Regorafenib diminishes the expression and secretion of angiogenesis and metastasis associated proteins and inhibits cell invasion via NF-κB inactivation in SK-Hep1 cells.
ABSTRACT: The aim of the present study was to investigate the effects of regorafenib on the nuclear factor κ-light-chain-enhancer of activated B cells (NF)-κB-modulated expression of angiogenesis- and metastasis-associated proteins and cell invasion in human hepatocellular carcinoma SK-Hep1 cells. The SK-Hep1 cells were treated with different concentrations of NF-κB inhibitor 4-N-[2-(4-phenoxyphenyl) ethyl] quinazoline-4,6-diamine (QNZ) or regorafenib for 24 or 48 h. The effects of QNZ and regorafenib on cell viability, NF-κB activation, expression and secretion levels of angiogenesis- and metastasis-associated proteins and cell invasion were evaluated with MTT assays, western blotting, ELISA, gelatin zymography and cell invasion assays. The results demonstrated that QNZ and regorafenib significantly reduced the expression and secretion levels of the angiogenesis- and metastasis-associated proteins vascular endothelial growth factor, tumor necrosis factor-α, interleukin (IL)-1β, IL-6, matrix metalloproteinase (MMP)-2 and MMP-9, NF-κB activation and cell invasion. In conclusion, the inhibition of NF-κB activation induces anti-angiogenic and antimetastatic effects in SK-Hep1 cells. Regorafenib reduces the level of expression and secretion of angiogenesis- and metastasis-associated proteins and cell invasion through the suppression of NF-κB activation in SK-Hep1 cells.
Project description:Numerous studies have shown that the levels of matrix metalloproteinase (MMP)-2 and/or MMP-9 are associated with the invasive phenotypes of cancer cells. This study investigated the effects of caffeic acid phenethyl ester (CAPE), a chemopreventive phytochemical derived from honeybee propolis, on the invasive phenotype of SK-Hep1 human hepatocellular carcinoma cells (SK-Hep1 cells). CAPE effectively suppressed SK-Hep1 cell invasion in a dose-dependent manner. The constitutive expression of MMP-2 and MMP-9 in SK-Hep1 cells was almost completely abolished by treatment with 12.5 muM CAPE. CAPE also significantly inhibited nuclear factor kappa B (NF-kappaB) DNA-binding activity in SK-Hep1 cells. These results taken together suggest that CAPE exerts antimetastatic potential through inhibition of MMP-2 and MMP-9 expression, possibly by targeting NF-kappaB in hepatocellular carcinoma.
Project description:<h4>Background</h4>Resistin, adipocyte-secreting adipokine, may play critical role in modulating cancer pathogenesis. The aim of this study was to investigate the effects of resistin on HCC adhesion to the endothelium, and the mechanism underlying these resistin effects.<h4>Methods</h4>Human SK-Hep1 cells were used to study the effect of resistin on intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) expressions as well as NF-?B activation, and hence cell adhesion to human umbilical vein endothelial cells (HUVECs). 5-Aminoimidazole-4-carboxamide 1-?-D-ribofuranoside (AICAR), an AMP-activated protein kinase (AMPK) activator, was used to determine the regulatory role of AMPK on HCC adhesion to the endothelium in regard to the resistin effects.<h4>Results</h4>Treatment with resistin increased the adhesion of SK-Hep1 cells to HUVECs and concomitantly induced NF-?B activation, as well as ICAM-1 and VCAM-1 expressions in SK-Hep1 cells. Using specific blocking antibodies and siRNAs, we found that resistin-induced SK-Hep1 cell adhesion to HUVECs was through NF-?B-regulated ICAM-1 and VCAM-1 expressions. Moreover, treatment with AICAR demonstrated that AMPK activation in SK-Hep1 cells significantly attenuates the resistin effect on SK-Hep1 cell adhesion to HUVECs.<h4>Conclusions</h4>These results clarify the role of resistin in inducing HCC adhesion to the endothelium and demonstrate the inhibitory effect of AMPK activation under the resistin stimulation. Our findings provide a notion that resistin play an important role to promote HCC metastasis and implicate AMPK may be a therapeutic target to against HCC metastasis.
