Cytokine-induced Killer Research Articles

Three-dimensional dynamics of mesothelin-targeted CAR.CIK lymphocytes against ovarian cancer peritoneal carcinomatosis

Intraperitoneal cellular immunotherapy with CAR-redirected lymphocytes is an intriguing approach to target peritoneal carcinomatosis (PC) from ovarian cancer (OC), which is currently evaluated in clinical trials. PC displays a composite structure with floating tumor cells within ascites and solid-like masses invading the peritoneum. Therefore, a comprehensive experimental model is crucial to optimize CAR-cell therapies in such a peculiar environment. Here, we explored the activity of cytokine-induced killer lymphocytes (CIK), redirected by CAR against mesothelin (MSLN-CAR.CIK), within reductionistic 3D models resembling the structural complexity of both liquid and solid components of PC. MSLN-CAR.CIK effectively killed and were functionally efficient against OC targets. In a “floating-like” 3D context with floating OC spheroids, both tumor localization and killing by MSLN-CAR.CIK were significantly boosted by fluid flow. In a “solid-like” context, MSLN-CAR.CIK were recruited through the extracellular matrix on embedded tumor aggregates, with variable kinetics depending on the effector-target distance. Furthermore, MSLN-CAR.CIK penetrated the inner levels of OC spheroids exerting effective tumor killing. Our findings provide currently unknown therapeutically relevant information on intraperitoneal approaches with CAR.CIK, supporting further developments and improvements for clinical studies in the context of locoregional cell therapy approaches for patients with PC from OC.

Immune Effect of Co-Culture of Dendritic Cells and Cytokine-Induced Killer Cells on Prostate Cancer Cells.

It was to explore the immune outcome of co-culture of dendritic cells (DC) and cytokine-induced killer cells (CIK) on prostate cancer (PCa) cells. Peripheral blood mononuclear cells (PBMCs) were extracted from healthy blood donors. DC and CIK cells were induced and co-cultured. The proliferation and phenotypic changes of DC, CIK, and DC-CIK cells/groups were observed. Model rats were constructed by injecting PC3 PCa cells into the abdominal cavity. The successful 50 cases were divided into a negative control group, a chemotherapy group, a DC group, a CIK group, and a DC-CIK treatment group (each consisting of 10 rats) to observe tumor progression. The secreted concentrations of interleukin-12 (IL-12) ((103.67 ± 2.77) pg/mL) and interferon-γ (IFN-γ) ((730.09 ± 23.52) pg/mL) were higher in DC-CIK group as against DC and CIK groups; the proliferation of CIK was higher in DC-CIK group as against CIK within 12 to 20 days of culture. The positive rate (PR) of CD3+/ CD56+ and CD8+ was higher and that of CD45RA+ was lower in DC-CIK group as against CIK.The killing rate of the DC-CIK group was higher than that of the DC and CIK groups at a target effect ratio of 10:1/20:1/50:1 (P < 0.05). After the treatment, the body weight of rats in the chemotherapy group, DC group, and CIK group was significantly reduced (P < 0.05), while no significant changes were observed in the negative control group and DC-CIK group (P > 0.05). After 25 days of treatment, the tumor size in the DC-CIK group was significantly smaller compared to the negative control group, chemotherapy group, DC group, and CIK group; the necrotic area of the tumor tissue in the DC-CIK group was also significantly larger than that in the negative control group, chemotherapy group, DC group, and CIK group (P < 0.05). Co-culture of DC and CIK is excellent in enhancing the proliferation of CIK cells, increasing the secretion of IL-12 and IFN-γ, and enhancing the activity of immune cells and anti-tumor ability, showing its potential in anti-PCa tumor immunotherapy.

CXCR4 up-regulation mediated by USP1 deubiquitination promotes the tumorigenesis and immune escape in esophageal squamous-cell carcinoma.

