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1.
Bioconjug Chem ; 18(6): 2045-53, 2007.
Artigo em Inglês | MEDLINE | ID: mdl-17939731

RESUMO

4-(Methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) is one of the most abundant and potent procarcinogens in tobacco smoke. In order to induce a strong and substained antibody response against NNK, we developed a functionalized derivative that closely mimicked its structural features, in particular, the pyridyloxobutyl moiety, the adjacent ketone, and the N-nitrosamino group. This hapten was conjugated via a C 2 linker to the highly immunogenic diphteria toxoid licensed as a vaccine in humans to induce polyclonal and monoclonal antibodies. Two monoclonal antibodies were obtained with Kd values of 45.8 nM (P9D5) and 37.6 nM (P7H3), respectively, for NNK-C 2. Both the monoclonal (P9D5 and P7H3) and polyclonal antibodies reacted strongly with NNK (IC 50 = 80 microM or 160 microM) and NNAL (IC 50 = 29 microM or 93 microM) and to a lesser extent with some of the metabolites of NNK. Interestingly, the mAbs did not react with the metabolites of the detoxification pathways such as NNK-N-Oxide and NNAL-N-Oxide (IC 50 > 512 microM). Therefore, such antibodies detect NNK and NNAL and may have the potential to modulate their redistribution in vivo, perhaps reducing some detrimental effects of smoking.


Assuntos
Anticorpos/imunologia , Haptenos/química , Nitrosaminas/química , Nitrosaminas/síntese química , Nitrosaminas/imunologia , Anticorpos/sangue , Proteínas de Transporte/química , Proteínas de Transporte/imunologia , Estrutura Molecular
2.
J Clin Virol ; 35(1): 95-8, 2006 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-16137922

RESUMO

BACKGROUND: Infection with wild-type (wt) measles virus strains induces high antibody levels believed to provide life-long protection against disease. OBJECTIVES: Humoral immunity was followed up in convalescent measles patients to assess the persistence of specific antibodies after measles disease in individuals without and with documented re-exposure to wt virus. STUDY DESIGN: Paired sera were collected from 43 late convalescents (LC) before re-exposure and 3.7-4.8 years after re-exposure to at least one measles patient (LC+ group). Antibody persistence in this group was compared to paired sera from 43 age- and sex-matched controls without documented exposure to wt virus (LC- group). Paired sera were also obtained from 26 measles patients 1.3-1.7 and 3.8-4.1 years after they had recovered from measles to observe the waning of antibodies in early convalescents (EC group). RESULTS: Antibody levels decreased by 12.1% (CI: 3.2-20.3%, p=0.01) within 6.3 years in the LC- group of late convalescent measles patients. In contrast, in the LC+ group GMT of first and second sera were virtually identical, indicating that exposure to wt virus stabilizes antibody levels even in absence of a detectable secondary immune response. In a subset of late convalescents of group LC+ with a secondary immune response, antibody waning after re-exposure was as high as 15.6%/year (CI: 13.0-17.7%/year), corresponding to a half-life of 4.1 years (CI: 3.5-5.0 years), but antibodies were still higher than before re-exposure. In the EC group GMT decreased by 6.5% (95% CI: -13.3% to +0.1%) during 2.5 years but significance was low (p=0.08). CONCLUSION: The maintenance of antibody levels in convalescent measles patients is at least partially dependant on recurrent exposure to circulating wt virus.


Assuntos
Anticorpos Antivirais/sangue , Especificidade de Anticorpos , Convalescença , Vírus do Sarampo/patogenicidade , Sarampo/imunologia , Sarampo/virologia , Adolescente , Adulto , Idoso , Criança , Pré-Escolar , Feminino , Humanos , Imunoglobulina G/sangue , Lactente , Vírus do Sarampo/imunologia , Pessoa de Meia-Idade , Fatores de Tempo
3.
Vaccine ; 24(2): 185-94, 2006 Jan 12.
Artigo em Inglês | MEDLINE | ID: mdl-16122851

RESUMO

Epitope-based peptide antigens have been under development for protection against measles virus. The immunogenicity of five peptides composed of the same B cell epitope (BCE) (H236-250 of the measles virus hemagglutinin), and different T cell epitopes of measles virus fusion protein (F421-435, F256-270, F288-302) and nucleoprotein (NP335-345) was studied in mice (subcutaneous immunisation). The adjuvant effects of peptidoglycan monomer (PGM), Montanide ISA 720 and 206 were also investigated. Results showed basic differences in peptide immunogenicity that were consistent with already described structural differences. PGM elevated peptide-specific IgG when applied together with four of five tested peptides. A strong synergistic effect was observed after co-immunisation of mice with a mixture containing all five chimeric peptides in small and equal amounts. Results revealed for the first time that immunisation with several peptides having the common BCE generated significantly higher levels of both anti-peptide and anti-BCE IgG in comparison to those obtained after immunisation with a single peptide in much higher quantity. Further improvement of immune response was obtained after incorporation of such a peptide mixture into oil-based adjuvants.


