Search

Chora and colleagues show that infection of the liver by Plasmodium modulates severity of disease in the experimental cerebral malaria (ECM) model by generating gamma delta (ɣδ) T cells that produce IL-17. This work calls into question the long-standing assumption that liver infection does not modulate severity of malaria. [ABSTRACT FROM AUTHOR]

Plasmodium parasites belong to the Apicomplexan phylum, which consists mostly of obligate intracellular pathogens that vary dramatically in host cell tropism. Plasmodium sporozoites are highly hepatophilic. The specific molecular mechanisms, which facilitate sporozoite selection and successful infection of hepatocytes, remain poorly defined. Here, we discuss the parasite and host factors which are critical to hepatocyte infection. We derive a model where sporozoites initially select host cells that constitute a permissive environment and then further refine the chosen hepatocyte during liver stage development, ensuring life cycle progression. While many unknowns of pre-erythrocytic infection remain, advancing models and technologies that enable analysis of human malaria parasites and of single infected cells will catalyze a comprehensive understanding of the interaction between the malaria parasite and its hepatocyte host. [ABSTRACT FROM AUTHOR]

Summary: Plasmodium parasites infect the liver and replicate inside hepatocytes before they invade erythrocytes and trigger clinical malaria. Analysis of host signaling pathways affected by liver-stage infection could provide critical insights into host–pathogen interactions and reveal targets for intervention. Using protein lysate microarrays, we found that Plasmodium yoelii rodent malaria parasites perturb hepatocyte regulatory pathways involved in cell survival, proliferation, and autophagy. Notably, the prodeath protein p53 was substantially decreased in infected hepatocytes, suggesting that it could be targeted by the parasite to foster survival. Indeed, mice that express increased levels of p53 showed reduced liver-stage parasite burden, whereas p53 knockout mice suffered increased liver-stage burden. Furthermore, boosting p53 levels with the use of the small molecule Nutlin-3 dramatically reduced liver-stage burden in vitro and in vivo. We conclude that perturbation of the hepatocyte p53 pathway critically impacts parasite survival. Thus, host pathways might constitute potential targets for host-based antimalarial prophylaxis. [ABSTRACT FROM AUTHOR]

Abstract: The human malaria parasite Plasmodium falciparum causes the most deadly parasitic disease worldwide, necessitating the development of interventions that block infection. Yet, preclinical assays to measure inhibition of infection date from the 1980s and are based on microscopy. Here, we describe the development of a simple flow cytometric assay that can be used to quantitatively assess P. falciparum sporozoite infection in vitro in low and medium throughput. We demonstrate the utility of this assay for assessing both drug inhibition of infection and measuring efficacy of antibodies in blocking parasite infection. This methodology will aid in assessing functional antibody responses to vaccination and novel drugs that prevent mosquito-to-man transmission of malaria. [Copyright &y& Elsevier]

Both subunit and attenuated whole-sporozoite vaccination strategies against Plasmodium infection have shown promising initial results in malaria-naive westerners but less efficacy in malaria-exposed individuals in endemic areas. Here, we demonstrate proof of concept by using a rodent malaria model in which non-neutralizing antibodies (nNAbs) can directly interfere with protective anti-circumsporozoite protein (CSP) humoral responses. We characterize a monoclonal antibody, RAM1, against Plasmodium yoelii sporozoite major surface antigen CSP. Unlike the canonical Py CSP repeat domain binding and neutralizing antibody (NAb) 2F6, RAM1 does not inhibit sporozoite traversal or entry of hepatocytes in vitro or infection in vivo. Although 2F6 and RAM1 bind non-overlapping regions of the CSP-repeat domain, pre-treatment with RAM1 abrogates the capacity of NAb to block sporozoite traversal and invasion in vitro. Importantly, RAM1 reduces the efficacy of the polyclonal humoral response against Py CSP in vivo. Collectively, our data provide a proof of concept that nNAbs can alter the efficacy of malaria vaccination. [Display omitted] • We isolate a non-neutralizing antibody from Py CSP-immunized BALB/cJ mice • A non-neutralizing Ab, RAM1, abrogates the blocking capacity of an αCSP NAb • A non-neutralizing Ab, RAM1, reduces the vaccine efficacy in mice in vivo • Further exploration for the role of pre-existing nNAbs in the field is warranted Plasmodium CSP is a complex protein with multiple unique domains and repetitive motifs. Vijayan et al. isolate a non-neutralizing CSP major repeat binding antibody that interferes with the protective efficacy of NAbs. This implicates direct antibody interference as a mechanism by which pre-existing nNAbs may interfere with vaccine-elicited NAbs. [ABSTRACT FROM AUTHOR]

