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Background: Ovarian cancer (OCa) peritoneal metastasis is the leading cause of cancer-related deaths in women with limited therapeutic options available for treating it and poor prognosis, as the underlying mechanism is not fully understood. Method: The clinicopathological correlation of G9a expression was assessed in tumor specimens of ovarian cancer patients. Knockdown or overexpression of G9a in ovarian cancer cell lines was analysed with regard to its effect on adhesion, migration, invasion and anoikis-resistance. In vivo biological functions of G9a were tested by i.p. xenograft ovarian cancer models. Microarray and quantitative RT-PCR were used to analyze G9a-regulated downstream target genes. Results: We found that the expression of histone methyltransferase G9a was highly correlated with late stage, high grade, and serous-type OCa. Higher G9a expression predicted a shorter survival in ovarian cancer patients. Furthermore, G9a expression was higher in metastatic lesions compared with their corresponding ovarian primary tumors. Knockdown of G9a expression suppressed prometastatic cellular activities including adhesion, migration, invasion and anoikis-resistance of ovarian cancer cell lines, while G9a over-expression promoted these cellular properties. G9a depletion significantly attenuated the development of ascites and tumor nodules in a peritoneal dissemination model. Importantly, microarray and quantitative RT-PCR analysis revealed that G9a regulates a cohort of tumor suppressor genes including CDH1, DUSP5, SPRY4, and PPP1R15A in ovarian cancer. Expression of these genes was also inversely correlated with G9a expression in OCa specimens. Conclusion: We propose that G9a contributes to multiple steps of ovarian cancer metastasis and represents a novel target to combat this deadly disease. [ABSTRACT FROM AUTHOR]

Angiogenesis and lymphangiogenesis are considered to play key roles in tumor metastasis. Targeting receptor tyrosine kinases essentially involved in the angiogenesis and lymphangiogenesis would theoretically prevent cancer metastasis. How ever, the optimal multikinase inhibitor for metastasis suppression has yet to be developed. In this study, we evaluated the effect of NSTPBP 0100194-A (194-A), a multikinase inhibitor of vascular endothelial growth factor receptors (VEGFRs)/fibroblast growth factor receptors (FGFRs), on lymphangiogenesis and angiogenesis in a mammary fat pad xenograft model of the highly invasive breast cancer cell line 4T1-Luc+. We investigated the biologic effect of 194-A on various invasive breast cancer cell lines as well as endothelial and lymphatic endothelial cells. Intriguingly, we found that 194-A drastically reduced the formation of lung, liver, and lymph node metastasis of 4T1-Luc+ and decreased primary tumor growth. This was associated with significant reductions in intratumoral lymphatic vessel length (LVL) and microvessel density (MVD). 194-A blocked VEGFRs mediated signaling on both endothelial and lymphatic endothelial cells. Moreover, 194-A significantly inhibited the invasive capacity induced by VEGF-C or FGF-2 in vitro in both 4T1 and MDA-MB231 cells. In conclusion, these experimental results demonstrate that simultaneous inhibition of VEGFRs/FGFRs kinases may be a promising strategy to prevent breast cancer metastasis. [ABSTRACT FROM AUTHOR]

Metastasis is a predominant cause of death in patients with cancer. It is a complex multistep process that needs to be better understood if we are to develop new approaches to managing tumor metastasis. Tumor cell invasion of the local stroma is suppressed by collapsin response mediator protein-1 (CRMP-1). Recently, we identified a long isoform of CRMP-1 (LCRMP-1), expression of which correlates with cancer cell invasiveness and poor clinical outcome in patients with non-small-cell lung cancer (NSCLC). Here, we report that LCRMP-1 overexpression in noninvasive human cell lines enhanced filopodia formation, cancer cell migration, and invasion via stabilization of actin. This effect required a highly conserved N-terminal region of LCRMP-1 as well as the WASP family verprolin-homologous protein-1/actin nucleation pathway (WAVE-1/actin nucleation pathway). Furthermore, LCRMP-1 appeared to act downstream of Cdc42, a Rho family protein known to be involved in actin rearrangement. In addition, LCRMP-1 associated with CRMP-1, which downregulated cancer cell metastasis by interrupting the association of LCRMP-1 and WAVE-1. Finally, we found that high-level expression of LCRMP-1 and low-level expression of CRMP-1 were associated with lymph node metastasis and poor survival in patients with NSCLC. In sum, we show that LCRMP-1 and CRMP-1 have opposing functions in regulating cancer cell invasion and metastasis and propose that this pathway may serve as a potential anticancer target. [ABSTRACT FROM AUTHOR]