Assuntos
Adjuvantes Imunológicos/farmacologia , Vírus do Sarampo/imunologia , Peptídeos/imunologia , Animais , Ensaio de Imunoadsorção Enzimática , Camundongos , Testes de Neutralização , Peptídeos/química
4.
Vaccine ; 23(17-18): 2074-7, 2005 Mar 18.
Artigo em Inglês | MEDLINE | ID: mdl-15755573

RESUMO

Chimeric molecules expressing multiple copies of the loop-forming hemagglutinin noose epitope (designated as "L"; aa386-400), a protective B cell epitope of the measles virus were generated by recombinant technology. The recombinant polyepitope [L4T4]2 combining two sets of four repeats of the L epitope and with two sets of four repeats of the human promiscuous T cell epitope of tetanus toxoid ("T", tt830-844) was produced in transgenic carrot plants. After intraperitoneal immunization of mice with plant membrane extract, sera neutralized all wild-type viruses. In a modified plaque reduction neutralization assay based on CD150-transfected Vero cells anti-[L4T4]2 sera neutralized all field isolates, irrespective of mutations in the L epitope. Even viruses with a mutation in the contact residues of a neutralizing L-specific monoclonal antibody or two mutations in other positions of the epitope were equally sensitive to neutralization. These results suggest that the multiple copies of the L epitope fold into different conformations that induce a repertoire of B cells diverse enough to overcome the genetic diversity of field viruses.


Assuntos
Anticorpos Antivirais/biossíntese , Vacina contra Sarampo/genética , Vacina contra Sarampo/imunologia , Vírus do Sarampo/genética , Vírus do Sarampo/imunologia , Sequência de Aminoácidos , Animais , Antígenos Virais/genética , Chlorocebus aethiops , Epitopos/genética , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Dados de Sequência Molecular , Mutação , Testes de Neutralização , Plantas Geneticamente Modificadas , Vacinas Sintéticas/genética , Vacinas Sintéticas/imunologia , Células Vero , Proteínas Virais/genética , Proteínas Virais/imunologia
5.
Carcinogenesis ; 26(4): 835-44, 2005 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-15637092

RESUMO

The influence of specific antibodies on molecular and cellular mechanisms of activation, detoxification and biological activity of the ubiquitous carcinogen benzo[a]pyrene (B[a]P) was investigated using a monoclonal antibody. The antibody was shown to decrease cellular uptake and metabolic activation of B[a]P as demonstrated by higher recovery of unmetabolized B[a]P and decreased formation of end-point phenol metabolites in two types of target cells. Furthermore, strong antibody reactivity with 7,8-diol-B[a]P provided a second chance for interrupting metabolic activation by sequestration of this intermediate metabolite in the extracellular space. The biological relevance of B[a]P and 7,8-diol-B[a]P redistribution by antibody was demonstrated by reversion of B[a]P-induced inhibition of proliferation of human peripheral blood lymphocytes and by inhibition of CYP 1A1 induction in HepG2 cells. Remarkably, the antibody was still protective against B[a]P-induced immunotoxicity even after delayed addition, suggesting a more important role of metabolites in immunotoxicity than has been appreciated so far. Although B[a]P is activated to 7,8-diol-B[a]P in the same cells that are inhibited by this metabolite, the antibody completely restored lymphocyte proliferation indicating that extracellular trapping of the 7,8-diol-B[a]P is biologically highly effective. Thus, repartitioning of both B[a]P and its metabolites by the antibody may reduce their effective concentration in susceptible target organs and therefore relieve overloaded DNA repair mechanisms and inhibit carcinogen-induced P450 induction. These in vitro data also suggest that a natural or prophylactic antibody response against carcinogens may be associated with a reduced risk of cancer.


Assuntos
Anticorpos Monoclonais/farmacologia , Benzo(a)pireno/metabolismo , Carcinógenos/efeitos adversos , Proliferação de Células/efeitos dos fármacos , Citocromo P-450 CYP1A1/antagonistas & inibidores , Di-Hidroxi-Di-Hidrobenzopirenos/efeitos adversos , Linfócitos/efeitos dos fármacos , Alquilação/efeitos dos fármacos , Animais , Carcinógenos/metabolismo , Carcinoma Hepatocelular/imunologia , Carcinoma Hepatocelular/metabolismo , Carcinoma Hepatocelular/patologia , Citocromo P-450 CYP1A1/imunologia , Citocromo P-450 CYP1A1/metabolismo , Adutos de DNA , Di-Hidroxi-Di-Hidrobenzopirenos/metabolismo , Feminino , Humanos , Neoplasias Hepáticas/imunologia , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/patologia , Microssomos Hepáticos/efeitos dos fármacos , Ratos , Ratos Wistar
6.
Int J Parasitol ; 33(5-6): 525-45, 2003 May.
Artigo em Inglês | MEDLINE | ID: mdl-12782053

RESUMO

Vaccination with the current live attenuated measles vaccine is one of the most successful and cost-effective medical interventions. However, as a result of persisting maternal antibodies and immaturity of the infant immune system, this vaccine is poorly immunogenic in children <9 months old. Immunity against the live vaccine is less robust than natural immunity and protection less durable. There may also be some concern about (vaccine) virus spread during the final stage of an eventual measles eradication program. Opinions may differ with respect to the potential threat that some of these concerns may be to the World Health Organisation goal of measles elimination, but there is a consensus that the development of new measles vaccines cannot wait. Candidate vaccines are based on viral or bacterial vectors expressing recombinant viral proteins, naked DNA, immune stimulating complexes or synthetic peptides mimicking neutralising epitopes. While some of these candidate vaccines have proven their efficacy in monkey studies, aerosol formulated live attenuated measles vaccine are evaluated in clinical trials.


