Rozprawy doktorskie na temat „Neutralizing antibodies”
Kliknij ten link, aby zobaczyć inne rodzaje publikacji na ten temat: Neutralizing antibodies.
Utwórz poprawne odniesienie w stylach APA, MLA, Chicago, Harvard i wielu innych
Sprawdź 50 najlepszych rozpraw doktorskich naukowych na temat „Neutralizing antibodies”.
Przycisk „Dodaj do bibliografii” jest dostępny obok każdej pracy w bibliografii. Użyj go – a my automatycznie utworzymy odniesienie bibliograficzne do wybranej pracy w stylu cytowania, którego potrzebujesz: APA, MLA, Harvard, Chicago, Vancouver itp.
Możesz również pobrać pełny tekst publikacji naukowej w formacie „.pdf” i przeczytać adnotację do pracy online, jeśli odpowiednie parametry są dostępne w metadanych.
Przeglądaj rozprawy doktorskie z różnych dziedzin i twórz odpowiednie bibliografie.
1
Ovcina, Emira. "An immune response to peptides selected with neutralizing antibodies". Thesis, National Library of Canada = Bibliothèque nationale du Canada, 2000. http://www.collectionscanada.ca/obj/s4/f2/dsk1/tape4/PQDD_0019/MQ59191.pdf.
Pełny tekst źródła
2
Bonsall, David George. "The role of neutralizing antibodies in HIV-1 infection". Thesis, Imperial College London, 2011. http://hdl.handle.net/10044/1/6360.
Pełny tekst źródłaStreszczenie:
Human immunodeficiency virus type 1 is a major cause of morbidity and mortality worldwide and there is urgent demand for a protective vaccine. A major goal of vaccine development is the elicitation of antibodies capable of neutralizing diverse strains. In order to achieve this goal it is necessary to understand the dynamic relationship between neutralizing antibodies (NAbs) and HIV-1, in vivo. In humans, HIV-1 rapidly escapes from NAbs, confirming that humoral responses inhibit virus replication. However, neutralizing responses are commonly detected in viraemic patients and the clinical impact of NAbs on HIV-1 control is unclear. To investigate this further, viral load (VL) and NAb activity were assessed longitudinally in patients enrolled into a clinical trial of short-course antiretroviral therapy (ART), administered in early infection. The aims of this study were two-fold: i) to understand the importance of VL in the control of NAb responses and ii) to assess whether NAbs contribute to durable control of VL set-point. A high-throughput pseudovirus neutralization assay was developed, using automated counting procedures to quantify infected TZM-bl reporter cells. The assay was used to assess NAb responses with autologous viruses derived from 22 patients. Seven patients with low VL set-points (<104 RNA copies/ml) failed to develop neutralizing responses throughout the 48-144 week follow-up period. In contrast, the remaining patients developed progressively-increasing neutralizing plasma titres (IC50) that correlated with the extent and timing of VL rebound after cessation of ART. This suggests that the production of NAbs depends on the duration and extent of viraemia in early infection. Viral load was poorly predictive of neutralizing responses against heterologous isolates assayed in 38 patients, suggesting that other factors are important in the production of antibodies with cross-neutralizing activity. Depletion of specific immunological compartments can yield crucial information as to their functional importance in vivo. We took advantage of a unique opportunity to investigate the role of NAbs and the consequences of their depletion in an HIV-1 infected human. Three years after cessation of short-course ART, the patient was treated for pre-existing low-grade lymphoplasmacytic lymphoma by antibodymediated depletion of CD20+ B cells using rituximab. This treatment was followed by a 1.7 log10 rise in HIV-1 VL which spontaneously reversed. Autologous NAb responses decreased as viraemia flared, and recovered as VL was controlled. Antibodies were found to target the CD4 binding site (CD4bs), as shown by competitive-binding assays. Sequence analysis revealed diversification through generation of new variants as NAbs decreased, with subsequent selection of NAb-resistant mutants at sites consistent with the binding data. These data suggest that B cell function contributed to long-term control of VL in this individual and that NAbs may be more important in controlling HIV-1 infection than previously suspected.
3
Zhang, Jianchao. "HIV-1/SIV neutralizing antibody gene delivery a novel vaccination approach /". Columbus, Ohio : Ohio State University, 2009. http://rave.ohiolink.edu/etdc/view?acc%5Fnum=osu1237924213.
Pełny tekst źródła
4
Lam, Sin Man. "Neutralizing antibodies and the biological response to interferon-beta therapy". Thesis, University of British Columbia, 2010. http://hdl.handle.net/2429/26977.
Pełny tekst źródłaStreszczenie:
Interferon-beta (IFN-beta) was the first disease modifying drug to be approved for the treatment of multiple sclerosis (MS). IFN-beta reduces relapse rates, relapse severity as well as slows the progression of disease burden. However, neutralizing antibodies (NAbs) occur in a proportion of patients receiving IFN-beta treatment. NAbs bind to IFN-beta, reduce drug bioavailability and high levels of NAbs reduce drug efficiency.
Our first objective was to develop and validate a sensitive and rapid method to measure NAbs. Existing methods to measure NAbs include the cytopathic effect assay (CPE) method and the myxovirus protein A (MxA) assay. However, these assays are time consuming and may be arduous to perform. We describe the optimization of a luciferase reporter gene assay to measure NAbs. To validate the assay, we assayed sera from IFN-beta treated MS patients with the optimized luciferase method and compared the results to those obtained with the reference CPE method. NAb status measured by the luciferase and the CPE method did not yield a significant difference. NAb titres obtained from the two methods correlated very well. The luciferase assay is reliable, appropriately sensitive and requires less time than the currently available NAb methods.
In addition to measuring NAbs, the biological activity of IFN-beta can be measured by monitoring IFN-inducible biomarkers, specifically MxA mRNA. Bioavailability measurements become especially valuable in patients with low to moderate NAb titres, “the grey zone”, and have been identified as a possible alternative for NAb measurements. Nonetheless, there is still controversy about how long should one wait after an IFN injection to draw blood for MxA mRNA measurements. Our objective was to identify the optimal time for blood draws to measure MxA mRNA. MxA mRNA was the most robust at 4-12 hours after IFN-beta injection and peaked at 8h post IFN injection. NAbs were evidently associated with an attenuation of IFN-beta bioactivity.
In conclusion, we characterized a technique to assess NAbs associated with IFN-beta therapy and a method to assess IFN-beta biological activity in treated patients. Altogether, this will help measure the effects of IFN-beta treatment and assist clinicians in tailoring therapy to the individual patient.
5
Burke, Karen L. "Antigen chimaeras of poliovirus". Thesis, University of Reading, 1989. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.328967.
Pełny tekst źródła
6
Wiley, James A. "Monoclonal anti-idiotypes induce neutralizing antibodies to enterovirus-70 conformational epitopes". Thesis, University of Ottawa (Canada), 1992. http://hdl.handle.net/10393/7649.
Pełny tekst źródłaStreszczenie:
The model pathogen used in the development of the anti-idiotypic antibodies produced in this project was enterovirus-70. This virus is the causative agent of acute hemorrhagic conjunctivitis. In the past twenty-five years, this virus has been responsible for two worldwide pandemics of acute hemorrhagic conjunctivitis. Monoclonal antibodies (MAbs), directed against the prototype enterovirus-70 strain, J670/71, were generated and characterized in order to produce a monoclonal anti-idiotypic antibodies (MAb2s) for use as surrogate immunogens. Radio-immunoprecipitation and western immunoblot assays suggested that all the monoclonal antibodies recognize conformational epitopes on the virion surface. A neutralizing monoclonal antibody, MAb/ev-12, was selected for the production of MAb2s. Five MAb2s were selected for their capacity to inhibit the interaction of MAb/ev-12 with EV-70 in dot immunobinding inhibition and immunofluorescence assays. These five MAb2s also inhibited virus neutralization mediated by MAb/ev-12 suggesting that each recognizes a paratope associated idiotope. In competition enzyme immunosorbent assays, none of the five MAb2s recognized other neutralizing and non-neutralizing enterovirus-70 specific MAbs, thus demonstrating that the MAb2s were specific for private idiotopes. Immunization with each of the MAb2s was carried out for the production of anti-anti-idiotypic antibodies (Ab3). All five MAb2s induced an immune response. Moreover, results suggested that they share idiotopes since MAb2:MAb/ev-12 binding could be inhibited by homologous as well as heterologous Ab3. Ab3 sera were shown to possess antibodies capable of immunoprecipitating $\sp{35}$S-labelled viral proteins in the same manner as MAb/ev-12. Nine of fifteen mice immunized with MAb2s demonstrated Ab3 neutralizing activity specific for the prototype EV-70 strain, J670/71. The potential application of MAb2s to serve as surrogate immunogens for conformational epitopes is substantiated by the results presented in this report.
7
Derby, Nina Rafterman. "Designing immunogens to elicit broadly reactive neutralizing antibodies to the HIV envelope /". Thesis, Connect to this title online; UW restricted, 2007. http://hdl.handle.net/1773/9302.
Pełny tekst źródła
8
Alpert, Michael. "Antibodies in Vaccine Protection against SIV and HIV-1 Infection". Thesis, Harvard University, 2011. http://dissertations.umi.com/gsas.harvard:10057.
Pełny tekst źródłaStreszczenie:
The properties of human immunodeficiency virus type 1 (HIV-1) and its simian counterpart SIV that enable persistent replication in the face of robust cellular, antibody, and innate immune responses have complicated efforts to develop a safe and effective vaccine. Vaccine protection against HIV-1 infection may require a combination of immune mechanisms. However, the types of immune responses that can be induced by vaccination to prevent HIV-1 infection remain unclear. The features of the viral envelope glycoprotein (Env) that confer inherent resistance to neutralization by antibodies also interfere with the development of antibody responses. We therefore vaccinated rhesus macaques with single-cycle SIV (scSIV) strains expressing Env proteins mutated to remove features that interfere with the induction of antibody responses. Antibodies capable of neutralizing Env-modified but not wild-type SIV were selectively enhanced. Identifying the immune responses underlying complete protection by live-attenuated SIV against pathogenic SIV challenge may provide guidance for HIV-1 vaccine design. To test the hypothesis that antibodies not measurable by assays for virus neutralization correlate with protection by live-attenuated SIV, we developed a novel assay for antibody-dependent cell-mediated cytotoxicity (ADCC). ADCC activity increased progressively over time after inoculation, and was measurable against viruses expressing heterologous Env proteins from independent SIV isolates when neutralization was undetectable. Two separate pathogenic \(SIV_{mac}251\) challenge experiments took advantage of either the strain specificity or the time-dependent development of immunity to overcome complete protection by live-attenuated SIV. In both experiments, macaques inoculated with live-attenuated SIV that remained uninfected by \(SIV_{mac}251\) had significantly higher ADCC activity than those that became infected. We also measured ADCC for the primary immune correlates analysis of a recent HIV-1 vaccine clinical trial in Thailand (RV144) that reported modest vaccine protection (31%). There was a nonsignificant trend towards lower risk of infection among vaccinees with high versus low relative ADCC activity. However, Env-specific IgA correlated with risk, prompting an analysis stratified by IgA levels. Among vaccinees with low Env-specific IgA, there was lower risk of infection among those with higher ADCC activity. These observations suggest that antibodies that direct ADCC may contribute to vaccine protection against SIV and HIV-1 infection.
9
Yari, Fayezeh. "Expression of recombinant neutralizing anti-HIV-1 antibodies in bacteria and eukaryotic cells /". Stockholm : Karolisnska institutet, 2007. http://diss.kib.ki.se/2007/978-91-7357-079-4/.
Pełny tekst źródła
10
Molinos, Albert Luis Manuel. "Targeting a major HIV-1 vulnerability region: the gp41 membrane proximal external region. Balance between neutralizing and non-neutralizing antibodies and implications for vaccine design". Doctoral thesis, Universitat Autònoma de Barcelona, 2016. http://hdl.handle.net/10803/392728.
Pełny tekst źródłaStreszczenie:
La generació d’anticossos àmpliament neutralitzants contra el VIH-1 es el principal objectiu d’una vacuna que sigui capaç de controlar l’epidèmia causada per aquest virus. La glicoproteïna de l’embolcall del VIH-1 es l’únic antigen viral exposat a la membrana del virus y la principal diana de respostes humorals protectores. Dins d’aquesta proteïna han sigut identificades regions funcionals reconegudes per anticossos àmpliament neutralitzants, fent possible la definició de punts de vulnerabilitat del virus, cap als que una vacuna hauria de dirigir-se. En aquest context, la regió externa pròxima a la membrana (MPER) de la gp41 es un dels llocs de vulnerabilitat més representatius del virus, ja que està altament conservat, té un paper crucial per a la infectivitat del virus i es reconegut per anticossos protectors àmpliament neutralitzants. Tot i així, la seva situació, prop a la membrana, fa que el MPER sigui un domini poc accessible, exposat de manera transitòria, hidrofòbic i altament influenciat pels lípids de la membrana. Per tant, la seva immunogenicitat presenta una alta complexitat que ha de ser explorada en major profunditat.
