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Direct technetium-99m labeling of anti-hepatoma monoclonal antibody fragment:a radioimmunoconjugate for hepatocellular carcinoma imaging

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World Journal of Gastroenterology 2000年第3期6卷 348-352页

作者： Hui Jie Bian zhi nan chen Jing Lan Deng Cell Engineering Research Center Basic Medical.DepartmentFourth Military Medical UniversityXi’an 710032Shaanxi ProvinceChina Department of Clinical Nuclear Medicine Xijing HospitalXi’an 710033Shaanxi ProvinceChina

AIM To directly radiolabel an anti-hepatomamAb fragment HAb18 F（ab’）2 with 99mTc bystannous-reduced method,and assess thestability,biodistribution and radioimmun-oimaging（RⅡ）.METHODS Immunoreactive fraction wasdetermined according to Lindmo’s ***’s reagent was used to determine thenumber of thiols in the reduced F（ab’）2.Labelingefficiency and homogeneity were measured bypaper chromatography,sodium dodecylsulphatepolyacrylamide gel electrophoresis（SDS-PAGE）and *** assay involvedthe incubation of aliquots of labeled antibody inethylenediaminetetraacetate（ EDTA ）and L-cysteine（L-cys）solutions with different molarratio at 37℃ for 1h,*** vivo utilized nude mice bearing humanhepatocellular carcinoma（HHCC）xenograftswith gamma camera imaging and tissuebiodistribution studies at regular *** The labeling procedure was finishedwithin 1.5 h compared with the'pretinning'method which would take at least *** vitrostudies demonstrated that the radiolabeled mAbfragment was homogeneous and retained *** studies indicatedthat 99mTc-labeled HAb18 F（ab’）2 in EDTA is morestable than in *** and biodistribution showed a significant tumor uptake at 24 h post-injection of 99mTc-labeled HAb18 F（ab’）2.Theblood,kidney,liver and tumor uptakes at 24hwere 0.56±0.09,56.45±11.36,1.43±0.27 and6.57±3.01（%ID/g）,*** 99mTc-HAb18 F（ab’）2 conjugateprepared by this direct method appears to be aneffective way to detect hepatoma in nude micemodel.

关键词： antibody monoclonal antibody fragments technetium-99m hepatocellular carcinoma liver neoplasms radioimmunoimaging

CD147 deficiency in T cells prevents thymic involution by inhibiting the EMT process in TECs in the presence of TGFβ

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Cellular & Molecular Immunology 2021年第1期18卷 171-181页

作者： Ruo chen Ke Wang Zhuan Feng Ming-Yang Zhang Jiao Wu Jie-Jie Geng zhi-nan chen Department of Cell Biology College of Life Science and TechnologyJinan UniversityGuangdongChina National Translational Science Center for Molecular Medicine&Department of Cell Biology Fourth Military Medical UniversityShaanxiChina

Thymic involution during aging is a major cause of decreased T-cell production and reduced ***,we show that the loss of CD147 on T cells prevents thymic senescence,resulting in slowed shrinkage of the thymus with age and increased production of naive T *** phenotype is the result of slowing of the epithelial-mesenchymal transition(EMT)process in thymic epithelial cells(TECs),which eventually leads to reduced adipocyte *** an in vitro coculture system,we found that TGFβ is an important factor in the EMT process in TECs and that it can reduce the expression of E-cadherin through p-5mad2/FoxC2 ***,CD147 on T cells can accelerate the decline in E-cadherin expression by interacting with Annexin A2 on *** the presence of TGFβ,Annexin A2 and E-cadherin colocalize on ***,CD147 on T cells competitively binds to Annexin A2 on TECs,leading to the isolation of ***,the isolated E-cadherin is easily phosphorylated by phosphorylated Src kinase,the phosphorylation of which was induced by TGFβ,and finally,p-E-cadherin is ***,in the thymus,the interaction between T ceils and TECs contributes to thymic involution with *** this study,we illuminate the mechanism underlying the triggering of the EMT process in TECs and show that inhibiting TGFβ and/or CD147 may serve as a strategy to hinder age-related thymic involution.

