The Experts below are selected from a list of 291 Experts worldwide ranked by ideXlab platform
Photini Sinnis - One of the best experts on this subject based on the ideXlab platform.
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halobacterium expression system for production of full length plasmodium falciparum Circumsporozoite Protein
2016Co-Authors: Priya Dassarma, Ram Karan, Photini Sinnis, Wolf T Pecher, Shiladitya DassarmaAbstract:We recently developed a novel expression system employing the halophilic Archaeon Halobacterium sp. NRC-1 for scaled-up production and nanoparticle-display of antigenic Proteins. Here, we have targeted the major human parasite Plasmodium falciparum Circumsporozoite Protein (CSP), which is of interest for formulation of a protective malaria vaccine. A codon-optimized synthetic gene coding the full-length CSP was inserted downstream of the strong promoter for gvpA, the major gas vesicle nanoparticle Protein gene, in the pDRK expression vector, and as a fusion to the gvpC Protein in the pSD expression vector for display on the surface of gas vesicle nanoparticles. We found that the pDRK-CSP expression plasmid programmed high-level production of full-length CSP and the pSD-CSP expression plasmid programmed production of a GvpC-CSP fusion Protein, for display on gas vesicle nanoparticles. The Halobacterium sp. expression system provides a novel approach and a potentially valuable technical advancement for the production of P. falciparum CSP for malaria vaccine development.
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proteolytic cleavage of the plasmodium falciparum Circumsporozoite Protein is a target of protective antibodies
The Journal of Infectious Diseases, 2015Co-Authors: Diego A Espinosa, Photini Sinnis, Gabriel M Gutierrez, Maricarmen Rojaslopez, Fidel P ZavalaAbstract:: Studies in animals and human volunteers demonstrate that antibodies against the repeat-region of the Plasmodium Circumsporozoite Protein (CSP) abrogate sporozoite infection. However, the realization that the N- and C- terminal regions flanking the repeats play essential roles in parasite infectivity raised the possibility that they could be targeted by protective antibodies. We characterized a monoclonal antibody (mAb5D5) specific for the N-terminus of the P. falciparum CSP, which inhibits the proteolytic cleavage of the CSP, a key requirement for parasite infection of hepatocytes. Adoptive transfer of mAb5D5 strongly inhibits the in vivo infection of sporozoites expressing the N-terminus of P. falciparum CSP, and this protection is greatly enhanced when combined with antirepeat antibodies. Our results show that antibodies interfering with molecular processes required for parasite infectivity can exert a strong in vivo protective activity and indicate that pre-erythrocytic vaccines against Plasmodium should include the CSP N-terminal region.
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the plasmodium Circumsporozoite Protein is proteolytically processed during cell invasion
Journal of Experimental Medicine, 2005Co-Authors: Alida Coppi, Consuelo Pinzonortiz, Christina Hutter, Photini SinnisAbstract:The Circumsporozoite Protein (CSP) is the major surface Protein of Plasmodium sporozoites, the infective stage of malaria. Although CSP has been extensively studied as a malaria vaccine candidate, little is known about its structure. Here, we show that CSP is proteolytically cleaved by a papain family cysteine protease of parasite origin. Our data suggest that the highly conserved region I, found just before the repeat region, contains the cleavage site. Cleavage occurs on the sporozoite surface when parasites contact target cells. Inhibitors of CSP processing inhibit cell invasion in vitro, and treatment of mice with E-64, a highly specific cysteine protease inhibitor, completely inhibits sporozoite infectivity in vivo.
