Jove-Journal of Visualized Experiments
·
2018
·
IF 1.184
Yanlan Wang,Jiayu Liu,Haitao Pan,Jieyu Xing,Xiaoqiong Wu,Qing Li,Zhong Wang
This protocol describes the construction and functional studies of a bispecific antibody (bsAb), GPC3-S-Fab. bsAbs can recognize two different epitopes through their two different arms. bsAbs have been actively studied for their ability to directly recruit immune cells to kill tumor cells. Currently, the majority of bsAbs are produced in the form of recombinant proteins, either as Fc-containing bsAbs or as smaller bsAb derivatives without the Fc region. In this study, GPC3-S-Fab, an antibody fragment (Fab) based bispecific antibody, was designed by linking the Fab of anti-GPC3 antibody GC33 with an anti-CD16 single domain antibody. The GPC3-S-Fab can be expressed in Escherichia coli and purified by two affinity chromatographies. The purified GPC3-S-Fab can specifically bind to and kill GPC3 positive liver cancer cells by recruiting natural killer cells, suggesting a potential application of GPC3-S-Fab in liver cancer therapy.
研究领域:
肿瘤学
·
免疫学
·
分子生物学
✓ 已验证
产品关联
DOI
PubMed
高影响因子
BIOCHEMICAL PHARMACOLOGY
·
2025
·
IF 5.6
Qianqian Wang,Xin Yuan,Cuijuan Liu,Ying Huang,Lin Li,Yimin Zhu
CAR-T cell therapy has been proven to be effective on hematological tumors, although graft-versus-host disease and cytokine release syndrome(CRS) limit its application to a certain extent. However, CAR-T therapy for solid tumors met challenges, among which the lack of tumor-specific antigens (TSA) and immunosuppressive tumor microenvironment (TME) are the most important factors. CAR-NK could be a good alternative to CAR-T in some ways since they can induce mild CRS and are independent of HLA-matching, but the efficacy of CAR-NKs remains limited in solid tumors. CAR cells armed with multiple tumor targeting molecules may obtain higher therapeutic efficacy against solid tumors. Due to large molecular weight, multivalent scFvs cannot be displayed efficiently on T cells and the high affinity of scFv to the target makes it easy to cause on-target, off-tumor(OTOT) toxicity. Peptides with low molecular weight and slightly lower affinity than scFvs allow immune cells to display multiple peptides to increase killing ability and reduce OTOT toxicity. In our study, peptide-based CAR-NK cells were designed to solve the dilemma of CAR-T in solid tumors. Firstly, the peptide-based CAR-NK92MI cells with A1 peptide were constructed and their inhibitory effects on the growth of A549 tumor cells were identified. Secondly, the tri-specific CAR-NK92MI cells with peptides that simultaneously targeted PD-L1, EGFR and VEGFR2 were developed for the combinatory therapy. Tri-specific CAR-NK92MI exhibited comparable killing activities to scFv-based CAR-NK92MI. Moreover, peptide-based CAR NK92MI mitigated OTOT toxicity. Our study implied that peptide-based CAR-NKs could behave as promising tools in solid tumor
研究领域:
免疫学
·
肿瘤学
·
细胞治疗
✓ 已验证
产品关联
DOI
PubMed
高影响因子
ACS Applied Materials & Interfaces
·
2024
·
IF 8.3
Wenyan Hao,Nan Sun,Yueyue Fan,Mengyu Chen,Qianqian Liu,Meiyan Yang,Yang Yang,Chunsheng Gao
Ferroptosis therapy and immunotherapy have been widely used in cancer treatment. However, nonselective induction of ferroptosis in tumors is prone to immunosuppression, limiting the therapeutic effect of ferroptosis cancer treatment. To address this issue, this study reports a customized hybrid nanovesicle composed of NK cell-derived extracellular versicles and RSL3-loaded liposomes (hNRVs), aiming to establish a positive cycle between ferroptosis therapy and immunotherapy. Thanks to the enhanced permeability and retention effect and the tumor homing characteristics of NK exosomes, our data indicate that hNRVs can actively accumulate in tumors and enhance cellular uptake. FASL, IFN-γ, and RSL3 are released into the tumor microenvironment, where FASL derived from NK cells effectively lyses tumor cells. RSL3 downregulates the expression of GPX4 in the tumor, leading to the accumulation of LPO and ROS, and promotes ferroptosis in tumor cells. The accumulation of IFN-γ and TNF-α stimulates the maturation of dendritic cells and effectively induces the inactivation of GPX4, promoting lipid peroxidation, making them sensitive to ferroptosis and indirectly promoting the occurrence of ferroptosis. This study highlights the role of the customized hNRV platform in enhancing the effectiveness of synergistic treatment with selective delivery of ferroptosis inducers and immune activation against glioma without causing additional side effects on healthy organs.
