1-furan-2-yl-3-pyridin-2-yl-propenone inhibits the invasion and migration of HT1080 human fibrosarcoma cells through the inhibition of proMMP-2 activation and down regulation of MMP-9 and MT1-MMP

Matrix metalloproteinases (MMPs) play important roles in solid tumor invasion and migration. In this study, we showed that 1-furan-2-yl-3-pyridin-2-yl-propenone (FPP-3) dose-dependently inhibited HT1080 cell invasion and migration, and decreased MMP-2 and MMP-9 activities. Furthermore, FPP-3 reduced MMP-2 expression at protein and mRNA levels, and suppressed 12-O-tetradecanoylphorbol-13-acetate (TPA)-enhanced expression of MT1-MMP without changing tissue inhibitors of metalloproteinase (TIMP)-2 level. FPP-3 also suppressed TPA-induced increases in MMP-9 protein and mRNA levels, but did not alter TIMP-1 level. Our results suggest that FFP-3 may be a valuable anti-invasive drug candidate for cancer therapy by suppressing MMP-2, MMP-9, and MT1-MMP.     

金荞麦提取物抑制肿瘤细胞侵袭、转移和HT-1080细胞产生Ⅳ型胶原酶的研究

目的探讨金荞麦提取物对肿瘤细胞侵袭和转移的影响。方法以人工重组基底膜及小鼠黑色素瘤高转移株自发性肺转移模型观察了金荞麦提取物对Ｂ１６－ＢＬ６细胞的体外抗侵袭活性和体内抗转移作用；用聚丙烯酰胺凝胶电泳法进一步观察了其对人纤维肉瘤ＨＴ－１０８０细胞Ⅳ型胶原酶的产生及活性的影响；同时用ＷＳＴ法观察了该药的细胞毒性。结果金荞麦提取物在１００ｍｇＬ－１剂量下能明显抑制Ｂ１６－ＢＬ６细胞侵袭；在２００ｍｇｋｇ－１剂量下能有效抑制Ｂ１６－ＢＬ６黑色素瘤细胞在Ｃ５７／ＢＬ６小鼠体内自发性肺转移。该药对Ｂ１６－ＢＬ６和ＨＴ－１０８０细胞无明显细胞毒作用。该药能抑制ＨＴ－１０８０细胞Ⅳ胶原酶的产生，但对酶的活性无明显影响。结论金荞麦提取物具有明显的抗癌侵袭和转移的作用。     

ZD6474 inhibits proliferation and invasion of human hepatocellular carcinoma cells

Hepatocellular carcinoma (HCC) is characterized by hypervascularization, neoangiogenesis formation and blood vessel invasion. Recently, it has been demonstrated that an inhibitor of the vascular endothelial growth factor (VEGF) receptor, ZD6474, may directly inhibit the growth of tumor cells. ZD6474 effectiveness was investigated on cell growth, apoptosis, adhesion, migration and invasion and related to the drug-dependent modulation of main molecular targets on HCC cells. ZD6474 inhibited HCC cell proliferation, however, such effect was reverted by Laminin-5 (Ln-5) but not by other extracellular matrix proteins (ECM). ZD6474 also inhibited HCC cell adhesion, migration and invasion, whereas the simultaneous treatment with the drug and Ln-5 strongly recovered those effects. Under the same experimental conditions, ZD6474 inhibited the expression of phosphorylated EGFR in all cell lines while the effect on p-Erk1/2 was dependent on cellular invasive characteristics. Nonetheless, co-incubation with Ln-5 completely recovered this effect. Our results support the hypothesis that ZD6474 could represent an interesting therapeutic opportunity for patients with HCC scarcely expressing the ECM protein, Ln-5.     

Anti-inflammatory activity of 20(S)-protopanaxadiol: enhanced heme oxygenase 1 expression in RAW 264.7 cells

20( S)-Protopanaxadiol (PPD) is one of the metabolites of ginsenosides from Panax ginseng. In this study, we demonstrate that PPD inhibits the increase in lipopolysaccharide (LPS)-induced inducible nitric oxide synthase (iNOS) expression through inactivation of nuclear factor-kappaB by preventing degradation of inhibitory factor-kappaBalpha. PPD also induces heme oxygenase 1 (HO-1) expression in RAW 264.7 cells, at the mRNA and protein levels, in the presence and absence of LPS. This effect is associated with suppression of LPS-induced nitric oxide (NO) production and iNOS expression. The HO-1 inducer hemin is associated with the suppression of LPS-induced NO production in a dose-dependent manner, and the HO-1 inhibitor tin protoporphyrin attenuates the inhibitory activity of PPD on LPS-induced NO production. These results provide evidence for the role of HO-1 in the inhibition of LPS-induced NO production by PPD.     

