scholarly journals Protective Humoral Immunity Elicited by a Needle-Free Malaria Vaccine Comprised of a Chimeric Plasmodium falciparum Circumsporozoite Protein and a Toll-Like Receptor 5 Agonist, Flagellin

2013 ◽  
Vol 81 (12) ◽  
pp. 4350-4362 ◽  
Author(s):  
Daniel Carapau ◽  
Robert Mitchell ◽  
Adéla Nacer ◽  
Alan Shaw ◽  
Caroline Othoro ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Humoral Immunity ◽  
Neutralizing Antibodies ◽  
Neutralizing Antibody ◽  
Content Type ◽  
Interleukin 5 ◽  
Toll Like Receptor 5 ◽  
Molecular Patterns

ABSTRACTImmunization withPlasmodiumsporozoites can elicit high levels of sterile immunity, and neutralizing antibodies from protected hosts are known to target the repeat region of the circumsporozoite (CS) protein on the parasite surface. CS-based subunit vaccines have been hampered by suboptimal immunogenicity and the requirement for strong adjuvants to elicit effective humoral immunity. Pathogen-associated molecular patterns (PAMPs) that signal through Toll-like receptors (TLRs) can function as potent adjuvants for innate and adaptive immunity. We examined the immunogenicity of recombinant proteins containing a TLR5 agonist, flagellin, and either full-length or selected epitopes of thePlasmodium falciparumCS protein. Mice immunized with either of the flagellin-modified CS constructs, administered intranasally (i.n.) or subcutaneously (s.c.), developed similar levels of malaria-specific IgG1 antibody and interleukin-5 (IL-5)-producing T cells. Importantly, immunization via the i.n. but not the s.c. route elicited sporozoite neutralizing antibodies capable of inhibiting >90% of sporozoite invasionin vitroandin vivo, as measured using a transgenic rodent parasite expressingP. falciparumCS repeats. These findings demonstrate that functional sporozoite neutralizing antibody can be elicited by i.n. immunization with a flagellin-modifiedP. falciparumCS protein and raise the potential of a scalable, safe, needle-free vaccine for the 40% of the world's population at risk of malaria.

scholarly journals Computational Chemogenomics Drug Repositioning Strategy Enables the Discovery of Epirubicin as a New Repurposed Hit for Plasmodium falciparum and P. vivax

2020 ◽  
Vol 64 (9) ◽  
Author(s):  
Letícia Tiburcio Ferreira ◽  
Juliana Rodrigues ◽  
Gustavo Capatti Cassiano ◽  
Tatyana Almeida Tavella ◽  
Kaira Cristina Peralis Tomaz ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
In Silico ◽  
Antimalarial Activity ◽  
Drug Repositioning ◽  
Dna Gyrase ◽  
Plasmodium Yoelii ◽  
Computer Assisted ◽  
Content Type

ABSTRACT Widespread resistance against antimalarial drugs thwarts current efforts for controlling the disease and urges the discovery of new effective treatments. Drug repositioning is increasingly becoming an attractive strategy since it can reduce costs, risks, and time-to-market. Herein, we have used this strategy to identify novel antimalarial hits. We used a comparative in silico chemogenomics approach to select Plasmodium falciparum and Plasmodium vivax proteins as potential drug targets and analyzed them using a computer-assisted drug repositioning pipeline to identify approved drugs with potential antimalarial activity. Among the seven drugs identified as promising antimalarial candidates, the anthracycline epirubicin was selected for further experimental validation. Epirubicin was shown to be potent in vitro against sensitive and multidrug-resistant P. falciparum strains and P. vivax field isolates in the nanomolar range, as well as being effective against an in vivo murine model of Plasmodium yoelii. Transmission-blocking activity was observed for epirubicin in vitro and in vivo. Finally, using yeast-based haploinsufficiency chemical genomic profiling, we aimed to get insights into the mechanism of action of epirubicin. Beyond the target predicted in silico (a DNA gyrase in the apicoplast), functional assays suggested a GlcNac-1-P-transferase (GPT) enzyme as a potential target. Docking calculations predicted the binding mode of epirubicin with DNA gyrase and GPT proteins. Epirubicin is originally an antitumoral agent and presents associated toxicity. However, its antiplasmodial activity against not only P. falciparum but also P. vivax in different stages of the parasite life cycle supports the use of this drug as a scaffold for hit-to-lead optimization in malaria drug discovery.

scholarly journals Identification of Synthetic Host Defense Peptide Mimics That Exert Dual Antimicrobial and Anti-Inflammatory Activities

