesunate in Asia - will Africa be next?

Fake artesunate could compromise the hope that artemisinin-based combination therapy offers for malaria control in Africa and Asia.

New ferrocenic pyrrolo[1,2-a]quinoxaline derivatives: synthesis, and in vitro antimalarial activity.

Following our search for antimalarial compounds, novel series of ferrocenic pyrrolo[1,2-a]quinoxaline derivatives 1-2 were synthesized from various substituted nitroanilines and tested for in vitro activity upon the erythrocytic development of Plasmodiumfalciparum strains with different chloroquine-resistance status. The pyrrolo[1,2-a]quinoxalines 1 were prepared in 6-8 steps through a regioselective palladium-catalyzed monoamination by coupling 4-chloropyrrolo[1,2-a]quinoxalines with 1,3-bis(aminopropyl)piperazine or -methylamine using Xantphos as the ligand. The ferrocenic bispyrrolo[1,2-a]quinoxalines 2 were prepared by reductive amination of previously described bispyrrolo[1,2-a]quinoxalines 9 with ferrocene-carboxaldehyde, by treatment with NaHB(OAc)(3). The best results were observed with ferrocenic pyrrolo[1,2-a]quinoxalines linked by a bis(3-aminopropyl)piperazine. Moreover, it was observed that a methoxy group on the pyrrolo[1,2-a]quinoxaline nucleus and no substitution on the terminal N-ferrocenylmethylamine function enhanced the pharmacological activity. Selected compounds 1b, 1f-h, 1l and 2a were tested for their ability to inhibit beta-haematin formation, the synthetic equivalent of hemozoin, by using the HPIA (heme polymerization inhibitory activity) assay. Of the tested compounds, only 2a showed a beta-haematin formation inhibition, but no inhibition of haem polymerization was observed with the other selected ferrocenic monopyrrolo[1,2-a]quinoxaline derivatives 1b, 1f-h and 1l, as the IC(50) values were superior to 10 equivalents.

In vitro activities of ferrochloroquine against 55 Senegalese isolates of Plasmodium falciparum in comparison with those of standard antimalarial drugs

The in vitro activities of ferrochloroquine, chloroquine, quinine, mefloquine, halofantrine, amodiaquine, artesunate, atovaquone, cycloguanil and pyrimethamine were evaluated against Plasmodium falciparum isolates from Senegal (Dielmo, Ndiop), using an isotopic micro‐drug susceptibility test. The IC50 values for ferrochloroquine ranged from 0.55 to 28.2 nM and the geometric mean IC50 for the 55 isolates was 7.9 nM (95% CI, 6.5–9.7 nM). Ferrochloroquine was 35 times more active than chloroquine (35‐fold greater against chloroquine‐resistant isolates), quinine, mefloquine, amodiaquine, cycloguanil and pyrimethamine. Weak positive correlations were observed between the responses to ferrochloroquine and that to chloroquine, quinine, and amodiaquine, but not compulsorily predictive of cross‐resistance. There was no significant correlation between the response to ferrochloroquine and that to mefloquine, halofantrine, artesunate, atovaquone, cycloguanil and pyrimethamine. Ferrochloroquine may be an important alternative drug for the treatment of chloroquine‐resistant malaria.

Evaluation of FRET real-time PCR assay for rapid detection and differentiation of Plasmodium species in returning travellers and migrants

BackgroundA simple real-time PCR assay using one set of primer and probe for rapid, sensitive and quantitative detection of Plasmodium species, with simultaneous differentiation of Plasmodium falciparum from the three other Plasmodium species (Plasmodium vivax, Plasmodium ovale and Plasmodium malariae) in febrile returning travellers and migrants was developed and evaluated.MethodsConsensus primers were used to amplify a species-specific region of the multicopy 18S rRNA gene, and fluorescence resonance energy transfer hybridization probes were used for detection in a LightCycler platform (Roche). The anchor probe sequence was designed to be perfect matches to the 18S rRNA gene of the fourth Plasmodium species, while the acceptor probe sequence was designed for P. falciparum over a region containing one mismatched, which allowed differentiation of the three other Plasmodium species. The performance characteristics of the real-time PCR assay were compared with those of conventional PCR and microscopy-based diagnosis from 119 individuals with a suspected clinical diagnostic of imported malaria.ResultsBlood samples with parasite densities less than 0.01% were all detected, and analytical sensitivity was 0.5 parasite per PCR reaction. The melt curve means Tms (standard deviation) in clinical isolates were 60.5°C (0.6°C) for P. falciparum infection and 64.6°C (1.8°C) for non-P. falciparum species. These Tms values of the P. falciparum or non-P. falciparum species did not vary with the geographic origin of the parasite. The real-time PCR results correlated with conventional PCR using both genus-specific (Kappa coefficient: 0.95, 95% confidence interval: 0.9 – 1) or P. falciparum-specific (0.91, 0.8 – 1) primers, or with the microscopy results (0.70, 0.6 – 0.8). The real-time assay was 100% sensitive and specific for differentiation of P. falciparum to non-P. falciparum species, compared with conventional PCR or microscopy. The real-time PCR assay can also detect individuals with mixed infections (P. falciparum and non-P. falciparum sp.) in the same sample.ConclusionThis real-time PCR assay with melting curve analysis is rapid, and specific for the detection and differentiation of P. falciparum to other Plasmodium species. The suitability for routine use of this assay in clinical diagnostic laboratories is discussed.

