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
Contrast Media & Molecular Imaging
·
2018
·
IF 2.934
Weiyue Li,Xiang Xiao,Xiaodan Li,Yikai Xu,Lichao Ma,Liuji Guo,Chenggong Yan,Yuankui Wu
Objective. To investigate the potential of L5 peptide-guided pretargeting approach to identify GPC3-expressing hepatocellular carcinoma (HCC) using ultrasmall superparamagnetic iron oxide (USPIO) as the MR probe. Methods. Immunofluorescence with carboxyfluorescein- (FAM-) labeled L5 peptide was performed in HepG2 cells. Polyethylene glycol-modified USPIO (PEG-USPIO) and its conjugation with streptavidin (SA-PEG-USPIO) were synthesized, and their hydrodynamic diameters, zeta potential, T2 relaxivity, and cytotoxicity were measured. In vitro and in vivo two-step pretargeting MR imaging was performed on HepG2 cells and tumor-bearing mice after the administration of biotinylated L5 peptide (first step), followed by SA-PEG-USPIO (second step). Prussian blue staining was performed to assess iron deposition in tumors. Results. The high specificity of L5 peptide for GPC3 was demonstrated. Generation of SA-PEG-USPIO nanoparticles with good biocompatibility (an average hydrodynamic diameter of 35.97 nm and a zeta potential of −7.91 mV), superparamagnetism (R2 = 0.1039 × 103 mM−1s−1), and low toxicity was achieved. The pretargeting group showed more enhancement than the nonpretargeting group both in vitro (60% vs 20%, ) and in vivo (32% vs 6%, ). Substantial iron deposition was only observed in HepG2 cells and tumors in the pretargeting group. Conclusion. L5 peptide-guided, two-step pretargeting approach with USPIO as the MR imaging probe is a lucrative strategy to specifically identify GPC3-expressing HCC.
研究领域:
医学影像学
·
肿瘤学
✓ 已验证
产品关联
DOI
PubMed
高影响因子
PROTEIN EXPRESSION AND PURIFICATION
·
2022
·
IF 2.025
Zhiting Lao,Shuanqi Li,Jinhui Liang,Jingyi Su,Xin Gong,Xiping Cui,Suqing Zhao
Glypican-3 (GPC3) has a promise to be the diagnostic biomarker as well as therapeutic target for hepatocellular carcinoma (HCC). Nanobody have the great potential in clinical diagnosis and treatment for their characteristics of small size, high solubility, stability, manipulability, binding advantages, and ease of production. In this study, the recombinant glypican-3-N terminal (GPC3-N) protein was expressed as inclusion body in E. coli BL21(DE3)pLysS cells and then purified, which is then used as the immunogen to construct nanobody phage library. The positive clone (named MF15) was obtained by four rounds of bio-panning, and then transformed into the E. coil TOP10F’ cells to express nanobody protein, with the molecular weight of 19 kDa. Both Western blot and immunofluorescence analysis revealed that bacterially expressed GPC3-N protein is biologically active, and MF15 could specifically recognized native GPC3 expressed in HepG2 cells. The results in this study would provide the technical support for the development of diagnostic kits and antibody drugs targeting GPC3.
研究领域:
分子生物学
·
癌症研究
·
生物技术
✓ 已验证
DOI
PubMed
高影响因子
Journal of Biomedical Nanotechnology
·
2021
·Vol. 17
·期 4
·677-690
·
IF 4.099
Zhou Shuping,Ma Yongfang,Liu Xueke,Yu Pan,Huang Ning,Song Li,Xu Ruyue,Huo Zhen,Zhu Tao,Tang Xiaolong
The miR let-7b-5p (a kind of microRNAs) has many pathophysiological regulation effects, including human hepatocellular carcinoma (HCC) pathogenesis. This study investigated whether nanoparticle-mediated miR let-7b-5p could jointly enhance the therapeutic effect of sorafenib on HCC by
inhibiting the proliferation of HCC cells, inducing apoptosis, and reversing drug resistance. We evaluated the level of miR let-7b-5p in sorafenib-resistant HepG2 cells (HepG2R) and HepG2 HCC cells by qRT-PCR and analyzed the biological effects of hepatocellular carcinoma treated
with sorafenib with miR let-7b-5p, and further studied the toxicity of nanoparticles (Ab-miR-NPs) that deliver miR let-7b-5p mimics and target GPC3 on the surface of hepatocellular carcinoma cells. Results showed that, in HepG2 cells, the expression level of miR let-7b-5p was significantly
higher than that in HepG2R cells. Targeted nanoparticle Ab-miR-NPs mediated the delivery of miR let-7b-5p to the HCC cytoplasm and released miRNA after being broken down, down-regulating the expression of IGF1R and inhibiting AKT/mTOR and Ras/Raf signal transmission. Ab-miR-NPs
not only enhanced the proliferation of sorafenib in cultured HepG2R cells and induced cell apoptosis efficiency, but they also improved the anti-tumor activity in the mouse models. These results indicated that GPC3 antibody-modified PLGA-PLL (polylactic acid-glycolic acetic copolymer
grafted hyper-branched polylysine) loaded miR let-7b-5p polymer nanoparticles combined with sorafenib may be a new treatment strategy for HCC resistant to sorafenib.
