Apigenin inhibits proliferation and invasion, and induces apoptosis and cell cycle arrest in human melanoma cells

  • Authors:
    • Guangming Zhao
    • Xiaodong Han
    • Wei Cheng
    • Jing Ni
    • Yunfei Zhang
    • Jingrong Lin
    • Zhiqi Song
  • View Affiliations

  • Published online on: February 14, 2017     https://doi.org/10.3892/or.2017.5450
  • Pages: 2277-2285
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Abstract

Malignant melanoma is the most invasive and fatal form of cutaneous cancer. Moreover it is extremely resistant to conventional chemotherapy and radiotherapy. Apigenin, a non-mutagenic flavonoid, has been found to exhibit chemopreventive and/or anticancerogenic properties in many different types of human cancer cells. Therefore, apigenin may have particular relevance for development as a chemotherapeutic agent for cancer treatment. In the present study, we investigated the effects of apigenin on the viability, migration and invasion potential, dendrite morphology, cell cycle distribution, apoptosis, phosphorylation of the extracellular signal-regulated protein kinase (ERK) and the AKT/mTOR signaling pathway in human melanoma A375 and C8161 cell lines in vitro. Apigenin effectively suppressed the proliferation of melanoma cells in vitro. Moreover, it inhibited cell migration and invasion, lengthened the dendrites, and induced G2/M phase arrest and apoptosis. Furthermore, apigenin promoted the activation of cleaved caspase-3 and cleaved PARP proteins and decreased the expression of phosphorylated (p)‑ERK1/2 proteins, p-AKT and p-mTOR. Consequently, apigenin is a novel therapeutic candidate for melanoma.

Introduction

Malignant melanoma has recently been reported to have one of the highest incidence rates among all types of cancer, with an increasing number of melanoma-related deaths each year (1). The principal cause of death in melanoma patients is attributed to widespread metastases to the lymphatic system and other organs (2). Following traditional therapy, the average survival time of patients with metastatic melanoma is only 6–12 months and the 5-year survival rate is consistently <10% in most cases (3). However, tremendous progress in both immunotherapy and molecular-targeted therapy has revolutionized the standard of care for terminal melanoma patients. In the meantime, some new challenges for clinicians have also surfaced (4). One such example is molecular-targeted therapy which often leads to fast-acting and significant responses in most patients with the targeted mutation, while the clinical benefit is usually transient due to the rapid emergence of drug resistance. Consequently, it is urgent to develop efficient agents that may be applied for melanoma treatment.

Apigenin, a natural plant flavonoid (4′,5,7-trihydroxyflavone), is widespread in common fruits and vegetables. According to the Biopharmaceutics Classification System, apigenin is categorized as a class II drug with poor solubility and high intestinal membrane permeability (5). The oral bioavailability of apigenin is relatively low due to its low solubility in water (~2.16 µg/ml) (5) and in high hydrophilic or nonpolar solvents (0.001–1.63 mg/ml) (6), which has extremely hampered its clinical development. Several formulation strategies have been investigated to improve the bioavailability for application, including liposome (7) and nanocrystals fabricated by high pressure homogenization (8).

Apigenin has been shown to have marked anti-inflammatory, antioxidant and anticarcinogenic properties (9). Recently researchers have demonstrated that apigenin has an anti-proliferative effect on a variety of cancer cells, such as bladder, ovarian, breast and prostate cancer (1015), including melanoma (16,17). Apoptosis plays a critical role in controlling cell proliferation and thus is pivotal for the prevention of cancer progression and oncogenesis (18). Extracellular signal-regulated kinase (ERK) is a crucial signaling molecule that regulates cell survival and proliferation. The ERK signaling pathway controls various pro- and anti-apoptotic mechanisms that determine cell viability (19). AKT serves as an anti-apoptotic signaling molecule and inhibits apoptosis through mitochondrial pathways (20). Consequently, in the present study, we investigated the effects of apigenin on the viability, migration and invasion potential, dendrite morphology, cell cycle distribution, apoptosis, ERK expression and the AKT/mTOR signaling pathway.

Materials and methods

Chemicals and reagents

Apigenin (no. A0113, CAS: 520-36-5, purity ≥98%) was purchased from Chengdu Must Bio-Technology Co., Ltd. (Chengdu, China). Dulbecco's modified Eagle's medium (DMEM), trypsin and fetal bovine serum (FBS) were purchased from Gibco BRL (Grand Island, NY, USA). Dimethyl sulfoxide (DMSO), 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT), Triton X-100 and anti-β-actin antibody were purchased from Sigma Chemical Co. (St. Louis, MO, USA). Trypsin free of ethylenediaminetetraacetic acid (EDTA) was purchased from Hyclone Co. (Logan, UT, USA). Matrigel was purchased from BD Biosciences (Franklin Lakes, NJ, USA). FITC-Annexin V kit was obtained from Nanjing KeyGen Biotech Co., Ltd. (Nanjing, China). Propidium iodide (PI) and RNase were purchased from Takara Bio, Inc. (Otsu, Shiga, Japan). Antibodies for ERK1/2 and phosphorylated (p)-ERK1/2 were purchased from Promega Corp. (Madison, WI, USA). The antibodies for poly(ADP-ribose) polymerase (PARP), caspase-3, AKT, p-AKT (Ser473), mTOR and p-mTOR (Ser2448) were purchased from Cell Signaling Technology Inc. (Beverly, MA, USA).