En aquesta tesi, aportem nou coneixement sobre l’immunogenicitat del MPER durant l´infecció natural i en models animals d’immunització. S’han avaluat miniproteïnes derivades de la gp41 que sobreexposen el MPER, per ser utilitzades com 1) nous plataformes per a la detecció d’anticossos anti-MPER i 2) prototipus d’immunògens presentats en proteoliposomes de diversa composició. Els resultats van revelar una alta immunogenicitat del MPER en humans, que correlaciona amb la resposta global contra la proteïna de l’embolcall, suggerint que el sistema immune no té cap restricció per a la generació d’aquest tipus d’anticossos. Tot i així, els anticossos detectats van mostrar una funcionalitat heterogènia, tant pel que fa a la capacitat neutralitzant com a la competició per diferents epítops, independentment de l’especificitat per MPER. A més, aquest resultats van ser reproduïts en animals immunitzats. En aquest últims, s’ha aconseguit generar un alt títol d’anticossos específics, que van ser potenciats per la incorporació als proteoliposomes de lípids complexes que mimetitzaven als de la partícula viral. Sorprenentment, es va generar una resposta immunodominant no neutralitzant contra un epítop solapant amb el de l’anticòs neutralitzant 2F5. En resum, la resposta humoral contra el MPER a humans i animals immunitzats no va correlacionar amb la capacitat neutralizant d’aquests i els anticossos generats són principalment no neutralitzants.
Aquests resultats suggereixen que el balanç de la resposta neutralitzant i no neutralitzant té una important rellevància en la resposta global contra el MPER. Conseqüentment, sembla necessari un major refinament d’immunògens capaços d’evitar respostes no neutralitzants. Aquest redisseny es veurà altament beneficiat pel coneixement generat a partir de nous anticossos monoclonals contra el MPER que recullin l’heterogeneïtat funcional observada als nostres estudis.La generación de anticuerpos ampliamente neutralizantes contra el VIH-1 es el principal objetivo de una vacuna que sea capaz de controlar la epidemia causada por el virus. La glicoproteína de la envuelta del VIH-1 es el único antígeno viral expuesto en la membrana del virus y la principal diana de respuestas humorales protectoras. Dentro de esta proteína se han identificado regiones funcionales que son reconocidas por anticuerpos ampliamente neutralizantes, lo cual ha permitido establecer puntos de vulnerabilidad del virus hacia los que una vacuna debería dirigirse. En este contexto, la región externa próxima a la membrana (MPER) de gp41 es uno de los sitios de vulnerabilidad del virus más representativos, ya que está altamente conservado, juega un papel crucial en la infectividad del virus y es reconocido por anticuerpos protectores ampliamente neutralizantes. Sin embargo, su localización, contigua a la membrana, hace del MPER un dominio poco accesible, expuesto de forma transitoria, hidrofóbico y altamente influenciado por lípidos de membrana. En consecuencia, su inmunogenicidad presenta una alta complejidad que debe ser explorada en mayor profundidad. En esta tesis, aportamos nuevo conocimiento sobre la inmunogenicidad del MPER a nivel de la infección natural y en modelos animales de inmunización. Hemos evaluado miniproteínas basadas en gp41 que sobreexponen el MPER, para ser utilizadas como 1) nuevas plataformas para la detección de anticuerpos anti-MPER y 2) prototipos de inmunógenos presentados en proteoliposomas de diversa composición. Los resultados han revelado una alta inmunogenicidad del MPER en humanos, que correlaciona con la respuesta global contra la proteína de la envuelta. Esto sugiere que el sistema inmune no tiene una especial restricción para la generación de este tipo de anticuerpos. Sin embargo, los anticuerpos detectados mostraron una funcionalidad heterogénea, en términos de capacidad neutralizante y competición por diferentes epítopos, independientemente de la especificidad por el MPER. Además, los resultados fueron reproducidos en animales inmunizados. En estos últimos, conseguimos generar un alto título de anticuerpos específicos, potenciados por la incorporación en los proteoliposomas de mezclas lipídicas complejas que mimetizaban las de la partícula viral. Sorprendentemente, se generó una respuesta inmunodominante no neutralizante que solapaba con un epítopo reconocido por el anticuerpo neutralizante 2F5. En resumen, la repuesta humoral natural contra el MPER en pacientes infectados por el VIH-1 y la generada en modelos animales mediante inmunización comparten ciertas características como son la especificidad y la escasa capacidad neutralizante. Estos resultados sugieren que el balance entre la respuesta neutralizante y no neutralizante podría tener una importante relevancia en la respuesta global contra el MPER. De este modo, es necesario un mayor refinamiento de inmunógenos que sean capaces de sortear respuestas no neutralizantes. Este rediseño se verá altamente beneficiado del conocimiento generado a partir de nuevos anticuerpos monoclonales contra el MPER que reflejen la heterogeneidad funcional observada en nuestros estudios.
The elicitation of broadly neutralizing antibodies (bNAbs) against HIV-1 constitutes the major goal of an effective vaccine able to control the epidemic caused by this virus. The HIV-1 envelope glycoprotein is the only viral antigen exposed on the surface of the virus and the main target of protective humoral responses. The identification within this protein of functional regions recognized by bNAbs has delineated an HIV-1 vulnerability map that has guided efforts for rational immunogen design. In this regard, the gp41 Membrane Proximal External Region (MPER) is a major HIV-1 vulnerability site because is highly conserved, plays a major role in viral infectivity and is targeted by protective bNAbs. However, its localization, next to the viral membrane, results in a hardly accessible, transiently exposed and hydrophobic domain, strongly influenced by membrane lipids. Therefore, the immunogenicity of the MPER offers a high level of complexity that needs to be further explored. In this thesis we provide new knowledge on the MPER immunogenicity in both natural HIV-1 infection and immunization in animal models. We generated gp41-based miniproteins which properly exposed the MPER region that have been used 1) as novel platforms for MPER antibody detection and 2) as immunogen candidates presented in different proteoliposome compositions. In humans, the results revealed a strong immunogenicity of the MPER that correlated with a global response against the envelope glycoprotein suggesting no special constraints for the immune system to target this region. However, the antibodies elicited showed heterogeneous functionality in terms of neutralizing capacity and epitope competition, regardless MPER specificity. Interestingly, these results were reproduced by immunization in animal models. High antibody titers were achieved that were specially enhanced by the addition of lipid mixtures mimicking the viral membrane. Interestingly, a non-neutralizing immunodominant response against an epitope that overlapped the 2F5 neutralizing antibody binding motif was identified. Overall, the anti-MPER response in both humans and animal model settings was not correlated with the neutralizing capacity and antibodies detected or induced by immunization were preferentially non-neutralizing. Our results suggest that the balance between neutralizing and non neutralizing responses may represent an important issue in the global response against MPER. Therefore, further redesign of immunogens able to skip non-neutralizing determinants will benefit from the knowledge derived from new anti-MPER antibodies that reflect the functional heterogeneous profile observed in our studies.
11
Fühner, Viola [Verfasser], Michael [Akademischer Betreuer] Hust i Stefan [Akademischer Betreuer] Dübel. "Development of neutralizing and non-neutralizing antibodies targeting known and novel epitopes on Clostridioides difficile Toxin B / Viola Fühner ; Michael Hust, Stefan Dübel". Braunschweig : Technische Universität Braunschweig, 2020. http://d-nb.info/1203299036/34.
Pełny tekst źródła
12
Penn-Nicholson, Adam Garth. "Characterization and evaluation of approaches to elicit Broadly Reactive Neutralizing Antibodies against HIV-1". Case Western Reserve University School of Graduate Studies / OhioLINK, 2008. http://rave.ohiolink.edu/etdc/view?acc_num=case1205433621.
Pełny tekst źródła
13
Ramey, Paul Christopher. "Population density and prevalence of rabies virus-neutralizing antibodies in a northern Ohio raccoon population". Connect to resource, 2005. http://rave.ohiolink.edu/etdc/view?acc%5Fnum=osu1173122630.
Pełny tekst źródła
14
Blay, Wendy Marie. "Human immunodeficiency virus type I (HIV-1) envelope evolution and the relationship to neutralizing antibodies /". Thesis, Connect to this title online; UW restricted, 2006. http://hdl.handle.net/1773/9296.
Pełny tekst źródła
15
Herrman, Marissa. "Neutralizing Antibodies to Epstein Barr Virus in the Rhesus Macaque Animal Model and in Humans". Thesis, Harvard University, 2015. http://nrs.harvard.edu/urn-3:HUL.InstRepos:23845409.
Pełny tekst źródłaStreszczenie:
Epstein-Barr virus (EBV) is associated with a number of human diseases and does not have a vaccine. It is believed that neutralizing antibodies are an important immune effector for an EBV vaccine, but it is unknown whether serum neutralizing antibodies can alter EBV infection through the oral mucosa. The studies presented in this dissertation were designed 1) to adapt the rhesus macaque animal model to allow testing of neutralizing antibodies in a biologically relevant system and 2) to better define the neutralizing antibody response of EBV infected humans.
Infection of rhesus macaques with the EBV related lymphocryptovirus, rhLCV, provides an accurate model system for studying EBV, but there were two hurdles that needed to be overcome before neutralizing antibodies could be tested in this model. First, there are no neutralizing antibodies specific to rhLCV and we found that a potent EBV neutralizing antibody, 72A1, did not cross-neutralize rhLCV. Second, murine monoclonal antibodies are inherently immunogenic in macaques and induce anti-antibody responses that limit their utility. To create a virus sensitive to 72A1 neutralization, the major membrane glycoprotein of rhLCV with replaced with EBV gp350. The data presented here show that this chimeric virus can use EBV gp350 to support entry into macaque cells in vitro and following oral inoculation of rhesus macaques. To reduce the immunogenicity of the murine antibodies, “rhesusized” antibody variants were generated and shown to retain antigen specificity. The combination of this novel, chimeric virus and the “rhesusized” antibodies will now allow testing of neutralizing antibodies in the macaque model.
Although multiple EBV glycoproteins have been shown to induce neutralizing antibodies in mice, studies of the human neutralizing antibody response have been narrowly focused on a single antibody binding epitope on gp350. Here we show that antibodies binding to this epitope do not represent all EBV neutralizing activity in human sera. Additionally, these data suggest that the neutralizing response is much broader than appreciated, with multiple glycoproteins inducing EBV neutralizing antibodies. Accurately defining the repertoire of viral glycoproteins targeted by human neutralizing antibodies can inform us of the naturally immunogenic proteins that may make good vaccine immunogens.Biology, Molecular and Cellular
16
Casa, Elisa. "Cell based assays used to quantify neutralizing antibodies against SARS-CoV-2 in human samples". Doctoral thesis, Università di Siena, 2022. http://hdl.handle.net/11365/1189903.
Pełny tekst źródłaStreszczenie:
Since the SARS-CoV-2 outbreak at the end of December 2019 in China, it has become
extremely important to have well‐established and validated diagnostic and research-use-only
assays for this new emerging virus. The microneutralization assay is a fundamentalserological
test in virology, immunology, vaccine assessment and epidemiological studies, and represents
one of the most used methods to evaluate the immune response induced by SARS-CoV-2
infection or vaccination.
In this Phd project different microneutralization methods are presented which can be used to
measure anti‐SARS‐CoV‐2 neutralizing antibodies (nAbs) in human serum samples. The aim of
the first project of this thesis (Project I) was to compare a microneutralization assay (MN) with
a read out based on the cytophatic effect (CPE) and a MN based on a colorimetric read out
for the detection of nAbs against SARS-CoV-2 Wild type strain. In the first method the cell
monolayers were microscopically inspected for inhibition of CPE at each serum dilution
(subjective method), while in the MN based on a colorimetric read out the healthy cell
monolayer was stained with neutral red solution, a vital dye. The plates were then read by a
spectrophotometer at 540 nm (objective method). A panel of 83 human serum samples were
previously tested in enzyme‐linked immunosorbent assay (ELISA) as a pre‐screening. All the
samples found to be positive, borderline, and negative in this ELISA were then tested to
determine the nAbs titers through the MN CPE and Colorimetric MN. The comparison
between log2-trasformed MN titers obtained through these two methods showed
comparable values, and the strong agreement in evaluating neutralizing antibodies against
SARS-CoV-2 Wild type strain was also confirmed by Correlation (r
2=0,9955), Bland-Altman,
and intra-class correlation (ICC) analysis (ICC value of 0.993, which is indicative of an excellent
agreement). This suggests the suitability of performing the MN assay using an ‘objective’
colorimetric-based read out method.
To better investigate if the classical MN CPE yielded similar results to those obtained with
other MN methods, we compared this “classical” MN to a new MN platform: the Virospot MN
assay (Project II). This method combines classic virus culture techniques with automated
sensitive detection of immunostained virus infected cells. In the Virospot MN, a virus-specific
immunostaining was used for plate reading and then the images of all wells were acquired by
a CTL ImmunoSpot analyzer. The 80% (MN80) or 90% (MN90) neutralization titers are
calculated according to the method described by Zielinska et al. 2005. This titer calculation is
based on the serum dilutions above and below the reduction point, 80% or 90%
neutralization. The MN CPE and Virospot methods were compared using a panel of 47 human
serum samples against SARS-CoV-2 Wild type and Alpha variant. The results of this project
showed that the these two different MN assays produce similar titer results against the Wild
Type virus, with good correlation values (correlation MN80 r2=0,9091; correlation MN90 r2=0,8900). A lower agreement between the MN CPE and Virospot MN assay was observed when SARS-CoV-2 Alpha variant was used (correlation MN80 r2=0,7226; correlation MN90 r2=0,6673).