CXC chemokine receptor 4 (CXCR4) and ubiquitin specific protease 1 (USP1) have been reported to involve in the tumorigenesis of esophageal squamous-cell carcinoma (ESCC). Here, we investigated whether USP1 induced CXCR4 deubiquitination in regulating ESCC progression. MTT assay, 5-ethynyl-2'-deoxyuridine assay, flow cytometry, transwell assay and ELISA analysis were used to detect cell oncogenic phenotypes, macrophage phenotypes, inflammatory cytokines production, the cytotoxicity of cytokine-induced killer (CIK) cells and CD8 + T cell apoptosis. Protein interaction was determined by immunoprecipitation assay. Cellular ubiquitination detected the ubiquitination effect on CXCR4. A mouse xenograft model was established for in vivo experiments. CXCR4 was highly expressed in ESCC tissues and cells. Functionally, CXCR4 silencing suppressed ESCC cell proliferation, invasion, and induced cell apoptosis. Moreover, CXCR4 deficiency suppressed cancer cell immune escape by suppressing macrophage M2 polarization, elevating inflammatory cytokines produced by PBMCs, enhancing the cytotoxicity of CIK cells, and suppressing CD8 + T cell apoptosis. A high USP1 expression was observed in ESCC, USP1 interacted with CXCR4 and enhanced its protein stability through deubiquitination. USP1 silencing suppressed ESCC cell proliferation, invasion, and immune escape, which were reversed by CXCR4 overexpression. In vivo assay showed that USP1 deficiency impeded tumor growth by regulating CXCR4. Besides, fused in sarcoma (FUS) was confirmed to bind to USP1 and stabilized its mRNA expression, and could regulate CXCR4 via USP1. In conclusion, USP1 stabilized CXCR4 by removing ubiquitination on CXCR4, thereby promoting ESCC cell proliferation, invasion, and immune escape in vitro, and tumor growth in vivo.

Intricate relationship between cancer stemness, metastasis, and drug resistance.

Cancer stem cells (CSCs) are widely acknowledged as the drivers of tumor initiation, epithelial-mesenchymal transition (EMT) progression, and metastasis. Originating from both hematologic and solid malignancies, CSCs exhibit quiescence, pluripotency, and self-renewal akin to normal stem cells, thus orchestrating tumor heterogeneity and growth. Through a dynamic interplay with the tumor microenvironment (TME) and intricate signaling cascades, CSCs undergo transitions from differentiated cancer cells, culminating in therapy resistance and disease recurrence. This review undertakes an in-depth analysis of the multifaceted mechanisms underlying cancer stemness and CSC-mediated resistance to therapy. Intrinsic factors encompassing the TME, hypoxic conditions, and oxidative stress, alongside extrinsic processes such as drug efflux mechanisms, collectively contribute to therapeutic resistance. An exploration into key signaling pathways, including JAK/STAT, WNT, NOTCH, and HEDGEHOG, sheds light on their pivotal roles in sustaining CSCs phenotypes. Insights gleaned from preclinical and clinical studies hold promise in refining drug discovery efforts and optimizing therapeutic interventions, especially chimeric antigen receptor (CAR)-T cell therapy, cytokine-induced killer (CIK)cell therapy, natural killer (NK) cell-mediated CSC-targeting and others. Ultimately use of cell sorting and single cell sequencing approaches for elucidating the fundamental characteristics and resistance mechanisms inherent in CSCs will enhance our comprehension of CSC and intratumor heterogeneity, which ultimately would inform about tailored and personalized interventions.

1322P A phase II trial of autologous cytokine-induced killer (CIK) therapy plus toripalimab with or without chemotherapy as first-line treatment in advance NSCLC

1322P A phase II trial of autologous cytokine-induced killer (CIK) therapy plus toripalimab with or without chemotherapy as first-line treatment in advance NSCLC

Functional Activity of Cytokine-Induced Killer Cells Enhanced by CAR-CD19 Modification or by Soluble Bispecific Antibody Blinatumomab.