Cell culture and apigenin treatment

Human malignant melanoma A375 and C8161 cell lines were obtained from Peking Union Cell Resource Center (Beijing, China). The cells were grown at 37°C in a humidified atmosphere containing 5% CO2. The cells were cultured and maintained in DMEM supplemented with 1% penicillin-streptomycin and 10% FBS. A375 and C8161 cells were treated with different concentrations of apigenin (dissolved in DMSO) whereas the control cells were treated with an equivalent volume of DMSO.

MTT assays

For cell proliferation assays, the A375 and C8161 cells were seeded in 96-well plates at a concentration of 1×104 cells/well. Cells were allowed to adhere for 24 h and subsequently exposed to different concentrations of apigenin (40, 80, 120, 160, 200, 240 and 280 µM) and incubated at 37°C for 24, 48, 72 and 96 h. MTT solution was added to each well at the specified time-point and incubated for an additional 4 h. The culture medium in each well was discarded and replaced with DMSO to dissolve the formazan crystals which were formed from the MTT. The absorbance value was evaluated using an automatic microplate reader (T17108U; PerkinElmer, Inc., Waltham, MA, USA) at 490 nm.

Cell migration assays in vitro

Cell migration was performed using the wound healing assay. A375 and C8161 cells were seeded in a 24-well plate at a concentration of 5×105 cells/well and allowed to form a confluent monolayer for 24 h. The monolayer was scratched with a sterile pipette tip (10 µl) then washed with serum-free medium to remove the floating and detached cells. After treatment with 40 and 80 µM of apigenin or DMSO, the cells were observed and photographed (time 0 h and 24 h) using an inverted microscope (Olympus Corporation, Tokyo, Japan). Moreover, the number of cells migrating to the wound was assessed. Data were obtained from three independent experiments.

Cell invasion assay

Cell culture inserts (24-well, pore size 8 µm; BD Biosciences) were seeded with 1×106 cells/ml in 100 µl of serum-free medium with 40 µM apigenin, or DMSO. Inserts were precoated with 10 µl of Matrigel (3 mg/ml; Becton-Dickinson, Mountain View, CA, USA). Medium with 10% FBS (500 µl) was added to the lower chamber and served as a chemotactic agent. After incubation for 72 h, non-invasive cells were wiped from the upper surface of the membrane. Cells on the lower side were fixed with chilled methanol, stained with crystal violet (dissolved in methanol) and counted using an inverted microscope. Each individual experiment had triplicate inserts and five random, non-overlapping fields at a magnification of ×200 were counted per insert.

Scanning electron microscopy

A375 and C8161 cells were plated at a concentration of 2×104 cells/well into a 60-mm culture dish. After treatment with 100 µM of apigenin or DMSO for 24 h, the cells were harvested, washed with PBS and fixed with 2.5% glutaraldehyde and 1% osmium tetraoxide, followed by an increasing gradient dehydration step using ethanol solutions of 50, 70, 95 and 100%. Samples were sputter-coated with platinum and palladium before being observed under a scanning electron microscope (Quanta 200F; FEI, Hillsboro, OR, USA).

Cell apoptosis

Cells were placed in 6-well culture plates (5×104 cells/ml) and allowed to attach for 8 h. A375 and C8161 cells were treated with apigenin (40 and 100 µM, respectively) or DMSO for 24 h. Following the manufacturers instructions, the cells were harvested by trypsinization free of EDTA, washed in cold PBS and resuspended in binding buffer at a concentration of 1×106 cells/ml. FITC-conjugated Annexin V (BioVision, Inc., Milpitas, CA, USA) and PI (5 µl each) (Becton-Dickinson) were added to the cells, gently mixed and then incubated for 15 min at room temperature in the dark. Afterwards binding buffer was added and the cells were analyzed by flow cytometry.

Cell cycle analysis

Cells were seeded in 60-mm culture dishes. After attachment, the cells were treated with 100 µM apigenin or DMSO for 24 h. Then cells were harvested and fixed with ice-cold 75% ethanol. The cell pellets were resuspended in binding buffer consisting of 480 µl PBS, 5 µl PI (5 mg/ml), 5 µl RNase (10 mg/ml) and 10 µl Triton X-100 (10%). After 30 min of incubation at room temperature in the dark, the DNA content of the cells was examined using a flow cytometer (Accuri C6; Becton-Dickinson) for cell cycle phase distribution.

