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Anticancer activity and underlying mechanism of neogambogic acid
Chinese Journal of Natural Medicines 2018, 16(9): 06410643
Chinese Journal of Natural Medicines
•Review•
Anticancer activity and underlying mechanism of neogambogic acid SUN Rui, ZHANG Hong-Ming, CHEN Bao-An* Department of Hematology and Oncology, Zhongda Hospital, School of Medicine, Southeast University, Nanjing 210009, China Available online 20 Sep., 2018
[ABSTRACT] Garcinia, a kind of dry resin secreted by Garcinia hanburyi Hook. F. G., is a traditional Chinese medicine with various biological functions such as detoxification, anti-inflammatory, and anthelmintic activities. Recent studies suggest that garcinia has potential anticancer activity. Increasing evidences indicate that the main active monomer gambogic acid isolated from garcinia can inhibit the growth of various cancer cells. Neogambogic acid is an isolated compound with a similar chemical structure as gambogic acid. Preliminary studies show that the neogambogic acid can selectively inhibit the growth of various cancer cells, and has a broader antitumor activity and lower toxicity than gambogic acid. In this review, we summarize the advances made in the investigation of the anti-tumor effect of neogambogic acid in recent years. [KEY WORDS] Chinese medicine; Neogambogic acid; Anticancer; Mechanism
[CLC Number] R965, Q5
[Document code] A
[Article ID] 2095-6975(2018)09-0641-03
Introduction Garcinia, a kind of dry resin secreted by Garcinia hanburyi Hook. F. G., is a traditional Chinese herbal medicine. In 1984, LU Gui-Bao and others [1], isolated a compound, whose chemical structure was similar to gambogic acid, and named it neogambogic acid. Neogambogic acid is one of the main component of garcinia and its content is up to 8.01%−37.8% [2]. The molar mass of neogambogic acid is 646.77 g·mol−1 and its molecular formula is C38H46O9. It appears as yellow crystalline, since it’s in large amount in the original ingredients, the extraction process is simple and low-cost, showing significant value of research and development [3].
Anticancer effects Xiao et al. [2] treated S180 cells with different concentrations of neogambogic acid for 48 h and used CCK-8 assay to [Received on] 02-Nov.-2017 [Research funding] This work is supported by the Fundamental Research Funds for the Central Universities (No. KYLX15-0186), the Key Medical Projects of Jiangsu Province (No. BL2014078), and the Key Discipline of Jiangsu Province (No. 2016-2020). [*Corresponding author] Tel/Fax: +86-25-83272006, E-mail: [email protected] These authors have no conflict of interest to declare. Published by Elsevier B.V. All rights reserved
detect the drug’s effect on cell proliferation. Its anti-tumor effect in vivo was evaluated by the survival time of the mice bearing S180 ascitic tumor, which were treated with 8.0, 4.0, or 2.0 mg·kg–1 (i.p.) of neogambogic acid once a day for a week, and continually observed for 45 days. And the mice bearing S180 xenograft tumors were treated with 16.0, 8.0, or 4.0 mg·kg–1 (i.g.) of neogambogic acid, once a day for 12 days. Their results indicate that the anti-tumor effects of neogambogic acid on S180 cells in vivo and in vitro are both observed. But these experiments did not determine the anticancer mechanism of neogambogic acid. In an in vitro study with LP-1 multiple myeloma cells, Ni and Kong [4] used CCK-8 assay to determine the effect of neogambogic acid on cell proliferation, Annexin V-EGFP fluorescent staining and flow cytometry to qualitatively detect its effect on apoptosis, and a cascade enzyme substrate method to detect its effect on pro-caspase-3 activation status. They also used a scarification assay to detect its effect on angiogenic activity of vascular endothelial cells in vitro. Their results showed that neogambogic acid can inhibit the proliferation of the multiple myeloma cells, induce apoptosis by the way of cascade of plasminogen activation, and inhibit angiogenesis of vascular endothelial cells in vitro. Dai and colleagues [5] treated HCT116 cells with 2.5, 5.0, and 7.5 μmol·L−1 neogambogic acid for 24 h, and the control HCT116 cells with 2.5, 5.0 and 7.5 μmol·L−1 of neogambogic acid plus 5 μmol·L−1 of endoplasmic reticulum stress (ERS)
