Inhibitory effects of isoliquiritigenin on the migration and invasion of human breast cancer cells

Introduction: Isoliquiritigenin (ISL) is a natural phenolic compound extracted from licorice. Previous studies have shown that ISL is a potent antioxidant with anti-inflammatory and antitumor activities. The anti-invasive activity of ISL was still unclear. The actual causes of death for most breast cancer patients were due to the tumor metastasis. Attenuating the expression of vascular endothelial growth factor (VEGF) and matrix metalloproteinases (MMPs) is well known to prevent tumor metastasis.

Objectives: The purpose of this study is to investigate the effects of ISL on VEGF and MMP expression in highly metastatic human breast cancer cell line, MDA-MB-231.

Results: ISL reduced the secretions and protein levels of VEGF. The VEGF upstream regulatory protein, hypoxia-inducible factor 1-alpha (HIF-1α), was also reduced after ISL treatment. Moreover, ISL inhibited the expression and gelatinolytic activity of MMP-2 and MMP-9 which were confirmed by western blot and gelatin zymography assay. Additionally, the anti-migratory activity of ISL was further confirmed by chamber migration assay and wound migration assay. Upstream signaling pathways, including the expression of phosphatidylinositol-3 kinase (PI3K), the phosphorylation of p38 and Akt kinase and NF-κB DNA binding activity, were suppressed by ISL.

Conclusion: These findings suggest that ISL suppresses the migration of MDA-MB-231 cells by inhibiting the upstream signaling pathways.     

Thymoquinone Suppresses Migration of Human Renal Carcinoma Caki-1 Cells through Inhibition of the PGE2-Mediated Activation of the EP2 Receptor Pathway

Renal cell carcinoma (RCC) is likely to metastasize to other organs, and is often resistant to conventional chemotherapies. Thymoquinone (TQ), a phytochemical derived from the seeds of Nigella sativa, has been shown to inhibit migration and metastasis in various cancers. In this study, we assessed the effect of TQ on the migratory activity of human RCC Caki-1 cells. We found that treatment with TQ reduced the proteolytic activity of matrix metalloproteinase-9 (MMP-9) in Caki-1 cells. TQ significantly repressed prostaglandin E₂ (PGE₂) production, its EP2 receptor expression as well as the activation of Akt and p38, the well-known upstream signal proteins of MMP-9. In addition, treatment with butaprost, a PGE₂ agonist, also induced MMP-9 activity and migration/invasion in Caki-1 cells. Moreover, pharmacological inhibitors of PI3K/Akt and p38 remarkably attenuated butaprost-induced Caki-1 cell migration and invasion, implying that activation of PI3K/Akt and p38 is a bridge between the PGE₂-EP2 axis and MMP-9-dependent migration and invasion. Taken together, these data suggest that TQ is a promising anti-metastatic drug to treat advanced and metastatic RCC.     

Evaluation of anti-invasion effect of cannabinoids on human hepatocarcinoma cells

Context: Cancer is a disease characterized by abnormal growth of cells. One of the most common types of liver cancers is called hepatocellular carcinoma (HCC) which is highly metastatic. As most of cannabinoids have shown anticancer effect against different cell lines in a number of reports, a biological investigation of two cannabinoids, CB65 (CB2 receptor agonist) and ACEA (CB1 receptor agonist) was carried out in this study.

Objective: In an attempt to find natural products as a new solution of cancer, this study was designed to investigate the potential antitumoral and anti-invasive activity of cannabinoids on HepG2 cells and the possible roles of matrix metalloproteinase-2 (MMP-2) and MMP-9 in its action.

Materials and methods: The researchers examined the effect of various concentrations of CB65 (CB2 receptor agonist) and ACEA (CB1 receptor agonist), on the cell proliferation, viability, and invasion as well as expression of MMP-2 and MMP-9 in HepG2 cells using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazoliumbromide (MTT) assay, matrigel invasion assay, and western blotting method.

Results: The results revealed that both cannabinoids reduce cell viability, cell invasion as well as MMP-2 and MMP-9 expression in higher dose of 20?nM. Furthermore, higher concentrations of examined cannabinoids were more effective.

Discussion: These data suggest ACEA and CB65 as an option for novel treatment of hepatocellular cancer.

