ABSTRACT
Tryptophan 2,3-dioxygnease 2 (TDO2) is specific for metabolizing tryptophan to kynurenine (KYN), which plays a critical role in mediating immune escape of cancer. Although accumulating evidence demonstrates that TDO2 overexpression is implicated in the development and progression of multiple cancers, its tumor-promoting role in esophageal squamous cell carcinoma (ESCC) remains unclear. Here, we observed that TDO2 was overexpressed in ESCC tissues and correlated significantly with lymph node metastasis, advanced clinical stage, and unfavorable prognosis. Functional experiments showed that TDO2 promoted tumor cell proliferation, migration, and colony formation, which could be prevented by inhibition of TDO2 and aryl hydrocarbon receptor (AHR). Further experimentation demonstrated that TDO2 could promote the tumor growth of KYSE150 tumor-bearing model, tumor burden of C57BL/6 mice with ESCC induced by 4-NQO, enhance the expression of phosphorylated AKT, with subsequent phosphorylation of GSK3
ABSTRACT
PD-1 and CTLA-4 antibodies offer great hope for cancer immunotherapy. However, many patients are incapable of responding to PD-1 and CTLA-4 blockade and show low response rates due to insufficient immune activation. The combination of checkpoint blockers has been proposed to increase the response rates. Besides, antibody drugs have disadvantages such as inclined to cause immune-related adverse events and infiltration problems. In this study, we developed a cyclic peptide C25 by using Ph.D.-C7C phage display technology targeting LAG-3. As a result, C25 showed a relative high affinity with human LAG-3 protein and could effectively interfere the binding between LAG-3 and HLA-DR (MHC-II). Additionally, C25 could significantly stimulate CD8 T cell activation in human PBMCs. The results also demonstrated that C25 could inhibit tumor growth of CT26, B16 and B16-OVA bearing mice, and the infiltration of CD8 T cells was significantly increased while FOXP3 Tregs significantly decreased in the tumor site. Furthermore, the secretion of IFN- by CD8 T cells in spleen, draining lymph nodes and especially in the tumors was promoted. Simultaneously, we exploited T cells depletion models to study the anti-tumor mechanisms for C25 peptide, and the results combined with MTT assay confirmed that C25 exerted anti-tumor effects CD8 T cells but not direct killing. In conclusion, cyclic peptide C25 provides a rationale for targeting the immune checkpoint, by blockade of LAG-3/HLA-DR interaction in order to enhance anti-tumor immunity, and C25 may provide an alternative for cancer immunotherapy besides antibody drugs.
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Background and purpose:Cytotoxic T lymphocyte (CTL) plays a vital role in the process of anti-tumor immunology. The aim of this study was to investigate whether changes in concentration of IL-2 (50, 200 and 1 000 U/mL) would affect the sub-population and cytotoxic function of cells cultivated by peptide-specific CTL induction systemin vitro and also observe whether using the concentration of IL-2 at a range of 50-1 000 U/mL isbeneifcial to regulatory cells (Tregs) enrichment.Methods:Peripheral blood from 10 healthy donors and 10 cancer patients that were HLA-A2 positive, were collected in the study. HLA-A2 restricted CTL epitope P321 (ILIGETIKI) derived from COX-2 pulsed with different concentrations of IL-2 were used to induce peptides-speciifc CTLin vitro. Flow cytometry was performed to analyze the proliferative capability, the proportion of different T-cell subsets, and secretion of perforin, granzyme B and IFN-γ. IFN-γ secretion was assessed by ELISpot assay.Results:High concentration of IL-2 increased the proliferative activity. The percentage of CD4+ T cells of cancer patient group was signiifcantly higher than that of healthy donor group, while the percentage of CD8+ T cells of cancer patient group was signiifcantly lower than that of healthy donor group. And there was no signiifcant difference in the percentages of CD4+ T cells, CD8+ T cells and Tregs among groups with different IL-2 concentrations. No difference was seen in cytokine (perforin, granzyme B, IFN-γ) secretion capacity of CD8+ T cells. ELISpot study revealed that high-dose IL-2 resulted in the increasing of IFN-γ secretion.Conclusion:The sub-population and the function of cells cultured by peptide-speciifc CTL induction systemin vitro are not affected by different concentrations of IL-2. Furthermore, high concentrations of IL-2 (50-1 000 U/mL) do not provide the enrichment for Tregs. Higher concentration of IL-2 is likely to cause high secretion of IFN-γ in ELISpot assay. In order to exclude the distraction of NK cells or NKT cells, the concentration of 50 U/mL is better choice.
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Objective To investigate the antitumor effect of specific cytotoxic T lymphocytes induced by renal cell carcinoma multiple antigen peptide dendritic cell (DC) vaccine in vitro. Methods Dendritic cells were induced by hGM-CSF,hIL-4 from blood.Peptide of renal cell carcinoma cell line (RCC786-0)was got by citrate-phosphate buffer elution.Peripheral blood mononuclear cell was cultured.Multiple antigen peptide DC cell vaccine was obtained by acid-eluted peptide pulsed DC.The tumor antigen specific CTL was generated from activated T cell by vaccine.Killing activity of the tumor antigen specific CTL was activated by vaccine. Results Antigen peptide DC cell vaccine could show a strong cytotoxic activity of CTLs(31.93±5.05%),which was much higher than control groups(5.88±2.26%,8.03±6.70%,9.70±2.09%,9.35±3.58%). Conclusion Renal cell carcinoma antigen peptide DC vaccine could show a high antitumor effect in vitro.
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The alkaline elution and fluorometrie DNA assay were adapted to the evaluation of DNA single-strand breaks in the cultured human fetal lung 2BS cell. the DNA damage was induced in vitro by the treatment with Alternariol monomethyl ether (AME) and Alternariol (AOH) which were the main mycotoxins of Alternariol alternata. The results showed that both AME and AOH could induce the DNA single-strand breaks in the cultured human fetal lung 2BS cells and there existed the dose-response association betweenthe fractions of DNA remaining on the filters and the doses. The fractions of DNA remaining on the filters were 0.57?0.04, 0.45?0.02, 0.30?0.02, 0.18?0.01 respetively under the AME concentration of 10, 25, 50, 100?g/ml and were 0.68?0.3, 0.54?0.01, 0.47?0.03, 0.34?0.01 respetively under the AOH concentration of 0.1, 1, 5, 10?g/ml when DNA was eluted for 6hr. It were significantly different from the fractions by solvent control (0.87?0.02) (P