Neglected diseases in the news: a content analysis of recent international media coverage focussing on leishmaniasis and trypanosomiasis.

Background Although the pharmaceutical industrys “neglect” of neglected tropical diseases (NTDs) has been investigated, no study evaluating media coverage of NTDs has been published. Poor media coverage exacerbates the neglect. This study aimed to investigate, describe, and analyse international media coverage of “neglected diseases” in general and three specific NTDs—African trypanosomiasis, leishmaniasis, and Chagas disease—from 1 January 2003 to 1 June 2007. Methods Archives of 11 leading international, English-language media were searched. A content analysis was done, coding for media organisation, date, author, type of report, slant, themes, and “frames”. Semi-structured interviews with journalists and key informants were conducted for further insight. Principal Findings Only 113 articles in a 53-month time period met the inclusion criteria, with no strong trends or increases in coverage. Overall, the BBC had the highest coverage with 20 results, followed by the Financial Times and Agence France Presse. CNN had the least coverage with one result. The term “neglected diseases” had good media currency and “sleeping sickness” was far more widely used than trypanosomiasis. The disease most covered was leishmaniasis and the least covered was Chagas. Academic researchers were most commonly quoted as a main source, while the World Health Organization (WHO) and pharmaceutical industry were the least quoted. Journalists generally agreed NTDs had not been adequately covered, but said a lack of real news development and the need to cater to domestic audiences were major obstacles for NTD reporting. All journalists said health agencies, particularly WHO, were not communicating adequately about the burden of NTDs. Conclusions Public health agencies need to raise priority for NTD advocacy. Innovative strategies, such as reporting grants or creating a network of voices, may be needed.

Effect of adjuvant formulations on the selection of B-cell epitopes expressed by a malaria peptide vaccine

Because subunit vaccines may create artificial epitopes, the goal of this study was to concentrate the immune response toward protective epitopes contained in a peptide, [(NAGG)5]Y. This peptide consisted of five repeat sequences of the immunodominant epitope of the circumsporozoite protein of the simian malaria parasite Plasmodium cynomolgi and a terminal tyrosine. It was conjugated to bovine serum albumin and mixed with various adjuvant formulations, including block copolymers L121, L141, L180.5 and a lipopolysaccharide (LPS). Outbred mice were vaccinated with seven vaccine formulations. Postvaccination sera from each group of mice were then pooled, and antibody responses against peptide [(NAGG)5]Y or (NAGG)5 were tested in an enzyme-linked immunosorbent assay and against sporozoites of P. cynomolgi in an indirect fluorescent antibody assay. Although all groups elicited a high antibody response to the peptide [(NAGG)5Y], the presence of the tyrosine induced a different antibody response to the peptide (NAGG)5, and formulations containing LPS alone did not induce an antibody response to either (NAGG)5 or P. cynomolgi sporozoites. The formulation including both LPS and the copolymer L121 was the only one to inhibit the development of P. cynomolgi sporozoites in rhesus monkey hepatocytes by 50%. These results suggest that vaccine formulations influence B-cell epitope selection.

Management of uncomplicated malaria in febrile under five-year-old children by community health workers in Madagascar: reliability of malaria rapid diagnostic tests

BackgroundEarly diagnosis, as well as prompt and effective treatment of uncomplicated malaria, are essential components of the anti-malaria strategy in Madagascar to prevent severe malaria, reduce mortality and limit malaria transmission. The purpose of this study was to assess the performance of the malaria rapid diagnostic tests (RDTs) used by community health workers (CHWs) by comparing RDT results with two reference methods (microscopy and Polymerase Chain Reaction, PCR).MethodsEight CHWs in two districts, each with a different level of endemic malaria transmission, were trained to use RDTs in the management of febrile children under five years of age. RDTs were performed by CHWs in all febrile children who consulted for fever. In parallel, retrospective parasitological diagnoses were made by microscopy and PCR. The results of these different diagnostic methods were analysed to evaluate the diagnostic performance of the RDTs administered by the CHWs. The stability of the RDTs stored by CHWs was also evaluated.ResultsAmong 190 febrile children with suspected malaria who visited CHWs between February 2009 and February 2010, 89.5% were found to be positive for malaria parasites by PCR, 51.6% were positive by microscopy and 55.8% were positive by RDT. The performance accuracy of the RDTs used by CHWs in terms of sensitivity, specificity, positive and negative predictive values was greater than 85%. Concordance between microscopy and RDT, estimated by the Kappa value was 0.83 (95% CI: 0.75-0.91). RDTs stored by CHWs for 24 months were capable of detecting Plasmodium falciparum in blood at a level of 200 parasites/μl.ConclusionIntroduction of easy-to-use diagnostic tools, such as RDTs, at the community level appears to be an effective strategy for improving febrile patient management and for reducing excessive use of anti-malarial drugs.