Silibinin activated p53 and induced autophagic death in human fibrosarcoma HT1080 cells via reactive oxygen species-p38 and c-Jun N-terminal kinase pathways

Our previous research demonstrated that hepatic-protectant silibinin induced autophagy in human fibro-sarcoma HT1080 cells through reactive oxygen species (ROS) pathway. Pifithrin-α (PFT-α), a specific inhibitor of p53, reduced autophagy and reversed silibinin's growth-inhibitory effect; besides, PFT-α decreased the activation of caspase-3, a crucial executor of apoptosis. Silibinin upregulated expression of p53/phosphorylated-p53 (p-p53) in a time-dependent manner. Catalase (scavenger of H(2)O(2)), superoxide dismutase (SOD) (scavenger of O(2)(?-)), and SB203580 (inhibitor of p38) attenuated upregulation of p53 expression, suggesting that p53 might be partially regulated by ROS-p38 pathway. On the other hand, c-Jun N-terminal kinase (JNK) increased autophagic death in silibinin-treated cells, and JNK/p-JNK expression was upregulated by silibinin time-dependently. Inhibition of JNK by SP600125 did not influence generation of ROS. Scavengers of H(2)O(2) or O(2)(?-) showed no effect on expression of JNK/p-JNK, indicating that JNK might not correlate with ROS in this process. However, activation of p53 was suppressed by SP600125; therefore the function of p53 was possibly controlled by JNK as well. Western blotting analysis showed that PFT-α reduced activation of extracellular regulated kinase1/2 (ERK1/2) and expression of protein kinase B (PKB, or Akt)/p-Akt. PD98059 (inhibitor of mitogen-activated protein kinase kinase (MEK)/ERK) and wortmannin (inhibitor of phosphoinositide 3-kinase (PI3K)/Akt) enhanced silibinin's cytotoxicity. Wortmannin augmented silibinin-induced autophagy, while PD98059 did not affect autophagic ratio. These results suggest that silibinin might induce p53-mediated autophagic cell death by activating ROS-p38 and JNK pathways, as well as inhibiting MEK/ERK and PI3K/Akt pathways.     

Inhibitory effect of aminoethyl-chitooligosaccharides on invasion of human fibrosarcoma cells

Chitooligosaccharides (COS) have been reported to show a variety of biological efficacies such as anti-bacterial activity, anti-tumor activity and immune activity. The purpose of this study is to investigate the inhibitory effect of aminoethyl-chitooligosaccharides (AE-COS) synthesized from COS that were substituted hydroxyl groups with aminoethyl group at C-6 position on cell invasion of human fibrosarcoma cells. First of all, the effect of AE-COS on cell viability was observed using MTT assay. The cytotoxicity of AE-COS was increased in a dose dependent manner. The inhibitory effects of AE-COS on the activity and expression level of MMP-2 and MMP-9 related to invasion of cancer cells were examined using gelatin zymography and western blot. It was found that AE-COS above 20 μg/ml showed the inhibitory effect on the activity and expression of MMP-9. Furthermore, AE-COS at 20 μg/ml reduced the expression level of p50, a part of NF-κB, compared with phorbol-12- myristate-13- acetate (PMA) group. The available data let us hypothesize that AE-COS could provide chemoprevention as an inhibitor against cell invasion associated with metastasis.     

1,3-Selenazine derivatives induce cytotoxicity and DNA fragmentation in human HT-1080 fibrosarcoma cells.

