Malignant tumors are complex structures composed of cancer cells and tumor microenvironmental cells.In this complex structure,cells cross-talk and interact,thus jointly promoting cancer development and metastasis.Rece...Malignant tumors are complex structures composed of cancer cells and tumor microenvironmental cells.In this complex structure,cells cross-talk and interact,thus jointly promoting cancer development and metastasis.Recently,immunoregulatory molecule-based cancer immunotherapy has greatly improved treatment efficacy for solid cancers,thus enabling some patients to achieve persistent responses or cure.However,owing to the development of drug-resistance and the low response rate,immunotherapy against the available targets PD-1/PD-L1 or CTLA-4 has limited benefits.Although combination therapies have been proposed to enhance the response rate,severe adverse effects are observed.Thus,alternative immune checkpoints must be identified.The SIGLECs are a family of immunoregulatory receptors(known as glyco-immune checkpoints)discovered in recent years.This review systematically describes the molecular characteristics of the SIGLECs,and discusses recent progress in areas including synthetic ligands,monoclonal antibody inhibitors,and Chimeric antigen receptor T(CAR-T)cells,with a focus on available strategies for blocking the sialylated glycan-SIGLEC axis.Targeting glyco-immune checkpoints can expand the scope of immune checkpoints and provide multiple options for new drug development.展开更多
Sialylated N-glycan isomers withα2-3 orα2-6 linkage(s)have distinctive roles in glycoproteins,but are difficult to distinguish.Wild-type(WT)and glycoengineered(mutant)therapeutic glycoproteins,cytotoxic T lymphocyte...Sialylated N-glycan isomers withα2-3 orα2-6 linkage(s)have distinctive roles in glycoproteins,but are difficult to distinguish.Wild-type(WT)and glycoengineered(mutant)therapeutic glycoproteins,cytotoxic T lymphocyte-associated antigen-4-immunoglobulin(CTLA4-Ig),were produced in Chinese hamster ovary cell lines;however,their linkage isomers have not been reported.In this study,N-glycans of CTLA4-Igs were released,labeled with procainamide,and analyzed by liquid chromatography-tandem mass spectrometry(MS/MS)to identify and quantify sialylated N-glycan linkage isomers.The linkage isomers were distinguished by comparison of 1)intensity of the N-acetylglucosamine ion to the sialic acid ion(Ln/Nn)using different fragmentation stability in MS/MS spectra and 2)retention time-shift for a selective m/z value in the extracted ion chromatogram.Each isomer was distinctively identified,and each quantity(>0.1%)was obtained relative to the total N-glycans(100%)for all observed ionization states.Twenty sialylated N-glycan isomers with onlyα2-3 linkage(s)in WT were identified,and each isomer's sum of quantities was 50.4%.Furthermore,39 sialylated N-glycan isomers(58.8%)in mono-(3 N-glycans;0.9%),bi-(18;48.3%),tri-(14;8.9%),and tetra-(4;0.7%)antennary structures of mutant were obtained,which comprised mono-(15 N-glycans;25.4%),di-(15;28.4%),tri-(8;4.8%),and tetra-(1;0.2%)sialylation,respectively,with onlyα2-3(10 N-glycans;4.8%),bothα2-3 andα2-6(14;18.4%),and onlyα2-6(15;35.6%)linkage(s).These results are consistent with those forα2-3 neuraminidase-treated N-glycans.This study generated a novel plot of Ln/Nn versus retention time to distinguish sialylated N-glycan linkage isomers in glycoprotein.展开更多
