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@# Objective: To evaluate the long-term clinical efficacy and follow-up of dendritic cell (DC) vaccines in combination with cytokine-induced killer cell (CIK) treatment in metastatic renal cell carcinoma. Methods: From January 2011 to December 2013, 29 patients with metastatic renal cell carcinoma (pathologically confirmed as renal clear cell carcinoma) were treated by DC vaccines-CIK at the Department of Hematopoietic Stem Cell Transplantation, the Fifth Medical Center of Chinese PLA General Hospital. The 29 patients included 24 male and 5 female, with a median age of 57(32-81) years old. Mature DC vaccine was obtained by gene transfection technology and CIK cells were obtained by i n v i t r o culture; and DC vaccine-CIK was infused back to patients through lymphatic drainage area and vein by each course. Twelve patients received first line treatment, 6 patients received second line treatment after the disease progression by targeted drug therapy or cytokine therapy, and 11 patients received third-linetreatment or above. The long-term clinical efficacy and overall survival rate were evaluated. Results: The median follow-up time was 5 (1-7) years. Treatment cycle was over 2 (2-23) cycles. One case (3.4%) achieved complete remission, 9 cases (31%) achieved partial responses, 13 cases (44.8%) demonstrated stable disease over 3 months and 6 patients (20.7%) developed progressive disease. The objective response rate was 34.4%,and the disease control rate was 79.2%. Stable disease for more than one year realized in 19 cases (65.5%). The 1-, 3- and 5-year survival rates were 93.1% (27/29), 65.5% (20/29) and 51.7% (15 / 29), respectively. Neither the median progression-free survival (PFS) nor the median survival time was achieved. No adverse reactions above grade 3 were observed during treatment. Conclusion: DC vaccines-CIK therapy for the treatment of metastatic renal cell carcinoma is affirmative; it achieved good disease control and long-term survival with controllable safety, and prolonged the survival time for advanced renal cell carcinoma patients.
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@# Objective: To investigate the function of CIK (cytokine induced killer) cells cultured using ATG-F (anti-human T lymphocyte rabbit immunoglobulin-Fresenius) and IFN- γ, IL-2 system and its feasibility in clinical practice. Methods: Peripheral blood mononuclear cells (PBMCs) were isolated from healthy donors and were used to culture CIK cells by different activating antibodies; the total cell count was calculated on Day 7 and 14. The CIK cell composition, cell surface activation and proportion of inhibitory receptor molecular in ATG-F group, CD3 group and TG (Thymoglobulin) group were analyzed by Flow cytometry, and the cytotoxicity of CIK cells against K562 cells were also determined by flow cytometry at day 14 in ATG-F high-dose group, CD3 group and TG group. Results: CIK cells were successfully cultured by ATG-F, IFN-γ, IL-2 system. The proliferation rate of ATGF high-dose group was significantly higher than that in TG group (27.25±1.25 vs 16.60±1.72, P <0.01), but the proportion of CD3+ CD56+ cells showed no statistical difference compare with the CD3 group ( P >0.05). The percentage of CD3-CD56+ NK cells in ATG-F high-dose group was significantly higher than that in TG group and CD3 group [(11.19±2.60）% vs（5.66±1.00）%,（1.42± 0.51）% ， P <0.01], while the proportion of CD4+T cells was significantly lower than that in CD3, TG group [(4.35±1.47）% vs （26.88±5.01）%,（14.52±6.22）%, P <0.01]; the proportion of CD56+CD94+, CD56+CD158a+, CD56+CD158b cells was significantly higher than those in CD3 group (all P <0.01). The ATG-F high does group showed significantly higher cytotoxicity against K562 cells than that of CD3 group at the target/effect ratio of 1∶10. Conclusion: CIK cells cultured by ATG-F culture system has higher NK cell proportion than other ordinary culture system, and its activated receptor has more stronger cytotoxicity against K562 cells.
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Objective To observe the efficacy of advanced non-small cell lung cancer patients treated with CIK cell combined with chemotherapy.Methods65 cases of advanced lung cancer patients admitted in our hospital from February 2013 to January 2015 were retrospectively analyzed.The patients were randomly divided into observation group (34 cases) and control group (n=31).The levels of vascular endothelial growth factor (VEGF), matrix metalloproteinase-9 (MMP-9) and carcinoembryonic antigen (CEA) in the two groups were observed before and after treatment.The patients in the control group were treated with chemotherapy on the basis of chemotherapy.Cytokeratin 19 (CYFRA21-1) levels, and clinical efficacy.Results①The patients with VEGF and MMP-9 values after treatment (298.67±36.49)ng/L, (1102.31±86.48)ng/mL, than the control group (459.19±45.28)ng/L, (1394.83±121.06)ng/mL, and the difference was significant (P<0.05).②CEA and CYFR21-1 value patients in the observation group were (14.16±1.08) ng/mL, (4.45±0.45) ng/mL, than the control group (17.86±1.84) ng/mL, (5.76±0.64) ng/mL, and the difference was significant (P<0.05).③The total observation group was significantly higher than patients 94.18% 74.19% efficiency, and the difference was statistically significant (P<0.05).ConclusionCIK cell biological therapy combined with chemotherapy for advanced lung cancer patients with a certain improvement is worthy of further research and application.
