The therapeutic effect of HSV1-hGM-CSF combined with doxorubicin on the mouse breast cancer model / 中华肿瘤杂志

Objective@#To evaluate the oncolytic effect of herpes simplex virus type 1 which carried recombined human granulocyte-macrophage colony-stimulating factor (HSV1-hGM-CSF) on the mouse breast cancer cell line 4T1 and compare the anticancer effects of HSV1-hGM-CSF, doxorubicin alone or combination on the breast cancer in mice.@*Methods@#We investigated the cytotoxic effect on 4T1 cells in vitro, the cell growth, cell apoptosis and cell cycle of 4T1 cells treated with oncolytic HSV1-hGM-CSF at different MOIs (0, 0.5, 1 and 2) and doxorubicin at different concentrations (0, 2, 4 and 8 μg/ml). The effects of oncolytic HSV1-hGM-CSF and doxorubicin on the tumor growth, survival time and their side effects on the mouse breast cancer model were observed.@*Results@#Both oncolytic HSV1-hGM-CSF and doxorubicin significantly inhibited the proliferation of 4T1 cells in vitro. Doxorubicin induced the G2/M phase arrest of 4T1 cells, while the cytotoxicity of oncolytic HSV1-hGM-CSF was no cell cycle-dependent.At day 16 after treatment with doxorubicin and HSV1-hGM-CSF, the tumor volume of 4T1 tumor bearing mice were (144.40±27.68)mm3, (216.80±57.18)mm3, (246.10±21.90)mm3, (327.50±44.24)mm3, (213.30±32.31)mm3 and (495.80±75.87)mm3 in the groups of doxorubicin combined with high dose HSV1-hGM-CSF, doxorubicin combined with low dose HSV1-hGM-CSF, doxorubicin alone, high dose HSV1-hGM-CSF alone, low dose HSV1-hGM-CSF alone and control, respectively.Compared with the control group, both doxorubicin and HSV1-hGM-CSF treatment exhibited significant reduction of primary tumor volume in vivo (P<0.001). The median survival times were 48, 50, 40, 42, 43 and 37 days in the six groups mentioned above, respectively. The median survival period of doxorubicin alone, high dose HSV1-hGM-CSF alone and low dose HSV1-hGM-CSF alone were significantly longer than that of control (P<0.05).@*Conclusion@#Synergistic effect of sequential treatment with doxorubicin and oncolytic HSV1-hGM-CSF is observed in 4T1 mouse breast cancer.

Radiofrequency ablation inhibits lung metastasis ofbreast cancer in mice / 中华肿瘤杂志

<p><b>OBJECTIVE</b>To explore the effects of radiofrequency ablation(RFA) on immune system and lung metastasis in a mouse model of triple negative breast cancer 4T1.</p><p><b>METHODS</b>Mouse breast cancer 4T1 cells were injected into the right hind limb of female Bal B/c mice. When the tumor size was 6-8 mm in diameter, RFA was used to treat the transplanted breast cancer in mice. We examined the splenic lymphocyte subsets by flow cytometry at different time points after RFA. Fourteen days after treatment, we sacrificed the mice of both control and treatment groups, counted the number of lung metastatic nodules, and detected the changes of splenic lymphocyte subsets by flow cytometry.</p><p><b>RESULTS</b>RFA basically eliminated the orthotopic carcinoma with a low local recurrence rate. After the RFA treatment, the amount of spleic CD4⁺ T cells, CD8⁺ T cells, B cells, NK and NKT cells was increased. Fourteen days after the RFA treatment, all mice were sacrificed, and the lung metastatic nodules were 24 ± 18 in the control group and 81 ± 35 in the RFA-treated group (P = 0.012). The mechanism of suppression of metastatic lung cancers was related to the increase of splenic CD4⁺ T cells, CD8⁺ T cells, B cells and NK cells, and the decrease of myeloid-derived suppressor cells.</p><p><b>CONCLUSIONS</b>RFA can enhance the anti-tumor immunity and effectively inhibit lung metastasis of 4T1 cell-induced breast cancer, and has a good potential effect in the treatment of triple-negative breast cancer and the control of distant metastasis.</p>

Construction and identification recombinant eukaryotic expressive vector of mouse GM-CSF gene / 中国免疫学杂志

Objective: Construction mouse GM-CSF gene effective eukatyotic expressive vector, selection high GM-CSF expressing mouse leukemia cell line RMA after transfected with the constructed vector, and study the method of treatment leukemia with tumor cells transfected with GM-CSF gene.Methods:770 bp of GM-CSF 3' end cDNA was amplified by PCR and inserted into pcDNAS vector.The constructed vector was transfected into RMA cells by electroporation. After screening by G418 and cloning by limiting dilution, a relative high GM-CSF expressing cell clone was selected by RT-PCR, hematopoietic progenitor cell proliferative assay and hematopoietic progenitor cell colony formation assay. The cells of this clone were inactivated by mitomycin-C and vaccinated mice to induce antitumor immune reaction. Results: The orientation and sequence of the insert was found to be correct, and a GM-CSF high expressing cell line RMA-GM was selected , which can induce mice obtain anti-tumor protective immune ability after inactivated by mitomycin-C.Conclusion:Tumor cells transfected with GM-CSF gene may be used an effective anti-T lymphycoma tumor vaccine.

Expression of Human T Cell Receptor ? Chain Gene in Insect Cell / 中国肿瘤生物治疗杂志

Objective: To clone human T cell receptor (TCR) ? chain gene and express its encoding protein by baculoviral expression system in insect cells. Methods: TCR ? chain cDNA was cloned from normal human peripheral blood mononuclear cells (PBMC) by RT-PCR and inserted into baculoviral transfer vector. This vector was co-transfected with baculovirus into insect sf 9 cells. The recombinant protein expressed was identified by SDS-PAGE and flow cytometry with mouse anti-human ? chain monoclonal antibody. Results: Human TCR ? chain cDNA cloned and inserted into baculoviral vector specifically expressed protein in the insect sf 9 cells, accounting for about 11% of the total protein yield in the supernatant of cell lysate. The molecular weight of the recombinant protein determined by SDS-PAGE was identical to what we anticipated. The insect cells transfected with recombinant baculovirus were demonstrated by intracellular labeling flow cytometry to express ? chain protein. Conclusion: Human T cell receptor ? chain gene was successfully cloned from human PBMC, and its encoding protein was highly expressed in insect cells. The TCR ? chain protein obtained by bioengineering technique is useful for in depth biological function study.

Studies of the augmenting effect of selenium (Se) on ceil-mediated immunity—The effect of Se on T cell function in mice / 中国免疫学杂志

In this paper,the in vitro and in vivo effect of Se on T cell function was presented. Mouse spleen lymphocytes cultured in the presence of sodium selenite(30～1,000ng/ml)showed increased proliferation in response to Concanavalin A,but sodium selenite itself was not mitogenic.When sodium selenite was added into one-way mixed lymphocyte culture (MLC),not only was the proliferation of the responder cells increased,the induction of cytotoxic T lymphocytes(CTL)was also enhanced.In mice given Se(2-4ppm)in drink- ing water,the activity of MLC-induced CTL was higher than in untreatedcontrols.The augmented cytotoxic activity of CTL was alloantigenspecific.