Strategies to increase the anti-tumor efficacy of cytokine-induced killer cells (CIKs) include genetic modification with chimeric antigen receptors (CARs) or the addition of soluble T-cell engaging bispecific antibodies (BsAbs). Here, CIKs were modified using a transposon system integrating two distinct anti-CD19 CARs (CAR-MNZ and CAR-BG2) or combined with soluble CD3xCD19 BsAb blinatumomab (CIK + Blina). CAR-MNZ bearing the CD28-OX40-CD3ζ signaling modules, and CAR-BG2, designed on the Tisagenlecleucel CAR sequence (Kymriah®), carrying the 4-1BB and CD3ζ signaling elements, were employed. After transfection and CIK expansion, cells expressed CAR-CD19 to a similar extent (35.9% CAR-MNZ and 17.7% CAR-BG2). In vitro evaluations demonstrated robust proliferation and cytotoxicity (~50% cytotoxicity) of CARCIK-MNZ, CARCIK-BG2, and CIK + Blina against CD19+ target cells, suggesting similar efficacy. All effectors formed an increased number of synapses, activated NFAT and NFkB, and secreted IL-2 and IFN-ɣ upon encountering targets. CIK + Blina displayed strongest NFAT and IFN-ɣ induction, whereas CARCIK-BG2 demonstrated superior synapse formation. All the effectors have shown therapeutic activity in vivo against the CD19+ Daudi tumor model, with CARCIK cells showing a more durable response compared to CIK + Blina, likely due to the short half-life of Blina in this model.

Generation of dual-attribute iTNK cells from hPSCs for cancer immunotherapy

Dual-attribute immune cells possess advantageous features of cytotoxic Tcells and natural killer (NK) cells and hold promise for advancing immunotherapy. Dual-attribute cell types such as invariant natural killer Tcells, induced T-to-NK cells, and cytokine-induced killer cells have demonstrated efficacy and safety in preclinical and clinical studies. However, their limited availability hinders their widespread application. Human pluripotent stem cells (hPSCs) offer an ideal source. Here, we generate dual-attribute induced T-NK (iTNK) cells from hPSCs, expressing markers of both cytotoxic T and NK cells. Single-cell RNA and Tcell receptor (TCR) sequencing analyses reveal that iTNK cells expressed signature genes associated with both NK and Tcells and displayed a diverse TCR repertoire. iTNK cells release cytotoxic mediators, exert cytotoxicity against diverse tumor cell lines, and inhibit tumor growth invivo. By harnessing adaptive and innate immune responses, hPSC-derived iTNK cells offer promising strategies for cancer immunotherapy.

Efficacy and Safety of Autologous Cytokine-Induced Killer (CIK) Cellular Immunotherapy Combined with Chemotherapy in Non-Small-Cell Carcinoma (NSCLC): A Meta-Analysis.

To investigate the efficacy and safety of autologous Cytokine-induced killer (CIK) cellular immunotherapy combined with chemotherapy for non-small-cell carcinoma (NSCLC). The literature related to the efficacy and safety of autologous CIK cellular immunotherapy combined with chemotherapy in NSCLC were collected. The Chinese literature databases were collected from CNKI, Wanfang and Weipu, and the English literature were retrieved from Cochrane Library and PubMed. All literature were retrieved until the year of 2022, and evaluated by its method quality and analyzed by RevMan5.3 software. A Meta-analysis on the 11 included literature showed that the treatment efficacy and disease control rate (DCR) of patients in the observation group were significantly higher than those in the control group (P < 0.05), and significantly increased the levels of CD3+, CD4+, CD4+/CD8+ indexes than the control group (P < 0.05), and the incidence of bone marrow suppression, liver injury and gastrointestinal symptoms were significantly lower than those of the control group (P < 0.05). Compared with single chemotherapy, CIK combined with chemotherapy in the treatment of patients with NSCLC can improve the efficacy of treatment and DCR, significantly improve their immune function, reduce the incidence of adverse reactions, and is beneficial to the recovery of prognosis.