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the binding of the Circumsporozoite Protein to cell surface heparan sulfate proteoglycans is required for plasmodium sporozoite attachment to target cells
Journal of Biological Chemistry, 2001Co-Authors: Consuelo Pinzonortiz, Jennifer F Friedman, Jeffrey D Esko, Photini SinnisAbstract:Abstract The major surface Protein of malaria sporozoites, the Circumsporozoite Protein, binds to heparan sulfate proteoglycans on the surface of hepatocytes. It has been proposed that this binding event is responsible for the rapid and specific localization of sporozoites to the liver after their injection into the skin by an infected anopheline mosquito. Previous in vitro studies performed under static conditions have failed to demonstrate a significant role for heparan sulfate proteoglycans during sporozoite invasion of cells. We performed sporozoite attachment and invasion assays under more dynamic conditions and found a dramatic decrease in sporozoite attachment to cells in the presence of heparin. In contrast to its effect on attachment, heparin does not appear to have an effect on sporozoite invasion of cells. When substituted heparins were used as competitive inhibitors of sporozoite attachment, we found that sulfation of the glycosaminoglycan chains at both the N- and O-positions was important for sporozoite adhesion to cells. We conclude that the binding of the Circumsporozoite Protein to hepatic heparan sulfate proteoglycans is likely to function during sporozoite attachment in the liver and that this adhesion event depends on the sulfated glycosaminoglycan chains of the proteoglycans.
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ANOPHELES STEPHENSI SALIVARY GLANDS BEAR RECEPTORS FOR REGION I OF THE Circumsporozoite Protein OF PLASMODIUM FALCIPARUM
Molecular and Biochemical Parasitology, 1997Co-Authors: Sacha Sidjanski, Jerome P. Vanderberg, Photini SinnisAbstract:In the mosquito, Plasmodium sporozoites rupture from oocysts found on the midgut wall, circulate in the hemolymph and invade salivary glands where they wait to be injected into a vertebrate host during a bloodmeal. The mechanisms by which sporozoites specifically attach to and invade salivary glands are not known but evidence suggests that it is a receptor-mediated process. Here we show that the major surface Protein of sporozoites, the Circumsporozoite Protein (CS), binds preferentially to salivary glands when compared to other organs exposed to the circulating hemolymph. In addition, we show that a peptide encompassing region I, a highly conserved sequence found in all rodent and primate Plasmodium CS Proteins, inhibits binding of CS to mosquito salivary glands.
Victor Nussenzweig - One of the best experts on this subject based on the ideXlab platform.
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the Circumsporozoite Protein is an immunodominant protective antigen in irradiated sporozoites
Nature, 2006Co-Authors: Kota Arun Kumar, Fidel P Zavala, Ruth S. Nussenzweig, Gen Ichiro Sano, Silvia Beatriz Boscardin, Michel C Nussenzweig, Victor NussenzweigAbstract:Malaria infection starts when mosquitoes inject sporozoites into the skin. The parasites enter the blood stream and make their way to the liver where they develop into the exo-erythrocytic forms (EEFs). Immunization with irradiated sporozoites (IrSp) leads to robust protection against malaria infection in rodents1, monkeys2 and humans3 by eliciting antibodies to Circumsporozoite Protein (CS) that inhibit sporozoite infectivity, and T cells that destroy the EEFs4. To study the role of non-CS antigens in protection, we produced CS transgenic mice that were tolerant to CS T-cell epitopes. Here we show that in the absence of T-cell-dependent immune responses to CS, protection induced by immunization with two doses of IrSp was greatly reduced. Thus, although hundreds of other Plasmodium genes are expressed in sporozoites5 and EEFs6, CS is a dominant protective antigen. Nevertheless, sterile immunity could be obtained by immunization of CS transgenics with three doses of IrSp.
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cutting edge a new tool to evaluate human pre erythrocytic malaria vaccines rodent parasites bearing a hybrid plasmodium falciparum Circumsporozoite Protein
Journal of Immunology, 2002Co-Authors: Cathrine Persson, Victor Nussenzweig, Giane A Oliveira, Ali A Sultan, Purnima Bhanot, Elizabeth NardinAbstract:Malaria vaccines containing the Plasmodium falciparum Circumsporozoite Protein repeat domain are undergoing human trials. There is no simple method to evaluate the effect of vaccine-induced responses on P. falciparum sporozoite infectivity. Unlike the rodent malaria Plasmodium berghei, P. falciparum sporozoites do not infect common laboratory animals and only develop in vitro in human hepatocyte cultures. We generated a recombinant P. berghei parasite bearing P. falciparum Circumsporozoite Protein repeats. These hybrid sporozoites are fully infective in vivo and in vitro. Monoclonal and polyclonal Abs to P. falciparum repeats neutralize hybrid parasite infectivity, and mice immunized with a P. falciparum vaccine are protected against challenge with hybrid sporozoites.