– 641 –
SUN Rui, et al. / Chin J Nat Med, 2018, 16(9): 641643
inhibitor 4-phenylbutyric acid (4-PBA) for 24 h. The methyl thiazolyl tetrazolium assay was used to measure the cellular proliferation inhibition rate; the HCT116 cells were stained by acridine orange (AO)/ethidium bromide (EB) and their morphological changes were observed under a fluorescence microscope; they used flow cytometry with annexin V-fluorescein isothiocyanate (FITC)/propidium iodide (PI) double staining to determine the cell apoptosis rate. Their results showed that the ERS inhibitor 4-PBA reduced the inhibitory effect of neogambogic acid on the proliferation of HCT116 cells. The AO/EB staining indicated the characteristics of apoptosis in the HCT116 cells treated by neogambogic acid under the fluorescence microscope. The flow cytometry with annexin V-FITC/PI double staining showed that the apoptosis rate of neogambogic acid-treated HCT116 cells were reduced by the 4-PBA. Therefore, the authors concluded that the mechanism by which neogambogic acid inhibits proliferation and induces apoptosis in HCT116 cells might be through regulating ERS response. To find out the mechanism of neogambogic acid in inducing the apoptosis of melanoma B16 cells, Chen and coworkers [6] measured the inhibitory effect of neogambogic acid on the proliferation of B16 cells by the methy thiszolyl tetrazolium (MTT) assay. The effect of neogambogic acid on B16 cells was detected by Hoechst 33258 staining. The ultrastructure changes of B16 cells were observed by the transmission electron microscopy. The changes in the levels of P13K, p-P13K, Akt, p-Akt, p-Mtor, and PTEN proteins were analyzed the Western blotting assay. Their results suggested that neogambogic acid was significant for inhibition of the growth and proliferation of melanoma B16 cells. The cell viability remarkably decreased as the neogambogic acid concentration increased and the extension of the action time increased. The results of the Hoechst 33258 staining indicated that cells processed with neogambogic acid demonstrated apoptotic characteristics. The B16 cells treated with neogambogic acid showed obvious morphological changes of apoptosis under a transmission electron microscope. The Western blotting resultsshowed a reduction in the p-PI3K and p-Akt protein expressions depended on time, without changes in amount of p-PI3K and p-Akt protein expression. The p-m TOR protein expression decreased with the extension of time and the PTEN protein expression showed an increase, dependent on time. So the conclusion was that neogambogic acid had the ability to inhibit the proliferation of melanoma B16 cells and within certain time and concentration ranges, neogambogic acid can induce apoptosis of B16 cells. Its mechanism in inducing the cell apoptosis may be related to PI3K/Akt/m TOR signaling pathways. Neogambogic acid has also been shown to significantly inhibit proliferation of HepG2 cell [7] and induce apoptosis [8]. Some researches [9] have shown that the expression level of phosphorylated p38 and downstream phosphorylated ERK1/2 is increased in HepG2 cells treated with neogambogic acid,.
Neogambogic acid plays its role by inducing oxidative stress of mitochondria and activating caspase proteins [10]. To determine the mechanisms of anticancer effect, Qu et al. [11] studied the impact of neogambogic acid on L1210 leukemia cell cycle by microspectrophotometry and scanning measurement. Their results showed that after the treatment with neogambogic acid (10 mg·kg−1, i.p.) for 6 h, the cells in S phase decreased from 31% in the control group to 17%, the cells in G2 phase fell to zero, the cells in G1 phase increased significantly and appeared low DNA contained cells, also appeared new cell peal at lower pathways. Meanwhile, neogambogic acid (10 mg·kg−1, i.p.) can make the content of RNA decrease significantly and slow down the progression from G2 phase to M phase. After using drugs, RNA recovered slower than DNA and the RNA/DNA ratio increased. It can last a long time, which indicated that RNA may be the sensitive site of the drug.
