Immunogenicity of the immunodominant cytotoxic T lymphocyte epitope E749-57 in HPV16 oncoprotein E7 chaperoned by HSP110 / 中华皮肤科杂志

Objective To investigate the immunogenicity of immunodominant cytotoxic T lymphocyte epitope E749-57 of human papilloma virus (HPV) 16 oncoprotein E7 chaperoned by heat shock protein (HSP)110. Methods Mouse HSP110 gene was cloned into prokaryotic expression vector pQE-80L for the expression of HSP110 protein, which was purified using Ni-NTA column. SDS-PAGE and Western-blot were conducted to confirm the purified mHSP110 protein, which was subsequently incubated with E749-57 peptide under heat shock condition, and high-performance liquid chromatography (HPLC) was used to evaluate the binding efficiency of the recombinant protein and E749-57 peptide. Twenty mice were divided into 4 groups to be immunized with mHSP110 protein, E749-57 peptide, mHSP110-E749-57 complex and phosphate buffered saline (PBS),respectively. Two weeks after the last immunization, spleen cells were collected from the immunized mice and divided into 2 parts: one were stimulated by E749-57 peptide followed by the detection of CD8+ INF-γ+ T cells with flow cytometry; the other one were subjected to MTT analysis for the estimation of cell proliferation. The mHSP110-E749-57 complex was also used to immunize TC-1 tumor bearing mice to observe its anti-tumor effect.Results The full-length 2577 bp-sized mHSP110 gene was amplified from mouse liver cDNA and cloned into pQE-80L vector. Direct sequencing confirmed the correctness of the cloning. SDS-PAGE and Western-blot demonstrated the successful purification of mHSP110. HPLC assay showed that the purified mHSP110 protein could bind with E749-57 to form a relatively stable protein complex. The percentage of IFN-γ+ CD8+ T cells in and proliferation index of spleen cells from the complex-immunized mice were statistically higher than those from the other 3 groups of mice. Moreover, the complex could obviously inhibit the growth of TC-1 tumor in mice. Conclusion The mHSP110-E749-57 complex could enhance the generation of specific cytotoxic T lymphocytes and exert anti-tumor effects in mice.

Therapeutic peptide based on CTL epitope of human melanoma antigen MART-1 can induce CD8~+ T cell response in vitro / 第三军医大学学报

Objective To explore how to improve the immunogenicity of short epitope peptides of triggering melanoma MART 1 specific CD8 +T cell responses Methods Therapeutic peptides based on the immunodominant MART 127 35, HIV Tat49 57CCP sequence and a tetanus toxoid universal Th epitope were designed and synthesized The immunological functions were studied in PBMCs from HLA A2 + melanoma patients Results The results demonstrated that the peptides could trigger vigorous MART 1 specific CD8 + CTL activities in vitro The function of peptide containing MART 127 35 and tetanus universal Th epitope was more vigorous than that of MART 127 35 peptide, and the immunogenicty of the peptides with HIV Tat49 57CCP sequence, MART 127 35 and tetanus universal Th epitope was the most vigorous Conclusion Linkage of HIV Tat49 57CCP sequence and a tetanus universal Th epitope could dramatically improve the immunogenictiy of the MART 127 35 epitope peptide

Polypeptides based on CTL, T helper and B-epitopes of HBV antigens induce HLA-A2~+ human PBMCs to produce CD8~+ CTL-mediated cytotoxicity / 第三军医大学学报

Objective To explore how to trigger an HLA Ⅰ restricted CD8 + T cell response to exogenously synthesized peptides in vitro . Methods A new panel of therapeutic peptides based on the immunodominant B and CTL epitopes of HBV PreS 2 region and HBcAg and the tetanus toxoid common T helper epitopes were synthesized by Merrifield solid phase peptide synthesis, and HLA A2 + human PBMCs were used to investigate the immunological properties of the mimetic peptides. Results The results demonstrated that the peptides could trigger vigorous CD8 + HBV specific CTL responses in vitro specifically and effectively. Conclusion The results reveal that T helper plus B epitopes designing with the introduction of short and flexible linker can remarkably improve the immunogenicity of short peptides and hence produce effective CTL responses in vitro .

Finding and identification of a novel HLA-A2-restricted CTL epitope derived fromthe tumor antigen MAGE-A3 / 中国免疫学杂志

To identify novel HLA-A2-restricted CTL epitopes derived from the tumor antigen MAGE-A3.Methods:The CTL epitope candidates were predicted by using the motif prediction method combined with the secondary anchor residues plot. It was established that the molecule model of CIL epitope candidates bound to the HLA-A2 molecule and of the HLA-A2-peptide complex by molecule modeling. Four selected candidates were assayed by the standard 51 Cr release assay to determine their abilities of inducing the generation of specific CTLs. Results: Among them, the pepnde MAGE-A3201-209 could effectively induce specific Oils and the CITs could lyse not only the melanoma cell line LB373-MEL but also the murine mastocytoma cell line genetically modified with the human HLA-A2 gene and pulsed with MAGE-A32oi.2cs ? Conclusion; MAGE-AS201-209 is a novel CTL epitope presented by HLA-A2. This study provides experimental basis for design and study of tumor therapeutic peptide vaccines based on the tumor antigen MAGE-A3.