Overall, these results showed a good agreement between the MN CPE assay and the two
different MN methods, Colorimetric MN and Virospot MN assay, in detecting neutralizing
antibodies against SARS-CoV-2 in human serum samples.
Despite the need for further standardization and/or the differences noticed during the
assessment of nAbs against SARS-COV-2 variants, the Colorimetric-based and Virospot MN
demonstrate to have advantages over the classical MN CPE, both being completely
automated methods, and hence offering a higher throughput, while inspection of each
dilution well by means of the optical microscope slows down the process. However, to ensure
that these correlation studies can provide meaningful results, further analysis with a bigger
number of samples and with other SARS-CoV-2 variants would be an added value. Moreover,
to make the data more comparable it would be necessary convert all the results to
international standard unit (IU/mL) allowing the accurate calibration of assays to an arbitrary
unit, thereby reducing inter-laboratory variation, and creating a common language for
reporting data.
The SARS-CoV-2 Virospot MN assay offers attractive advantages over the MN assay with a
read out based on the cytophatic effect, including the relative insensitivity to variation in
amount of infectious virus used in the test, independence from virus replication kinetics and
suitability for high throughput analyses.
Since many new SARS-CoV-2 variants occurred during the last two years, to make the Virospot
more sensitive and robust in detecting neutralizing antibodies against these new variants, the
third project (Project III) focused on the optimization study of this MN assay. Several new
conditions were adapted to optimize the method and make it more sensitive for the analysis
of samples against the Wild Type, Alpha, Beta, and Gamma SARS-CoV-2 variants.
Carboxymethyl cellulose (CMC) overlay was introduced to make the spot count more accurate
avoiding the viral spread after the first infection. Moreover, different sample matrices (serum
and plasma), culture media with and without the CO2 supplementation, and different
incubation time points and temperatures were assessed to evaluate and improve the assay
performance and robustness.
This optimization study has a planned follow-up, which can possibly include samples not only
from infected/convalescent individuals but also from vaccinated donors (with two or more
doses) or from people with hybrid immunity (such as breakthrough infections). Additionally,
further analyses with additional SARS-CoV-2 variants to strengthen these finding will also be
part of the next study. This project is worth to be conducted as the Virospot MN assay is likely
to have importance for the pre-clinical evaluation and eventual licensing of the SARS-CoV-2
vaccines.
17
Jackson, Nicholas A. C. "Properties and mechanism of action of a 17 amino acid, V3-loop-specific HIV-1 neutralizing miniantibody". Thesis, University of Warwick, 1998. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.340227.
Pełny tekst źródła
18
Belmonte, Antonietta. "The effects of antiviral therapy on the levels of neutralizing antibodies and antibodies mediating antibody-dependent cellular cytotoxicity in HI-1 seropositive patients /". Thesis, McGill University, 1992. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=56983.
Pełny tekst źródłaStreszczenie:
This study consists of evaluating the effects of zidovudine or ribavirin treatment on the humoral response to human immunodeficiency syndrome (HIV-1) in a cohort of 36 HIV-1 seropositive patients. Viral neutralization antibodies were demonstrated against HTLV$ sb{ rm IIIb}$ virus while titers of circulating antibody-dependent cellular cytotoxicity (ADCC) antibodies were measured against the HIV-1 envelope protein, gp120, using the vaccinia virus expression system which has been successfully used to express foreign viral proteins in target cells. Virologic (viral isolation) and immunologic (CD4$ sp+$ cells) parameters were also monitored pre and post antiviral therapy.The results indicate that patients receiving zidovudine for 36 weeks, have a diminished anti-HIV-1-ADCC directing antibody response, while the levels of these antibodies in patients receiving ribavirin or placebo remain constant. The titers of neutralizing antibodies and CD4 counts remain stable regardless of the treatment except for patients receiving ribavirin, where a decline in CD4 cells is observed. The decrease in the HIV-1 specific ADCC during zidovudine treatment parallels with the decrease in the amount of viral burden. This suggests that the two effects are somehow correlated. The impact of a washout period was also assessed. An increase in viral burden during cessation of AZT was reported which may reflect the inability of the treated host to mount a rapid immune response. As a consequence, this may lead to the deterioration of the immune status and the progression of the disease. The implications of these findings will be discussed in this study.
19
Reid, Phillip. "Functional Neutralizing Monoclonal Antibodies F-2-1 Against gp42 Ameliorates Disease Progression in Experimental Autoimmune Encephalomyelitis". Scholarship @ Claremont, 2018. http://scholarship.claremont.edu/cmc_theses/1822.
Pełny tekst źródłaStreszczenie:
Multiple Sclerosis (MS) is a chronic inflammatory demyelinating and neurodegenerative disease of the central nervous system (CNS), occurring in isolated attacks or progressive forms. Many observations implicate Epstein-Barr virus (EBV) in the pathogenesis of MS. With the relentless accumulation of evidence for a significant pathogenic role of EBV in MS, I believe it may be possible to prevent and cure MS by effectively controlling EBV infection. Currently, monoclonal antibodies (MAb) are used as therapeutics for a molecular targeted approach to slowing disease progression in MS. However, to my knowledge, there have been no antibodies targeted against EBV infection in any model of MS. The objective of this study is to determine whether or not a MAb against EBV could be a therapeutic target for EAE. In this study, I will propose an experiment that will examine the effects of intraperitoneal injection of MAb F-2-1 in 2-month-old new humanized BALB/c Rag2-/- ll2rg-/- (BRG) adult EBV/EAE male mice. My expected results suggest that mice with EBV/EAE + MAb F-2-1 may have an attenuated clinical disease course. Through immunohistochemical studies, I will also propose that MAb F-2-1 may decrease inflammation, demyelination and axonal loss in the CNS of mice with EAE. I believe that this novel treatments success would depend on MAb F-2-1’s ability to inhibit clonal expansion of EBV-infected autoreactive B cell in the CNS. Ultimately, my proposed experiment suggests that inhibition of virus-cell fusion of EBV to the B cell membrane might attenuate neuropathology in EAE. I hope that my prospective study highlights the importance of controlling EBV in patients with MS and provides grounds for optimism on how to successfully treat MS by controlling EBV infection. In conclusion, by proposing an alternative therapeutic approach, I hope that this hypothetical experiment will aid in future investigations that could further our knowledge on treatment and prevention of multiple sclerosis
20
Yuan, Tingting, i 袁婷婷. "Identification of intermediate antibodies of broadly neutralizing HIV-1 human monoclonal antibody b12 and characterization of variable loops of HIV-1 envelop glycoprotein". Thesis, The University of Hong Kong (Pokfulam, Hong Kong), 2013. http://hdl.handle.net/10722/196445.
Pełny tekst źródła
21
tw, spchen@mail atit org, i Shih-Ping Chen. "Epidemiology, pathogenesis and surveillance of the pig adapted strain of foot and mouth disease in Taiwan". Murdoch University, 2008. http://wwwlib.murdoch.edu.au/adt/browse/view/adt-MU20080813.104029.
Pełny tekst źródłaStreszczenie:
Foot-and-mouth disease (FMD) is one of the most contagious infectious diseases of domestic and wild cloven-hoofed animals, particular in cattle, sheep, pigs, goats and domestic buffalo, as well as wild ruminants such as deer. In Taiwan, there was a severe outbreak of FMD after more than 60 years freedom from the disease. The virus strain, O Taiwan 97 from the March 1997 outbreak of FMD in Taiwan, however, has been shown to have a species-specific adaptation to pigs. Although there are 7 distinct serotypes of FMD found in different regions of the world, this study focuses on the pig-adapted type O strain of FMD.
After the FMD outbreak commenced in Taiwan, the spread of disease was very rapid and the whole of the western parts of Taiwan was affected within a few days after the diagnosis of FMD was confirmed. In some situations airborne transmission of FMD virus was suspected and it was speculated that this was the explanation for such rapid spread in Taiwan. Therefore, studies were conducted to investigate the transmissibility of O Taiwan/97 FMDV to susceptible pigs by direct and indirect spread including airborne spread in an enclosed animal house. This study showed that pigs in direct contact with challenged pigs became infected but none of the close-contact pigs became infected. These experiments clearly demonstrated that the pig adapted strain O Taiwan/97 was only efficiently transmitted by direct contact. This indicates that effective control against future outbreaks of pig adapted FMDV strains could be achieved by restriction of pig movement and stamping out if the outbreak has been detected in the early stages and prior to the movements of pigs from the infected premises.
The measures used to control the Taiwanese FMD outbreak in 1997 were initially the slaughter of whole herds in the infected premises. However, with the rapid spread and large numbers of cases, the decision was taken to use universal compulsory vaccination of pig herds to control the outbreak when sufficient supply of vaccines was organized. Type O FMD vaccines were imported from a number of major FMD vaccine manufactures from around the world. Initially, vaccine efficacy for the imported vaccines was tested by measurement of neutralizing antibody titers in vaccinated pigs. To establish the relationship between serum neutralizing titers and protection from foot and mouth disease in pigs after vaccination, challenge studies were conducted with O/Taiwan/97 FMD in vaccinated pigs. Additionally, antibody responses to structural (neutralizing antibody) and non-structural proteins (NSP) were evaluated in vaccinated pig herds after primary and secondary vaccination in herds infected before and after vaccination.
In order to be able to monitor the circulation of virus in vaccinated pig populations, valid diagnostic kits based on the detection of antibody against NSP were required. These tests needed to be evaluated against pig sera derived from challenge studies and natural FMD outbreaks. Three commercially available ELISAs (Cedi, UBI and Checkit), which were available to differentiate infected from vaccinated pigs, were tested and results showed that the Cedi test had the optimal sensitivity and specificity for pig adapted type O FMD testing. This test was used to retrospectively evaluate the sera collected from infected and non-infected pig herds collected sequentially in the year after the 1997 FMD outbreak in Taiwan. These studies also showed that the early vaccines used, stimulated NSP antibody production in swine herds that were vaccinated but not infected. This resulted in the requirement for purified FMD vaccines to be used when monitoring programs for FMD infection by NSP testing were in place. In these studies, it was also demonstrated that the purified FMD vaccines used later in the control program did not induce NSP antibody after multiple double dosage to pigs.
Although clinical FMD appeared to be successfully controlled with vaccination program in Taiwan it was essential for the eradication plan to maintain active surveillance for NSP reactors in the pig population. The UBI and Cedi NSP kits were applied as screening and confirmatory tests, respectively, to pig sera collected in auction markets distributed around Taiwan to monitor for evidence of the circulation of FMD virus. Herds with positive reactors were followed-up by clinical inspection and 15 sera from suspected herds were further sampled. Negative results were obtained from all these investigation. With the absence of clinical outbreaks and the lack of evidence of FMDV circulation in the field from the NSP reactor surveillance, the Taiwanese government has progressed the eradication plan to a progressive cessation of vaccination, commencing with banning of vaccination on one isolated island in December 2006. The absence of outbreaks on that island, paved the way for further cessation of FMD vaccination in Taiwan from July 2008.
22
Gardner, Matthew Ryan. "Targeting the CD4- and Coreceptor-Binding Sites of the HIV-1 Envelope Glycoprotein". Thesis, Harvard University, 2014. http://dissertations.umi.com/gsas.harvard:11644.
Pełny tekst źródłaStreszczenie:
The HIV-1 envelope glycoprotein, Env, facilitates the translocation of the viral capsid across the cellular membrane. Env is a trimer of hetero-dimers composed of a gp120 subunit and gp41 transmembrane protein. The gp120 subunit binds the primary receptor, CD4, leading to conformational changes of Env that then promote binding to the coreceptor, principally CCR5 or CXCR4. As the sole protein on the surface of the virion, Env is under continuous pressure from the host's antibody response. Two classes of antibodies target the highly conserved receptor-binding sites of gp120: CD4-binding site (CD4bs) and CD4-induced (CD4i) antibodies.
23
Suphaphiphat, Karunasinee. "Anti-viral immune response in the semen of cynomolgus macaques and inhibition of cell to cell transmission by broadly neutralizing antibodies in an SIV/SHIV model of infection SHIV162P3 transmission by semen 1 leukocytes is efficiently 2 inhibited by broadly neutralizing antibodies". Thesis, Université Paris-Saclay (ComUE), 2019. http://www.theses.fr/2019SACLS599.