Western blot analysis

Cells were plated in 6-well culture plates at concentrations determined to yield 60–70% confluence within 24 h. Next, the cells were left untreated or treated with 100 µM apigenin for 24 h. After preparing appropriate protein concentrations of 25 µg, sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) was performed. Proteins were separated by electrophoresis and transferred onto nitrocellulose membranes and afterwards blocked for 1 h with 5% non-fat dry milk in TBS-T. The membranes were incubated with respective primary antibodies at appropriate concentrations overnight at 4°C. After being washed to remove unbound primary antibodies, they were incubated with the corresponding secondary antibodies. Proteins were visualized by image scanning and the optical density for each band was assessed using Image Lab software (version 4.0; Bio-Rad, Hercules, CA, USA) after data were normalized to β-actin as an internal reference.

Statistical analysis

All the experiments were carried out in triplicate and the values are expressed as the mean ± standard deviation (SD). SPSS v17.0 software (SPSS, Inc., Chicago, IL, USA) was used for statistical analysis. The repeated experiments used analysis of variance, Dunnetts test and Student's t-test for the assessment of differences between groups. A probability value of ≤0.05 was deemed statistically significant. *P<0.05, **P<0.01 and ***P<0.001 as indicated in the figures, are relative to the controls.

Results

Apigenin inhibits A375 and C8161 cell proliferation

To investigate the growth inhibitory effect of apigenin, A375 and C8161 cells were treated with different concentrations (40, 80, 120, 160, 200, 240 and 280 µM) of apigenin for different periods of time (24, 48, 72 and 96 h). The cell viability was assessed by MTT assay. As shown in Fig. 1, cell growth inhibition caused by apigenin was relatively marked in a dose-dependent, as well as a time-dependent manner (ranging from 40–160 µM within 48 h). The IC50 value at 24 h was estimated to be 100 µM.

Apigenin inhibits A375 and C8161 cell migration potential

To assess whether or not apigenin has an effect on the metastasis of A375 and C8161 cell lines, we examined the number of migratory cells using a wound-healing approach. Migration was significantly inhibited in the A375 and C8161 cell lines after treatment with apigenin (40 and 80 µM) for 24 h (P<0.001) (Fig. 2). When the cells were exposed to 100 µM of apigenin for 24 h, no migrating cells were observed (data not shown).

Apigenin suppresses the invasion of A375 and C8161 cells

We further investigated cell motility by invasion assays. Both the density of the invasive cells on the membrane and the number of invasive cells/field are shown in Fig. 3. Treatment with 40 µM of apigenin for 72 h significantly decreased the invasive ability of the A375 and C8161 melanoma cells compared with the control cells (P<0.001). Following treatment with 80 µM of apigenin, no invading cells were observed (data not shown). These findings demonstrated that apigenin decreased the invasion of melanoma cells in vitro.

Apigenin affects the dendrite morphology of A375 and C8161 cells

Following treatment with 100 µM of apigenin for 24 h, both the A375 and C8161 cells changed their cellular morphology as visualized using scanning electron microscopy. The dendrites became thinner and longer compared to those noted in the untreated control cells (P<0.001) (Fig. 4).

Apigenin promotes the apoptosis of A375 and C8161 cells

To ascertain the underlying mechanism which leads to apigenin-induced inhibition of cell proliferation, we observed the effects of apigenin on the A375 and C8161 cells by detecting their apoptotic rates. Significant apoptosis was found in both the A375 and C8161 cell lines. Treatment with apigenin (40 and 100 µM) for 24 h, resulted in higher apoptosis rates of the A375 and C8161 cells than the rates noted in the untreated control cells, and the effects occurred in a dose-dependent manner (P<0.01, P<0.001) (Fig. 5).

Apigenin arrests A375 and C8161 cells at the G2/M phase of the cell cycle

Cell cycle analysis was also performed by flow cytometry. A375 and C8161 cells were treated with 100 µM of apigenin for 24 h. As shown in Fig. 6, apigenin treatment resulted in a noticeable accumulation of cells in the G2/M phase with a decrease in the G0/G1 phase during the cell cycle (P<0.05, P<0.01, P<0.001). This blockage of cell progression may be one of the mechanisms by which apigenin exerts its anti-proliferative effect on melanoma cell lines.

Apigenin alters ERK protein expression

Western blot analysis showed that treatment with 100 µM of apigenin significantly increased the expression of cleaved caspase-3 and cleaved PARP in the A375 and C8161 cells, while it decreased the expression of p-ERK1/2 but did not alter the total ERK1/2 level as compared to the DMSO controls (P<0.05, P<0.01, P<0.001) (Fig. 7).

