Ginsenoside compound K inhibits nuclear factor-kappa B by targeting Annexin A2

Wang, Yu-Shi,Zhu, Hongyan,Li, He,Li, Yang,Zhao, Bing,Jin, Ying-Hua The Korean Society of Ginseng 2019 Journal of Ginseng Research Vol.43 No.3

Background: Ginsenoside compound K(C-K), a major metabolite of ginsenoside, exhibits anticancer activity in various cancer cells and animal models. A cell signaling study has shown that C-K inhibited nuclear factor-kappa B ($NF-{\kappa}B$) pathway in human astroglial cells and liver cancer cells. However, the molecular targets of C-K and the initiating events were not elucidated. Methods: Interaction between C-K and Annexin A2 was determined by molecular docking and thermal shift assay. HepG2 cells were treated with C-K, followed by a luciferase reporter assay for $NF-{\kappa}B$, immunofluorescence imaging for the subcellular localization of Annexin A2 and $NF-{\kappa}B$ p50 subunit, coimmunoprecipitation of Annexin A2 and $NF-{\kappa}B$ p50 subunit, and both cell viability assay and plate clone formation assay to determine the cell viability. Results: Both molecular docking and thermal shift assay positively confirmed the interaction between Annexin A2 and C-K. This interaction prevented the interaction between Annexin A2 and $NF-{\kappa}B$ p50 subunit and their nuclear colocalization, which attenuated the activation of $NF-{\kappa}B$ and the expression of its downstream genes, followed by the activation of caspase 9 and 3. In addition, the overexpression of Annexin A2-K320A, a C-K binding-deficient mutant of Annexin A2, rendered cells to resist C-K treatment, indicating that C-K exerts its cytotoxic activity mainly by targeting Annexin A2. Conclusion: This study for the first time revealed a cellular target of C-K and the molecular mechanism for its anticancer activity.

Auranofin Suppresses Plasminogen Activator Inhibitor-2 Expression through Annexin A5 Induction in Human Prostate Cancer Cells

신동원,권여정,예동진,백형석,이주은,천영진 한국응용약물학회 2017 Biomolecules & Therapeutics(구 응용약물학회지) Vol.25 No.2

Auranofin has been developed as antirheumatic drugs, which is currently under clinical development for the treatment of chronic lymphocytic leukemia. Previous report showed that auranofin induced apoptosis by enhancement of annexin A5 expression in PC-3 cells. To understand the role of annexin A5 in auranofin-mediated apoptosis, we performed microarray data analysis to study annexin A5-controlled gene expression in annexin A5 knockdown PC-3 cells. Of differentially expressed genes, plasminogen activator inhibitor (PAI)-2 was increased by annexin A5 siRNA confirmed by qRT-PCR and western blot. Treatment with auranofin decreased PAI-2 and increased annexin A5 expression as well as promoting apoptosis. Furthermore, auranofin-induced apoptosis was recovered by annexin A5 siRNA but it was promoted by PAI-2 siRNA. Interestingly, knockdown of annexin A5 rescued PAI-2 expression suppressed by auranofin. Taken together, our study suggests that induction of annexin A5 by auranofin may enhance apoptosis through suppression of PAI-2 expression in PC-3 cells.

Ginsenoside compound K inhibits nuclear factor-kappa B by targeting Annexin A2

Yushi Wang,Hongyan Zhu,He Liu,Yang Li,Bing Zhao,Ying-Hua Jin 고려인삼학회 2019 Journal of Ginseng Research Vol.43 No.3

Background: Ginsenoside compound K(C-K), a major metabolite of ginsenoside, exhibits anticancer activityin various cancer cells and animal models. A cell signaling study has shown that C-K inhibitednuclear factor-kappa B (NF-kB) pathway in human astroglial cells and liver cancer cells. However, themolecular targets of C-K and the initiating events were not elucidated. Methods: Interaction between C-K and Annexin A2 was determined by molecular docking and thermalshift assay. HepG2 cells were treated with C-K, followed by a luciferase reporter assay for NF-lB,immunofluorescence imaging for the subcellular localization of Annexin A2 and NF-lB p50 subunit,coimmunoprecipitation of Annexin A2 and NF-lB p50 subunit, and both cell viability assay and plateclone formation assay to determine the cell viability. Results: Both molecular docking and thermal shift assay positively confirmed the interaction betweenAnnexin A2 and C-K. This interaction prevented the interaction between Annexin A2 and NF-lB p50subunit and their nuclear colocalization, which attenuated the activation of NF-lB and the expression ofits downstream genes, followed by the activation of caspase 9 and 3. In addition, the overexpression ofAnnexin A2-K320A, a C-K binding-deficient mutant of Annexin A2, rendered cells to resist C-K treatment,indicating that C-K exerts its cytotoxic activity mainly by targeting Annexin A2. Conclusion: This study for the first time revealed a cellular target of C-K and the molecular mechanismfor its anticancer activity.

