Apigenin inhibits epithelial-mesenchymal transition of ... · Human colon carcinoma cell lines...

Bioscience Reports (2019) 39 BSR20190452https://doi.org/10.1042/BSR20190452

*These authors contributeequally to this work

Received: 28 February 2019Revised: 02 April 2019Accepted: 02 April 2019

Accepted Manuscript Online:09 April 2019Version of Record published:14 May 2019

Research Article

Apigenin inhibits epithelial-mesenchymal transitionof human colon cancer cells through NF-κB/Snailsignaling pathwayJiafeng Tong1,*, Ying Shen2,*, Zhenghua Zhang3, Ye Hu2, Xu Zhang1 and Li Han1

1Department of Traditional Chinese Medicine, Huadong Hospital, Shanghai 200040, China; 2Department of Gastrology, Shanghai Municipal Hospital of Traditional ChineseMedicine, Shanghai University of Traditional Chinese Medicine, Shanghai 200071, China; 3Department of Oncology, Jing’an District Central Hospital of Shanghai (HuashanHospital, Fudan University, Jing’an branch200040), China

Correspondence: Li Han ([email protected])

Colon cancer is a leading cause of cancer-related deaths worldwide. Theepithelial-mesenchymal transition (EMT) plays an important role in tumor metastasisof colon cancer. We first evaluated the effects of EMT-related transcription factors on theprognosis of colon cancer through analysis the data obtained from The Cancer GenomeAtlas (TCGA). And then we screened a series of Chinese medicine monomers to find effectEMT inhibitors. First, Snail is a more important EMT transcription factors for colon cancerprognosis, compared with Twist and Slug. Then, we found that apigenin effectively inhibitsthe activity of Snail. Apigenin could inhibit the EMT, migration, and invasion of human coloncancer cells in vitro and in vivo through the NF-κB/Snail pathway. Snail is a key regulatorof EMT in colon cancer and Snail inhibitor apigenin may be a therapeutic application forpatients with colon cancer.

IntroductionColon cancer is a leading cause of cancer-related deaths worldwide. This disease is usually diagnosed atits advanced stage [1]. The 5-year survival rate of stage IV colon cancer is less than 10%. Colon metastasesreadily develop through the epithelial-mesenchymal transition (EMT) of epithelial cells, a key process thatsupports tumor metastasis.

In the process of EMT, epithelial cells lose their typical epithelial characteristics and acquire mesenchy-mal traits. Cancer cells lose their cell–cell connection, cell-matrix contact, and normal epithelial polaritywhile gaining mesenchymal characteristics, which enhance the migratory and invasive ability of cancercells. EMT is an integral component of colorectal cancer progression. So targetting EMT will be beneficialfor prognosis and therapy of colorectal cancer [2,3].

Natural products (animals, plants, and other pharmacologically active ingredients present in nature)are important sources for prevention and treatment of diseases. The drugs used in various traditionalmedicines in the world almost belong to natural sources. The chemical substances from natural chemi-cals are also more than 30% in modern medicine applications. And more drugs are produced by structuralmodification of the natural products (paclitaxel, vincristine, artemisinin, etc.). According to a review byNewman et al., about 61% of the 877 new small chemical drugs introduced in the world between 1981and 2002 were derived from natural products or the derivatives of natural products [4]. At present, as thedevelopment of synthetic drugs becomes more and more difficult, scientists have refocussed the develop-ment of new drugs on natural products. Chinese herbal monomers, such as the flavonoid monomers, have

© 2019 The Author(s). This is an open access article published by Portland Press Limited on behalf of the Biochemical Society and distributed under the Creative Commons AttributionLicense 4.0 (CC BY).

1

Dow

nloaded from https://portlandpress.com

/HTTPH

andlers/ArticlePdfHandler.ashx?partialdoi=BSR

20190452&journal=bioscirep by guest on 01 January 2020

http://orcid.org/0000-0002-6842-7620

mailto:[email protected]

https://crossmark.crossref.org/dialog/?doi=10.1042/BSR20190452&domain=pdf&date_stamp=2019-05-14


extensive pharmacological activity [5] (antitumor, anti-inflammatory, immune regulation, etc.), which has attractedmore and more attention.

Some transcription factors, such as zinc-finger proteins of the Snail/Slug family and Twist Family BHLH Transcrip-tion Factor 1 (Twist1), have been proved to regulate the EMT [6]. In order to find effect EMT inhibitor in colorectalcancer, we first evaluated the effects of these transcription factors on the prognosis of colon cancer through analy-sis the data obtained from The Cancer Genome Atlas (TCGA). And then we screened a series of Chinese medicinemonomer to find effect EMT inhibitors through detecting the promoter activity of the prognosis-related transcriptionfactor. The antitumor effects of the screened inhibitor (apigenin) were further evaluated in vitro and in vivo.