Discussion In terms of cell cycle progression [12], neogambogic acid could arrest cells in S phase and further arrest in G0/G1 phase with extension of exposure time. In terms of the MAPK signal transduction pathway, neogambogic acid could reduce the signals of ERK signaling pathway to arrest cell cycle, resulting in its anti-proliferative effect [12]. And the targeting point may be in the upstream of MEK in the Ras/Raf/MEK/ERK cascade pathway because the level of p-MEK1/2, the actie form of MEK, is reduced [12]. In terms of telomerase activity, neogambogic acid is able to inhibit telomerase activity dependent on drug concentration [13]. In terms of the angiogenesis of tumor, neogambogic acid could reduce the levels of CD31 in solid tumor tissue to some extent [14]. Chang and co-workders [13] used the MTT method to analyze the effect of folate-modified vesicles loaded neogambogic acid on cell proliferation and apoptosis of HepG2 cells and proved folate-modified vesicles loaded neogambogic acid is more effective than unprocessed neogambogic acid. Future studies are needed to illuminate the specific mechanisms by which neogambogic acid exerts its anticancer effects.
References [1]
[2]
[3]
[4]
– 642 –
Lu GB, Yang XX, Huang QS. Isolation and structure of neo-gambogic acid from gamboges [J]. Acta Pharmaceutica Sinica, 1984, 19(8): 636-639. Xiao GL, Zhao XJ, Liu WH, et al. Anti-tumor effects of neogambogic acid on S180 cell line both in vivo and in vitro [J]. Chin J Exp Tradit Med Formul, 2012, 18(13): 193-196. Wang K, Tang Y, Sun M, et al. The mechanism of neogambogic acid-induced apoptosis in human MCF-7 cells [J]. Acta Biochim Biophys Sin (Shanghai), 2011, 43(9): 698-702. Ni HW, Kong XT. Research on the neogambogic acid inducing LP-1 cell apoptosis in human multiple myeloma tumor and inhibition of angiogenesis effect [J]. Clin J Tradit Chin Med, 2012, 24(12): 1139-1142.
SUN Rui, et al. / Chin J Nat Med, 2018, 16(9): 641643
[5]
[6]
[7] [8]
[9]
Dai TT, Cheng H, Su JJ, et al. Neogambogic acid induces apoptosis of human colon cancer HCT116 cells through endoplasmic reticulum stress [J]. Anhui Univ Chin Med, 2014, 33(1): 63-66. Chen H, Zhang X, Su JJ, et al. Study of gambogenic acidinduced apoptosis of melanoma B16 cells through P13K/ AKt/mTOR signaling pathways [J]. China J Chin Mat Med, 2014, 39(9): 1666-1669. Chen RZ, Chen BA. Advances of anti-tumor effects of neogambogic acid [J]. E J Transl Med, 2016, 3(3): 48-49. Wang X, Chen Y, Han QB, et al. Proteomic identification of molecular targets of gambogie acid: role of stathmin in hepatocellular carcinoma [J]. Proteomics, 2009, 9(2): 242-53. Fu WM, Zhang JF, Wang H, et al. Apoptosis induced by 1, 3, 6, 7-tetrahydroxyxanthone in hepatocellular carcinoma and proteomic analysis [J]. Apoptosis, 2012, 17(8): 842-851.
[10] Yan F, Wang M, Li J, et a1. Gambogeuic acid induced mitochondrial—dependent apoptosis and referred to phospho-Erkl/2 and phospho-p38 MAPK in human hepatoma HepG2 cells [J]. Environ Toxicol Pharm, 2012, 33(2): 181-190. [11] Qu BX, Hao G, Li DH, et al. Study on the anticancer effect of garcinia Ⅱ [J]. Chin J Clin Oncol, 1991, 18(1): 50. [12] Liu WH. Research on the anti-tumor effect and mechanism of neogambogic acid [D]. Guangzhou Univ Chin Med, 2011: 7-9. [13] Chang JL, Fang QY, Zhang B, et al. Effect of folate-modified vesicles loaded neogambogic acid on inhibit cell proliferation and induct apoptosis of HepG2 cell [J]. J Jiangxi Univ Tradit Chin Med, 2018, 30(1): 84-88. [14] Liu WH, Lai XP, Zhou XT. Advances in anti-tumor effects of neogambogic acid [J]. Lishizhen Med Mat Res, 2010, 21(9): 2347.
Cite this article as: SUN Rui, ZHANG Hong-Ming, CHEN Bao-An. Anticancer activity and underlying mechanism of neogambogic acid [J]. Chin J Nat Med, 2018, 16(9): 641-643.