2012 ◽  
Vol 19 (11) ◽  
pp. 1784-1791 ◽  
Author(s):  
Abhigyan Som ◽  
Nicolás Navasa ◽  
Avital Percher ◽  
Richard W. Scott ◽  
Gregory N. Tew ◽  
...  
Keyword(s):  
Antimicrobial Peptides ◽  
Lipoteichoic Acid ◽  
Antibacterial Activities ◽  
Innate Immune Responses ◽  
Content Type ◽  
Host Defense Peptide ◽  
Anti Inflammatory ◽  
Molecular Patterns

ABSTRACTA group of synthetic antimicrobial oligomers, inspired by naturally occurring antimicrobial peptides, were analyzed for the ability to modulate innate immune responses to Toll-like receptor (TLR) ligands. These synthetic mimics of antimicrobial peptides (SMAMPs) specifically reduced cytokine production in response toStaphylococcus aureusand theS. aureuscomponent lipoteichoic acid (LTA), a TLR2 agonist. Anti-inflammatory SMAMPs prevented the induction of tumor necrosis factor (TNF), interleukin 6 (IL-6), and IL-10 in response toS. aureusor LTA, but no other TLR2 ligands. We show that these SMAMPs bind specifically to LTAin vitroand prevent its interaction with TLR2. Importantly, the SMAMP greatly reduced the induction of TNF and IL-6in vivoin mice acutely infected withS. aureuswhile simultaneously reducing bacterial loads dramatically (4 log10). Thus, these SMAMPs can eliminate the damage induced by pathogen-associated molecular patterns (PAMPs) while simultaneously eliminating infectionin vivo. They are the first known SMAMPs to demonstrate anti-inflammatory and antibacterial activitiesin vivo.

scholarly journals Antibodies to Cryptic Epitopes in Distant Homologues Underpin a Mechanism of Heterologous Immunity between Plasmodium vivax PvDBP and Plasmodium falciparum VAR2CSA

mBio ◽  
2019 ◽  
Vol 10 (5) ◽  
Author(s):  
Catherine J. Mitran ◽  
Angie Mena ◽  
Sedami Gnidehou ◽  
Shanna Banman ◽  
Eliana Arango ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Plasmodium Vivax ◽  
Molecular Mechanism ◽  
Neutralizing Antibodies ◽  
Natural Infection ◽  
Placental Malaria ◽  
Content Type ◽  
Heterologous Immunity ◽  
Cryptic Epitopes

ABSTRACT Many pathogens evolve extensive genetic variation in virulence proteins as a strategy to evade host immunity. This poses a significant challenge for the host to develop broadly neutralizing antibodies. In Plasmodium falciparum, we show that a mechanism to circumvent this challenge is to elicit antibodies to cryptic epitopes that are not under immune pressure. We previously discovered that antibodies to the Plasmodium vivax invasion protein, PvDBP, cross-react with P. falciparum VAR2CSA, a distantly related virulence factor that mediates placental malaria. Here, we describe the molecular mechanism underlying this cross-species immunity. We identified an epitope in subdomain 1 (SD1) within the Duffy binding-like (DBL) domain of PvDBP that gives rise to cross-reactive antibodies to VAR2CSA and show that human antibodies affinity purified against a synthetic SD1 peptide block parasite adhesion to chondroitin sulfate A (CSA) in vitro. The epitope in SD1 is subdominant and highly conserved in PvDBP, and in turn, SD1 antibodies target cryptic epitopes in P. falciparum VAR2CSA. The epitopes in VAR2CSA recognized by vivax-derived SD1 antibodies (of human and mouse origin) are distinct from those recognized by VAR2CSA immune serum. We mapped two peptides in the DBL5ε domain of VAR2CSA that are recognized by SD1 antibodies. Both peptides map to regions outside the immunodominant sites, and antibodies to these peptides are not elicited following immunization with VAR2CSA or natural infection with P. falciparum in pregnancy, consistent with the cryptic nature of these target epitopes. IMPORTANCE In this work, we describe a molecular mechanism of heterologous immunity between two distant species of Plasmodium. Our results suggest a mechanism that subverts the classic parasite strategy of presenting highly polymorphic epitopes in surface antigens to evade immunity to that parasite. This alternative immune pathway can be exploited to protect pregnant women from falciparum placental malaria by designing vaccines to cryptic epitopes that elicit broadly inhibitory antibodies against variant parasite strains.