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levels of Circumsporozoite Protein in the plasmodium oocyst determine sporozoite morphology
The EMBO Journal, 2002Co-Authors: Victor Nussenzweig, Ruth S. Nussenzweig, Vandana Thathy, Hisashi Fujioka, Soren Gantt, Robert MenardAbstract:The sporozoite stage of the Plasmodium parasite is formed by budding from a multinucleate oocyst in the mosquito midgut. During their life, sporozoites must infect the salivary glands of the mosquito vector and the liver of the mammalian host; both events depend on the major sporozoite surface Protein, the Circumsporozoite Protein (CS). We previously reported that Plasmodium berghei oocysts in which the CS gene is inactivated do not form sporozoites. Here, we analyzed the ultrastructure of P.berghei oocyst differentiation in the wild type, recombinants that do not produce or produce reduced amounts of CS, and corresponding complemented clones. The results indicate that CS is essential for establishing polarity in the oocyst. The amounts of CS Protein correlate with the extent of development of the inner membranes and associated microtubules underneath the oocyst outer membrane, which normally demarcate focal budding sites. This is a first example of a Protein controlling both morphogenesis and infectivity of a parasite stage.
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Circumsporozoite Protein is required for development of malaria sporozoites in mosquitoes
Nature, 1997Co-Authors: Robert Menard, Ruth S. Nussenzweig, Ali A Sultan, Claudio Cortes, Rita Altszuler, Melissa R Van Dijk, Chris J Janse, Andrew P Waters, Victor NussenzweigAbstract:Malaria parasites undergo a sporogonic cycle in the mosquito vector. Sporozoites, the form of the parasite injected into the host during a bloodmeal, develop inside oocysts in the insect midgut, then migrate to and eventually invade the salivary glands. The Circumsporozoite Protein (CS), one of the major Proteins synthesized by salivary gland sporozoites1, is a surface-associated molecule which is important in sporozoite infectivity to the host2. Here, by gene targeting, we created Plasmodium berghei lines in which the single-copy CS gene was disrupted. The CS( − ) and wild-type parasites produced similar numbers of oocysts of comparable size in the mosquito midgut. In the CS( − ) oocysts, however, sporozoite formation was profoundly inhibited. CS therefore appears to have a pleiotropic role and to be vital for malaria parasites in both the vector and the host: in mosquitoes, CS is essential for sporozoite development within oocysts, and in the vertebrate host it promotes sporozoite attachment to hepatocytes3–7.
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structural and functional properties of region ii plus of the malaria Circumsporozoite Protein
Journal of Experimental Medicine, 1994Co-Authors: Photini Sinnis, Carla Cerami, Pedro Clavijo, David Fenyo, Brian T Chait, Victor NussenzweigAbstract:Stlmmary During feeding, infected mosquitos inject malaria sporozoites into the host circulation. Within minutes, the parasites are found in the liver where they initiate the first stage of malaria infection. All species of malaria sporozoites are uniformly covered by the Circumsporozoite Protein (CS), which contains a conserved COOH-terminal sequence called region II-plus. We have previously shown that region II-plus is the parasite's hepatocyte-binding ligand and that this ligand binds to heparan sulfate proteoglycans (HSPGs) on the hepatocyte membrane. Using a series of substituted region II-plus peptides, we show here that the downstream basic amino acids as well as the interdispersed hydrophobic residues are required for binding of CS to hepatocyte HSPGs. We also show that this positively charged stretch of amino acids must be aggregated in order to bind to the receptor. On the basis of this information, we have synthesized a multiple antigen peptide that mimics the hepatocyte-binding ligand. This construct inhibits both CS binding to HepG2 cells in vitro as well as CS clearance in mice.
Fidel P Zavala - One of the best experts on this subject based on the ideXlab platform.
