Introduction
The bioactive constituents of Huachansu
Cytostatic effects of Huachansu and Bufalin on tumors
Cancer type | Cell line | 24H IC50 | 48H IC50 | 72H IC50 | Refs. |
---|---|---|---|---|---|
Breast | MCF-7 | 317.9 ± 1.5 nM | 46.5 ± 1.4 nM | Yan et al. [125] | |
3.2 nM | Wang et al. [56] | ||||
MCF-7/ADR | 100.17 ± 30.16 nmol/L | Wang et al. [126] | |||
MCF-10A | 465.2 ± 25.9 nM | Clifford et al. [31] | |||
MCF10CA1 | 635.2 ± 171.8 nM | Clifford et al. [31] | |||
MDA-MB-231 | 934.1 ± 2.0 nM | 513.3 ± 1.6 nM | Yan et al. [125] | ||
263.3 ± 68.24 nmol/L | Wang et al. [126] | ||||
936.4 ± 4.9 nM | Clifford et al. [31] | ||||
20.0 nM | Song et al. [55] | ||||
MDA-MB-231-LM3-3 | 16.6 nM | Song et al. [55] | |||
SUM149PT | 16.6 nM | Song et al. [55] | |||
SUM159PT | 15.9 nM | Song et al. [55] | |||
Cervical | Hela | 154 ± 21.5 nM | 37.5 ± 2.15 nM | 11.8 ± 2.13 nM | Pan et al. [127] |
Colorectal | SW620 | 76.72 ± 6.21 nmol/L | 34.05 ± 4.21 nmol/L | 16.7 ± 6.37 nmol/L | Zhu et al. [128] |
287.35 ± 4.34 nM | 57.63 ± 7.80 nM | 20.39 ± 1.95 nM | Zhang et al. [129] | ||
HCT116 | 82.6 μmol/L | Qiu et al. [130] | |||
0.243 μM | 0.024 μM | Wang et al. [131] | |||
LoVo | 56.778 ± 7.34 nM | 11.48 ± 2.89 nM | 6.64 ± 2.79 nM | Zhang et al. [129] | |
Endometrial | Ishikawa | 0.7 ng/mL | Takai et al. [132] | ||
HHUA | 0.5 ng/mL | Takai et al. [132] | |||
HEC-1B | 0.6 ng/mL | Takai et al. [132] | |||
Esophageal | Eca-109 | 1.0 μM | Lv et al. [133] | ||
EC9706 | 1.0 μM | Lv et al. [133] | |||
TE5 | 2.6 μM | Lv et al. [133] | |||
TE11 | 4.9 μM | Lv et al. [133] | |||
Hec2 | 3.8 μM | Lv et al. [133] | |||
Gallbladder | GBC-SD | 48.12 ± 2.03 nM | 28.23 ± 1.78 nM | Jiang et al. [45] | |
SGC996 | 125.03 ± 5.16 nM | 102.78 ± 3.21 nM | Jiang et al. [45] | ||
Gastric | SGC7901 | > 800 nmol/L | Li et al. [134] | ||
MGC803 | 160 ± 0.87 nmol/L | Li et al. [134] | |||
Liver | HepG2 | 182.30 ± 13.78 nM | Li et al. [27] | ||
307 ± 87 nM | Ozdemir et al. [84] | ||||
33.65 nmol/L | Gao et al. [60] | ||||
143.2 nM | Miao et al. [49] | ||||
PLC/PRF/5 | 52.20 ± 14.16 nM | Li et al. [27] | |||
157.87 nmol/L | Gao et al. [60] | ||||
SMMC7721 | 97.74 ± 8.83 nM | Li et al. [27] | |||
SK-HEP-1 | 110.33 ± 5.32 nM | Tsai et al. [135] | |||
Leukemia | NB4 | 40 nmol/L | 27 nmol/L | 17 nmol/L | Zhai et al. [136] |
K562 | 0.0943 μmol/L | Zhai et al. [136] | |||
K562/VCR | 0.0401 μmol/L | Zhai et al. [136] | |||
HEL | 0.046 μmol/L | Wang et al. [137] | |||
Lung | A549 | 4.5 nM | Wang et al. [56] | ||
56.14 ± 6.72 nmol/L | 15.57 ± 4.28 nmol/L | Zhu et al. [138] | |||
8.15 ± 0.69 nmol/L | Sun et al. [92] | ||||
22.00 ± 3.53 nM | 10.20 ± 1.01 nM | Liu et al. [36] | |||
Melanoma | A375.S2 | 450.38 nM | Hsiao et al. [44] | ||
Oral | CAL27 | 125 nM | Tsai et al. [139] | ||
122.6 nM | Tian et al. [140] | ||||
Osteosarcoma | U-2OS | 0.297 μM | Zhang et al. [141] | ||
8.49 ± 2.1 μg/L | Yin et al. [142] | ||||
U-2OS/MTX300 | 10.19 ± 1.7 μg/L | Yin et al. [142] | |||
Saos-2 | 0.318 μΜ | Zhang et al. [141] | |||
Ovarian | SK-OV-3 | 1.0 ng/mL | Takai et al. [132] | ||
OMC-3 | 0.6 ng/mL | Takai et al. [132] | |||
Tongue | SCC-4 | 300 nM | Chou et al. [143] | ||
Pancreatic | Capan-2 | 159.2 nM | Tian et al. [140] | ||
Non-cancerous | Breast epithelial cell 184D | 384.6 ± 36.5 nM | Clifford et al. [31] | ||
Breast epithelial cell 184A | 295.9 ± 10.5 nM | Clifford et al. [31] | |||
Mouse normal hepatocyte | No cytotoxicity at 10 μM | Song et al. [55] | |||
HUVECs | 53.1 nmol/L | Qiu et al. [130] | |||
Het-1Aa | 10.1 μM | Lv et al. [133] | |||
PBMCsb | 16.5 ± 4.7 ng/mL | Yuan et al. [144] | |||
Primary normal human endometrial epithelial cells | Little sensitivity to BF from 0.1 to 10 ng/mL | Takai et al. [132] | |||
PBMCs | 16.5 ± 4.7 ng/mL | Yuan et al. [144] |
In vivo evaluation of Huachansu and its derivative Bufalin on tumor inhibition
Cancer type | Drugs | Animal | Tumor models | Transplantation | Treatment | Results | Refs. | Year |
---|---|---|---|---|---|---|---|---|
Bone | Bufalin | Athymic nude mice; male; 7–8 weeks old | Rat breast sarcocarcinoma Walker 256 cell | Inoculate 2 × 105 cell into the intramedullary space of the mouse femur | 1 day after tumor inoculation, given by i.p. 0.5, 1, 1.5, or 2 mg/kg/day for 21 days | Cancer-induced pain relief; 50% reduction in bone tissue injury | Ji et al. [83] | 2017 |