scholarly journals LACK OF IDENTITY IN NEUTRALIZING AND HEMAGGLUTINATION-INHIBITING ANTIBODIES AGAINST INFLUENZA VIRUSES

1950 ◽  
Vol 91 (1) ◽  
pp. 65-86 ◽  
Author(s):  
Duard L. Walker ◽  
Frank L. Horsfall
Keyword(s):  
Influenza Virus ◽  
Hemagglutination Inhibition ◽  
Neutralizing Antibodies ◽  
Neutralizing Antibody ◽  
Immune Serum ◽  
Influenza Viruses ◽  
In Ovo ◽  
Neutralization Line

There is an exponential linear relationship between the quantity of influenza virus neutralized and the quantity of immune serum employed in in ovo neutralization. The slope of the neutralization line is extremely steep. The concentration of neutralizing antibody can be measured with considerable precision in ovo if the constant virus-varying serum technique is utilized. The amounts of hemagglutination-inhibiting and neutralizing antibodies which are absorbed by a given quantity of influenza virus (PR8) were found to be predictable and the degree of reactivity of these two antibodies was shown to be directly related to the extent of immunization. It was demonstrated that there are marked discrepancies in correlation between antibody titers obtained by in vitro hemagglutination-inhibition and in vivo neutralization techniques and that neutralizing antibody is preferentially absorbed by a given quantity of virus. Inasmuch as the results were found not to be attributable to peculiarities of the techniques employed, it appears that the antibodies measured by hemagglutination-inhibition in vitro and by neutralization in vivo are not identical.

scholarly journals The Mu Subunit of Plasmodium falciparum Clathrin-Associated Adaptor Protein 2 ModulatesIn VitroParasite Response to Artemisinin and Quinine

2015 ◽  
Vol 59 (5) ◽  
pp. 2540-2547 ◽  
Author(s):  
Gisela Henriques ◽  
Donelly A. van Schalkwyk ◽  
Rebekah Burrow ◽  
David C. Warhurst ◽  
Eloise Thompson ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Adaptor Protein ◽  
Content Type ◽  
Artemisinin Derivatives ◽  
Combination Treatments ◽  
Antimalarial Resistance ◽  
Transgenic Parasites ◽  
Multiple Drugs

ABSTRACTThe emergence of drug-resistant parasites is a serious threat faced by malaria control programs. Understanding the genetic basis of resistance is critical to the success of treatment and intervention strategies. A novel locus associated with antimalarial resistance,ap2-mu(encoding the mu chain of the adaptor protein 2 [AP2] complex), was recently identified in studies on the rodent malaria parasitePlasmodium chabaudi(pcap2-mu). Furthermore, analysis in Kenyan malaria patients of polymorphisms in thePlasmodium falciparumap2-muhomologue,pfap2-mu, found evidence that differences in the amino acid encoded by codon 160 are associated with enhanced parasite survivalin vivofollowing combination treatments which included artemisinin derivatives. Here, we characterize the role ofpfap2-muin mediating thein vitroantimalarial drug response ofP. falciparumby generating transgenic parasites constitutively expressing codon 160 encoding either the wild-type Ser (Ser160) or the Asn mutant (160Asn) form ofpfap2-mu. Transgenic parasites carrying thepfap2-mu160Asn allele were significantly less sensitive to dihydroartemisinin using a standard 48-hin vitrotest, providing direct evidence of an altered parasite response to artemisinin. Our data also provide evidence thatpfap2-muvariants can modulate parasite sensitivity to quinine. No evidence was found thatpfap2-muvariants contribute to the slow-clearance phenotype exhibited byP. falciparumin Cambodian patients treated with artesunate monotherapy. These findings provide compelling evidence thatpfap2-mucan modulateP. falciparumresponses to multiple drugs. We propose that this gene should be evaluated further as a potential molecular marker of antimalarial resistance.

scholarly journals In VitroandIn VivoAntimalarial Activities of T-2307, a Novel Arylamidine

2012 ◽  
Vol 56 (4) ◽  
pp. 2191-2193 ◽  
Author(s):  
Akiko Kimura ◽  
Hiroshi Nishikawa ◽  
Nobuhiko Nomura ◽  
Junichi Mitsuyama ◽  
Shinya Fukumoto ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Body Weight ◽  
Broad Spectrum ◽  
Antimalarial Activity ◽  
Liver Stage ◽  
Rodent Malaria ◽  
Content Type ◽  
Clinically Significant

