Objective To clarify the effect and mechanism of tumor antigen-loaded dendritic cells (Ag-DCs) combined with cytokine-induced killers (CIKs) on the killing of esophageal cancer tumor cells. Methods Peripheral blood DCs and CIKs were induced and cultured, and the DCs were loaded with tumor antigen to obtain Ag-DCs, and Ag-DCs were co-cultured with CIKs. The experiment was divided into CIK group, DC combined with CIK group, Ag-DC combined with CIK group. Flow cytometry was used to detect the phenotype of cells. MTT assay was employed to determine the killing activity against EC9706 cells. Annexin V-FITC/PI double staining was used to detect the apoptosis rate of cells, immunofluorescence staining to detect the expression of phosphorylated apoptotic signal-regulated kinase 1 (p-ASK1) and Western blot analysis to detect the expression of ASK1 pathway related proteins. A nude mouse model of esophageal cancer transplantation tumor was constructed and divided into control group, DC combined with CIK group and Ag-DC combined with CIK group. The corresponding immune cells were injected into the tail vein for treatment and the tumor volume was measured every 2 days. After 21 days, all nude mice were sacrificed with the tumors taken out. HE staining was used to observe the tumor pathological changes and immunohistochemical staining was performed to detect the expression of ki67 and ASK1 in the tumor tissue. Results Comparedwith the CIK group alone and the DC combined with CIK group, the ratio of CD3⁺ CD8⁺ and CD3⁺ CD56⁺ in the cells significantly increased after Ag-DCs and CIKs co-culture, along with the increased killing rate of EC9706 cells, increased apoptosis rate of EC9706 cells, and the improved activation level of ASK1. Compared with the CIK group and the DC combined with CIK group, the growth of the transplanted tumor in nude mice treated with Ag-DCs combined with CIKs was significantly inhibited, and after 21 days, it was observed that the tumor tissue mass in this group was relatively smaller, with sparsely arranged cells in the tumor tissue and a decline in the positive rate of ki67 in tumor tissue, while the positive rate of ASK1 was significantly increased. Conclusion Co-cultivation of tumor antigen-loaded DCs with CIKs can significantly increase the killing activity of esophageal cancer tumor cells. The mechanism of action may be related to the activation of the ASK1 pathway.
Animals ; Humans ; Mice ; Antigens, Neoplasm ; Cytokine-Induced Killer Cells ; Cytokines/metabolism* ; Cytotoxicity, Immunologic ; Dendritic Cells ; Esophageal Neoplasms/therapy* ; Ki-67 Antigen ; Mice, Nude

BACKGROUND: Asbestos fibers possess tumorigenicity and are thought to cause mesothelioma. We have previously reported that exposure to asbestos fibers causes a reduction in antitumor immunity. Asbestos exposure in the mixed lymphocyte reaction (MLR) showed suppressed induction of cytotoxic T lymphocytes (CTLs), accompanied by a decrease in proliferation of CD8 METHODS: For MLR, human peripheral blood mononuclear cells (PBMCs) were cultured with irradiated allogenic PBMCs upon exposure to chrysotile B asbestos at 5 μg/ml for 7 days. After 2 days of culture, IL-15 was added at 1 ng/ml. After 7 days of MLR, PBMCs were collected and analyzed for phenotypic and functional markers of CD8 RESULTS: IL-15 addition partially reversed the decrease in CD3 CONCLUSION These findings indicate that CTLs induced upon exposure to asbestos possess dysfunctional machinery that can be partly compensated by IL-15 supplementation, and that IL-15 is more effective in the recovery of proliferation and granzyme B levels from asbestos-induced suppression of CTL induction compared with IL-2.
Asbestos/adverse effects* ; CD8-Positive T-Lymphocytes/metabolism* ; Humans ; Interleukin-15/pharmacology* ; Lymphocyte Activation/immunology* ; T-Lymphocytes, Cytotoxic/metabolism*

