Anti-Tumor Effect of Ginsenoside Rh2 and Rg5 in Xenograft Hepatoma Animal Model: Comparison with Sorafenib

( Han Seul Park ),( Jae Young Jang ),( Soung Won Jeong ),( Jeong-ju Yoo ),( Sae Hwan Lee ),( Sang Gyune Kim ),( Sang-woo Cha ),( Young Seok Kim ),( Young Deok Cho ),( Hong Soo Kim ),( So Young Jin ),( 대한간학회 2018 춘·추계 학술대회 (KASL) Vol.2018 No.1

Aims: Animal & cell models of hepatoma give a crucial information, not only pathogenesis of liver cancer but also therapeutic effects of various agents. In this study, we investigate therapeutic effects of ginsenoside Rh2 and Rg5 using animal & cell models of hepatoma comparing with sorafenib. Methods: Huh7 & huh7.5.1 cells were harvested from 2-D petri-dish and cultured in 1.5 % soft agarose gels for 10-14 days. After 10-14 days, 3-D hepatic structure was formed and treated with ginsenoside Rh2 (100, 200uM) and Rg5 (10, 50, 100uM) for 72h. Hep3b cells (2×10⁵) in matrigel were suspended in 100 μl of phosphate-buffered saline (PBS) and then injected into the flanks of BALB/C nude mouse at 6 weeks. Sorafenib (10mg/kg), ginsenoside Rh2 (100 uM) and Rg5 (100 uM) was injected to intra-peritoneum twice a week. After 4 weeks, all mice were sacrificed and tumor tissue was collected. The tissue was stained with hematoxylin and eosin and histological evaluation was conducted by blindly pathologist. Area of necrosis and vascular change in tumor tissue was calculated using the lasso tool to encircle the area in ZEN 2011 Imaging Software. Results: Both ginsenoside Rh2 and Rg5 induced cell necrosis in hepatocellular carcinoma (HCC) cell lines, and more necrosis occurred in 2D models. The expression of cleaved PARP protein was increased in both 2D and 3D cells with exposure to ginsenoside Rh2 and Rg5. The hepatocellular carcinoma was confirmed in hepatoma mouse models by H&E stain. Increased necrosis and telangiectasia were observed in mice treated with sorafenib, Rh2, and Rg5 compared to control mouse. Conclusions: Our findings provide insight into the use of xenograft mouse as models of HCC and ginsenoside Rh2 & Rg5 might be a potential treatment candidate of liver cancer.

Anti-breast cancer activity of Fine Black ginseng (Panax ginseng Meyer) and ginsenoside Rg5

Kim, Shin-Jung,Kim, An Keun The Korean Society of Ginseng 2015 Journal of Ginseng Research Vol.39 No.2

Background: Black ginseng (Ginseng Radix nigra, BG) refers to the ginseng steamed for nine times and fine roots (hairy roots) of that is called fine black ginseng (FBG). It is known that the content of saponin of FBG is higher than that of BG. Therefore, in this study, we examined antitumor effects against MCF-7 breast cancer cells to target the FBG extract and its main component, ginsenoside Rg5 (Rg5). Methods: Action mechanism was determined by MTT assay, cell cycle assay and western blot analysis. Results: The results from MTT assay showed that MCF-7 cell proliferation was inhibited by Rg5 treatment for 24, 48 and 72 h in a dose-dependent manner. Rg5 at different concentrations (0, 25, 50 and $100{\mu}M$), induced cell cycle arrest in G0/G1 phase through regulation of cell cycle-related proteins in MCF-7 cells. As shown in the results from western blot analysis, Rg5 increased expression of p53, $p21^{WAF1/CIP1}$ and $p15^{INK4B}$ and decreased expression of Cyclin D1, Cyclin E2 and CDK4. Expression of apoptosiserelated proteins including Bax, PARP and Cytochrome c was also regulated by Rg5. These results indicate that Rg5 stimulated cell apoptosis and cell cycle arrest at G0/G1 phase via regulation of cell cycle-associated proteins in MCF-7 cells. Conclusion: Rg5 promotes breast cancer cell apoptosis in a multi-path manner with higher potency compared to 20(S)-ginsenoside Rg3 (Rg3) in MCF-7 (HER2/ER+) and MDA-MB-453 (HER2+/ER) human breast cancer cell lines, and this suggests that Rg5 might be an effective natural new material in improving breast cancer.