Materials and methodsChemicals and antibodiesPrimary antibodies including vimentin (1:1000), E-cadherin (1:1000) and GAPDH (1:5000), NF-κB (1:1000), andSnail (1:1000) were supplied by Affinty (CO, U.S.A.). Secondary antibodies included FITC-labeled goat-antimouselgG and TRITC-labeled goat anti-rabbit IgG were obtained from EarthOx (San Francisco, CA, U.S.A.). Apigenin andother flavanoid derivates were obtained from PUSH BIO-TECHNOLOGY (Chengdu, China). Matrigel and transwellchambers were purchased from BD Biosciences (San Jose, CA, U.S.A.).

Cell cultureHuman colon carcinoma cell lines HCT-116 and LOVO, purchased from KeyGen Biotech (Nanjing, China), weremaintained according to ATCC’s recommendation. The cells were grown in DMEM (for HCT-116) or RPMI 1640(for LOVO) medium supplemented with 10% FBS (Hyclone, U.S.A.) in a humidified atmosphere of 37◦C, 5% CO2.

Wound-healing assayHCT-116 and LOVO cells were seeded in six-well plates. The next day, a sterile 200 μl pipette tip was used to drawa straight line when the whole plates were covered with cells. Then the cells were washed with PBS for three times.After that different concentration of apigenin were added to DMEM mediums for 24 h. The distance of the scratchwas calculated at different time points with microscope (Olympus CX31).

Cell invasion assaysHCT-116 and LOVO cells were cultured to near 90–100% confluence. Then the cells were digested and seeded onthe upper chamber (8.0 μm) of the transwell cultured with containing different concentrations of apigenin, whichwere coated with matrigel. The lower compartment of the chamber was added 0.5 ml of medium supplemented with10% FBS. After treatment for 48 h, the medium was removed and the cells were washed with PBS for three times.Subsequently, the cells were fixed with precooled methanol and then the cells were stained with crystal violet for10 min at room temperature. Finally, the stained cells were evaluated with the microscope.

Immunofluorescent stainingHCT-116 cells (4 × 104 cells/ml) were pretreated with 10 or 20 μM apigenin for 48 h and then the cells were seededon the chamber slides of 24-well culture plates. After 24 h, the cells were fixed with 4% paraformaldehyde about 30min, following permeation with 1% Trition X-100 dissolved in PBS. Then the nonspecific binding was blocked with5% BSA in TBS and incubated with primary antibodies (E-Cadherin and vimentin were 1:100 dilution respectively)for 1 h at 4◦C overnight. Slides were washed with PBS and incubated at 37◦C for 45 min with secondary antibodiesconjugated to fluorescein isothiocyanate. Then the cells were washed with PBS. DAPI were used to monitor nuclearintegrity. Finally, the cells were examined using Nikon confocal microscopy.

Animal studiesThe BALB/c nu/nu mice were well taken care under pathogen-free conditions in accordance with institutional guide-lines. Studies were carried out in accordance with National Institutes of Health Animal Use Guidelines and the currentChinese Regulations and Standards for the Use of Laboratory Animals. The mice were subcutaneously injected with100 μl resuspended HCT-116 cells (2 × 106 cells) via a sterile microsyringe. After the tumor volume reached about40 mm3, the mice were randomly assigned to control or apigenin treatment groups. Control group was treated withequal volume saline, and treatment groups were intragastrically administered with different concentrations apigenin(low dose of 200 mg/kg and high dose of 300 mg/kg). Mice weight and tumor size were recorded every 2 days. The

2 © 2019 The Author(s). This is an open access article published by Portland Press Limited on behalf of the Biochemical Society and distributed under the Creative Commons AttributionLicense 4.0 (CC BY).

Dow


/HTTPH




tumor volumes were measured using calipers and calculated with the formula V = length × width2/2. After treatmentfor 2 weeks, the mice were killed. Tumor weight was recorded and the tissues were fixed in 10% formlinata solutionand then embedded by paraffin.

