Caspase-8 mediates inflammation and disease in rodent malaria

Earlier studies indicate that either the canonical or non-canonical pathways of inflammasome activation have a limited role on malaria pathogenesis. Here, we report that caspase-8 is a central mediator of systemic inflammation, septic shock in the Plasmodium chabaudi-infected mice and the P. berghei-induced experimental cerebral malaria (ECM). Importantly, our results indicate that the combined deficiencies of caspases-8/1/11 or caspase-8/gasdermin-D (GSDM-D) renders mice impaired to produce both TNFalpha and IL-1beta and highly resistant to lethality in these models, disclosing a complementary, but independent role of caspase-8 and caspases-1/11/GSDM-D in the pathogenesis of malaria. Further, we find that monocytes from malaria patients express active caspases-1, -4 and -8 suggesting that these inflammatory caspases may also play a role in the pathogenesis of human disease

Preclinical characterization and target validation of the antimalarial pantothenamide MMV693183.

Drug resistance and a dire lack of transmission-blocking antimalarials hamper malaria elimination. Here, we present the pantothenamide MMV693183 as a first-in-class acetyl-CoA synthetase (AcAS) inhibitor to enter preclinical development. Our studies demonstrate attractive drug-like properties and in vivo efficacy in a humanized mouse model of Plasmodium falciparum infection. The compound shows single digit nanomolar in vitro activity against P. falciparum and P. vivax clinical isolates, and potently blocks P. falciparum transmission to Anopheles mosquitoes. Genetic and biochemical studies identify AcAS as the target of the MMV693183-derived antimetabolite, CoA-MMV693183. Pharmacokinetic-pharmacodynamic modelling predict that a single 30 mg oral dose is sufficient to cure a malaria infection in humans. Toxicology studies in rats indicate a \u3e 30-fold safety margin in relation to the predicted human efficacious exposure. In conclusion, MMV693183 represents a promising candidate for further (pre)clinical development with a novel mode of action for treatment of malaria and blocking transmission

The efficacy of dihydroartemisinin-piperaquine and artemether-lumefantrine with and without primaquine on Plasmodium vivax recurrence: A systematic review and individual patient data meta-analysis

BACKGROUND Artemisinin-based combination therapy (ACT) is recommended for uncomplicated Plasmodium vivax malaria in areas of emerging chloroquine resistance. We undertook a systematic review and individual patient data meta-analysis to compare the efficacies of dihydroartemisinin-piperaquine (DP) and artemether-lumefantrine (AL) with or without primaquine (PQ) on the risk of recurrent P. vivax. METHODS AND FINDINGS Clinical efficacy studies of uncomplicated P. vivax treated with DP or AL and published between January 1, 2000, and January 31, 2018, were identified by conducting a systematic review registered with the International Prospective Register of Systematic Reviews (PROSPERO): CRD42016053310. Investigators of eligible studies were invited to contribute individual patient data that were pooled using standardised methodology. The effect of mg/kg dose of piperaquine/lumefantrine, ACT administered, and PQ on the rate of P. vivax recurrence between days 7 and 42 after starting treatment were investigated by Cox regression analyses according to an a priori analysis plan. Secondary outcomes were the risk of recurrence assessed on days 28 and 63. Nineteen studies enrolling 2,017 patients were included in the analysis. The risk of recurrent P. vivax at day 42 was significantly higher in the 384 patients treated with AL alone (44.0%, 95% confidence interval [CI] 38.7-49.8) compared with the 812 patients treated with DP alone (9.3%, 95% CI 7.1-12.2): adjusted hazard ratio (AHR) 12.63 (95% CI 6.40-24.92), p < 0.001. The rates of recurrence assessed at days 42 and 63 were associated inversely with the dose of piperaquine: AHRs (95% CI) for every 5-mg/kg increase 0.63 (0.48-0.84), p = 0.0013 and 0.83 (0.73-0.94), p = 0.0033, respectively. The dose of lumefantrine was not significantly associated with the rate of recurrence (1.07 for every 5-mg/kg increase, 95% CI 0.99-1.16, p = 0.0869). In a post hoc analysis, in patients with symptomatic recurrence after AL, the mean haemoglobin increased 0.13 g/dL (95% CI 0.01-0.26) for every 5 days that recurrence was delayed, p = 0.0407. Coadministration of PQ reduced substantially the rate of recurrence assessed at day 42 after AL (AHR = 0.20, 95% CI 0.10-0.41, p < 0.001) and at day 63 after DP (AHR = 0.08, 95% CI 0.01-0.70, p = 0.0233). Results were limited by follow-up of patients to 63 days or less and nonrandomised treatment groups. CONCLUSIONS In this study, we observed the risk of P. vivax recurrence at day 42 to be significantly lower following treatment with DP compared with AL, reflecting the longer period of post-treatment prophylaxis; this risk was reduced substantially by coadministration with PQ. We found that delaying P. vivax recurrence was associated with a small but significant improvement in haemoglobin. These results highlight the benefits of PQ radical cure and also the provision of blood-stage antimalarial agents with prolonged post-treatment prophylaxis

Plasmodium vivax and Plasmodium falciparum ex vivo susceptibility to anti-malarials and gene characterization in Rondônia, West Amazon, Brazil.