Ginsenoside Rg5 overcomes chemotherapeutic multidrug resistance mediated by ABCB1 transporter: in vitro and in vivo study

Feng, Sen-Ling,Luo, Hai-Bin,Cai, Liang,Zhang, Jie,Wang, Dan,Chen, Ying-Jiang,Zhan, Huan-Xing,Jiang, Zhi-Hong,Xie, Ying The Korean Society of Ginseng 2020 Journal of Ginseng Research Vol.44 No.2

Background: Multidrug resistance (MDR) to chemotherapy drugs remains a major challenge in clinical cancer treatment. Here we investigated whether and how ginsenoside Rg5 overcomes the MDR mediated by ABCB1 transporter in vitro and in vivo. Methods: Cytotoxicity and colon formation as well as the intracellular accumulation of ABCB1 substrates were carried out in MDR cancer cells A2780/T and A549/T for evaluating the reversal effects of Rg5. The expressions of ABCB1 and Nrf2/AKT pathway were determined by Western blotting. An A549/T cell xenograft model was established to investigate the MDR reversal activity of Rg5 in vivo. Results: Rg5 significantly reversed ABCB1-mediated MDR by increasing the intracellular accumulation of ABCB1 substrates without altering protein expression of ABCB1. Moreover, Rg5 activated ABCB1 ATPase and reduced verapamil-stimulated ATPase activity, suggesting a high affinity of Rg5 to ABCB1 binding site which was further demonstrated by molecular docking analysis. In addition, co-treatment of Rg5 and docetaxel (TXT) suppressed the expression of Nrf2 and phosphorylation of AKT, indicating that sensitizing effect of Rg5 associated with AKT/Nrf2 pathway. In nude mice bearing A549/T tumor, Rg5 and TXT treatment significantly suppressed the growth of drug-resistant tumors without increase in toxicity when compared to TXT given alone at same dose. Conclusion: Therefore, combination therapy of Rg5 and chemotherapy drugs is a strategy for the adjuvant chemotherapy, which encourages further pharmacokinetic and clinical studies.

Ginsenoside Rg5 overcomes chemotherapeutic multidrug resistance mediated by ABCB1 transporter: in vitro and in vivo study

Sen-Ling Feng,Hai-Bin Luo,Liang Cai,Jie Zhang,Dan Wang,Ying-Jiang Chen,Huan-Xing Zhan,Zhi-Hong Jiang,Ying Xie 고려인삼학회 2020 Journal of Ginseng Research Vol.44 No.2

Background: Multidrug resistance (MDR) to chemotherapy drugs remains a major challenge in clinicalcancer treatment. Here we investigated whether and how ginsenoside Rg5 overcomes the MDR mediatedby ABCB1 transporter in vitro and in vivo. Methods: Cytotoxicity and colon formation as well as the intracellular accumulation of ABCB1 substrateswere carried out in MDR cancer cells A2780/T and A549/T for evaluating the reversal effects of Rg5. Theexpressions of ABCB1 and Nrf2/AKT pathway were determined by Western blotting. An A549/T cellxenograft model was established to investigate the MDR reversal activity of Rg5 in vivo. Results: Rg5 significantly reversed ABCB1-mediated MDR by increasing the intracellular accumulation ofABCB1 substrates without altering protein expression of ABCB1. Moreover, Rg5 activated ABCB1 ATPaseand reduced verapamil-stimulated ATPase activity, suggesting a high affinity of Rg5 to ABCB1 bindingsite which was further demonstrated by molecular docking analysis. In addition, co-treatment of Rg5 anddocetaxel (TXT) suppressed the expression of Nrf2 and phosphorylation of AKT, indicating that sensitizingeffect of Rg5 associated with AKT/Nrf2 pathway. In nude mice bearing A549/T tumor, Rg5 and TXTtreatment significantly suppressed the growth of drug-resistant tumors without increase in toxicitywhen compared to TXT given alone at same dose. Conclusion: Therefore, combination therapy of Rg5 and chemotherapy drugs is a strategy for the adjuvantchemotherapy, which encourages further pharmacokinetic and clinical studies.

