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Epitope Peptide (epitope + peptide)
Selected AbstractsHigh epitope density in a single protein molecule significantly enhances antigenicity as well as immunogenicity: a novel strategy for modern vaccine development and a preliminary investigation about B,cell discrimination of monomeric proteinsEUROPEAN JOURNAL OF IMMUNOLOGY, Issue 2 2005Wanli Liu Abstract Although early studies have shown a close correlation between epitope density and epitope-specific humoral immune responses, few attempts have been made to quantitatively compare the antigenic and immunogenic differences between protein molecules bearing low or high degrees of epitope density, nor have studies quantitatively investigated the mechanism of B,cell discrimination of monomeric antigens. In this study, we prepared glutathione S-transferase (GST) fusion proteins bearing various copies of the M2e epitope from the influenza virus M2,protein [GST-(M2e)8, GST-(M2e)4 and GST-(M2e)1], which were used to detect and compare the real-time kinetic binding with M2e-specific mAb by surface plasma resonance. Our data show clearly that fusion proteins bearing higher M2e epitope density resulted in higher average avidity for M2e-specific mAb. Furthermore, it was observed that fusion proteins bearing high M2e epitope density could induce polyclonal antibodies (pAb) with enhanced an average affinity constant (KA) for M2e epitope peptide compared to fusion proteins bearing low epitope density. The average KA of pAb induced by GST-(M2e)8 (3.08 × 108,M,1 or 9.96 × 108,M,1) was up to two orders of magnitude greater than the average KA of pAb induced by GST-(M2e)1 (2.00 × 106,M,1 or 3.43 × 106,M,1). Thus, the data presented here demonstrate that high epitope density in a single protein molecule significantly enhances antigenicity and immunogenicity. These findings enrich our knowledge of how epitope density might relate to the recognition, activation and antibody production processes of epitope-specific immature B,cells. [source] ,-Galactosylceramide-loaded, antigen-expressing B cells prime a wide spectrum of antitumor immunityINTERNATIONAL JOURNAL OF CANCER, Issue 12 2008Yeon-Jeong Kim Abstract Most of the current tumor vaccines successfully elicit strong protection against tumor but offer little therapeutic effect against existing tumors, highlighting the need for a more effective vaccine strategy. Vaccination with tumor antigen-presenting cells can induce antitumor immune responses. We have previously shown that NKT-licensed B cells prime cytotoxic T lymphocytes (CTLs) with epitope peptide and generate prophylactic/therapeutic antitumor effects. To extend our B cell vaccine approach to the whole antigen, and to overcome the MHC restriction, we used a nonreplicating adenovirus to transduce B cells with antigenic gene. Primary B cells transduced with an adenovirus-encoding truncated Her-2/neu (AdHM) efficiently expressed Her-2/neu. Compared with the moderate antitumor activity induced by vaccination with adenovirus-transduced B cells (B/AdHM), vaccination with ,-galactosylceramide-loaded B/AdHM (B/AdHM/,GalCer) induced significantly stronger antitumor immunity, especially in the tumor-bearing mice. The depletion study showed that CD4+, CD8+ and NK cells were all necessary for the therapeutic immunity. Confirming the results of the depletion study, B/AdHM/,GalCer vaccination induced cytotoxic NK cell responses but B/AdHM did not. Vaccination with B/AdHM/,GalCer generated Her-2/neu -specific antibodies more efficiently than B/AdHM immunization. More importantly, B/AdHM/,GalCer could prime Her-2/neu -specific cytotoxic T cells more efficiently and durably than B/AdHM. CD4+ cells appeared to be necessary for the induction of antibody and CTL responses. Our results demonstrate that, with the help of NKT cells, antigen-transduced B cells efficiently induce innate immunity as well as a wide range of adaptive immunity against the tumor, suggesting that they could be used to develop a novel cellular vaccine. © 2008 Wiley-Liss, Inc. [source] Epitope mapping of the neuronal growth inhibitor Nogo-A for the Nogo receptor and the cognate monoclonal antibody IN-1 by means of the SPOT techniqueJOURNAL OF MOLECULAR RECOGNITION, Issue 3 2007Hilke Zander Abstract Nogo-A is a potent inhibitor of axonal outgrowth in the central nervous system of adult mammals, where it is expressed as a membrane protein on