研究领域:
肿瘤学
·
分子生物学
·
药理学
✓ 已验证
DOI
PubMed
高影响因子
International Journal of Nanomedicine
·
2024
·
IF 6.6
Juying Zhang,Xia Luo,Xin Yang,Hanmei Li,Qiong Jiang,You Yang,Menglin Luo,Zijun Ma,Ping He,Linli Feng,Ling Li,Maochun Zhang,Yang Li,Jinhong Yu
Purpose The incidence of hepatocellular carcinoma (HCC) is continuously increasing, and the mortality rate remains high. Thus, more effective strategies are needed to improve the treatment of HCC.Methods In this study, we report the use of a visualized glypican-3 (GPC3)-targeting nanodelivery system (named GC-NBs) in combination with sonodynamic therapy (SDT) to enhance the therapeutic efficacy for treating HCC. The obtained nanodelivery system could actively target hepatocellular carcinoma cells and achieve ultrasound imaging through phase changes into nanobubbles under low-intensity ultrasound irradiation. Meanwhile, the released chlorine e6 (Ce6) after the nanobubbles collapse could lead to the generation of reactive oxygen species (ROS) under ultrasound irradiation to induce SDT.Results Both in vitro and in vivo experiments have shown that GC-NBs can accumulate in tumour areas and achieve sonodynamic antitumour therapy under the navigation action of glypican-3-antibody (GPC3-Ab). Furthermore, in vitro and in vivo experiments did not show significant biological toxicity of the nanodelivery system. Moreover, GC-NBs can be imaged with ultrasound, providing personalized treatment monitoring.Conclusion GC-NBs enable a visualized antitumour strategy from a targeted sonodynamic perspective by combining tumour-specific targeting and stimuli-responsive controlled release into a single system.
研究领域:
肿瘤学
·
纳米技术
·
诊疗学
·
生物医学工程
✓ 已验证
产品关联
DOI
PubMed
高影响因子
MOLECULAR PHARMACEUTICS
·
2021
·
IF 4.939
Haoran Xu,Yongjia Tang,Yue Zhao,Fang Wang,Xin Gao,Dawei Deng,Yueqing Gu
The glypican-3 (GPC3) receptor is a membrane protein that is highly expressed in tumor tissues but rarely expressed in the normal liver and can be used as a target for early diagnosis of hepatocellular carcinoma (HCC). Herein, we developed a GPC3-targeted 99mTc-labeled probe for SPECT imaging in HCC. 99mTc-HPG was rapidly radiosynthesized within 20 min with an excellent radiochemical purity (>98%), possessing good stability. Results from in vitro cell binding assays indicated that the binding specificity of 99mTc-HPG to GPC3-positive HepG2 cells was acceptable. For SPECT/CT imaging, the HepG2 tumors were clearly visualized with the highest tumor/muscle ratio (11.55 ± 0.54) at 1 h post-injection, and the tumor uptake of 99mTc-HPG reduced from 2.99 ± 0.15 to 1.17 ± 0.09% ID/g in the blocking study. Convenient preparation, excellent GPC3 specificity in HCC, rapid clearance from normal organs, and good biosafety profiles of 99mTc-HPG warrant further investigations for clinical translation.