흑삼의 진세노사이드 Rg3, Rk1 및 Rg5 분석법 검증을 위한 가이드라인

최보람,윤다혜,반재숙,신우철,장진규,나현선,이대영 한국응용생명화학회 2024 Journal of Applied Biological Chemistry (J. Appl. Vol.67 No.-

This study was conducted to establish the manufacturing and quality control of standards for black ginseng and propose a standard analytical method to present marker components different from red ginseng. Changes in the content of ginsenosides Rg3(s), Rk1, and Rg5 were investigated according to the number of steaming and drying processes of black ginseng. In the industrially manufactured black ginseng, the sum of Rg3(s), Rk1, and Rg5 was more than 3 mg/g. In addition, through Ultra-performance liquid chromatography quadrupole time of flight mass spectrometry analysis-based metabolomic analysis, ginsenosides Rg3(s), Rk1, and Rg5 of black ginseng were identified as components differentiated from red ginseng in the results of multivariate statistical analysis and metabolic profiling. The analytical method was validated through HPLC to standardize the marker components. For method validation, the analytical method was tested for specificity, linearity, accuracy, precision, limit of detection, and limit of quantitation, and it was found to be suitable for standardization of marker components. 본 연구는 새로운 가공 인삼 중 하나인 흑삼의 구체적인 제조기준과 검사기준을 마련하고 홍삼과 차별화되는 지표성분을 제시하기위한 표준 분석법을 제안하고자 수행되었다. 흑삼의 증숙및 건조 횟수에 따른 진세노사이드 Rg3(s), Rk1, 및 Rg5의 함량 변화를 조사하였으며, 산업용 규모로 제조된 3증숙 흑삼에서는 Rg3(s), Rk1, 및 Rg5합은 총 3mg/g 이상으로 나타났다. UPLC-QTOF/MS를 활용하여 대사체 프로파일링 및 다변량통계분석을 수행한 결과, 홍삼과 차별화되는 흑삼의 지표성분으로진세노사이드 Rg3(s), Rk1, 및 Rg5로 제안되었으며, HPLC를이용하여 지표성분 표준화를 위한 분석법에 대한 밸리데이션을실시하였다. 유효성 검증을 위해 특이성, 직선성, 정확성, 정밀성, 검출한계, 정량한계 항목에 대한 검증한 결과, 규격 범위 이내의 유효한 결과값을 얻었으며 지표성분 표준화를 위한 적합한 시험법임이 검증되었다.

Anti-breast cancer activity of Fine Black ginseng (Panax ginseng Meyer) and ginsenoside Rg5

Shin-Jung Kim,An Keun Kim 고려인삼학회 2015 Journal of Ginseng Research Vol.39 No.2

Background: Black ginseng (Ginseng Radix nigra, BG) refers to the ginseng steamed for nine times andfine roots (hairy roots) of that is called fine black ginseng (FBG). It is known that the content of saponin ofFBG is higher than that of BG. Therefore, in this study, we examined antitumor effects against MCF-7breast cancer cells to target the FBG extract and its main component, ginsenoside Rg5 (Rg5). Methods: Action mechanism was determined by MTT assay, cell cycle assay and western blot analysis. Results: The results from MTT assay showed that MCF-7 cell proliferation was inhibited by Rg5 treatmentfor 24, 48 and 72 h in a dose-dependent manner. Rg5 at different concentrations (0, 25, 50 and 100 mM),induced cell cycle arrest in G0/G1 phase through regulation of cell cycle-related proteins in MCF-7 cells. As shown in the results from western blot analysis, Rg5 increased expression of p53, p21WAF1/CIP1 andp15INK4B and decreased expression of Cyclin D1, Cyclin E2 and CDK4. Expression of apoptosiserelatedproteins including Bax, PARP and Cytochrome c was also regulated by Rg5. These results indicate that Rg5stimulated cell apoptosis and cell cycle arrest at G0/G1 phase via regulation of cell cycle-associatedproteins in MCF-7 cells. Conclusion: Rg5 promotes breast cancer cell apoptosis in a multi-path manner with higher potencycompared to 20(S)-ginsenoside Rg3 (Rg3) in MCF-7 (HER2/ER+) and MDA-MB-453 (HER2+/ER) humanbreast cancer cell lines, and this suggests that Rg5 might be an effective natural new material inimproving breast cancer.