oligodendrocytes and in myelin. Here we describe an attempt to identify linear peptide epitopes in its sequence that are responsible for the interaction either with the Nogo receptor (NgR) or with the neutralizing monoclonal antibody IN-1. Analysis of an array of immobilized overlapping 15,mer peptides covering the entire amino acid sequence of human Nogo-A (1192 residues) revealed a single epitope with prominent binding activity both towards the recombinant NgR and the IN-1 Fab fragment. Further truncation and substitution analysis yielded the minimal epitope sequence 'IKxLRRL' (x,,,P), which occurs within the so-called Nogo66 region (residues 1054,1120) of Nogo-A. The bacterially produced Nogo66 fragment exhibited binding activity both for the recombinant NgR and for the IN-1 Fab fragment on the Western blot as well as in ELISA. Unexpectedly, the synthetic epitope peptide and the recombinant Nogo66 showed cross-reactivity with the 8-18C5 Fab fragment, which is directed against myelin oligodendrocyte glycoprotein (MOG) as a structurally unrelated target. On the other hand, the recombinant N-terminal domain of Nogo-A (residues 334,966) was shown to specifically interact on the Western blot and in an ELISA with the IN-1 Fab fragment but not with the recombinant NgR, which is in agreement with previous results. Hence, our data suggest that there is a distinct binding site for the Nogo receptor in the Nogo66 region of Nogo-A, whereas its interaction with NgR is less specific than anticipated before. Although there probably exists a non-linear epitope for the neutralizing antibody IN-1 in the N-terminal region of Nogo-A, which is likely to be accessible from outside the cell, a previously postulated second binding site for NgR in this region (called Nogo-A-24) remains elusive. Copyright © 2007 John Wiley & Sons, Ltd. [source] Peptide based vaccine design: Synthesis and immunological characterization of branched polypeptide conjugates comprising the 276,284 immunodominant epitope of HSV-1 glycoprotein DJOURNAL OF PEPTIDE SCIENCE, Issue 3 2002Gábor Mez Abstract The importance of the length and conjugation site of a protective epitope peptide (276SALLEDPVG284) from glycoprotein D of herpes simplex virus in branched polypeptide conjugates has been investigated. A new set of peptides, with a single attachment site and truncated sequences, was prepared. The immunogenicity of conjugates and the specificity of antibody responses elicited were investigated in BALB/c, C57/Bl/6 and CBA mice. It was found that the covalent coupling of the peptide comprising the 276,284 sequence of gD through its Asp residue at position 281 did not influence the immunogenic properties of the epitope, while involvement of the side chain of Glu at position 280 almost completely abolished immunogenicity. These results clearly indicated that the conjugation site of the epitope peptide influenced the intensity and specificity of antibody responses. Comparison of the immunological properties of conjugates containing truncated gD peptides revealed the presence of two epitopes within the 276,284 region. One of the proposed epitopes is situated at the N -terminal (276,281) region, while the other is located at the C -terminal end of the sequence (279,284). Binding data demonstrated that some of the peptides comprising these epitopes induced gD-specific responses in their conjugated form and also elicited an immune response that conferred protection against lethal HSV-1 infection. The correlation of peptide- and gD-specific antibody responses with the protective effect of the immune response is discussed. Copyright © 2002 European Peptide Society and John Wiley & Sons, Ltd. [source] Phase I clinical trial using peptide vaccine for human vascular endothelial growth factor receptor 2 in combination with gemcitabine for patients with advanced pancreatic cancerCANCER SCIENCE, Issue 2 2010Motoki Miyazawa Vascular endothelial growth factor receptor 2 (VEGFR2) is an essential factor in tumor angiogenesis and in the growth of pancreatic cancer. Immunotherapy using epitope peptide for VEGFR2 (VEGFR2-169) that we identified previously is expected to improve the clinical outcome. Therefore, a phase I clinical trial combining of VEGFR2-169 with gemcitabine was conducted for patients with advanced pancreatic cancer. Patients with metastatic and unresectable pancreatic cancer were eligible for the trial. Gemcitabine was administered at a