目的:了解P-选择素及其配体sLeA、sLeX在食管癌组织及区域淋巴结中的表达,探讨其与食管癌侵袭转移的关系。方法:应用组织芯片技术结合免疫组化检测食管鳞癌86例及其淋巴结P-选择素和sialy lewis A(sLeA)、sialylewis X(sLe X)的表达情况...目的:了解P-选择素及其配体sLeA、sLeX在食管癌组织及区域淋巴结中的表达,探讨其与食管癌侵袭转移的关系。方法:应用组织芯片技术结合免疫组化检测食管鳞癌86例及其淋巴结P-选择素和sialy lewis A(sLeA)、sialylewis X(sLe X)的表达情况,并与16例正常食管鳞状上皮对照。结果:P-选择素在食管癌组织、正常食管鳞状上皮表达率分别为59.3%、12.5%(P<0.05),在转移淋巴结、非转移淋巴结中表达率分别为92.9%、31.3%(P<0.05)。sLeA在食管癌组织,正常食管组织中表达率分别为77.9%、6.25%(P<0.05),在转移淋巴结和非转移淋巴结中表达率分别为97.6%、20.3%(P<0.05)。sLe X食管癌组织、正常食管组织中表达率分别为15.1%、6.25%(P>0.05)。P-选择素和配体sLeA过度表达与食管鳞癌病理分级,TNM分期及淋巴结转移成显著相关性;而配体sLeX在食管癌组织中无过度表达。结论:P-选择素和配体sLeA在食管鳞癌组织、转移淋巴结中表达率明显升高,其与食管癌的侵袭转移有一定关系;配体sLeX与食管癌侵袭转移无关。展开更多
目的探讨大肠腺癌组织粘蛋白MUC2和sialyl lewis X(sLex)的分型与临床各个病理参数之间的关系。方法应用免疫组织化学双染法对34例远切端大肠黏膜、18例大肠腺瘤及55例大肠腺癌组织进行粘蛋白MUC2、sLex检测。结果远切端大肠黏膜、大肠...目的探讨大肠腺癌组织粘蛋白MUC2和sialyl lewis X(sLex)的分型与临床各个病理参数之间的关系。方法应用免疫组织化学双染法对34例远切端大肠黏膜、18例大肠腺瘤及55例大肠腺癌组织进行粘蛋白MUC2、sLex检测。结果远切端大肠黏膜、大肠腺瘤及大肠腺癌中,黏蛋白MUC2阳性表达率分别为100%、94.4%和58.2%(P<0.01);sLex阳性表达率分别为5.9%、50.0%和80.0%(P<0.05)。根据MUC2和sLex在大肠腺癌中的表达把大肠腺癌分为四型:MUC2+/sLex+、MUC2+/sLex-、MUC2-/sLex+、MUC2-/sLex-。MUC2+/sLex+型与患者的性别具有相关性;MUC2+/sLex-型与淋巴结转移、Duke’s分期具有相关性;MUC2-/sLex+型与发生部位、分化程度具有相关性;MUC2-/sLex-型与大肠腺癌临床病理参数均不具有相关性。结论 MUC2的下调表达或sLex的上调表达可能参与了大肠肿瘤的发生及恶性转化,各型大肠腺癌不同程度与其分化、生物学行为相关,对临床上判断预后具有较大的意义。展开更多
AIM: High levels of serum sialyl Lewisa (sLea) are frequently found in cholangiocarcinoma (CCA) patients and have been suggested to be a serum marker for CCA. However, the significance of this antigen in CCA is unknow...AIM: High levels of serum sialyl Lewisa (sLea) are frequently found in cholangiocarcinoma (CCA) patients and have been suggested to be a serum marker for CCA. However, the significance of this antigen in CCA is unknown. In this study, the clinical significance of sLea expression in CCA tissues and the possible role of sLea in vascular invasion in vitro were elucidated. METHODS: Expression of sLea in tumor tissues of 77 patients with mass-forming CCA and 33 with periductal infiltrating CCA was determined using immunohistochemistry. The in vitro assays on adhesion and transmigration of CCA cells to human umbilical vein endothelial cells were compared between CCA cell lines with and without sLea expression. RESULTS: sLea was aberrantly expressed in 60% of CCA tumor tissues. A significant relationship was found between the frequency of sLea expression and the mass-forming type CCA (P= 0.041), well differentiated histological grading (P=0.029), and vascular invasion (P=0.030). Patients with positive sLea expression had a significantly poorer prognosis (21.28 wk, 95% CI=16.75-25.81 wk) than those negative for sLea (37.30 wk, 95% CI=27.03-47.57 wk) (P<0.001). Multivariate analysis with adjustment for all covariates showed that patients positive for sLea possessed a 2.3-fold higher risk of death than patients negative for sLea (P<0.001). The role of sLea in vascular invasion was demonstrated using in vitro adhesion and transmigration assays. KKU-M213, a human CCA cell-line with a high expression of sLea, adhered and transmigrated to IL-1β-activated endothelial cells of the human umbilical vein more than KKU-100, the line without sLea expression (P<0.001). These processes were significantly diminished when the antibodies specific to either sLea or E-selectin were added to the assays (P<0.001) CONCLUSION: This study demonstrates the clinical significance of sLea expression in vascular invasion, and an unfavorable outcome in CCA. The role of sLea in vascular invasion which may lead to poor prognosis is supported by the in vitro adhesion and transmigration studies.展开更多