– 643 –
Chinese Journal of Natural Medicines
•Review•
Anticancer activity and underlying mechanism of neogambogic acid SUN Rui, ZHANG Hong-Ming, CHEN Bao-An* Department of Hematology and Oncology, Zhongda Hospital, School of Medicine, Southeast University, Nanjing 210009, China Available online 20 Sep., 2018
[ABSTRACT] Garcinia, a kind of dry resin secreted by Garcinia hanburyi Hook. F. G., is a traditional Chinese medicine with various biological functions such as detoxification, anti-inflammatory, and anthelmintic activities. Recent studies suggest that garcinia has potential anticancer activity. Increasing evidences indicate that the main active monomer gambogic acid isolated from garcinia can inhibit the growth of various cancer cells. Neogambogic acid is an isolated compound with a similar chemical structure as gambogic acid. Preliminary studies show that the neogambogic acid can selectively inhibit the growth of various cancer cells, and has a broader antitumor activity and lower toxicity than gambogic acid. In this review, we summarize the advances made in the investigation of the anti-tumor effect of neogambogic acid in recent years. [KEY WORDS] Chinese medicine; Neogambogic acid; Anticancer; Mechanism
[CLC Number] R965, Q5
[Document code] A
[Article ID] 2095-6975(2018)09-0641-03
Introduction Garcinia, a kind of dry resin secreted by Garcinia hanburyi Hook. F. G., is a traditional Chinese herbal medicine. In 1984, LU Gui-Bao and others [1], isolated a compound, whose chemical structure was similar to gambogic acid, and named it neogambogic acid. Neogambogic acid is one of the main component of garcinia and its content is up to 8.01%−37.8% [2]. The molar mass of neogambogic acid is 646.77 g·mol−1 and its molecular formula is C38H46O9. It appears as yellow crystalline, since it’s in large amount in the original ingredients, the extraction process is simple and low-cost, showing significant value of research and development [3].
Anticancer effects Xiao et al. [2] treated S180 cells with different concentrations of neogambogic acid for 48 h and used CCK-8 assay to [Received on] 02-Nov.-2017 [Research funding] This work is supported by the Fundamental Research Funds for the Central Universities (No. KYLX15-0186), the Key Medical Projects of Jiangsu Province (No. BL2014078), and the Key Discipline of Jiangsu Province (No. 2016-2020). [*Corresponding author] Tel/Fax: +86-25-83272006, E-mail: [email protected] These authors have no conflict of interest to declare. Published by Elsevier B.V. All rights reserved
detect the drug’s effect on cell proliferation. Its anti-tumor effect in vivo was evaluated by the survival time of the mice bearing S180 ascitic tumor, which were treated with 8.0, 4.0, or 2.0 mg·kg–1 (i.p.) of neogambogic acid once a day for a week, and continually observed for 45 days. And the mice bearing S180 xenograft tumors were treated with 16.0, 8.0, or 4.0 mg·kg–1 (i.g.) of neogambogic acid, once a day for 12 days. Their results indicate that the anti-tumor effects of neogambogic acid on S180 cells in vivo and in vitro are both observed. But these experiments did not determine the anticancer mechanism of neogambogic acid. In an in vitro study with LP-1 multiple myeloma cells, Ni and Kong [4] used CCK-8 assay to determine the effect of neogambogic acid on cell proliferation, Annexin V-EGFP fluorescent staining and flow cytometry to qualitatively detect its effect on apoptosis, and a cascade enzyme substrate method to detect its effect on pro-caspase-3 activation status. They also used a scarification assay to detect its effect on angiogenic activity of vascular endothelial cells in vitro. Their results showed that neogambogic acid can inhibit the proliferation of the multiple myeloma cells, induce apoptosis by the way of cascade of plasminogen activation, and inhibit angiogenesis of vascular endothelial cells in vitro. Dai and colleagues [5] treated HCT116 cells with 2.5, 5.0, and 7.5 μmol·L−1 neogambogic acid for 24 h, and the control HCT116 cells with 2.5, 5.0 and 7.5 μmol·L−1 of neogambogic acid plus 5 μmol·L−1 of endoplasmic reticulum stress (ERS)