Cervical | Bufalin | BALB/c nude mice; female; 4–5 weeks old | Human cervical squamous cell carcinoma Siha cell | Subcutaneously inoculate 3 × 106 cell | After reaching an average tumor volume of 100 mm3, given by i.p. Bufalin 10 mg/kg ± paclitaxel (10 mg/kg per 4 days) for 32 days | Bufalin synergizes with paclitaxel to inhibit tumor growth without apparent loss of body weight | Liu et al. [80] | 2016 |
Breast | Bufalin-NP | SCID mice; female; 6–7 weeks old | Human breast MDA-MB-231-LM3.3 cell | Inject 0.75 × 106 cell into one of the second mammary fat pads | 6 days after tumor injections, given by i.v. Bufalin-NP (1.5 mg/kg) three times per week for 20 days | Bufalin sensitizes cancer cells to MK-2206 and blocks tumor growth | Wang et al. [56] | 2014 |
Bufalin-BCS-NP | Balb/c nude mice; female; 4 weeks old | Human breast MCF-7 cell | Subcutaneously implant 1 × 107 cell into the right axilla skin | 7–10 days after tumor inoculation, (1) given by i.p. BF-BCS-NPs (1 mg/kg BF equivalent in PBS) or free BF (1 mg/kg in ethanol) every 2 days for 20 days; (2) give by i.v. BF-BCS-NPs (5 mg/kg BF equivalent in PBS) at D1, D3 | Free BF suppresses tumor growth accompanying with a significantly decreased body weight. BF-BCS-NPs suppresses tumor growth without apparent loss of body weight | Tian et al. [104] | 2014 | |
3-Phospho-bufalin | Nude mice; female; 4–6 weeks old | Human breast MDA-MB-231-LM3-3 cell | Inject 0.75 × 106 cell into one of the second mammary fat pads | 14 days after injection, given by s.c. phospho-BF (0.75 mg/kg per dose, 3 doses per week) for 3 weeks | 2.4 times reduction in tumor weight; no cardiotoxicity observed | Song et al. [55] | 2015 | |
Bufalin | Athymic nude mice; female; 5 weeks old | Human breast MDA-MB-231 cell | Subcutaneously inject 5 × 106 into both dorsal flank regions | 4 weeks after injection, (1) given by intra-tumoral injection BF 10 μL (1 mM, in 0.9% normal saline) to the left flank for 4 weeks; (2) 10 μL volume of normal saline to the right flank for 4 weeks | Significantly enhances breast cancer xenograft growth; promote the inflammatory response | Chen et al. [38] | 2017 | |
Colorectal | Bufalin-loaded mPEG-PLGA-PLL-cRGD nanoparticles (BNPs) | Athymic nude mice; female; 4–6 weeks old | Human colon SW620 cell | Inject 5 × 106 cell into the dorsal subcutaneous space | Bufalin-loaded mPEG-PLGA-PLL-cRGD nanoparticles (BNPs) containing 1 mg/kg bufalin, given by injection through the vena caudalis once a day for 14 days | Suppresses tumor growth; bufalin-loaded NPs significantly enhanced treatment efficacy compared to that of a bufalin water solution | Yin et al. [105] | 2012 |
Bufalin | Male athymic nude mice | HCT116-luc-vector and HCT116-luc-miR-497 | Inject 1 × 106 cells intravenously via the tail vein | 1 week after injection, given by Bufalin for 1 mg/kg via the tail vein (three times a week) for 5 weeks | Inhibited colorectal cancer metastasis; improved life of survival. Improved physiological characteristics in terms of body weight, skin roughness, mental status, and survival rate | Qiu et al. [130] | 2014 | |
Bufalin-loaded pluronic polyetherimide nanoparticles | Male athymic nude mice | HCT116 | Injecte 1 × 106 cells intravenously via the tail vein. | 2 weeks after injection, (1) given by of Bufalin 1 mg/kg; (2) given by 20 mg/kg of Bufalin-loaded pluronic PEI nanoparticles via the tail vein (0.2 mL per mouse, three times per week) for 3 weeks | Inhibited colorectal cancer metastasis. Improved quality of life and physiological characteristics in terms of body weight, skin roughness, and mental status | Hu et al. [93] | 2014 | |
Bufalin | BALB/c mice; male; 5–6 weeks old | HCT116 | Inject 2 × 1010 cells s.c. into the right axillary region. Tumor mode of the second generation: 2 weeks after injection, harvest the s.c. xenogra tumors, cut into pieces (1.5 mm in diameter), implant into the axillary region s.c. Orthotopic xenogra model: harvest third generation s.c. tumors and cut into pieces (1.5 mm in diameter) | NS group (treated with 0.2 mL normal saline); 5-Fu group (treated with 5-FU, 25 mg/kg); low Bufalin group (0.5 mg/kg); medium Bufalin group (1.0 mg/kg); high Bufalin group (1.5 mg/kg). NS, 5-FU, and Bufalin were administrated by intraperitoneal injection, once per day from day 15 to day 21 (12 mice in each group) | Inhibit cell growth. Lower tumor volume. Prolong survival time | Wang et al. [131] | 2015 | |