dose of 1000 mg/m2 on days 1, 8, and 15 in a 28-day cycle. The VEGFR2-169 peptide was subcutaneously injected weekly in a dose-escalation manner (doses of 0.5, 1, and 2 mg/body, six patients/one cohort). Safety and immunological parameters were assessed. No severe adverse effect of grade 4 or higher was observed. Of the 18 patients who completed at least one course of the treatment, 15 (83%) developed immunological reactions at the injection sites. Specific cytotoxic T lymphocytes (CTL) reacting to the VEGFR2-169 peptide were induced in 11 (61%) of the 18 patients. The disease control rate was 67%, and the median overall survival time was 8.7 months. This combination therapy for pancreatic cancer patients was tolerable at all doses. Peptide-specific CTL could be induced by the VEGFR2-169 peptide vaccine at a high rate, even in combination with gemcitabine. From an immunological point of view, the optimal dose for further clinical trials might be 2 mg/body or higher. This trial was registered with ClinicalTrial.gov (no. NCT 00622622). (Cancer Sci 2009) [source] Induction of a Melanoma-Specific Antibody Response by a Monovalent, but not a Divalent, Synthetic GM2 NeoglycopeptideCHEMMEDCHEM, Issue 4 2009S. Bay Dr. Abstract Human tumor cell-specific antibodies were induced in mice after immunization with a synthetic glycopeptide, which is based on the GM2 ganglioside carbohydrate moiety produced on a gram scale in bacteria. Such neoglycopeptides represent a promising cancer vaccine strategy for active immunotherapy targeting carbohydrates. The GM2 ganglioside represents an important target for specific anticancer immunotherapy. We designed and synthesized a neoglycopeptide immunogen displaying one or two copies of the GM2 tetrasaccharidic moiety. These glycopeptides were prepared using the Huisgen cycloaddition, which enables the efficient ligation of the alkyne-functionalized biosynthesized GM2 with an azido CD4+ T,cell epitope peptide. It is worth noting that the GM2 can be produced on a gram scale in bacteria, which can be advantageous for a scale-up of the process. We show here for the first time that a fully synthetic glycopeptide, which is based on a ganglioside carbohydrate moiety, can induce human tumor cell-specific antibodies after immunization in mice. Interestingly, the monovalent, but not the divalent, form of GM2 peptide construct induced antimelanoma antibodies. Unlike traditional vaccines, this vaccine is a pure chemically-defined entity, a key quality for consistent studies and safe clinical evaluation. Therefore, such carbohydrate,peptide conjugate represents a promising cancer vaccine strategy for active immunotherapy targeting gangliosides. [source] Identification of SPARC as a candidate target antigen for immunotherapy of various cancersINTERNATIONAL JOURNAL OF CANCER, Issue 6 2010Mitsuhiro Inoue Abstract To establish efficient anticancer immunotherary, it is important to identify tumor-associated antigens (TAAs) directing the immune system to attack cancer. A genome-wide cDNA microarray analysis identified that secreted protein acidic and rich in cysteine (SPARC) gene is overexpressed in the gastric, pancreatic and colorectal cancer tissues but not in their noncancerous counterparts. This study attempted to identify HLA-A24 (A*2402)-restricted and SPARC-derived CTL epitopes. We previously identified H-2Kd -restricted and SPARC-derived CTL epitope peptides in BALB/c mice, of which H-2Kd -binding peptide motif is comparable with that of HLA-A24 binding peptides. By using these peptides, we tried to induce HLA-A24 (A*2402)-restricted and SPARC-reactive human CTLs and demonstrated an antitumor immune response. The SPARC-A24-1143,151 (DYIGPCKYI) and SPARC-A24-4225,234 (MYIFPVHWQF) peptides-reactive CTLs were successfully induced from peripheral blood mononuclear cells by in vitro stimulation with these two peptides in HLA-A24 (A*2402) positive healthy donors and cancer patients, and these CTLs exhibited cytotoxicity specific to cancer cells expressing both SPARC and HLA-A24 (A*2402). Furthermore, the adoptive transfer of the SPARC-specific CTLs could inhibit the tumor growth in nonobese diabetic/severe combined immunodeficient mice bearing human cancer cells expressing both HLA-A24 (A*2402) and SPARC. These findings suggest that SPARC is a potentially useful target candidate for cancer immunotherapy. [source] Identification of a dengue virus-specific HLA-A*0201-restricted