Therapeutic effect of small extracellular vesicles from cytokine-induced memory-like natural killer cells on solid tumors

Small extracellular vesicles (sEV) derived from diverse natural killer (NK) cell lines have proven their exceptional antitumor activities. However, sEV from human primary NK cells, especially memory-like NK cells, are rarely utilized for cancer treatment. In this study, we obtained sEV from IL-12, IL-15 and IL-18 cultured human memory-like NK cells (mNK-sEV) that showed strong cytokine-secretory ability. It was uncovered that mNK-sEV entered cancer cells via macropinocytosis and induced cell apoptosis via caspase-dependent pathway. Compared to sEV from conventionally cultured NK cells (conNK-sEV), mNK-sEV inhibited tumor growth to a greater extent. Concomitantly, pharmacokinetics and biodistribution results validated a higher accumulation of mNK-sEV than conNK-sEV in tumors of xenografted murine models. Notably, elevated containment of granulysin (GNLY) within mNK-sEV, at least in part, may contribute to the enhanced therapeutic effect. Herein our results present that mNK-sEV can be a novel class of therapeutic reagent for effective cancer treatment.Graphical

Advances and clinical applications of immune checkpoint inhibitors in hematological malignancies.

Immune checkpoints are differentially expressed on various immune cells to regulate immune responses in tumor microenvironment. Tumor cells can activate the immune checkpoint pathway to establish an immunosuppressive tumor microenvironment and inhibit the anti-tumor immune response, which may lead to tumor progression by evading immune surveillance. Interrupting co-inhibitory signaling pathways with immune checkpoint inhibitors (ICIs) could reinvigorate the anti-tumor immune response and promote immune-mediated eradication of tumor cells. As a milestone in tumor treatment, ICIs have been firstly used in solid tumors and subsequently expanded to hematological malignancies, which are in their infancy. Currently, immune checkpoints have been investigated as promising biomarkers and therapeutic targets in hematological malignancies, and novel immune checkpoints, such as signal regulatory protein α (SIRPα) and tumor necrosis factor-alpha-inducible protein 8-like 2 (TIPE2), are constantly being discovered. Numerous ICIs have received clinical approval for clinical application in the treatment of hematological malignancies, especially when used in combination with other strategies, including oncolytic viruses (OVs), neoantigen vaccines, bispecific antibodies (bsAb), bio-nanomaterials, tumor vaccines, and cytokine-induced killer (CIK) cells. Moreover, the proportion of individuals with hematological malignancies benefiting from ICIs remains lower than expected due to multiple mechanisms of drug resistance and immune-related adverse events (irAEs). Close monitoring and appropriate intervention are needed to mitigate irAEs while using ICIs. This review provided a comprehensive overview of immune checkpoints on different immune cells, the latest advances of ICIs and highlighted the clinical applications of immune checkpoints in hematological malignancies, including biomarkers, targets, combination of ICIs with other therapies, mechanisms of resistance to ICIs, and irAEs, which can provide novel insight into the future exploration of ICIs in tumor treatment.

Cytokine-induced killer cells-mediated chlorin e6-loaded gold nanostars for targeted NIR imaging and immuno-photodynamic combination therapy for lung cancer

Recently, cytokine-induced killer (CIK) cells have a broad application prospect in the comprehensive diagnosis and treatment of tumors owing to their unique characteristics of killing and targeting malignant tumors. Herein, we report a facile strategy for synthesis of monodisperse gold nanostars (GNSs) based on PEGylation and co-loaded with the photosensitizer chlorin e6 (Ce6) to form GNSs-PEG@Ce6 NPs. Then employing CIK cells loading the as-prepared GNSs-PEG@Ce6 NPs to fabricate a CIK cells-based drug delivery system (GNSs-PEG@Ce6-CIK) for lung cancer. Among them, GNSs was functioned as transport media, Ce6 acted as the near-infrared (NIR) fluorescence imaging agent and photodynamic therapy (PDT), and CIK cells served as targeting vectors for immunotherapy, which can increase the efficiency of tumor enrichment and treatment effect. The results of cellular experiments demonstrated that GNSs-PEG@Ce6 NPs had good dispersibility, water solubility and low toxicity under physiological conditions, and the cultured CIK cells had strong anti-tumor properties. Subsequently, GNSs-PEG@Ce6-CIK could effectively inhibit the growth of A549 cells under the exposure of 633 nm laser, which showed stronger killing effect than that of GNSs-PEG@Ce6 NPs or CIK cells. In addition, they showed good tumor targeting and tumor synergistic killing activity in vivo. Therefore, GNSs-PEG@Ce6-CIK was constructed for targeted NIR fluorescence imaging, enhanced PDT and immunotherapy of lung cancer.

