Effect Of Cytokine-induced Killer Cells Research Articles

Analysis the effect of cytokine-induced killer cells combined with mFOLFOX6 regimen in chemotherapy for advanced colorectal cancer

Objective: To investigate the effect of cytokine-induced killer cells (CIK) combined with mFOLFOX6 regimen (fluorouracil+oxaliplatin+calcium folinate) on advanced colorectal cancer. Methods: 161 patients with advanced colon cancer admitted to the Fourth Clinical College of Xinxiang Medical College, Xinxiang Central Hospital of Henan Province from 2019 to 2021 were selected and divided into the study group (mFOLFOX6 regimen chemotherapy+CIK therapy) with 80 cases and the control group (mFOLFOX6 regimen chemotherapy) with 81 cases. The KPS score, median survival time, progression-free survival time, the proportion of Treg in peripheral blood mononuclear cells, the levels of CD4+, CD4+/CD8+, the expression levels of Foxp3 and CD127 mRNA, the quality-of-life score and the occurrence of toxic and side effects were compared between the two groups before and after chemotherapy. Results: The ages of patients in the study group and the control group were (64.8±7.5) and (66.1±7.0) years old, respectively, and the proportions of males were 62.5% (50 cases) and 50.6% (41 cases), respectively (both P values>0.05). After 2 cycles of chemotherapy and at the end of chemotherapy, the proportions of Treg, Foxp3 and CD127 mRNA in the study group were lower than those in the control group [2 cycles of chemotherapy: (4.33±0.86)% vs (4.89±1.20)%, (0.61±0.22) vs (0.73±0.20), (0.58±0.13) vs (0.70±0.15); after chemotherapy: (2.66±0.70)% vs (3.31±0.84)%, (0.43±0.18) vs (0.51±0.16), (0.41±0.10) vs (0.50±0.13)] (all P values<0.05). The KPS scores of the study group were higher than those of the control group [2 cycles of chemotherapy: (68.41±5.41) points vs (65.38±5.11) points; after chemotherapy: (72.08±5.94) points vs (67.95±5.51) points] (all P values<0.05). The median survival time of the study group was (16.8±2.2) months, which was higher than that of the control group [(11.2±1.8) months]. The progression-free survival time of the study group was also higher than that of the control group [(9.5±1.1) months vs (6.4±1.2) months], and the mortality rate was lower than that of the control group [11.3% (9 cases) vs 31.3% (25 cases)] (all P values<0.001). After chemotherapy, the scores of physical function, emotional function and role function in the study group were higher than those in the control group, and the scores of pains, fatigue and insomnia in the study group were lower than those in the control group (all P values<0.05). Conclusions: CIK combined with mFOLFOX6 regimen for advanced colorectal cancer can improve and regulate the immune function of patients, prolong the survival time of patients, and enhance the quality of life of patients.

Increase of Antitumoral Effects of Cytokine-Induced Killer Cells by Antibody-Mediated Inhibition of MICA Shedding.

Natural killer group 2D (NKG2D) receptor plays a pivotal role in cytokine-induced killer (CIK) cell-mediated cytotoxicity against malignancies, and the expression of NKG2D ligands might allow targets to be more susceptible to the CIK cell-mediated destruction. In this study, we investigated the synergistic effects of CIK cells antitumor activity and antibody-mediated inhibition of MICA/B shedding. This monoclonal antibody (7C6) has been previously shown to be able to specifically target MICA/B a3 domain on tumor cells, resulting in the increase in cell surface MICA/B expression by inhibition of their shedding. In the current study, we show that 7C6 antibody could substantially inhibit MICA shedding and stabilize the expression of MICA/B on Hela cells and MDA-MB-231 cells. In combination with 7C6, CIK cells showed higher degranulation rate, more IFN-γ production and elevated cytotoxic capacity against tumor cells. Furthermore, we demonstrate that NKG2D-MICA/B ligation could lead to activation of both CD3+ CD56− T cells and CD3+CD56+ NKT subset cells of CIK culture and NKT subset was more sensitive to NKG2D signaling than the counterpart T cells. 7C6-mediated inhibition of MICA shedding could strengthen this signal and eventually enhance the antitumor activity of CIK cells. With multiple advantages of easy ex vivo expansion, minor GVHD, natural tumor trafficking and non-MHC restricted, CIK cell-based therapy may serve as a potent combination partner with MICA antibody-mediated immunotherapy.