关键词： CD147 TGFβ thymic involution EMT

Modified Therapeutic Antibodies: Improving Efficacy

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Engineering 2021年第11期7卷 1529-1540页

作者： Ji-Min Dai Xue-Qin Zhang Jing-Yao Dai Xiang-Min Yang zhi-nan chen Department of Cell Biology National Translational Science Center for Molecular MedicineState Key Laboratory of Cancer BiologyThe Fourth Military Medical UniversityXi’an 710032China Department of Hepatobiliary Surgery Chinese PLA General HospitalBeijing 100853China Department of Hepatobiliary Surgery Air Force Medical CenterBeijing 100142China

The prosperity of the biotherapeutics market reflects the feasibility and effectiveness of therapeutic antibodies for the treatment of cancers,inflammatory disorders,and refractory *** drawbacks emerge in clinical trials and practice,such as impeded binding,reduced effector functions,and frequent adverse reactions,modifications of therapeutic antibodies are unprecedently burgeoning in research and development(R&D).These modifications include:①modified glycosylation;②fragment of crystallizable domain(Fc)amino acid alterations;③cross-isotype or cross-subclass exchanges;④antibody–drug conjugates(ADCs);⑤single chain of variable region fragment(scFv)for chimeric antigen receptor T(CAR-T)cells;and⑥bispecific antibodies(bsAbs)in order to promote binding affinity,half-life in circulation,effectiveness toward target cells and,ultimately,to achieve overall improved *** many achievements have been made around the world in the past decades,China has been playing an active role in this realm,with its great demand for biotherapeutics with R&D *** review recapitulates the international progress that has been achieved with modified therapeutic antibodies,and then focuses on that of China in an independent section.

关键词： Therapeutic antibody Modification Efficacy Antigen Antibody-drug conjugate Bispecific antibody

Monoclonal antibody targeting mu-opioid receptor attenuates morphine tolerance via enhancing morphine-induced receptor endocytosis

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Journal of Pharmaceutical Analysis 2023年第10期13卷 1135-1152页

作者： Jia-Jia Zhang Chang-Geng Song Miao Wang Gai-Qin Zhang Bin Wang Xi chen Peng Lin Yu-Meng Zhu zhi-Chuan Sun Ya-Zhou Wang Jian-Li Jiang Ling Li Xiang-Min Yang zhi-nan chen National Translational Science Center for Molecular Medicine&Department of Cell Biology Fourth Military Medical UniversityXi'an710032China Department of Neurology Xijing HospitalFourth Military Medical UniversityXi'an710032China Key Laboratory of Synthetic and Natural Functional Molecule of the Ministry of Education College of Chemistry and Materials ScienceNorthwest UniversityXi'an710032China Department of Neurosurgery Xi'an Daxing HospitalXi'an710032China Department of Neurobiology and Institute of Neurosciences Collaborative Innovation Center for Brain ScienceSchool of Basic MedicineFourth Military Medical UniversityXi'an710032China

Morphine is a frequently used analgesic that activates the mu-opioid receptor(MOR),which has prominent side effects of *** the inefficiency of morphine in inducing the endocytosis of MOR underlies the development of morphine tolerance,currently,there is no effective therapy to treat morphine *** the current study,we aimed to develop a monoclonal antibody(mAb)precisely targeting MOR and to determine its therapeutic efficacy on morphine tolerance and the underlying molecular *** successfully prepared a mAb targeting MOR,named 3A5C7,by hybridoma technique using a strategy of deoxyribonucleic acid immunization combined with cell immunization,and identified it as an immunoglobulin G mAb with high specificity and affinity for MOR and binding ability to antigens with spatial *** of two cell lines,HEK293T and SH-SY5Y,with 3A5C7 enhanced morphine-induced MOR endocytosis via a G protein-coupled receptor kinase 2(GRK2)/b-arrestin2-dependent mechanism,as demonstrated by immunofluorescence staining,flow cytometry,Western blotting,coimmunoprecipitation,and small interfering ribonucleic acid(siRNA)-based *** mAb also allowed MOR recycling from cytoplasm to plasma membrane and attenuated morphine-induced phosphorylation of *** established an in vitro morphine tolerance model using differentiated SH-SY5Y cells induced by retinoic *** blot,enzyme-linked immunosorbent assays,and siRNA-based knockdown revealed that 3A5C7 mAb diminished hyperactivation of adenylate cyclase,the in vitro biomarker of morphine tolerance,via the GRK2/b-arrestin2 ***,in vivo hotplate test demonstrated that chronic intrathecal administration of 3A5C7 significantly alleviated morphine tolerance in mice,and withdrawal jumping test revealed that both chronic and acute 3A5C7 intrathecal administration attenuated morphine ***,intrathecal electroporation of silencing short hairpin RNA illustrated that the in v