Optimization of Cytokine-Induced Killer Cell Generation from Mouse Spleen Cells

Optimization of Cytokine-Induced Killer Cell Generation from Mouse Spleen Cells

APPLICATION OF BIOMEDICAL CELL PRODUCTS BASED ON NATURAL KILLER CELLS AND CYTOKINE-INDUCED KILLER CELLS FOR THE TREATMENT OF ONCOLOGICAL DISEASES

Cellular immunotherapy has become an essential part of modern concepts of oncology treatment. One of the promising tool of immunotherapy in the treatment of cancer is the use of biomedical cell products (BMCP) based on natural killer cells (NK-cells) and cytokine-induced killer cells (CIKc). NK-cells are a unique population of innate lymphoid cells that identify tumor cells and play a key role in anticancer immunity through their cytotoxicity mechanisms. CIKc share the immunofunctional properties both of T-lymphocytes and NK cells; they do not require antigen-specific priming to recognize tumor cells and can be quickly cultured in vitro. The purpose of this review is to evaluate the available literature describing adoptive immunotherapy for BMCP based on NK-cells and CIKc.

Circ_0027791 contributes to the growth and immune evasion of hepatocellular carcinoma via the miR-496/programmed cell death ligand 1 axis in an m6A-dependent manner.

Emerging evidence indicates the critical roles of circular RNAs in the development of multiple cancers, containing hepatocellular carcinoma (HCC). Herein, our present research reported the biological function and mechanism of circ_0027791 in HCC progression. Circ_0027791, microRNA-496 (miR-496), programmed cell death ligand 1 (PDL1), and methyltransferase-like 3 (METTL3) levels were detected by real-time quantitative polymerase chain reaction (RT-qPCR). Cell viability, proliferation, invasion, and sphere formation ability were detected using 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide, 5-ethynyl-2'-deoxyuridine, transwell, and sphere formation assays. Macrophage polarization was detected using flow cytometry assay. To understand the role of circ_0027791 during the immune escape, HCC cells were cocultured with peripheral blood mononuclear cells or cytokine-induced killer (CIK) cells in vitro. A xenograft mouse model was applied to assess the function of circ_0027791 in vivo. After prediction using circinteractome and miRDB, the binding between miR-496 and circ_0027791 or PDL1 was validated based on a dual-luciferase reporter assay. Interaction between METTL3 and circ_0027791 was determined using methylated RNA immunoprecipitation (MeRIP)-qPCR, RIP-qPCR, and RNA pull-down assays. Circ_0027791, PDL1, and METTL3 expression were upregulated, and miR-496 was decreased in HCC patients and cells. Moreover, circ_0027791 knockdown might repress proliferation, invasion, sphere formation, M2 macrophage polarization, and antitumor immune response. Circ_0027791 knockdown repressed HCC tumor growth in vivo. In mechanism, circ_0027791 functioned as a sponge for miR-496 to increase PDL1 expression. In addition, METTL3 mediated the m6A methylation of circ_0027791 and stabilized its expression. METTL3-induced circ_0027791 facilitated HCC cell progression partly regulating the miR-496/PDL1 axis, which provided a new prognostic and therapeutic marker for HCC.

Abstract 3972: Personalized immune-organoid co-culture system for modeling the anti-tumor immune response in breast cancer patients

