RESUMEN
Purified subunit viral antigens are weakly immunogenic and stimulate only the antibody but not the T cell-mediated immune response. An alternative approach to inducing protective immunity with small viral peptides may be the targeting of viral epitopes to immunocompetent cells by DNA and protein-engineered vaccines. This review will focus on DNA and protein-generated chimeric molecules carrying engineered fragments specific for activating cell surface co-receptors for inducing protective antiviral immunity. Adjuvanted protein-based vaccine or DNA constructs encoding simultaneously T- and B-cell peptide epitopes from influenza viral hemagglutinin, and scFvs specific for costimulatory immune cell receptors may induce a significant increase of anti-influenza antibody levels and strong CTL activity against virus-infected cells in a manner that mimics the natural infection. Here we summarize the development of several DNA and protein chimeric constructs carrying influenza virus HA317-41 fragment. The generated engineered molecules were used for immunization in intact murine and experimentally humanized NSG mouse models.
Asunto(s)
Vacunas contra la Influenza , Gripe Humana , Orthomyxoviridae , Animales , Ratones , Humanos , Gripe Humana/prevención & control , Vacunas contra la Influenza/genética , Epítopos de Linfocito B , ADN , Orthomyxoviridae/genéticaRESUMEN
Antiviral DNA vaccines are a novel strategy in the vaccine development field, which basically consists of the administration of expression vectors coding viral antigen sequences into the host's cells. Targeting of conserved viral epitopes by antibody fragments specific to activating cell surface co-receptor molecules on antigen-presenting cells could be an alternative approach for inducing protective immunity. It has been shown that FcγRI on human monocytes enhances antigen presentation in vivo. Various DNA constructs, encoding a Single-chain variable antibodies (scFv) from mouse anti-human FcγRI monoclonal antibody, coupled to a sequence encoding a T- and B-cell epitope-containing influenza A virus hemagglutinin inter-subunit peptide were inserted into the eukaryotic expression vector system pTriEx-3 Neo. The constructed chimeric DNA molecules were expressed by transfected Chinese hamster ovary cells and the ability of the engineered proteins to interact with FcγRI-expressing cells was confirmed by flow cytometry. The fusion protein induced a strong signal transduction on human monocytes via FcγRI. The expression vector pTriEx-3 Neo containing the described construct was used as a naked DNA vaccine and introduced directly to experimental humanized NOD SCID gamma mice with or without boosting with the expressed fusion protein. Immunization with the generated DNA chimeric molecules and prime-boost with the expressed recombinant proteins induced significant serum levels of anti-influenza immunoglobulin G antibodies and strong cytotoxic T lymphocyte activity against influenza virus-infected cells in humanized animals.
Asunto(s)
Anticuerpos Monoclonales/inmunología , Epítopos/inmunología , Vacunas contra la Influenza/inmunología , Gripe Humana/inmunología , Vacunas de ADN/inmunología , Animales , Anticuerpos Monoclonales/biosíntesis , Anticuerpos Monoclonales Humanizados/genética , Anticuerpos Monoclonales Humanizados/inmunología , Células Presentadoras de Antígenos/inmunología , Células CHO , Cricetulus , Epítopos/biosíntesis , Citometría de Flujo , Regulación de la Expresión Génica , Ingeniería Genética , Humanos , Fragmentos de Inmunoglobulinas/genética , Fragmentos de Inmunoglobulinas/inmunología , Gripe Humana/prevención & control , Gripe Humana/virología , Ratones , Orthomyxoviridae/inmunología , Orthomyxoviridae/patogenicidad , Linfocitos T Citotóxicos/inmunologíaRESUMEN
BACKGROUND: Recurrent respiratory papillomatosis (RRP) is a rare manifestation of human papilloma virus (HPV) infection with extremely high relapse frequency, poorly understood immunopathogenesis, and lack of efficient treatment. Immunotherapy with Calgevax (BCG) in combination with CO2 surgery significantly improves the outcome of RRP. The present study investigates cellular immunity parameters in RRP patients, and the effects of 20-month Calgevax immunomodulation. MATERIALS AND METHODS: RRP patients (n = 15) subjected to combined therapy were tested before, 6, 12 and 20 months after the start of immunomodulation. Absolute counts and percentage of T, B and NK cells, effector T1 (CD8 + IFNgamma+); Th1 (CD4+IFNgamma+), Th17 (CD4+IL-17+) and regulatory (CD4+FoxP3+) T lymphocytes, as well as the in vitro stimulated secretion of IL-2, IL-4, IL-5, IL-10, IFNgamma and TNFalpha were determined by flow cytometry (FACSCanto II, BD). RESULTS: While no significant changes were detected in the circulating T, B and NK subsets, RRP patients presented increased proportions of Tc1, Th1 and Th17 cells, and significantly reduced IFNgamma/IL-4 and IFNgamma/IL-10 ratios as compared to healthy controls (15% vs. 8%), (58 vs. 139 and 15 vs. 26, respectively), p < 0.05 for all comparisons. Increased Treg (9% vs. 4%), and decreased Th17 effectors share (0.7% vs. 0.4%) were observed at 12 months, while IFNgamma/IL-4 and IFNgamma/IL-10 ratios were restored after 20 months of Calgevax application. CONCLUSIONS: Antiviral response closely depends on cytokine background. Calgevax potentiates Treg differentiation at the expense of proinflammatory Th17, limits hyperactivation and virus-specific T cell clones depletion, and restores a Th1 cytokine background.
Asunto(s)
Vacuna BCG/uso terapéutico , Factores Inmunológicos/uso terapéutico , Infecciones por Papillomavirus/tratamiento farmacológico , Infecciones del Sistema Respiratorio/tratamiento farmacológico , Adulto , Femenino , Humanos , Interferón gamma/biosíntesis , Interleucina-10/biosíntesis , Interleucina-4/biosíntesis , Masculino , Infecciones por Papillomavirus/inmunología , Infecciones del Sistema Respiratorio/inmunología , Células Th17/inmunologíaRESUMEN
We investigated the circulating cytotoxic CD160+ CD8(high) subset in correlation to antiviral immunity and response to highly active antiretroviral therapy (HAART) in HIV+ subjects. The study included 45 treatment-naive patients receiving HAART for 18 months, retrospectively defined as good (n=29) and transient (n=16) responders. HIV-specific CD8 T lymphocyte levels were measured by IFNgamma production in response to p17 Gag, in the presence of immobilized anti-CD160 mAb. We report a significantly increased baseline level of CD160+ CD8(high) subset in good therapy responders. CD160+ CD8(high) subset correlates with CD4+ T cell count, immune activation, and viral load. CD160+ CD8(high) lymphocytes contain a high amount of Granzyme B and include virus-specific T lymphocytes in HIV-1+ subjects. Co-stimulation through CD160 molecules enhances IFNgamma production in response to p17 Gag. Therefore, the CD160+ CD8(high) subset may be useful for monitoring of virus-specific cellular immunity and predicting response to antiretroviral therapy in chronic HIV-1 infection.