Submitted by Nuzia Santos ([email protected]) on 2015-02-09T18:31:12Z No. of bitstreams: 1 2014_064.pdf: 497580 bytes, checksum: 316e9d474b23be75c14c54a2ed6320ab (MD5)Approved for entry into archive by Nuzia Santos ([email protected]) on 2015-02-09T18:31:20Z (GMT) No. of bitstreams: 1 2014_064.pdf: 497580 bytes, checksum: 316e9d474b23be75c14c54a2ed6320ab (MD5)Approved for entry into archive by Nuzia Santos ([email protected]) on 2015-02-09T18:35:37Z (GMT) No. of bitstreams: 1 2014_064.pdf: 497580 bytes, checksum: 316e9d474b23be75c14c54a2ed6320ab (MD5)Made available in DSpace on 2015-02-09T18:35:37Z (GMT). No. of bitstreams: 1 2014_064.pdf: 497580 bytes, checksum: 316e9d474b23be75c14c54a2ed6320ab (MD5) Previous issue date: 2014Fundação Oswaldo Cruz. Centro de Pesquisas René Rachou. Laboratório de Malária.Belo Horizonte, MG, Brazil/Universidade Federal de Minas Gerais. Faculdade de Medicina. Belo Horizonte, MG, BrazilCentro de Pesquisas em Medicina Tropical de Rondônia. Porto Velho, RO, BrazilUniversidade Federal de Minas Gerais. Faculdade de Medicina. Belo Horizonte, MG, BrazilUniversidade Federal de Minas Gerais. Faculdade de Medicina. Belo Horizonte, MG, BrazilFundação Oswaldo Cruz. Centro de Pesquisas René Rachou. Laboratório de Malária. Belo Horizonte, MG, Brazil/Universidade Federal de Minas Gerais. Faculdade de Medicina. Belo Horizonte, MG, BrazilBackground: Chloroquine (CQ), a cost effective antimalarial drug with a relatively good safety profile and therapeutic index, is no longer used by itself to treat patients with Plasmodium falciparum due to CQ-resistant strains. P. vivax, representing over 90% of malaria cases in Brazil, despite reported resistance, is treated with CQ as well as with primaquine to block malaria transmission and avoid late P. vivax malaria relapses. Resistance to CQ and other antimalarial drugs influences malaria control, thus monitoring resistance phenotype by parasite genotyping is helpful in endemic areas. Methods: A total of 47 P. vivax and nine P. falciparum fresh isolates were genetically characterized and tested for CQ, mefloquine (MQ) and artesunate (ART) susceptibility in vitro. The genes mdr1 and pfcrt, likely related to CQ resistance, were analyzed in all isolates. Drug susceptibility was determined using short-term parasite cultures of ring stages for 48 to 72 hour and thick blood smears counts. Each parasite isolate was tested with the antimalarials to measure the geometric mean of 50% inhibitory concentration. Results; The low numbers of P. falciparum isolates reflect the species prevalence in Brazil; most displayed low sensitivity to CQ (IC50 70 nM). However, CQ resistance was rare among P. vivax isolates (IC50 of 32 nM). The majority of P. vivax and P. falciparum isolates were sensitive to ART and MQ. One hundred percent ofP. falciparum isolates carried non-synonymous mutations in the pfmdr1 gene in codons 184, 1042 and 1246, 84% in codons 1034 and none in codon 86, a well-known resistance mutation. For the pfcrt gene, mutations were observed in codons 72 and 76 in all P. falciparum isolates. One P. falciparum isolate from Angola, Africa, showing sensitivity to the antimalarials, presented no mutations. In P. vivax, mutations of pvmdr1 and the multidrug resistance gene 1 marker at codon F976 were absent. Conclusion: All P. falciparum Brazilian isolates showed CQ resistance and presented non-synonymous mutations inpfmdr1 and pfcrt. CQ resistant genotypes were not present among P. vivax isolates and the IC50 values were low in all samples of the Brazilian West Amazon