Project description:CONTEXT:Development of novel strategies in the treatment of advanced thyroid cancer are needed. Our laboratory has previously identified a role for nuclear factor κB (NF-κB) signaling in human thyroid cancer cell growth, survival, and invasion. OBJECTIVE:Our goal was to establish the role of NF-κB signaling on thyroid cancer growth and metastases in vivo and to begin to dissect mechanisms regulating this effect. SETTING AND DESIGN:We examined tumor formation of five thyroid cancer cell lines in an in vivo model of thyroid cancer and observed tumor establishment in two of the cell lines (8505C and BCPAP). RESULTS:Inhibition of NF-κB signaling by overexpression of a dominant-negative IκBα (mIκBα) significantly inhibited thyroid tumor growth in tumors derived from both cell lines. Further studies in an experimental metastasis model demonstrated that NF-κB inhibition impaired growth of tumor metastasis and prolonged mouse survival. Proliferation (mitotic index) was decreased in 8505C tumors, but not in BCPAP tumors, while in vitro angiogenesis and in vivo tumor vascularity were significantly inhibited by mIkBα only in the BCPAP cells. Cytokine antibody array analysis demonstrated that IL-8 secretion was blocked by mIκBα expression. Interestingly, basal NF-κB activity and IL-8 levels were significantly higher in the two tumorigenic cell lines compared with the nontumorigenic lines. Furthermore, IL-8 transcript levels were elevated in high-risk human tumors, suggesting that NF-κB and IL-8 are associated with more aggressive tumor behavior. CONCLUSIONS:These studies suggest that NF-κB signaling is a key regulator of angiogenesis and growth of primary and metastatic thyroid cancer, and that IL-8 may be an important downstream mediator of NF-κB signaling in advanced thyroid cancer growth and progression.
Project description:Highlights • The cellular activity of USPs is functionally implicated in cancer progression.• USP8 negatively regulates the TRAF6-mediated signals for NF-κB and autophagy.• USP8KO SK-HEP-1 cells reveal increases of cancer progression in vivo and in vitro.• TCGA and transcriptome data support the functional role of USP8 in liver cancers. Herein, we aimed to elucidate the molecular and cellular mechanism in which ubiquitin-specific protease 8 (USP8) is implicated in liver cancer progression via TRAF6-mediated signal. USP8 induces the deubiquitination of TRAF6, TAB2, TAK1, p62, and BECN1, which are pivotal roles for NF-κB activation and autophagy induction. Notably, the LIHC patient with low USP8 mRNA expression showed markedly shorter survival time, whereas there was no significant difference in the other 18-human cancers. Importantly, the TCGA data analysis on LIHC and transcriptome analysis on the USP8 knockout (USP8KO) SK-HEP-1 cells revealed a significant correlation between USP8 and TRAF6, TAB2, TAK1, p62, and BECN1, and enhanced NF-κB-dependent and autophagy-related cancer progression/metastasis-related genes in response to LPS stimulation. Furthermore, USP8KO SK-HEP-1 cells showed an increase in cancer migration and invasion by TLR4 stimulation, and a marked increase of tumorigenicity and metastasis in xenografted NSG mice. The results demonstrate that USP8 is negatively implicated in the LIHC progression through the regulation of TRAF6-mediated signal for the activation of NF-κB activation and autophagy induction. Our findings provide useful insight into the LIHC pathogenesis of cancer progression.