Immunohistochemical analysisThe fixed tissues were used for immunohistochemistry staining. About 3 μm thick paraffin sections were deparaf-finized with xylene, dehydrated in ethanol in a concentration gradient, and followed by incubation with 3% hydrogenperoxide at room temperature for 10 min to quench endogeneous peroxidase. Total 10 mM citrate buffer (pH 6.0)was used to antigen retrieval in a microwave about 10 min. Nonspecific-binding sites were blocked with normal goatserum for 30 min. The paraffin slices were incubated with the primary antibodies anti-E-cadherin (rabbit polyclonal,1;100) and antivimentin (mouse polyclonal, 1:100) in 1% BSA overnight at 4◦C. The tissues were washed in PBS forthree times. Then antimouse IgG and antirabbit lgG (diluted 1:200) was added to the paraffin slices for 30 min. DABsubstrate was utilized for color development.

Statistical analysesData are represented as means +− S.D. by three independent experiments. The statistical significance of differentgroups was analyzed by one-way ANOVA following SPSS 17.0 software. The P values <0.05 were considered signif-icant difference existence.

ResultsSnail is associated with prognosis of colon cancerThe survival analysis of EMT-related transcription factors, Snail, Slug, Twist1, and Twist2, is shown in Figure 1A).Patients with higher expression of Snail had shorter survival time (***P<0.0005) AMONGST all the factors, whichshowed that Snail is the key driver of EMT in colon cancer. Snail as a poor prognosis gene were highly expressed incolon cancer tissues from the results of immunohistochemical staining (Figure 1B). The patient data were all derivedfrom the TCGA database.

Apigenin effectively inhibits the transcription activity of snailIn order to find the inhibitor of Snail, we then screened dozens of Chinese herbal monomers. The inhibitors of snailtranscription activity were screened using the dual luciferase reporter assay system. We found several Chinese herbalmonomers had regulatory effect on Snail and apigenin had the best effect (Figure 1C)

Apigenin inhibits the proliferation of human colon cancer cellsThe cytotoxicity of apigenin was detected by MTT. The results showed that apigenin can inhibit the proliferation ofHCT116 and LOVO in a dose-dependent manner (Figure 2A,B). The half-maximal inhibitory concentrations (IC50)of HCT-116 and LOVO were 47.33 and 34.45 μM respectively. The morphology of colon cancer cells changed sig-nificantly after treatment with apigenin, and the cells exhibited low viability after exposed to apigenin at differentconcentrations.

Apigenin inhibits migration and invasion in human colon cancer cellsWe detected the effect of apigenin on invasion and metastasis activity of cancer cells through wound-healing andtranswell experiments. The wound-healing experiments showed that both 10 and 20 μM apigenin could inhibit themigration of HCT-116 and LOVO cells comparing with the control groups (Figure 2C,D).

Matrigel-coated transwell chambers were used to verify the invasive ability of colon cancer cells after apigenin treat-ment. The potential metastatic capacity of tumor cells can be represented by its invasiveness. In the apigenin-treatedgroup, the number of cells that invading the basement membrane of the transwell chamber is significantly reducedcompared with the control group, and it was concentration dependent in HCT-116 and LOVO cells (Figure 2E,F).

Apigenin reversed changes in E-cadherin and vimentin expression inhuman colon cancer cellsWhen EMT occurs in tumor cells, the expression of E-Cadherin is down-regulated and the expression of vimentinis up-regulated. Cells treated with apigenin displayed distinct morphological changes compared with untreated cells.Pseudopodia increased significantly after apigenin treatment. We performed immunofluorescence double staining on


3

Dow


/HTTPH




Figure 1. Snail is a key regulator of EMT in colon cancer

(A) The survival curves of Snail, Slug, Twist1, and Twist2 analysis in colon cancer patients. The patient data were obtained from

the TCGA database (*P<0.05, **P<0.0001). (B) The immunohistochemical microarray staining of Snail in normal and colon cancer

tissues. The data were obtained from the TCGA database. (C) The inhibitors of Snail transcription activity were screened using the

dual luciferase reporter assay system.

EMT markers E-Cadherin and vimentin, and found that E-Cadherin levels increased and vimentin levels decreasedin the apigenin treatment groups (Figure 3A,B).

Apigenin inhibits NF-κB expression and Snail transcriptional activation incolon cancerBased on the above experimental results, we learned that apigenin could inhibit EMT by inhibiting the activity of Snailin colon cancer cells. NF-κB could regulate the activity of Snail. In order to detect the effect of apigenin on NF-κB, weperformed a dual luciferase reporter gene assay experiment. The results showed that apigenin could also inhibit thetranscriptional activity of NF-κB (Figure 4A). We also validated the inhibit effect of apigenin on the transcriptionalactivity of Snail (Figure 4A). Apigenin inhibited the transcriptional activity of Snail in a dose dependent manner.