Proteomic Analysis of Urine from Patients with Plasmodium vivax Malaria Unravels a Unique Plasmodium vivax Protein That Is Absent from Plasmodium falciparum

Five species of Plasmodium cause malaria in humans and two of them, P. vivax and P. falciparum, pose the greatest threat. Rapid antigen detection tests (RADT) have been used for many years to diagnose and distinguish malaria caused by these two parasites. P. falciparum malaria can single-handedly be diagnosed using an RADT, which detects the unique P. falciparum specific histidine-rich protein 2 (HRP2). Unfortunately, there is no RADT that can single-handedly diagnose P. vivax malaria because no specific marker of this parasite has yet been described. Here, we report the discovery of a unique P. vivax protein (Vir14, NCBI Reference Sequence: XP_001612449.1) that has no sequence similarity with proteins of P. falciparum and no significant similarities with proteins of other species of Plasmodium. We propose that this protein could be an outstanding candidate molecule for the development of a promising RADT that can single-handedly and specifically diagnose P. vivax malaria

Malaria primes the innate immune response due to interferon-γ induced enhancement of toll-like receptor expression and function

Malaria-induced sepsis is associated with an intense proinflammatory cytokinemia for which the underlying mechanisms are poorly understood. It has been demonstrated that experimental infection of humans with Plasmodium falciparum primes Toll-like receptor (TLR)-mediated proinflammatory responses. Nevertheless, the relevance of this phenomenon during natural infection and, more importantly, the mechanisms by which malaria mediates TLR hyperresponsiveness are unclear. Here we show that TLR responses are boosted in febrile patients during natural infection with P. falciparum. Microarray analyses demonstrated that an extraordinary percentage of the up-regulated genes, including genes involving TLR signaling, had sites for IFN-inducible transcription factors. To further define the mechanism involved in malaria-mediated “priming,” we infected mice with Plasmodium chabaudi. The human data were remarkably predictive of what we observed in the rodent malaria model. Malaria-induced priming of TLR responses correlated with increased expression of TLR mRNA in a TLR9-, MyD88-, and IFNγ-dependent manner. Acutely infected WT mice were highly susceptible to LPS-induced lethality while TLR9−/−, IL12−/− and to a greater extent, IFNγ−/− mice were protected. Our data provide unprecedented evidence that TLR9 and MyD88 are essential to initiate IL12 and IFNγ responses and favor host hyperresponsiveness to TLR agonists resulting in overproduction of proinflammatory cytokines and the sepsis-like symptoms of acute malaria

Plasmodium vivax infection impairs regulatory T-Cell suppressive function during acute malaria

The balance between pro- and antiinflammatory mechanisms is essential to limit immune-mediated pathology, and CD4+ forkhead box P3 (Foxp3+) regulatory T cells (Treg) play an important role in this process. The expression of inhibitory receptors regulates cytokine production by Plasmodium vivax-specific T cells. Our goal was to assess the induction of programmed death-1 (PD-1) and cytotoxic T-lymphocyte antigen (CTLA-4) on Treg during malaria and to evaluate their function. We found that P. vivax infection triggered an increase in circulating Treg and their expression of CTLA-4 and PD-1. Functional analysis demonstrated that Treg from malaria patients had impaired suppressive ability and PD-1+Treg displayed lower levels of Foxp3 and Helios, but had higher frequencies of T-box transcription factor+ and interferon-gamma+ cells than PD-1−Treg. Thus malaria infection alters the function of circulating Treg by triggering increased expression of PD-1 on Treg that is associated with decreased regulatory function and increased proinflammatory characteristics

CD4+ T cells apoptosis in Plasmodium vivax infection is mediated by activation of both intrinsic and extrinsic pathways

