The combination of Dendritic cells-cytotoxic T lymphocytes/cytokine-induced killer (DC-CTL/CIK) therapy exerts immune and clinical responses in patients with malignant tumors

Detection of CIKs,CTLs and intracellular IFN-Y in T cells by flow cytometry

Detection of CIKs,CTLs and intracellular IFN-Y in T cells by flow cytometry

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Summary

The purpose of this study is to investigate the role of DC-CTL/CIK therapy and evaluate the changes of immune indicators and tumor serological markers.As a result The study demonstrated that DC-CTL/CIK therapy could reduce Tregs, MDSCs, and several crucial serological tumor markers in particular tumors, and improve the function of T cells immune systems and the quality of life in patients with malignant tumor..

Patients characteristics

Median age of the patient was 54 years with a range of 27–81 years, 18 were female and 42 were male. The pathologic stage was Stage I in 3 patients (5 %), Stage II in 4 patients (6.7 %), Stage IIIA in 12 patients (20 %), Stage IIIB in 2 patient (3.3 %), Stage IIIC in 9 patient (15 %) and Stage IV in 27 patients (45 %).

Methodology

Peripheral blood mononuclear cells (PBMCs) were harvested by blood cell separator. About 1 × 109 –3 × 109 PBMC cells/40–60 ml white cells were collected from each patient and were cultured in AIM-V cell culture solution for 2 hours. The adherent cells were initiated into culture for DC maturation by 100 ng/ml GM-CSF and 100 ng/ml IL-4 in AIM-V cell culture solution. Part of the non-adherent cells were stored at −80 °C and thawed for co-cultivation, and the rest were applied for inducing CIKs.
On Day 6, the recombinant non-proliferative adenovirus (serotype 35) carrying the expression cassettes of tumor-associated antigens (TAAs) were used to infect the DC cells with a multiplicity of infection (MOI) of 5. After 24 h cultivation, IL-1β (25 ng/ml) and TNF-α (100 ng/ml) were added into the culture for DC- maturation for another 24 h.
On day 8, about 2 × 106 DC cells were harvested and co-cultured with T cells (about 4 × 107 cells) at a DC/T cell ratio of 1:20 for another 4 days to induce antigen-specific CTL cells which were stimulated with CD3 monoclonal antibody (mAb) pre-coated onto plastic plates and amplified by IL-2 (500 IU/ml). The rest cells were applied as DC vaccine. The applied TAAs included p53, survivin, hTERT and additional AFP/CEA for AFP/CEA positive patients.
Additionally, CIKs were cultured in 4 × 40 Ml serum-free medium supplemented with 1000 U/mL IL-2, 5 μg/mL CD3 monoclonal antibodies, 12.5 μg/ml RetroNectin and 1000 U/mL IFN-γ.

Treatment

Consecutively analyzed cohorts of 83 patients in three subgroups. The first subgroup (cohort 1) contained 60 patients received DC-CTL/CIK therapy. In Cohort 2, 14 patients with negative for all serological tumor indexes were reviewed. Cohort 3 included nine patients for myeloid-derived suppressor cells (MDSC) detection.
For each treatment, patients were treated with 3 intravenous infusions. About 1 × 107 DC vaccine, 1 × 109 CIKs and both DC vaccine and CTLs (about 1 × 109 cells) were infused intravenously on day 8, day 10 and day 12 respectively.

Results

Side effects
The side-effects were also recorded in all cases. There were no severe or unusual side-effects recorded that are common in chemotherapy and radiotherapy during or after the transfusion, except for temporary fever. In total, 40 % patients developed a fever during the immunotherapy, and the body temperature was 37.5– 38. These patients recovered without any treatment. Six out of 60 (10 %) cases had temperatures >38 °C treated with drugs to lower body temperature.
Quality of life
The quality of life was measured for patients undergoing DC-CIK/CTL treatment. It is inspiring that the QoL score of 25 (41.7 %) patients improved obviously post treatment, and 12 (20 %) patients did not take a turn for the worse. Thus, it indicated that DC-CIK/CTL therapy could at least partially block the deterioration of cancers in the patients with malignant tumors with elevated quality of life.
Comparison of MDSC levels pre‐ and post‐treatment of DC‐CTL/CIK therapy
To analyze the changes of MDSCs in DC-CTL/CIK therapy, other nine patients were included for the detection of MDSCs and Tregs. The results showed a significant decrease in the proportion of MDSC and Treg after treatment, which provided an additional proof of therapeutic effect of DC-CTL/ CIK therapy.

Conclusion

This study demonstrated that DC-CTL/CIK therapy could reduce Tregs, MDSCs, and several crucial serological tumor markers in particular tumors, and improve the function of T cells immune systems and the quality of life in patients with malignant tumor.

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Scientific article publishing date : 11/10/2015

Immucura identifier BSC21_180EN

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