Induction of anti-tumour immunity by dendritic cells transduced with hTERT recombinant adenovirus in mice.

Dendritic cells (DCs) transfected with recombinant, replication-defective adenovirus (Ad) vectors encoding the human telomerase reverse transcriptase (hTERT) are potent inducers of cytotoxic T lymphocytes (CTLs) and anti-tumour immunity. However, previous studies have mostly been in vitro. In this study, we sought to determine whether DCs transfected with hTERT (DC/Ad-hTERT) could elicit a potent anti-tumour immunogenic response in vivo. We found that murine DCs transfected with recombinant adenovirus encoding the hTERT gene (DC/Ad-hTERT) induced hTERT-specific CTLs in vivo effectively, compared with Ad-LacZ-transduced DC (DC/Ad-LacZ) controls. These hTERT-specific CTLs lysed various tumour cell lines in an hTERT-specific and MHC-I molecule-restricted fashion. We also found that DC/Ad-hTERT could increase antigen-specific T-cell proliferation and augment the number of IFN-gamma secreting T-cells in mice. These data suggest that the DC/Ad-hTERT vaccine may induce anti-tumour immunity against tumour cells expressing hTERT in an MHC-I molecule-restricted fashion in vivo through the augmentation of the hTERT-specific CTL response. The DC/Ad-hTERT vaccine may thus be used as an efficient DC-based tumour vaccine in clinical applications.

Dendritic cells reconstituted with a human heparanase gene induce potent cytotoxic T-cell responses against gastric tumor cells in vitro.

BACKGROUND AND AIMS: Dendritic cell-based tumor vaccination is a promising approach in the treatment of cancer. Strategies to modify dendritic cells (DCs) with tumor-associated antigens (TAAs) can elicit specific immune responses against tumors. Heparanase is overexpressed in gastric cancer, especially in invasive and metastatic cells, but is downregulated in differential normal tissue. Therefore, heparanase is a potential target in immunotherapy for patients with advanced gastric cancer who are not candidates for surgery. The present paper was designed to investigate the immune response of a heparanase gene-modified DC-based vaccine against gastric cancer cell lines in vitro. METHODS: DCs from peripheral blood mononuclear cells of healthy HLA-A2-positive donors were transfected with recombinant adenovirus containing the full-length cDNA of heparanase (rAd-Hpa) to generate heparanase gene-modified DC vaccine. T lymphocytes from the same donors were repeatedly activated by genetically modified DC vaccine to generate heparanase-specific cytotoxicity T lymphocytes (CTLs). CTL-mediated cell lysis of gastric cancer cells lines (KATO-III and SGC-7901) was analyzed in vitro by a standard (51)Cr releasing assay. IFN-gamma secretion was measured by ELISA in heparanase-specific CTLs cocultured with those gastric cancer cell lines. RESULTS: Our results showed that the expression of heparanase in DCs transfected with rAd-Hpa was significantly increased. Furthermore, DCs transfected with rAd-Hpa could induce heparanase-specific CTLs against HLA-matched and heparanase-positive gastric cancer cells in vitro, while there were no killing effects on autologous lymphocytes. Meanwhile, these rAd-Hpa-modified DCs could increase IFN-gamma secretion of effector cells when cocultured with KATO-III cells. CONCLUSIONS: These findings demonstrate for the first time that the transduction of DCs with rAd-Hpa can induce CTLs that specifically lyse heparanase-positive gastric cancer cells and increase IFN-gamma secretion in an MHC-restricted fashion. Heparanase gene-modified DC vaccine offers a great opportunity for immunotherapy in patients with advanced gastric cancer and possibly also with other malignancies.

In vitro anti-tumor immune response induced by dendritic cells transfected with hTERT recombinant adenovirus.

Transduction with recombinant, replication-defective adenoviral (Ad) vectors encoding a transgene is an efficient method for gene transfer into human dendritic cells (DC). Several studies have demonstrated that epitopes of the human telomerase reverse transcriptase gene (hTERT) can produce CTLs specific for malignant tumors. In this study, we constructed an hTERT recombinant adenovirus (rAd-hTERT) using DNA recombination. We found that human dendritic cells transduced with rAd-hTERT could effectively induce hTERT-specific cytotoxic T lymphocytes (CTLs) in vitro against various tumor cell lines, which were hTERT-positive and HLA-A2 matched. We also found that these hTERT-specific CTLs could not lyse autologous lymphocytes with low telomerase activity. Further studies revealed that rAd-hTERT transduced DCs could increase secretion of IFN-gamma by effector cells when they were co-cultured with hTERT-positive and HLA-A2 matched tumor cell lines. These data suggest that an hTERT vaccine can induce anti-tumor immunity against various tumor cells expressing hTERT in a HLA-A2-restricted fashion in vitro. The transduction of DCs with rAd-hTERT offers a great opportunity in cancer immunotherapy.