Ginsenoside Rg5 promotes wound healing in diabetes by reducing the negative regulation of SLC7A11 on the efferocytosis of dendritic cells

Wei Xia,Zongdong Zhu,Song Xiang,Yi Yang 고려인삼학회 2023 Journal of Ginseng Research Vol.47 No.6

Background: ginsenoside Rg5 is a rare ginsenoside with known hypoglycemic effects in diabetic mice. This study aimed to explore the effects of ginsenoside Rg5 on skin wound-healing in the Leprdb/db mutant(db/db) mice (C57BL/KsJ background) model and the underlying mechanisms. Methods: Seven-week-old male C57BL/6J, SLC7A11-knockout (KO), the littermate wild-type (WT), and db/db mice were used for in vivo and ex vivo studies. Results: Ginsenoside Rg5 provided through oral gavage in db/db mice significantly alleviated the abundanceof apoptotic cells in the wound areas and facilitated skin wound healing. 50 mM ginsenoside Rg5treatment nearly doubled the efferocytotic capability of bone marrow-derived dendritic cells (BMDCs)from db/db mice. It also reduced NF-kB p65 and SLC7A11 expression in the wounded areas of db/db micedose-dependently. Ginsenoside Rg5 physically interacted with SLC7A11 and suppressed the cystineuptake and glutamate secretion of BMDCs from db/db and SLC7A11-WT mice but not in BMDCs fromSLC7A11-KO mice. In BMDCs and conventional type 1 dendritic cells (cDC1s), ginsenoside Rg5 reducedtheir glycose storage and enhanced anaerobic glycolysis. Glycogen phosphorylase inhibitor CP-91149almost abolished the effect of ginsenoside Rg5 on promoting efferocytosis. Conclusion: ginsenosideRg5 can suppress the expression of SLC7A11 and inhibit its activity via physical binding. These effectscollectively alleviate the negative regulations of SLC7A11 on anaerobic glycolysis, which fuels theefferocytosis of dendritic cells. Therefore, ginsenoside Rg5 has a potential adjuvant therapeutic reagent tosupport patients with wound-healing problems, such as diabetic foot ulcers.

Changes of Prosapogenin Components in Tienchi Seng (Panax notoginseng) by Ultrasonic Thermal Fusion Process

이재범,양병욱,김도형,Dezhong Jin,고성권 한국생약학회 2021 Natural Product Sciences Vol.27 No.1

The purpose of this study is to develop a new method of producing tienchi seng (notoginseng, Panax notoginseng) extracts featuring high concentrations of the ginsenoside Rg3, Rg5, and Rg6, special components of Korean red ginseng. The chemical transformation from ginseng saponin glycosides to prosapogenin was analyzed by HPLC. Tienchi seng was heat-processed at 100oC and the optimum conditions were identified. The highest concentrations of total saponin (29.723%) and the ginsenoside Rg3 (1.769%), Rg5 (5.979%), and Rg6 (13.473%) were produced at 48 hours. Also, when tienchi seng was subjected to the ultrasonic thermal fusion (100oC) process, the concentrations of total saponin (30.578%), ginsenoside Rg3 (2.392%), Rg5 (6.614%), and Rg6 (13.017%) were highest at 36 hours. On the other hand, the 2-hour heat-processed extract and 2-hour ultrasonic thermal fusion-processed extract did not contain ginsenoside Rg3, Rg5, and Rg6. The ultrasonic thermal fusion process had an extraction yield that was approximately 1.26 times greater than that of the heat process. These results indicate that the highly functional tienchi seng extracts created through the ultrasonic thermal fusion process are more industrially useful than those produced using the heat process.