Summary Plasmodium parasites are highly selective when infecting hepatocytes and induce many changes within the host cell upon infection. While several host cell factors have been identified that are important for liver infection, our understanding of what facilitates the maintenance of infection remains incomplete. Here, we describe a role for phosphorylated ribosomal protein S6 (Ser235/236) (p-RPS6) in Plasmodium yoelii -infected hepatocytes. Blocking RPS6 phosphorylation prior to infection decreases the number of liver stage parasites within 24 h. Infected hepatocytes exhibit elevated levels of p-RPS6 while simultaneously abrogating the induction of phosphorylation of RPS6 in response to insulin stimulation. This is in contrast with the regulation of p-RPS6 by Toxoplasma gondii, which elevates levels of p-RPS6 after infection but does not alter the response to insulin. Our data support a model in which RPS6 phosphorylation is uncoupled from canonical regulators in Plasmodium- infected hepatocytes and is relied on by the parasite to maintain infection. Graphical Abstract Highlights • p-RPS6 is elevated in hepatocyte populations susceptible to Plasmodium (Py) • Inhibiting RPS6 phosphorylation prior to infection decreases Py infection • In Py- infected hepatocytes, canonical Akt/ RPS6 signaling is disrupted • Response to insulin is abrogated in Py- infected hepatocytes After mosquito-to-human transmission, Plasmodium parasites infect hepatocytes. Glennon et al. demonstrate that infected cells exhibit elevated levels of ribosomal protein S6 phosphorylation, and this phosphorylation appears uncoupled from canonical regulators. This work raises the possibility that Plasmodium -infected hepatocytes are governed by non-canonical, re-wired signal transduction cascades. [ABSTRACT FROM AUTHOR]

Despite the recent successes of artemisinin-based antimalarial drugs, many still die from severe malaria, and eradication efforts are hindered by the limited drugs currently available to target transmissible gametocyte parasites and liver-resident dormant Plasmodium vivax hypnozoites. Host-targeted therapy is a new direction for infectious disease drug development and aims to interfere with host molecules, pathways, or networks that are required for infection or that contribute to disease. Recent advances in our understanding of host pathways involved in parasite development and pathogenic mechanisms in severe malaria could facilitate the development of host-targeted interventions against Plasmodium infection and malaria disease. This review discusses new opportunities for host-targeted therapeutics for malaria and the potential to harness drug polypharmacology to simultaneously target multiple host pathways using a single drug intervention. Highlights Host-targeted therapy is a strategy for eliminating or reducing symptoms of infectious disease, and it holds promise for application to malaria. New insights into the malaria parasite–host interaction may lead to novel interventions to target liver-dormant forms called hypnozoites or to counteract the complications associated with cerebral malaria. Systematic approaches that exploit the polyphamacology of drugs developed for other indications could facilitate repurposing drugs that impact one or more points in the malaria parasite life cycle and also address severe complications of the disease. [ABSTRACT FROM AUTHOR]

Eliminating malaria parasites during the asymptomatic but obligate liver stages (LSs) of infection would stop disease and subsequent transmission. Unfortunately, only a single licensed drug that targets all LSs, Primaquine, is available. Targeting host proteins might significantly expand the repertoire of prophylactic drugs against malaria. Here, we demonstrate that both Bcl-2 inhibitors and P53 agonists dramatically reduce LS burden in a mouse malaria model in vitro and in vivo by altering the activity of key hepatocyte factors on which the parasite relies. Bcl-2 inhibitors act primarily by inducing apoptosis in infected hepatocytes, whereas P53 agonists eliminate parasites in an apoptosis-independent fashion. In combination, Bcl-2 inhibitors and P53 agonists act synergistically to delay, and in some cases completely prevent, the onset of blood stage disease. Both families of drugs are highly effective at doses that do not cause substantial hepatocyte cell death in vitro or liver damage in vivo. P53 agonists and Bcl-2 inhibitors were also effective when administered to humanized mice infected with Plasmodium falciparum. Our data demonstrate that host-based prophylaxis could be developed into an effective intervention strategy that eliminates LS parasites before the onset of clinical disease and thus opens a new avenue to prevent malaria. [ABSTRACT FROM AUTHOR]