Apigenin inhibits the AKT signaling pathway

Western blot analysis revealed that the expression levels of phosphorylated AKT (p-AKT) and p-mTOR were decreased after treatment with 100 µM of apigenin, whereas no significant difference was observed in total AKT and mTOR, in comparison to the DMSO controls (P<0.001) (Fig. 8).

Discussion

Melanoma is one of the most malignant cancers with a propensity for metastases. The well-established conventional treatments for melanoma, such as cryotherapy, surgery, and chemotherapy (21) and some nonsurgical treatments are usually limited to adjuvant therapies. Therefore, increasing interest has focused on the search for natural dietary phytochemicals both safe and effective against melanoma. It is generally known that many compounds from natural plants possess chemopreventative and chemotherapeutic efficacy in human cancers including melanoma (22). Apigenin, a flavonoid belonging to the flavone subgroup, is present in various vegetables, herbs, fruits and Chinese traditional medications (9,23) and has been shown to suppress tumor growth through inhibition of cell proliferation (24).

In the present study, we investigated the chemotherapeutic capacity of apigenin against human melanoma. We selected the human melanoma A375 and C8161 cell lines which have a different BRAF mutation status. A375 cells harbor the BRAFV600E mutation while C8161 cells contain the BRAF wild-type gene. Apigenin, as shown in this study, significantly suppressed the growth of A375 and C8161 cells. These data suggest that apigenin possesses chemotherapeutic potential against human melanoma.

Dysregulation of the cell cycle is a hallmark of tumorigenesis. The cell cycle is controlled at multiple checkpoints. The G2/M checkpoint inhibits cells from entering mitosis when DNA is impaired, enabling cell repair. Pathways that result in apoptosis may be activated when the damage is irreparable (25). The G2/M checkpoint is controlled by Cdc2/cyclin B, as well as their negative regulators including p21Cip1 and p27 (26). Regulating these G2/M checkpoint proteins may enhance the sensitivity of cancer cells to radiotherapy and chemotherapy (27). Therefore, the G2/M checkpoint is a potential target for cancer therapy. It has been reported that apigenin arrested human colon cancer HCT116 cells in the G2/M phase. Moreover, it suppressed the expression of both cyclin B1 and its activating partners, Cdc2 and Cdc25c (28). In addition, apigenin treatment led to a significant accumulation of cells in the G2/M phase via the downregulation of Cdc25c expression in human papillary thyroid carcinoma BCPAP cells (29). We found that apigenin suppressed the growth of human melanoma A375 and C8161 cells by inducing G2/M phase arrest of the cell cycle. Furthermore, apigenin treatment decreased the expression of p-AKT and p-mTOR. Previous studies have indicated that the AKT/mTOR pathway could influence the progression of G2 to the mitosis phase through the regulation of the expression of G2/M-related proteins (30). Expression of the active form of AKT increases Cdk1 both at the protein and mRNA level, while its predominant negative mutation suppresses cell proliferation by inducing G2/M arrest (31). Consequently, apigenin may inhibit proliferation of A375 and C8161 cells by arresting G2/M transition in the cell cycle probably via the AKT/mTOR pathway.

Invasion and metastasis are major concerns in the prognosis and progression of cancer. The AKT/mTOR pathway is pivotal in modulating the invasion and migration of tumor cells (32). This pathway promotes resistance to chemotherapy-induced apoptosis in a variety of cancers including melanoma (33). We found that 40 µM of apigenin significantly inhibited cell migration and invasion. Furthermore, western blot analysis demonstrated that the expression levels of p-AKT and p-mTOR were decreased after apigenin treatment, while no significant difference was observed in total AKT and mTOR. These results indicate that the AKT/mTOR pathway plays an important role in the apigenin-induced inhibition of migration and invasion of A375 and C8161 cells. Erdogan et al (34) also showed that apigenin reduced prostate cancer CD44+ stem cell survival and migration through PI3K/AKT/NF-κB signaling.