关键词： Morphine tolerance Mu-opioid receptor Endocytosis Monoclonal antibody Physical dependence

CD147 regulates antitumor CD8^(+)T-cell responses to facilitate tumor-immune escape

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Cellular & Molecular Immunology 2021年第8期18卷 1995-2009页

作者： Yatong chen Jing Xu Xiaodong Wu Hui Yao Zhou Yan Ting Guo Wenjing Wang Peixiao Wang Yu Li Xiangmin Yang Hao Li Huijie Bian zhi-nan chen National Translational Science Center for Molecular Medicine&Department of Cell Biology Fourth Military Medical University710032Xi’anChina Medical Research Center Southern University of Science and Technology Hospital518055ShenzhenChina Center of Anesthesiology&Operation Chinese PLA General Hospital100853BeijingChina Department of Radiation Oncology First Peoples’Hospital of ChangzhouThird Affiliated Hospital of Soochow University213000ChangzhouChina Department of Gastroenterology Henan Children’s Hospital450018ZhengzhouChina

Negative regulation of antitumor T-cell-immune responses facilitates tumor-immune ***,we show that deletion of CD147,a type I transmembrane molecule,in T cells,strongly limits in vivo tumor growth of mouse melanoma and lung cancer in a CD8^(+)T-cell-dependent *** mouse tumor models,CD147 expression was upregulated on CD8^(+)tumor-infiltrating lymphocytes(TILs),and CD147 was coexpressed with two immune-checkpoint molecules,Tim-3 and *** publicly available gene-profiling data for CD8^(+)TILs in tumor biopsies from metastatic melanoma patients showed a higher level of CD147 expression in exhausted CD8^(+)TILs than in other subsets of CD8^(+)TILs,along with expression of PD-1 and ***,CD147 deletion increased the abundance of TILs,cytotoxic effector function of CD8^(+)T cells,and frequency of PD-1^(+)CD8^(+)TILs,and partly reversed the dysfunctional status of PD-1^(+)Tim-3^(+)CD8^(+)*** cytotoxic transcription factors Runx3 and T-bet mediation enhanced antitumor responses by CD147^(-/-)CD8^(+)T ***,CD147 deletion in T cells increased the frequency of TRM-like cells and the expression of the T-cell chemokines CXCL9 and CXCL10 in the tumor *** of tumor tissue samples from patients with non-small-cell lung cancer showed negative correlations between CD147 expression on CD8^(+)TILs and the abundance of CD8^(+)TILs,histological grade of the tumor tissue samples,and survival of patients with advanced ***,we found a novel function of CD147 as a negative regulator of antitumor responses mediated by CD8^(+)TILs and identified CD147 as a potential target for cancer immunotherapy.

关键词： CD147 CD8+T cells Anti-tumor immunity

CD98-induced CD147 signaling stabilizes the Foxp3 protein to maintain tissue homeostasis

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Cellular & Molecular Immunology 2021年第12期18卷 2618-2631页

作者： JieJie Geng Ruo chen Feng-fan Yang Peng Lin Yu-meng Zhu Xianghui Fu Ke Wang Zhuan Feng Jiao Wu Hai Zhang Qi-jing Li zhi-nan chen Ping Zhu Department of Clinical Immunology Xijing HospitalFourth Military Medical UniversityXi’anP.R.China National Translational Science Center for Molecular Medicine&Department of Cell Biology Fourth Military Medical UniversityXi’anP.R.China Department of Immunology Duke University Medical CenterDurhamNCUSA