研究领域:
肿瘤学
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药理学
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纳米医学
✓ 已验证
DOI
PubMed
高影响因子
BMC CANCER
·
2025
·Vol. 25
·期 1
·1-10
·
IF 3.4
Liu Shiyuan,Guan Tian,Yang Futian,Zhang Honglei,Yao Yao,Huang Jiman,Zhao Pengfei,Huang Wei,Wu Tailiang,Lin Hui,Fu Ting,Chen Shaoyu,Dai Pengzhan,Ding Zhechun,Chen Dongmei,Li Wei,Zeng Haoyu,Zhang Xianjun
Human oral tongue squamous cell carcinoma (OTSCC) is a prevalent form of head and neck squamous cell carcinoma (HNSCC), often presenting at an advanced stage with a grim prognosis. Traditional therapeutic approaches such as surgery, adjuvant radiotherapy, and chemoradiotherapy have shown limited efficacy in treating advanced OTSCC, underscoring the urgent need for innovative treatment strategies. Our bioinformatics analysis identified CD276 as a significant biomarker in OTSCC, with high protein expression levels correlating to a markedly reduced survival rate in late-stage patients. This discovery has led us to develop chimeric antigen receptor-natural killer (CAR-NK) cells derived from umbilical cord blood cells (UCBCs), specifically targeting CD276. Our aim is to investigate this novel therapeutic approach for its potential to combat OTSCC under pre-clinical conditions. Our in vitro and ex vivo studies have demonstrated that CD276-targeted CAR-NK cells exhibit remarkable efficacy in lysing OTSCC cell lines and primary cells, as well as in eliminating OTSCC organoids. These promising results underscore the pivotal role of CD276 in OTSCC pathogenesis and highlight the potential of CAR-NK cell therapy as a groundbreaking treatment option for advanced-stage OTSCC, offering new hope for translational medicine in the field of stomatology.
研究领域:
肿瘤学
·
免疫学
·
分子生物学
✓ 已验证
DOI
PubMed
高影响因子
Bioresources and Bioprocessing
·
2022
·Vol. 9
·期 1
·1-11
·
IF 4.983
Liu Yangyang,Sun Qihao,Hao Mengyang,Tan Wen‑Song,Cai Haibo
The application of natural killer (NK) cells as potential antitumor effector cells appears to be valuable for immunotherapies. However, the clinical use of NK cells is limited because the technical difficulties associated with mass production NK cells at sufficiently high numbers represents a great challenge. Ex vivo expansion of NK cells is a key technology for cell therapy. Bioreactor systems can generate homogeneous culture condition and modulate the environmental and biochemical cues. In this study, a novel magnetically controlled bioreactor was developed for supporting NK cells ex vivo expansion. Using synthetic magnetic beads, the stirring device of the magnetically controlled bioreactor generated reduced shearing force. The intermittent magnetic field was applied for magnetic beads movement to homogenize the culture system. NK-92 cells were cultured in the magnetically controlled bioreactor and the expansion and function of expanded cells were investigated on day 8. The results showed that the expansion of NK-92 cells in the bioreactor was 67.71 ± 10.60-fold, which was significantly higher than that of the T25 culture flask (P < 0.05). Moreover, the proportions of CD3−CD56+ cells and cell killing activity of expanded cells in the bioreactor did not reveal any differences compared to T25 flasks. Taken together, this study demonstrated the possibility of magnetically controlled bioreactor as a potent strategy in NK cells production for facilitating cancer immunotherapy.