Apoptosis, a type of programmed cell death, is a physiological process essential for normal tissue development (35). In mammals, there are two primary apoptotic pathways: the extrinsic pathway (death receptor-mediated pathway) and the intrinsic pathway (mitochondrial-mediated pathway) (36). Caspase-3 is a key executioner caspase and its activation leads to the cleavage of PARP during cell death (37). Cleavage of PARP is regarded as a central indicator of apoptosis. The ERK and AKT signaling pathways are related to cell biological functions and cancer malignancies which could also play an important role in the proliferation and apoptosis of cancers. In our study, we confirmed that apigenin treatment resulted in higher apoptosis rates of the A375 and C8161 cells compared to the control cells in a dose-dependent manner and significantly increased the expression of cleaved caspase-3 and cleaved PARP, while it decreased the expression of p-ERK1/2. The cell death occurring in the A375 and C8161 cells treated with apigenin was probably induced by apoptosis, which was possibly involved with ERK phosphorylation. Pretreatment of A375 and C8161 cells with ERK inhibitors is necessary to reveal whether apigenin-induced apoptosis is dependent on ERK activity and we will investigate the relevant mechanism in future research. Seo et al demonstrated that apigenin induced apoptosis via the extrinsic pathway, increasing p53 and inhibiting STAT3 and NF-κB signaling in HER2-overexpressing breast cancer cells (38). Shukla et al observed that apigenin induced apoptosis by targeting inhibitor of apoptosis proteins and Ku70-Bax interaction in prostate cancer (39). Das et al (40) found that apigenin induced apoptosis in A375 and A549 cells through selective action and dysfunction of mitochondria, suggesting the activation of the intrinsic apoptosis pathway. However, we did not ascertain whether apigenin induced cell apoptosis through the intrinsic or extrinsic pathway and this relevant research will be carried out in the future. In addition, our western blot analysis showed that the expression of p-AKT and p-mTOR was decreased after apigenin treatment, while the expression of total AKT and mTOR was not altered. This indicates that the AKT/mTOR pathway plays a vital role in the apigenin-induced apoptosis observed in A375 and C8161 cells. Zhu et al (30) demonstrated that apigenin induced apoptosis via the PI3K/AKT pathway, the regulation of the Bcl-2 family and activation of caspase-3 and PARP.

Recent studies have implicated glutamate signaling in the development of melanoma (4143). The antagonists of ionotropic glutamate receptors (iGluRs) have been demonstrated to cause a rapid and reversible change in melanocyte morphology. Metabotropic glutamate 1 (mGlu1) receptor has been proposed as a target for metastatic melanoma therapy (44). It is expressed aberrantly in over half of human melanoma cell lines and biopsies (45). In our previous study (46), we found that the antagonists mGlu1 receptor and N-methyl-D-aspartate (NMDA) receptor increased dendritic branching and inhibited the motility, migration and proliferation of the human metastatic melanoma cell line WM451. We also demonstrated that the invasion and motility effects were significantly inhibited by the combination of increased microtubule-associated protein (MAP)2a (MAP2a) expression and either an mGlu1 receptor or NMDA receptor antagonist. One plausible explanation for this phenomenon is that the blockade of the glutamate-mediated signaling pathway via the ERK1/2 pathway suppresses cell motility and invasion through a tubulin-dependent mechanism (47). In the present study, we found that the main cytodendrites of human melanoma A375 and C8161 cells following treatment with apigenin became thinner and longer than those of the controls. Moreover, treatment with apigenin significantly suppressed cell invasion and migration. We deduced that the aforementioned effects of apigenin may be induced by blocking the glutamate-mediated signaling pathway, leading to cytoskeletal protein reorganization and tumor cell differentiation. These results suggest that the blockade of glutamate signaling is a promising novel therapy for the treatment of melanoma.

In conclusion, apigenin is a potent suppressor of cell viability, migration and invasion. Concomitantly, it induces apoptosis in human melanoma A375 and C8161 cells, via activation of caspase-3 and PARP, inhibition of ERK phosphorylation and the AKT/mTOR pathway. Furthermore, it affects the dendrite morphology of the A375 and C8161 cells, which might be involved with the blockade of the glutamate signaling pathway. These findings need to be supported by further experimental evidence. Consequently, apigenin exhibits effective antineoplastic potency and provides a hopeful treatment paradigm for melanoma.

Acknowledgements

We would like to thank Quentin Liu for guidance in our study. The present study was supported by grants from the National Natural Science Foundation of China (nos. 81472865 and 81171491) and the Natural Science Foundation of Liaoning Province (no. 201102056).

References

1 

Liu J, Gu J, Feng Z, Yang Y, Zhu N, Lu W and Qi F: Both HDAC5 and HDAC6 are required for the proliferation and metastasis of melanoma cells. J Transl Med. 14:72016. View Article : Google Scholar : PubMed/NCBI

2 

Agarwala SS: Current systemic therapy for metastatic melanoma. Expert Rev Anticancer Ther. 9:587–595. 2009. View Article : Google Scholar : PubMed/NCBI

3 

Balch CM, Gershenwald JE, Soong SJ, Thompson JF, Atkins MB, Byrd DR, Buzaid AC, Cochran AJ, Coit DG, Ding S, et al: Final version of 2009 AJCC Melanoma Staging and Classification. J Clin Oncol. 27:6199–6206. 2009. View Article : Google Scholar : PubMed/NCBI