[Immune response of heparinase gene modified dendritic cell-based vaccine on gastric cancer cells].

OBJECTIVE: To explore the feasibility of heparinase vaccine in active immunity for gastric cancer. METHODS: Dendritic cells originated from the peripheral blood mononuclear cells (PBMC) of healthy HLA-A2 positive donors were transfected with recombinant adenovirus containing heparinase full length cDNA of heparanase to generate heparanase gene modified DC vaccine. T lymphocytes from the same donors were activated by those genetically modified DC vaccine repeatedly to generate heparanase specific cytotoxicity T lymphocytes (CTL). CTL-mediated cell lysis to gastric cancer cells of the lines KATO-III and SGC-7901 was analyzed in vitro by standard (51)Cr releasing assay. Heparinase specific CTL were co-cultured with KATO-III and SGC-7901 cells, and then ELISA was used to detect the IFN-gamma release. RESULTS: Expression of heparanase was significantly increased in the DCs transfected with heparinase recombinant adenovirus. Heparanase specific CTL generated from the genetically modified DC vaccine exhibited potent lysis to the KATO-III gastric cancer cells positive in both heparinase and HLA-A2 at each E/T ratio, whereas, these heparinase specific CTL could not lyse the SGC-7901 cells positive to heparinase but negative to HLA-A2, with a specific lysis rate of only 11.1% +/- 4.6% even at an E:T ratio of 40:1. Further study showed that heparanase vaccination had no detectable lysis on the autologous lymphocytes in vitro with a specific lysis rate of only 11.4% +/- 7.9% even at an E:T ratio of 40:1. The IFN-gamma release amount when the heparanase specific CTL were co-cultured with the KATO-III cells was 280.4 pg/ml +/- 23.5 pg/ml, significantly higher than that when the heparanase specific CTL were co-cultured with the rAd5-Lacz modified DC (120.6 pg/ml +/- 18.9 pg/ml), and that of the IL-2 stimulated T cells (60.0 pg/ml +/- 10.6 pg/ml, both P < 0.05). In contrast the IFN-gamma release amounts of the SGC-7901 cells and autologous lymphocytes remained unchanged when they were co-cultured with either above-mentioned effector cells (both P > 0.05). CONCLUSION: DC genetically modified by heparanase gene activate heparanase specific CTL and induce potent immune response against HLA-matched and heparinase positive gastric cancer cells in vitro, whereas they have no killing effect on autologous lymphocytes. Heparanase is an effective and safe target for immunogen therapy of tumor, thus providing a new biotherapy method for advanced gastric cancer.

[Increased expression of heparanase mRNA is correlated with metastasis of gastric carcinoma and is probably correlated with c-met protein].

OBJECTIVE: To explore the expression of human heparanase mRNA and its relationship with the clinicopathological characteristics and angiogenesis and its possible mechanism in gastric carcinomas. METHODS: Expression of heparanase mRNA in gastric cancerous tissues and its corresponding adjacent tissues were detected by RT-PCR in forty-seven patients. Expression of c-met protein and microvessel density (MVD) were examined by immunohistochemcal staining. RESULTS: Expression of heparanse mRNA was significantly higher in gastric cancerous tissues than in its adjacent tissues (74.5% vs. 19.1%, P < 0.01). MVD was significantly higher in heparanase positive group than its counterpart (51 +/- 15 vs. 31 +/- 10, P < 0.01). Expression of heparanase mRNA was correlated with expression of c-met (r = 0.714, P < 0.01). The positive rate of heparanse expression in gastric carcinomas were correlated with tumor diameter, invasive depth, regional lymph nodes metastasis and TNM stage, whereas there were no relation with gender, age, tumor differentiation and remote metastasis of the patient. CONCLUSION: Increased heparanase expression contributes to metastasis and angiogenesis in gastric cancer. Heparanase mRNA expression correlates with poor clinicopathologic characteristics in its prognosis. C-met is correlated with heparanase expression.