Abstract Recent studies have reported the effectiveness of immune checkpoint blockade (ICB) therapy in breast cancer patients. However, only a subset of patients benefits from ICB therapy due to various immunosuppressive mechanisms. The development of personalized predictive models for the anti-tumor immune response is essential to broaden the indication of ICB therapy. Here, we suggest a novel model platform using a co-culture system with patient tumor-derived organoids and matched immune cells to predict the immune response of individual patients. We successfully established fifty-nine cases of patient-derived breast cancer organoids with matched peripheral blood mononuclear cells (PBMCs) (subtype: Luminal A, 18.6%; Luminal B, 28.8%; Triple-negative breast cancer, 35.6%; HER2-positive, 17.0%). Three rapidly proliferating organoids (one Luminal B and two TNBC organoids) were selected to develop an immune-organoid co-culture (IOC) system. To mimic the anti-tumor immune response, cytotoxic immune cells were expanded from PBMCs by modified cytokine-induced killer (CIK) cell expansion protocol utilizing IL-2/IL-15. The CIK cells were co-cultured with patient-derived tumor organoids for killing assay to enable the prediction of ICB responsiveness in vitro. The CIK cells were expanded in the IOC system by stimulation of PBMC with IL-2/IL-15 in the presence of matched tumor organoids for 11 to 14 days. The CIK cells exhibited tumor organoid-specific killing (measured by Caspase-3, 15.22% ± 8.58, mean ± SEM) on organoid co-culture and showed induction of CD107a and CD137 expression in CD8+ T cells and NK cells with the PD-1 blocking antibody. The CIK-mediated tumor organoid deaths and immune cell activation were reduced by HLA class I blocking antibody (W6/32) treatment, confirming HLA-TCR mediated immune response and killing of tumor organoids in the IOC system (-14.85% in organoid-specific lysis and -7.46% in CD107a expression). In the live cell imaging of the IOC system, the interaction between T cells and tumor organoid cells and subsequent induction of Caspase 3 was noted. Our IOC system could directly measure the immunogenic tumor organoid killing by immune cells on ICB treatment in individual breast cancer patients. Moreover, the analysis of immune receptor profiles may reveal immune modulating mechanisms in each patient. We expect the IOC system to serve as a useful platform for personalized immunotherapy and exploration of combinatory therapy to enhance the anti-tumor efficacy of ICB in breast cancer patients. Citation Format: Cheol-Hwa Hong, Won-Ji Ryu, Shinyoung Park, Tae Yeong Kim, Yumi Hwang, Hyunju Han, Seongyeon Jo, Taeyong Kweon, Gun Min Kim, Min Hwan Kim, Hyung Seok Park, Joo Hyuk Sohn. Personalized immune-organoid co-culture system for modeling the anti-tumor immune response in breast cancer patients [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 3972.

Abstract 5253: Epigenetic drugs partially restored anti-cholangiocarcinoma activity of dysfunctional CD3+CD56+ cells

Abstract The cholangiocarcinoma cell-exposed cytokine-induced killer (CIK) cells or their most efficacious CD3+CD56+ subset displayed weaken anti-tumor cytotoxicity upon subsequent exposure. RNAseq analysis of the resistant phenotype gave rise to a list of drugs that could potentially counter the activity of genes associated with resistance to the anti-tumor activity of CIK cells. The top-ranking drugs appeared to be epigenetic drugs. Sublethal concentration of these drugs were assayed for their synergistic activity with CIK cell or the CD3+CD56+ subset for cytolysis of resistant CCA cells in tissue culture model. These drugs, although in low concentration, contained anti-tumor activity. The tumor cytolysis was greater after the combination with either CIK cells or the CD3+CD56+ subset. An in vivo model of CCA-implanted mice exhibited the synergistic activity of subtherapeutic dose of these drugs and CD3+CD56+ subset in suppressing tumor growth. It is concluded that epigenetic drugs in subtherapeutic dose could prevent the dysfunction of CCA-exposed CIK cells. Citation Format: Adisak Wongkajornsilp, Porncheera Chusorn, Kriengkrai Phongkitkarun, Khin Su Su Htwe, Sunisa Duangsa-ard, Kanda Kasetsinsombat, Nathawadee Sawatpiboon. Epigenetic drugs partially restored anti-cholangiocarcinoma activity of dysfunctional CD3+CD56+ cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 5253.