Annexin A6 is highly abundant in monocytes of obese and type 2 diabetic individuals and is downregulated by adiponectin in vitro

Fabian Stögbauer,Johanna Weigert,Markus Neumeier,Josef Wanninger,Daniela Sporrer,Markus Weber,Andreas Schäffler,Carlos Enrich,Peta Wood,Thomas Grewal,Charalampos Aslanidis,Christa Buechler 생화학분자생물학회 2009 Experimental and molecular medicine Vol.41 No.7

Adiponectin stimulates cholesterol efflux in macrophages and low adiponectin may in part contribute to disturbed reverse cholesterol transport in type 2 diabetes. Monocytes express high levels of annexin A6 that could inhibit cholesterol efflux and it was investigated whether the atheroprotective effects of adiponectin are accompanied by changes in annexin A6 levels. Adiponectin reduces annexin A6 protein whereas mRNA levels are not affected. Adiponectin-mediated activation of peroxisome proliferator-activated receptor α (PPARα) and AMP-activated protein kinase (AMPK) does not account for reduced annexin A6 expression. Further, fatty acids and lipopolysaccharide that are elevated in obesity do not influence annexin A6 protein levels. Annexin A6 in monocytes from overweight probands or type 2 diabetic patients is significantly elevated compared to monocytes of normal- weight controls. Monocytic annexin A6 positively correlates with body mass index and negatively with systemic adiponectin of the blood donors. Therefore, the current study demonstrates that adiponectin reduces annexin A6 in monocytes and thereby may enhance cholesterol efflux. In agreement with these in vitro finding an increase of monocytic annexin A6 in type 2 diabetes monocytes was observed. Adiponectin stimulates cholesterol efflux in macrophages and low adiponectin may in part contribute to disturbed reverse cholesterol transport in type 2 diabetes. Monocytes express high levels of annexin A6 that could inhibit cholesterol efflux and it was investigated whether the atheroprotective effects of adiponectin are accompanied by changes in annexin A6 levels. Adiponectin reduces annexin A6 protein whereas mRNA levels are not affected. Adiponectin-mediated activation of peroxisome proliferator-activated receptor α (PPARα) and AMP-activated protein kinase (AMPK) does not account for reduced annexin A6 expression. Further, fatty acids and lipopolysaccharide that are elevated in obesity do not influence annexin A6 protein levels. Annexin A6 in monocytes from overweight probands or type 2 diabetic patients is significantly elevated compared to monocytes of normal- weight controls. Monocytic annexin A6 positively correlates with body mass index and negatively with systemic adiponectin of the blood donors. Therefore, the current study demonstrates that adiponectin reduces annexin A6 in monocytes and thereby may enhance cholesterol efflux. In agreement with these in vitro finding an increase of monocytic annexin A6 in type 2 diabetes monocytes was observed.

Auranofin Suppresses Plasminogen Activator Inhibitor-2 Expression through Annexin A5 Induction in Human Prostate Cancer Cells

( Dong-won Shin ),( Yeo-jung Kwon ),( Dong-jin Ye ),( Hyoung-seok Baek ),( Joo-eun Lee ),( Young-jin Chun ) 한국응용약물학회 2017 Biomolecules & Therapeutics(구 응용약물학회지) Vol.25 No.2