The inhibitory effects of a series of 5,6-dihydro-4H-1,3-selenazine derivatives, 1,3-selenazole, and 5,6-dihydro-4H-1,3-thiazine derivatives on the proliferation of human HT-1080 fibrosarcoma cells were investigated. The compounds 4-ethyl-4-hydroxy-2-p-tolyl-5, 6-dihydro-4H-1,3-selenazine (TS-2) and 4-hydroxy-4-methyl-6-propyl-2-p-tolyl-5,6-dihydro-4H-1,3-selenazine++ + (TS-6) exhibited the strongest inhibitory effect among 1, 3-selenazine derivatives, and the EC(50) of TS-2 and TS-6 was 7.76 and 8.40 microM, respectively. On the other hand, 1,3-selenazole and 5,6-dihydro-4H-1,3-thiazines had no inhibitory effects. TS-2 and TS-6 inhibited the proliferation of HT-1080 cells time- and dose-dependently. They induced dose-dependent DNA fragmentation in HT-1080 cells, revealing a typical apoptosis characteristics. The present study demonstrated that TS-2 and TS-6 inhibited HT-1080 proliferation through the induction of DNA fragmentation.     

A ginsenoside metabolite, 20-O-beta-D-glucopyranosyl-20(S)-protopanaxadiol, triggers apoptosis in activated rat hepatic stellate cells via caspase-3 activation

We investigated the apoptotic effects of the protopanaxadiol ginsenosides, Rb (1) and Rb (2), and their intestinal bacterial metabolite, 20-O-beta-D-glucopyranosyl-20(S)-protopanaxadiol (M1), and of the protopanaxatriol ginsenoside, Rg (1), and its intestinal bacterial metabolite, 20(S)-protopanaxatriol, in activated rat hepatic stellate cells (HSCs) transformed by Simian virus 40 (T-HSC/Cl-6). As HSCs play a central role in liver fibrosis, agents that selectively induce apoptosis of HSCs could be used to treat this disease. Apoptosis was measured using cell viability tests, DNA fragmentation analysis, and immunoblot analysis of poly(ADP-ribose) polymerase cleavage. M1 (40 microM for 24 h) significantly induced apoptosis in activated rat HSCs. M1 induced apoptosis in a dose-dependent manner as shown by DNA fragmentation, an increased population of cells in the sub-G1 phase, and reduced mitochondrial transmembrane potential. M1 induced caspase-3 activity in a dose- and time-dependent manner. A specific inhibitor of caspase-3 prevented induction of apoptosis by M1 as shown by DNA fragmentation analysis. It is concluded that M1 induces apoptosis in T-HSC/Cl-6 cells via caspase-3 activation.     

20(S)-原人参二醇4种剂型在体透皮实验的初步研究

目的 研究20(S)-原人参二醇(PPD)4种剂型的制备及大鼠在体PPD透皮吸收情况.方法 用薄膜超声法制备PPD药质体和醇质体,用超声法制备环糊精包合物,用增溶法制备PPD溶液;建立在体透皮试验中PPD的HPLC测定方法;利用皮肤剩余量法计算PPD 4种剂型透皮18 h后残留在皮肤内的药量.色谱条件:COSMO-SIL 5 C18-MS-Ⅱ色谱柱(250 mm×4.6 mm,5 μm);流动相为甲醇-水(90:10);体积流量1.0 mL/min;检测波长203 nm;柱温25℃;进样量50μL.结果 豆磷脂、羟丙基-β-环糊精(HP-β-CD)空白皮肤洗液和试剂对药物的测定无干扰,PPD质量浓度在0.1～0.5 mg/mL与峰面积的线性关系良好(r=0.999 9),回收率均在96.93%～104.78%,日内及日间RSD均小于1.54%(n=3).PPD药质体、PPD醇质体、PPD β-环糊精包合物、PPD原药液的皮肤剩余量分别为69.6%、86.36%、100%、73.55%.结论 RP-HPLC法能很好地测定PPD的量,且回归方程线性均较好,表明建立的在体透皮试验方法准确、稳定,适用于PPD经皮给药系统制剂的在体透皮试验研究.     