– 641 –
SUN Rui, et al. / Chin J Nat Med, 2018, 16(9): 641643
inhibitor 4-phenylbutyric acid (4-PBA) for 24 h. The methyl thiazolyl tetrazolium assay was used to measure the cellular proliferation inhibition rate; the HCT116 cells were stained by acridine orange (AO)/ethidium bromide (EB) and their morphological changes were observed under a fluorescence microscope; they used flow cytometry with annexin V-fluorescein isothiocyanate (FITC)/propidium iodide (PI) double staining to determine the cell apoptosis rate. Their results showed that the ERS inhibitor 4-PBA reduced the inhibitory effect of neogambogic acid on the proliferation of HCT116 cells. The AO/EB staining indicated the characteristics of apoptosis in the HCT116 cells treated by neogambogic acid under the fluorescence microscope. The flow cytometry with annexin V-FITC/PI double staining showed that the apoptosis rate of neogambogic acid-treated HCT116 cells were reduced by the 4-PBA. Therefore, the authors concluded that the mechanism by which neogambogic acid inhibits proliferation and induces apoptosis in HCT116 cells might be through regulating ERS response. To find out the mechanism of neogambogic acid in inducing the apoptosis of melanoma B16 cells, Chen and coworkers [6] measured the inhibitory effect of neogambogic acid on the proliferation of B16 cells by the methy thiszolyl tetrazolium (MTT) assay. The effect of neogambogic acid on B16 cells was detected by Hoechst 33258 staining. The ultrastructure changes of B16 cells were observed by the transmission electron microscopy. The changes in the levels of P13K, p-P13K, Akt, p-Akt, p-Mtor, and PTEN proteins were analyzed the Western blotting assay. Their results suggested that neogambogic acid was significant for inhibition of the growth and proliferation of melanoma B16 cells. The cell viability remarkably decreased as the neogambogic acid concentration increased and the extension of the action time increased. The results of the Hoechst 33258 staining indicated that cells processed with neogambogic acid demonstrated apoptotic characteristics. The B16 cells treated with neogambogic acid showed obvious morphological changes of apoptosis under a transmission electron microscope. The Western blotting resultsshowed a reduction in the p-PI3K and p-Akt protein expressions depended on time, without changes in amount of p-PI3K and p-Akt protein expression. The p-m TOR protein expression decreased with the extension of time and the PTEN protein expression showed an increase, dependent on time. So the conclusion was that neogambogic acid had the ability to inhibit the proliferation of melanoma B16 cells and within certain time and concentration ranges, neogambogic acid can induce apoptosis of B16 cells. Its mechanism in inducing the cell apoptosis may be related to PI3K/Akt/m TOR signaling pathways. Neogambogic acid has also been shown to significantly inhibit proliferation of HepG2 cell [7] and induce apoptosis [8]. Some researches [9] have shown that the expression level of phosphorylated p38 and downstream phosphorylated ERK1/2 is increased in HepG2 cells treated with neogambogic acid,.
Neogambogic acid plays its role by inducing oxidative stress of mitochondria and activating caspase proteins [10]. To determine the mechanisms of anticancer effect, Qu et al. [11] studied the impact of neogambogic acid on L1210 leukemia cell cycle by microspectrophotometry and scanning measurement. Their results showed that after the treatment with neogambogic acid (10 mg·kg−1, i.p.) for 6 h, the cells in S phase decreased from 31% in the control group to 17%, the cells in G2 phase fell to zero, the cells in G1 phase increased significantly and appeared low DNA contained cells, also appeared new cell peal at lower pathways. Meanwhile, neogambogic acid (10 mg·kg−1, i.p.) can make the content of RNA decrease significantly and slow down the progression from G2 phase to M phase. After using drugs, RNA recovered slower than DNA and the RNA/DNA ratio increased. It can last a long time, which indicated that RNA may be the sensitive site of the drug.