Project description:Protein phosphatase of regenerating liver‑3 (PRL‑3) is considered to be metastasis‑associated phosphatase and is associated with a poor prognosis. Additionally, tumor‑associated macrophages (TAMs) participate in cancer progression. A previous study demonstrated that PRL‑3 promotes invasion and metastasis by inducing TAM infiltration. However, the underlying mechanism has not been elucidated. In the present study, western blot analysis, polymerase chain reaction, immunohistochemistry, ELISA, mouse model experiments and functional experiments were performed to confirm that the interaction between TAMs and colorectal cancer (CRC) cells induced epithelial‑mesenchymal transition (EMT)‑associated features in CRC cells by activating mitogen‑activated protein kinase (MAPK) pathways in TAMs and upregulating the expression of interleukin (IL)‑6 and IL‑8. The neutralization of IL‑6 and IL‑8 reduced EMT and the invasive and migratory abilities of CRC cells. Therefore, IL‑6 and IL‑8 were considered important factors in EMT, and in CRC invasion and metastasis. In addition, increased angiogenesis was observed after TAMs were co‑cultured with CRC cells that overexpress PRL‑3. Vascular endothelial growth factor‑A was significantly upregulated, and the nuclear factor‑κB (NF‑κB) signaling pathway was activated in CRC cells after co‑culture. Moreover, nude mice injected with CRC cells with high PRL‑3 expression levels tended to generate larger xenografts. Immunohistochemistry results from xenografted CRC cells overexpressing PRL‑3 also confirmed the activation of MAPK pathways in xenografts. Overall, the findings indicate that PRL‑3 promotes CRC cell invasion and metastasis by activating MAPK pathways in TAMs to initiate the EMT, and PRL‑3 promotes angiogenesis by activating the NF‑κB pathway in CRC cells.
Project description:<b>Background:</b> Tumor-infiltrating immune cells are closely associated with tumor occurrence and progression. The present study explored the potential mechanism of tumor-infiltrating plasmacytoid dendritic cells (pDC) mediating the proliferation and metastasis of cancer cells in oral squamous cell carcinoma (OSCC). <b>Methods:</b> pDC distribution was detected by immunofluorescence and flow cytometry. chemotaxis cytokine receptor-4/7 (CXCR-4/7) expression was detected by quantitative polymerase chain reaction and immunohistochemistry. Cell proliferation and migration were measured by CCK-8, colony formation, wound healing and transwell assay. ELISA and western blotting were used to investigate cytokines secretion and NF-κB pathway activity. <b>Results:</b> Tumor-infiltrating pDC in OSCC was significantly increased and associated with tumor size, lymph node (LN) metastasis (<i>P</i> <0.05). Tumor-infiltrating-pDC-conditioned medium from OSCC patients significantly promoted tumor cell proliferation and invasion, which was at least partly mediated via enhancing the CXCR-4 expression of tumor cell. In addition, the activation of NF-κB pathway played a decisive role in the overexpression of CXCR-4, which was further regulated by pDC-derived TNF-α secretion. <b>Conclusions:</b> Tumor-infiltrating pDC promoted oral cancer proliferation and invasion via activating the TNF-α/NF-κB/CXCR-4 pathway, which may serve as a potential immunological target for the treatment of OSCC in the future.
Project description:Potential roles of euchromatic histone methyltransferase 2 (<i>EHMT2</i> or G9a) in invasion and metastasis are not well understood in non-small cell lung cancer (NSCLC). Here, we investigated the effect and underlying mechanisms of G9a and therapeutic implications of targeting G9a in the invasion and metastasis of NSCLC. Overexpression of G9a significantly enhanced <i>in vitro</i> proliferation and invasion, while knockdown of G9a drastically suppressed <i>in vivo</i> growth and metastasis of A549 and H1299 NSCLC cells. Knockdown or inhibition of G9a significantly decreased the expression of focal adhesion kinase (FAK) protein and activation of FAK pathway. In addition, defactinib, a potent FAK inhibitor, partially abolished the G9a-enhanced invasion in these NSCLC cells. Furthermore, targeting G9a was found to suppress NF-κB transcriptional activity in NSCLC cells through stabilizing NF-κB inhibitor alpha (IκBα), while an NF-κB inhibitor Parthenilide partially abolished the G9a-enhanced FAK activation, which suggests that G9a-enhanced invasion and activation of FAK is mediated by elevated NF-κB activity. Notably, a strong positive correlation between the IHC staining of G9a and phosphorylated FAK proteins was identified in H1299 xenografts and 159 cases of NSCLC tissues (R = 0.408). IMPLICATIONS: The findings of this study strongly demonstrate that G9a may promote invasion and metastasis of NSCLC cells by enhancing FAK signaling pathway via elevating NF-κB transcriptional activity, indicating potential significance and therapeutic implications of these pathways in the invasion and metastasis of NSCLCs that overexpress G9a protein.