4 

Zhu Z, Liu W and Gotlieb V: The rapidly evolving therapies for advanced melanoma - Towards immunotherapy, molecular targeted therapy, and beyond. Crit Rev Oncol Hematol. 99:91–99. 2015. View Article : Google Scholar : PubMed/NCBI

5 

Zhang J, Liu D, Huang Y, Gao Y and Qian S: Biopharmaceutics classification and intestinal absorption study of apigenin. Int J Pharm. 436:311–317. 2012. View Article : Google Scholar : PubMed/NCBI

6 

Ding SM, Zhang ZH, Song J, Cheng XD, Jiang J and Jia XB: Enhanced bioavailability of apigenin via preparation of a carbon nanopowder solid dispersion. Int J Nanomedicine. 9:2327–2333. 2014. View Article : Google Scholar : PubMed/NCBI

7 

Arsić I, Tadić V, Vlaović D, Homšek I, Vesić S, Isailović G and Vuleta G: Preparation of novel apigenin-enriched, liposomal and non-liposomal, antiinflammatory topical formulations as substitutes for corticosteroid therapy. Phytother Res. 25:228–233. 2011.PubMed/NCBI

8 

Al Shaal L, Shegokar R and Müller RH: Production and characterization of antioxidant apigenin nanocrystals as a novel UV skin protective formulation. Int J Pharm. 420:133–140. 2011. View Article : Google Scholar : PubMed/NCBI

9 

Patel D, Shukla S and Gupta S: Apigenin and cancer chemoprevention: Progress, potential and promise (Review). Int J Oncol. 30:233–245. 2007.PubMed/NCBI

10 

Shi MD, Shiao CK, Lee YC and Shih YW: Apigenin, a dietary flavonoid, inhibits proliferation of human bladder cancer T-24 cells via blocking cell cycle progression and inducing apoptosis. Cancer Cell Int. 15:332015. View Article : Google Scholar : PubMed/NCBI

11 

Suh YA, Jo SY, Lee HY and Lee C: Inhibition of IL-6/STAT3 axis and targeting Axl and Tyro3 receptor tyrosine kinases by apigenin circumvent taxol resistance in ovarian cancer cells. Int J Oncol. 46:1405–1411. 2015.PubMed/NCBI

12 

Shukla S, Bhaskaran N, Babcook MA, Fu P, Maclennan GT and Gupta S: Apigenin inhibits prostate cancer progression in TRAMP mice via targeting PI3K/Akt/FoxO pathway. Carcinogenesis. 35:452–460. 2014. View Article : Google Scholar : PubMed/NCBI

13 

Shukla S, Kanwal R, Shankar E, Datt M, Chance MR, Fu P, MacLennan GT and Gupta S: Apigenin blocks IKKα activation and suppresses prostate cancer progression. Oncotarget. 6:31216–31232. 2015.PubMed/NCBI

14 

Scherbakov AM and Andreeva OE: Apigenin inhibits growth of breast cancer cells: The role of ERα and HER2/neu. Acta naturae. 7:133–139. 2015.PubMed/NCBI

15 

Seo HS, Ku JM, Choi HS, Woo JK, Jang BH, Go H, Shin YC and Ko SG: Apigenin induces caspase-dependent apoptosis by inhibiting signal transducer and activator of transcription 3 signaling in HER2-overexpressing SKBR3 breast cancer cells. Mol Med Rep. 12:2977–2984. 2015.PubMed/NCBI

16 

Caltagirone S, Rossi C, Poggi A, Ranelletti FO, Natali PG, Brunetti M, Aiello FB and Piantelli M: Flavonoids apigenin and quercetin inhibit melanoma growth and metastatic potential. Int J Cancer. 87:595–600. 2000. View Article : Google Scholar : PubMed/NCBI

17 

Ye Y, Chou GX, Wang H, Chu JH and Yu ZL: Flavonoids, apigenin and icariin exert potent melanogenic activities in murine B16 melanoma cells. Phytomedicine. 18:32–35. 2010. View Article : Google Scholar : PubMed/NCBI

18 

Ghobrial IM, Witzig TE and Adjei AA: Targeting apoptosis pathways in cancer therapy. CA Cancer J Clin. 55:178–194. 2005. View Article : Google Scholar : PubMed/NCBI

19 

Pang W, Leng X, Lu H, Yang H, Song N, Tan L, Jiang Y and Guo C: Depletion of intracellular zinc induces apoptosis of cultured hippocampal neurons through suppression of ERK signaling pathway and activation of caspase-3. Neurosci Lett. 552:140–145. 2013. View Article : Google Scholar : PubMed/NCBI

20 

Chan TO, Rittenhouse SE and Tsichlis PN: AKT/PKB and other D3 phosphoinositide-regulated kinases: Kinase activation by phosphoinositide-dependent phosphorylation. Annu Rev Biochem. 68:965–1014. 1999. View Article : Google Scholar : PubMed/NCBI