Dow


/HTTPH




Figure 2. Effect of apigenin on cell viability, invasion, and metastasis of HCT-116 and LOVO cells

(A) HCT-116 cells viability was inhibited after reincubation in medium containing 0, 10, or 20 μM apigenin for 24 h. (B) LOVO cells

viability was inhibited after reincubation in medium containing 0, 10, or 20 μM apigenin for 24 h. (C,D) Effects of apigenin on

migration ability of HCT-116 and LOVO cells. Cells were treated with 0, 10, or 20 μM apigenin (*P<0.05, **P<0.01). (E,F) Effects of

apigenin on the invasion ability of HCT-116 and LOVO cells (*P<0.05, **P<0.01). Cells were treated with 0, 10, or 20 μM apigenin.

Representative images of cells at the bottom of the membrane stained with crystal violet were visualized.


5

Dow


/HTTPH




Figure 3. Apigenin reverses changes of EMT biomarkers in HCT-116 cells

(A,B) In HCT-116 cells, vimentin expression decreased whereas E-cadherin expression increased after apigenin treatment. Typical

images of immunofluorescent double staining for E-cadherin and vimentin in HCT-116 cells. Each experiment was performed

in triplicate. Results are shown as the means of three experiments, and error bars represent the standard deviation (*P<0.05,

**P<0.01).

The expression of transcription factors, NF-κB and Snail which related to EMT, were significantly up-regulated intumor cells. After apigenin treatment, the transcription activity of NF-κB and Snail (Figure 4B-D) were significantlydown-regulated. We also examined the expression of NF-κB and Snail in the nucleus, and it was proved that thechange of protein level consistent with the trend of NF-κB and Snail in the cytoplasm. After apigenin treatment, theexpression of both NF-κB and Snail decreased in the cytosol and nucleus. These results showed that apigenin couldinhibit EMT of colon cancer cells through NF-κB/Snail pathway.

Apigenin inhibits the metastasis of HCT-116 cells in xenograft modelnude miceThe effect of apigenin on animal body weight was not significant (Figure 5A). Our results showed that after drugtreatment, the tumor volume of mice treated with apigenin was decreased compared with control group (Figure5B,C). In vivo experiments showed that apigenin could effectively reduce the tumor volume.


Dow


/HTTPH




Figure 4. Apigenin inhibits NF-κB and Snail in human colon cancer cells

(A,B) The effect of apigenin on NF-κB and Snail expression in HCT-116 and LOVO cells tested by dual-luciferase reporter assay

(*P<0.05, **P<0.01). (C,D) Effect of apigenin on NF-κB and Snail expression in HCT-116 and LOVO cells detected by western blot.

Apigenin reduced the expression of NF-κB and Snail (*P<0.05, **P<0.01).

We further performed immunohistochemical staining on the fixed tissue cells. The expression of the EMT markersin colon tissue was evaluated. Our results demonstrated that E-Cadherin and occludin expression were up-regulatedafter apigenin treatment, while vimentin and N-cadherin expression were down-regulated after apigenin treatment(Figure 6A,B). We then performed immunohistochemical staining analysis of EMT-related transcription factors andfound that the expression of NF-κB and Snail was down-regulated after apigenin treatment, which indicated thatapigenin treatment could inhibit the expression of EMT-related transcription factors. Besides, apigenin treatmentcould also inhibit the expression of Ki-67 and NF-kB signaling proteins such as p65 and IKK (Figure 6C,D), suggestingthat apigenin might inhibit EMT of colon cancer cells through NF-kB/Snail signaling pathways.

DiscussionSince colon cancer is a leading cause of cancer-related deaths worldwide [7], search for effective drugs is crucialfor the treatment of colon carcinoma [8–10]. EMT promotes epithelial tumor invasion and transformation towardmalignant disease [11]. The phenotype of EMT is mainly embodied in the cell–cell adhesion systems down-regulation,loss of epithelial polarity, and the acquisition of a mesenchymal phenotype, which lead to migration and movement ofepithelial cells [12]. Thus, targetting EMT is beneficial for inhibition of carcinoma progression [13]. So in this work wescreened the EMT inhibitors for colon carcinoma. In this work, we found that the expression of Snail1 has correlationwith the prognosis of colon cancer patients. And we screened a series of flavonoid Chinese herbal monomers to find


7

Dow


/HTTPH




Figure 5. Effect of apigenin on HCT-116 xenograft model

(A) Body weight changes of animals with HCT-116 xenografts (*P<0.05, **P<0.01). (B,C) Changes in the tumor volume of HCT-116

xenografts after being treated with apigenin. Apigenin treatment inhibited xenograft growth in a dose-dependent manner (*P<0.05,

**P<0.01). The results were shown as the means of three experiments, and error bars represent standard deviation.

Snail1 transcription activity inhibitors. We found that apigenin had good inhibitory activity for Snail1, so we furtherevaluated its antitumor effect on human colon cancer cells.