顺铂联合人类白细胞介素-24基因对于宫颈癌Siha细胞增殖和侵袭力的影响

目的研究人类白细胞介素(hIL)-24基因联合顺铂对人宫颈癌Siha细胞的生长抑制作用以及hIL-24基因对Siha宫颈癌细胞侵袭、迁移能力的影响。方法构建重组质粒pDC316-hIL-24,使用脂质体携带pDC316-hIL-24质粒转染Siha细胞;聚合酶链反应(PCR)法检测hIL-24基因在宫颈癌细胞中的表达情况。Siha细胞在转染hIL-24基因后,添加不同剂量的顺铂,用CCK-8试剂检测hIL-24基因联合顺铂对Siha细胞增殖抑制效果;Transwell实验检测hIL-24基因对Siha细胞迁移侵袭能力的影响。结果重组质粒pDC316-hIL-24成功转染Siha宫颈癌细胞;hIL-24联合顺铂对Siha细胞的增殖抑制作用大于单独使用顺铂或hIL-24基因组(P<0.05);Transwell实验中,hIL-24干预组肿瘤细胞的穿膜细胞数最少(P<0.05)。结论 hIL-24基因的表达可增加Siha宫颈癌细胞对顺铂的敏感性;hIL-24基因可以明显抑制Siha宫颈癌细胞的迁移侵袭能力。     

Geranylated flavanone tomentodiplacone B inhibits proliferation of human monocytic leukaemia (THP-1) cells

BACKGROUND AND PURPOSE

Paulownia tomentosa is a rich source of geranylated flavanones, some of which we have previously shown to have cytotoxic activity. To identify members of this class of compounds with cytostatic effects, we assessed the effects of the geranylated flavanone tomentodiplacone B (TOM B) on cell cycle progression and cell cycle regulatory pathways of THP-1 human monocytic leukaemia cells.

EXPERIMENTAL APPROACH

Cell viability was measured by dye exclusion and proliferation by WST-1 assays; cell cycle was monitored by flow cytometry. Regulatory proteins were assessed by immunoprecipitation and kinase assays, and Western blotting.

KEY RESULTS

Tomentodiplacone B had no effect during the first 24 h of cell growth at concentrations between 1 and 2.5 µM, but inhibited cell growth in a dose-dependent manner at concentrations of 5 µM or higher. Growth inhibition during the first 24 h of exposure to TOM B was not accompanied by cytotoxicity as cells were accumulated in G1 phase dose-dependently. This G1 phase accumulation was associated with down-regulation of cyclin-dependent kinase 2 activity and also protein levels of cyclins E1 and A2. However, key stress-related molecules (γ-H2AX, p53 and p21) were not induced, suggesting that TOM B acts by directly inhibiting the cyclin-dependent kinase 2 signalling pathway rather than initiating DNA damage or cellular stress.

CONCLUSIONS AND IMPLICATIONS

Our study provides the first evidence that TOM B directly inhibits proliferation of human monocytic leukaemia cells, and thus is a potential anticancer agent, preventing leukaemia cells from progressing from G1 phase into DNA synthesis.     

Oligomannurarate sulfate inhibits CXCL12/SDF-1-mediated proliferation and invasion of human tumor cells in vitro

Aim:

JG6 is a novel marine-derived oligosaccharide that has shown to inhibit angiogenesis and tumor metastasis. In this study, we sought to identify the potential target responsible for the anti-cancer activity of JG6.

Methods:

Human liver cancer cell line Bel-7402 and human cervical cancer cell line HeLa were examined. CXCL12-stimulated cell proliferation and migration were determined using a CCK-8 kit and a transwell assay, respectively. Western blotting was performed to examine the changes in CXCL12/CXCR4 axis. Molecular docking and surface plasmon resonance (SPR) were performed to characterize the possible interaction between JG6 and the CXCL12/CXCR4 axis.

Results:

Treatment with CXCL12 potently stimulated the proliferation and migration in both Bel-7402 and HeLa cells. Co-treatment of the cells with JG6 (10, 50 and 100 μg/mL) dose-dependently impeded the CXCL12-stimulated cell proliferation and migration. Furthermore, CXCL12 rapidly induced phosphorylation of AKT, ERK, FAK and Paxillin in Bel-7402 and HeLa cells, whereas pretreatment with JG6 dose-dependently inhibited the CXCL12-induced phosphorylation of these proteins. The SPR assay showed that JG6 bound to CXCL12 with a high affinity. In molecular docking study, JG6 appeared to interact with CXCL12 via multiple polar interactions, including 6 ionic bonds and 7 hydrogen bonds.

Conclusion:

Inhibition of the CXCL12/CXCR4 axis by JG6 may account for its anticancer activity.     