研究领域:
医学成像
·
肿瘤学
·
分子生物学
·
核医学
✓ 已验证
DOI
PubMed
高影响因子
International Journal of Nanomedicine
·
2023
·
IF 8
Jiahe Hu,Jialan Shi,Yingqian Gao,Wei Yang,Ping Liu,Qinghao Liu,Fei He,Chunxu Wang,Tao Li,Rui Xie,Jiuxin Zhu,Piaoping Yang
Background It is important to explore effective treatment for liver cancer. Photodynamic therapy (PDT) is a novel technique to treat liver cancer, but its clinical application is obstructed by limited depth of visible light penetration into tissue. The near-infrared (NIR) photosensitizer is a potential solution to the limitations of PDT for deep tumor tissue treatment.Purpose We aimed to investigate 808 nm NIR light-excited UCNPs@mSiO2-Ce6-GPC3 nanocomposites for PDT in liver cancer.Methods In our study, 808 nm NIR light-excited upconversion nanoparticles (UCNPs) were simultaneously loaded with the photosensitizer chlorin e6 (Ce6) and the antibody glypican-3 (GPC3), which is overexpressed in hepatocellular carcinoma cells. The multitasking UCNPs@mSiO2-Ce6-GPC3 nanoparticles under 808 nm laser irradiation with enhanced depth of penetration would enable the effective targeting of PDT.Results We found that the UCNPs@mSiO2-Ce6-GPC3 nanoparticles had good biocompatibility, low toxicity, excellent cell imaging in HepG2 cancer cells and high anti-tumor effect in vitro and in vivo.Conclusion We believe that the utilization of 808 nm NIR excited UCNPs@mSiO2-Ce6-GPC3 nanoparticles for PDT is a safe and potential therapeutic option for liver cancer.
研究领域:
生物医学工程
·
癌症研究
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纳米技术
·
治疗学
✓ 已验证
DOI
PubMed
高影响因子
ANALYTICAL AND BIOANALYTICAL CHEMISTRY
·
2022
·Vol. 414
·期 22
·6441-6453
·
IF 4.478
Li Guiyin,Chen Wei,Mi Danhong,Wang Bo,Li HaiMei,Wu Guangxiong,Ding Ping,Liang Jintao,Zhou Zhide
Hepatocellular carcinoma (HCC) is the second leading cause of cancer-related deaths in China. Glypican-3 (GPC3) is a specific antigen related to HCC, which is widely used in clinical detection as a reliable marker of HCC. In this paper, a highly sensitive homogeneous apatasensor was designed for GPC3 detection based on fluorescence resonance energy transfer (FRET) where the GPC3 aptamer labelled gold carbon dots (AuCDs-GPC3 Apt ) are used as a donor and magnetic graphene oxide (Fe 3 O 4 /GO) nanosheets are used as an acceptor. A one-step hydrothermal method was used to synthesize AuCDs to provide sufficient fluorescence. The FRET phenomenon exists between AuCDs-GPC3 Apt and Fe 3 O 4 /GO, which weakens the fluorescence intensity of the whole system. When the target GPC3 is added to the FRET system, the fluorescent AuCDs-GPC3 Apt binds to the GPC3 and forms a folded structure, which leads to AuCDs-GPC3 Apt separation from Fe 3 O 4 /GO nanosheets. The Fe 3 O 4 /GO is then magnetically separated so that the fluorescence of free labelled AuCDs-GPC3 Apt is restored. Under the optimum conditions, the fluorescence recovery rate is linearly correlated with the concentration of GPC3 (5–100 ng·mL −1 ) and the detection limit is 3.01 ng·mL −1 (S/N = 3). This strategy shows recoveries from 98.76 to 101.29% in real human serum samples and provides an immediate and effective detection method for the quantification of GPC3 with great potential applications for early diagnosis of HCC. Graphical abstract A sensitive homogeneous FRET-based apatasensor was designed for GPC3 detection where the AuCDs-GPC3 Apt is a donor and Fe 3 O 4 /GO nanosheets are an acceptor. The GPC3 fluorescent aptasensor combines wider output range with low cost, high specificity, and good anti-interference.