Snail, Slug, and Twist1 are important transcription factors for the regulation of the EMT in tumor cells. The firstdiscovered transcriptional repressor of E-cadherin was Snail [14,15], which plays a fundamental role in EMT. Thedata analysis obtained from TCGA database revealed that the expression of Snail1 has correlation with the prognosisof colon cancer patients. Patients with high expression of Snail have shorter survival. The transcriptional factors of


Dow


/HTTPH




Figure 6. Effect of apigenin on NF-κB, Snail, Ki-67, NF-κB p65 IKK, and EMT protein levels

(A) Effect of apigenin on EMT biomarkers in HCT-116 xenograft model (*P<0.05, **P<0.01). (B) Expression level of EMT biomarkers.

(C) Effect of apigenin on expression of NF-κB, Snail, Ki-67, NF-κB p65, IKK in HCT-116 xenograft model (*P<0.05, **P<0.01). (D)

Expression of NF-κB, Snail, Ki-67, NF-κB p65, and IKK.

Slug and Twist1 have no correlation with the prognosis of colon cancer patients. These findings reveal the importanceof Snail1 in the malignant evolution of colon cancer. And Snail1 is a marker of poor prognosis of colon cancer.

E-cadherin and vimentin are important markers of EMT. Studies on E-cadherin behavior provided considerableinsights into carcinoma progression [16]. In addition, altered E-cadherin expression is associated with tumor stageand increased lymph node metastasis [17,18]. Vimentin acts as an intermediate filament protein, which forms the cy-toskeleton along with microtubules and actin filaments. In this work, we found that apigenin increased the expressionlevels of E-cadherin but decreased the expression levels of vimentin. These results suggest that apigenin suppressesEMT of colon carcinoma cells. The results of the HCT-116 xenografts in nude mice also validated the effect of api-genin on EMT. Apigenin also inhibited the migration and invasion of tumor cells, which could be promoted by EMTof tumor cells. The mechanism of apigenin in the treatment of colon cancer can be explained by EMT.

Studies show that the nuclear factor-κB (NF-κB)-mediated activation of Snail transcription plays an important rolein the regulation of EMT in tumor cells [19–21]. NF-κB pathway regulates Snail expression through transcriptionaland post-translational mechanisms, and NF-κB increases the transcription of Snail [22]. In this work, we found thatNF-κB expression and activity decreased following apigenin treatment in colon carcinoma cells. Apigenin also inhib-ited the relocation of NF-κB to the nucleus. NF-κB expression and activity decreased following apigenin treatmentin colon carcinoma cells. Apigenin also inhibited the relocation of NF-κB and Snail to the nucleus. This result isconsistent with the earlier finding that NF-κB/Snail is crucial to the ability of apigenin to inhibit tumor progression.

Apigenin is a nontoxic, naturally occurring dietary flavonoid, which is abundant in vegetables, fruits, and beans[23]. Many researches show that apigenin has anti-inflammatory, antioxidant, and anticancer properties [24,25]. Be-cause apigenin is plant-derived natural compound, it may be an ideal and safe antitumor agent [26]. Studies showthat apigenin has good antitumor activities in vitro and in vivo [27–30] and inhibits the growth of various cancer celllines [31]. Different studies have found various molecular mechanisms underlying the anticancer effect of apigenin


9

Dow


/HTTPH




[32]. Apigenin can strongly inhibit the growth of breast cancer cells, including HER2-positive cells [33] and suppressthe metastasis of human hepatocellular carcinoma by inhibiting EMT [34]. Although some researchers indicate thatapigenin can induce apoptosis and autophagy in HCT116 colon cancer cells [35], the effect of apigenin on the EMTof colon cancer remains unclear to date. Accordingly, the present study evaluated the effect of apigenin on EMT ofcolon cancer cells in vitro and in vivo and clarified the mechanism of apigenin regulating EMT of colon cancer cells.We found that apigenin inhibited migration and invasion of colon carcinoma cells. Apigenin inhibited the EMT ofHCT-116 and LOVO human colon cancer cells through NF-κB/Snail signaling pathway. The present study evaluatedthe efficacy of apigenin in the treatment of colorectal cancer at the animal level by establishing xenografts on Balbcnude mice. Actually, PDX (patient derived xenografts) model is closer to the characteristics of clinical tumor sam-ples. In the future, we will further evaluate the efficacy of apigenin in the treatment of colorectal cancer based on PDXmodel.