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proteolytic cleavage of the plasmodium falciparum Circumsporozoite Protein is a target of protective antibodies
The Journal of Infectious Diseases, 2015Co-Authors: Diego A Espinosa, Photini Sinnis, Gabriel M Gutierrez, Maricarmen Rojaslopez, Fidel P ZavalaAbstract:: Studies in animals and human volunteers demonstrate that antibodies against the repeat-region of the Plasmodium Circumsporozoite Protein (CSP) abrogate sporozoite infection. However, the realization that the N- and C- terminal regions flanking the repeats play essential roles in parasite infectivity raised the possibility that they could be targeted by protective antibodies. We characterized a monoclonal antibody (mAb5D5) specific for the N-terminus of the P. falciparum CSP, which inhibits the proteolytic cleavage of the CSP, a key requirement for parasite infection of hepatocytes. Adoptive transfer of mAb5D5 strongly inhibits the in vivo infection of sporozoites expressing the N-terminus of P. falciparum CSP, and this protection is greatly enhanced when combined with antirepeat antibodies. Our results show that antibodies interfering with molecular processes required for parasite infectivity can exert a strong in vivo protective activity and indicate that pre-erythrocytic vaccines against Plasmodium should include the CSP N-terminal region.
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adenovirus particles that display the plasmodium falciparum Circumsporozoite Protein nanp repeat induce sporozoite neutralizing antibodies in mice
Vaccine, 2011Co-Authors: Christopher Palma, Fidel P Zavala, Michael G Overstreet, Jean Marc Guedon, Egbert Hoiczyk, Cameron Ward, Kasey A Karen, Gary KetnerAbstract:Adenovirus particles can be engineered to display exogenous peptides on their surfaces by modification of viral capsid Proteins, and particles that display pathogen-derived peptides can induce protective immunity. We constructed viable recombinant adenoviruses that display B-cell epitopes from the Plasmodium falciparum Circumsporozoite Protein (PfCSP) in the major adenovirus capsid Protein, hexon. Recombinants induced high-titer antibodies against CSP when injected intraperitoneally into mice. Serum obtained from immunized mice recognized both recombinant PfCSP Protein and P. falciparum sporozoites, and neutralized P. falciparum sporozoites in vitro. Replicating adenovirus vaccines have provided economical protection against adenovirus disease for over three decades. The recombinants described here may provide a path to an affordable malaria vaccine in the developing world.
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the Circumsporozoite Protein is an immunodominant protective antigen in irradiated sporozoites
Nature, 2006Co-Authors: Kota Arun Kumar, Fidel P Zavala, Ruth S. Nussenzweig, Gen Ichiro Sano, Silvia Beatriz Boscardin, Michel C Nussenzweig, Victor NussenzweigAbstract:Malaria infection starts when mosquitoes inject sporozoites into the skin. The parasites enter the blood stream and make their way to the liver where they develop into the exo-erythrocytic forms (EEFs). Immunization with irradiated sporozoites (IrSp) leads to robust protection against malaria infection in rodents1, monkeys2 and humans3 by eliciting antibodies to Circumsporozoite Protein (CS) that inhibit sporozoite infectivity, and T cells that destroy the EEFs4. To study the role of non-CS antigens in protection, we produced CS transgenic mice that were tolerant to CS T-cell epitopes. Here we show that in the absence of T-cell-dependent immune responses to CS, protection induced by immunization with two doses of IrSp was greatly reduced. Thus, although hundreds of other Plasmodium genes are expressed in sporozoites5 and EEFs6, CS is a dominant protective antigen. Nevertheless, sterile immunity could be obtained by immunization of CS transgenics with three doses of IrSp.