Conclusion: Our findings may contribute to design of new therapeutic strategies for the management of HCC.     

Curcumin synergistically increases effects of β-interferon and retinoic acid on breast cancer cells in vitro and in vivo by up-regulation of GRIM-19 through STAT3-dependent and STAT3-independent pathways

Objective: The study aimed to investigate the effects of combination treatment of curcumin and β-interferon (IFN-β)/retinoic acid (RA) on breast cancer cells, including cell viability, apoptosis and migration, and to determine the mechanisms related to GRIM-19 through STAT3-dependent and STAT3-independent pathways.

Methods: The following groups were used for the in vitro experiment: control siRNA, GRIM-19 siRNA, IFN-β/RA and IFN-β/RA?+?curcumin. Cell viability is by the MTT method, cell apoptosis by flow cytometry and cell migration by wound healing experiment; GRIM-19, STAT3, survivin, Bcl-2, GADD153 and COX-2 expression was measured by Western blot. In vivo experiment, MCF-7 cells were subcutaneously injected into nude mice.

Results: GRIM-19 siRNA promoted MCF-7 cell proliferation and migration; inhibited cell apoptosis; and promoted the expression of STAT3, survivin, Bcl-2 and MMP-9. IFN-β/RA inhibited cell proliferation and migration; promoted cell apoptosis; up-regulated GRIM-19; and inhibited the expression of STAT3, survivin, Bcl-2 and MMP-9. Combination treatment of curcumin and IFN-β/RA had a stronger effect than that of the IFN-β/RA group. In addition, curcumin and IFN-β/RA combination inhibited the expression of COX-2 and up-regulated GADD153.

Conclusion: Curcumin synergistically increases the effects of IFN-β/RA on breast cancer cells. The mechanism may be related to the up-regulation of GRIM-19 through STAT3-dependent and STAT3-independent pathways.     

miR-578调控长链脂酰辅酶A合成酶4表达对胃癌细胞增殖、侵袭的影响

目的 探讨miR-578调控长链脂酰辅酶A合成酶4（ACSL4）对胃癌细胞增殖及侵袭的影响。方法 选取人胃癌细胞MKN-45、SNU-1、MKN-7、KTAO3、N87及人正常胃黏膜细胞GES-1为研究对象,将MKN-45细胞分为对照组、miR-578 NC组、miR-578 mimic组、miR-578 mimic+pc-ACSL4组,测定各组MKN-45细胞活力及凋亡、侵袭、迁移能力,测定细胞中miR-578、ACSL4、凋亡相关蛋白（Bax、Caspase-3、Bcl-2）、侵袭相关蛋白（MMP-9、MMP-2）的表达,验证miR-578与ACSL4的靶向关系。结果 miR-578在人胃癌细胞中低表达;与对照组相比,miR-578 mimic组MKN-45细胞活力、侵袭、迁移能力、MMP-2、MMP-9、Bcl-2蛋白表达水平均显著降低（P<0.05）,凋亡能力、Caspase-3、Bax蛋白表达水平均显著升高（P<0.05）;与miR-578 mimic组相比,miR-578 mimic+pc-ACSL4组MKN-45细胞活力、侵袭、迁移能力、MMP-2、MMP-9、Bcl-2蛋白表达水平均显著升高（P<0.05）,凋亡能力、Caspase-3、Bax蛋白表达水平均显著降低（P<0.05）;荧光素酶实验结果显示,miR-578与ACSL4存在靶向调节关系。结论 miR-578可抑制人胃癌细胞MKN-45的增殖及迁移能力,可能是通过抑制ACSL4表达实现的。     