Submitted by Repositório Arca ([email protected]) on 2019-04-24T17:44:00Z No. of bitstreams: 1 license.txt: 1748 bytes, checksum: 8a4605be74aa9ea9d79846c1fba20a33 (MD5)Approved for entry into archive by Janaína Nascimento ([email protected]) on 2019-08-22T12:31:14Z (GMT) No. of bitstreams: 2 ve_Hojo-Souza_Natália_etal_INI_2015.pdf: 673291 bytes, checksum: 5de19dfdb1440f05f52dfb3676dbc81a (MD5) license.txt: 1748 bytes, checksum: 8a4605be74aa9ea9d79846c1fba20a33 (MD5)Made available in DSpace on 2019-08-22T12:31:14Z (GMT). No. of bitstreams: 2 ve_Hojo-Souza_Natália_etal_INI_2015.pdf: 673291 bytes, checksum: 5de19dfdb1440f05f52dfb3676dbc81a (MD5) license.txt: 1748 bytes, checksum: 8a4605be74aa9ea9d79846c1fba20a33 (MD5) Previous issue date: 2015Federal University of Minas Gerais. Institute of Biological Science. Department of Parasitology. Belo Horizonte, MG, Brasil.Research Centre in Tropical Medicine. Porto Velho, RO, Brazil.Federal University of Minas Gerais. Institute of Biological Science. Department of Parasitology. Belo Horizonte, MG, Brasil.Federal University of Minas Gerais. Institute of Biological Science. Department of Parasitology. Belo Horizonte, MG, Brasil.Federal University of Minas Gerais. Institute of Biological Science. Department of Parasitology. Belo Horizonte, MG, Brasil.Federal University of Minas Gerais. Institute of Biological Science. Department of Parasitology. Belo Horizonte, MG, Brasil.Research Centre in Tropical Medicine. Porto Velho, RO, Brazil.Fundação Oswaldo Cruz. Instituto de Pesquisa Clínica Evandro Chagas. Rio de Janeiro, RJ, Brasil.Federal University of Minas Gerais. Institute of Biological Science. Department of Parasitology. Belo Horizonte, MG, Brasil.Federal University of Minas Gerais. Institute of Biological Science. Department of Parasitology. Belo Horizonte, MG, Brasil.Federal University of Minas Gerais. Institute of Biological Science. Department of Parasitology. Belo Horizonte, MG, Brasil.Background: Reduction in the number of circulating blood lymphocytes (lymphocytopaenia) has been reported during clinical episodes of malaria and is normalized after treatment with anti-malaria drugs. While this phenomenon is well established in malaria infection, the underlying mechanisms are still not fully elucidated. In the present study, the occurrence of apoptosis and its pathways in CD4+ T cells was investigated in naturally Plasmodium vivax-infected individuals from a Brazilian endemic area (Porto Velho – RO). Methods: Blood samples were collected from P. vivax-infected individuals and healthy donors. The apoptosis was characterized by cell staining with Annexin V/FITC and propidium iodide and the apoptosis-associated gene expression profile was carried out using RT2 Profiler PCR Array–Human Apoptosis. The plasma TNF level was determined by ELISA. The unpaired t-test or Mann–Whitney test was applied according to the data distribution. Results: Plasmodium vivax-infected individuals present low number of leukocytes and lymphocytes with a higher percentage of CD4+ T cells in early and/or late apoptosis. Increased gene expression was observed for TNFRSF1B and Bid, associated with a reduction of Bcl-2, in individuals with P. vivax malaria. Furthermore, these individuals showed increased plasma levels of TNF compared to malaria-naive donors. Conclusions: The results of the present study suggest that P. vivax infection induces apoptosis of CD4+ T cells mediated by two types of signaling: by activation of the TNFR1 death receptor (extrinsic pathway), which is amplified by Bid, and by decreased expression of the anti-apoptotic protein Bcl-2 (intrinsic pathway). The T lymphocytes apoptosis could reflect a strategy of immune evasion triggered by the parasite, enabling their persistence but also limiting the occurrence of immunopathology

An engineered vaccine of the Plasmodium vivax Duffy binding protein enhances induction of broadly neutralizing antibodies

Abstract Plasmodium vivax invasion into human reticulocytes is a complex process. The Duffy binding protein (DBP) dimerization with its cognate receptor is vital for junction formation in the invasion process. Due to its functional importance, DBP is considered a prime vaccine candidate, but variation in B-cell epitopes at the dimer interface of DBP leads to induction of strain-limited immunity. We believe that the polymorphic residues tend to divert immune responses away from functionally conserved epitopes important for receptor binding or DBP dimerization. As a proof of concept, we engineered the vaccine DEKnull to ablate the dominant Bc epitope to partially overcome strain-specific immune antibody responses. Additional surface engineering on the next generation immunogen, DEKnull-2, provides an immunogenicity breakthrough to conserved protective epitopes. DEKnull-2 elicits a stronger broadly neutralizing response and reactivity with long-term persistent antibody responses of acquired natural immunity. By using novel engineered DBP immunogens, we validate that the prime targets of protective immunity are conformational epitopes at the dimer interface. These successful results indicate a potential approach that can be used generally to improve efficacy of other malaria vaccine candidates

DNA-Containing Immunocomplexes Promote Inflammasome Assembly and Release of Pyrogenic Cytokines by CD14+ CD16+ CD64high CD32low Inflammatory Monocytes from Malaria Patients