Pełny tekst źródłaStreszczenie:
La transmission sexuelle du VIH-1 se fait principalement via du sperme infecté contaminé, contenant à la fois des virions libres et des leucocytes infectés. Les facteurs présents dans le liquide séminal peuvent aussi moduler l’infectivité du sperme et la réponse immunitaire de l’hôte. Ainsi, le macaque infecté par le virus de l’immunodéficience simienne (VIS) sera utilisé comme modèle expérimental afin d’étudier l’infectivité des cellules séminales, les réponses immunitaires anti-virales et évaluer l’effet inhibiteur d’anticorps neutralisant à large spectre (bNAbs) sur la transmission du virus intercellulaire (TVI).Chez les macaques infectés avec le SIVmac251, les réponses immunitaires innées et adaptatives spécifiques du VIS ont été étudiées dans le sperme, en se focalisant sur les réponses T CD8, humorale et sur l’expression des cytokines, chimiokines et facteurs de croissance. L’infection VIS induit l’expression de cytokines pro-inflammatoires et immuno-régulatrices dans le sperme concordant avec une augmentation des cellules activées T CD8+ CD69+ et T CD8+ CXCR3+ CCR5+. Ni les cellules T CD8+ spécifiques du VIS, ni la réponse humorale ne permettent le contrôle de la dissémination du virus dans le sperme. L’absence de contrôle de la réplication virale dans le sperme infecté VIS est associée à une inflammation générale et à une activation immunitaire, pouvant refléter ce qui se produit dans le tractus reproducteur masculin, et qui pourrait mener à l’augmentation de la transmission VIH-1/VIS.De plus, nous avons développé un modèle d’étude in vitro de TVI en utilisant soit la lignée cellulaire TZM-bl, soit des PBMC humains comme cellules cibles, et des splénocytes ou des leucocytes CD45+ du spermes infectés avec le SHIV163P3 comme cellules infectieuses. Ce modèle nous a permis d’évaluer l’inhibition de TVI par des bNAbs. Nous avons testé 4 bNabs de 1ère génération et 8 bNabs de 2nde génération. La combinaison de bNabs de 1ère génération (2F5 + 2G12 + 4E10) permet d’inhiber TVI, alors qu’en combinaison double ou seule, aucune inhibition de la transmission n’est observée. Cependant, les bNabs de 2nde génération peuvent, seules, induire une inhibition de TVI aussi efficacement qu’en combinaison. Ainsi, un bNabs de 2nde génération anti-V3 a été sélectionné afin de tester son effet inhibiteur de TVI dans un modèle in vivoHIV-1 sexual transmission occurs mostly through contaminated semen, which contains both free virions and infected leukocytes. Moreover, factors in seminal plasma (SP) can influence both semen infectivity and host’s response. Therefore, we used the experimental model of Simian Immunodeficiency Virus (SIV) infection of macaques, to investigate semen cells infectivity and the antiviral immune responses and to evaluate the potency of broadly neutralizing antibodies (bNAbs) to block cell-to-cell virus transmission.In SIVmac251 infected cynomolgus macaques, we investigated SIV-specific innate and adaptive responses in semen, including CD8+ T cell response, humoral response and levels of cytokines, chemokines and growth factors. SIV infection induced pro-inflammatory and immunoregulatory cytokines in semen and a concomitant upregulation of activated CD69+ CD8+ T cells and CCR5+ CXCR3+ CD8+ T cells. Neither SIV-specific CD8+ T-cell responses nor humoral responses controlled seminal viral shedding. Failure to control viral replication in SIV-infected semen is related to a general inflammation and immune activation, which possibly mirrors what happen in the male genital tract and which could lead to enhanced HIV/SIV transmission.Moreover, we developed cell-to-cell transmission assays, using either TZM-bl or human PBMC as target cells and SHIV162P3-infected splenocytes and CD45+ semen leukocytes as donor cells, and evaluated bNAbs-mediated inhibition. The bNAb panel included four 1st generation bNAbs and eight 2nd generation bNAbs. A combination of 1st generation bNAbs (2F5+2G12+4E10) was able to efficiently inhibit CAV transmission, while double combination or single bNAbs showed reduced potency. Of note, individual 2nd generation bNAbs inhibited transmission as efficiently as bNAbs combinations. An anti-V3 bNAb has been selected to evaluate its potential to block cell-to-cell transmission in vivo
24
Wenzel, Esther Veronika Verfasser], Michael [Akademischer Betreuer] [Hust i Stefan [Akademischer Betreuer] Dübel. "Development of recombinant human anti-diphtheria toxin neutralizing antibodies for diphtheria therapy / Esther Veronika Wenzel ; Michael Hust, Stefan Dübel". Braunschweig : Technische Universität Braunschweig, 2019. http://nbn-resolving.de/urn:nbn:de:gbv:084-2019050215003.
Pełny tekst źródła
25
Corti, Davide. "Analysis of the human B cell memory repertoire against infectious pathogens and isolation of broadly neutralizing human monoclonal antibodies /". Bern : [s.n.], 2008. http://www.ub.unibe.ch/content/bibliotheken_sammlungen/sondersammlungen/dissen_bestellformular/index_ger.html.
Pełny tekst źródła
26
Ianni, Elvira <1981>. "Use of E. coli OMVs as delivery system to elicit neutralizing antibodies against Chlamydia infectivity: the HtrA protein example". Doctoral thesis, Alma Mater Studiorum - Università di Bologna, 2011. http://amsdottorato.unibo.it/3542/1/Ianni_Elvira_tesi.pdf.
Pełny tekst źródłaStreszczenie:
The obligate intracellular pathogen Chlamydia trachomatis is a gram negative bacterium which infects epithelial cells of the reproductive tract.
C. trachomatis is the leading cause of bacterial sexually transmitted disease worldwide and a vaccine against this pathogen is highly needed. Many evidences suggest that both antigen specific-Th1 cells and antibodies may be important to provide protection against Chlamydia infection. In a previous study we have identified eight new Chlamydia antigens inducing CD4-Th1 and/or antibody responses that, when combined properly, can protect mice from Chlamydia infection. However, all selected recombinant antigens, upon immunization in mice, elicited antibodies not able to neutralize Chlamydia infectivity in vitro.
With the aim to improve the quality of the immune response by inducing effective neutralizing antibodies, we used a novel delivery system based on the unique capacity of E. coli Outer Membrane Vesicles (OMV) to present membrane proteins in their natural composition and conformation. We have expressed Chlamydia antigens, previously identified as vaccine candidates, in the OMV system. Among all OMV preparations, the one expressing HtrA Chlamydia antigen (OMV-HtrA), showed to be the best in terms of yield and quantity of expressed protein, was used to produce mice immune sera to be tested in neutralization assay in vitro.
We observed that OMV-HtrA elicited specific antibodies able to neutralize efficiently Chlamydia infection in vitro, indicating that the presentation of the antigens in their natural conformation is crucial to induce an effective immune response. This is one of the first examples in which antibodies directed against a new Chlamydia antigen, other than MOMP (the only so far known antigen inducing neutralizing antibodies), are able to block the Chlamydia infectivity in vitro.
Finally, by performing an epitope mapping study, we investigated the specificity of the antibody response induced by the recombinant HtrA and by OMV-HtrA. In particular, we identified some linear epitopes exclusively recognized by antibodies raised with the OMV-HtrA system, detecting in this manner the antigen regions likely responsible of the neutralizing effect.
27
Ianni, Elvira <1981>. "Use of E. coli OMVs as delivery system to elicit neutralizing antibodies against Chlamydia infectivity: the HtrA protein example". Doctoral thesis, Alma Mater Studiorum - Università di Bologna, 2011. http://amsdottorato.unibo.it/3542/.
Pełny tekst źródłaStreszczenie:
The obligate intracellular pathogen Chlamydia trachomatis is a gram negative bacterium which infects epithelial cells of the reproductive tract.
C. trachomatis is the leading cause of bacterial sexually transmitted disease worldwide and a vaccine against this pathogen is highly needed. Many evidences suggest that both antigen specific-Th1 cells and antibodies may be important to provide protection against Chlamydia infection. In a previous study we have identified eight new Chlamydia antigens inducing CD4-Th1 and/or antibody responses that, when combined properly, can protect mice from Chlamydia infection. However, all selected recombinant antigens, upon immunization in mice, elicited antibodies not able to neutralize Chlamydia infectivity in vitro.
With the aim to improve the quality of the immune response by inducing effective neutralizing antibodies, we used a novel delivery system based on the unique capacity of E. coli Outer Membrane Vesicles (OMV) to present membrane proteins in their natural composition and conformation. We have expressed Chlamydia antigens, previously identified as vaccine candidates, in the OMV system. Among all OMV preparations, the one expressing HtrA Chlamydia antigen (OMV-HtrA), showed to be the best in terms of yield and quantity of expressed protein, was used to produce mice immune sera to be tested in neutralization assay in vitro.
We observed that OMV-HtrA elicited specific antibodies able to neutralize efficiently Chlamydia infection in vitro, indicating that the presentation of the antigens in their natural conformation is crucial to induce an effective immune response. This is one of the first examples in which antibodies directed against a new Chlamydia antigen, other than MOMP (the only so far known antigen inducing neutralizing antibodies), are able to block the Chlamydia infectivity in vitro.
Finally, by performing an epitope mapping study, we investigated the specificity of the antibody response induced by the recombinant HtrA and by OMV-HtrA. In particular, we identified some linear epitopes exclusively recognized by antibodies raised with the OMV-HtrA system, detecting in this manner the antigen regions likely responsible of the neutralizing effect.
28
Rojas, Expósito Luis Alfonso. "Blood barriers for oncolytic adenovirus efficacy: study of binding to erythrocytes via CAR and albumin‐mediated evasion of neutralizing antibodies". Doctoral thesis, Universitat de Barcelona, 2017. http://hdl.handle.net/10803/404054.
Pełny tekst źródłaStreszczenie:
Cancer virotherapy with oncolytic adenoviruses represents a promising therapeutic approach due to the capacity of these viruses to infect and selectively kill tumor cells without damaging normal tissues. Although the intravenous is the preferred route of administration in order to reach disseminated metastasis, several interactions with blood components cause the neutralization of the virus. Thus, improving the delivery of such adenoviruses to tumors by systemic injection is crucial for the success of the therapy. In this work we have studied the interaction of the adenovirus serotype 5 (Ad5) with human erythrocytes through the receptor CAR, which was described to sequester and inactivate the virus. Although erythrocyte binding was observed, it did not reduce viral transduction of tumor cells in vitro. Since mouse erythrocytes do not express CAR, human erythrocytes were transferred into nude mice to analyze the impact of erythrocyte binding after systemic administration. However, adenovirus extravasation and transduction of liver and tumors was not reduced, suggesting that this binding is reversible and does not neutralize the virus.
On the other hand, the high prevalence of anti-Ad5 neutralizing antibodies (NAbs) is a major obstacle for the intravenous administration of adenoviruses. To protect adenovirus against NAbs we inserted an albumin-binding domain (ABD) in the main adenovirus capsid protein, the hexon. This domain binds serum albumin to shield the virus upon systemic administration. The ABD-modified adenoviruses bind human and mouse albumin, which allow them to maintain the infectivity and replication capacity in presence of NAbs. Non-modified adenoviruses are completely neutralized after systemic administration in pre-immune mice, whereas ABD-modified viruses preserve the ability to transduce target organs and induce oncolysis. The data presented in this thesis supports the use of this strategy to treat patients systemically with oncolytic adenoviruses.
In summary, this thesis focused on improving the intravenous delivery of oncolytic adenoviruses, which is one of the main limitations of this therapy. The results presented in this work demonstrate that while erythrocyte binding via CAR does not inactivate the virus, NAbs represent a major obstacle for efficacy. In this regard, albumin coating of the virus capsid represents an effective approach to evade pre-existing NAbs. This strategy has translational relevance in the use of adenovirus by systemic injection not only for cancer virotherapy, but also for gene therapy and vaccination.Els adenovirus oncolítics són agents terapèutics prometedors, degut a la seva capacitat d’infectar i eliminar selectivament les cèl·lules tumorals, sense afectar les cèl·lules normals. Tot i que la ruta preferida d’administració és la intravenosa per tal d’arribar a totes les metàstasis, la interacció del virus amb diversos components de la sang provoca la seva neutralització. Per tant, millorar l’arribada dels virus als tumors per via sistèmica és un aspecte clau per a l’èxit d’aquesta teràpia. En aquest treball s’ha estudiat la interacció de l’adenovirus serotip 5 amb els eritròcits humans a través del receptor CAR, la qual es va descriure que provocava el segrest i la inactivació del virus. Malgrat es va observar unió als eritròcits, aquesta no va reduir la transducció de cèl·lules tumorals in vitro. Degut a que els eritròcits murins no expressen CAR, es van transferir eritròcits humans a ratolins immunodeprimits per tal d’analitzar l’efecte de la interacció després de la injecció sistèmica. Tot i així, aquesta unió als eritròcits no va alterar la extravasació ni la transducció del fetge per part del virus, suggerint que la interacció és reversible i no neutralitzant. Per altra banda, l’alta prevalença d’anticossos neutralitzants contra l’adenovirus 5 en la població humana representa un obstacle molt important per la injecció intravenosa d’aquest. Per protegir l’adenovirus contra els anticossos neutralitzants s’ha inserit un domini d’unió a albúmina (ABD) a la proteïna principal de la càpside viral, la proteïna hexó. Aquest domini s’uneix a l’albúmina sèrica, recobrint el virus amb aquesta després de l’administració sistèmica. Els virus modificats amb ABD són capaços d’unir-se tant a l’albúmina humana com a la murina, fet que els permet mantenir la infectivitat i la capacitat replicativa en presència d’anticossos neutralitzants. Els adenovirus no modificats són completament neutralitzats després de la administració sistèmica en ratolins pre-immunes, mentre que els virus modificats amb ABD mantenen la capacitat de transduïr els òrgans i controlar el creixement tumoral. Els resultats presentats en aquesta tesi recolzen l’ús d’aquesta estratègia per a tractar pacients amb adenovirus oncolítics per via sistèmica.