Cancer Chemopreventive Compounds of Red Ginseng Produced from panax ginseng C.A. Meyer

Yun, Taik-Koo,Lee, Yun-Sil,Lee, You-Hui,Yun, Hyo-Yung The Korean Society of Ginseng 2001 Journal of Ginseng Research Vol.25 No.3

Fresh Panax gineng C.A. cultivated in Korea(Korean red ginseng) was found to be ineffective as anticarcinogenic or cancer preventive in experimental animal model or in human case-control and cohort study. However, when treated with heat, the fresh ginseng, white ginseng were highly effective cancer preventives. Four compounds including 20(S)-ginsenoside Rh$_1$(Rh$_1$), 20(S)-ginsenoside Rh$_2$(Rh$_2$), 20(S)0-siwenoside Rg$_3$(Rg$_3$) and sinsenoside Rg$\sub$5/ were consequently purified from Korean red ginseng, and they were tested by Yun\`s 9 week medium-term anticarcinogenicity test model. Rg$_3$ and Rg$\sub$5/ statistically significantlydecreased the incidence of benzo(a)pyrene-induced mouse lung tumor, Rh$_2$showed tendency of decrease, and Rh1 showed no effect. It is, therefore, concluded that Rg$_3$ and Rg$\sub$5/ are active anticarcinogenic components in res ginseng and they either singularly or synergistically act in the prevention of cancer. 발암물질을 투여하여 발생하는 마우스 폐선종은 홍삼추출믈의 투여에 의하여 그 발생율이 억제되나 수삼을 투여하면 발생율이 억제되지 않는다. 또한 암환자-대조군연구 결과에 있어서도 수삼즙 또는 수삼절편을 복용한 사람에서는 암의 위험비가 감소되지 않으나 수삼열탕 또는 홍삼을 복용하면 현저한 위험비의 감소를 볼 수 있었다. 이와 같은 결과는 열로 처리된 홍삼중에 암예방 유효성분이 있을 것이라고 추정되어 왔다. 저자들은 4종의 홍삼중의 진세노사이드 즉 Rh$_1$, Rh$_2$, Rg$_3$ 및 Rg$_{5}$ 를 고려홍삼으로부터 분리합성하여 윤의 9주 중기 항발암실험법에 의하여 항발암성을 관찰한바 진세노사이드 Rg$_3$와 Rg$_{5}$의 투여시에는 통계학적으루 유의한 폐선종 발생율이 감소되었으나 Rh$_2$에서는 폐선종발생율이 약간 감소되는 경향을 보였고 Rh$_1$에서는 전혀 영향을 주지 않았다. 이와 같은 소견으로 홍삼에 의한 항발암작용 또는 암예방작용은 홍삼중의 진세노사이드 Rg$_3$및 Rg$_{5}$가 유효성분임을 파악하였으며 이들 진세노사이드 Rg$_3$ Rg$_{5}$ 및 Rh$_2$가 단독 또는 복합적으로 작용할 것으로 추정된다.

흑삼의 신생혈관 억제활성에 대한 연구

신영진,이숙영,서영배,이계원,정규진,송규용 대한본초학회 2011 大韓本草學會誌 Vol.26 No.3

Objectives: This study was performed to investigate the influence of black ginseng radix extracts (BG) and ginsenoside Rg3, Rg5 on basic fibroblast growth factor (bFGF) induced proliferation, migration and capillary tubule-like formation of human umbilical vein endothelial cells (HUVECs). Methods: HUVECs were cultured with BG and ginsenoside Rg3, Rg5 at different concentrations (60, 125, 250, 500, 1,000 ㎍/㎖) for 2 day in the presence of bFGF, respectively. XTT was used to detect the proliferation. Migration and tube formations were examined to detect the antiangiogenesis. Also, the chick embryo chorioallantoic membrane (CAM) assay was performed to detect the antiangiogenesis. Results: BG and ginsenoside Rg3, Rg5 significantly inhibited bFGF-induced endothelial cell proliferation and migration in a dose-dependent manner. Tube formation in bFGF-induced HUVECs were suppressed by BG and ginsenoside Rg3, Rg5. Moreover, BG and ginsenoside Rg3, Rg5 (30-50 ㎍/egg) inhibited new blood vessel formation on the growing CAM. Conclusions: Based on the present results, it can be suggested that BG has a potential chemopreventive agent via antiangiogenesis.