研究领域:
生物工程
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免疫学
·
肿瘤学
✓ 已验证
DOI
高影响因子
Future Science OA
·
2024
·
IF 2.4
Yan Zhu,Luanhong Wang,Biyu Jiang,Yini Wang,Qing Wu,Sihua Hong,Xiaojing Wang,Yuancheng Li,Tian Guan,Haoyu Zeng,Congzhu Li
Aim: This study aimed to explore using peripheral blood mononuclear cell (PBMC)-derived chimeric antigen receptor (CAR) NK cells targeting ROBO1 as a personalized medicine approach for ovarian cancer. Methods: A two-step strategy generated ROBO1-targeted CAR NK cells from PBMCs of ovarian cancer patients. Efficacy was evaluated using xCELLigence RTCA, CCK-8 and Live/Dead fluorescence assays. Results: ROBO1-NK cells exhibited higher efficiency in eradicating primary ovarian cancer cells and lysing ovarian tumor organoids compared with primary NK cells without ROBO1-CAR modification. Conclusion: These findings highlight the potential of developing ROBO1-targeted CAR-NK cells from patients' PBMCs as a personalized treatment option for ovarian cancer.
研究领域:
肿瘤学
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免疫治疗
·
细胞生物学
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分子生物学
✓ 已验证
产品关联
DOI
PubMed
高影响因子
Bioactive Materials
·
2024
·
IF 18
Zilin Tan,Liangjie Tian,Yang Luo,Kexin Ai,Xuehua Zhang,Haitao Yuan,Jinfan Zhou,Guangyao Ye,Shuofei Yang,Ming Zhong,Gaohua Li,Yanan Wang
Colorectal cancer (CRC) recurrence post-surgery remains a major challenge. While Chimeric Antigen Receptor (CAR)-engineered natural killer (NK) cells hold immense therapeutic potential, their intratumoral infiltration ability remains limited, hampering efficacy. Building upon prior research suggesting that chemokines like C-X-C motif chemokine ligand 9 (CXCL9) and C-X-C motif chemokine ligand 10 (CXCL10) recruit CAR-NK cells, we hypothesized that tumor cell m6A methylation, regulated by Methyltransferase-like 3 (METTL3), influences chemokine secretion. This study aims to elucidate the underlying mechanisms and improve METTL3 inhibition efficiency. We designed an adhesive hemostasis hydrogel loaded with STM2457, a METTL3 inhibitor, aimed at sustained release in the acidic tumor microenvironment. In vitro, the hydrogel promoted CAR-NK cell recruitment and tumor killing via sustained METTL3 inhibition. The hydrogel's Schiff base bonds further enabled intestinal adhesion and hemostasis in an incomplete tumor resection model of CRC. Combining the hydrogel with CAR-NK cell therapy significantly reduced CRC recurrence in vivo. Overall, our study reveals the crucial role of METTL3 in CRC recurrence and proposes a promising, multimodal strategy using STM2457-loaded hydrogel and CAR-NK cells for enhanced therapeutic efficacy.
研究领域:
肿瘤学
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分子生物学
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免疫疗法
·
生物医学工程
✓ 已验证
产品关联
DOI
PubMed
高影响因子
BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS
·
2020
·
IF 2.985
Min Liu,Yuan Meng,Leisheng Zhang,Zhongchao Han,Xiaoming Feng
Natural killer (NK) cells are pivotal effector lymphocytes characterized for the innate immune response to pathogenic microorganism and tumor cells without priming and sensitization. Despite emerging knowledge has highlighted the rosy prospects in tumor immunosurveillance , yet the large-scale clinical application of NK cell-based therapy is hindered largely attributes to the defects in generating sufficient and high-quality cellular products. Herein, on the basis of 16 kinds of candidate combinations, we investigated the feasibility of cytokine cocktail-based strategy for convenient and standardized NK cell cultivation as well as the multifaceted characteristics and cytotoxicity against tumor cells. Our results revealed that joint utilization of Interleukin (IL)-2, IL-15, IL-18 manifested the optimal facilitation upon the ex vivo expansion and proportion of NK cells in peripheral blood mononuclear cells (PBMCs). Meanwhile, the obtained NK cell population expressed high levels of activating molecules (CD16 and NKG2D) and exhibited splendid cytotoxicity against K562 cell line. Collectively, with the aid of cytokine-based programming, we established an alternative strategy for facilitating the large-scale persistence and activation of NK cells from peripheral blood, which would benefit the NK cell- and chimeric antigen receptor-modified NK (CAR-NK) cell-based autologous or allogeneic tumor immunotherapy.