miRNA-381靶向HMGB1对IHH-4细胞生长侵袭及迁移的影响

目的 探究微小RNA-381（miR-381）调控高迁移率族蛋白B1（HMGB1）对甲状腺乳头状瘤IHH-4细胞生长、侵袭及迁移的影响。方法 购置甲状腺乳头状瘤IHH-4细胞及正常甲状腺细胞Nthy-ori-3各1株,取第3代细胞培养48 h后,采用实时定量PCR （qRT-PCR）检测miR-381和HMGB1在甲状腺乳头状瘤IHH-4细胞和正常甲状腺Nthy-ori-3细胞中的表达水平。利用Lipofectamine 2000转染miR-381 mimic后,检测IHH-4细胞中miR-381和HMGB1的表达水平。生物信息学预测miR-381和HMGB1的靶向关系,并用荧光素酶报告实验验证两者的靶向关系。每孔按2×10⁵/mL细胞密度接种IHH-4细胞,分为IHH-4组（甲状腺乳头状瘤细胞IHH-4组）、miR-381 mimic组（miR-381 mimic转染组）、pc-HMGB1组（HMGB1过表达转染组）、mimic+pc-HMGB1组（miR-381 mimic和pc-HMGB1共同转染组）,每组设3个复孔,采用CCK8法检测各组IHH-4细胞活性,Transwell及划痕实验分别检测IHH-4细胞侵袭及迁移能力,免疫印迹法检测IHH-4细胞中Ki67、基质金属蛋白酶-2（MMP-2）、MMP-9和晚期糖基化终产物受体（RAGE）的表达水平。结果 与Nthy-ori-3细胞比较,IHH-4细胞中miR-381 mRNA水平降低,HMGB1 mRNA水平升高,差异有统计学意义（P<0.01）。与IHH-4组相比,miR-381 mimic组miR-381表达增加（P<0.01）。miR-381和HMGB1存在靶向关系。与IHH-4组相比,miR-381 mimic组细胞增殖倍数、Ki67表达、侵袭细胞数、划痕闭合率、MMP-2、MMP-9和RAGE表达均降低（P均<0.01）,pc-HMGB1组上述指标均升高（P均<0.01）;与miR-381 mimic组相比,mimic+pc-HMGB1组细胞增殖倍数、Ki67表达、侵袭细胞数、划痕闭合率、MMP-2、MMP-9和RAGE水平也明显上升（P均<0.01）。结论 miR-381通过靶向下调HMGB1表达,抑制IHH-4细胞增殖、侵袭和迁移。     

S-adenosylmethionine inhibits the activated phenotype of human hepatic stellate cells via Rac1 and Matrix metalloproteinases

ObjectiveTo investigate the effects of S-adenosylmethionine (SAM) on the proliferation, adhesion, migration, invasion and apoptosis of activated human hepatic stellate cells (HSCs) and to explore the relevant potential mechanisms.MethodsHuman HSCs LX-2 were cultured with SAM. The proliferation and adhesion were detected by CCK-8. Cell apoptosis rate were analyzed by flow cytometry, and cell migration and invasion were tested by the transwell assay. The expression of Rac1 and MMP-2 was identified by real-time PCR or Western blotting, and activated Rac1 was detected by GST pull-down assay. The activity of MMP-2 and MMP-9 was analyzed by substrate zymography.ResultsThe proliferation of LX-2 cells was inhibited by SAM, exhibiting a dose-dependent manner. Cell apoptosis rate induced by SAM was remarkably increased. SAM decreased the adhesion, migration and invasion of LX-2 cells. The expression and activation of Rac1 in LX-2 cells were significantly suppressed by SAM. In contrast, the activity of MMP-2 and MMP-9 was enhanced by SAM. SAM attenuated the up-regulated expression of Smad3/4 and Rac1 induced by TGF-β1.ConclusionSAM inhibits the proliferation, adhesion, migration and invasion of LX-2 cells in vitro probably via attenuating the expression and activation of Rac1 and up-regulating MMP-2 and MMP-9 expression, which possibly provide a molecular basis for potential application of SAM in the therapy of liver fibrosis.     

Inhibition of invasion and metastasis of human liver cancer HCCLM3 cells by portulacerebroside A

Abstract

Context: Portulacerebroside A (PCA) is a novel cerebroside compound isolated from Portulaca oleracea L. (Portulacaceae), an edible and medicinal plant distributed in the temperate and tropical zones worldwide.

Objective: This study investigates the effects of PCA in human liver cancer HCCLM3 cells on metastasis and invasion.

Materials and methods: After the cells were treated with PCA (2.5, 5, and 10?μg/ml) for 6, 12, 24, or 48?h, adhesion, transwell invasion, and scratch tests were conducted and cell functions were evaluated. Western blot and FQ-RT-PCR assays explored the mechanism of PCA-inhibited invasion and metastasis in the cells.