Assuntos
Vacina contra Sarampo/uso terapêutico , Sarampo/prevenção & controle , Animais , Modelos Animais de Doenças , Epitopos/imunologia , Humanos , Sarampo/epidemiologia , Sarampo/imunologia , Vacina contra Sarampo/imunologia , Plantas/imunologia , Primatas/imunologia , Ratos , Proteínas Recombinantes/imunologia , Vacinação/métodos , Vacinas Atenuadas/imunologia , Vacinas Atenuadas/uso terapêutico , Vacinas Sintéticas/imunologia , Vacinas Sintéticas/uso terapêutico
7.
Vaccine ; 21(17-18): 2065-72, 2003 May 16.
Artigo em Inglês | MEDLINE | ID: mdl-12706696

RESUMO

Transgenic carrot plants were developed expressing a designer polyepitope combining tandem repeats of a protective loop-forming B cell epitope (H386-400) of the measles virus hemagglutinin protein with a human promiscuous, measles-unrelated T cell epitope (tt830-844). Despite the sensitivity of the loop conformation to its molecular environment, proper folding was confirmed by conformation-dependent monoclonal antibodies. The antibodies also reacted with the boiled antigen in Western blot. Immunisation of mice peritoneally with carrot plant extracts induced high titers of antibodies that crossreacted strongly with the virus. Furthermore, the sera neutralised field isolates of different geographic origins and genotypes in a modified plaque reduction neutralisation assay performed on CD150-transfected Vero cells. These results demonstrate that transgenic carrot plants can serve as an efficient expression system to produce highly immunogenic, randomly assembled polyepitope antigens. The combined features of the selected epitopes and the potential of the plant expression system may pave the way towards new vaccines against measles.


Assuntos
Epitopos/imunologia , Vacina contra Sarampo/imunologia , Sarampo/imunologia , Plantas Comestíveis/imunologia , Plantas Geneticamente Modificadas/imunologia , Sequência de Aminoácidos , Animais , Anticorpos Antivirais/sangue , Formação de Anticorpos , Antígenos Virais/imunologia , Daucus carota , Epitopos/química , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Dados de Sequência Molecular , Testes de Neutralização , Reação em Cadeia da Polimerase , Conformação Proteica , Dobramento de Proteína , Linfócitos T/imunologia
8.
Vaccine ; 21(7-8): 816-9, 2003 Jan 30.
Artigo em Inglês | MEDLINE | ID: mdl-12531367

RESUMO

Vaccine-induced immunity against measles is less robust than natural immunity. Waning of immunity in vaccines may eventually require a revaccination of adults. Measles antigens expressed in plants have been shown to be antigenic and immunogenic both after invasive and oral vaccination. Strategies for the vaccination of adults, the potential of an oral measles vaccine produced in edible plants and the design of suitable antigens are discussed.


Assuntos
Antígenos Virais/biossíntese , Vacina contra Sarampo/administração & dosagem , Plantas Geneticamente Modificadas/metabolismo , Administração Oral , Adulto , Animais , Antígenos Virais/genética , Humanos , Sarampo/prevenção & controle , Vacina contra Sarampo/imunologia , Plantas Geneticamente Modificadas/genética , Vacinas de Plantas Comestíveis/administração & dosagem , Vacinas de Plantas Comestíveis/imunologia
9.
Viral Immunol ; 15(3): 451-71, 2002.
Artigo em Inglês | MEDLINE | ID: mdl-12479395

RESUMO

Co-evolving mechanisms of immune clearance and of immune suppression are among the hallmarks of measles. B cells are major targets cells of measles virus (MV) infection. Virus interactions with B cells result both in immune suppression and a vigorous antibody response. Although antibodies fully protect against (re)infection, their importance during the disease and in the presence of a potent cellular response is less well understood. Specific serum IgM appears with onset of rash and confirms clinical diagnosis. After isotype switching, IgG1 develops and confers life-long protection. The most abundant antibodies are specific for the nucleoprotein, but neutralizing and protective antibodies are solely directed against the two surface glycoproteins, the hemagglutinin and the fusion protein. Major neutralizing epitopes have been mapped mainly on the hemagglutinin protein with monoclonal antibodies, producing an increasingly comprehensive map of functional domains.


Assuntos
Anticorpos Antivirais/imunologia , Linfócitos B/imunologia , Hemaglutininas Virais/imunologia , Vírus do Sarampo/imunologia , Sarampo/imunologia , Animais , Anticorpos Antivirais/biossíntese , Antígenos de Superfície/isolamento & purificação , Linfócitos B/virologia , Humanos , Imunoglobulina A , Imunoglobulina E , Imunoglobulina G
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