Bufalin-DOX | Athymic nude mice (BALB/c-nu/nu) of 6–8 weeks | HCT8/ADR | Inject 1 × 107 cells s.c. under the shoulder in the nude mice | Mice were randomized into six groups (6 in each group) when the tumor volumes reached 150–200 mm3: control; BF (0.1 mg/kg, i.p., q3d × 5); DOX (0.1 mg/kg, i.p., q3d × 5); DOX (0.5 mg/kg, i.p., q3d × 5); DOX (1.0 mg/kg, i.p., q3d × 5); DOX (0.1 mg/kg, i.p., q3d × 5) plus BF (0.1 mg/kg, i.p., q3d × 5, given 1 h before DOX administration) | BF remarkable increased the effect of DOX against the ABCB1 resistant HCT8/ADR colorectal cell xenografts in nude mice | Yuan et al. [145] | 2015 | |
Bufalin | Male nude mice (BALB/c nu/nu, 5-week-old) | HCT116 | Inject 2 × 106 cells into the subcutaneous tissues. | 2 weeks after injection, (1) given by cisplatin (10 mg/kg body weight) i.p. every 3 days for 4 weeks.; (2) given by bufalin (1 mg/kg body weight) i.p. every 3 days for 4 weeks | Inhibition of tumor growth and tumor tissue weights are greater with the combination of cisplatin and bufalin than with cisplatin alone. Tumors treated with the combination of cisplatin and bufalin showed more cell vacuolization and nuclear shrinkage than with cisplatin alone | Sun et al. [146] | 2017 | |
Gallbladder | Bufalin | Male athymic nude mice (5 week-old) | GBC-SD | Xenograft model: Inoculate 1 × 106 GBC-SD cells into the left axillary region | 24 h postinoculation, given by PBS i.p. and bufalin with (0.1, 0.2, and 0.4 mg/kg) i.p. every 2 days for up to 20 days | Suppression of tumor growth | Jiang et al. [45] | 2014 |
Liver | Bufalin | BALB/c nu/nu mice (18–20 g, 5 week-old) | HCCLM3 | Inject 5 × 106 cells s.c. into the upper left flank region of nude mice. Revmove the tumors when reached approximately 1 cm in length (approximately 4 weeks after injection) and mince into small pieces of equal volume (1.5–2 × 2 × 2 mm3), then transplant into the livers of 24 nude mice | From day 8 to 38, given by1 mg/kg Bufalin; 1.5 mg/kg Bufalin; 100 mg/kg LY294002 and saline i.p. thrice weekly, respectively | 1.5 mg/kg Bufalin decreased the sizes and qualities of orthotopic transplanted tumors as well as pulmonary metastasis. Orthotopic transplanted tumor tissues were necrotic and the apoptotic cell number was markedly higher in 1.5 mg/kg Bufalin group. Inhibition of AKT/GSK3β/β-catenin/E-cadherin signaling pathways | Zhang et al. [72] | 2014 |
Bufalin–sorafenib | BALBc nu/nu mice (6 week-old) | Human HCC cell lines SMMC7721 | Inoculate 5 × 106 cells s.c. into the abdominal intraderma | Control group: inject with the vehicle i.p. (5 days/week, 2 weeks). Experimental group: (1) injection of 1 mg/kg bufalin i.p. (5 days/week, 2 weeks); (2) oral uptake of 30 mg/kg/day sorafenib (5 days/week, 2 weeks); (3) the combination of both injections of bufalin i.p. and oral uptake of sorafenib (5 days/week, 2 weeks) | Inhibit blood vessel formation in the intradermal tumors, manifested by the vessel numbers and branches and attenuate tumor weight in nude mice with the combination treatment | Wang et al. [64] | 2016 | |
Bufalin | BALBc nu/nu mice (6 week-old) | Human HCC cell lines SMMC7721 | Inoculate 5 × 106 cells s.c. into the right flank | Tumor size was measured every 4 days after the treatment. Tumor-bearing mice were sacrificed after 16 days of treatment, and the tumor weight was evaluated | Combination treatment inhibits tumor growth and tumor angiogenesis in vivo. The combination treatment group showed more reduced microvessel density than any other group | Wang et al. [64] | 2016 | |
Bufalin | BALBc nu/nu mice (6 week-old) | SMMC7721-GFP | Inoculate 5 × 106 cells s.c. into the right flanks. 4 weeks later, cut the non-necrotic tumor tissue into 1 mm3 pieces and orthotopically implanted into the liver. In addition, inject 2 × 106 via mouse tail veins | (1) Inject 1 mg/kg bufalin i.p. (5 days/week for six weeks); (2) Inject PBS i.p. (5 days/week for 6 weeks) | Bufalin reduced liver/lung metastases. Bufalin inhibited invasion through EMT | Wang et al. [76] | 2016 | |
Myeloma | Bufalin-MK2206 | BALB-c nu/nu female mice (4–6 week-old); NOD-SCID female mice (4–6 week-old) | MOPC315; H929 | Inject 2 × 107 MOPC315 cells s.c. in the right flanks of the BALB-c nu/nu mice. Inject 1 × 107 cells H929 cells s.c. in the right hind leg of NOD/SCID mice | Mice bearing MOPC315 MM tumors were treated with bufalin (1 mg/kg; intraperitoneally) daily in the presence and/or absence of MK2206 (120 mg/kg orally) for 10 days. Mice injected with H929 MM cells were treated with 1 mg/kg bufalin daily with or without 120 mg/kg MK2206 for 12 days | Bufalin combined with MK2206 blocked MM tumor growth, decreased tumor cell proliferation and increased the percentage of apoptotic cells | Xiang et al. [147] | 2017 |