Results: The adhesion rate of the cells was suppressed at 0.5?h (79.4?±?1.0, 68.7?±?1.3, and 58.1?±?1.3%, versus 100?±?1.5% in the control), 1?h (78.2?±?1.2, 70.9?±?1.6, and 55.4?±?1.9%, versus 100?±?1.2% in the control), and 1.5?h (71.6?±?1.1, 62.3?±?0.9, and 50.4?±?0.9%, versus 100?±?1.1% in the control). The 24?h invasion ability was decreased (356.6?±?11.2, 204.0?±?17.6, and 113.0?±?9.5%, versus 443.6?±?15.4% in the control). The migration capability was also restrained by PCA for 24?h (324.8?±?25.4, 250.4?±?21.0, and 126.3?±?10.1, versus 381.6?±?30.6 in the control) and 48?h (470.3?±?34.3, 404.0?±?19.7, and 201.0?±?15.4, versus 752.0?±?63.6 in the control). There was an increase in the mRNA and protein expression levels of TIMP-2 and nm23-H1, inhibition in the mRNA expression of MTA1, MMP-2, and MMP-9, and suppression in the protein expression of MTA1, RhoA, Rac1/Cdc42, MMP-2, but not RhoC and MMP-9.

Conclusion: PCA suppresses the invasion and metastasis of HCCLM3 cells possibly by modulation of the mRNA and protein expression of related parameters. This is the first study to reveal a new potential therapeutic application of PCA in antimetastatic therapy for liver cancer.     

Status of research on matrix metalloproteinases (MMPs) in India

Introduction: MMPs are extracellular matrix (ECM)-degrading enzymes that play a crucial role in embryogenesis, tissue remodeling, inflammation and angiogenesis. MMP-2 and -9 (also called type 2 and type 4 collagenase, or gelatinase A and B) are the key molecules that control invasion, tumor growth and metastasis. Tissue inhibitor of matrix metalloproteinase (TIMP) -2 and -1 are specific inhibitors of MMP-2 and MMP-9 respectively, and play a crucial role in regulation of MMP-2 and -9 activation, during pathophysiological processes. MMPs can specifically degrade native gelatin, collagens, fibronectin, ectactin and elastin. MMP-2 and -9 are overexpressed in almost all types of cancers, and so act as important therapeutic targets.

Areas covered: The status of MMP research in India from 1998 to 2010. In this review, the authors cover the role of matrix metalloproteinase inhibitors in cancer therapy.

Expert opinion: As compared with other parts of the world, Indian scientists have not generated a significant number of specific MMP inhibitors for the treatment of cancer, or other diseases. MMPs and membrane type (MT)-MMPs are potentially important therapeutic targets in many diseases, including cancers, therefore, designing specific inhibitors from natural products or through synthetic routes, is crucial.     

黄芪通过微小 RNA-133a抑制 Janus激酶 /信号转导和转录激活因子信号通路抑制膀胱癌的迁移和侵袭

目的探讨黄芪对膀胱癌细胞迁移和侵袭的影响及分子机制。方法 2019年 8月至 2020年 2月,从美国模式培养物研究所( ATCC)购入膀胱癌 T24细胞,体外培养并分为对照( NC)组、黄芪组、 miR-133a模拟物( mimic)阴性对照( miR-NC)组、 miR-133a mimic(miR-133a)组、 miR-133a抑制剂( inhibitor)阴性对照( anti-miR-NC)组、 miR-133a inhibitor(anti-miR-133a)组、芪+anti-miR-NC组、黄芪 +anti-miR-133a组、黄芪 +anti-miR-133a+AG490组。蛋白质印迹法( western blotting)检测基质金属蛋白黄酶-2(MMP-2)、基质金属蛋白酶 -9(MMP-9)磷酸化 Janus激酶 1(p-JAK1)、磷酸化信号转导和转录激活因子 6(p-STAT6)蛋白表达; Transwell检测细胞迁移和侵袭;实时荧光、定量 PCR(RT-qPCR)检测 miR-133a表达水平。结果与对照组相比,黄芪组 MMP-2、MMP-9蛋白表达水平降低,细胞迁移［(81.42±8.05)比( 175.43±16.02)］和侵袭［(71.23±8.01)比( 142.55±14.03)］数量降低, miR-133a表达水平［( 2.36±0.21)比( 1.00±0.11)］升高, p-JAK1［( 0.12±0.01)比( 0.55±0.04)］、 p-STAT6［( 0.08±0.01)比( 0.38±     