The β2m (Beta-2-microglobin) gene encodes a non-glycosylated protein that functions as an important component of major histocompatibility complexⅠ(MHCⅠ) for antigen presentation. To evade immune mediated clearance, human tumors and pathogens have adopted different strategies, including loss of MHCⅠexpression. Appropriate animal models are essential for understanding the mechanisms underpinning the clinical treatment of tumor and other human diseases. We constructed β2m knockout mice using CRISPR/Cas9 gene editing tool through embryo microinjection. Subsequently, genotyping and phenotyping of knockout mice were performed by PCR, qPCR, and flow cytometry. Mice genotyping showed that the coding region of the target gene was absent in the knockout mice. Real time PCR showed that mRNA level of β2m was significantly downregulated. Flow cytometry showed that the proportions of CD8+ killer T cells was significantly reduced in a variety of tissues and organs of the immune system. Taken together, we have successfully constructed a strain of β2m knockout mice, which will facilitate subsequent in vivo study on the function and mechanism of the β2m gene.
Animals ; Histocompatibility Antigens Class I ; Mice ; Mice, Inbred C57BL ; Mice, Knockout ; T-Lymphocytes, Cytotoxic ; beta 2-Microglobulin/genetics*

Objective: To evaluate the safety and effectiveness of a vaccine based on latent membrane protein 2 (LMP2) modified dendritic cells (DCs) that boosts specific responses of cytotoxic T lymphocytes (CTLs) to LMP2 before and after intradermal injection in patients with nasopharyngeal carcinoma (NPC). Methods: DCs were derived from peripheral blood monocytes of patients with NPC. We prepared LMP2-DCs infected by recombinant adenovirus vector expressing LMP2 (rAd-LMP2). NPC patients were immunized with 2 × 10 Results: We demonstrated that DCs derived from monocytes displayed typical DC morphologies; the expression of LMP2 in the LMP2-DCs vaccine was confirmed by immunocytochemical assay. Twenty-nine patients with NPC were enrolled in this clinical trial. The LMP2-DCs vaccine was well tolerated in all of the patients. Boosted responses to LMP2 peptide sub-pools were observed in 18 of the 29 patients with NPC. The follow-up data of 29 immunized patients from April, 2010 to April 2015 indicated a five-year survival rate of 94.4% in responders and 45.5% in non-responders. Conclusion In this pilot study, we demonstrated that the LMP2-DCs vaccine is safe and effective in patients with NPC. Specific CTLs responses to LMP2 play a certain role in controlling and preventing the recurrence and metastasis of NPC, which warrants further clinical testing.
Adult ; Aged ; Cancer Vaccines/therapeutic use* ; China ; Dendritic Cells/immunology* ; Female ; Humans ; Immunotherapy/methods* ; Injections, Intradermal ; Male ; Middle Aged ; Nasopharyngeal Carcinoma/therapy* ; Nasopharyngeal Neoplasms/therapy* ; T-Lymphocytes, Cytotoxic/immunology* ; Viral Matrix Proteins/therapeutic use* ; Young Adult

PURPOSE: The influenza B virus diverges into two antigenically distinct lineages: B/Yamagata and B/Victoria. Influenza B is the dominant circulating virus during some influenza seasons, and recent data demonstrated that influenza A and B infection similarly cause severe clinical symptoms in hospitalized patients. Nucleoprotein (NP) is a good target for a universal influenza vaccine. This study investigated whether NP epitope variation within two lineages affects the dominant cytotoxic T lymphocyte (CTL) responses induced by vaccination and the resultant protective immunity. MATERIALS AND METHODS: The NP of B/Yamagata/16/1988, the representative strain of the Yamagata lineage, includes a dominant CTL epitope, FSPIRITFL, while B/Shangdong/7/1997 from the Victoria lineage has one amino acid difference in this sequence, FSPIRVTFL. Two recombinant replication-deficient adenovirus (rAd)-vectored vaccines expressing either NP were prepared (rAd/B-NP(I) and rAd/B-NP(V), respectively) and administered to BALB/c mice intranasally. To examine the efficacy of vaccination, antibody responses, CTL responses, and morbidity/mortality after challenge were measured. RESULTS: Both vaccines induce similar antibody and CD8 T-cell responses cross-reacting to both epitopes, and also confer cross-protection against both lineages regardless of amino acid difference. CONCLUSION: The rAd-vectored vaccine expressing the NP could be developed as universal influenza B vaccine which provides broader protection.
Adenoviridae ; Animals ; Antibody Formation ; Epitopes ; Humans ; Influenza B virus* ; Influenza Vaccines ; Influenza, Human* ; Lymphocytes ; Mice ; Nucleoproteins* ; Seasons ; T-Lymphocytes ; T-Lymphocytes, Cytotoxic* ; Vaccination ; Vaccines ; Victoria