Osteosarcoma | Bufalin | BALB/c nude mice; female; 6 week-old | U2OS/MTX300 | Inject 5.6 × 106 cells s.c. into the axilla of the mice | 10 days after injection, (1) control group: 100 mL of vehicle i.p.; MTX group: MTX (250 mg/kg) with calcium leucovorin rescue (24 mg/kg at 16, 20, or 24 h after MTX) i.p. per week; (3) low Bufalin group: 0.75 mg/kg i.p.; (4) High Bufalin group:1.5 mg/kg i.p. | Bufalin inhibited tumor growth | Xie et al. [86] | 2012 |
Pancreas | Bufalin | BALB/c nu/nu mice; male; 4-week-old | Mia PaCa-2 | Inject 6 × 106 cells s.c. into the back of mice | When tumors reached the size of 100 mm3: (1) vehicle alone (control); (2) Bufalin (0.1 mg/kg, for 10 days); (3) Gemcitabine (125 mg/kg, 3 times/week for 2 weeks); (4) Bufalin and Gemcitabine in combination | Bufalin potentiates the anti-tumor effect of gemcitabine in vivo. Combination treatment with gemcitabine signicantly reduced the tumor volume and cell proliferation activity | Chen et al. [148] | 2012 |
Bufalin | BALBc nu/nu mice; female; 6-week-old | MiaPaCa2/GEM | Inoculated 2 × 105 cells into the right flanks of mice | Given by (1) injections of 1.5 mg/kg bufalin (5 days/week) for 4 weeks; (2) injections with vehicle (20 μL saline) for 4 weeks. In addition, MiaPaCa2/GEM cells (2 × 106) was given to one of another two groups pre-treated with bufalin via the tail veins for 6 weeks | Inhibit pancreatic tumor growth | Wang et al. [82] | 2016 | |
Bufalin | BALBc nude mice; male; 5-week-old) | BxPC3-luc2 | Injected 1 × 107 cells s.c. into the left buttock of mice | 7 days after inoculation, control group: inject saline i.p.; Bufalin groups: inject 1 mg/kg and 2 mg/kg for 14 days; Positive control: DDP 2 mg/kg every other day i.p. for 14 days | Bufalin treatment inhibits tumor growth | Liu et al. [149] | 2016 | |
Lung | Bufalin | BALB/c nu/nu mice; male; 6–8 week-old | NCI-H460 | Inject 1.3 × 107 cells s.c. into flank of each mouse | When tumor volume exceeded 100 mm3, given by (1) vehicle (0.1% DMSO); (2) Bufalin: 0.1, 0.2, or 0.4 mg/kg for 14 days | Bufalin suppresses tumor growth | Wu et al. [150] | 2017 |
Molecular mechanisms of anti-tumor activity
Cancer type | Study type | Pharmacological action | Pathways involved | Refs. | Year |
---|---|---|---|---|---|
Bladder | In vitro | Growth inhibition (50–300 nM, 24H); G2/M arrest; apoptosis induction (T24, EJ) [BF] | Bax/Bcl-2 ratio Fas, DR4, DR5, TRAIL, PARP cleavage↑; pro-caspase-3, -8, -9, Bcl-xL, Bid, XIAP, cIAP-1, cIAP-2↓ | Hong et al. [43] | 2012 |
In vitro | Growth inhibition (50–200 nM, 24H); G0/G1 arrest; mitochondrial/apoptosis (T24) [BF] | ROS production, cytochrome c, Apaf-1, AIF, caspase-3, -7, -9, Bax↑; ΔΨm, cyclin D, CDK4, cyclin E, CDK2, phospho-Rb, phospho-AKT Bcl-2↓ | Huang et al. [29] | 2012 | |
In vitro | Migration and invasion inhibition (5–100 nM, 24H) (T24) [BF] | TIMP-1, -2, phospho-ERK↑; claudin-2, -3, -4, MMP-2, -9, Active-MMP-2, -9↓ | Hong et al. [151] | 2013 | |
In vitro | Sensitization of TRAIL-mediated apoptosis (2.5–10 nM, 24H) (T24) [BF] | Caspase-9, pro-caspase-9, DR5, degradation of poly (ADP-ribose) polymerase↑ | Kang et al. [152] | 2017 | |
Bone | In vivo | Relieves cancer-induced pain and bone destruction (Walker 256 cells) [BF] | OPG↑; RANKL, serum TRACP5b, ICTP, PINP↓ | Ji et al. [83] | 2017 |
Breast | In vitro | Sensitization of TRAIL-mediated apoptosis (0.02–0.25 μM, 24H) (MCF-7, MDA-MB-231) [BF] | Mcl-1, Bcl-xL, p-STAT3↓ | Dong et al. [153] | 2011 |
In vitro | Growth inhibition (50 nM, 24H); apoptosis induction; enhanced TRAIL-induced apoptosis (MCF-7, MDA-MB-231) [BF] | DR4, DR5, p-ERK, p-JNK, p-p38, cleaved-PARP↑; pro-caspase-8, Cbl-b↓ | Yan et al. [125] | 2012 | |
In vitro and in vivo | Growth inhibition (1–5 nM, 24H); promote coactivators’ protein degradation (MCF-7) [BF] | SRC-3 mRNA expression, SRC-3 protein degradation↑; intrinsic transcriptional activities of SRC-1 and SRC-3↓ | Wang et al. [56] | 2014 | |
In vitro | Sensitization of TRAIL-mediated apoptosis (50 nmol/L, 24H); redistributing death receptors in lipid rafts (MCF-7, MDA-MB-231) [BF] | DR4, DR5, cleaved-PARP, cleaved-caspase-8↑; caspase-8↓ | Yan et al. [154] | 2014 | |
In vitro and in vivo | Growth inhibition (100 ng/mL, 8H); mitochondrial/apoptosis (MCF-7) [BF, BF-BCS-NPs] | Intracellular ROS, BAX, cleaved-caspase-3↑; ΔΨm, Bcl-2, survivin↓ | Tian et al. [104] | 2014 | |
In vitro | Growth inhibition; apoptosis (10–50 nmol/L, 48H) (MCF-7/ADR, MDA-MB-231)[BF] | PARP cleavage, miR-155-5p expression↑; DNMT1, DNMT3a, FOXO3a↓ | Wang et al. [126] | 2016 | |