Auranofin has been developed as antirheumatic drugs, which is currently under clinical development for the treatment of chronic lymphocytic leukemia. Previous report showed that auranofin induced apoptosis by enhancement of annexin A5 expression in PC-3 cells. To understand the role of annexin A5 in auranofin-mediated apoptosis, we performed microarray data analysis to study annexin A5-controlled gene expression in annexin A5 knockdown PC-3 cells. Of differentially expressed genes, plasminogen activator inhibitor (PAI)-2 was increased by annexin A5 siRNA confirmed by qRT-PCR and western blot. Treatment with auranofin decreased PAI-2 and increased annexin A5 expression as well as promoting apoptosis. Furthermore, auranofin-induced apoptosis was recovered by annexin A5 siRNA but it was promoted by PAI-2 siRNA. Interestingly, knockdown of annexin A5 rescued PAI-2 expression suppressed by auranofin. Taken together, our study suggests that induction of annexin A5 by auranofin may enhance apoptosis through suppression of PAI-2 expression in PC-3 cells.

Auranofin Suppresses Plasminogen Activator Inhibitor-2 Expression through Annexin A5 Induction in Human Prostate Cancer Cells

Shin, Dong-Won,Kwon, Yeo-Jung,Ye, Dong-Jin,Baek, Hyoung-Seok,Lee, Joo-Eun,Chun, Young-Jin The Korean Society of Applied Pharmacology 2017 Biomolecules & Therapeutics(구 응용약물학회지) Vol.25 No.2

Auranofin has been developed as antirheumatic drugs, which is currently under clinical development for the treatment of chronic lymphocytic leukemia. Previous report showed that auranofin induced apoptosis by enhancement of annexin A5 expression in PC-3 cells. To understand the role of annexin A5 in auranofin-mediated apoptosis, we performed microarray data analysis to study annexin A5-controlled gene expression in annexin A5 knockdown PC-3 cells. Of differentially expressed genes, plasminogen activator inhibitor (PAI)-2 was increased by annexin A5 siRNA confirmed by qRT-PCR and western blot. Treatment with auranofin decreased PAI-2 and increased annexin A5 expression as well as promoting apoptosis. Furthermore, auranofin-induced apoptosis was recovered by annexin A5 siRNA but it was promoted by PAI-2 siRNA. Interestingly, knockdown of annexin A5 rescued PAI-2 expression suppressed by auranofin. Taken together, our study suggests that induction of annexin A5 by auranofin may enhance apoptosis through suppression of PAI-2 expression in PC-3 cells.

Annexin A2 and CD105 Expression in Pancreatic Ductal Adenocarcinoma is Associated with Tumor Recurrence and Prognosis

Huang, Ya-Kai,Liu, Hong,Wang, Xin-Zheng,Zhu, Shan Asian Pacific Journal of Cancer Prevention 2014 Asian Pacific journal of cancer prevention Vol.15 No.22

To investigate the value of expression of annexin A2, microvessel density (MVD) and CD105 in pancreatic ductal adenocarcinoma (PDAC) tissues and adjacent normal tissues, immunohistochemical staining was used. The positive expression rate of Annexin A2 and the MVD in pancreatic ductal adenocarcinoma tissues was higher than that in in adjacent normal tissues (p<0.005). Expression of Annexin A2 and MVD correlated with histological grade (p<0.05). MVD of cancers in TNM stage IIb was higher than that in TNM stageI~IIa (p<0.026). Cancerous tissues with Annexin A2 staining grade 3+ had lower MVD than the tissues with the other Annexin A2 staining grade (p<0.05). Patients with high MVD had worse prognosis. However, our study did not confirm Annexin A2 was an independent risk factor for patients with PDAC. We confirmed MVD labeled by CD105 was an independent risk factor for patients with PDAC and had moderate predictive value of prognosis.

Molecular cloning and expression analysis of annexin A2 gene in sika deer antler tip

Xia, Yanling,Qu, Haomiao,Lu, Binshan,Zhang, Qiang,Li, Heping Asian Australasian Association of Animal Productio 2018 Animal Bioscience Vol.31 No.4