Quinine-Resistant Malaria in Traveler Returning from Senegal, 2007

We describe clinical and parasitologic features of in vivo and in vitro Plasmodium falciparum resistance to quinine in a nonimmune traveler who returned to France from Senegal in 2007 with severe imported malaria. Clinical quinine failure was associated with a 50% inhibitory concentration of 829 nmol/L. Increased vigilance is required during treatment follow-up.

Treatment of malaria from monotherapy to artemisinin-based combination therapy by health professionals in rural health facilities in southern Cameroon

BackgroundOne year after the adoption of artesunate-amodiaquine (AS/AQ) as first-line therapy for the treatment of uncomplicated malaria, this study was designed to assess the treatment practices regarding anti-malarial drugs at health facilities in four rural areas in southern Cameroon.MethodsBetween April and August 2005, information was collected by interviewing fifty-two health professionals from twelve rural health facilities, using a structured questionnaire.ResultsIn 2005, only three anti-malarial drugs were used in rural health facilities, including: amodiaquine, quinine and sulphadoxine-pyrimethamine. Only 2.0% of the health professionals prescribed the recommended AS/AQ combination. After reading the treatment guidelines, 75.0% were in favour of the treatment protocol with the following limitations: lack of paediatric formulations, high cost and large number of tablets per day. Up to 21.0% of professionals did not prescribe AS/AQ because of the level of adverse events attributed to the use of amodiaquine as monotherapy.ConclusionThe present study indicates that AS/AQ was not available in the public health facilities at the time of the study, and health practitioners were not informed about the new treatment guidelines. Results of qualitative analysis suggest that prescribers should be involved as soon as possible in projects related to the optimization of treatment guidelines and comply with new drugs. Adapted formulations should be made available at the international level and implemented locally before new drugs and treatments are proposed through a national control programme. This baseline information will be useful to monitor progresses in the implementation of artemisinin-based combination therapy in Cameroon.

The initial pharmaceutical development of an artesunate/amodiaquine oral formulation for the treatment of malaria: a public-private partnership

BackgroundArtemisinin-based combination therapy is currently recommended worldwide for the treatment of uncomplicated malaria. Fixed-dose combinations are preferred as they favour compliance. This paper reports on the initial phases of the pharmaceutical development of an artesunate-amodiaquine (ASAQ) bilayer co-formulation tablet, undertaken following pre-formulation studies by a network of scientists and industrials from institutions of both industrialized and low income countries.MethodsPharmaceutical development was performed by a research laboratory at the University Bordeaux Segalen, School of Pharmacy, for feasibility and early stability studies of various drug formulations, further transferred to a company specialized in pharmaceutical development, and then provided to another company for clinical batch manufacturing. The work was conducted by a regional public-private not-for-profit network (TropiVal) within a larger Public Private partnership (the FACT project), set up by WHO/TDR, Médecins Sans Frontières and the Drugs for Neglected Disease initiative (DNDi).ResultsThe main pharmaceutical goal was to combine in a solid oral form two incompatible active principles while preventing artesunate degradation under tropical conditions. Several options were attempted and failed to provide satisfactory stability results: incorporating artesunate in the external phase of the tablets, adding a pH regulator, alcoholic wet granulation, dry granulation, addition of an hydrophobic agent, tablet manufacturing in controlled conditions. However, long-term stability could be achieved, in experimental batches under GMP conditions, by physical separation of artesunate and amodiaquine in a bilayer co-formulation tablet in alu-alu blisters. Conduction of the workplan was monitored by DNDi.ConclusionsCollaborations between research and industrial groups greatly accelerated the process of development of the bi-layered ASAQ tablet. Lack of public funding was the main obstacle hampering the development process, and no intellectual property right was claimed. This approach resulted in a rapid technology transfer to the drug company Sanofi-Aventis, finalizing the process of development, registration and WHO pre-qualification of the fixed-dose co-formulation together with DNDi. The bi-layered tablet is made available under the names of Coarsucam® and Artesunate amodiaquine Winthrop®, Sanofi-Aventis. The issue related to the difficulty of public institutions to valorise their participation in such initiative by lack of priority and funding of applied research is discussed.