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Studies using a recombinant vaccinia virus expressing the Circumsporozoite Protein of Plasmodium berghei
Molecular and Biochemical Parasitology, 1991Co-Authors: Vijaya Satchidanandam, Fidel P Zavala, Bernard MossAbstract:A recombinant vaccinia virus was constructed which expressed the Circumsporozoite Protein of Plasmodium berghei. Four different strains of mice belonging to different haplotypes were immunized with the recombinant virus. The antibody response to the Circumsporozoite Protein as well as to vaccinia virus varied among the strains, independently of each other. The antiCircumsporozoite Protein titers were comparable to that obtained on immunization with irradiated sporozoites. Spleen cells from H2d mice immunized with P. berghei sporozoites showed a significant proliferative response when cultured in vitro with a low multiplicity of the recombinant vaccinia virus. A weak cytotoxic T lymphocyte response specifically targeting the Circumsporozoite Protein could be identified in spleens of BALB/c (H2d) mice immunized with vaccinia virus when BALB 3T3 cells transformed with a plasmid expressing the Circumsporozoite Protein under control of the simian virus 40 promoter were used as target cells in the cytotoxic T lymphocyte assay. However, none of the recombinant virus-immunized animals could be protected from a challenge of sporozoites even at the lowest dose of parasite used.
Srdjan M Dragovic - One of the best experts on this subject based on the ideXlab platform.
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anopheles gambiae Circumsporozoite Protein binding Protein facilitates plasmodium infection of mosquito salivary glands
The Journal of Infectious Diseases, 2013Co-Authors: Jiuling Wang, Yue Zhang, Yang O Zhao, Michelle W M Li, Lili Zhang, Srdjan M DragovicAbstract:Malaria, a mosquito-borne disease caused by Plasmodium species, causes substantial morbidity and mortality throughout the world. Plasmodium sporozoites mature in oocysts formed in the mosquito gut wall and then invade the salivary glands, where they remain until transmitted to the vertebrate host during a mosquito bite. The Plasmodium Circumsporozoite Protein (CSP) binds to salivary glands and plays a role in the invasion of this organ by sporozoites. We identified an Anopheles salivary gland Protein, named CSP-binding Protein (CSPBP), that interacts with CSP. Downregulation of CSPBP in mosquito salivary glands inhibited invasion by Plasmodium organisms. In vivo bioassays showed that mosquitoes that were fed blood with CSPBP antibody displayed a 25% and 90% reduction in the parasite load in infected salivary glands 14 and 18 days after the blood meal, respectively. These results suggest that CSPBP is important for the infection of the mosquito salivary gland by Plasmodium organisms and that blocking CSPBP can interfere with the Plasmodium life cycle.
Emily Locke - One of the best experts on this subject based on the ideXlab platform.
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characterization of two in vivo challenge models to measure functional activity of monoclonal antibodies to plasmodium falciparum Circumsporozoite Protein
Malaria Journal, 2020Co-Authors: Rama Raghunandan, Bryan T Mayer, Yevel Floresgarcia, Monica W Gerber, Raphael Gottardo, Hugo Jhun, Sonia M Herrera, Daniel W Perezramos, Emily LockeAbstract:New strategies are needed to reduce the incidence of malaria, and promising approaches include the development of vaccines and monoclonal antibodies (mAbs) that target the Circumsporozoite Protein (CSP). To select the best candidates and speed development, it is essential to standardize preclinical assays to measure the potency of such interventions in animal models. Two assay configurations were studied using transgenic Plasmodium berghei expressing Plasmodium falciparum full-length Circumsporozoite Protein. The assays measured (1) reduction in parasite infection of the liver (liver burden) following an intravenous (i.v) administration of sporozoites and (2) protection from parasitaemia following mosquito bite challenge. Two human CSP mAbs, AB311 and AB317, were compared for their ability to inhibit infection. Multiple independent experiments were conducted to define assay variability and resultant impact on the ability to discriminate differences in mAb functional activity. Overall, the assays produced highly consistent results in that all individual experiments showed greater functional activity for AB317 compared to AB311 as calculated by the dose required for 50% inhibition (ID50) as well as the serum concentration required for 50% inhibition (IC50). The data were then used to model experimental designs with adequate statistical power to rigorously screen, compare, and rank order novel anti-CSP mAbs. The results indicate that in vivo assays described here can provide reliable information for comparing the functional activity of mAbs. The results also provide guidance regarding selection of the appropriate experimental design, dose selection, and group sizes.