21 

Lopez RF, Lange N, Guy R and Bentley MV: Photodynamic therapy of skin cancer: Controlled drug delivery of 5-ALA and its esters. Adv Drug Deliv Rev. 56:77–94. 2004. View Article : Google Scholar : PubMed/NCBI

22 

Eggler AL, Gay KA and Mesecar AD: Molecular mechanisms of natural products in chemoprevention: Induction of cytoprotective enzymes by Nrf2. Mol Nutr Food Res. 52:(Suppl 1). S84–S94. 2008.PubMed/NCBI

23 

Shukla S and Gupta S: Apigenin: A promising molecule for cancer prevention. Pharm Res. 27:962–978. 2010. View Article : Google Scholar : PubMed/NCBI

24 

Kim MA, Kang K, Lee HJ, Kim M, Kim CY and Nho CW: Apigenin isolated from Daphne genkwa Siebold et Zucc. inhibits 3T3-L1 preadipocyte differentiation through a modulation of mitotic clonal expansion. Life Sci. 101:64–72. 2014. View Article : Google Scholar : PubMed/NCBI

25 

Wang Y, Ji P, Liu J, Broaddus RR, Xue F and Zhang W: Centrosome-associated regulators of the G2/M checkpoint as targets for cancer therapy. Mol Cancer. 8:82009. View Article : Google Scholar : PubMed/NCBI

26 

Dash BC and El-Deiry WS: Phosphorylation of p21 in G2/M promotes cyclin B-Cdc2 kinase activity. Mol Cell Biol. 25:3364–3387. 2005. View Article : Google Scholar : PubMed/NCBI

27 

Stewart ZA, Westfall MD and Pietenpol JA: Cell-cycle dysregulation and anticancer therapy. Trends Pharmacol Sci. 24:139–145. 2003. View Article : Google Scholar : PubMed/NCBI

28 

Lee Y, Sung B, Kang YJ, Kim DH, Jang JY, Hwang SY, Kim M, Lim HS, Yoon JH, Chung HY, et al: Apigenin-induced apoptosis is enhanced by inhibition of autophagy formation in HCT116 human colon cancer cells. Int J Oncol. 44:1599–1606. 2014.PubMed/NCBI

29 

Zhang L, Cheng X, Gao Y, Zheng J, Xu Q, Sun Y, Guan H, Yu H and Sun Z: Apigenin induces autophagic cell death in human papillary thyroid carcinoma BCPAP cells. Food Funct. 6:3464–3472. 2015. View Article : Google Scholar : PubMed/NCBI

30 

Zhu Y, Mao Y, Chen H, Lin Y, Hu Z, Wu J, Xu X, Xu X, Qin J and Xie L: Apigenin promotes apoptosis, inhibits invasion and induces cell cycle arrest of T24 human bladder cancer cells. Cancer Cell Int. 13:542013. View Article : Google Scholar : PubMed/NCBI

31 

Lee SR, Park JH, Park EK, Chung CH, Kang SS and Bang OS: Akt-induced promotion of cell-cycle progression at G2/M phase involves upregulation of NF-Y binding activity in PC12 cells. J Cell Physiol. 205:270–277. 2005. View Article : Google Scholar : PubMed/NCBI

32 

Hennessy BT, Smith DL, Ram PT, Lu Y and Mills GB: Exploiting the PI3K/AKT pathway for cancer drug discovery. Nat Rev Drug Discov. 4:988–1004. 2005. View Article : Google Scholar : PubMed/NCBI

33 

Lin HP, Jiang SS and Chuu CP: Caffeic acid phenethyl ester causes p21Cip1 induction, Akt signaling reduction, and growth inhibition in PC-3 human prostate cancer cells. PLoS One. 7:e312862012. View Article : Google Scholar : PubMed/NCBI

34 

Erdogan S, Doganlar O, Doganlar ZB, Serttas R, Turkekul K, Dibirdik I and Bilir A: The flavonoid apigenin reduces prostate cancer CD44+ stem cell survival and migration through PI3K-Akt/NF-κB signaling. Life Sci. 162:77–86. 2016. View Article : Google Scholar : PubMed/NCBI

35 

Renehan AG, Booth C and Potten CS: What is apoptosis, and why is it important? BMJ. 322:1536–1538. 2001. View Article : Google Scholar : PubMed/NCBI

36 

Hassen S, Ali N and Chowdhury P: Molecular signaling mechanisms of apoptosis in hereditary non-polyposis colorectal cancer. World J Gastrointest Pathophysiol. 3:71–79. 2012. View Article : Google Scholar : PubMed/NCBI

