We report a large multicenter genome-wide association study of Plasmodium falciparum resistance to artemisinin, the frontline antimalarial drug. Across 15 locations in Southeast Asia, we identified at least 20 mutations in kelch13 (PF3D7_1343700) affecting the encoded propeller and BTB/POZ domains, which were associated with a slow parasite clearance rate after treatment with artemisinin derivatives. Nonsynonymous polymorphisms in fd (ferredoxin), arps10 (apicoplast ribosomal protein S10), mdr2 (multidrug resistance protein 2) and crt (chloroquine resistance transporter) also showed strong associations with artemisinin resistance. Analysis of the fine structure of the parasite population showed that the fd, arps10, mdr2 and crt polymorphisms are markers of a genetic background on which kelch13 mutations are particularly likely to arise and that they correlate with the contemporary geographical boundaries and population frequencies of artemisinin resistance. These findings indicate that the risk of new resistance-causing mutations emerging is determined by specific predisposing genetic factors in the underlying parasite population.

The widespread distribution and relapsing nature of Plasmodium vivax infection present major challenges for malaria elimination. To characterise the genetic diversity of this parasite within individual infections and across the population, we performed deep genome sequencing of >200 clinical samples collected across the Asia-Pacific region, and analysed data on >300,000 SNPs and 9 regions of the genome with large copy number variations. Individual infections showed complex patterns of genetic structure, with variation not only in the number of dominant clones but also in their level of relatedness and inbreeding. At the population level, we observed strong signals of recent evolutionary selection both in known drug resistance genes and at novel loci, and these varied markedly between geographical locations. These findings reveal a dynamic landscape of local evolutionary adaptation in P. vivax populations, and provide a foundation for genomic surveillance to guide effective strategies for control and elimination.

SUMMARY Through in-vivo selection of human cancer cell populations, we uncover a convergent and cooperative miRNA network that drives melanoma metastasis. We identify miR-1908, miR-199a-5p, and miR-199a-3p as endogenous promoters of metastatic invasion, angiogenesis, and colonization in melanoma. These miRNAs convergently target Apolipoprotein E (ApoE) and the heat-shock factor DNAJA4. Cancer-secreted ApoE suppresses invasion and metastatic endothelial recruitment (MER) by engaging melanoma-cell LRP1 and endothelial-cell LRP8 receptors, respectively–while DNAJA4 promotes ApoE expression. Expression levels of these miRNAs and ApoE correlate with human metastatic progression outcomes. Treatment of cells with locked nucleic acids (LNAs) targeting these miRNAs inhibits metastasis to multiple organs, while therapeutic delivery of these LNAs strongly suppresses melanoma metastasis. We thus identify miRNAs with dual cell-intrinsic/cell-extrinsic roles in cancer, reveal convergent cooperativity in a metastatic miRNA network, identify ApoE as an anti-angiogenic and metastasis-suppressive factor, and uncover multiple prognostic miRNAs with synergistic combinatorial therapeutic potential in melanoma.

BackgroundThere is currently no licensed antiviral drug for treatment of dengue. Chloroquine (CQ) inhibits the replication of dengue virus (DENV) in vitro.Methods and FindingsA double-blind, randomized, placebo-controlled trial of CQ in 307 adults hospitalized for suspected DENV infection was conducted at the Hospital for Tropical Diseases (Ho Chi Minh City, Vietnam) between May 2007 and July 2008. Patients with illness histories of 72 hours or less were randomized to a 3-day course of CQ (n = 153) or placebo (n = 154). Laboratory-confirmation of DENV infection was made in 257 (84%) patients. The primary endpoints were time to resolution of DENV viraemia and time to resolution of DENV NS1 antigenaemia. In patients treated with CQ there was a trend toward a longer duration of DENV viraemia (hazard ratio (HR) = 0.80, 95% CI 0.62–1.05), but we did not find any difference for the time to resolution of NS1 antigenaemia (HR = 1.07, 95% CI 0.76–1.51). Interestingly, CQ was associated with a significant reduction in fever clearance time in the intention-to-treat population (HR = 1.37, 95% CI 1.08–1.74) but not in the per-protocol population. There was also a trend towards a lower incidence of dengue hemorrhagic fever (odds ratio = 0.60, PP 95% CI 0.34–1.04) in patients treated with CQ. Differences in levels of T cell activation or pro- or anti-inflammatory plasma cytokine concentrations between CQ- and placebo-treated patients did not explain the trend towards less dengue hemorrhagic fever in the CQ arm. CQ was associated with significantly more adverse events, primarily vomiting.ConclusionsCQ does not reduce the durations of viraemia and NS1 antigenaemia in dengue patients. Further trials, with appropriate endpoints, would be required to determine if CQ treatment has any clinical benefit in dengue.Trial RegistrationCurrent Controlled Trials number ISRCTN38002730.

BackgroundDengue is a major public health problem in tropical and subtropical countries. Exploring the relationships between virological features of infection with patient immune status and outcome may help to identify predictors of disease severity and enable rational therapeutic strategies.MethodsClinical features, antibody responses and virological markers were characterized in Vietnamese adults participating in a randomised controlled treatment trial of chloroquine.ResultsOf the 248 patients with laboratory-confirmed dengue and defined serological and clinical classifications 29 (11.7%) had primary DF, 150 (60.5%) had secondary DF, 4 (1.6%) had primary DHF and 65 (26.2%) had secondary DHF. DENV-1 was the commonest serotype (57.3%), then DENV-2 (20.6%), DENV-3 (15.7%) and DENV-4 (2.8%). DHF was associated with secondary infection (Odds ratio = 3.13, 95% CI 1.04–12.75). DENV-1 infections resulted in significantly higher viremia levels than DENV-2 infections. Early viremia levels were higher in DENV-1 patients with DHF than with DF, even if the peak viremia level was often not observed because it occurred prior to enrolment. Peak viremias were significantly less often observed during secondary infections than primary for all disease severity grades (P = 0.001). The clearance of DENV viremia and NS1 antigenemia occurs earlier and faster in patients with secondary dengue (P<0.0001). The maximum daily rate of viremia clearance was significantly higher in patients with secondary infections than primary (P<0.00001).ConclusionsCollectively, our findings suggest that the early magnitude of viremia is positively associated with disease severity. The clearance of DENV is associated with immune status, and there are serotype dependent differences in infection kinetics. These findings are relevant for the rational design of randomized controlled trials of therapeutic interventions, especially antivirals.