In vitro and in vivo | Growth inhibition (12–200 nM) (LM3-3) [BF, phospho-BFs] | Synergizes with Gefitinib; SRC-3↓ | Song et al. [55] | 2015 | |
In vitro | Growth inhibition (4–8 nM, 48H) (MDA-MB-231); enhanced HDAC inhibitors induced apoptosis [BF] | ↑;SRC-3p-Akt, Bcl-2↓ | Zou et al. [155] | 2016 | |
In vitro and in vivo | Promote inflammatory response (0.001–1 μM, 12H) (MDA-MB-231) [BF] | p65 translocation; PKC-induced COX-2 and IL-8, PGE2, p-JNK, p-p38, p-ERK, TPA-induced MMP-3 protein and mRNA expression↑ | Chen et al. [38] | 2017 | |
Cervical | In vitro and in vivo | Growth inhibition (0.05–0.2 μM, 24H); G2/M arrest; apoptosis; migration and invasion inhibition (0.01–0.04 μM, 24H) (Siha, Hela) [BF] | BAX, P21, p27, E-cadherin, GSK3β↑;Bcl-2, Bcl-xL, cyclin A, cyclin B1, CDK2, MMP-9, SNAIL1, integrin α2, integrin β5, FAK, p-FAK(Tyr397), p-GSK3β(Ser389), AKT1, p-AKT(Ser473)↓ | Liu et al. [80] | 2016 |
In vitro | Growth inhibition (0–50 nM, 24H); apoptosis induction (Hela) [BF] | HSP27, vimentin, HNRPK↓ | Pan et al. [127] | 2012 | |
Colorectal | In vitro | Growth inhibition (25–100 nM, 48H); G2/M arrest; autophagy induction (HT-29, Caco-2) [BF] | LC3-II, ROS, Atg-5, Beclin-1, p-JNK2↑ | Xie et al. [59] | 2011 |
In vitro | Growth inhibition (20, 80 nmol/L, 24H); G2/M arrest; apoptosis (SW620) [BF] | PARP cleavage, cleaved-caspase-3, BAX/BCL-2 ratio↑;p-Stat3, p-ERK, livin↓ | Zhu et al. [128] | 2012 | |
In vitro | Growth inhibition (100 nM, 9H); mitotic arrest; G2/M arrest (HT-29, HCT-116) [BF] | p-H3↑;HIF-1α, NF-κB, Plk1 expression↓ | Xie et al. [156] | 2013 | |
In vitro and in vivo | Anti-migration and anti-metastasis (3.12–50 nM, 12H) (HCT116), ex vivo micro vessel sprouting (HUVECs) [BF] | miR-497 expression↑; VEGFA expression↓ | Qiu et al. [130] | 2014 | |
In vitro and in vivo | Growth inhibition (0.03–3 μΜ, 24–48H); G2/M arrest; apoptosis (HT116) [BF] | PTEN, Bad, caspase-3 phosphorylation, caspase-3 cleavage↑; p-PTEN, p-AKT↓ | Wang et al. [131] | 2015 | |
In vitro | Anti-proliferation and anti-migration (10–50 nM, 48H); induce apoptosis; G2/M arrest (LoVo, SW620) [BF] | Cyclin B1, p–cdc2, p21, cleaved-PARP, Bax, cleaved-caspase-7, -9, E-cadherin↑; Bcl-2, N-cadherin, β-catenin, CPSF4, hTERT↓; inhibit hTERT by down-regulating CPSF4 | Zhang et al. [129] | 2016 | |
In vitro and in vivo | Suppress growth of cisplatin-resistant cell rather than sensitive-one; reverse ABCB1-mediated multi-drug resistance (5–20 nM, 48H) (LoVo/ADR, HCT8/ADR, HCT8/ABCB1) [BF] | ATPase activity of ABCB1↑; ABCB1↓ | Yuan et al. [145] | 2015 | |
In vitro and in vivo | Reverse cisplatin drug resistance (5 nM, 48H) (HCT116, LoVo HCT116-STSCscis, LoVo-STSCscis) [BF] | CD133, CD44, OCT4, SOX2, NANOG, ABCG2 ↓ | Sun et al. [146] | 2017 | |
Gallbladder | In vitro | Apoptosis induction (25–200 nmol/L, 48H); S-phase arrest; mitochondrial dysfunction induction (GBC-SD, SGC996) [BF] | Cleaved-caspase-3, -9, cleaved-PARP ↑; Cyclin A, Cyclin B1, Cyclin D1, CDK1, NF-κB, Bcl-2, ΔΨm↓ | Jiang et al. [45] | 2014 |
Gastric | In vitro | Growth inhibition (100, 200 nmol/L, 24H); G2/M phase arrest; intrinsic apoptosis induction (SGC7901, MGC803) [BF] | SPARC antagonizes bufalin-induced apoptosis; p-Src, p-Akt, p-ERK↑; ΔΨm, cyclin B1, cyclin A↓ | Li et al. [134] | 2015 |
In vitro | Growth inhibition (25–200 nmol/L, 48H); apoptosis induction (SGC7901, MGC803)[BF] | Bax, cleaved-PARP↑; Bcl-2, pro-caspase-3, miR-298 suppressed apoptosis↓ | Zhao et al. [157] | 2015 | |
In vitro | Proliferation inhibition (100 nM, 48H); cisplatin-sensitization; apoptosis induction (SGC7901, MKN-45, BGC823) [BF] | p-AKT, p-GSK3β, p-mTOR, p-4EBP1, p-S6 K↓ | Zhao et al. [158] | 2016 | |
In vitro | G0/G1 phase arrest (50, 80 nmol/L, 48Η); caspase and mitochondrial-mediated apoptosis induction; ER stress induction; protective autophagy activation (SGC7901, BGC823) [BF] | Cleaved-caspase-3, cleaved-PARP, Bax/Bcl-2, CHOP, p-eIF2a, p-JNK, LC3-II, Atg5, Beclin-1↑; LC3-I, p62↓
| Zhao et al. [159] | 2017 | |
Glioma | In vitro | Growth inhibition (20–80 nM, 24H); mitochondria-mediated apoptosis induction; autophagy activation (U87MG, LN229) [BF] | ROS production, Bax, cytosolic cytochrome c, cleaved-PARP, cleaved-caspase-3, -4, LC3-II, p-AMPK, p-ACC, ATF6f, p-PERK, p-IRE1α, p-eIF2α, GRP78, GRP994, CHOP↑; Bcl-2, p-mTOR, p-4EBP1, p-p70S6K, PERK↓ | Shen et al. [160] | 2014 |
In vitro | Proliferation inhibition (5–40 μM, 24H); cancer stem cell-like phenotypes inhibition; apoptosis induction (U251, U87) [BF] | miR-203↑;OCT4, SOX2, SPARC↓ | Liu et al. [161] | 2017 | |