Submitted by Nuzia Santos ([email protected]) on 2016-04-05T17:25:39Z No. of bitstreams: 1 DNA-Containing Immunocomplex (...) Monocytes from Malaria Patients.pdf: 4339793 bytes, checksum: 1b4586b816c6971792523290aa455335 (MD5)Approved for entry into archive by Nuzia Santos ([email protected]) on 2016-04-05T17:31:16Z (GMT) No. of bitstreams: 1 DNA-Containing Immunocomplex (...) Monocytes from Malaria Patients.pdf: 4339793 bytes, checksum: 1b4586b816c6971792523290aa455335 (MD5)Made available in DSpace on 2016-04-05T17:31:16Z (GMT). No. of bitstreams: 1 DNA-Containing Immunocomplex (...) Monocytes from Malaria Patients.pdf: 4339793 bytes, checksum: 1b4586b816c6971792523290aa455335 (MD5) Previous issue date: 2015Fundação Oswaldo Cruz. Centro de Pesquisa René Rachou. Belo Horizonte, MG, Brasil.University of Massachusetts Medical School. Division of Infectious Diseases and Immunology. Worcester, MA, USA / Centro Internacional de Entrenamiento e Investigaciones Medicas. Cali, Colombia.Fundação Oswaldo Cruz. Centro de Pesquisa René Rachou. Belo Horizonte, MG, Brasil / Universidade Federal de Minas Gerais. Instituto de Ciências Biológicas. Departamento de Bioquímica e Imunologia. Belo Horizonte, MG, Brasil.Fundação Oswaldo Cruz. Centro de Pesquisa René Rachou. Belo Horizonte, MG, Brasil / University of Massachusetts Medical School. Division of Infectious Diseases and Immunology. Worcester, MA, USA.Universidade Federal de Minas Gerais. Instituto de Ciências Biológicas. Departamento de Bioquímica e Imunologia. Belo Horizonte, MG, Brasil.Fundação Oswaldo Cruz. Centro de Pesquisa René Rachou. Belo Horizonte, MG, Brasil / Universidade Federal de Minas Gerais. Instituto de Ciências Biológicas. Departamento de Bioquímica e Imunologia. Belo Horizonte, MG, Brasil.University of Massachusetts Medical School. Division of Infectious Diseases and Immunology. Worcester, MA, USA.Centro de Pesquisa em Medicina Tropical. Porto Velho, RO, Brasil.University of California San Diego. School of Medicine. San Diego, CA, USA.The Feinstein Institute for Medical Research. Center for Autoimmune and Musculoskeletal Diseases. Manhasset, NY, USA.University of Massachusetts Medical School. Division of Infectious Diseases and Immunology. Worcester, MA, USA.Fundação Oswaldo Cruz. Centro de Pesquisa René Rachou. Belo Horizonte, MG, Brasil / University of Massachusetts Medical School. Division of Infectious Diseases and Immunology. Worcester, MA, USA.Fundação Oswaldo Cruz. Centro de Pesquisa René Rachou. Belo Horizonte, MG, Brasil / University of Massachusetts Medical School. Division of Infectious Diseases and Immunology. Worcester, MA, USA / Universidade Federal de Minas Gerais. Instituto de Ciências Biológicas. Departamento de Bioquímica e Imunologia. Belo Horizonte, MG, Brasil.High levels of circulating immunocomplexes (ICs) are found in patients with either infectious or sterile inflammation. We report that patients with either Plasmodium falciparum or Plasmodium vivax malaria have increased levels of circulating anti-DNA antibodies and ICs containing parasite DNA. Upon stimulation with malaria-induced ICs, monocytes express an NF-κB transcriptional signature. The main source of IC-induced proinflammatory cytokines (i.e., tumor necrosis factor alpha [TNF-α] and interleukin-1β [IL-1β])in peripheral blood mononuclear cells from acute malaria patients was found to be a CD14+ CD16 (FcγRIIIA)+ CD64 (FcγRI)high CD32 (FcγRIIB)low monocyte subset. Monocytes from convalescent patients were predominantly of the classical phenotype (CD14+ CD16−) that produces high levels of IL-10 and lower levels of TNF-α and IL-1β in response to ICs. Finally, we report a novel role for the proinflammatory activity of ICs by demonstrating their ability to induce inflammasome assembly and caspase-1 activation in human monocytes. These findings illuminate our understanding of the pathogenic role of ICs and monocyte subsets and may be relevant for future development of immunity-based interventions with broad applications to systemic inflammatory diseases