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Objective To explore the enhancing effect of monoclonal antibody coated with anti-human CD3 and CD28 on acti-vation and transformation of peripheral blood mononuclear cell (PBMC)in vitro.Methods Human peripheral blood mononuclear cells were separated.Cells was cultured in vitro,and determined by flow cytometry.The solid phase with CD3 and CD28 antibody was coated and added in.The mature CIK cells were obtained after 12 days culturing.Results The CD4 + cells was lower in group C than those in group A(P <0.05).The CD8 + cells was higher in group C than that in group A and B(P <0.05).There was signifi-cant difference of T4/T8 between group C and group A and B(P <0.05 ).There was significant difference of NK cells between group B and group C(P <0.05).The CD25 + cells was lower in group C than that in group A (P <0.01).Conclusion CD3 antibody solid coated combined with CD28 antibody added to the suspension has more strong activation than both CD3 antibody and CD28 antibody solid coating on peripheral blood mononuclear cell.
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Objective To observe whether hepatitis B vaccine enhance the treating effect of cyto-kine induced kill(CIK) cells on hepatitis B virus transgenic(HBV-Tg) mice. Methods The HBV-Tg mice were treated with CIK cells by peritoneal injection and hepatitis B vaccine by hypodermic injection. The HBV DNA level were tested by real-time PCR,T lymphocyte subgroup were detected by flow cytometry and the pathological diversify of hepatic tissue were observed by HE staining. Results The HBV DNA loading in peripheral blood of HBV-Tg mice decreased after CIK cells were treated and CD3~+ , CD4~+ and CD8~+ cells increased which were enhanced after CIK cells combined with hepatitis B vaccine. Conclusion Hepa-titis B vaccine enhanced the treating effect of CIK on HBV-Tg mice which may be implemented by increased the blood level of CD3~+, CD4~+ and CD8~+ cells, especially CD8~+ cells level.
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Objective To study the antitumor effect and mechanism of co-cultured cytokine-induced killer(CIK) cells and autologous DC modified with IL-24 gene on A549 cells in vitro. Methods DC and CIK cells were prepared routinely from human peripheral blood mononuclear cells(PBMC). Recombinant adenovirus vector pAdEasy-1-pTrack-CMV-IL-24 was extracted from DH5α, it was lineared with Pac I and transfected into A293 cells, and then the IL-24 recombined adenovirus(Ad-IL-24) was obtained. Ad-IL-24 was used to infect DC. The cells obtained were named DC-IL-24. RT-PCR and ELISA were used to evaluate the expression of IL-24 gene in transfected DC. The phenotypes change of DC were identified by flow cytometry analysis, the concen-tration of IL-12 and TNF-α in supernatant of DC were determined by EIJSA. The ability of CIK producing per-forin was measured by homolysis method. FCM was used to determine the cytotoxicity of cocultured CIK cells and autologous DC modified with IL-24 gene to A549 cells. Results We obtained the high titre of Ad-IL-24.IL-24 gene was transfered into DC successfully via Ad-IL-24. The green fluorescence was observed on DC by fluorescence microscope. The expression rate of CD80, CD83, HI.A-DR, CD40, CXCR4 on DC-IL-24 was sig-nificantly increased compared with that of the control group. DC-IL-24 produced markedly higher levels of IL-12 and TNF-α as compared with DC. DC-IL-24 can enhance the ability of CIK cells producing perforin. On com-parison with non-transfected DC co-cultured with CIK cells, transfected DC co-cultured with CIK cells had a sig-nificantly higher lytic activity against A549 cells. Conclusion IL-24 gene modification can enhance the anti-tu-moral immunity of DC. The mechanism of which might be related to the increased secretion of IL-12 and TNF-α, up-regulation expression of co-stimulatory molecules and MHC Ⅱ class molecules on DC, promoting the acti-vation and maturation of DC, and then enhancing CIK cells to generate specific anti-tumoral immunity.
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Objective:To study the antitumor effect and mechanism of cocultured CIK cells modified with IL-24 gene and autologous DCs on HL-60 cells in vitro.Methods:DCs and CIK cells were prepared routinely from human peripheral blood mononuclear cells ( PBMC).IL-24 gene was transferred into CIK cells via electroporation.The cells obtained were named CIK-IL24.RT-PCR and ELISA were used to evaluate expression of IL-24 gene in transfected CIK cells.The phenotypic changes of CIK cells were identified by flowcytometry analysis.The concentration of IFN-γ and TNF-α in supernatant of CIK was determined by ELISA.FCM was used to determine the cytotoxicity of cocultured CIK cells modified with IL-24 gene and autologous DCs against HL-60 cells.Results:Eukaryotic expressing plasmid pcDNA3.0-IL24 was transferred into CIK cells successfully via electroporation.The expressing rate of CD3~+、CD3~+CD56~+ cells had no significant change in CIK cells.However,the rate of CD4~+CD25~+ cells was significantly decreased compared with that of the control group.Expression of adhesion molecules CD54,CXCR4 were significantly increased on CD3+CD56+ cells.CIK-IL24 cells produced markedly higher levels of IFN-γ and TNF-α as compared with the CIK cells.By comparison with non-transfected CIK cells co-cultured with DCs,transfected CIK cells co-cultured with DCs had a significantly higher lytic activity against HL-60 cells.Conclusion:IL-24 gene modification can enhance the anti-tumoral immunity of CIK cells,the mechanism of which might be related to the increased secretion of IFN-γ,TNF-α,up-regulation of adhesion molecule expression,and reduction of the rate of CD4~+CD25~+ cells in CIK cells.