Regulatory T cell(Treg)stability is necessary for the proper control of immune activity and tissue ***,it remains unclear whether Treg stability must be continually reinforced or is established during development under physiological ***3 has been characterized as a central mediator of the genetic program that governs Treg ***,we demonstrate that to maintain Foxp3 protein expression,Tregs require cell-to-cell contact,which is mediated by the CD147-CD98 *** Tregs are produced,CD147,which is expressed on their surface,is stimulated by CD98,which is widely expressed in the physiological *** a result,CD147’s intracellular domain binds to CDK2 and retains it near the membrane,leading to Foxp3 dephosphorylation and the prevention of Foxp3 *** addition,the optimal distribution of Foxp3+Tregs under both pathological and physiological conditions depends on CD98 ***,our study provides direct evidence that Foxp3-dependent Treg stability is reinforced in the periphery by the interaction between CD147 and CD98 in the surrounding *** importantly,Tregs with high CD147 expression effectively inhibit inflammatory responses and maintain Foxp3 stability,which has guiding significance for the application of Tregs in immunotherapy.

关键词： Treg Stability CD147 CD98 CDK2 Immune therapy

Preparation and activity of conjugate of monoclonal antibody HAb18 against hepatoma F( ab′ )_2 fragment and staphylococcal enterotoxin A

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World Journal of Gastroenterology 2001年第2期7卷 216-221页

作者： Lian Jun Yang Yan Fang Sui zhi nan chen Department of Pathology, Fourth Military Medical University. Xi’an 710032, Shaanxi Province, China Department of Pathology Fourth Military Medical University Xi'an 710032 Shaanxi ProvinceChina. yangsi@***

AIM To prepare the conjugate of staphylococcal enterotoxin A (SEA) protein which is a bacterial SAg and the F(ab')2 fragment of mAb HAbl8 against human hepatocellular carcinoma (HCC), and identify its activity in order to use SAg in the targeting therapy of *** MAb HAbl8 was extracted from the abdominal dropsy of Balb/ c mice, and was purified through chromatography column SP-40HR with Fast protein liquid chromatography (FPLC) system. The F(ab')2 fragment of mAb HAb18 was prepared by papainic digestion method. The conjugate of mAb HAb18 F(ab')2fragment and SEA was prepared with chemical conjugating reagent N-succinimidyl-3-( 2-pyridyldithio) propionate (SPDP) and purified through chromatography column Superose 12with FPLC system. The molecular mass and purity of each collected peak were identified with SDS-PAGE assay. The protein content was assayed by Lowry's method. The antibody activity of HAb18 F (ab')2 against HCC in the conjugate was identified by indirect immunocytochemical ABC method, and the activity of SEA in the conjugate to activate peripheral blood mononuclear cells (PBMC) was identified with MTT *** The lgG mAb HAb18 was extracted,and purified successfully. Immunocytochemical staining demonstrated that it reacted with most of HHCC cells of human HCC cell line. There were two peaks in the process of purification of the prepared HAb18 F(ab)2-SEA conjugate. SDS-PAGE assay demonstrated that the molecular mass of the first peak was about 130 ku, and the second peak was the mixture of about 45 ku and a little 100 ku proteins. The immunocytochemical staining was similar in HAb18 F (ab ')2-SEAconjugate and HAb18 F (ab ')2, i.e., thecytoplasm and/or cell membranes of most HHCC cells were positively stained. The MTT assay showed that the optical absorbance (A) value at 490 nm of HAb18 F (ab')2-SEA conjugate was 0.182 ± 0.012, that of negative control was 0.033± 0.009, and there was significant difference between them ( P < 0.05).CONCLUSION SPDP

关键词： carcinoma, hepatocellular/immunology liver neoplasms/immunology superantigens enterotoxins antibodies, monoclonal

PDGFA-associated protein 1 is a novel target of c-Myc and contributes to colorectal cancer initiation and progression

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Cancer Communications 2022年第8期42卷 750-767页

作者： Hong-Yong Cui Wei Wei Mei-Rui Qian Ruo-Fei Tian Xin Fu Hong-Wei Li Gang nan Ting Yang Peng Lin Xi chen Yu-Meng ZhuBin Wang Xiu-Xuan Sun Jian-Hua Dou Jian-Li Jiang Ling Li Shi-Jie Wang zhi-nan chen National Translational Science Center for Molecular Medicine&Department of Cell Biology Fourth Military Medical UniversityXi’anShaanxi 710032P.R.China State Key Laboratory of Cancer Biology and National Clinical Research Center for Digestive Diseases Xijing Hospital of Digestive DiseasesFourth Military Medical UniversityXi’anShaanxi 710032P.R.China Department of Clinical Medicine Medical College of Yan’an UniversityYan’anShaanxi 716000P.R.China Key Laboratory of Synthetic and Natural Functional Molecule of the Ministry of Education College of Chemistry and Materials ScienceNorthwest UniversityXi’anShaanxi 710127P.R.China