37 

Boulares AH, Yakovlev AG, Ivanova V, Stoica BA, Wang G, Iyer S and Smulson M: Role of poly(ADP-ribose) polymerase (PARP) cleavage in apoptosis. Caspase 3-resistant PARP mutant increases rates of apoptosis in transfected cells. J Biol Chem. 274:22932–22940. 1999. View Article : Google Scholar : PubMed/NCBI

38 

Seo HS, Choi HS, Kim SR, Choi YK, Woo SM, Shin I, Woo JK, Park SY, Shin YC and Ko SG: Apigenin induces apoptosis via extrinsic pathway, inducing p53 and inhibiting STAT3 and NFκB signaling in HER2-overexpressing breast cancer cells. Mol Cell Biochem. 366:319–334. 2012. View Article : Google Scholar : PubMed/NCBI

39 

Shukla S, Fu P and Gupta S: Apigenin induces apoptosis by targeting inhibitor of apoptosis proteins and Ku70-Bax interaction in prostate cancer. Apoptosis. 19:883–894. 2014. View Article : Google Scholar : PubMed/NCBI

40 

Das S, Das J, Samadder A, Boujedaini N and Khuda-Bukhsh AR: Apigenin-induced apoptosis in A375 and A549 cells through selective action and dysfunction of mitochondria. Exp Biol Med (Maywood). 237:1433–1448. 2012. View Article : Google Scholar : PubMed/NCBI

41 

Choi KY, Chang K, Pickel JM, Badger JD II and Roche KW: Expression of the metabotropic glutamate receptor 5 (mGluR5) induces melanoma in transgenic mice. Proc Natl Acad Sci USA. 108:15219–15224. 2011. View Article : Google Scholar : PubMed/NCBI

42 

Khan AJ, Wall B, Ahlawat S, Green C, Schiff D, Mehnert JM, Goydos JS, Chen S and Haffty BG: Riluzole enhances ionizing radiation-induced cytotoxicity in human melanoma cells that ectopically express metabotropic glutamate receptor 1 in vitro and in vivo. Clin Cancer Res. 17:1807–1814. 2011. View Article : Google Scholar : PubMed/NCBI

43 

Lee HJ, Wall BA, Wangari-Talbot J, Shin SS, Rosenberg S, Chan JLK, Namkoong Jin, Goydos JS and Chen S: Glutamatergic pathway targeting in melanoma: Single-agent and combinatorial therapies. Clin Cancer Res. 17:7080–7092. 2011. View Article : Google Scholar : PubMed/NCBI

44 

Gelb T, Pshenichkin S, Hathaway HA, Grajkowska E, Dalley CB, Wolfe BB and Wroblewski JT: Atypical signaling of metabotropic glutamate receptor 1 in human melanoma cells. Biochem Pharmacol. 98:182–189. 2015. View Article : Google Scholar : PubMed/NCBI

45 

Lee HJ, Wall BA, Wangari-Talbot J and Chen S: Regulation of mGluR1 expression in human melanocytes and melanoma cells. Biochim Biophys Acta. 1819:1123–1131. 2012. View Article : Google Scholar : PubMed/NCBI

46 

Song Z, He CD, Liu J, Sun C, Lu P, Li L, Gao L, Zhang Y, Xu Y, Shan L, et al: Blocking glutamate-mediated signalling inhibits human melanoma growth and migration. Exp Dermatol. 21:926–931. 2012. View Article : Google Scholar : PubMed/NCBI

47 

Tanimura S, Uchiyama A, Watanabe K, Yasunaga M, Inada Y, Kawabata T, Iwashita K, Noda S, Ozaki K and Kohno M: Blockade of constitutively activated ERK signaling enhances cytotoxicity of microtubule-destabilizing agents in tumor cells. Biochem Biophys Res Commun. 378:650–655. 2009. View Article : Google Scholar : PubMed/NCBI

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April 2017
Volume 37 Issue 4

Print ISSN: 1021-335X
Online ISSN:1791-2431

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APA
Zhao, G., Han, X., Cheng, W., Ni, J., Zhang, Y., Lin, J., & Song, Z. (2017). Apigenin inhibits proliferation and invasion, and induces apoptosis and cell cycle arrest in human melanoma cells. Oncology Reports, 37, 2277-2285. https://doi.org/10.3892/or.2017.5450
MLA
Zhao, G., Han, X., Cheng, W., Ni, J., Zhang, Y., Lin, J., Song, Z."Apigenin inhibits proliferation and invasion, and induces apoptosis and cell cycle arrest in human melanoma cells". Oncology Reports 37.4 (2017): 2277-2285.
Chicago
Zhao, G., Han, X., Cheng, W., Ni, J., Zhang, Y., Lin, J., Song, Z."Apigenin inhibits proliferation and invasion, and induces apoptosis and cell cycle arrest in human melanoma cells". Oncology Reports 37, no. 4 (2017): 2277-2285. https://doi.org/10.3892/or.2017.5450