Compound Kushen Injection Promoted the Killing Effect of Cytokine-Induced Killer Cells Which Was Activated by Dendritic-Colon Cancer Stem Cell Fusion Cells on Colon Cancer Stem Cells

Colon cancer stem cells (cCSCs) are highly tumorigenic and resistant to traditional chemotherapeutic drugs. Therefore, they are an essential factor in colorectal cancer (CRC) metastasis and recurrence. Dendritic cells (DCs) could bind to the tumor cells and form the fusion cells (FCs). And these FCs could inhibit the development of malignant tumors. Furthermore, the cytokine induced killer (CIK) cells (CD3+ /CD56+ T lymphocytes) could also apply for the immunotherapy of cancer. And compound Kushen injection (CKI) is a traditional Chinese medicine (TCM) which has been used for the treatment of various tumors. However, whether the dendritic-colon cancer stem cell fusion cells (DC-cCSC FCs) could activate the CIK cells and kill the colon cancer stem cells is unknown. And whether the CKI could enhance the lethal effect is still unclear. In this study, we collected peripheral blood samples from healthy participants to acquire mononuclear cells and induced DC and CIK cells. Meanwhile, the CD44+ cells (cCSCs) were screened from SW480 cells. Next, the DC-cCSC FCs were established for the next experiments. At last, CCK-8 assays were performed to determine the effect of DC-cCSC FCs and CKI on the viability of cCSCs. We found that DC-cCSC FCs enhanced the proliferation of CIK cells and induce the CIK cells to secrete more IL-12. The DC-cCSC FCs enhanced the inhibitory effect of CIK cells on cCSCs. Furthermore, application of CKI enhanced the killing rates of DC-cCSC FCs and CIK cells on cCSCs. CIK cells activated by the DC-cCSC FCs had the lethal effect on the cCSCs. Furthermore, CKI enhanced this lethal effect of DC-cCSC FCs and CIK cells.

Abstract 3180: Targeting peptide-loaded-DC CIK cells induce a specific antitumor response

Abstract Currently, immunotherapy has become an effective alternative therapeutic approach for cancers. Cytokine-induced killer (CIK) cells have a higher proliferation rate, increased efficacy with few side-effects, and non- MHC-restricted killing after co-culturing with dendritic cells (DCs). However, the specificity and efficacy of DC-CIK treatment still need to be improved. In our study, the antitumor effects of CIK cells co-culturing with DCs pulsed with non-cell derived targeting peptides, which could specifically bind to certain tumor cells, were evaluated. Our results indicated that targeting peptide-loaded DCs could enhance the differentiation and cytotoxicity of CIK cells. CIK cells, which were treated with specific targeting peptide-loaded DCs, could effectively and specifically kill tumor cells in vitro and in vivo, as long as tumor cells were pre-treated with these targeting peptides. Moreover, the CIK cytotoxic effect of this type acquired a certain properties of memory and resistance to the tolerance. In conclusion, targeting peptides could guide DC-CIK more effectively and specifically kill tumor cells and this non-cell derived targeting peptide-loaded-DC CIK may work as a novel means for cancer therapy. Citation Format: Yimin Zhu, Xueyuan Cui, Cuijuan Liu. Targeting peptide-loaded-DC CIK cells induce a specific antitumor response [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 3180.