The sialyl Lewis X（SLe;） antigen encoded by the FUT7 gene is the ligand of endotheliam-selectin（E-selectin）. The combination of SLe;antigen and E-selectin represents an important way for malignant tumor metastasis...The sialyl Lewis X（SLe;） antigen encoded by the FUT7 gene is the ligand of endotheliam-selectin（E-selectin）. The combination of SLe;antigen and E-selectin represents an important way for malignant tumor metastasis. In the present study, the effect of the SLe;-binding DNA aptamer on the adhesion and metastasis of hepatocellular carcinoma HepG2 cells in vitro was investigated. Reverse transcription-polymerase chain reaction（RT-PCR） and immunofluorescence staining were conducted to detect the expression of FUT7 at both transcriptional and translational levels. The SLe;expression in HepG2 cells treated with different concentrations of SLe;-binding DNA aptamer was detected by flow cytometry. Besides, the adhesion, migration, and invasion of HepG2 cells were measured by cell adhesion assay, and the Transwell migration and invasion assay. The results showed that the FUT7 expression was up-regulated at both mR NA and protein levels in HepG2 cells. SLe;-binding DNA aptamer could significantly decrease the expression of SLe;in HepG2 cells. The cell adhesion assay revealed that the SLe;-binding DNA aptamer could effectively inhibit the interactions between E-selectin and SLe;in the HepG2 cells. Additionally, SLe;-binding DNA aptamers at 20 nmol/L were found to have the similar effect to the monoclonal antibody CSLEX-1. The Transwell migration and invasion assay revealed that the number of penetrating cells on the down-side of Transwell membrane was significantly less in cells treated with 5, 10, 20 nmol/L SLe;-binding DNA aptamer than those in the negative control group（P<0.01）. Our study demonstrated that the SLe;-binding DNA aptamer could significantly inhibit the in vitro adhesion, migration, and invasion of HepG2 cells, suggesting that the SLe;-binding DNA aptamer may be used as a potential molecular targeted drug against metastatic hepatocellular carcinoma.展开更多
基金supported by the Shanghai Science and Technology Committee (Grant Nos. 20DZ2201900 to Y.Y. and 23ZR1432500 to W.P.)National Natural Science Foundation of China (Grant Nos. 82072602 to Y.Y.+4 种基金91853121, 21977066, and 22177069 to W.P.)Innovation Foundation of Translational Medicine of Shanghai Jiao Tong University School of Medicine(Grant No. TM202001 to Y.Y.)Collaborative Innovation Center for Clinical and Translational Science by Chinese Ministry of Education&Shanghai (Grant No. CCTS-2022202 to Y.Y.)Shanghai Pilot Program for Basic Research-Shanghai Jiao Tong University (Grant No. 21TQ1400210 to W.P.)Medical-Engineering Interdisciplinary Research Foundation of Shanghai Jiao Tong University (Grant No. YG2022ZD001 to W.P.)