Project description:Hydrogen sulfide (H<sub>2</sub> S), an endogenous signaling gaseous molecule, is involved in various physiological activities, including vessel relaxation, regulation of cellular bioenergetics, inflammation, and angiogenesis. By using xenograft orthotopic implantation of prostate cancer PC3 cells and subsequently comparing bone metastatic with primary tumor-derived cancer cells, we find that H<sub>2</sub> S-producing enzyme cystathionine γ-lyase (CTH) is upregulated in bone-metastatic PC3 cells. Clinical data further reveal that the expression of CTH is elevated in late-stage prostate cancer patients, and higher CTH expression correlates with poor survival from The Cancer Genome Atlas (TCGA) prostate cancer RNA-seq datasets. CTH promotes NF-κB nuclear translocation through H<sub>2</sub> S-mediated sulfhydration on cysteine-38 of the NF-κB p65 subunit, resulting in increased IL-1β expression and H<sub>2</sub> S-induced cell invasion. Knockdown of CTH in PC3 cells results in the suppression of tumor growth and distant metastasis, while overexpression of CTH in DU145 cells promotes primary tumor growth and lymph node metastasis in the orthotopic implanted xenograft mouse model. Together, our findings provide evidence that CTH generated H<sub>2</sub> S promotes prostate cancer progression and metastasis through IL-1β/NF-κB signaling pathways.
Project description:GNA13 has been found overexpressed in various types of cancer, which is related to tumor metastasis and progression. However, the biological functions of GNA13 in colorectal cancer (CRC) progression remain unclear. This study aimed to explore the role of GNA13 in CRC and investigate the mechanism of how GNA13 promotes tumor growth. Interestingly, our findings showed that GNA13 is commonly upregulated in CRC, where these events are associated with a worse histologic grade and poor survival. Increased expression levels of GNA13 promoted cell growth, migration, invasion, and epithelial-mesenchymal transition, whereas GNA13 silencing abrogated these malignant phenotypes. In addition, overexpressing GNA13 in cancer cells increased the levels of the chemokines CXCL1, CXCL2, and CXCL4, which contributed to CRC proliferation and colony formation. Moreover, our mechanistic investigations suggest that the NF-κB/p65 signaling pathway was activated by the increase in GNA13 levels. Inhibiting the NF-κB/p65 pathway with an inhibitor decreased GNA13-induced migration, invasion and CXCL chemokine level increases, indicating the critical role of NF-κB/p65 signaling in mediating the effects of GNA13 in CRC. Together, these results demonstrate a key role of GNA13 overexpression in CRC that contributes to malignant behavior in cancer cells, at least in part through stimulating angiogenesis and increasing the levels of the NF-κB-dependent chemokines CXCL1, CXCL2, and CXCL4.
Project description:The high mortality in breast cancer is often associated with metastatic progression in patients. Previously we have demonstrated that testes-specific protease 50 (TSP50), an oncogene overexpressed in breast cancer samples, could promote cell proliferation and tumorigenesis. However, whether TSP50 also has a key role in cell invasion and cancer metastasis, and the mechanism underlying the process are still unclear. Here we found that TSP50 overexpression greatly promoted cell migration, invasion, adhesion and formation of the stellate structures in 3D culture system in vitro as well as lung metastasis in vivo. Conversely, TSP50 knockdown caused the opposite changes. Mechanistic studies revealed that NF-κB signaling pathway was required for TSP50-induced cell migration and metastasis, and further results indicated that TSP50 overexpression enhanced expression and secretion of MMP9, a target gene of NF-κB signaling. In addition, knockdown of MMP9 resulted in inhibition of cell migration and invasion in vitro and lung metastasis in vivo. Most importantly, immunohistochemical staining of human breast cancer samples strongly showed that the coexpression of TSP50 and p65 as well as TSP50 and MMP9 were correlated with increased metastasis and poor survival. Furthermore, we found that some breast cancer diagnosis-associated features such as tumor size, tumor grade, estrogen receptors (ER) and progesterone receptors (PR) levels, were correlated well with TSP50/p65 and TSP50/MMP9 expression status. Taken together, this work identified the TSP50 activation of MMP9 as a novel signaling mechanism underlying human breast cancer invasion and metastasis.