The priming role of dendritic cells on the cancer cytotoxic effects of cytokine-induced killer cells

Introduction: In vitro cultivation of DCs and cytokine-induced killer cells (CIK cells) - a special phenotype of T lymphocyte populations — for cancer treatment has gained significant research interest. The goal of this study is to understand whether the priming from DCs helps CIK cells to exert their toxic function and kill the cancer cells.&#x0D; Methods: In this research, DCs were differentiated from mononuclear cells in culture medium supplemented with Granulocyte-macrophage colony-stimulating factor (GM-CSF), and Interleukin-4 (IL-4), and were induced to mature with cancer cell antigens. Umbilical cord blood mononuclear cells were induced into CIK cells by Interferon-γ (IFN-γ), anti-CD3 antibody and IL-2. After 4-day exposure (with DC:CIK = 1:10), DCs and CIK cells interacted with each other.&#x0D; Results: Indeed, DCs interacted with and secreted cytokines that stimulated CIK cells to proliferate up to 133.7%. In addition, DC-CIK co-culture also stimulated strong expression of IFN-γ. The analysis of flow cytometry data indicated that DC-CIK co-culture highly expressed Granzyme B (70.47% ± 1.53, 4 times higher than MNCs, twice higher than CIK cells) and CD3+CD56+ markers (13.27% ± 2.73, 13 times higher than MNCs, twice higher than CIK cells). Particularly, DC-CIK co-culture had the most specific lethal effects on cancer cells after 72 hours.&#x0D; Conclusion: In conclusion, co-culture of DCs and CIK cells is capable of increasing the expression of CIK-specific characteristics and CIK toxicity on cancer cells.&#x0D; &#x0D;

Epithelial ovarian cancer stem‑like cells are resistant to the cellular lysis of cytokine‑induced killer cells via HIF1A‑mediated downregulation of ICAM‑1.

Epithelial ovarian cancer(EOC) is the most lethal of all gynecologic tumors. Cancer spheroid culture is a widely used model to study cancer stem cells. Previous studies have demonstrated the effectiveness of cytokine‑induced killer(CIK) cell‑based therapies against cancer and cancer stem cells. However, it is not clear how EOC spheroid cells respond to CIK‑mediated cellular lysis, and the mechanisms involved have never been reported before. A flow cytometry‑based method was used to evaluate the anti‑cancer effects of CIK cells against adherent A2780 cells and A2780 spheroids. To demonstrate the association between hypoxia inducible factor‑1α(HIF1A) and intercellular adhesion molecule‑1(ICAM‑1), two HIF1A short hairpin RNA(shRNA) stable transfected cell lines were established. Furthermore, the protein expression levels of hypoxia/HIF1A‑associated signaling pathways were evaluated, including transforming growth factor‑β1(TGF‑β1)/mothers against decapentaplegic homologs(SMADs) and nuclear factor‑κB(NF‑κB) signaling pathways, comparing A2780 adherent cells and cancer spheroids. Flow cytometry revealed that A2780 spheroid cells were more resistant to CIK‑mediated cellular lysis, which was partially reversed by an anti‑ICAM‑1 antibody. HIF1A was significantly upregulated in A2780 spheroids compared with adherent cells. Using HIF1A shRNA stable transfected cell lines and cobalt chloride, it was revealed that hypoxia/HIF1A contributed to downregulation of ICAM‑1 in A2780 spheroid cells and adherent cells. Furthermore, hypoxia/HIF1A‑associated signaling pathways, TGF‑β1/SMADs and NF‑κB, were activated in A2780 spheroid cells by using western blotting. The findings indicate that EOC stem‑like cells resist the CIK‑mediated cellular lysis via HIF1A‑mediated downregulation of ICAM‑1, which may be instructive for optimizing and enhancing CIK‑based therapies.

Selective effect of cytokine-induced killer cells on survival of patients with early-stage melanoma.