29
Kaiser, Fabian Marc Philipp [Verfasser], i Axel [Akademischer Betreuer] Rethwilm. "Analysis of Cross-Clade Neutralizing Antibodies against HIV-1 Env Induced by Immunofocusing / Fabian Marc Philipp Kaiser. Betreuer: Axel Rethwilm". Würzburg : Universitätsbibliothek der Universität Würzburg, 2013. http://d-nb.info/1030475695/34.
Pełny tekst źródła
30
Davis, Katie L. "Analysis of HIV-1 variable loop 3-specific neutralizing antibody responses by HIV-2/HIV-1 envelope chimeras". Thesis, Birmingham, Ala. : University of Alabama at Birmingham, 2008. https://www.mhsl.uab.edu/dt/2009r/davis.pdf.
Pełny tekst źródła
31
Wiegers, Anna-Katharina [Verfasser], Michael [Akademischer Betreuer] Mach i Thomas H. [Akademischer Betreuer] Winkler. "Neutralizing Antibodies to Antigenic Domain 5 of Glycoprotein B of Human Cytomegalovirus / Anna-Katharina Wiegers. Gutachter: Michael Mach ; Thomas H. Winkler". Erlangen : Friedrich-Alexander-Universität Erlangen-Nürnberg (FAU), 2014. http://d-nb.info/1065270526/34.
Pełny tekst źródła
32
Hashem, Anwar. "Targeting the Highly Conserved Sequences in Influenza A Virus". Thèse, Université d'Ottawa / University of Ottawa, 2013. http://hdl.handle.net/10393/24058.
Pełny tekst źródłaStreszczenie:
All challenges associated with influenza A viruses including antigenic variation in hemagglutinin (HA) and neuraminidase (NA), the evolving drug resistance and the drawbacks of current vaccines hinder our ability to control this constant threat. Furthermore, gene reassortment as well as the direct transmission of highly pathogenic avian viruses to humans can result in an occasional emergence of novel influenza strains with devastating pandemic potential. Therefore, it is crucial to investigate alternative approaches to better control these viruses and to develop new prophylactic and treatment options.
Targeting highly conserved epitopes or antigens among the different subtypes of influenza A virus could offer protection against broad range of influenza viruses, including emerging strains. In my research, I have investigated the potential of broadly neutralizing antibodies against HA and conducted mechanistic study of a prototype vaccine based on the highly conserved nucleoprotein (NP).
We recently found that the 14 amino acids of the amino-terminus of the fusion peptide of influenza HA2 subunit is the only universally conserved sequence in all HA subtypes of influenza A and the two lineages of influenza B viruses. Here, I show that universal antibodies targeting this linear sequence in the viral HA (Uni-1 antibodies) can cross-neutralize multiple subtypes of influenza A virus by inhibiting the pH-dependant fusion of viral and cellular membranes.
It is noted that the influenza NP is a highly conserved antigen and has the potential to induce heterosubtypic immunity against divergent subtypes of influenza A virus. However, NP-based vaccination only affords weak protective immunity compared to HA. This is mostly due to the non-sterilizing immunity induced by NP. Using CD40 ligand (CD40L), a key regulator of the immune system, as both a targeting ligand and a molecular adjuvant, I show that single immunization with recombinant adenovirus carrying a fused gene encoding the secreted NP-CD40L fusion protein provided robust and long-lasting protection against influenza in normal mice. It enhanced both B-cell and T-cell responses and augmented the role of both NP-specific antibodies and CTLs in protection. Importantly, it afforded effective protection in CD40L and CD4 deficient mice, confirming that the induced protection is CD40L-mediated and CD4+ T cell-independent.
The rapid evolution of the influenza A viruses necessitates the development of new alternatives to contain this medically important pathogen. The results of these studies could significantly contribute to future vaccine development and avert the necessity of yearly vaccine updates.
33
Olsen, Ole Andrew. "The selected lymphocyte antibody method : a novel approach for the generation and genetic analysis of human cytomegalovirus (HCMV) neutralizing monoclonal IgG antibodies". Thesis, National Library of Canada = Bibliothèque nationale du Canada, 1999. http://www.collectionscanada.ca/obj/s4/f2/dsk1/tape8/PQDD_0019/NQ46403.pdf.
Pełny tekst źródła
34
Kim, Dhohyung. "Mechanism of Maternal Antibody Inhibition and Vaccination Strategies in the presence of Maternal Antibodies". The Ohio State University, 2012. http://rave.ohiolink.edu/etdc/view?acc_num=osu1330968863.
Pełny tekst źródła
35
Muharemagic, Darija. "Aptamers as Enhancers of Oncolytic Virus Therapy". Thesis, Université d'Ottawa / University of Ottawa, 2015. http://hdl.handle.net/10393/32170.
Pełny tekst źródłaStreszczenie:
Oncolytic viruses promise to significantly improve current cancer treatments through their tumour-selective replication and multimodal attack against cancer cells. However, one of the biggest setbacks for oncolytic virus therapies is the intravenous delivery of the virus, as it can be cleared by neutralizing antibodies (nAbs) from the bloodstream before it reaches the tumour cells. In our group, we have succeeded in developing aptamers to vesicular stomatitis virus (VSV), as well as to rabbit anti-VSV polyclonal neutralizing antibodies (nAbs). We tested these aptamers’ biological activity with a cell-based plaque forming assay and found that the aptamers prevented in vitro neutralization of VSV by nAbs and increased the virus infection rate of transformed cells up to 77%.
In line with this approach, we enhanced the delivery of oncolytic viruses by selecting aptamers to the CT26 colon carcinoma cell line. The binding of aptamer pools has been tested on flow cytometry and the best pools were subjected to high throughput sequencing. Selected aptamers were linked to anti-VSV aptamers and applied for target delivery of the virus to cancer cells. Development of this aptamer-based technology aims to improve viral anti-cancer therapies, with a potential to be applied as treatment for patients affected with cancer.
Finally, in collaboration with a group from Erlangen University, we performed an aptamer selection using capillary electrophoresis and cell-SELEX. The target, the extracellular domain of human CD83, is a maturation marker for dendritic cells and is involved in the regulation of the immune system. Selected aptamer sequences bound selectively to mature dendritic cells, in comparison to immature dendritic cells, and thus hold promise to be applied for further studies leading to a better understanding of CD83’s mechanism of action.
36
Otterstrom, Jason John. "Visualizing Influenza Virus Membrane Fusion: Inhibition and Kinetics". Thesis, Harvard University, 2013. http://dissertations.umi.com/gsas.harvard:11251.
Pełny tekst źródłaStreszczenie:
The influenza virus hemagglutinin (HA) surface protein is a primary antigenic target for neutralization of viral infection. HA also mediates membrane fusion between the virus and a cell, which is the first critical step during infection. Traditional techniques to study infection neutralization by antibodies or the membrane fusion process rely on ensemble measurements, confounding the precise mechanism of infection neutralization and obscuring transient conformational intermediates. This dissertation describes advances made in a fluorescence microscopy-based single-particle fusion assay to overcome the limitations of ensemble measurements in these types of studies. Virus particles are labeled to visualize lipid mixing between a virus and a target membrane formed upon a glass or polymer support. Optionally, the viral lumen can be labeled to visualize the subsequent release of viral contents.
37
Cheng, Yuxing. "Elicitation of antibody responses against the HIV-1 gp41 Membrane Proximal External Region (MPER)". Thesis, Harvard University, 2014. http://dissertations.umi.com/gsas.harvard:11427.
Pełny tekst źródłaStreszczenie:
An effective vaccine to protect against HIV-1/AIDS remains elusive due to the extensive mechanisms employed by the HIV-1 virus to evade immune attack. Highly potent broadly neutralizing antibodies isolated from chronically infected individuals, however, show that such relevant antibodies can be naturally produced, implying that their elicitation through vaccination is a realistic possibility. These broadly neutralizing antibodies target different regions on the trimeric spikes formed by three protomers of the envelope (Env) protein. Each Env protein is comprised of the gp120 surface subunit in non-covalent association with the gp41 transmembrane subunit. Four regions have been identified: the CD4 binding site, the V1/V2 segment and the V3/glycan area all on the gp120 subunit as well as the MPER segment on the gp41 subunit. This dissertation focuses on the gp41 MPER segment given its highly conserved amino acid sequence among all HIV-1 clades and viral strain isolates and essential function in Env-mediated fusion and HIV entry. Of note, the MPER segment contains several adjacent epitopes targeted by broadly neutralizing antibodies, suggesting that the immune system is capable of producing neutralizing antibodies against this specific region. Analysis of both clade B and C MPER segments shows them to be L-shaped, consisting of two helices separated by a hinge. We have found that the hinge region of the MPER segment provides the conformational flexibility necessary for the Env-mediated hemifusion and fusion processes. A significant reduction in virus infectivity is observed when the hinge region is disrupted by introduction of two amino acid mutations that eliminate -helical capping residues and the tandem hinge joints. The importance of the hinge region of the MPER segment is further supported by the action of four MPER-specific neutralizing antibodies 2F5, 4E10, 10E8 and Z13E1. These neutralizing antibodies block virus infection by disrupting MPER hinge-related function.
38
Mehta, Dhwani. "Development and Evaluation of an Antibody-Dependent Cellular Cytotoxicity (ADCC) Assay for Influenza A Virus". University of Cincinnati / OhioLINK, 2020. http://rave.ohiolink.edu/etdc/view?acc_num=ucin1613742682998356.
Pełny tekst źródła
39
Miyazaki, Yasuyuki. "Analyses of sensitivity to antibody-mediated neutralization and induction of neutralizing antibodies to HIV-1 Env by simian/human immunodeficiency virus-macaque model". Kyoto University, 2002. http://hdl.handle.net/2433/149868.
Pełny tekst źródłaStreszczenie:
Kyoto University (京都大学)0048
新制・課程博士
博士(人間・環境学)
甲第9660号
人博第144号
13||129(吉田南総合図書館)
新制||人||35(附属図書館)
UT51-2002-G418
京都大学大学院人間・環境学研究科人間・環境学専攻
(主査)教授 速水 正憲, 教授 津田 謹輔, 教授 松村 道一, 助教授 三浦 智行
学位規則第4条第1項該当
40
Miles, Brodie, Shannon M. Miller i Elizabeth Connick. "CD4 T Follicular Helper and Regulatory Cell Dynamics and Function in HIV Infection". FRONTIERS MEDIA SA, 2016. http://hdl.handle.net/10150/622733.
Pełny tekst źródłaStreszczenie:
T follicular helper cells (T-FH) are a specialized subset of CD4 T cells that reside in B cell follicles and promote B cell maturation into plasma cells and long-lived memory B cells. During chronic infection prior to the development of AIDS, HIV-1 (HIV) replication is largely concentrated in T-FH. Paradoxically, T-FH numbers are increased in early and midstages of disease, thereby promoting HIV replication and disease progression. Despite increased T-FH numbers, numerous defects in humoral immunity are detected in HIV-infected individuals, including dysregulation of B cell maturation, impaired somatic hypermutation, and low quality of antibody production despite hypergammaglobulinemia. Clinically, these defects are manifested by increased vulnerability to bacterial infections and impaired vaccine responses, neither of which is fully reversed by antiretroviral therapy (ART). Deficits in T-FH function, including reduced HIV-specific IL-21 production and low levels of co-stimulatory receptor expression, have been linked to these immune impairments. Impairments in T-FH likely contribute as well to the ability of HIV to persist and evade humoral immunity, particularly the inability to develop broadly neutralizing antibodies. In addition to direct infection of T-FH, other mechanisms that have been linked to T-FH deficits in HIV infection include upregulation of PD-L1 on germinal center B cells and augmented follicular regulatory T cell responses. Challenges to development of strategies to enhance T-FH function in HIV infection include lack of an established phenotype for memory T-FH as well as limited understanding of the relationship between peripheral T-FH and lymphoid tissue T-FH. Interventions to augment T-FH function in HIV-infected individuals could enhance immune reconstitution during ART and potentially augment cure strategies.
41
Tabata, Rosana. "Proteção cruzada entre bacterinas antileptospirose produzidas com três representantes do Sorogrupo Sejroe. Ensaio experimental em hamsters (Mesocricetus auratus)". Universidade de São Paulo, 2002. http://www.teses.usp.br/teses/disponiveis/10/10134/tde-03052004-144546/.