Objective: Molecular cloning and bioinformatics analysis of annexin A2 (ANXA2) gene in sika deer antler tip were conducted. The role of ANXA2 gene in the growth and development of the antler were analyzed initially. Methods: The reverse transcriptase polymerase chain reaction (RT-PCR) was used to clone the cDNA sequence of the ANXA2 gene from antler tip of sika deer (Cervus Nippon hortulorum) and the bioinformatics methods were applied to analyze the amino acid sequence of Anxa2 protein. The mRNA expression levels of the ANXA2 gene in different growth stages were examined by real time reverse transcriptase polymerase chain reaction (real time RT-PCR). Results: The nucleotide sequence analysis revealed an open reading frame of 1,020 bp encoding 339 amino acids long protein of calculated molecular weight 38.6 kDa and isoelectric point 6.09. Homologous sequence alignment and phylogenetic analysis indicated that the Anxa2 mature protein of sika deer had the closest genetic distance with Cervus elaphus and Bos mutus. Real time RT-PCR results showed that the gene had differential expression levels in different growth stages, and the expression level of the ANXA2 gene was the highest at metaphase (rapid growing period). Conclusion: ANXA2 gene may promote the cell proliferation, and the finding suggested Anxa2 as an important candidate for regulating the growth and development of deer antler.

Annexin A2 gene interacting with viral matrix protein to promote bovine ephemeral fever virus release

Lihui Chen,Xingyu Li,Hongmei Wang,Peili Hou,Hongbin He 대한수의학회 2020 Journal of Veterinary Science Vol.21 No.2

Bovine ephemeral fever virus (BEFV) causes bovine ephemeral fever, which can produce considerable economic damage to the cattle industry. However, there is limited experimental evidence regarding the underlying mechanisms of BEFV. Annexin A2 (AnxA2) is a calcium and lipid-conjugated protein that binds phospholipids and the cytoskeleton in a Ca2+-dependent manner, and it participates in various cellular functions, including vesicular trafficking, organization of membrane domains, and virus proliferation. The role of the AnxA2 gene during virus infection has not yet been reported. In this study, we observed that AnxA2 gene expression was up-regulated in BHK-21 cells infected with the virus. Additionally, overexpression of the AnxA2 gene promoted the release of mature virus particles, whereas BEFV replication was remarkably inhibited after reducing AnxA2 gene expression by using the small interfering RNA (siRNA). For viral proteins, overexpression of the Matrix (M) gene promotes the release of mature virus particles. Moreover, the AnxA2 protein interaction with the M protein of BEFV was confirmed by GST pull-down and co-immunoprecipitation assays. Experimental results indicate that the C-terminal domain (268–334 aa) of AxnA2 contributes to this interaction. An additional mechanistic study showed that AnxA2 protein interacts with M protein and mediates the localization of the M protein at the plasma membrane. Furthermore, the absence of the AnxA2-V domain could attenuate the effect of AnxA2 on BEFV replication. These findings can contribute to elucidating the regulation of BEFV replication and may have implications for antiviral strategy development.

Efficacy of Annexin A1 Immunostaining in Bone Marrow for the Diagnosis of Hairy Cell Leukemia

박창훈,김현영,신상용,김희진,정철원,김선희 대한진단검사의학회 2019 Laboratory Medicine Online Vol.9 No.4

Splenic B-cell lymphomas (SBCLs) show characteristically pronounced splenomegaly without significant lymphadenopathy. Distinguishing hairy cell leukemia (HCL) from other SBCLs (splenic marginal zone lymphoma [SMZL], variant HCL [v-HCL], and splenic diffuse red pulp small B-cell lymphoma [SDRPL]) is essential to determine suitable treatments and prognoses. With advances in diagnostic modalities and therapies, splenectomy is not commonly performed, and thus diagnosis of HCL must be based on the results obtained using blood and bone marrow samples. Annexin A1 is known as the most specific marker for HCL. There has yet been no report of the assessment of annexin A1 immunostaining from Korea. In this study we analyzed samples from 13 Korean patients with SBCLs (three HCL, three v-HCL, six SMZL, and one SDRPL) from May 2001 to December 2016. Immunohistochemical analyses for annexin A1 and CD20 were performed using bone marrow sections; molecular analyses for detection of the BRAF V600E mutation were also performed. All HCL patients showed positive results for annexin A1 immunostaining and the presence of the BRAF V600E mutation, and negative results for other SBCLs. Our results confirmed the high specificity of annexin A1 and the BRAF V600E mutation as HCL markers. Molecular analysis requires expensive equipment and substantial manpower. Annexin A1 is a better alternative as an HCL marker than the BRAF V600E mutation in terms of cost-effectiveness.