Adoptive immunotherapy using cytokine-induced killer (CIK) cells has shown potential antitumor ability against several kinds of cancers, including melanoma. However, little is known about the achievable outcome of CIK cells in melanoma patients at different pathological stages. Here we recruited 55 patients treated with conventional therapy plus CIK cells as the CIK group, and 49 patients treated with conventional therapy alone as the control group. The pathological characteristics were comparable between two groups, with a follow-up period up to 40months. Survival data and immune responses were evaluated after CIK cell treatment. In this study, CIK cells were successfully generated from peripheral blood of melanoma patients after in vitro culture for 14days. The cultured CIK cells not only produced high levels of pro-inflammatory cytokines upon in vitro stimulation but also efficiently killed human melanoma cell lines. No serious side events were observed in all patients treated with CIK cells. Furthermore, infusions of CIK cells improved the quality of life in some patients, including advanced cases. More importantly, the CIK group exhibited better survival rates compared to the control group among early-stage melanoma patients, in consistent with the increased frequency of peripheral CD4+ T cells. However, the patients with advanced-stage melanoma did not benefit from the CIK cell therapy in terms of survival rate. In conclusion, CIK cells combined with conventional treatments may prolong the survival of early-stage melanoma patients and improve the quality of life for some advanced cases in a safe way.

Immunotherapeutic effects of cytokine-induced killer cells combined with CCL21/IL15 armed oncolytic adenovirus in TERT-positive tumor cells

The effective antitumor immune responses are dependent on coordinate interaction of various effector cells. Thus, the combination of adoptive immunotherapy and target gene therapy is capable of efficiently generating a productive antitumor immune response. We investigated whether combination of cytokine-induced killer (CIK) cells adoptive immunotherapy and CCL21/IL15 armed oncolytic adenovirus could induce the enhanced antitumor activity. The CCL21/IL15 co-expression oncolytic adenoviruses were constructed by using the AdEasy system, which uses homologous recombination with shuttle plasmids and full length Ad backbones. This conditionally replicating adenoviruses CRAd-CCL21-IL15 could induce apoptosis in TERTp-positive tumor cells for viral propagation, but do not replicate efficiently in normal cells, because the E1A promoter was replaced by telomerase reverse transcriptase promoter (TERTp). Our results showed that the combination of CIK cells and CRAd-CCL21-IL15 could induce higher antitumor activity than either CIK cells or CRAd-CCL21-IL15 alone. This combined treatment could induce the tumor specific cytotoxicity of CTLs (cytotoxic T lymphocytes) in vitro. Moreover, the treatment of established tumors with the combined therapy of CIK cells and CRAd-CCL21-IL15 resulted in tumor regression. This study suggests that the combined treatment by adoptive immunotherapy and gene therapy is a promising strategy for the therapy of tumor.

Effect of cytokine-induced killer cells on immune function in patients with lung cancer.

Cytokine-induced killer (CIK) cells have been used as adoptive immunotherapy in cancer. The present study evaluated the effect of CIK cells on immune function in patients with lung cancer. Patients were divided into three groups, according to the treatment received prior to CIK cell treatment: CIK group (no prior treatment), Che-Sur group (prior chemotherapy and surgery) and Che-Rad group (prior chemotherapy and radiotherapy). Following treatment, the average percentage of cluster of differentiation (CD)3+CD4+, CD3+, natural killer (NK) and NKT cells in peripheral blood was significantly higher than that prior to CIK treatment in the Che-Sur and CIK groups, and the levels of interferon-γ in serum were significantly higher than those prior to CIK treatment in the Che-Sur and CIK groups. On the contrary, the levels of interleukin-10 had decreased in these groups following CIK treatment. Subsequently, patients were divided into three groups according to the percentage of CD3+CD56+ CIK cells that were administered to the patients. The number of NK and NKT cells increased with increasing number of CD3+CD56+ cells. The patients in the CIK and Che-Sur groups were the most benefited ones following CIK treatment, contrarily to those in the Che-Rad group, since the increase in the number of CD3+CD56+ CIK cells in the aforementioned patients enhanced the number of NK cells, which exhibit antitumor activity.