Pełny tekst źródłaStreszczenie:
Foi investigada a existência de proteção cruzada entre bacterinas bivalentes formuladas com um de três representantes do Sorogrupo Sejroe: hardjo (bacterina A), wolffi (bacterina B) e guaricura (bacterina C), e uma estirpe do sorovar pomona, empregada por ser patogênica para hamsters (Mesocricetus auratus) e possibilitar a realização do teste de potência com desafio. Os ensaios foram efetuados em hamsters machos, comparando-se os níveis de anticorpos aglutinantes e neutralizantes, respectivamente obtidos nos testes de soroaglutinação microscópica (SAM) e inibição de leptospiras in vitro (ICL). Os animais receberam duas doses de vacinas via subcutânea; aos dez dias da segunda dose, foram inoculados com culturas não inativadas dos respectivos sorovares do Sorogrupo Sejroe. Aos 21 dias pós-infecção (d.p.i.), os animais foram sangrados, os soros (n=8) foram agrupados em pools e submetidos aos testes de SAM e ICL. O teste de potência com desafio para o sorovar pomona foi adaptado do protocolo preconizado pelo United States Department of Agriculture, mas as vacinas não foram diluídas e o esquema de imunização empregou duas aplicações de 1,0mL pela via subcutânea em intervalo de dez dias; o desafio foi efetuado aos dez dias da segunda aplicação; os óbitos por leptospirose foram registrados e, aos 21 d.p.i., os sobreviventes foram sacrificados e a condição de portadores renais foi investigada através de cultivos de tecido renal para isolamento de leptospiras. No teste de potência com o sorovar pomona, o número de doses infectantes empregado para desafio (100) situou-se dentro da faixa preconizada (10 a 100); respectivamente para as bacterinas A, B e C, as proporções de mortes por leptospirose entre os animais vacinados foram de 1/10, 0/10 e 3/10, e as de portadores renais de leptospiras entre os sobreviventes foram 2/9, 1/10 e 2/7. Os resultados do teste SAM revelaram que a bacterina A induziu reações para os sorovares hardjo e wolffi; a bacterina B, para hardjo, wolffi e guaricura, e a bacterina C, apenas para a guaricura, e do teste de ICL, que animais vacinados com as bacterinas B ou C apresentaram proteção para hardjo, wolffi e guaricura; entretanto, a bacterina A conferiu proteção apenas para wolffi. Apesar das variações no poder imunogênico segundo a estirpe de leptospira empregada para a produção das bacterinas, houve proteção cruzada entre os sorovares hardjo, wolffi e guaricura.The existence of cross-protection among bivalent bacterins, produced with one of three leptospires belonging to Serogroup Sejroe: hardjo (bacterin A), wolffi (bacterin B) and guaricura (bacterin C), and a strain of serovar pomona (included because of its pathogenicity to hamsters and the possibility of performing potency assay with challenge), was investigated in male hamsters (Mesocricetus auratus) by comparison of agglutinating and neutralizing antibodies titers, respectively measured by microscopic agglutination (MAT) and in vitro growth inhibition (GIT) tests. All animals received two doses of bacterins by subcutaneous route; after ten days from the second dose, they were inoculated with non-inactivated cultures of respective serovars of Serogroup Sejroe. At 21 post-challenge day (p.c.d.), all animals were bled and their sera (n=8) were joined in pools and tested by MAT and GIT. The potency assay with challenge performed only with serovar pomona was modified from protocol of USDA, but vaccines were not diluted and the immunization schedule employed two 1.0 mL vaccine doses by subcutaneous route with 10?day interval; the challenge was performed after ten days from the second dose; the number of deaths due to leptospirosis was registered; at 21 p.c.d., the survivors were sacrificed and their renal carrier state was investigated by culture of renal tissue for leptospires isolation. In the potency assay with serovar pomona, the number of infectious doses employed for challenge (100 infective units) was in accordance with the recommended range (10-1,000 infective units); respectively to bacterins A, B and C, proportions of deaths due to leptospirosis among vaccinated animals were 1/10, 0/10 and 3/10, and proportions of leptospires renal carrier among survivors were 2/9, 1/10 and 2/7. Results of MAT showed that bacterin A induced reactions against serovars hardjo and wolffi; bacterin B, against hardjo, wolffi and guaricura, and bacterin C, against guaricura; results of GIT revealed that vaccinated animals with bacterins B or C presented protection against serovar hardjo, wolffi and guaricura; however, bacterin A induced protection only against wolffi. A cross?protection was observed among serovars hardjo, wolffi and guaricura, although variations exist in the immunogenic capacity according to the strain of leptospires used for the bacterins production.
42
Pötzsch, Sonja [Verfasser], i Thomas [Akademischer Betreuer] Winkler. "A comprehensive antibody repertoire analysis reveals novel antigenic domains on glycoprotein B of human cytomegalovirus that induce potently neutralizing antibodies / Sonja Pötzsch. Betreuer: Thomas Winkler". Erlangen : Friedrich-Alexander-Universität Erlangen-Nürnberg (FAU), 2011. http://d-nb.info/107758220X/34.
Pełny tekst źródła
43
Miyaji, Karina Takesaki. "Persistência de anticorpos neutralizantes anti-febre amarela em pessoas com 60 anos ou mais previamente vacinadas". Universidade de São Paulo, 2015. http://www.teses.usp.br/teses/disponiveis/5/5134/tde-06082015-113813/.
Pełny tekst źródłaStreszczenie:
INTRODUÇÃO: A Febre Amarela (FA) é uma doença viral aguda endêmica em grande parte do Brasil. A principal medida de prevenção é a vacinação. O presente estudo avaliou a prevalência e os títulos de anticorpos neutralizantes em pessoas com 60 anos ou mais que haviam recebido anteriormente a vacina de FA 17DD, em comparação a adultos saudáveis com 18 a 59 anos. Além disso, foram avaliadas a correlação entre os títulos de anticorpos e o tempo decorrido desde a vacinação, nos participantes que receberam apenas uma dose da vacina, e a prevalência de anticorpos nos vacinados há menos e há mais de dez anos. MÉTODOS: Os participantes foram recrutados entre pessoas que procuraram o Centro de Referência para Imunobiológicos Especiais (CRIE) do Hospital das Clínicas da Faculdade de Medicina da Universidade de São Paulo (HCFMUSP) para receber diferentes vacinas e que referiam ter recebido a vacina de FA anteriormente. Os seguintes dados foram coletados: idade, etnia, sexo, número de doses da vacina de FA recebidas, data da última vacinação de FA. Foi realizada contagem de linfócitos TCD4+ usando citometria de fluxo. Os anticorpos neutralizantes contra FA foram dosados pelo teste de neutralização por redução de 50% das placas de lise (PRNT50). RESULTADOS: Foram incluídos 94 indivíduos, 46 com idade de 60 anos ou mais (Grupo 1) e 48 com 18 a 59 anos (Grupo 2). Não houve diferença significativa entre os dois grupos na distribuição de gênero, etnia, número de doses de vacina de FA recebidas anteriormente, tempo desde a última dose e contagem de linfócitos TCD4+. Não houve diferença na prevalência de anticorpos neutralizantes anti-FA entre os dois grupos (87% e 93,8% nos Grupos 1 e 2, respectivamente, p=0,263). O título médio geométrico (GMT) dos anticorpos neutralizantes foi maior no grupo mais jovem (3,77 log10mUI/mL) comparado ao grupo mais velho (3,64 log10mUI/mL) e essa diferença foi estatisticamente significante (p=0,022). Não foi encontrada correlação entre os títulos de anticorpos neutralizantes e o tempo decorrido desde a vacinação entre os participantes que receberam apenas uma dose de vacina, tendo sido analisados os dois grupos conjuntamente. Também não foi encontrada diferença estatisticamente significativa na prevalência de anticorpos neutralizantes entre os participantes que receberam apenas uma dose da vacina de FA há mais de 10 anos ou há menos de 10 anos. CONCLUSÕES: São necessários outros estudos de persistência de anticorpos na população idosa devido à possibilidade de resposta vacinal alterada pela imunosenescênciaINTRODUCTION. Yellow Fever (YF) is an acute viral disease endemic in large parts of Brazil. The main preventive measure is vaccination. This study aimed to assess the prevalence and titers of neutralizing antibodies in persons aged 60 years and older who had previously received YF 17DD vaccine, in comparison to healthy adults aged 18 to 59 years. The study also evaluated the correlation between the antibodies titers and the time elapsed since vaccination, in participants who had received a single dose of the vaccine, and the prevalence of antibodies in participants vaccinated within ten years and more than ten years before enrollment. METHODS. Participants were recruited among persons who came to the Reference Center for Special Immunobiologicals (Centro de Referência para Imunobiológcos Especiais, CRIE) of the Hospital das Clínicas da Faculdade de Medicina da Universidade de São Paulo (HC-FMUSP) to receive any vaccine and who had previously received the YF vaccine. The following data were collected: age, ethnicity, gender, number of YF vaccine doses taken and date of last YF vaccination. CD4 T cells counts were performed using flow cytometry. YF neutralizing antibodies were measured using test of neutralization by 50% reduction of lysis plaques (PRNT50). RESULTS. Ninety-four subjects were enrolled: 46 persons aged 60 years and older (Group 1) and 48 persons aged 18 to 59 years (Group 2). There was no significant difference between the groups regarding gender, ethnicity, number of YF vaccine doses previously received, time since the last dose and CD4+T cells count. There was no difference in the prevalence of YF neutralizing antibodies between the groups (87% and 93.8% in Groups 1 and 2, respectively, p=0.263). The log-transformed Geometric Mean Titer (GMT) of YF neutralizing antibodies was higher in the younger group (3.77 log10mUI/mL) in comparison to the older group (3.64 log10mUI/mL), and this difference was statistically significant (p=0.022). There was no correlation between YF neutralizing antibodies titers and time elapsed since vaccination among the participants who had previously received a single dose of YF vaccine, with the two groups analyzed together. There was no significant difference in the prevalence of neutralizing antibodies among participants who received a single dose of YF vaccine within ten years or more than 10 years before enrollment. CONCLUSIONS. Further studies on antibodies persistence in the elderly are necessary, considering the possibility of compromised immune response to vaccines due to immunosenescence
44
Lucar, Olivier. "Implication des cellules Natural Killer dans la physiopathologie des infections chroniques VIH et VHC : application à des stratégies thérapeutiques". Thesis, Paris 6, 2017. http://www.theses.fr/2017PA066471/document.
Pełny tekst źródłaStreszczenie:
Les infections chroniques de l’Immunodéficience Humaine et de l’Hépatite C (VIH et VHC) sont à l’origine de pandémies. Malgré des traitements avancés, leurs relations avec le système immunitaire ne sont pas résolues et restent nécessaires pour établir de nouvelles stratégies thérapeutiques. Les cellules Natural Killer (NK) sont des effecteurs majeurs antiviraux et sont importants pour l’immunité innée et adaptative. Ils contrôlent leur cytotoxicité et leur fonction immunorégulatrice via de multiples récepteurs activateurs et inhibiteurs qui sont enclenchés par interaction avec leurs ligands respectifs. Parmi tous les récepteurs, je me suis particulièrement intéressé aux Natural Cytotoxicity Receptors NKp30 et NKp44. Il est intéressant de noter que le laboratoire a précédemment identifié un épitope conservé de la gp41 du VIH-1 qui induit l’expression du ligand de NKp44 sur des LT CD4+ les rendant susceptibles à la lyse par des cellules NK-NKp44+. Après plusieurs études, le laboratoire a mis en place une stratégie vaccinale basée sur un peptide de l’épitope conservé de la gp41 qui induit chez la souris des Anticorps Neutralisants (AcNs W614A-3S) contre l’infection VIH-1. Alors que le VIH-2 est considéré comme un modèle unique d’une infection VIH contrôlée, les données sur les cellules NK y sont très limitées. Nous avons observé une dérégulation du récepteur NKp30 et une augmentation de ses ligands qui conduisent à des déficiences de leurs fonctions et représentent un nouveau mécanisme de persistance virale. Ensuite, nous avons observé pendant l’infection chronique par le VHC une forte proportion de cellules NK intra-hépatiques exprimant NKp44 qui corrèle avec la fibrose et la charge virale. De plus, nous avons identifié un épitope conservé de la protéine Core du VHC qui induit le ligand de NKp44 sur des lignées hépatiques. Ces données suggèrent que la déplétion des hépatocytes passe par un mécanisme similaire à celui observé pendant l’infection VIH-1. Enfin, une étude sur des patients VIH-1 contrôleurs nous a permis d’identifier la présence d’AcNs W614A-3S qui sont associées au contrôle viral et au maintien de LT CD4+ fonctionnelles. Ces données ont confirmé le potentiel de ces AcNs et, dont leur production par vaccination, ont été confirmée chez le lapin et le singe. Ainsi, ces études apportent de nouvelles données dans les relations entre les cellules NK et le VIH ou le VHC ainsi que de nouvelles stratégies thérapeutiques. Ces études ont notamment confirmé le pouvoir des AcNs W614A-3S dans un vaccin contre le VIH-1Human Immunodeficiency and Hepatitis C (HIV and HCV) chronic infections are at the origin of pandemics. Despite advance drug treatments, their relationship with the immune system is not resolved and is still required to establish new therapeutic strategies. Natural Killer (NK) cells are major antiviral effectors of the immune system and are important for innate and adaptive immune processes. They mediate cytotoxicity and immunoregulation via various activator and inhibitor receptors that are triggered upon interaction with their cognate ligands. Among all receptors, I particularly took an interest in Natural Cytotoxicity Receptors NKp30 and NKp44. Interestingly, the lab previously identified a conserved HIV-1 gp41 épitope that induce expression of NKp44 ligand on CD4+ T cells making them susceptible to lyses by NK-NKp44+ cells. After various studies, the lab established a vaccine strategy based on a peptide from the conserved gp41 épitope that induced in mice Neutralizing Antibodies (Nab W614A-3S) against HIV-1 infection. Whereas HIV-2 infection could be considered as a HIV control infection unique model, data on NK cells are very limited. We found a down-modulation of NKp30 receptor and an increased of its ligands that lead to functional impairments of NK cells and could represent a new viral persistence mechanism. Then, during the HCV chronic infection we found a high proportion of intrahepatic NK cells expressing NKp44 that correlates with fibrosis and viral load. Furthermore we identified a conserved épitope of HCV core protein that induced NKp44 ligand on hepatic cell lines. These data suggest that destruction of hepatocyte might occur by a similar mechanism observed during HIV-1 infection. Finally, a study on HIV-1 controllers patients allow us to identify the presence of Nab W614A-3S that correlates with viral control and the preservation of functional CD4+ T cells. These data confirm the potency of this Nab and their induction by vaccination has been also confirmed in rabbit and macaques. Thus, these studies highlight new data regarding relationship between NK cells and HIV or HCV that could represent new therapeutic approaches. These studies especially confirm the potency of Nab W614A-3S to implement a vaccine against HIV-1
45
Argolo, Angela Ferreira Lopes de Teive e. "Circulação dos vírus dengue no estado de Goiás: vigilância laboratorial (1994-2013) e perfil de anticorpos neutralizantes sorotipo específico durante o surto de 2013 em Goiânia". Universidade Federal de Goiás, 2014. http://repositorio.bc.ufg.br/tede/handle/tede/8774.