Thrombopoietin (TPO) is the primary regulator of platelet production, affecting cell survival, proliferation, and differentiation through binding to and stimulation of the cell surface receptor the cellular myeloproliferative leukemia virus oncogene (c-Mpl). Activating mutations in c-Mpl constitutively stimulate downstream signaling pathways, leading to aberrant hematopoiesis, and contribute to development of myeloproliferative neoplasms. Several studies have mapped the tyrosine residues within the cytoplasmic domain of c-Mpl that mediate these cellular signals; however, secondary signaling pathways are incompletely understood. In this study, we focused on c-Mpl tyrosine 591 (Y591). We found Y591 of wild-type c-Mpl to be phosphorylated in the presence of TPO. Additionally, eliminating Y591 phosphorylation by mutation to Phe resulted in decreased total receptor phosphorylation. Using a Src homology 2/phosphotyrosine-binding (SH2/PTB) domain binding microarray, we identified novel c-Mpl binding partners for phosphorylated Y591, including Src homology region 2 domain-containing phosphatase-1 (SHP-1), spleen tyrosine kinase (SYK) and Bruton's tyrosine kinase (BTK). The functional significance of binding partners was determined through small interfering RNA treatment of Ba/F3-Mpl cells, confirming that the increase in pERK1/2 resulting from removal of Y591 may be mediated by spleen tyrosine kinase. These findings identify a novel negative regulatory pathway that controls TPO-mediated signaling, advancing our understanding of the mechanisms required for successful maintenance of hematopoietic stem cells and megakaryocyte development. [ABSTRACT FROM AUTHOR]

Immunization with live-attenuated Plasmodium sporozoites completely protects against malaria infection. Genetic engineering offers a versatile platform to create live-attenuated sporozoite vaccine candidates. We previously generated a genetically attenuated parasite (GAP) by deleting the P52 and P36 genes in the NF54 wild-type (WT) strain of Plasmodium falciparum (Pf p52−/p36− GAP). Preclinical assessment of p52−/p36− GAP in a humanized mouse model indicated an early and severe liver stage growth defect. However, human exposure to >200 Pf p52−/p36− GAP-infected mosquito bites in a safety trial resulted in peripheral parasitemia in one of six volunteers, revealing that this GAP was incompletely attenuated. We have now created a triple gene deleted GAP by additionally removing the SAP1 gene (Pf p52−/p36−/sap1− GAP) and employed flippase (FLP)/flippase recognition target (FRT) recombination for drug selectable marker cassette removal. This next-generation GAP was indistinguishable from WT parasites in blood stage and mosquito stage development. Using an improved humanized mouse model transplanted with human hepatocytes and human red blood cells, we show that despite a high-dose sporozoite challenge, Pf p52−/p36−/sap1− GAP did not transition to blood stage infection and appeared to be completely attenuated. Thus, clinical testing of Pf p52−/p36−/sap1− GAP assessing safety, immunogenicity, and efficacy against sporozoite challenge is warranted. [ABSTRACT FROM AUTHOR]

Abstract: Whole-parasite malaria vaccines have shown promise in clinical trials. We recently reported the first human trial of a malaria vaccine based on Plasmodium falciparum genetically attenuated parasites (PfGAP). Herein we report for the first time that PfGAP induces prolonged functional humoral responses in humans. Six volunteers were exposed to 5 bites of PfGAP-infected mosquitoes followed by approximately 200 bites. Plasma collected from all volunteers 3 months after the last exposure efficiently inhibits invasion of hepatocytes by P. falciparum sporozoites. The level of inhibition observed is comparable to that attained using plasma collected after 4–5 intravenously administrations of high numbers of irradiated sporozoites, validating the potential of PfGAP malaria vaccines. Our data highlight the role of antibody responses in pre-erythrocytic stages of human malaria, and suggests that to be protective, malaria vaccines might need to elicit long-lasting functional antibodies in addition to cellular responses. [Copyright &y& Elsevier]