Phenotypic characterization and anti-tumor effects of cytokine-induced killer cells derived from cord blood

Background aimsCytokine-induced killer (CIK) cell therapy represents a feasible immunotherapeutic option for treating malignancies. However, the number of anti-tumor lymphocytes cannot be easily obtained from the cancer patients with poor immunity status, and older patients cannot tolerate repeated collection of blood. Cord blood–derived CIK (CB-CIK) cells have shown efficacy in treating the patients with cancer in several clinical trials. This study was conducted to evaluate the biological characteristics and anti-tumor function of CB-CIK cells. MethodsThe immunogenicity, chemokine receptors and proliferation of CB-CIK cells were analyzed by flow cytometry. The CIK cells on day 13 were treated with cisplatin and the anti-apoptosis capacity was analyzed. The function of CB-CIK cells against the human cancer was evaluated both in vitro and in vivo. ResultsCompared with peripheral blood–derived CIK (PB-CIK) cells, CB-CIK cells demonstrated lower immunogenicity and increased proliferation rates. CB-CIK cells also had a higher percentage of main functional fraction CD3+CD56+. The anti-apoptosis ability of CB-CIK cells after treatment with cisplatin was higher than that of PB-CIK cells. Furthermore, CB-CIK cells were effective for secreting interleukin-2 and interferon-γ and a higher percentage of chemokine receptors CCR6 and CCR7. In addition, tumor growth was greatly inhibited by CB-CIK treatment in a nude mouse xenograft model. ConclusionsCB-CIK cells exhibit more efficient anti-tumor activity in in vitro analysis and in the preclinical model and may serve as a potential therapeutic approach for the treatment of cancer.

The Role of Mir-155 in the Apoptosis of Human Lymphoma Cell Induced By Cik Cells

ABSTRACT Aim: To observe the effect of cytokine-induced killer cells (CIK) on the apoptosis of human lymphoma cells (Raji and BJAB), and explore the role of microRNA-155(miR-155) in this process. Methods: MiR-155 was determined by real-time quantitative PCR, and the apoptosis was detected by flow cytometry in Raji and BJAB cells. Psi CHECK2-CEBPB 3′-UTR containing the binding site of miR-155 was constructed, and then transfected into Raji and BJAB cells. Luciferase activity of CEBPB (CCAAT/enhancer binding protein beta) was determined with the assistance of a dual luciferase report system. Results: CIK cells could promote Raji and BJAB cells apoptosis (t = 3.634, P Conclusions: CIK cells could enhance human lymphoma Raji and BJAB cells apoptosis by upregulating miR-155, which may provide a new database to elucidate lymphoma cell therapy using CIK cells. Disclosure: All authors have declared no conflicts of interest.

Effect of cytokine-induced killer cells expressing hIL-21 on hepatocellular carcinoma in nude mice

Objective To investigate the feasibility of using cytokine-induced killer(CIK) cells as targeted vehicle to deliver hIL-21 gene for hepatocellular carcinoma(HCC) treatment. Methods We constructed pDC759-hIL-21 and pDC759-CopGFP for cotransfection of HEK293 cells with pAd5 and pAd5F35 which can express hIL-21 and CopGFP proteins for viral packaging.We isolated CIK cells and plotted the growth curve.Then Ad5/AAV-CopGFP and Ad5F35/AAV-CopGFP of different MOIs(0,1,5,10,50,100,and 1 000) were used to infect CIK cells;Ad5/AAV-hIL-21 and Ad5F35/AAV-hIL-21 of different MOIs(0,1,5,10,20,and 50) were used to infect SMMC-7721 cells and the hIL-21 protein levels were examined by ELISA assay 48 h after infection. Ad5F35/AAV-hIL-21 of different MOIs(0,1,5,10,50,and 100) was used to infect CIK cells and hIL-21 protein levels were also examined 48 h after infection. SMMC-7721 cells were subcutaneously injected to nude mice for tumor forming,and then the tumor-bearing mice were treated with CIK cells carrying hIL-21 and the therapeutic effects were observed. Results Plasmids for viral packaging were constructed.After cotransfection of HEK293 cells,Ad5 and Ad5F35 which can express hIL-21 and CopGFP proteins were successfully packaged.Ad5 and Ad5F35 which can express CopGFP were used in this study.It was found that Ad5F35 was superior to Ad5 in CIK infection,with the suitable MOI being 50.Ad5/AAV-hIL-21 and Ad5F35/AAV-hIL-21 had similar effect on IL-21 expression in SMMC-7721 cells.Results of animal study showed that mice in group CIK/hIL-21 had a better curative effect. Conclusion hIL-21 and CIK cells have synergistic anti-tumor effect in nude mice bearing HCC.