Abstract 3595: A retrospective analysis of the clinical characteristics of metastastic solid cancer patients with cytokine-induced killer cells combined immune checkpoint inhibitor immunotherapy

Abstract Background: Cytokine-induced killer cells (CIKs), obtained through ex vivo expasion from peripheral blood mononuclear cells, are phenotypically and functionally shared with both T and NK cells. Within the T cell group, it can be classified into a group that simultaneously expresses CD3 and CD56 molecules (range: 30% to 70%) and a group representing CD3+ CD56- phenotype (range: 20% to 60%). It also contains a small population (&amp;lt; 30%) of CD3 CD56+ NK cells. They can exhibit potent MHC-unrestricted cytotoxicity for both hematologic and solid tumors, but they are not harmful in hematopoietic precursors and normal tissues. Anti-programmed death-1 (PD-1) antibodies reinvigorate T cell-mediated antitumor immunity. PD-1 blockade antibody might significantly increase the cytotoxic potency of CIK cells via increasing proliferative capacity and IFN-r secretion. This retrospective study is the first to combine CIK cells with PD-1/PD-L1/CTLA-4 blockade(Immune checkpoint inhibitors, ICIs) in patients with metastatic solid cancers. Methods: We analyzed 225 heavily treated metastastic solid cancer patients receiving CIK cell (Immunecell-LC, GC cell inc provide.) and with ICIs. Blood Sampling for CIK cells production was performed after 3 days after ICIs administration. Patients received a single intravenous infusion of activated autologous CIK cells every 2 weeks. ICIs was administered according to the protocol of each drug regardless of the CIK cells administration date. The primary end points were safety and adverse event (AE) profiles. Objective responses, overall survival (OS), progression-free survival (PFS), and diseae control rate were secondary end points. Results: Treatment-related AEs occurred in 154/225 patients. Grade 3 or 4 toxicities, including fever and chills, were observed in 28 patients. All treatment-related AEs were tolerable and no death due to autoimmune disease. When administration ICIs, CIK cells exhibited excellent antitumor properties and increased IFN-γ secretion. Objective responses (complete or partial responses) were observed in 67 of the 225 patients. One patients with hepatocellular carcionma had complete response. In most patients who showed an objective response, the effectiveness was maintained for more than 6 months. The overall disease control rate in the patients was 58.9%. At the time of this report, the median OS and PFS were 18.6 and 7.1 months in hepatocellular carcinoma patients and were 13.1 and 6.5 months in triple negative breast cancer patients, respectively. Conclusion: Treatment with activated autologous CIK cells and ICIs was safe and exerted encouraging antitumor activity in metastastic solid cancers. Phase 2 clinical studies for each tumor type will be conducted. Keywords: Cytokine-induced killer cells, Immune check inhibitors, Safety, Objective response Citation Format: Jonggwon Choi, Jong Yeup Kim, Dong Won Yang, Gun He Nam. A retrospective analysis of the clinical characteristics of metastastic solid cancer patients with cytokine-induced killer cells combined immune checkpoint inhibitor immunotherapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 3595.

Discovering single cannabidiol or synergistic antitumor effects of cannabidiol and cytokine-induced killer cells on non-small cell lung cancer cells.

A multitude of findings from cell cultures and animal studies are available to support the anti-cancer properties of cannabidiol (CBD). Since CBD acts on multiple molecular targets, its clinical adaptation, especially in combination with cancer immunotherapy regimen remains a serious concern. Considering this, we extensively studied the effect of CBD on the cytokine-induced killer (CIK) cell immunotherapy approach using multiple non-small cell lung cancer (NSCLC) cells harboring diverse genotypes. Our analysis showed that, a) The Transient Receptor Potential Cation Channel Subfamily V Member 2 (TRPV2) channel was intracellularly expressed both in NSCLC cells and CIK cells. b) A synergistic effect of CIK combined with CBD, resulted in a significant increase in tumor lysis and Interferon gamma (IFN-g) production. c) CBD had a preference to elevate the CD25+CD69+ population and the CD62L_CD45RA+terminal effector memory (EMRA) population in NKT-CIK cells, suggesting early-stage activation and effector memory differentiation in CD3+CD56+ CIK cells. Of interest, we observed that CBD enhanced the calcium influx, which was mediated by the TRPV2 channel and elevated phosphor-Extracellular signal-Regulated Kinase (p-ERK) expression directly in CIK cells, whereas ERK selective inhibitor FR180204 inhibited the increasing cytotoxic CIK ability induced by CBD. Further examinations revealed that CBD induced DNA double-strand breaks via upregulation of histone H2AX phosphorylation in NSCLC cells and the migration and invasion ability of NSCLC cells suppressed by CBD were rescued using the TRPV2 antagonist (Tranilast) in the absence of CIK cells. We further investigated the epigenetic effects of this synergy and found that adding CBD to CIK cells decreased the Long Interspersed Nuclear Element-1 (LINE-1) mRNA expression and the global DNA methylation level in NSCLC cells carrying KRAS mutation. We further investigated the epigenetic effects of this synergy and found that adding CBD to CIK cells decreased the Long Interspersed Nuclear Element-1 (LINE-1) mRNA expression and the global DNA methylation level in NSCLC cells carrying KRAS mutation. Taken together, CBD holds a great potential for treating NSCLC with CIK cell immunotherapy. In addition, we utilized NSCLC with different driver mutations to investigate the efficacy of CBD. Our findings might provide evidence for CBD-personized treatment with NSCLC patients.