Pełny tekst źródłaStreszczenie:
Submitted by Marlene Santos (marlene.bc.ufg@gmail.com) on 2018-08-08T19:03:16Z
No. of bitstreams: 2
Tese - Angela Ferreira Lopes de Teive e Argolo - 2014.pdf: 1773502 bytes, checksum: 393b63ce506fdd2be70f95e14be99755 (MD5)
license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5)Approved for entry into archive by Luciana Ferreira (lucgeral@gmail.com) on 2018-08-09T11:52:01Z (GMT) No. of bitstreams: 2 Tese - Angela Ferreira Lopes de Teive e Argolo - 2014.pdf: 1773502 bytes, checksum: 393b63ce506fdd2be70f95e14be99755 (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5)
Made available in DSpace on 2018-08-09T11:52:01Z (GMT). No. of bitstreams: 2 Tese - Angela Ferreira Lopes de Teive e Argolo - 2014.pdf: 1773502 bytes, checksum: 393b63ce506fdd2be70f95e14be99755 (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) Previous issue date: 2014-12-19
Conselho Nacional de Pesquisa e Desenvolvimento Científico e Tecnológico - CNPq
Fundação de Amparo à Pesquisa do Estado de Goiás - FAPEG
Dengue is an endemic disease in Goias state with current circulation of the four serotypes. In the last five years has been occurred a significant increase in cases, hospitalizations and deaths caused by dengue in the region. In this thesis the first manuscript analyzed data from the virological surveillance of DENV in Goias during 1994-2013. Virus isolation in C6/36 (A.albopictus) cell culture followed by indirect immunofluorescence using monoclonal antibodies were performed in LACEN-GO. During the study period the four serotypes have been identified with introduction of DENV-1 in 1994, DENV-2(1998), DENV-3(2002) and DENV-4(2011). Among 21,684 virus isolation tests performed 4,839(22.3%) were positive. DENV-1 was isolated in 3,023(62.5%), DENV-3 in 1,051 (21.7%), DENV-4 in 413(8.5%) and DENV-2 in 352(7.3%) samples respectively. There were changes in the predominance of serotypes throughout the study period with dominance of DENV-1 between 1994-2002 and 2009-2011, DENV-3 in 2003-2008 and DENV-4 in 2013. The DENV-2 was never prevalent among serotypes isolated in LACEN-GO. The time series of the virological surveillance of dengue in Goias showed the introduction and the temporal sequence of circulation of DENV in the state. This study contributes to characterize the epidemiological scenario of dengue in Goias with possible implications for epidemiological surveillance of the disease in the region. The second manuscript evaluated the presence of dengue serotype-specific neutralizing antibodies (NAb) in patients involved in a clinical dengue cohort established during epidemic of 2012-2013 in Goiania, state of Goias. Among 452 participants of the cohort we analyzed the paired samples from 60 patients classified as severe dengue (n=40) or dengue fever (n=20). The eligibility criteria were confirmation of the infection by IgM, NS1Ag and /or RT-PCR; IgG positive in at least one of the samples. Monotypic or multitypic response were defined: PRNT50≥1/20 for only one serotype or ≥1/20 for two or more serotypes simultaneously. The infecting serotype was defined by ≥4 fold increase in the title of NAb in paired samples. Overall 73.3% of patients had NAb against DENV-4, 68.3% against DENV-1, 68.3% against DENV-2 or 61.6% against DENV-3. The patients' ages ranged from 3 to 81 years. Regardless of age group 85% of patients had multitypic response while 15% had monotypic response. Patients infected with DENV-4 showed the greatest difference in NAb titers in comparison to other serotypes indicating predominance of seroconversion for serotype 4. There was no correlation among preexistent NAb and disease severity. The characterization of dengue serotype-specific immune response is important to study the relationship of preexistence of neutralizing antibodies with clinical outcomes of disease also facing the perspective of introducing of antidengue vaccines to identify protective immunity.
A dengue é uma doença endêmica em Goiás com atual circulação dos quatro sorotipos virais com aumento expressivo no número de casos, hospitalizações e óbitos pela doença de forma acentuada nos últimos cinco anos. Na presente tese o primeiro manuscrito analisou os dados da vigilância virológica dos DENV em Goiás no período de 1994-2013. Os testes de isolamento viral em cultura celular C6/36(A.albopictus) seguido de imunofluorescência indireta utilizando anticorpos monoclonais foram realizados no LACEN-GO. No período do estudo, os quatro sorotipos virais foram identificados no estado com introdução do DENV-1 em 1994, DENV-2(1998), DENV-3(2002) e DENV-4(2011). Do total de 21.684 testes de isolamento viral 4.839(22,3%) foram positivos. DENV-1 foi isolado em 3.023(62,5%), DENV-3 em 1.051(21,7%), DENV-4 em 413(8,5%) e DENV-2 em 352(7,3%) amostras. A predominância dos sorotipos virais alternou ao longo do período com DENV-1 dominante entre os anos de 1994-2002 e 2009-2011, DENV-3 entre 2003-2008 e DENV-4 em 2013. O DENV-2 nunca foi predominante entre os sorotipos isolados no LACEN-GO. A série histórica da vigilância virológica da dengue em Goiás mostrou a introdução e a sequência temporal da circulação dos DENV no estado. Este estudo contribui para a caracterização do cenário epidemiológico da dengue em Goiás com possíveis implicações nas ações de vigilância epidemiológica da doença na região. O segundo manuscrito avaliou a presença de anticorpos neutralizantes (AcN) antidengue sorotipo específico em pacientes de uma coorte clínica de dengue estabelecida durante a epidemia de 2012-2013, em Goiânia-GO. Entre os 452 participantes da coorte, analisamos amostras pareadas de 60 pacientes classificados como dengue grave (n=40) ou dengue (n=20). Os critérios de elegibilidade foram a confirmação da infecção por IgM, NS1Ag e/ou RT-PCR; IgG positivo em pelo menos uma das amostras. Resposta monotípica ou multitípica foram definidas por PRNT50≥1/20 para apenas um sorotipo viral ou ≥1/20 para dois ou mais sorotipos virais simultaneamente. O sorotipo viral infectante foi definido por um aumento ≥4 vezes no título dos AcN em amostras pareadas. No total 73,3% dos pacientes apresentaram AcN contra DENV-4, 68,3% contra DENV-1, 68,3% contra DENV-2 ou 61,6% contra DENV-3. A idade dos pacientes variou de 3 a 81 anos. Independente da faixa etária 85% dos pacientes apresentaram resposta multitípica enquanto 15% tiveram resposta monotípica. Pacientes infectados por DENV-4 apresentaram a maior diferença de títulos de AcN em comparação com outros sorotipos virais indicando predominância de soroconversão para o sorotipo 4. Não houve correlação entre preexistência de AcN com a gravidade da doença. A caracterização de resposta imune sorotipo específica é importante para estudar a relação de anticorpos neutralizantes preexistentes com a gravidade clínica da doença, bem como frente à perspectiva de introdução de vacinas antidengue para identificar a imunidade protetora.
46
Grundner, Christoph. "HIV-1 immune escape and neutralizing antibodies solid phase proteoliposomes containing HIV-1 envelope glycoproteins as antigens and immunogens and HIV-1 core envelope glycoproteins deficient in T-helper epitopes /". [S.l.] : [s.n.], 2002. http://deposit.ddb.de/cgi-bin/dokserv?idn=965908992.
Pełny tekst źródła
47
Henzel, Andréia. "Experimental pathogenesis of acute and latent genital infection by bovine herpesvirus type 1.2 in heifers". Universidade Federal de Santa Maria, 2012. http://repositorio.ufsm.br/handle/1/4062.
Pełny tekst źródłaStreszczenie:
Coordenação de Aperfeiçoamento de Pessoal de Nível SuperiorThe feline calicivirus (FCV) and the felid herpesvirus type 1 (FeHV-1) are the main agents of the respiratory tract diseases of felines. Both viruses are distributed worldwide, however its distribution and prevalence in Brazil are not well known. In the present thesis we describe the isolation and one serosurvey of FCV and FeHV-1 in some counties of the Rio Grande do Sul State, Brazil; and the molecular characterization of the capsid protein gene of FCV isolates is also described. In Chapter 1, we describe the epidemiologic survey of FCV and FeHV-1 through investigation of the conjunctival, nasal, oral and oropharyngeal swabs from 302 domestic cats. The viral isolation was performed in Crandell-Reese feline kidney cells and the isolates were submitted to PCR and RT-PCR for confirmation of the presence of the FeHV-1 and FCV, respectively. Fifty five (18.2%) of the 302 cats analyzed were positive for the viral isolation; when tested by PCR, 29 cats (52.7%) were shedding the FCV, 21 (38.2%) shedding FeHV-1, and 5 cats (9.1%) shedding both viruses. In addition, isolates of FCV and FeHV-1 were standardized to be used as control in the PCR and virus neutralization (VN) assays (serological technical used in chapter 3 of the thesis). The SV65/90 (FCV) and the SV534/00 (FeHV-1) isolates were defined as standard viruses for the assays. In Chapter 2, the molecular characterization of 13 FCV isolates is described; ten isolates came from the survey performed in chapter 1, two were deposited in the virology laboratory of the UFSM and one was obtained from a cat with gingivitis-stomatitis syndrome. The ORF2 (open reading frame) region that codifies the major protein of the viral capsid was sequenced, which includes an immunodominant region of the virus. The ORF 2 was analyzed at the nucleotide and amino acid levels and these data was used for phylogenetic studies of the 13 Brazilian isolates of FCV. These isolates were compared to ten reference strains of FCV and to the San Miguel sea lion virus type 1 (SMSV-1) as an outgroup in phylogenetic study. The primers were designed using the complete sequence of FCV-F9. The comparison of the 13 Brazilian isolates with the F9 strain revealed a large molecular diversity among them, concentrated mainly along the linear epitopes of the region. The Chapter 3 describes a serosurvey against FCV and FeHV-1 in serum samples from domestic feline. From the 630 samples tested by the VN assay, 53.6% (338/630) were positive to one or both viruses with a prevalence of 39.2% for neutralizing antibodies against FCV and 30.6% for FeHV-1. The results from the present study demonstrated that the FCV and FeHV-1 are circulating among the feline population examined, and also, the presence of carriers of the viruses among these cats. Besides, the molecular characterization of the FCV evidenced the great genetic variability of the Brazilian isolates when compared to vaccine and reference strains.
O calicivírus felino (feline calicivirus FCV) e o herpesvírus felino tipo 1 (felid herpesvirus type 1 FeHV-1) são os principais agentes das doenças do trato respiratório dos felinos. Ambos os vírus são mundialmente distribuídos, mas no Brasil sua distribuição e prevalência são ainda pouco conhecidas. Na presente tese descrevemos o isolamento e sorologia do FCV e do FeHV-1 em alguns municípios do estado do Rio Grande do Sul (RS), Brasil; e a caracterização molecular do gene da proteína do capsídeo de isolados de FCV. No Capítulo 1, descreve-se o isolamento do FCV e do FeHV-1 por meio da coleta de suabes (conjuntival, nasal, oral e orofaringeano) de 302 gatos domésticos. O material coletado foi inoculado em cultivo celular de origem felina (CRFK) e submetidos a PCR e RT-PCR para confirmação em FeHV-1 e FCV, respectivamente. Dos 302 gatos analisados, 55 (18,2%) foram positivos no isolamento; e quando testados por PCR, 29 dos 55 gatos (52,7%) excretavam o FCV, 21 (38,2%) excretavam o FeHV-1 e 5 gatos (9,1%) apresentavam ambos os vírus. Além dessa descrição, realizou-se a padronização de um isolado de FCV e de FeHV-1, para serem utilizados como controle nas técnicas de PCR e na vírus neutralização [VN] (técnica sorológica utilizada no capítulo 3 da tese). O SV65/90 (FCV) e o SV534/00 (FeHV-1) foram os isolados definidos como vírus-padrão. No Capítulo 2, incluímos o estudo sobre a análise molecular de 13 isolados de FCV; dez isolados foram obtidos a partir do material descrito no capítulo 1, dois estavam armazenados no laboratório de virologia da UFSM e um foi obtido de um gato acometido pela síndrome gengivo-estomatite. A região analisada foi o gene da proteína do capsídeo, codificado pela ORF2 (open reading frame), na qual se localizam as regiões imunodominantes do FCV. A ORF2 foi analisada em nível de nucleotídeos e aminoácidos; e uma análise filogenética dos 13 isolados brasileiros de FCV, incluindo dez cepas de referência e o vírus dos leões marinhos de San Miguel tipo 1 (San Miguel sea lion virus type 1 SMSV-1) como um outgroup, foi também realizada. Os primers foram desenhados com base na sequência completa do genoma da cepa de referência a FCV-F9. Quando os 13 isolados brasileiros de FCV foram comparados a cepa F9, detectou-se grande diversidade molecular nas regiões variáveis do gene e nos epitopos lineares mapeados. O Capítulo 3 contem um estudo sorológico contra FCV e FeHV-1 em amostras de soro de felinos domésticos. Das 630 amostras testadas pela VN, 53,6% (338/630) foram positivas para um ou ambos os vírus; sendo a prevalência de anticorpos neutralizantes contra o FCV de 39,2% e do FeHV-1 de 30,6%. Os resultados aqui apresentados demonstram a circulação do FCV e do FeHV-1 na população de gatos domésticos analisados, assim como, a presença de gatos portadores para ambos os vírus. Além disso, a caracterização molecular do FCV demonstrou a grande variabilidade genética dos isolados brasileiros em comparação às cepas vacinais e às de referência.