Tandem therapy for retinoblastoma: immunotherapy and chemotherapy enhance cytotoxicity on retinoblastoma by increasing apoptosis

The goal of this study was to provide an experimental basis for the clinical application of cell immunotherapy on RB in combination with chemotherapy treatment and to explore the mechanism of their combined cytotoxicity. We investigated the antitumor effect of cytokine-induced killer cells (CIK), co-cultivated with dendritic cells pulsed with tumor antigens (DC-Ag) and/or with carboplatin. Cytotoxicity was evaluated on a retinoblastoma cell line (RB-Y79) by FCM and immunofluorescence microscopy. Time-lapse video microscopy was used to follow the sequence of events during the carboplatin and CIK cytotoxicity. Our results showed that a small proportion of RB-Y79 cells died after a low-dose carboplatin application. The cell population recovered 5 days after carboplatin was removed from the culture medium. Three times fewer normal epithelium retina cell lines (hTERT-RPE1) died at the same carboplatin dose. CIK achieved 5 times more cytotoxicity against RB cells pre-treated with low dose of carboplatin, showing the highest antitumor activity in the tandem carboplatin-DC-Ag-CIK-carboplatin treatment. Time-lapse video microscopy revealed that carboplatin-preconditioned RB cells are more avidly engaged by CIK cells, increasing RB mortality and resulting in an overall increment in apoptosis. This study provides evidence that carboplatin combined with cell immunotherapy is superior to carboplatin alone to kill RB cells in vitro. We propose that a primary application of a low dose of a chemotherapeutic drug that is able to attack the tumor, and a subsequent treatment with highly effective immunotherapy based on DC-Ag-CIK cells could be a safe and selective treatment for RB.

Assessment of CIK cells in adoptive cellular therapy

Cytokine-induced killer (CIK) cells therapy plays an important role in cancer adjuvant treatment.Researches show that the number of peripheral blood immunocytes will change if patients with cancer accept the treatment of CIK cells.The changes of T cell subsets,regulatory T cells are relatively obvious,which may be one of the standards that can evaluate the curative effect of CIK cells. Key words: Neoplasms; Cytokine-induced killer cells; Immune response

The effects of cytokine-induced killer cells for the treatment of patients with solid tumors: a clinical retrospective study

Cytokine-induced killer (CIK) cells exert high impact on adoptive immunotherapeutic approaches for malignant tumors. This study aimed to evaluate the effect of adjuvant immunotherapy with CIK cells on the prognosis of solid tumor. Peripheral blood mononuclear cells were collected by a blood cell separator from 40 patients, then expanded by priming them with interferon-gamma followed by monoclonal antibody against CD3 and interleukin-2 the next day. The phenotypic patterns of CIK cells were characterized by flow cytometry on days 0, 7, 10, 14 and 21 of incubation, respectively. Then, 5 ml of venous blood was obtained from 40 patients before and after CIK cells were transfused into patients to assess the influence of CIK cells on the percentages of effector cells. After 14 days of incubation in vitro, the percentages of CD3(+), CD8(+), CD3(+) and CD56(+) increased significantly (P < 0.05). The clinical symptoms of 40 patients were improved apparently. The 6-month, 1-year and 3-year overall survival rates were 70.0, 60.0 and 57.5%, respectively. Our results indicated that CIKs immunotherapy can be an effective adjuvant instrument of the routine therapy of malignancy.

