Search

Background Resistance to front-line antimalarials (artemisinin combination therapies) is spreading, and development of new drug treatment strategies to rapidly kill Plasmodium spp. malaria parasites is urgently needed. Azithromycin is a clinically used macrolide antibiotic proposed as a partner drug for combination therapy in malaria, which has also been tested as monotherapy. However, its slow-killing ‘delayed-death’ activity against the parasite’s apicoplast organelle and suboptimal activity as monotherapy limit its application as a potential malaria treatment. Here, we explore a panel of azithromycin analogues and demonstrate that chemical modifications can be used to greatly improve the speed and potency of antimalarial action. Results Investigation of 84 azithromycin analogues revealed nanomolar quick-killing potency directed against the very earliest stage of parasite development within red blood cells. Indeed, the best analogue exhibited 1600-fold higher potency than azithromycin with less than 48 hrs treatment in vitro. Analogues were effective against zoonotic Plasmodium knowlesi malaria parasites and against both multi-drug and artemisinin-resistant Plasmodium falciparum lines. Metabolomic profiles of azithromycin analogue-treated parasites suggested activity in the parasite food vacuole and mitochondria were disrupted. Moreover, unlike the food vacuole-targeting drug chloroquine, azithromycin and analogues were active across blood-stage development, including merozoite invasion, suggesting that these macrolides have a multi-factorial mechanism of quick-killing activity. The positioning of functional groups added to azithromycin and its quick-killing analogues altered their activity against bacterial-like ribosomes but had minimal change on ‘quick-killing’ activity. Apicoplast minus parasites remained susceptible to both azithromycin and its analogues, further demonstrating that quick-killing is independent of apicoplast-targeting, delayed-death activity. Conclusion We show that azithromycin and analogues can rapidly kill malaria parasite asexual blood stages via a fast action mechanism. Development of azithromycin and analogues as antimalarials offers the possibility of targeting parasites through both a quick-killing and delayed-death mechanism of action in a single, multifactorial chemotype.

Aim Individual Placement and Support is an effective vocational intervention for increasing competitive employment for people with severe mental illness. Little is known, however, about its effectiveness in the context of early psychosis. This study assesses improvements in clients' employment in a phase of illness during which functional abilities often decline. Methods The trial design is an assessor-blinded randomized clinical trial, set in the context of a population-based Early Psychosis Intervention program in British Columbia, Canada. Participants were randomized either to 1 year of employment support added to treatment-as-usual, or the latter alone. Interviews at intake captured data regarding demographics, symptom severity, and employment; assessments at 6 and 12 months repeated queries about employment activities. Results A total of 109 clients were recruited. Employment rates in the Individual Placement and Support group increased over time, unlike the control group. Further, the number of days worked over the 12-month intervention period, compared to the 6 months prior to the study, improved for both groups, but the increase was greater among clients receiving IPS. Sensitivity analysis indicated the advantage in days worked was evident in the second half of the intervention period (6-12 months), but not the first half. Conclusions Employment rates, for younger clients in both early-psychosis groups, were high compared to older clients in later stages of illness. In this study, use of the Individual Placement and Support strategy further increased employment, despite the high baseline rates. Further research is needed to identify the optimal timing of employment support for these clients.

Plasmodium spp. parasites that cause malaria disease remain a significant global-health burden. With the spread of parasites resistant to artemisinin combination therapies in Southeast Asia, there is a growing need to develop new antimalarials with novel targets. Invasion of the red blood cell by Plasmodium merozoites is essential for parasite survival and proliferation, thus representing an attractive target for therapeutic development. Red blood cell invasion requires a co-ordinated series of protein/protein interactions, protease cleavage events, intracellular signals, organelle release and engagement of an actin-myosin motor, which provide many potential targets for drug development. As these steps occur in the bloodstream, they are directly susceptible and exposed to drugs. A number of invasion inhibitors against a diverse range of parasite proteins involved in these different processes of invasion have been identified, with several showing potential to be optimised for improved drug-like properties. In this review, we discuss red blood cell invasion as a drug target and highlight a number of approaches for developing antimalarials with invasion inhibitory activity to use in future combination therapies., Malaria invasion of red blood cells is an essential step in parasite replication and this review discusses targets and drug chemotypes being developed to stop invasion and growth.