Abstract: MicroRNA-10b (miR-10b) has been reported to facilitate the metastasis of breast cancer. However, little is known about the role of miR-10b in the metastasis of nasopharyngeal carcinoma (NPC). Here, we show that high levels of miR-10b expression in Epstein-Barr virus (EBV)-positive latent membrane protein-1 (LMP1)-expressing NPC cells, and its expression is down-regulated by silencing LMP1 or Twist. Induction of miR-10b over-expression in LMP1-silent C666-1 cells promoted significant wound healing and transmembrane invasiveness in vitro. More importantly, miR-10b over-expression promoted the metastasis of NPC and accelerated the death of tumor-bearing nude mice. These findings strongly suggest that miR-10b positively regulates the metastasis of NPC. [ABSTRACT FROM AUTHOR]

Fibroblast growth factor (FGF)-9 belongs to the FGF family which modulate cell proliferation, differentiation, and motility. Benzo(a)pyrene is a polycyclic aromatic hydrocarbon (PAH) and ubiquitous environmental carcinogen present in automobile exhaust, cigarette smoke, and foods. The major purposes of this study were to explore the roles of FGF-9 in the benzo(a)pyrene-induced lung cancer invasion in vitro and the metastatic development of lung adenocarcinoma in human. The data of RT-PCR analysis indicated that treatments of human lung adenocarcinoma CL5 cells with benzo(a)pyrene and a PAH mixture motorcycle exhaust particulate (MEP) extracts increased FGF-9 mRNA expression. The increased expression was blocked by cotreatments with a p38 mitogen-activated protein kinase inhibitor SB202190 and an extracellular signal-regulated kinase inhibitor PD98059. The results of immunoblot analysis and Matrigel assay showed that benzo(a)pyrene and MEP extracts produced a concomitant induction of FGF-9 protein and invasive ability of CL5 cells. The benzo(a)pyrene- and MEP-induced invasion was suppressed by FGF-9 neutralizing antibodies. The results of immunohistochemistry analysis of human lung adenocarcinoma specimens showed that FGF-9 protein was detected in the adenocarcinoma cells but not in normal epithelium. FGF-9 staining intensity was positively correlated with status of disease and degree of lymph node metastasis in these lung adenocarcinomas. These present findings suggest that FGF-9 has potential roles in benzo(a)pyrene-induced CL5 cell invasion and human lung adenocarcinoma metastasis. [ABSTRACT FROM AUTHOR]