The Therapeutic Effect of Cytokine-Induced Killer Cells on Pancreatic Cancer Enhanced by Dendritic Cells Pulsed with K-Ras Mutant Peptide

Objective. This study is to investigate the role of the CIKs cocultured with K-ras-DCs in killing of pancreatic cancer cell lines, PANC-1 (K-ras+) and SW1990 (K-ras−). Methods. CIKs induced by IFN-γ, IL-2, and anti-CD3 monoantibody, K-ras-DCCIKs obtained by cocultivation of k-ras-DCs and CIKs. Surface markers examined by FACS. IFN-γ IL-12 ,CCL19 and CCL22 detected by ELISA. Proliferation of various CIKs tested via 3H-TdR. Killing activities of k-ras-DCCIKs and CTLs examined with 125IUdR. Results. CD3+CD56+ and CD3+CD8+ were highly expressed by K-ras-DCCIKs. In its supernatant, IFN-γ, IL-12, CCL19 and CCL22 were significantly higher than those in DCCIK and CIK. The killing rate of K-ras-DCCIK was greater than those of CIK and CTL. CTL induced by K-ras-DCs only inhibited the PANC-1 cells. Conclusions. The k-ras-DC can enhance CIK's proliferation and increase the killing effect on pancreatic cancer cell. The CTLs induced by K-ras-DC can only inhibit PANC-1 cells. In this study, K-ras-DCCIKs also show the specific inhibition to PANC-1 cells, their tumor suppression is almost same with the CTLs, their total tumor inhibitory efficiency is higher than that of the CTLs.

Synergistic therapeutic effects of cytokine-induced killer cells and temozolomide against glioblastoma

The presence of active immunity within the brain supports the possibility of effective immunotherapy for glioblastoma (GBM). To provide a clinically-relevant adoptive immunotherapy for GBM using ex vivo expanded cytokine-induced killer (CIK) cells, the treatment capability of CIK cells, either alone or in combination with temozolomide (TMZ) were evaluated. Human CIK (hCIK) cells were cultured from PBMC using activating anti-CD3 antibody and IL-2, which were 99% CD3+, 91% CD3+CD8+ and 29% CD3+CD56+. In vitro, hCIK cells showed tumor-specific cytotoxicity against U-87MG human GBM cells. When hCIK cells were injected into tail veins of immune-compromised mice bearing U-87MG tumors in their brains, numerous CIK cells infiltrated into the brain tumors. CIK treatments (1x10(5), 1x10(6) or 1x10(7), once a week for four weeks) inhibited the tumor growth significantly in a dose-dependent manner; 44, 54 and 72% tumor volume reduction, respectively, compared with the control group (P<0.05). Moreover, hCIK cells (1x10(7), once a week for four weeks) and TMZ (2.5 mg/kg, daily for 5 days) combination treatment further increased tumor cell apoptosis and decreased tumor cell proliferation and vessel density (P<0.05), creating a more potent therapeutic effect (95% reduction in tumor volume) compared with either hCIK cells or TMZ single therapy (72% for both, P<0.05). Taken together, CIK cell-immunotherapy and TMZ chemotherapy have synergistic therapeutic effects and could be combined for a successful treatment of GBM.

A clinical trial of cytokine induced killer cells treating patients with malignant tumor after radiochemotherapy

Objective Our study investigated the immunity changes and life quality changes after the treatment of cytokine induced killer (CIK) cells for patients with malignant tumor after radiochemotherapy, and explored the therapeutic effects of CIK cells on these patients.