Membrane solubilization by sodium dodecyl sulfate (SDS) is indispensable for many established biotech-nological applications, including viral inactivation and protein extraction. Although the ensemble thermo-dynamics have been thoroughly explored, the underlying molecular dynamics have remained inaccessible, owing to major limitations of traditional measurement tools. Here, we integrate multiple advanced biophysical approaches to gain multi-angle insight into the time-dependence and fundamental kinetic steps associated with the solubilization of single sub-micron sized vesicles in response to SDS. We find that the accumulation of SDS molecules on in-tact vesicles triggers biphasic solubilization kinetics comprising an initial vesicle expansion event followed by rapid lipid loss and micellization. Our findings support a general mechanism of detergent-induced membrane solubilization and we expect the framework of correlative biophysical technologies presented here will form a general platform for elucidating the complex kinetics of membrane perturbation induced by a wide variety of surfactants and disrupting agents.

Intracellular pathogens mobilize host signaling pathways of their host cell to promote their own survival. Evidence is emerging that signal transduction elements are activated in a-nucleated erythrocytes in response to infection with malaria parasites, but the extent of this phenomenon remains unknown. Here, we fill this knowledge gap through a comprehensive and dynamic assessment of host erythrocyte signaling during infection with Plasmodium falciparum. We used arrays of 878 antibodies directed against human signaling proteins to interrogate the activation status of host erythrocyte phospho-signaling pathways at three blood stages of parasite asexual development. This analysis reveals a dynamic modulation of many host signalling proteins across parasite development. Here we focus on the hepatocyte growth factor receptor (c-MET) and the MAP kinase pathway component B-Raf, providing a proof of concept that human signaling kinases identified as activated by malaria infection represent attractive targets for antimalarial intervention., Plasmodium infection activates signaling pathways in a-nucleated erythrocytes. Here, Adderley et al. use a comprehensive antibody microarray to show that infection extensively modulates host cell signalling and that the host receptor tyrosine kinase c-MET supports Plasmodium falciparum proliferation.

Neuronal nitric oxide synthase (nNOS) catalyses the production of the neurotransmitter nitric oxide. nNOS is expressed in the dorsal raphe nucleus (DRN), a source of ascending serotonergic projections. In this study, we examined the distribution nNOS and the function of nitric oxide in the DRN and adjacent median raphe nucleus (MRN) of the rat. We hypothesized that nNOS is differentially expressed across the raphe nuclei and that nitric oxide influences the firing activity of a subgroup of 5-HT neurons. Immunohistochemistry revealed that, nNOS is present in around 40% of 5-HT neurons, throughout the DRN and MRN, as well as in some non-5-HT neurons immediately adjacent to the DRN and MRN. The nitric oxide receptor, soluble guanylyl cyclase, was present in all 5-HT neurons examined in the DRN and MRN. In vitro extracellular electrophysiology revealed that application of the nitric oxide donor, diethylamine NONOate (30-300 mu M) inhibited 60%-70% of putative 5-HT neurons, excited approximately 10% of putative 5-HT neurons and had no effect on the rest. The inhibitory response to nitric oxide was blocked by [1H-[1,2,4]oxadiazolo-[4, 3-a]quinoxalin-1-one (ODQ, 30 or 100 mu M), indicating mediation by soluble guanylyl cyclase. Juxtacellular labelling revealed that nitric oxide inhibits firing in both putative 5-HT neurons which express nNOS and those which do not express nNOS. Our data are consistent with the notion that nitric oxide acts as both a trans-synaptic and autocrine signaller in 5-HT neurons in the DRN and MRN and that its effects are widespread and primarily inhibitory.