Background:The mechanism underlying colorectal cancer(CRC)initiation and progression remains elusive,and overall survival is far from *** studies have shown that PDGFA-associated protein 1(PDAP1)is upregulated in several cancers including ***,we aimed to identify the cause and consequence of PDAP1 dysregulation in CRC and evaluate its role as a potential therapeutic ***:Multi-omics data analysis was performed to identify potential key players in CRC initiation and ***(IHC)staining was applied to determine the expression pattern of PDAP1 in CRC ***1 conditional knockout mice were used to establish colitis and CRC mouse *** sequencing,a phosphoprotein antibody array,western blotting,histological analysis,5-bromo-2′-deoxyuridine(BrdU)incorporation assay,and interactome analysis were applied to identify the underlying mechanisms of PDAP1.A human patient-derived xenograft(PDX)model was used to assess the potential of PDAP1 as a therapeutic ***:PDAP1 was identified as a potential key player in CRC development using multi-omics data ***1 was overexpressed in CRC cells and correlated with reduced overall *** investigation showed that PDAP1 was critical for the regulation of cell proliferation,migration,invasion,and ***,depletion of Pdap1in intestinal epithelial cells impaired mucosal restitution in dextran sulfate sodium salt-induced colitis and inhibited tumor initiation and growth in colitis-associated *** studies showed that c-Myc directly transactivated PDAP1,which contributed to the high PDAP1 expression in CRC ***1 interacted with the juxtamembrane domain of epidermal growth factor receptor(EGFR)and facilitated EGFRmitogen-activated protein kinase(MAPK)signaling activation,which resulted in FOS-related antigen 1(FRA-1)expression,thereby facilitating CRC progres***,silencing of PDAP1 could hinder the

关键词： carcinogenesis colorectal cancer FRA-1 PDAP1 c-Myc

Inhibitory effects of antisense RNA of HAb18G/CD147 on invasion of hepatocellular carcinoma cells in vitro

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World Journal of Gastroenterology 2003年第10期9卷 2174-2177页

作者： Yu Li Peng Shang Ai-Rong Qian Li Wang Yong Yang zhi-nan chen, Department of Cell Biology, Fourth Military Medical University, Xi’an 710032, Shaanxi Province, China Department of Cell Biology Fourth Military Medical University Xi'an 710032 Shaanxi Province China.

AIM: To study the inhibitory effects of antisense RNA of HAb18G/CD147 on invasion of hepatocellular carcinoma (HCC) cells in ***: Antisense RNA of HAb18G/CD147 vector PCIasHAb18G was constructed by reversely inserting HAb18G/CD147 cDNA to eukaryotic expression vector PCI-neo. The HCC cell line HHCC was transfected by PCI-asHAb18G via cation liposome. Expression of HAb18G/CD147 of transfected cells selected by G418 (geneticin) was observed by immunohistochemical SP staining and FACS (fluorescence activated cell sorting). Gelatin zymography was used to determine the effect of PCI-asHAb18G on reducing secretions of MMP2 and MMP-9 of the transfected cells. Boyden chamber was employed to test the invasion of HCC cells in ***: The construction of antisense RNA vector PCIasHAb18G was verified correct by partial nucleotide sequencing and restricted endonuclease digestion. The expression of HAb18G/CD147 in transfected HHCC was inhibited by PCI-asHAb18G. Secretions of MMP-2 and MMP9 of transfected HHCC were reduced and the invasion of transfected HHCC was inhibited compared to HHCC,***: Invasion of HCC cells can be inhibited by antisense RNA of HAb18G/CD147. HAb18G/CD147 may be used as a potential target of drugs for anti-invasion and metastasis of HCC.

关键词：肝细胞癌肿瘤侵袭抑制效应 HAb18G/CD147 体外试验