Identification of the C1qDC gene family in grass carp (Ctenopharyngodon idellus) and the response of C1qA, C1qB, and C1qC to GCRV infection in vivo and in vitro

Proteins from the C1q domain-containing (C1qDC) family recognize self-, non-self-, and altered-self ligands and serves as an initiator molecule for the classical complement pathway as well as recognizing immune complexes. In this study, C1qDC gene family members were identified and analyzed in grass carp (Ctenopharyngodon idellus). Members of the C1q subfamily were cloned, and their response to infection with the grass carp virus was investigated. In the grass carp genome, 54 C1qDC genes and 67 isoforms have been identified. Most were located on chromosome 3, with 52 shared zebrafish homologies. Seven substantially differentially expressed C1qDC family genes were identified in the transcriptomes of cytokine-induced killer (CIK) cells infected with grass carp reovirus (GCRV), all of which exhibited sustained upregulation. The opening reading frames of grass carp C1qA, C1qB, and C1qC, belonging to the C1q subfamily, were determined to be 738, 732, and 735 base pairs, encoding 245, 243, and 244 amino acids with molecular weights of 25.81 kDa, 25.63 kDa and 26.16 kDa, respectively. Three genes were detected in the nine collected tissues, and their expression patterns were similar, with the highest expression levels observed in the spleen. In vivo after GCRV infection showed expression trends of C1qA, C1qB, and C1qC in the liver, spleen, and kidney. An N-type pattern in the liver and kidney was characterized by an initial increase followed by a decrease, with the highest expression occurring during the recovering period, and a V-type pattern in the spleen with the lowest expression levels during the death period. In vitro, after GCRV infection showed expression trends of C1qA, C1qB, and C1qC, and this gradually increased within the first 24 h, with a notable increase observed at the 24 h time point. After CIK cells incubation with purified recombinant proteins, rC1qA, rC1qB, and rC1qC for 3 h, followed by GCRV inoculation, the GCRV replication indicated that rC1qC exerted a substantial inhibitory effect on viral replication in CIK cells after 24 h of GCRV inoculation. These findings offer valuable insights into the structure, evolution, and function of the C1qDC family genes and provide a foundational understanding of the immune function of C1q in grass carp.

The Therapeutic Potential of Cytokine-Induced Killer in Patients with Cancer.

Despite the promising results of immunotherapy, further experiments need to be considered because of several factors ranging from physical barriers to off-tumor adverse effects. It is surprising that adoptive cellular immunotherapy, particularly dendritic cell and cytokine-induced killer (DC-CIK) therapy, is far less emphasized in the treatment of cancer diseases. DC-CIK therapy in cancer patients presents auspicious results with low or no side effects, which should not be overlooked. More interestingly, almost all DC-CIK clinical trials are ongoing in China that highlight the limitations of therapeutic strategies and require large-scale research. To date, it is advisable to consider combination therapy with chemotherapy since it has shown promising outcomes with higher efficacy. In this article, the efficacy of DC-CIK therapy in patients with cancer is summarized by underscoring the lack of experiments on soft cancers on an unprecedented scale. In brief, DC-CIK therapy is a safe and effective therapeutic agent for malignant and nonmalignant diseases that enhances short-term and long-term effects.