In summary, we found that Snail is a more important EMT transcription factors for colon cancer prognosis, com-pared with Twist1 and Slug. After screening a series of flavonoid Chinese herbal monomers, we found that apigenineffectively inhibited the activity of Snail. And we found that apigenin suppressed the EMT, migration, and invasionof human colon cancer by inhibiting the NF-κB/Snail pathway. Our results provide a new mechanistic basis for thetherapeutic application of apigenin in patients with colon cancer. Further exploration of apigenin and its mechanismscould lead to the development of a new therapeutic approach to treating colon cancer.

FundingThe present study was funded by Foundation of Shanghai Municipal Science and Technology Commission Project: Preventionand Treatment of Colorectal Cancer by TCM syndrome differentiation [grant number 17dz2307500] and Clinical Study on ChangJi Tai Combined with Auricular Acupuncture in Treating Postoperative Cancer-related fatigue of Postoperative colorectal cancer,Special item of Shanghai Integrative Medicine [grant number ZY(2018-2020)-FWTX-3016].

Competing InterestsThe authors declare that there are no competing interests associated with the manuscript.

Author ContributionJ.T. was focussed on the integrity of the entire study, study concepts, manuscript editing and manuscript review. Y.S. and Z.Z.were involved in the study design, data acquisition, and statistical analysis. Y.H. and X.Z. were dedicated to the literature researchand data analysis. L.H. carried out the manuscript preparation and manuscript review. All authors have read and approved thisarticle.

AbbreviationsEMT , epithelial-mesenchymal transition; NF-κB, nuclear factor-κB; PDX , patient derived xenograft; TCGA, The CancerGenome Atlas.

References1 Weekes, J., Lam, AK-Y, Sebesan, S. and Ho, Y.-H. (2009) Irinotecan therapy and molecular targets in colorectal cancer: a systemic review. World J.

Gastroenterol. 15, 3597–3602, https://doi.org/10.3748/wjg.15.35972 Bates, R.C. and Mercurio, A.M. (2005) The epithelial-mesenchymal transition (EMT) and colorectal cancer progression. Cancer Biol. Ther. 4, 365–370,

https://doi.org/10.4161/cbt.4.4.16553 Lefort, E.C. and Blay, J. (2011) The dietary flavonoid apigenin enhances the activities of the anti-metastatic protein CD26 on human colon carcinoma

cells. Clin. Exp. Metastasis 28, 337–349, https://doi.org/10.1007/s10585-010-9364-64 Newman, D.J. and Cragg, G.M. (2007) Natural products as sources of new drugs over the last 25 years. J. Nat Prod. 70, 461–477,

https://doi.org/10.1021/np068054v5 Nie, J., Zhao, C., Deng, L.I. et al. (2016) Efficacy of traditional Chinese medicine in treating cancer. Biomed. Rep. 4, 3–14,

https://doi.org/10.3892/br.2015.5376 Yu, Q., Zhang, K., Wang, X., Liu, X. and Zhang, Z. (2010) Expression of transcription factors snail, slug, and twist in human bladder carcinoma. J. Exp.

Clin. Cancer Res. 29, 119, https://doi.org/10.1186/1756-9966-29-1197 Rashtak, S., Rego, R., Sweetser, S.R. and Sinicrope, F.A. (2017) Sessile serrated polyps and colon cancer prevention. Cancer Prev. Res. (Phila.) 10,

270–278, https://doi.org/10.1158/1940-6207.CAPR-16-02648 Siveen, K.S., Ahn, K.S., Ong, T.H. et al. (2014) Y-tocotrienol inhibits angiogenesis-dependent growth of human hepatocellular carcinoma through

abrogation of AKT/mTOR pathway in an orthotopic mouse model. Oncotarget 5, 1897–1911, https://doi.org/10.18632/oncotarget.18769 Cai, W., Xiong Chen, Z., Rane, G. et al. (2017) Wanted DEAD/H or alive: helicases winding up in cancers. J. Natl Cancer Inst. 109,

https://doi.org/10.1093/jnci/djw278

10 © 2019 The Author(s). This is an open access article published by Portland Press Limited on behalf of the Biochemical Society and distributed under the Creative CommonsAttribution License 4.0 (CC BY).