阻断Wnt信号通路对人绒毛膜癌细胞株JEG-3迁移及侵袭能力的影响

目的 观察阻断Wnt信号通路后，人绒毛膜癌细胞株JEG-3细胞迁移及侵袭能力的影响。方法 将人绒毛膜癌细胞株JEG-3细胞传代后，按完全随机法分为实验组和对照组，每组6孔。实验组在常规细胞培养基础上，给予外源性Wnt信号通路阻断剂DKK-1（100 ng/mL），对照组加入同体积细胞培养基。运用蛋白印迹法（Western blot法）、逆转录聚合酶链反应（RT-PCR）法检测两组细胞中β-catenin、Wnt、E-cadherin、Vimentin等蛋白及mRNA的表达；采用划痕实验、Transwell实验观测两组细胞迁移及侵袭能力的变化。结果 与对照组相比，实验组β-catenin、Wnt、Vimentin蛋白及mRNA的表达均降低，E-cadherin蛋白及mRNA的表达增高，差异均有统计学意义（P<0.05）；实验组肿瘤细胞迁移及穿膜细胞数较对照组均降低，差异有统计学意义（P<0.05）。结论 阻断Wnt信号通路可抑制肿瘤细胞上皮-间充质转化过程，进而降低人绒毛膜癌细胞株JEG-3细胞的迁移及侵袭能力。     

Progress and contrasts of the development of tivozanib for therapy of kidney cancer

Introduction: Targets for drug development for the treatment of kidney cancer (renal cell carcinoma; RCC) include vascular endothelial growth factor (VEGF) and its receptors and mammalian target of rapamycin. Currently available oral multitargeted VEGF tyrosine kinase inhibitors (TKIs) that have been approved by the US Food and Drug Administration for advanced RCC, include sunitinib, sorafenib and pazopanib. Off-target TKI inhibition can potentially preclude full-dose combination with other targeted and chemotherapeutic agents. There is a need to develop more potent and selective targeted agents for RCC therapy, which are more effective and have minimal off-target effects.

Areas covered: This drug evaluation review addresses the ongoing development for the treatment of RCC with tivozanib: a potent, selective and long-half-life VEGF TKI. The testing for clinical efficacy alone or in combination with other therapies for RCC and for other tumor types, and the clinical and market relevance of introducing another RCC therapy are discussed.

Expert opinion: Tivozanib is distinguished by its high potency, selectivity, long-half-life and its potential to be effectively combined with other agents in RCC. This may offer more effective, yet well-tolerated treatment options. The relative clinical and market relevance remain to be seen, both for RCC therapy and other tumor types.     

In vitro effect of IL-2 in combination with pazopanib or sunitinib on lymphocytes function and apoptosis of RCC cells

Objective: Combination of immunotherapy with tyrosine kinase inhibitors (TKIs) has been used with some success for the treatment of metastatic renal cell carcinoma. Herein we evaluate the in vitro effect of high-dose interleukin-2 (HDIL-2) and pazopanib or sunitinib on the lymphocyte function and on induction of apoptosis in renal cell carcinoma (RCC) cell lines.

Methods: Peripheral blood mononuclear cells (PBMCs) isolated from healthy donors or RCC patients were treated with different HDIL-2/TKI combinations. Effects of different combinations on proliferation and cytotoxic activity of PBMCs were evaluated, in addition to their effect on apoptosis of three different RCC cell lines.

Results: While sunitinib did not inhibit the proliferation of various immune cells induced by HDIL-2, pazopanib appeared to inhibit the HDIL-2–induced proliferation of these cells. Interestingly, none of the HDIL-2/TKI combinations appeared to compromise the functional properties of these cells. Additionally, significant proportion of RCC cell lines treated with pazopanib alone underwent apoptosis, while the proportions of apoptotic cells post-HDIL-2 or sunitinib were not different from the background. Furthermore, the combination of HDIL-2/pazopanib did not inhibit the pazopanib-induced RCC apoptosis.

Conclusion: The combination of HDIL-2 with either pazopanib or sunitinib exerts different anticancer mechanisms that could enhance the treatment efficacy.