研究领域:
医学诊断
·
癌症研究
·
纳米技术
·
分子生物学
✓ 已验证
产品关联
DOI
PubMed
高影响因子
JOURNAL OF PHOTOCHEMISTRY AND PHOTOBIOLOGY B-BIOLOGY
·
2023
·
IF 5.4
Jianying Yang,Zhao Wang,Chunhong Mo,Haikun Luo,Shuting Li,Qian Mo,You Qin,Fan Yang,Xinchun Li
Efficiently synergistic therapy of hepatocellular carcinoma (HCC) by chemotherapeutic drug and photothermal agent remains a considerable challenge. Here, we report a nanodrug that integrates specific hepatoma-targeted delivery, pH-triggered drug release, and cooperative photothermal–chemotherapy function. By grafting the easily self-assembled [email protected] ( [email protected] ) nanocapsulation with polyacrylic acid (PAA), an inorganic-organic-polymeric hybrid nanovehicle was developed as a dual photothermal agent and carrier for loading antitumor drug–doxorubicin (DOX) through electrostatic adsorption and chemical linking antibody against GPC3 commonly overexpressed in HCC, resulting in the nanodrug, [email protected] /PAA/DOX/GPC3. The multifunctional nanovehicle had excellent biocompatibility , stability, and high photothermal conversion efficiency, due to the rationally designed binary [email protected] photothermal agent. The 72-h accumulative drug release in pH 5.5 tumor microenvironment can reach up to 84%, far higher than 15% measured in pH 7.4 condition. Notably, in contrast to the merely 20% survival rate of H9c2 and HL-7702 cells exposed to free DOX, their viabilities in the nanodrug circumstance can maintain 54% and 66%, respectively, suggesting the abated toxicity to the normal cell lines. When exposed to the hepatoma-targeting nanodrug, the viability of HepG2 cells was found to be 36%, which further drastically declined to 10% plus 808-nm NIR irradiation. Moreover, the nanodrug is potent to cause tumor ablation in HCC-modeled mice, and the therapeutic efficacy can be greatly enhanced under NIR stimulus. Histology analyses reveal that the nanodrug can effectively alleviate the chemical damage to heart and liver, as compared to free DOX. This work thus offers a facile strategy for design of targeting anti-HCC nanodrug toward combined photothermal-chemotherapy.
研究领域:
癌症研究
·
纳米技术
·
药学
·
生物医学工程
✓ 已验证
产品关联
DOI
PubMed
高影响因子
HUMAN MUTATION
·
2025
·Vol. 2025
·期 1
·5319444
·
IF 3.7
Xin Wu,Linde Sun,Peifa Liu,Qiong Wang,Yu Wang
Background PANoptosis, as an inflammatory programmed cell death, is involved in tumor development. This study set out to discover novel PANoptosis-correlated prognostic signatures in stomach adenocarcinoma (STAD), a prevalent malignancy of the digestive system. Methods STAD samples were derived from a public database, and PANoptosis-related genes (PRGs) were acquired from existing reports. Prognosis-related PRGs were screened by univariate Cox regression analysis. Molecular subtypes of STAD were identified by the “ConsensusClusterPlus” package. The “Limma” package was employed to filter differentially expressed genes (DEGs) between different subtypes. PANoptosis-related prognostic signatures in STAD were identified to establish the RiskScore model. The RiskScore and some of the clinical features were integrated to establish a nomogram. Immune cell infiltration and TIDE score in different risk groups were compared. Correlation between immune checkpoint genes, drug sensitivity, and RiskScore was analyzed by the Spearman method. The biological function of PANoptosis-related signature genes in STAD was preliminarily explored by in vitro cell experiments. Results Based on 18 prognosis-related PRGs, two molecular subtypes of STAD were recognized, and the C1 subtype showed a lower overall survival (OS) rate than the C2 subtype. Further, three PANoptosis-related signature genes ( APOD , GPC3 , and SERPINE1 ) were determined to establish a RiskScore model that could accurately assess the prognostic outcomes for STAD patients. Then, by integrating RiskScore with clinical features, a nomogram was established. The high-risk group had higher immune cell infiltration and TIDE score and lower OS rate than those with a low risk. RiskScore was positively correlated with nine immune checkpoint genes. Besides, we screened 23 drugs that significantly correlated with RiskScore. In vitro cell experiments showed that the mRNA and protein levels of APOD , GPC3 , and SERPINE1 were upregulated in the STAD cell line and that APOD knockout significantly reduced cancer cell proliferation, migration, and invasion levels and increased the apoptotic capacity of the STAD cell line. Conclusion This study established a PANoptosis-related RiskScore model for assessing STAD patient prognosis, which could contribute to the personalized treatment of STAD.