Dow


/HTTPH



https://doi.org/10.3748/wjg.15.3597

https://doi.org/10.4161/cbt.4.4.1655

https://doi.org/10.1007/s10585-010-9364-6

https://doi.org/10.1021/np068054v

https://doi.org/10.3892/br.2015.537

https://doi.org/10.1186/1756-9966-29-119

https://doi.org/10.1158/1940-6207.CAPR-16-0264

https://doi.org/10.18632/oncotarget.1876

https://doi.org/10.1093/jnci/djw278


10 Siveen, K.S., Nguyen, A.H., Lee, J.H. et al. (2014) Negative regulation of signal transducer and activator of transcription-3 signalling cascade by lupeolinhibits growth and induces apoptosis in hepatocellular carcinoma cells. Br. J. Cancer 111, 1327, https://doi.org/10.1038/bjc.2014.422

11 Hay, E.D. (1995) An overview of epithelio-mesenchymal transformation. Acta Anat. (Basel) 154, 8–20, https://doi.org/10.1159/00014774812 Bates, R.C. (2005) Colorectal cancer progression: integrin alphavbeta6 and the epithelial-mesenchymal transition (EMT). Cell Cycle 4, 1350–1352,

https://doi.org/10.4161/cc.4.10.205313 Bellovin, D.I., Bates, R.C., Muzikansky, A., Rimm, D.L. and Mercurio, A.M. (2005) Altered localization of p120 catenin during epithelial to mesenchymal

transition of colon carcinoma is prognostic for aggressive disease. Cancer Res. 65, 10938–10945, https://doi.org/10.1158/0008-5472.CAN-05-194714 Wu, Y. and Zhou, B.P. (2010) TNF-alpha/NF-kappaB/Snail pathway in cancer cell migration and invasion. Br. J. Cancer 102, 639–644,

https://doi.org/10.1038/sj.bjc.660553015 Brzozowa, M., Michalski, M., Wyrobiec, G. et al. (2015) The role of Snail1 transcription factor in colorectal cancer progression and metastasis.

Contemp. Oncol. (Pozn.) 19, 265–27016 Wijnhoven, B.P., Dinjens, W.N. and Pignatelli, M. (2000) E-cadherin-catenin cell-cell adhesion complex and human cancer. Br. J. Surg. 87, 992–1005,

https://doi.org/10.1046/j.1365-2168.2000.01513.x17 Chao, Y.L., Shepard, C.R. and Wells, A. (2010) Breast carcinoma cells re-express E-cadherin during mesenchymal to epithelial reverting transition. Mol.

Cancer 9, 179, https://doi.org/10.1186/1476-4598-9-17918 Bae, K.-M., Parker, N.N., Dai, Y., Vieweg, J. and Siemann, D.W. (2011) E-cadherin plasticity in prostate cancer stem cell invasion. Am. J. Cancer Res. 1,

71–8419 Julien, S., Puig, I., Caretti, E. et al. (2007) Activation of NF-kappaB by Akt upregulates Snail expression and induces epithelium mesenchyme transition.

Oncogene 26, 7445–7456, https://doi.org/10.1038/sj.onc.121054620 Liu, Y., Tergaonkar, V., Krishna, S. and Androphy, E.J. (1999) Human papillomavirus type 16 E6-enhanced susceptibility of L929 cells to tumor necrosis

factor alpha correlates with increased accumulation of reactive oxygen species. J. Biol. Chem. 274, 24819–24827,https://doi.org/10.1074/jbc.274.35.24819

21 Chakrabarti, O., Veeraraghavalu, K., Tergaonkar, V., Liu, Y., Androphy, E.J., Stanley, M.A. et al. (2004) Human papillomavirus type 16 E6 amino acid 83variants enhance E6-mediated MAPK signaling and differentially regulate tumorigenesis by notch signaling and oncogenic Ras. J. Virol. 78,5934–5945, https://doi.org/10.1128/JVI.78.11.5934-5945.2004

22 Barbera, M.J., Puig, I., Dominguez, D. et al. (2004) Regulation of Snail transcription during epithelial to mesenchymal transition of tumor cells.Oncogene 23, 7345–7354, https://doi.org/10.1038/sj.onc.1207990

23 Wang, W., VanAlstyne, P.C., Irons, K.A., Chen, S., Stewart, J.W. and Birt, D.F. (2004) Individual and interactive effects of apigenin analogs on G2/Mcell-cycle arrest in human colon carcinoma cell lines. Nutr. Cancer 48, 106–114, https://doi.org/10.1207/s15327914nc4801˙14

24 Lefort, E.C. and Blay, J. (2013) Apigenin and its impact on gastrointestinal cancers. Mol. Nutr. Food Res. 57, 126–144,https://doi.org/10.1002/mnfr.201200424