HCC | In vitro | Growth inhibition (0.1–1 μΜ, 2H) (HepG2, PLC/PRF/5, SMMC7721) [BF] | p-ERK, p-Akt, ATP1A3↑; FoxO3a↓ | Li et al. [27] | 2011 |
In vitro | Growth inhibition (0.001–0.1 μmol/L, 24–72H); induction of fas- and mitochondria-mediated apoptosis (HepG2) [BF] | Bax, cytochrome c, cleaved-caspase-3, -9, PARP cleavage ↑; ΔΨm, Bcl-2, pro-caspase-3, -9, -10, Bid ↓ | Qi et al. [162] | 2011 | |
In vitro | Anti-proliferation (50–250 nM, 48Η); enhances the anti-cancer effects of Sorafenib (6.25lM) (PLC/PRF/5, HepG2) [BF] | Enhanced apoptotic cell death in combination with Sorafenib; p-Akt↑;p-ERK↓ | Gao et al. [60] | 2012 | |
In vitro | Growth inhibition (50–100 nM, 24H); G2/M arrest; autophagy induction (SK-HEP-1)[CS] | Chk1, Wee1, LC3-II, Atg5, Atg7, Atg12, Beclin-1↑; Cyclin A, cyclin B, CDK1, p-CDK1(Thr161), Cdc25c, p-CDC25c(Ser198), p-Akt(Ser308), p-AKT(Ser473), p-mTOR(Ser2481), AKT kinase activity↓ | Tsai et al. [135] | 2012 | |
In vitro | Growth inhibition (10–100 nM, 48H); autophagy induction; apoptosis induction (HepG2) [BF] | p-APMK, Beclin-1, LC3-II, p-p70S6K↑; p-mTOR, p62↓ | Miao et al. [49] | 2013 | |
In vitro | Growth inhibition; G2/M arrest (0.04 μM, 4–12H); autophagy induction (Huh7, Hep3B, HA22T) [BF] | TNF, BECN-1, MAPK, ATG8↑; Bcl-2, Bid↓ | Hsu et al. [30] | 2013 | |
In vitro | Growth inhibition; anti-migration (10, 100 nmol/L, 48H); anti-invasion; anti-adhesion (HCCLM3, HepG2) [BF] | GSK3β, E-cadherin↑; p-Akt, p-GSK3β, MMP-9, -2, β-catenin nuclear translocation↓ | Qiu et al. [71] | 2013 | |
In vitro | Growth inhibition (5, 10 nM, 48H); anti-migration and invasion (SK-Hep1) [BF] | PI3K, p-Akt, NF-κB translocation, MMP-2, -9, FAK, Rho A, VEGF, MEKK3, MKK7, uPA↓ | Chen et al. [68] | 2013 | |
In vitro | Growth inhibition (50–100 nmol/L, 48H); apoptosis induction; ER stress induction; autophagy induction (Huh-7, HepG2) [BF] | Beclin-1, p-JNK1, p-JNK2, IRE1, ATG5, LC3-I, LC3-II↑; p62↓ | Hu et al. [163] | 2014 | |
In vitro | Reverse multidrug resistance (1 nM, 48H); G0/G1 phase arrest (BEL-7402/5-FU) [BF] | drug efflux pump activity, TS, MRP1, Bcl-xL/Bax ratio ↓ | Gu et al. [164] | 2014 | |
In vivo | Orthotopic growth inhibition, anti-metastasis (1 mg/kg and 1.5 mg/kg) (HCCLM3-R) [BF] | p-Akt, GSK3β, E-cadherin↑; p-GSK3β, β-catenin, MMP-9, -2↓ | Zhang et al. [72] | 2014 | |
In vitro | Reverse Sorafenib resistance (50–200 nM, 48H); synergies with sorafenib to induce apoptosis (HepG2, HepG2-Sora, Huh7, Huh7-Sora) [BF] | IRE1, CHOP, P-eIF2α↑; p-Akt↓ | Zhai et al. [165] | 2015 | |
In vitro | Anti-invasion and metastasis (0.085 μg/mL, 72H) (BEL-7402) [BF] | E-cadherin, ALB↑; β-catenin, p-GSK-3β Ser9, MMP-7, COX-2, Cyclin D1, AFP↓ | Gai et al. [66] | 2015 | |
In vitro and in vivo | Synergies anti-angiogenic effect of sorafenib (2.5–10 nM, 48H); anti-migration; S-phase arrest (HUVECs, SMMC7721) [BF] | p-ERK↑; VEGF, p-Akt, p-mTOR↓ | Wang et al. [64] | 2016 | |
In vitro and in vivo | Inhibits TGF-β1 induced EMT and invasion (10 nM, 24–72H) (SMMC7721) [BF] | E-cadherin↑; N-cadherin, Vimentin, Snail, HIF-1α↓ | Wang et al. [76] | 2016 | |
In vitro | Growth inhibition (40–200 nmol/L, 24–48H); apoptosis induction (Hep3B)[BF] | Apolipoprotein E (APOE) knockdown induced Na+/K+-ATPase, caveolin, PI3K/AKT/GSK3b and apoptosis signal cascades↑;Cyclin D1, Cdc25c, Cdc2↓ | Liu et al. [166] | 2016 | |
In vitro | Growth inhibition (0.04 μg/mL, 72H); S- and G2-phase arrest; apoptosis induction; anti-migration and invasion; adhesion inhibition (HCC-LM3) [BF] | E-cadherin↑; β-catenin, MMP-2, -9, VEGF↓ | Sheng et al. [67] | 2016 | |
In vitro | Growth inhibition (0.1 mg/mL, 24H); apoptosis induction (HepG2, HLE) [HCS] | Bcl-2↑;ΔΨm, Bax, Bid, cytochrome c↓ | Xia et al. [167] | 2017 | |
Leukemia | In vitro | Vitamin D-induced cell differentiation enhancing (7.5, 10, 12.5 nM, 24H); VDR transactivation activity enhancing (HL-60, THP-1, U937) [BF] | 1,25(OH)2D3-induced CYP24A1, CD14, CAMP, PTGS1, CD11b, CDKN1A expression, nuclear VDR expression, histone acetylation and VDR recruitment to the CYP24A1 promoter, Erk MAP kinase activation↑ | Amano et al. [168] | 2009 |
In vitro | Proliferation inhibition (5–80 nmol/L, 12–72H); apoptosis induction (NB4) [BF] | Synergized with PD98059; caspase-3 activation↑; survivin expression↓ | Zhu et al. [169] | 2012 | |
In vitro | Reverse multidrug resistance (0.001–0.1 μM, 48H); S-phase arrest (K562, K562/VCR) [BF] | Bax↑; MRP1, Bcl-xL↓ | Zhai et al. [136] | 2014 | |
In vitro | Growth inhibition (0.01–0.5 μmol/L, 48H); apoptosis induction; cell cycle arrest (HEL) [BF] | WT1 gene methylation, DNMT3a, DNMT3b protein↑; WT1 mRNA expression↓ | Wang et al. [137] | 2017 | |