ABSTRACTT-2307, a novel arylamidine, has been shown to exhibit broad-spectrum antifungal activities against clinically significant pathogens. Here, we evaluated thein vitroandin vivoantimalarial activity of T-2307. The 50% inhibitory concentrations (IC50s) of T-2307 againstPlasmodium falciparumFCR-3 and K-1 strains were 0.47 and 0.17 μM, respectively. T-2307 at 2.5 to 10 mg/kg of body weight/day exhibited activity against blood stage and liver stage parasites in rodent malaria models. In conclusion, T-2307 exhibitedin vitroandin vivoantimalarial activity.

scholarly journals Fluorometric High-Throughput Assay for Measuring Chlamydial Neutralizing Antibody

2012 ◽  
Vol 19 (11) ◽  
pp. 1864-1869 ◽  
Author(s):  
Timothy Southern ◽  
Leah Bess ◽  
Jillian Harmon ◽  
Lacey Taylor ◽  
Harlan Caldwell
Keyword(s):  
High Throughput ◽  
Neutralizing Antibodies ◽  
Chlamydial Infection ◽  
Transmitted Disease ◽  
Neutralizing Antibody ◽  
Human Populations ◽  
High Throughput Analysis ◽  
Content Type ◽  
Fluorometric Analysis

ABSTRACTChlamydia trachomatisis an obligate intracellular mucosotropic pathogen that causes human infections of global importance.C. trachomatiscauses trachoma, the leading cause of preventable blindness worldwide, and is the most common cause of bacterial sexually transmitted disease. Although oculogenital infections are treatable with antibiotics, a vaccine is needed to controlC. trachomatisinfection. Ideally, a vaccine would provide coverage against most, if not all, naturally occurring antigenically distinct serovariants. The development of a subunit vaccine to prevent oculogenital disease could be advanced by identifying chlamydial antigens that elicit pan-neutralizing antibodies, particularly among infected human populations of known risk factors. There is currently no objective high-throughputin vitroassay to screen human sera for neutralization to aid in identification of these antigens. This report describes an objective, high-throughputin vitroassay that measuresC. trachomatis-neutralizing antibodies. Antibody-mediated neutralization of chlamydial infection was performed in a 96-well microtiter format, and neutralization was quantified by immunostaining fixed cells followed by automated fluorometric analysis. This report shows that fluorometric analysis ofC. trachomatisinfection directly correlates to labor-intensive manual inclusion counts. Furthermore, this report shows that fluorometry can be used to identifyC. trachomatis serovar- and serocomplex-specific neutralization. This objective, high-throughput analysis of serum neutralization is amenable to epidemiological studies of human chlamydial infection, human clinical vaccine trials, and preclinical animal model experiments ofChlamydiainfection.

scholarly journals Aminoindoles, a Novel Scaffold with Potent Activity against Plasmodium falciparum

2011 ◽  
Vol 55 (6) ◽  
pp. 2612-2622 ◽  
Author(s):  
Robert H. Barker ◽  
Sameer Urgaonkar ◽  
Ralph Mazitschek ◽  
Cassandra Celatka ◽  
Renato Skerlj ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Parent Compound ◽  
Therapeutic Index ◽  
Preclinical Development ◽  
Drug Pressure ◽  
Content Type ◽  
Novel Scaffold ◽  
Trough Levels

ABSTRACTThis study characterizes aminoindole molecules that are analogs of Genz-644442. Genz-644442 was identified as a hit in a screen of ∼70,000 compounds in the Broad Institute's small-molecule library and the ICCB-L compound collection at Harvard Medical School. Genz-644442 is a potent inhibitor ofPlasmodium falciparum in vitro(50% inhibitory concentrations [IC50s], 200 to 285 nM) and inhibitsP. berghei in vivowith an efficacy of >99% in an adapted version of Peters' 4-day suppressive test (W. Peters, Ann. Trop. Med. Parasitol. 69:155–171, 1975). Genz-644442 became the focus of medicinal chemistry optimization; 321 analogs were synthesized and were tested forin vitropotency againstP. falciparumand forin vitroabsorption, distribution, metabolism, and excretion (ADME) properties. This yielded compounds with IC50s of approximately 30 nM. The lead compound, Genz-668764, has been characterized in more detail. It is a single enantiomer with IC50s of 28 to 65 nM againstP. falciparum in vitro. In the 4-dayP. bergheimodel, when it was dosed at 100 mg/kg of body weight/day, no parasites were detected on day 4 postinfection. However, parasites recrudesced by day 9. Dosing at 200 mg/kg/day twice a day resulted in cures of 3/5 animals. The compound had comparable activity againstP. falciparumblood stages in a human-engrafted NOD-scidmouse model. Genz-668764 had a terminal half-life of 2.8 h and plasma trough levels of 41 ng/ml when it was dosed twice a day orally at 55 mg/kg/day. Seven-day rat safety studies showed a no-observable-adverse-effect level (NOAEL) at 200 mg/kg/day; the compound was not mutagenic in Ames tests, did not inhibit the hERG channel, and did not have potent activity against a broad panel of receptors and enzymes. Employing allometric scaling and usingin vitroADME data, the predicted human minimum efficacious dose of Genz-668764 in a 3-day once-daily dosing regimen was 421 mg/day/70 kg, which would maintain plasma trough levels above the IC90againstP. falciparumfor at least 96 h after the last dose. The predicted human therapeutic index was approximately 3, on the basis of the exposure in rats at the NOAEL. We were unable to select for parasites with >2-fold decreased sensitivity to the parent compound, Genz-644442, over 270 days ofin vitroculture under drug pressure. These characteristics make Genz-668764 a good candidate for preclinical development.