OBJECTIVE: To investigate the safety and clinical efficacy of autologous DC-CIK cells combined with other immune cells for patients with hematological malignancies and analyze patient prognosis. METHODS: 50 patients with hematological malignancies who received cellular immunotherapy from September 2014 to April 2016 were retrospectively studied in the First Affiliated Hospital of Xi'an Jiaotong University, 115 cases times of cellular immunotherapy were performed. According to the selected treatment, the patients were divided into the dual cell group (DC-CIK cell treatment) and the multi-cell group (DC-CIK cell combined with other immune cells); According to the treatment course, the patients were divided into the single course group (completed by ＜3 times) and the multiple course group. The changes of T lymphocyte subsets, blood routine indicators and KPS scores as well as the overall survival time before and after treatment were compared and analyzed. RESULTS: [WTB1]The difference of general conditions before treatment including the number of patients, sex, age, T lymphocyte subsets, blood routine indicators, KPS scores and so on in 2 groups divided according to 2 kinds of treatment methods were not statistically significant, indicating that the 2 groups were comparable. Grouped by selected treatment, the CD4/CD8 ratio, Hb and Plt levels decreased in the dual cell group, compared with those before treatment(P＜0.05). The CD3CD4 ratio after treatment in multiple cell group decreased, compared with that before treatment (P＜0.05). The 3-year survival rate of patients in dual cell and multiple cell groups was 61.3% vs 69.8%, the overall survival time of patients in 2 groups was 32.4 months vs 39.6 months, there were no statisticall differences between 2 groups(P>0.05). Grouped by treatment course, the CD3 ratio after treatment increased, while the Hb level after treatment decreased in single course group, compared with level before treatment(P＜0.05). The CD3CD4 ratio, Plt level decreased, while the KPS scores increased after treatment in multiple course group, compared with those before treatment(P＜0.05). The 3-year survival rate in single course and multiple course groups was 52% vs 76.4%, the overall survival time was 28.7 months vs 40.9 months respectively, statistically significant with difference (P＜0.05). CONCLUSION Autologous DC-CIK cells combined with other immune cells in the treatment of hematological malignancies can change the immune function of the patients and improve the antitumor activity. The multi-course treatment can improve the quality of life, prolong the overall survival time, thus worthing clinical promotion.
Cytokine-Induced Killer Cells ; Dendritic Cells ; Hematologic Neoplasms ; Humans ; Immunotherapy, Adoptive ; Quality of Life ; Retrospective Studies ; Treatment Outcome

Systemic target therapeutic drugs, such as sorafenib, lenvatinib, or regorafenib are the only drugs that are known to be effective against advanced hepatocellular carcinoma (HCC). However, these agents show a limited efficacy in killing residual tumors. Immunotherapy is an alternative approach to this treatment and has been used to successfully treat different cancers, including HCC. HCC is an inflammation-induced cancer and represents a very interesting target for immunotherapeutics. Immunotherapies aim to reverse the immune tolerance and suppression found in tumor microenvironments and include approaches, such as adoptive cell therapy, immune checkpoint inhibition, and cancer vaccination. Adoptive cell therapy uses autologous natural killer or cytokine-induced killer cells by cultivating them ex vivo and subsequently reinfusing them into the patient. Immune checkpoint inhibitors reactivate tumor-specific T cells by suppressing checkpoint-mediated inhibitory signaling. Cancer vaccination induces a tumor-specific immune response by activating effector T lymphocytes. A wide range of potential immunotherapy-related adverse events occur; therefore, a multidisciplinary collaborative management is required across the clinical spectrum. This review summarizes the current status of immunotherapy for HCC and provides a perspective on its future applications.
Carcinoma, Hepatocellular ; Cell- and Tissue-Based Therapy ; Cytokine-Induced Killer Cells ; Homicide ; Humans ; Immune Tolerance ; Immunotherapy ; Neoplasm, Residual ; Oncolytic Viruses ; T-Lymphocytes ; Tumor Microenvironment ; Vaccination