研究领域:
免疫学
·
肿瘤学
·
细胞生物学
✓ 已验证
产品关联
DOI
PubMed
高影响因子
Separations
·
2024
·Vol. 11
·期 4
·105
·
IF 2.5
Dan Yu,Xiaoming Cai,Shuo Wang,Yi Li,Yuguang Du,Zhuo A. Wang,Siming Jiao,Zhenquan Yang
Phlebopus portentosusis an edible and medicinal mushroom with a delicious taste and high nutritional value. The oligosaccharides derived fromP. portentosusmay be the material basis for its biological activity. The degradation of polysaccharide and the maintenance of its activity after degradation are key steps in related research. This study applied an acid degradation method to prepareP. portentosusrefined polysaccharide (PPRP) with a smaller molecular weight, and the optimal hydrolysis conditions determined were a temperature of 80 °C, an acid concentration of 2 mol/L, and a hydrolysis time of 2 h. The polysaccharide structure and immune activity were then further investigated. The results showed that the PPRP comprised two fractions with approximate weights of 61,600 Da and 5500 Da. The monosaccharide composition of PPRP was mannose, rhamnose, glucose, and galactose, with a molar ratio of 1.00: 22.24: 2.93: 1.03. The major functional groups included O-H, C-H, C-O, and C-O-C. The glycosidic bond types were mainly α- and β-glycosidic bonds. Cell experiments demonstrated that PPRP could significantly increase the proliferation of macrophages and enhance the cytotoxicity of NK cells. Moreover, PPRP also significantly promoted the proliferation of B lymphocytes and T lymphocytes, especially at a concentration of 200 μg/mL. This study furnishes scientific evidence underlining the significant potential of PPRP in immune activity, thereby serving as a material basis and scientific bedrock for further investigations into the mechanism ofP. portentosusoligosaccharide activity.
研究领域:
真菌学
·
免疫学
·
生物化学
·
药理学
✓ 已验证
产品关联
DOI
高影响因子
Cell Reports Medicine
·
2025
·
IF 10.6
Yong Liu,Zefan Du,Lindi Li,Junbin Huang,Su Liu,Bo Lu,Yifei Duan,Yucai Cheng,Tianwen Li,Jing Zhang,Jiani Mo,Yalin Yang,Wengqing Wang,Hailin Zou,Tianqi Liang,Meng Jiang,Mo Yang,Yun Chen,Cheng Ouyang,Chun Chen
T cell acute lymphoblastic leukemia (T-ALL) is a heterogeneous disease characterized by a high relapse rate. By single-cell transcriptome analysis, we characterize the bone marrow immune microenvironment in patients with T-ALL, identifying 13 major cell clusters. These patients exhibited abnormally expanded hematopoietic stem cells (HSCs) and granulocyte-monocyte progenitors (GMPs), immunosuppressive traits in CD4 + T, CD8 + T, and natural killer (NK) cells. Subdividing CD4 + T cells reveal two subsets transitioning between T helper (Th)1/Th2, Annexin-A1 (ANXA1) − GATA3 − CD4 + T, and ANXA1 + GATA3 + CD4 + T. Additionally, NK cells demonstrate exhaustion in the tumor microenvironment of patients with relapsed T-ALL, with JUN identified as a critical factor. Additionally, JUN is also highly expressed in T-ALL and is crucial for maintaining its proliferation. The JUN inhibitor exhibited successful lethality toward leukemia cells and ameliorated NK cell exhaustion in relapsed T-ALL cell line, as well as in cell-derived tumor xenograft (CDX), patient-derived tumor xenograft (PDX), and NOTCH1-mutant mouse models. In summary, our findings enhance the understanding of T-ALL relapse mechanisms and support the development of innovative immunotherapies for patients with relapsed T-ALL.