文摘Malignant tumors are complex structures composed of cancer cells and tumor microenvironmental cells.In this complex structure,cells cross-talk and interact,thus jointly promoting cancer development and metastasis.Recently,immunoregulatory molecule-based cancer immunotherapy has greatly improved treatment efficacy for solid cancers,thus enabling some patients to achieve persistent responses or cure.However,owing to the development of drug-resistance and the low response rate,immunotherapy against the available targets PD-1/PD-L1 or CTLA-4 has limited benefits.Although combination therapies have been proposed to enhance the response rate,severe adverse effects are observed.Thus,alternative immune checkpoints must be identified.The SIGLECs are a family of immunoregulatory receptors(known as glyco-immune checkpoints)discovered in recent years.This review systematically describes the molecular characteristics of the SIGLECs,and discusses recent progress in areas including synthetic ligands,monoclonal antibody inhibitors,and Chimeric antigen receptor T(CAR-T)cells,with a focus on available strategies for blocking the sialylated glycan-SIGLEC axis.Targeting glyco-immune checkpoints can expand the scope of immune checkpoints and provide multiple options for new drug development.
基金supported by the Basic Science Research Program through the National Research Foundation of Korea(NRF)and funded by the Ministry of Education,Korea(Grant No.:2021R1A6A1A03044296)This study was supported by the Chung-Ang University Graduate Research Scholarship in 2022.
文摘Sialylated N-glycan isomers withα2-3 orα2-6 linkage(s)have distinctive roles in glycoproteins,but are difficult to distinguish.Wild-type(WT)and glycoengineered(mutant)therapeutic glycoproteins,cytotoxic T lymphocyte-associated antigen-4-immunoglobulin(CTLA4-Ig),were produced in Chinese hamster ovary cell lines;however,their linkage isomers have not been reported.In this study,N-glycans of CTLA4-Igs were released,labeled with procainamide,and analyzed by liquid chromatography-tandem mass spectrometry(MS/MS)to identify and quantify sialylated N-glycan linkage isomers.The linkage isomers were distinguished by comparison of 1)intensity of the N-acetylglucosamine ion to the sialic acid ion(Ln/Nn)using different fragmentation stability in MS/MS spectra and 2)retention time-shift for a selective m/z value in the extracted ion chromatogram.Each isomer was distinctively identified,and each quantity(>0.1%)was obtained relative to the total N-glycans(100%)for all observed ionization states.Twenty sialylated N-glycan isomers with onlyα2-3 linkage(s)in WT were identified,and each isomer's sum of quantities was 50.4%.Furthermore,39 sialylated N-glycan isomers(58.8%)in mono-(3 N-glycans;0.9%),bi-(18;48.3%),tri-(14;8.9%),and tetra-(4;0.7%)antennary structures of mutant were obtained,which comprised mono-(15 N-glycans;25.4%),di-(15;28.4%),tri-(8;4.8%),and tetra-(1;0.2%)sialylation,respectively,with onlyα2-3(10 N-glycans;4.8%),bothα2-3 andα2-6(14;18.4%),and onlyα2-6(15;35.6%)linkage(s).These results are consistent with those forα2-3 neuraminidase-treated N-glycans.This study generated a novel plot of Ln/Nn versus retention time to distinguish sialylated N-glycan linkage isomers in glycoprotein.
文摘目的探讨大肠腺癌组织粘蛋白MUC2和sialyl lewis X(sLex)的分型与临床各个病理参数之间的关系。方法应用免疫组织化学双染法对34例远切端大肠黏膜、18例大肠腺瘤及55例大肠腺癌组织进行粘蛋白MUC2、sLex检测。结果远切端大肠黏膜、大肠腺瘤及大肠腺癌中,黏蛋白MUC2阳性表达率分别为100%、94.4%和58.2%(P<0.01);sLex阳性表达率分别为5.9%、50.0%和80.0%(P<0.05)。根据MUC2和sLex在大肠腺癌中的表达把大肠腺癌分为四型:MUC2+/sLex+、MUC2+/sLex-、MUC2-/sLex+、MUC2-/sLex-。MUC2+/sLex+型与患者的性别具有相关性;MUC2+/sLex-型与淋巴结转移、Duke’s分期具有相关性;MUC2-/sLex+型与发生部位、分化程度具有相关性;MUC2-/sLex-型与大肠腺癌临床病理参数均不具有相关性。结论 MUC2的下调表达或sLex的上调表达可能参与了大肠肿瘤的发生及恶性转化,各型大肠腺癌不同程度与其分化、生物学行为相关,对临床上判断预后具有较大的意义。
基金Supported by research grants of Faculty of Medicine (#146007)Graduate School (#4432201)Khon Kaen University, Thailand