Ginsenoside Rg1 inhibits proliferation of vascular smooth muscle cells stimulated by tumor necrosis factor-α

AIM: To investigate the proliferation of vascular smooth muscle cells (VSMC) affected by ginsenoside Rg1 and further explore the molecular mechanism of ginsenoside Rg1 using proteomics. METHODS: The proliferation of VSMC was measured by MTS assay kit and flow cytometry. Proteomic alterations were analyzed using two-dimensional electrophoresis and peptide mass fingerprinting. Differential proteins found in proteomics were confirmed by RT-PCR. RESULTS: The proliferation of VSMC was enhanced significantly after tumor necrosis factor-alpha (TNF-alpha) treatment, and ginsenoside Rg1 treatment inhibited proliferation in a dose-dependent manner. Proteomic analysis showed 24 protein spots were changed, including 17 spots that were increased and 7 spots that were decreased. Ginsenoside Rg1 could restore the expression levels of these proteins, at least partly, to basic levels of untreated cells. The expression of G-protein coupled receptor kinase, protein kinase C (PKC)-zeta, N-ras protein were decreased, while cycle related protein p21 was increased by ginsenoside Rg1 in TNF-alpha treated VSMC. CONCLUSION: PKC-zeta and p21 pathway might be the mechanism for inhibitory effects of ginsenoside Rg1 on proliferation of VSMC.     

Oligomannurarate sulfate inhibits CXCL12/SDF-1- mediated proliferation and invasion of human tumor cells in vitro

Aim： JG6 is a novel marine-derived oligosaccharide that has shown to inhibit angiogenesis and tumor metastasis. In this study, we sought to identify the potential target responsible for the anti-cancer activity of JG6. Methods： Human liver cancer cell line Bel-7402 and human cervical cancer cell line HeLa were examined. CXCL12-stimulated cell proliferation and migration were determined using a CCK-8 kit and a transwell assay, respectively. Western blotting was performed to examine the changes in CXCL12/CXCR4 axis. Molecular docking and surface plasmon resonance （SPR） were performed to characterize the possible interaction between JG6 and the CXCL12/CXCR4 axis. Results： Treatment with CXCL12 potently stimulated the proliferation and migration in both Bel-7402 and HeLa cells. Co-treatment of the cells with JG6 （10, 50 and 100 pp=VmL） dose-dependently impeded the CXCL12-stimulated cell proliferation and migration. Furthermore, CXCL12 rapidly induced phosphorylation of AKT, ERK, FAK and Paxillin in Bel-7402 and HeLa cells, whereas pretreatment with JG6 dose-dependently inhibited the CXCL12-induced phosphorylation of these proteins. The SPR assay showed that JG6 bound to CXCL12 with a high affinity. In molecular docking study, JG6 appeared to interact with CXCL12 via multiple polar interactions, including 6 ionic bonds and 7 hydrogen bonds. Conclusion： Inhibition of the CXCL12/CXCR4 axis by JG6 may account for its anticancer activity.     

A nanoparticulate drug-delivery system for 20(S)-protopanaxadiol: formulation,characterization, increased oral bioavailability and anti-tumor efficacy

As with many other hydrophobic anticancer agents, 20(S)-protopanaxadiol (PPD) has a very low oral bioavailability. In this study, a precipitation-combined ultrasonication technique was used to prepare PPD nanosuspensions. The mean particle size of the nanosuspensions was approximately 222?±?12?nm, the drug payload achieved 50% after lyophilization and the maximum PPD concentration can reach 100?mg/ml, which is over 30?000 times the solubility of PPD in aqueous solution (3?μg/ml). After oral administration, the C_max and AUC_last values of PPD nanosuspensions were approximately 3.66-fold and 3.48-fold as those of PPD coarse suspensions, respectively. In contrast to the free drug solution, PPD nanosuspensions showed higher in vitro anti-tumor activity against HepG-2 cells (an IC₅₀ value of 1.40 versus 5.83?μg/ml at 24?h, p?<?0.01). The in vivo study in H22-tumor-bearing mice demonstrated that PPD nanosuspensions showed good anti-tumor efficacy with an inhibition rate of 79.47% at 100?mg/kg, while 50?mg/kg of cyclophosphamide was displayed as positive control, and the inhibition rate was 87.81%. Considering the highest drug payload, oral bioavailability reported so far, significant anti-tumor efficacy and excellent safety of encapsulated drugs, PPD nanosuspensions could be used in potential effective strategies for anticancer therapy; further investigation is ongoing.