25 Janssen, K., Mensink, R.P., Cox, F.J., Harryvan, J.L., Hovenier, R., Hollman, P.C. et al. (1998) Effects of the flavonoids quercetin and apigenin onhemostasis in healthy volunteers: results from an in vitro and a dietary supplement study. Am. J. Clin. Nutr. 67, 255–262,https://doi.org/10.1093/ajcn/67.2.255

26 Patel, D., Shukla, S. and Gupta, S. (2007) Apigenin and cancer chemoprevention: progress, potential and promise (review). Int. J. Oncol. 30, 233–24527 Wang, Q.R., Yao, X.Q., Wen, G. et al. (2011) Apigenin suppresses the growth of colorectal cancer xenografts via phosphorylation and up-regulated FADD

expression. Oncol. Lett. 2, 43–47, https://doi.org/10.3892/ol.2010.21528 Lepley, D.M. and Pelling, J.C. (1997) Induction of p21/WAF1 and G1 cell-cycle arrest by the chemopreventive agent apigenin. Mol. Carcinog. 19,

74–82, https://doi.org/10.1002/(SICI)1098-2744(199707)19:2%3c74::AID-MC2%3e3.0.CO;2-L29 Hu, X.-W., Meng, D. and Fang, J. (2008) Apigenin inhibited migration and invasion of human ovarian cancer A2780 cells through focal adhesion kinase.

Carcinogenesis 29, 2369–2376, https://doi.org/10.1093/carcin/bgn24430 Shukla, S. and Gupta, S. (2010) Apigenin: a promising molecule for cancer prevention. Pharm. Res. 27, 962–978,

https://doi.org/10.1007/s11095-010-0089-731 Salmani, J.M.M., Zhang, X.-P., Jacob, J.A. and Chen, B.-A. (2017) Apigenin’s anticancer properties and molecular mechanisms of action: Recent

advances and future prospectives. Chin. J. Nat. Med. 15, 321–32932 Liu, L.-Z., Fang, J., Zhou, Q., Hu, X., Shi, X. and Jiang, B.-H. (2005) Apigenin inhibits expression of vascular endothelial growth factor and angiogenesis

in human lung cancer cells: implication of chemoprevention of lung cancer. Mol. Pharmacol. 68, 635–64333 Scherbakov, A.M. and Andreeva, O.E. (2015) Apigenin inhibits growth of breast cancer cells: the role of ERalpha and HER2/neu. Acta. Naturae 7,

133–13934 Qin, Y., Zhao, D., Zhou, H.-G. et al. (2016) Apigenin inhibits NF-kappaB and snail signaling, EMT and metastasis in human hepatocellular carcinoma.

Oncotarget 7, 41421–41431, https://doi.org/10.18632/oncotarget.940435 Shao, H., Jing, K., Mahmoud, E., Huang, H., Fang, X. and Yu, C. (2013) Apigenin sensitizes colon cancer cells to antitumor activity of ABT-263. Mol.

Cancer Ther. 12, 2640–2650, https://doi.org/10.1158/1535-7163.MCT-13-0066


11

Dow


/HTTPH



https://doi.org/10.1038/bjc.2014.422

https://doi.org/10.1159/000147748

https://doi.org/10.4161/cc.4.10.2053

https://doi.org/10.1158/0008-5472.CAN-05-1947

https://doi.org/10.1038/sj.bjc.6605530

https://doi.org/10.1046/j.1365-2168.2000.01513.x

https://doi.org/10.1186/1476-4598-9-179

https://doi.org/10.1038/sj.onc.1210546

https://doi.org/10.1074/jbc.274.35.24819

https://doi.org/10.1128/JVI.78.11.5934-5945.2004

https://doi.org/10.1038/sj.onc.1207990

https://doi.org/10.1207/s15327914nc4801_14

https://doi.org/10.1002/mnfr.201200424

https://doi.org/10.1093/ajcn/67.2.255

https://doi.org/10.3892/ol.2010.215

https://doi.org/10.1002/(SICI)1098-2744(199707)19:2%3c74::AID-MC2%3e3.0.CO;2-L

https://doi.org/10.1093/carcin/bgn244

https://doi.org/10.1007/s11095-010-0089-7

https://doi.org/10.18632/oncotarget.9404

https://doi.org/10.1158/1535-7163.MCT-13-0066

Apigenin inhibits epithelial-mesenchymal transition of ... · Human colon carcinoma cell lines...

Documents

Transcript of Apigenin inhibits epithelial-mesenchymal transition of ... · Human colon carcinoma cell lines...