Coculturing Dendritic Cells with Zoledronate Acid Efficiently Enhance the Anti-Tumor Effects of Cytokine-Induced Killer Cells

Dendritic cells (DCs) have greater stimulating activity on innate and adaptive immunity following short-term sensitization with zoledronate acid (DCs(Zol)). We identified the phenotype, cytotoxicity, and mechanisms of killing of cytokine-induced killer (CIK) cells which were cocultured with DCs(Zol). Adherent and nonadherent cells of peripheral blood mononuclear cell from myeloma patients were incubated for DCs and CIK cells. Then, the CIK cells were cocultured with DCs(Zol) (DCs(Zol)-CIK). Expression of markers for DCs(Zol)-CIK cells was measured using flow cytometry. Cytotoxicity was evaluated by against human myeloma cell lines and mechanisms of killing were tested by selectively blocking NKG2D receptor. The anti-tumor activity of these effector cells was further evaluated using a nude mice tumor model. gammadelta TCR expression of CIK cells significantly increased after coculture with immature or mature DCs(Zol) (iDCs/mDCs(Zol)-CIK) and these cells aggressively lysed myeloma cells compared with mDCs-CIK and zoledronate acid pulsed CIK cells (CIK(Zol); 50.8 +/- 7.9% and 48.2 +/- 4.7% versus 31.9 +/- 5.1% and 20.5 +/- 3.6%, effector versus target ratio was 60:1). Both alphabeta T and gammadelta T cells in the iDCs(Zol)-CIK cells performed the majority of lysis. The iDCs/mDCs(Zol)-CIK cells greatly increased NKG2D expression compared with mDCs-CIK and CIK(Zol) during culture (71.5 +/- 11.3% and 67.7 +/- 9.3% versus 51.3 +/- 6.2% and 47.1 +/- 5.7%). iDCs(Zol)-CIK cell-mediated lysis dropped 69.21% when the NKG2D receptor was blocked and the cytotoxicity correlated with NKG2D ligand-MICA expression on the target cells. In a human myeloma bearing nude mice model, iDCs(Zol)-CIK and mDCs(Zol)-CIK cells treatment groups obtained 75% and 62.5% long-term survival (>120 days) respectively, as compared with none of the control animals or 37.5% treated with mDCs-CIK cells. Large numbers of CIK cells with greater anti-tumor activities are rapidly generated by Zol-treated iDCs/mDCs. This strategy is worthy of further investigation to improve adoptive cell therapy against tumors.

Efficient lysis of rhabdomyosarcoma cells by cytokine-induced killer cells: implications for adoptive immunotherapy after allogeneic stem cell transplantation

Rhabdomyosarcoma is the most common soft tissue sarcoma in childhood and has a poor prognosis. Here we assessed the capability of ex vivo expanded cytokine-induced killer cells to lyse both alveolar and embryonic rhabdomyosarcoma cell lines and investigated the mechanisms involved. Peripheral blood mononuclear cells from six healthy donors were used to generate and expand cytokine-induced killer cells. The phenotype and composition of these cells were determined by multiparameter flow cytometry, while their cytotoxic effect against rhabdomyosarcoma cells was evaluated by a europium release assay. Cytokine-induced killer cells efficiently lysed cells from both rhabdomyosarcoma cell lines. Antibody-mediated masking of either NKG2D molecule on cytokine-induced killer cells or its ligands on rhabdomyosarcoma cells (major histocompatibility antigen related chain A and B and UL16 binding protein 2) diminished this effect by 50%, suggesting a major role for the NKG2D molecule in rhabdomyosarcoma cell killing. No effect was observed after blocking CD11a, CD3 or TCRalphabeta molecules on cytokine-induced killer cells or CD1d on rhabdomyosar-coma cells. Remarkably, cytokine-induced killer cells used tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) to activate caspase-3, as the main caspase responsible for the execution of apoptosis. Accordingly, blocking TRAIL receptors on embryonic rhabdomyosarcoma cell lines significantly reduced the anti-tumor effect of cytokine-induced killer cells. About 50% of T cells within the cytokine-induced killer population had an effector memory phenotype, 20% had a naïve phenotype and approximately 30% of the cells had a central memory phenotype. In addition, cytokine-induced killer cells expressed low levels of activation-induced markers CD69 and CD137 and demonstrated a low alloreactive potential. Our data suggest that cytokine-induced killer cells may be used as a novel adoptive immunotherapy for the treatment of patients with rhabdomyosarcoma after allogeneic stem cell transplantation.