Lung | In vitro | Growth inhibition (0.1 μM, 24–48H); apoptosis induction; ROS-dependent mitochondrial dysfunction (ASTC-a-1) [BF] | ROS production, Bax translocation from cytosol to mitochondria, caspase-3 activation↑ | Sun et al. [50] | 2011 |
In vitro | Growth inhibition (2.5–10 μM, 48–72H); apoptosis induction; G1-phase arrest (A549) [BF] | Cyto C (Cytosol)/Cyto C (mitochondrial), cleaved-caspase-3, cleaved-PARP, p53, p21Waf↑; Bcl-2/Bax ratio, Cyclin D1, COX-2, p-VEGFR2, p-VEGFR1, p-EGFR, p-Akt, p-p38MAPK, p-NKκB, p-ERK1/2↓ | Jiang et al. [46] | 2010 | |
In vitro | Growth inhibition (20–100 nM, 48–72H); apoptosis induction (A549) [BF] | Bax, cleaved-caspase-3↑; Bcl-2, livin, p-Akt↓ | Zhu et al. [138] | 2012 | |
In vitro | Growth inhibition (20 nM, 72H); reverses HGF-resistance to EGFR-TKIs; apoptosis induction (HCC827, PC-9, H1975) [BF] | Cleaved-caspase-3, -9, cleaved-PARP↑; blockage of Met/PI3k/Akt pathway | Kang et al. [170] | 2013 | |
In vitro | Growth inhibition (1–4 μM, 24H); induce DNA condensation (NCI-H460) [BF] | DNA-PK, BRCA1, 14-3-3-σ, MDC1, MGMT, P53↓ | Wu et al. [70] | 2014 | |
In vitro | Growth inhibition (2–8 ng/mL, 48H); apoptosis induction (A549) [BF] | Cleaved-caspase-3, cytosol cytochrome c↑; ΔΨm↓ | Ding et al. [41] | 2014 | |
In vitro | TGF-β induced epithelial-to-mesenchymal transition and migration inhibition (50 nM, 24H) (A549) [BF] | TGF-β receptor, I and II TGF-β induced E-cadherin, p-Smad2, p-Smad3↓ | Zhao et al. [79] | 2015 | |
In vitro | Invasion and migration inhibition (25–100 nM, 24–48H); cell adhesion inhibition (NCI-H460) [BF] | RhoA, MMP-2, -9 expression, TIMP1 expression↑; MMP-2, -9 activity, NF-κB, PKC, GRB2, p-AKT, p-ERK, p-P38, p-JNK1/2, ROCK1, FAK, TIMP2↓ | Wu et al. [70] | 2015 | |
In vitro | Growth inhibition (5–20 nmol/L, 72H); cellular proteasome activity inhibition; ubiquitinated proteins accumulation (A549) [BF211] | PSMB6 β1↓ | Sun et al. [92] | 2016 | |
In vitro | Growth inhibition (1–2 nM, 48H); enhanced HDAC inhibitors induced apoptosis (A549) [BF] | SRC-3p-Akt, Bcl-2↓ | Zou et al. [155] | 2016 | |
In vitro and in vivo | Growth inhibition (1–4 μM, 12–48H); cell morphological changes induction; DNA condensation; apoptosis induction (H460) [BF] | Cytochrome C, Apaf-1, active caspase-3, FasL/CD95, FasL, AIF, Endo G, caspase-9 activity, GADD153 mRNA expression, ROS production↑; pro-caspase-3, Bcl-2, GRP78 mRNA expression, ΔΨm↓ | Wu et al. [150] | 2017 | |
In vitro and in vivo | Growth inhibition (12.5–50 nM, 1–12H); apoptosis induction (A549) [BF] | Cleaved-caspase-3, cleaved-PARP, Caspase-3 activity↑; p-Src↓ | Liu et al. [36] | 2016 | |
In vitro | Inhibits gefitinib resistant cell migration and invasion (2.5–10 nM, 24–48H) (NCI-H460) [BF] | p-p38, p65↑; SOS-1, MMP-2, RhoA, N-Cadherin, E-Cadherin↓ | Huang et al. [74] | 2016 | |
In vitro | Growth inhibition (25–100 nM, 6–48H); apoptosis induction (H1975) [BF] | Mcl-1 protein degradation, cleaved-PARP, cleaved-caspase-3, Bax, Bak↑; p-GSK-3β, Bcl-1, Bcl-2, Bcl-xL↓ | Kang et al. [171] | 2017 | |
In vitro | Growth inhibition (1–100nmoL/L, 24–72H); apoptosis induction; S-phase arrest (A549) [BF] | Caspase-3↑ | Zhang et al. [172] | 2017 | |
Melanoma | In vitro | Growth inhibition (150–550 nM, 24–48H); apoptosis induction (A375.S2) [BF] | ROS production, intracellular Ca2+ production, NO formation; cleaved-caspase-3, -8, -9, cytochrome c, AIF, Endo G, Bax, Fas, FasL, GRP78; ΔΨm, Bcl-xL↓ | Hsiao et al. [44] | 2012 |
Myeloma | In vitro | Growth inhibition (20 nM, 48H); apoptosis induction; G2/M arrest (U929, U266) [BF] | Chemosensitivity↑; PARP1↓ | Huang et al. [173] | 2013 |
In vitro and in vivo | Growth inhibition (12 nM, 12–48H); synergistic with MK2206 surpassed bortezomib resistance (U929, U266) [BF] | p-Akt, p-mTOR↑; p-P70, IL-6 secretion↓ | Xiang et al. [147] | 2017 | |
Oral | In vitro | Growth inhibition (125 nM, 24H); G0/G1 phase arrest; apoptosis induction (CAL27)[BF] | Cytochrome c, Apaf-1, AIF, cleaved-caspase-3, -9↑; p-Akt, Cyclin D1, p-Bad↓ | Tsai et al. [139] | 2012 |
In vitro | Growth inhibition (50–150 nM, 24–48H); apoptosis induction (CAL27) [BF] | Intracellular ROS accumulation, p-JNK, p-p38, p-c-Jun↑; hTERT expression ↓ | Tian et al. [140] | 2015 | |
Osteosarcoma | In vitro | Growth inhibition (25–50 nM, 24–48H); anti-invasion and migration (U2OS) [BF] | SOS-1, JNK1/2, ERK1/2, p-38, MMP-7, -9 enzyme activity↓ | Chueh et al. [69] | 2011 |