Discussion In terms of cell cycle progression [12], neogambogic acid could arrest cells in S phase and further arrest in G0/G1 phase with extension of exposure time. In terms of the MAPK signal transduction pathway, neogambogic acid could reduce the signals of ERK signaling pathway to arrest cell cycle, resulting in its anti-proliferative effect [12]. And the targeting point may be in the upstream of MEK in the Ras/Raf/MEK/ERK cascade pathway because the level of p-MEK1/2, the actie form of MEK, is reduced [12]. In terms of telomerase activity, neogambogic acid is able to inhibit telomerase activity dependent on drug concentration [13]. In terms of the angiogenesis of tumor, neogambogic acid could reduce the levels of CD31 in solid tumor tissue to some extent [14]. Chang and co-workders [13] used the MTT method to analyze the effect of folate-modified vesicles loaded neogambogic acid on cell proliferation and apoptosis of HepG2 cells and proved folate-modified vesicles loaded neogambogic acid is more effective than unprocessed neogambogic acid. Future studies are needed to illuminate the specific mechanisms by which neogambogic acid exerts its anticancer effects.
References [1]
[2]
[3]
[4]
– 642 –
Lu GB, Yang XX, Huang QS. Isolation and structure of neo-gambogic acid from gamboges [J]. Acta Pharmaceutica Sinica, 1984, 19(8): 636-639. Xiao GL, Zhao XJ, Liu WH, et al. Anti-tumor effects of neogambogic acid on S180 cell line both in vivo and in vitro [J]. Chin J Exp Tradit Med Formul, 2012, 18(13): 193-196. Wang K, Tang Y, Sun M, et al. The mechanism of neogambogic acid-induced apoptosis in human MCF-7 cells [J]. Acta Biochim Biophys Sin (Shanghai), 2011, 43(9): 698-702. Ni HW, Kong XT. Research on the neogambogic acid inducing LP-1 cell apoptosis in human multiple myeloma tumor and inhibition of angiogenesis effect [J]. Clin J Tradit Chin Med, 2012, 24(12): 1139-1142.
SUN Rui, et al. / Chin J Nat Med, 2018, 16(9): 641643
[5]
[6]
[7] [8]
[9]
Dai TT, Cheng H, Su JJ, et al. Neogambogic acid induces apoptosis of human colon cancer HCT116 cells through endoplasmic reticulum stress [J]. Anhui Univ Chin Med, 2014, 33(1): 63-66. Chen H, Zhang X, Su JJ, et al. Study of gambogenic acidinduced apoptosis of melanoma B16 cells through P13K/ AKt/mTOR signaling pathways [J]. China J Chin Mat Med, 2014, 39(9): 1666-1669. Chen RZ, Chen BA. Advances of anti-tumor effects of neogambogic acid [J]. E J Transl Med, 2016, 3(3): 48-49. Wang X, Chen Y, Han QB, et al. Proteomic identification of molecular targets of gambogie acid: role of stathmin in hepatocellular carcinoma [J]. Proteomics, 2009, 9(2): 242-53. Fu WM, Zhang JF, Wang H, et al. Apoptosis induced by 1, 3, 6, 7-tetrahydroxyxanthone in hepatocellular carcinoma and proteomic analysis [J]. Apoptosis, 2012, 17(8): 842-851.
[10] Yan F, Wang M, Li J, et a1. Gambogeuic acid induced mitochondrial—dependent apoptosis and referred to phospho-Erkl/2 and phospho-p38 MAPK in human hepatoma HepG2 cells [J]. Environ Toxicol Pharm, 2012, 33(2): 181-190. [11] Qu BX, Hao G, Li DH, et al. Study on the anticancer effect of garcinia Ⅱ [J]. Chin J Clin Oncol, 1991, 18(1): 50. [12] Liu WH. Research on the anti-tumor effect and mechanism of neogambogic acid [D]. Guangzhou Univ Chin Med, 2011: 7-9. [13] Chang JL, Fang QY, Zhang B, et al. Effect of folate-modified vesicles loaded neogambogic acid on inhibit cell proliferation and induct apoptosis of HepG2 cell [J]. J Jiangxi Univ Tradit Chin Med, 2018, 30(1): 84-88. [14] Liu WH, Lai XP, Zhou XT. Advances in anti-tumor effects of neogambogic acid [J]. Lishizhen Med Mat Res, 2010, 21(9): 2347.
Cite this article as: SUN Rui, ZHANG Hong-Ming, CHEN Bao-An. Anticancer activity and underlying mechanism of neogambogic acid [J]. Chin J Nat Med, 2018, 16(9): 641-643.
– 643 –