Background: Connective tissue growth factor (CTGF) inhibits the metastatic activity of human lung cancer cells in a mouse model; however, the mechanism of this modulation is unclear. We investigated the role of angiogenesis in this process. Methods: CL1-5 and A549 human lung adenocarcinoma cells were stably transfected with vectors containing CTGF or hypoxia-inducible factor (HIF) la or with vector controls. Transfected cells were injected into nude mice (n = 10 per group), and tumor growth, metastasis, and mouse survival were measured. Excised xenograft tumors and primary human lung adenocarcinomas (n = 24) were subjected to immunohistochemistry with antibodies to the endothelial cell marker CD31 and to CTGF. Expression of HIF-1α and vascular endothelial growth factor (VEGF) A was assessed in vitro by using reporter gene assays. Cells were transiently transfected with HIF-1α mutant and antisense arrest-defective 1 protein (ARD-1), and HIF-1α acetylation was assayed by immunoprecipitation. All statistical tests were two-sided. Results: Xenograft tumors derived from CTGF transfectants grew more slowly than those from control-transfected cells and had reduced expression of HIF-1α and VEGF-A, vascularization (as assessed by CD31 expression), and metastasis (all P<.001). Xenograft tumors derived from CTGF-overexpressing cells that were transfected with HIF-1α had higher VEGF-A expression than CTGF-overexpressing xenografts. Mice with CTGF/HIF-1α xenografts had lower survival than mice carrying CTGF-overexpressing xenografts (CL1-5/Neo, mean = 69.6 days, 95% confidence interval [CI ] = 53.9 to 85.3 days versus CL1-5/CTGF, mean = 102.1 days, 95% CI = 92.1 to 112.1 days; P = .001, CL1-5/CTGF, mean = 102.1 days, 95% CI = 92.1 to 112.1 days versus CLI-5/CTGF/HIF-la, mean = 81.7 days, 95% CI = 66.5 to 96.9 days; P = .011, CLI-5/Neo, mean = 69.6 days, 95% CI = 53.9 to 85.3 days versus CL1-5/CTGF/HI F-1α, mean = 81.7 days, 95% CI = 66.5 to 96.9 days; P = .122). Tumors of patients with the same disease stage but with high CTGF protein expression had reduced microvessel density compared with tumors with low expression. Transfection with antisense-ARD1 decreased the level of acetylated HIF-1α and restored HIF-1α and VEGF-A expression in CTGF-overexpressing cells. Conclusion: CTGF inhibition of metastasis involves the inhibition of VEGF-A-dependent angiogenesis, possibly by promoting HIF-1α protein degradation. [ABSTRACT FROM AUTHOR]

Summary: Flt-4, a VEGF receptor, is activated by its specific ligand, VEGF-C. The resultant signaling pathway promotes angiogenesis and/or lymphangiogenesis. This report provides evidence that the VEGF-C/Flt-4 axis enhances cancer cell mobility and invasiveness and contributes to the promotion of cancer cell metastasis. VEGF-C/Flt-4-mediated invasion and metastasis of cancer cells were found to require upregulation of the neural cell adhesion molecule contactin-1 through activation of the Src-p38 MAPK-C/EBP-dependent pathway. Examination of tumor tissues from various types of cancers revealed high levels of Flt-4 and VEGF-C expression that correlated closely with clinical metastasis and patient survival. The VEGF-C/Flt-4 axis, through upregulation of contactin-1, may regulate the invasive capacity in different types of cancer cells. [Copyright &y& Elsevier]

Arsenic trioxide (ATO) has been implicated as a promising anticancer agent by inhibiting cell growth and inducing apoptosis in certain types of cancer cells. This study explored the antimetastasis property of arsenic, drew potential link between arsenic use and radiotherapy, and uncovered the specific mechanisms underlying these remarkable responses. Using gelatin invasion assay and intravasation assay, we report the novel finding that low-dose ATO (1?µM) reduced the intrinsic migration ability of HeLa cells and significantly inhibited radiation-promoted tumor invasive potential of CaSki cells without inducing apoptotic cell death. Using the murine Lewis lung carcinoma model, the control animals and ATO treatment animals (1?mg/kg i.p., twice weekly) displayed similar in vivo growth kinetics, whereas the radiation (30?Gy in one fraction) and concurrent treatment groups showed considerable growth inhibition. Importantly, although concurrent treatment did not enhance the effectiveness of radiation therapy to the primary tumor, further examination of the lungs revealed that all animals succumbed to radiation-accelerated lung metastases could be effectively treated by combination of ATO and radiation. Radiation-induced matrix metalloproteinase-9 (MMP-9) expression was significantly inhibited by ATO using sequential analysis of the expression of MMPs in xenografts. Supporting this observation, ATO directly downregulates radiation-induced MMP-9 mRNA expression by inhibiting nuclear factor?B activity in human cervical cancer cells. In sum, concurrent arsenic-radiation therapy not only achieves local tumor control but also inhibits distant metastasis. Experimental results of this study highlight a novel strategy in cancer treatment.Oncogene (2005) 24, 390-398. doi:10.1038/sj.onc.1208192 Published online 8 November 2004 [ABSTRACT FROM AUTHOR]