文摘AIM: High levels of serum sialyl Lewisa (sLea) are frequently found in cholangiocarcinoma (CCA) patients and have been suggested to be a serum marker for CCA. However, the significance of this antigen in CCA is unknown. In this study, the clinical significance of sLea expression in CCA tissues and the possible role of sLea in vascular invasion in vitro were elucidated. METHODS: Expression of sLea in tumor tissues of 77 patients with mass-forming CCA and 33 with periductal infiltrating CCA was determined using immunohistochemistry. The in vitro assays on adhesion and transmigration of CCA cells to human umbilical vein endothelial cells were compared between CCA cell lines with and without sLea expression. RESULTS: sLea was aberrantly expressed in 60% of CCA tumor tissues. A significant relationship was found between the frequency of sLea expression and the mass-forming type CCA (P= 0.041), well differentiated histological grading (P=0.029), and vascular invasion (P=0.030). Patients with positive sLea expression had a significantly poorer prognosis (21.28 wk, 95% CI=16.75-25.81 wk) than those negative for sLea (37.30 wk, 95% CI=27.03-47.57 wk) (P<0.001). Multivariate analysis with adjustment for all covariates showed that patients positive for sLea possessed a 2.3-fold higher risk of death than patients negative for sLea (P<0.001). The role of sLea in vascular invasion was demonstrated using in vitro adhesion and transmigration assays. KKU-M213, a human CCA cell-line with a high expression of sLea, adhered and transmigrated to IL-1β-activated endothelial cells of the human umbilical vein more than KKU-100, the line without sLea expression (P<0.001). These processes were significantly diminished when the antibodies specific to either sLea or E-selectin were added to the assays (P<0.001) CONCLUSION: This study demonstrates the clinical significance of sLea expression in vascular invasion, and an unfavorable outcome in CCA. The role of sLea in vascular invasion which may lead to poor prognosis is supported by the in vitro adhesion and transmigration studies.
基金supported by grants from the National Natural Science Foundation of China(No.81072152)the Natural Science Foundation of Hubei Province(No.2015CFA027)+1 种基金the Research Foundation of Health and Family Planning Commission of Hubei Province(No.WJ2015MA010)the Clinical Medical Research Center of Peritoneal Cancer of Wuhan(No.2015060911020462)
文摘The sialyl Lewis X（SLe;） antigen encoded by the FUT7 gene is the ligand of endotheliam-selectin（E-selectin）. The combination of SLe;antigen and E-selectin represents an important way for malignant tumor metastasis. In the present study, the effect of the SLe;-binding DNA aptamer on the adhesion and metastasis of hepatocellular carcinoma HepG2 cells in vitro was investigated. Reverse transcription-polymerase chain reaction（RT-PCR） and immunofluorescence staining were conducted to detect the expression of FUT7 at both transcriptional and translational levels. The SLe;expression in HepG2 cells treated with different concentrations of SLe;-binding DNA aptamer was detected by flow cytometry. Besides, the adhesion, migration, and invasion of HepG2 cells were measured by cell adhesion assay, and the Transwell migration and invasion assay. The results showed that the FUT7 expression was up-regulated at both mR NA and protein levels in HepG2 cells. SLe;-binding DNA aptamer could significantly decrease the expression of SLe;in HepG2 cells. The cell adhesion assay revealed that the SLe;-binding DNA aptamer could effectively inhibit the interactions between E-selectin and SLe;in the HepG2 cells. Additionally, SLe;-binding DNA aptamers at 20 nmol/L were found to have the similar effect to the monoclonal antibody CSLEX-1. The Transwell migration and invasion assay revealed that the number of penetrating cells on the down-side of Transwell membrane was significantly less in cells treated with 5, 10, 20 nmol/L SLe;-binding DNA aptamer than those in the negative control group（P<0.01）. Our study demonstrated that the SLe;-binding DNA aptamer could significantly inhibit the in vitro adhesion, migration, and invasion of HepG2 cells, suggesting that the SLe;-binding DNA aptamer may be used as a potential molecular targeted drug against metastatic hepatocellular carcinoma.