In recent years, with the incidence of malignant tumors increasing year by year, its treatment has been made great progress on the basis of traditional treatments such as surgery, radiotherapy and chemotherapy, it mainly focuses on rapid development of cellular immunotherapy. The efficacy of dendritic cells combined with cytokine induced killer cell (DC-CIK) immunotherapy for cancer patients is positive, it shows good antitumor activities and the abilities to reconstruct and enhance the immune system of tumor patients in clinical application, it has potential application value. In the treatment of hematologic malignancies which are chemotherapy-based and high-grade malignant, what is the effect of DC-CIK cells immunotherapy for them? In this review, we searched Chinese and abroad literatures from 2012 to 2018 and further focused on 18 clinical research articles about hematologic malignancies in order to analyze the characteristics of DC-CIK cells immunotherapy. It was found that DC-CIK cells can be an effective and promising treatment for patients with hematologic malignancies, and thus, if the process and unified plan are further standardized and improved, the efficacy will be more obvious. For this purpose, this paper reviews the biological characteristic of DC cells, CIK cells and the clinical research progress of DC-CIK cell immunotherapy for hematological malignancies.
Combined Modality Therapy ; Cytokine-Induced Killer Cells ; Dendritic Cells ; Hematologic Neoplasms ; Humans ; Immunotherapy ; Immunotherapy, Adoptive

POEMS syndrome is a rare multiple organ involvement of the parasympathetic syndrome associated with abnormal plasma cells, mostly with high-dose chemotherapy and stem cell transplantation for the treatment. Recently, more treatment attempts to treat POEMS syndrome have been utilized so as to improve the efficacy and safety for the patients with POEMS syndrome, such as immunomodulator, alkylating agent, cytokine-induced killer cells and so on. Lenalidomide has a significant effect on relapse/refractory POEMS syndrome and patients with endocrinopathy. Cytokine-induced killer cells are also a safe and effective regimen for the treatment of POEMS syndrome. This review described the efficacy and safety of immunomodulatos, alkylators, cytokine-induced killer cells, ASCT, proteasome inhibitors and monoclonal antibodies for POEMS syndrome, and the newest clinical research and progress of POEMS syndrome ware summarized briefly.
Cytokine-Induced Killer Cells ; Humans ; Immunologic Factors ; POEMS Syndrome ; Stem Cell Transplantation ; Transplantation, Autologous

OBJECTIVETo investigate the efficiency of inducing CIK from peripheral blood mononuclear cells(PBMNC) by using immune cell serum replacement(immune cell SR), so as to provide a new strategy for the industrialized production of immune cells.

METHODSThe PBMNC of healthy volunteers were collected, and these cells were thawed after short-term cryopreservation and cultured to induce CIK cells. The cells viability was measured by trypan blue exclusion, the phenotypes were analyzed by flow cytometry, and the cytotoxicity was determined by Calcein-AM/PI double staining.

RESULTSIn cryopreserved PBMNC, the control group cells failed to normally proliferate. Cell proliferation ratio was low in 2% SR group in comparison with the fresh group, and the difference was significant (P<0.05), however, differences were not statistically significant between 5% SR and fresh group or between 10% AP and fresh group. CD3, CD3CD8 and CD3CD56 cell subsets were not significantly different before and after cryopreservation (P>0.05). After being cultured, CD3, CD3CD4, CD3CD8, CD3CD56 and CD3CD56 subsets and the cytotoxicity in vitro were not significantly different among all group(P>0.05).

CONCLUSION5% SR without the protein of animal origin can be safely used as a substitute for autologous plasma in CIK induced from cryopreserved PBMNC by culture, thus providing a basis for the application of cryopreservation technique of immune cells to cell therapy.