48
Rousset, Claire. "Coévolution entre les glycoprotéines d'enveloppe du VIH et les anticorps neutralisants à large spectre ciblant la région du glycane N332". Thesis, Université Grenoble Alpes (ComUE), 2018. http://www.theses.fr/2018GREAV071/document.
Pełny tekst źródłaStreszczenie:
Le VIH est la cause de la pandémie de SIDA depuis les années 1980. Avec plus d’un million de nouvelles infections chaque année, un vaccin prophylactique est indispensable pour bloquer de façon définitive la propagation du virus. Parmi les stratégies vaccinales, l’induction d’anticorps neutralisants à large spectre est une des plus prometteuses, car ceux-ci pourraient protéger contre l’infection par la grande diversité génétique des souches de VIH circulantes dans le monde. A ce jour, aucun immunogène n’a permis l’induction de tels anticorps, mais ils ont été isolés à partir de personnes infectées par le VIH. En effet, une faible fraction d’individus infectés développe des anticorps neutralisants à large spectre qui ciblent des régions vulnérables et conservées de la glycoprotéine d’enveloppe. La région du patch riche en mannose, centrée autour du glycane en position N332 de la gp120, est la plus fréquemment ciblée, et est à cet égard attractive d’un point de vue vaccinal.Afin de mieux comprendre comment se développent les anticorps ciblant le patch riche en mannose, nous avons étudié un donneur sélectionné de la cohorte du Protocole C de l’International AIDS Vaccine Initiative, et ayant une activité neutralisante sérique exceptionnelle. Nous avons isolé, à partir des cellules sanguines de cet individu, deux lignées d’anticorps ciblant la région N332, que nous avons caractérisées pour leur activité neutralisante et dont nous avons cartographié l’épitope. Nous avons également cartographié le paratope d’une lignée d’anticorps issue d’un autre donneur du Protocole C ciblant également la région N332. Nos résultats font apparaître la diversité de solutions adoptées pour atteindre une neutralisation à large spectre contre cette région. Les études de lignées, telles que nous l’avons entrepris, permettent d’appréhender comment la coévolution anticorps-virus conduit à la sélection d’anticorps neutralisants à large spectre. Le but ultime est d’utiliser les connaissances ainsi générées, pour mettre au point des immunogènes et des protocoles d’immunisations, visant à induire des lignées d’anticorps spécifiques et à conduire leur évolution vers la neutralisation à large spectreHIV has been the cause of the AIDS pandemic since the 1980s. With over a million new infections each year, a prophylactic vaccine is needed to stop the virus spread. Among vaccine strategies, the induction of broadly neutralizing antibodies is one of the most promising, as they could protect against infection by the huge genetic diversity of circulating HIV strains. To date, no immunogen has induced such antibodies, but they have been isolated from HIV infected people. Indeed, a small fraction of infected individuals eventually develops broadly neutralizing antibodies that target vulnerable and conserved sites of the envelope glycoprotein. The region of the high-mannose patch, centred around a glycan at position N332 of gp120, is the most frequently targeted, and is therefore attractive from a vaccination standpoint.In order to better understand how antibodies targeting the high-mannose patch develop, we studied a donor selected from the International AIDS Vaccine Initiative Protocol C cohort with exceptional serum neutralizing activity. We isolated two antibody lineages targeting the N332 region from this individual's blood cells, which we characterized for their neutralizing activity and mapped their epitope. We also mapped the paratope of an antibody lineage from another Protocol C donor, also targeting the N332 region. Our results show the great diversity of solutions to achieve broad neutralization against this region. Lineage studies, as we have undertaken, provide an understanding of how antibody-virus coevolution leads to the selection of broadly neutralizing antibodies. The ultimate goal is to use this knowledge to develop immunogens and immunization protocols, to induce specific antibody lineage and drive their evolution towards broad neutralization
49
Reinhard, Henrike Christiane. "Struktur-Funktions-Beziehung der HCMV-kodierten Fcgamma-Rezeptoren gp34 und gp68". Doctoral thesis, Humboldt-Universität zu Berlin, Mathematisch-Naturwissenschaftliche Fakultät I, 2010. http://dx.doi.org/10.18452/16124.
Pełny tekst źródłaStreszczenie:
Neutralisierende Antikörper sind entscheidend in der Eindämmung der Virusinfektion, indem sie den Eintritt in die Wirtszelle hemmen bzw. die Aktivierung der Komplementkaskade initiieren. Distinkte wirtseigene Oberflächenrezeptoren für die Fc-Domäne von IgG (FcyR) sind für die Kommunikation von humoraler und zellulärer Immunantwort verantwortlich. Auch Mitglieder der Herpesviren kodieren für Fc-bindende Proteine, die Kandidaten für immunevasive Funktionen darstellen könnten. Der Nachweis der HCMV-kodierten Fc-bindenden Proteine gp34 und gp68 als Bestandteil der Virushülle lies auf eine immunevasive Funktion hinsichtlich neutralisierendem IgG und Komplement-vermittelter Virolyse schließen, wie für den HSV-1-kodierten FcyR gE beschrieben. Weder für gp34 noch für gp68 konnte in vitro ein hemmender Effekt auf Neutralisation und Virolyse beobachtet werden. In unserem Labor wurde jedoch gezeigt, dass gp34 und gp68 selektiv die IgG-abhängige Aktivierung zellulärer FcyR inhibieren. Die glykosylierungsunabhängige Ligandenbindung von gp34 und gp68 wies auf unterschiedliche Interaktionsmechanismen zwischen den zellulären und den viralen FcyR hin. Mithilfe eines mutierten Fc-Fragments konnte für gp68 eindeutig eine mit HSV-1 gE überlappende Bindestelle an IgG identifiziert werden. Die für die Ligandenbindung erforderlichen Aminosäuren 71-292 von gp68 binden Fc in einer 2:1 Stöchiometrie, wobei die N-, nicht aber die O-Glykosylierung des vFcyRs essentiell sind. Darüber hinaus formt gp34 auf infizierten Zellen und auf der Virushülle kovalente Homooligomere. gp34-Cysteinpunktmutanten auf Basis der für die Bindung notwendigen Aminosäuren 24-140 lassen vermuten, dass die Oligomerisierung Voraussetzung für die Fc-Bindung ist. Im Gegensatz zu gp68 scheint der Mechanismus der Fc-Bindung von gp34 einzigartig unter den bekannten Fcy-Rezeptoren zu sein. Diese Ergebnisse lassen vermuten, dass trotz redundanter Expression der HCMV-FcyR der Bindungs- und Wirkungsmechanismus selektiv ist.Neutralizing IgGs play a key role in diminishing virus infectivity by inhibiting the entry into host cells. Additionally, IgG-bound particles may be inactivated by virolysis through the activation of complement. Surface receptors specific for the Fc domain of IgG represent host proteins, connecting humoral and cellular immune responses. Also members of the herpes virus family code for proteins with Fc binding properties, implying functions that could intervene with antibody-dependent effector mechanisms. The presence of gp34 and gp68 on the virion membrane raised the question whether they are able to inhibit neutralising IgG and complement-mediated virolysis. The HSV-1-encoded FcyR gE was described to affect neutralisation and virolysis. Despite extensive analysis, there were no implications found that gp34 or gp68 interfere with neutralising IgG or virolysis in vitro. However, our lab could demonstrate that gp34 and gp68 selectively inhibit the IgG-dependent activation of the different host FcyRs. In contrast to the cFcyRs Fc recognition by gp34 and gp68 occurs independently of N-linked glycosylation of IgG, which points to a different binding mechanism among host and viral FcyRs. By taking advantage of a mutated Fc fragment, overlapping binding regions of the HSV-1 gE and gp68 were identified. For gp68 the amino acids 71-292 including the N-glycans are strictly required for Fc binding in a 2:1 stoichiometry. Interestingly, gp34 forms covalently linked homo-oligomers in infected cells and on the virion. Based on the minimal binding domain comprising the amino acids 24-140 of gp34, targeted cysteine exchange mutants revealed that oligomer formation by gp34 is absolutely required for Fc binding. In contrast to gp68, the Fc binding characteristics of gp34 appears to be unique among the known FcyRs. These findings allow us to postulate that even if the HCMV-encoded FcyRs are redundantly expressed the mechanistic details and binding properties are selective.
50
Fiebig, Uwe. "Induktion neutralisierender Antikörper gegen transmembrane Hüllproteine von Retroviren". Doctoral thesis, Humboldt-Universität zu Berlin, Mathematisch-Naturwissenschaftliche Fakultät I, 2008. http://dx.doi.org/10.18452/15746.
Pełny tekst źródłaStreszczenie:
Die Transplantation von porzinen Organen könnte eine Lösung des akuten Mangels von Allotransplantaten in der Transplantationsmedizin darstellen. Bevor die Xenotransplantation klinische Realität werden kann, sind jedoch zahlreiche Hürden zu überwinden. Insbesondere die mögliche Übertragung porziner endogener Retroviren (PERVs), die integraler Bestandteil des porzinen Genoms sind, stellt ein mikrobiologisches Risiko dar. PERVs können humane Zellen in vitro produktiv infizieren. Mögliche Strategien zur Abwehr von Xenosen sind die Verwendung von PERV-knockout Tieren oder die Entwicklung eines effektiven Impfstoffes, durch den der Transplantatrezipient vor einer möglichen Übertragung geschützt werden kann. Dazu wurden Antiseren gegen die Hauptstrukturproteine von PERV generiert. Es konnte gezeigt werden, dass in Ziegen und Ratten durch Immunisierung mit der rekombinant generiereten Ektodomäne des transmembranen Hüllprotein p15E neutralisierende Antikörper induziert werden konnten. Die Epitopkartierung der induzierten Antikörper zeigt eine Bindung an eine Domäne im N-terminalen, nahe des Fusionspeptids (E1, GPQQLEK) und eine Domäne im C-terminalen, membranproximalen (E2, FEGWFN) Bereich der p15E-Ektodomäne. Diese Sequenzen sind in allen PERVs identisch und innerhalb der Gammaretroviren hochkonserviert. Aus AIDS-Patienten isolierte neutralisierende Antikörper (mAb2F5: ELDKWA, mAb4E10: LWNWFN) binden ebenfalls an den C-terminalen Bereich der Ektodomäne des Transmembranproteins gp41. Der Bindungsmechannismus dieser Antikörper wurde in ELISA-Experimenten und in vitro-Inhibitionsassays analysiert. Die Ergebnisse legen die Bindung eines Konformationsepitopes nahe, das aus der E1 und der E2 Domäne gebildet wird. Die Aufklärung des Bindungsmechnismus breit neutralisierender Antikörper gegen Transmembranproteine von Retroviren könnte die Basis für neue Impfstoffansätze darstellen.Porcine xenotransplants may offer a potential solution to the problem posed by the limited supply of allotransplants. However, xenotransplantation may be associated with the risk of transmission of microorganisms, in particular of porcine endogenous retroviruses (PERVs) that are an integral part of the porcine genome and able to infect human cells in vitro. Possible strategies to prevent virus transmission include the development of PERV knockout animals or of effective vaccines. When antisera prepared against the main structural proteins of PERV were screened, goat and rat antisera against the recombinant ectodomain of the transmembrane envelope protein p15E were found to neutralize PERV infectivity. Epitope mapping using overlapping peptides revealed two epitopes, one near the fusion peptide (E1, GPQQLEK) and the other near the transmembrane domain (E2, FEGWFN). These sequences are identical for all PERVs and are highly conserved among all gammaretroviruses. Interestingly, neutralizing antibodies isolated from AIDS patients that recognize regions partially homologous with E2 (mAb4E10, LWNWFN) or located in close proximity to E2 (mAb2F5, ELDKWA) are known to neutralize a broad range of HIV-1 strains. The binding mechanisms of these HIV neutralizing antibodies were analyzed in ELISA experiments and in vitro inhibition assays. The results indicate that the two most broadly reactive HIV-1 envelope gp41 human mAbs are specific for a discontinuous epitope composed of the E1 and the E2 domain. If so, these two transmembrane protein domains in different retroviruses act as effective targets for neutralizing antibodies and may provide the basis for effective antiretroviral vaccines.