| Objective:The therapy of cytokine-induced killer(CIK) cells is one of biotherapies. Statistics showed that there were some differences in application of CIK cells treatment for malignant tumor in different clinical research, including widely select of indications, different intervention time of cell therapy, different dosages of infused cells and different ratio of effector cell, so clinical therapeutic effect also have certain differences. Therefore, the research selected several indications which have effective to the therapy in report of clinical application, intervention time was 2 month after standard chemotherapy, assessed patients who with malignant solid tumor, and isolated their peripheral blood mononuclear cells(PBMNCs), cultured with interferon-?(INF-?), anti-CD3 monoclonal antibody(Mab), anti-CD28 Mab and interleukin-2(IL-2), gradually induced and expanded and obtained the CIK cells finally. The culture system reliability was verified, the relationships between CIK cells yield and leukocyte count?lymphocyte count?volumes of blood-drawing ?initial number of PBMNCs were studied to find the initial factors which most influenced the number of CIK cells, in order to ensure steady amount of CIK cells. Through CIK cells therapy, QOL of patients were evaluated by Karnofsky(KPS) score. According to solid tumor curative effect evaluation standard, therapeutical effects were evaluated by objective response rate(ORR), clinical benefic rate(CBR), progression free survival(PFS) and overall survival OS, to investigate the clinical treatment effects of this scheme.Methods: 1Patient inclusion and therapeutic schedule 41 patients with clinical evaluation as stable disease(SD), partial response(PR), and complete remission(CR) after standard chemotherapy in two months were enrolled in this study. The CIK group contained 22 patientsand the other 19 patients enrolled in control group. There were no significant difference in gender, age, body weight, tumor stage, and disease status in the two groups(P > 0.05). The patient in CIK group would receive CIK cells infusion treatment one course a month in the future three months.Every course drew blood one time, infused CIK cells three times continuously(time/day). 2CIK cells culture and infused 60-120 m L peripheral blood was draw by antecubital vein from the CIK group patients. PBMNCs were separated by the method of density gradient centrifugation; CIK cells were gradually induced and expanded with INF-?, anti-CD3 Mab, anti-CD28 Mab and IL-2 from PBMNCs in vitro. In the 14 d, 15 d, 16 d during the interval of CIK cells cultured in vitro, CIK cell injections were prepared respectively and on the condition of quality tests passed, infused to the patient. 3Curative effect evaluation Patients were followed up and reexamined after chemotherapy on a regular basis. In this research, two groups were checked as mentioned above at 2?4?6 and 12 months after chemotherapy. The follow-up ended at the time of disease progression(PD) occurred or patents died. Quality of life(QOL) were evaluated by KPS score, compared the control group with CIK group by the first two follow-up, in CIK group KPS score of 3d?7d?15d after the first course of infusion were compared with the first two follow-up. According to solid tumor curative effect evaluation standard, ORR and CBR of four follow-up were compared, PFS and OS were assessed, that in order to evaluate the clinical curative effect of patients.Result: 1CIKcells culture The results of mycoplasma tests and sterility tests were negative, all samples no pollution. Endotoxin check of CIK injections were qualified. Flow cytometric of cell phenotype showed that CD3+CD56+ was(27.02±3.58)%. 2The relationship between the number of CIK cells and leukocyte count?lymphocyte count?volumes of blood-drawing?PBMNCs count66 samples of peripheral blood were drew from CIK group, the mean of leukocyte count(×109/L) and lymphocyte count(×109/L) were 6.74±4.27 and 1.68±0.52 respectively;the mean of blood-drawing volumes(m L) and initial PBMNCs count(×106) were 68.94±14.37 and 101.64±21.48respectively;the mean number of CIK cells(×109) in bulk culture was 10.43 ± 2.16. There was a highest positive correlation(P<0.05) between the number of CIK cells and PBMNCs count by person correlation test(R=0.804,P<0.0001). Linear regression statistics showed that an equation could be used to estimate the relationship, that was Y= 0.080 X + 2.233(Y represented CIK number cells(×109); X represented PBMNC count(×106)). 3Adverse reactions 198 times infusions of CIK cells were carried out in this study, no nausea, vomiting, dizziness and other adverse reactions occurred during transfusion. After the first infusion, 4 patients suffered from slight fever, and 2 patients felt thirst, soon after spontaneous remission. No rash, shock, embolism, liver and kidney damage occurred during the interval of follow-up. There were no cases that terminate or quit treatment? 4Evaluation of QOL QOL was evaluated with KPS score. At the time of the first follow-up, patients were divided into two groups, KPS score of control group and CIK group were 50.53±10.79 and 56.82±16.44 respectively that there was no significant difference between the two groups(P>0.05). At the time of the second follow-up, the patients of CIK group have finished two courses treatment, KPS score of control group and CIK group were 44.74±22.70 and 70.45±15.88 respectively, that the control group was significantly lower than CIK group(P<0.05). In the CIK group KPS score of 3d?7d?15d afterthe first course of infusion were 57.27±14.86 ? 63.64±16.20 and 64.09±15.32 respectively, which showed a rising trend but no significant difference(P>0.05) between them. This rising trend was continuous along CIK cells treatment in the first two follow-up, the KPS score of the second follow-up was significantly higher than the first follow-up and the third day after the firsttreatment course of infusion(P<0.05). 5Clinical curative effect evaluation During the follow-up period, two groups of patients gradually appeared PD and death cases. ORR and CBR were compared between the two groups in every follow-up period, it showed that of two groups declined with time went by, ORR and CBR of CIK group higher than those of control group, but no significant difference(P>0.05). 1 years survival rate of control group and CIK group were 68.4% and 77.3%, CIK group was a little higher than control group, but there was no statistical difference between the two groups(P>0.05). PFS of control group and CIK group were average 7.74 ± 4.28 and 9.91 ± 2.67(months), OS were average 9.95± 2.81 and 11.09 ± 1.40(months) respectively. Log-rank test showed that no significant differences were observed on PFS and OS between the two groups(P> 0.05).Conclusion: 1There was a positive correlation between the number of CIK cells and PBMNCs count(volume of blood-drawing × lymphocyte count). Therefore, the yield of CIK cells could be guaranteed by controling volumes of blood-drawing individually. 2CIK cells therapy had high security for clinical and no serious adverse events occurred. 3CIK cells therapy intervened after the standard chemotherapy could improve the QOL of patients obviously in a short period, but it couldn't raise ORR and CBR of patients significantly, and also couldn't prolong PFS and OS significantly. |
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