CD8+ T cell epitopeJOURNAL OF MEDICAL VIROLOGY, Issue 4 2010Jinsheng Wen Abstract In this study, a combination of epitope-prediction programs and in vitro assays was used to identify dengue virus (DENV)-specific CD8+ T cell epitopes. Peripheral blood mononuclear cells (PBMCs) isolated from patients who recovered from dengue fever were stimulated with candidate epitope peptides derived from DENV, which were predicted by using SYFPEITHI and RANKpep epitope-prediction programs. The IFN-, ELISpot results and the results of intracellular staining of IFN-, showed that peptides NS4b_40 (TLYAVATTI), E_256 (QEGAMHTAL), NS3_205 (LPAIVREAI), NS5_210 (SRNSTHEMY), and NS3_207 (AIVREAIKR) could induce the recall response of CD8+ T cells. Furthermore, the results of the MHC,peptide complex stabilization assay revealed that peptide NS4b_40 (TLYAVATTI) has a high affinity for HLA-A*0201 molecules. The IFN-, ELISpot results and staining of intracellular IFN-, confirmed that this peptide could induce high-level CD8+ T cell response in HLA-A*0201 positive PBMCs. Peptide NS4b_40 (TLYAVATTI) was identified as a novel DENV-specific HLA-A*0201-restricted CD8+ T cell epitope. J. Med. Virol. 82:642,648, 2010. © 2010 Wiley-Liss, Inc. [source] Mass spectrometric and chemical stability of the Asp-Pro bond in herpes simplex virus epitope peptides compared with X-Pro bonds of related sequencesJOURNAL OF PEPTIDE SCIENCE, Issue 8 2002Zsolt Skribanek Abstract The mass spectrometric analysis of the immunodominant epitope region (273,284) of herpes simplex virus type 1 (HSV-1) glycoprotein D (gD) showed a favoured fission at the Asp-Pro peptide bond. The fast atom bombardment collision induced dissociation (FAB-CID) study of closely related X-Pro peptides documented that neither the length nor the amino acid composition of the peptide has a significant influence on this preferential cleavage. At the same time the DP bond proved to be sensitive to acidic conditions in the course of peptide synthesis. These observations prompted us to compare the chemical and mass spectrometric stability of a new set of nonapeptides related to the 273,284 epitope region of gD, i.e. SALLEDPVG and SALLEXPVG peptides, where X = A, K, I, S, F, E or D, respectively. The chemical stability of these peptides during acidic hydrolysis was investigated by electrospray ionization mass spectrometry (ESI-MS) and the products were identified by ESI-MS and on-line high performance liquid chromatography,mass spectrometry (HPLC-MS). The mass spectrometric fragmentation and bond stability of the untreated peptide samples were also studied using ESI-MS and liquid secondary ion mass spectrometry (LSIMS). Both the chemical hydrolysis and the mass spectrometric fragmentation showed that the Asp-Pro bond could easily be cleaved, while the KP bond proved to be stable under both circumstances. On the other hand, the XP bond (X = A, I, S, F or E) fragmented easily under the mass spectrometric conditions, but was not sensitive to the acidolysis. Copyright © 2002 European Peptide Society and John Wiley & Sons, Ltd. [source] Expression, refolding, crystallization and preliminary crystallographic study of MHC H-2Kk complexed with octapeptides and nonapeptidesACTA CRYSTALLOGRAPHICA SECTION D, Issue 7 2004Christine Kellenberger Major histocompatibility complex (MHC) molecules are heterodimeric cell-surface receptors that play a crucial role in the cellular immune response by presenting epitope peptides to T-cell antigen receptors (TCR). Although the structural basis of the peptide,MHC binding mechanism is becoming better understood, it is still difficult to predict a binding mode for an MHC of unknown structure. Therefore, as the first stage of a TCR,MHC interaction study, the crystal structures of the mouse H-2Kk molecule in complex with both an octapeptide from Influenza A virus and a nonapeptide from simian virus SV40 were solved. Here, the expression, refolding, purification and crystallization of the two complexes are reported. For the H-2Kk,HA(259,266) complex, crystals were obtained via an extensive screen using a nanodrop-dispensing robot and diffracted to 2.5,Å resolution. For the H-2Kk,SV40(560,568) complex, microscopic needles were initially obtained and their size was improved by macroseeding and a stepwise increase in precipitant concentration. Diffraction data to a resolution of 3.0,Å were collected at a synchrotron facility. [source] | |||||||||||||