研究领域:
血液学
·
肿瘤学
·
免疫学
✓ 已验证
产品关联
DOI
PubMed
高影响因子
INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES
·
2025
·Vol. 26
·期 23
·
IF 4.9
Huiqiang Wang,Ya Wang,Shuo Wu,Lijun Qiao,Wen Sheng,Haiyan Yan,Kun Wang,Ge Yang,Jiandong Jiang,Yuhuan Li
Previous studies have reported that Enterovirus A71 (EV-A71) infection could activate STING-related signaling pathways in vitro, but the role of STING in EV-A71 infection in vivo and the associated immune regulatory mechanisms remain unclear. Here, we used the STING-specific agonist diABZI to activate STING and STING-knockout mice to jointly study the role and mechanism of regulating STING on EV-A71 infection in vivo. The results showed that activating STING could inhibit the in vivo replication of EV-A71, alleviate clinical symptoms in infected mice, and increase the survival rate. Conversely, STING knockout significantly promoted viral replication in vivo and increased the lethality and severity of EV-A71 infection. Mechanistic studies further revealed that STING activation exerts its antiviral effects by stimulating interferon signaling pathways, upregulating the expression of interferon-stimulated genes (ISGs). Additionally, STING activation also modulated the serum cytokine response profile. Moreover, STING activation drove the expansion of diverse immune cell populations, including T cells, natural killer (NK) cells and myeloid cells. In contrast, STING knockout not only reduced the proportion of thymic T cells and impeded T cell developmental progression from double-positive (DP) to single-positive (SP) stages, but also impaired the effector functions of CD8+ T cells and NK cells during viral infection. In summary, this study demonstrates that STING activation effectively suppresses EV-A71 replication and mitigates infection symptoms by modulating immune and inflammatory responses. These findings provide a foundational framework for understanding how STING coordinates antiviral immunity and inform future investigations into STING-targeted therapies for viral infections.
✓ 已验证
产品关联
DOI
PubMed
高影响因子
COLLOIDS AND SURFACES B-BIOINTERFACES
·
2025
·
IF 5.6
Xiaolong Li,Lanlan Ma,Jueshuo Guo,Yaya Wei,Shijie Ma,Yaping Mai,Guojing Gou,Wenbao Zuo,Jianhong Yang
Melanoma is a highly aggressive form of skin cancer. mRNA vaccines deliver genetic material encoding specific antigens into cells, thereby triggering the host immune system to produce the antigen. Gp-100, an antigenic protein expressed on the surface of melanoma cells, serves as a target mRNA to stimulate the cytotoxic T lymphocyte (CTL) response. However, the absence of natural killer (NK) cells can lead to significant tumor cell proliferation. Gardiquimod, a TLR7 agonist, enhances NK cell cytotoxicity, promoting tumor clearance. In advanced melanoma, the unfolded protein response (UPR) often becomes dysregulated. By inhibiting protein kinase R-like ER kinase (PERK), the UPR can be disrupted, inducing apoptosis in cancer cells and shifting the tumor microenvironment (TME) towards an increased M1/M2 macrophage ratio. This study developed a cationic liposome-based mRNA vaccine (GD-LPR) using DOTMA to co-deliver gp-100 mRNA and the TLR7 agonist Gardiquimod, combined with the PERK inhibitor GSK2656157 (GSK), for synergistic melanoma immunotherapy. GD-LPR achieved 95 % mRNA encapsulation efficiency and demonstrated enhanced dendritic cell maturation and NK cell activation both in vitro and in vivo. In subcutaneous melanoma models, GD-LPR+GSK reduced tumor volume and prolonged survival by modulating the tumor microenvironment (TME): increasing CD8 + T cells (Fig. 3 f), repolarizing M2 to M1 macrophages (Fig. 4 f), and suppressing IL-10 while elevating pro-inflammatory cytokines (IL-2, IFN-γ, TNF-α). Mechanistically, GSK inhibited PERK/ATF-4 signaling, synergizing with GD-LPR to suppress lung metastasis. The combination of the GD-LPR vaccine and GSK provides new potential strategies for treating melanoma, particularly in subcutaneous tumors and lung metastases.
研究领域:
肿瘤学
·
免疫学
·
分子生物学
·
转化医学
·
药理学
✓ 已验证
产品关联
DOI
PubMed
高影响因子
危险品化学品经营许可证（带存储）