研究领域:
肿瘤学
·
分子生物学
·
生物信息学
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免疫治疗
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精准医学
✓ 已验证
DOI
PubMed
高影响因子
NEW JOURNAL OF CHEMISTRY
·
2021
·Vol. 45
·期 19
·8608-8618
·
IF 3.591
Min Chen,Haimei Li,Xueming Su,Runqiang Wu,Huafu Feng,Xiaohang Shi,Jintao Liang,Jiejing Chen,Guiyin Li
Glypican-3 (GPC3) is a potential hepatocellular carcinoma (HCC) serum marker with good sensitivity and specificity. Herein, a label-free electrochemical aptasensor for the detection of GPC3 was developed using a reduced graphene oxide–hemin–chitosan (RGO–H–CS) nanocomposite-modified screen-printed electrode (SPE) as a biosensing platform and GPC3 aptamer as the recognition element. When GPC3 was present, the aptamer could specifically bind with the target GPC3 by specific recognition reaction and form GPC3–aptamer conjugations on the sensing surface, which could alter the electrochemical redox signal of hemin (Fe(III)/hemin(Fe(II))) in the RGO–H–CS nanocomposite recorded by differential pulse voltammetry (DPV). The electrochemical GPC3 aptasensor was characterized via scanning electron microscopy (SEM), Raman spectroscopy, cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS). Under optimal conditions, currents originating from the oxidation of hemin had a linear relationship with the concentration of GPC3 in the range of 0.01 µg mL−1 to 10.0 µg mL−1 with a detection limit of 7.9 ng mL−1 (S/N = 3). The developed aptasensor was applied to detect the GPC3 level in the human serum samples with the recovery rate of 101.04–104.9%. Moreover, the label-free aptasensor exhibited considerable anti-interfering ability, acceptable stability and good reproducibility. Therefore, this study provides a reliable and objective analysis method to achieve high screening for the early diagnosis of HCC.
研究领域:
生物传感器
·
癌症诊断
·
电化学
·
纳米技术
·
分析化学
✓ 已验证
产品关联
DOI
高影响因子
JOURNAL OF THE ELECTROCHEMICAL SOCIETY
·
2022
·Vol. 169
·期 8
·087517
·
IF 4.386
Shengnan Li,Xinhao Li,Liping Cao,Chaoxian Wang,Jintao Liang,Zhide Zhou,Guiyin Li
Glypican-3 (GPC3) is a serum marker for hepatocellular carcinoma (HCC) and has excellent specificity and sensitivity. Herein, the construction of an electrochemical aptasensor based on reduced graphene oxide-carboxymethyl chitosan-hemin-palladium nanoparticles (RGO-CMCS-Hemin/Pd NPs) and GPC3 aptamer (GPC3apt) has the function of GPC3 identification and analysis. RGO-CMCS-Hemin/Pd NPs has excellent biocompatibility, large specific surface area and good electrical conductivity, which is not only used as advanced sensing materials to modify the electrode surface to reduce the resistance and increase the signal amplitude, but also used as an electroactive substance to provide electrical signals. GPC3apt as a biometric element makes the aptasensor has good specificity and sensitivity. The GPC3apt can combine with GPC3 and form GPC3-aptamer conjugation, which increases the electron transfer impedance and changes the electrical signal. Under the optimum conditions, the aptasensor had a good linear relationship with the GPC3 concentration in the range of 1.0–100.0 ng ml−1 with R2 of 0.9573. The lower limit of detection was 1.0 ng ml−1. When the aptasensor was applied to GPC3 detection in human serum samples, the recovery was 93.73%–118.83%. In addition, the GPC3 aptasensor has excellent specificity, reproducibility and stability, which is anticipated to be applied for the clinical detection of GPC3.
研究领域:
生物医学工程
·
分析化学
·
纳米技术
·
癌症诊断
✓ 已验证
产品关联
DOI
高影响因子
危险品化学品经营许可证（带存储）