scholarly journals Effect of Preexisting Immunity on an Adenovirus Vaccine Vector: In Vitro Neutralization Assays Fail To Predict Inhibition by Antiviral Antibody In Vivo

2009 ◽  
Vol 83 (11) ◽  
pp. 5567-5573 ◽  
Author(s):  
Susan L. Pichla-Gollon ◽  
Shih-Wen Lin ◽  
Scott E. Hensley ◽  
Marcio O. Lasaro ◽  
Larissa Herkenhoff-Haut ◽  
...  
Keyword(s):  
Neutralizing Antibodies ◽  
Neutralizing Antibody ◽  
Human Adenovirus ◽  
T Cell Response ◽  
Fundamental Properties ◽  
High Prevalence ◽  
Antiviral Antibody ◽  
Adenovirus Vectors

ABSTRACT A major obstacle to the use of adenovirus vectors derived from common human serotypes, such as human adenovirus 5 (AdHu5), is the high prevalence of virus-neutralizing antibodies in the human population. We previously constructed a variant of chimpanzee adenovirus 68 (AdC68) that maintained the fundamental properties of the carrier but was serologically distinct from AdC68 and resisted neutralization by AdC68 antibodies. In the present study, we tested whether this modified vector, termed AdCDQ, could induce transgene product-specific CD8+ T cells in mice with preexisting neutralizing antibody to wild-type AdC68. Contrary to our expectation, the data show conclusively that antibodies that fail to neutralize the AdCDQ mutant vector in vitro nevertheless impair the vector's capacity to transduce cells and to stimulate a transgene product-specific CD8+ T-cell response in vivo. The results thus suggest that in vitro neutralization assays may not reliably predict the effects of virus-specific antibodies on adenovirus vectors in vivo.

scholarly journals Laboratory Detection of Artemisinin-Resistant Plasmodium falciparum

2014 ◽  
Vol 58 (6) ◽  
pp. 3157-3161 ◽  
Author(s):  
Kesinee Chotivanich ◽  
Rupam Tripura ◽  
Debashish Das ◽  
Poravuth Yi ◽  
Nicholas P. J. Day ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Parasite Clearance ◽  
Ring Stage ◽  
Clearance Time ◽  
Content Type ◽  
Reduced Ring ◽  
Susceptibility Tests ◽  
Low Sensitivity

ABSTRACTConventional 48-hin vitrosusceptibility tests have low sensitivity in identifying artemisinin-resistantPlasmodium falciparum, defined phenotypically by lowin vivoparasite clearance rates. We hypothesized originally that this discrepancy was explained by a loss of ring-stage susceptibility and so developed a simple field-adapted 24-h trophozoite maturation inhibition (TMI) assay focusing on the ring stage and compared it to the standard 48-h schizont maturation inhibition (WHO) test. In Pailin, western Cambodia, where artemisinin-resistantP. falciparumis prevalent, the TMI test mean (95% confidence interval) 50% inhibitory concentration (IC50) for artesunate was 6.8 (5.2 to 8.3) ng/ml compared with 1.5 (1.2 to 1.8) ng/ml for the standard 48-h WHO test (P= 0.001). TMI IC50s correlated significantly with thein vivoresponses to artesunate (parasite clearance time [r= 0.44,P= 0.001] and parasite clearance half-life [r= 0.46,P= 0.001]), whereas the standard 48-h test values did not. On continuous culture of two resistant isolates, the artemisinin-resistant phenotype was lost after 6 weeks (IC50s fell from 10 and 12 ng/ml to 2.7 and 3 ng/ml, respectively). Slow parasite clearance in falciparum malaria in western Cambodia results from reduced ring-stage susceptibility.

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