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Characterization of two in vivo challenge models to measure functional activity of monoclonal antibodies to Plasmodium falciparum Circumsporozoite Protein
2020Co-Authors: Rama Raghunandan, Emily Locke, Bryan T Mayer, Monica W Gerber, Raphael Gottardo, Hugo Jhun, Sonia M Herrera, Yevel Flores-garcia, Daniel W Perez-ramos, C Richter KingAbstract:Abstract Background New strategies are needed to reduce the incidence of malaria, and promising approaches include the development of vaccines and monoclonal antibodies (mAbs) that target the Circumsporozoite Protein (CSP). To select the best candidates and speed development, it is essential to standardize preclinical assays to measure the potency of such interventions in animal models. Methods Two assay configurations were studied using transgenic Plasmodium berghei expressing Plasmodium falciparum full-length Circumsporozoite Protein. The assays measured 1) reduction in parasite infection of the liver (liver burden) following an intravenous (i.v) administration of sporozoites and 2) protection from parasitaemia following mosquito bite challenge. Two human CSP mAbs, AB311 and AB317, were compared for their ability to inhibit infection. Multiple independent experiments were conducted to define assay variability and resultant impact on the ability to discriminate differences in mAb functional activity. Results Overall, the assays produced highly consistent results in that all individual experiments showed greater functional activity for AB317 compared to AB311 as calculated by the dose required for 50% inhibition (ID50) as well as the serum concentration required for 50% inhibition (IC50). The data were then used to model experimental designs with adequate statistical power to rigorously screen, compare, and rank order novel anti-CSP mAbs. Conclusion The results indicate that in vivo assays described here can provide reliable information for comparing the functional activity of mAbs. The results also provide guidance regarding selection of the appropriate experimental design, dose selection, and group sizes.
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diverse antibody responses to conserved structural motifs in plasmodium falciparum Circumsporozoite Protein
Journal of Molecular Biology, 2020Co-Authors: T Pholcharee, David Oyen, Jonathan L. Torres, Emily Locke, Yevel Floresgarcia, G M Martin, Gonzalo E Gonzalezpaez, Daniel Emerling, Wayne Volkmuth, C R KingAbstract:Abstract Malaria vaccine candidate RTS,S/AS01 is based on the central and C-terminal regions of the Circumsporozoite Protein (CSP) of P. falciparum. mAb397 was isolated from a volunteer in an RTS,S/AS01 clinical trial, and it protects mice from infection by malaria sporozoites. However, mAb397 originates from the less commonly used VH3-15 germline gene compared to the VH3-30/33 antibodies generally elicited by RTS,S to the central NANP repeat region of CSP. The crystal structure of mAb397 with an NPNA4 peptide shows that the central NPNA forms a type I β-turn and is the main recognition motif. In most anti-NANP antibodies studied to date, a germline-encoded Trp is used to engage the Pro in NPNA β-turns, but here the Trp interacts with the first Asn. This “conserved” Trp, however, can arise from different germline genes and be located in the heavy or the light chain. Variation in the terminal ψ angles of the NPNA β-turns results in different dispositions of the subsequent NPNA and, hence, different stoichiometries and modes of antibody binding to rsCSP. Diverse protective antibodies against NANP repeats are therefore not limited to a single germline gene response or mode of binding.
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A comparison of Plasmodium falciparum Circumsporozoite Protein-based slot blot and ELISA immuno-assays for oocyst detection in mosquito homogenates
Malaria Journal, 2015Co-Authors: Will Stone, Bryan Grabias, Kjerstin Lanke, Hong Zheng, Emily Locke, Diadier Diallo, Ashley Birkett, Merribeth J. Morin, Teun Bousema, Sanjai KumarAbstract:Background The infectivity of Plasmodium gametocytes is typically determined by microscopically examining the midguts of mosquitoes that have taken a blood meal containing potentially infectious parasites. Such assessments are required for the development and evaluation of transmission-reducing interventions (TRI), but are limited by subjectivity, technical complexity and throughput. The detection of Circumsporozoite Protein (CSP) by enzyme-linked immunosorbent assay (ELISA) and enhanced chemiluminescent slot-blot (ECL-SB) may be used as objective, scalable alternatives to microscopy for the determination of infection prevalence.