In vitro and in vivo | Growth inhibition (25 nM, 24H); apoptosis induction; G2/M arrest (U2OS, U2OS/MTX300) [BF] | Cleaved-PARP↑; Hsp27, p-Akt, P65↓ | Xie et al. [86] | 2012 | |
In vitro | Inhibition of differentiation and proliferation (10 μM, 72Η) (hMG63-derived cancer stem cell) [BF] | CD133↓ | Chang et al. [174] | 2014 | |
In vitro | Growth inhibition (10–40 nM, 24H) (MG-63) [BF] | ROS production, mitochondrial membrane hyperpolarization, Apaf-1, cleaved-PARP, cleaved-caspase-3, -7, -9↑; Bcl-2/Bax ratio↓ | Wang et al. [51] | 2014 | |
In vitro | Inhibition of differentiation and proliferation (10 μΜ, 8 days) (Primary osteosarcoma stem cell C1OS) [BF] | Cleaved-caspase-3, miR-148a↑; ALDH1, hTERT, Nanog. CD133, Notch, Bmi-1↓ | Chang et al. [175] | 2015 | |
In vitro | Growth inhibition (10–50 μg/L, 6–24H); apoptosis induction (U-2OS) [BF] | Cytosol/mitochondrial cytochrome c, cleaved-PARP, cleaved-caspase-3, -9, Bax↑; ΔΨm, PARP, Bcl-2↓ | Chen et al. [40] | 2016 | |
In vitro | Growth inhibition (0.05–10 μM, 24H); apoptosis induction (U-2OS, Saos-2) [BF] | ROS production, BBC3↑; miR221↓ | Zhang et al. [141] | 2016 | |
In vitro | Growth inhibition (200 nM, 6–48H); apoptosis induction (U-2OS) [BF] | Ca2+ release, caspase-3, -8, -9 activity, cytochrome c, Fas-L, cleaved-PARP, Calpain 1, ATF-6α, GRP-78, caspase-4↑; ΔΨm, Bcl-2, Bcl-xL↓ | Lee et al. [176] | 2017 | |
Ovarian | In vitro | Growth inhibition (1 ng/mL, 48H); G0/G1 arrest; apoptosis induction [BF] (SK-OV-3, OMC-3) | p21, cleaved-caspase-9↑; Cyclin A, Cyclin D3, Bcl-2, Bcl-xL↓ | Takai et al. [132] | 2008 |
In vitro | Growth inhibition (1–100 ng/mL, 48H); apoptosis induction (SKOV3, ES-2) [BF] | miR-183 downregulation enhanced bufalin-induced growth inhibition and apoptosis | Chen et al. [177] | 2016 | |
Pancreatic | In vitro and in vivo | Growth inhibition (0.001–0.1 μM, 48H); apoptosis induction; enhance sensitivity to gemcitabine (Bx-PC3, MiaPaCa2, Panc-1) [BF] | ASK1, p-JNK, cleaved-caspase-3↑; Ki-67, Bcl-2↓ | Chen et al. [148] | 2012 |
In vitro | Growth inhibition (50–200 nM,48H); G2/M arrest; apoptosis induction; enhanced gemcitabine chemosensitivity (Panc-1, CFPAC-1) [BF] | Bax, P21↑; Bcl-2, pro-caspase-3, -9, CyclinB1, CDK1↓ | Li et al. [47] | 2014 | |
In vitro | Growth inhibition (50–150 nM, 24–48H); mitochondria-dependent apoptosis induction; (Capan-2)[BF] | p-JNK, p-p38, p-c-Jun, intracellular ROS accumulation↑; hTERT expression↓ | Tian et al. [140] | 2015 | |
In vitro and in vivo | Cancer stem cells formation inhibition (50 nM, 24H) (MiaPaCa2/GEM) [BF] | CD24 expression, ESA expression, PTCH1, PTCH2, Gli1↓ | Wang et al. [82] | 2016 | |
In vitro and in vivo | Growth inhibition (0.1–10 μM, 24H); S-phase arrest (SW1990, BxPc3) [BF] | c-Myc, NF-κB↓ | Liu et al. [149] | 2016 | |
Prostate | In vitro | Growth inhibition (0.1–10 μM, 24H); apoptosis induction (LNCaP, PC3, DU145) [BF] | Caspase-3 activity, caspase-9 activity, intracellular Ca2+↑ | Yeh et al. [178] | 2003 |
In vitro | Growth inhibition (15 μM, 24H); apoptosis induction (PC3) [BF] | miR-181a induction enhanced bufalin-induced growth inhibition and apoptosis; miR-181a expression, caspase-3 activity↑; Bcl-2↓ | Zhai et al. [179]. | 2013 | |
Renal | In vitro | Induce new high density glycogen-microtubule structures formation (1–20 nM, 0–4.5H) (ACHN) [BF] | K−ATPase-induced ERK1/2 phosphorylation↑ | Fridman E et al. [180] | 2012 |
Tongue | In vitro | Growth inhibition (100–500 nM, 48H); G2/M arrest; mitochondria-dependent apoptosis (SCC-4) [BF] | Ca2+, NO production, DR5 expression, caspase-9 expression↑; Bcl-2, Bid, calpain 1, ATF-6β, ΔΨm, ROS production↓ | Chou et al. [143] | 2017 |
Induce cell death other than apoptosis
Metastasis inhibition (angiogenesis, MMPs, EMT, others)
Angiogenesis
MMPs
EMT
Others
Omics approach in the study of antineoplastic effects of Bufalin and Huachansu
Drug delivery and its derivatives
Potential cardiotoxic property of Bufalin and the side-effects of toad extract and HCS in clinical studies
Clinical trials
Discussion and future perspectives
Concluding remarks
Highlights
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Huachansu and Bufalin possess anti-cancer effects both in vitro and in vivo.
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The multi-target and multi-pathway pharmacological actions are promising.
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Potential drug–drug interactions and multi-target interaction lacks studies.
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Further large-scale clinical trials are warranted.