Background: Tumor invasion and metastasis cause most deaths in cancer patients. Connective tissue growth factor (CTGF), a secreted protein that binds to integrins, modulates the invasive behavior of certain human cancer cells, but few mechanistic details are known. We investigated the roles of CTGF and collapsin response mediator protein 1 (CRMP-1) in metastasis and invasion of human lung adenocarcinoma. Methods: We compared vector control-transfected cells with corresponding CTGF gene-transfected cells. Invasive activity was measured with a modified Boyden chamber assay, and metastatic activity was measured in an animal model. We used CTGF deletion mutants, CTGF and CRMP-1 antisense oligonucleotides, and anti-integrin and anti-CRMP-1 antibodies to investigate the functional relationship between CTGF and CRMP-1. Expression of CTGF protein in 78 lung adenocarcinoma specimens was investigated immunohistochemically. All statistical tests were twosided. Results: Invasive (both P<.001) and metastatic (P<.001 and P = .003, respectively) activities were lower in cells that overexpress CTGF than in vector control cells. Expression of CRMP-1 was higher in CTGF-transfected clones than in vector control cells, and its level decreased after cells were treated with anti-integrin αvβ3 and αvβ5 antibodies. Reduced levels of CRMP-1 protein after the transfection of CRMP-1-specific antisense oligonucleotides, but not sense oligonucleotides, increased the invasiveness of CTGF-transfected cells (mean numbers of invasive CTGFtransfected cells treated with 20 µM CRMP-1-specific sense and antisense oligonucleotides were 327 and 516 cells, respectively [difference = 189 cells, 95% confidence interval {CI} = 156 to 221 cells; P<.001]). The CT module of CTGF was the region primarily responsible for the increased expression of CRMP-1 and the inhibition of invasion (mean numbers of invasive cells expressing full-length CTGF and CT module-deleted mutant were 148 and 385 cells, respectively [difference = 237 cells, 95% CI = 208 to 266 cells; P<.001]). Reduced expression of CTGF in lung cancer specimens was statistically significantly associated with the risk of more advanced-stage disease (stages III and IV versus stages I and II; P = .001), lymph node metastasis (P = .014), and shorter survival (median survival with high levels of CTGF = 66.7 months and median survival for low levels = 18.2 months; difference = 48.5 months, 95% CI = 33.5 to 63.5 months; P = .02). Conclusion: CTGF inhibits metastasis and invasion of human lung adenocarcinoma by a CRMP1-dependent mechanism. [ABSTRACT FROM AUTHOR]

A correction is presented to the article "Inhibition of TGF-β with neutralizing antibodies prevents radiation-induced acceleration of metastatic cancer progression" which appeared in the previous issue.

Laryngeal and hypopharyngeal squamous cell carcinomas (LHSCCs) are common head and neck cancers with a high propensity for lymph node (LN) and lung metastasis. Here, we report that LHSCCs express high levels of functional CXCR4 receptors, native for chemokine stromal cell-derived factor-1 (SDF-1/CXCL12). Primary tumor immunohistochemistry from LHSCC patients has revealed significant expression of CXCR4 and CXCL12. Greater expression of CXCR4 but not that of CXCL12 is correlated with LN and distant metastasis. Reverse transcription–polymerase chain reaction and western blots have demonstrated that CXCR4 messenger RNA (mRNA) and protein were expressed in LHSCC cell lines as well, but failed to detect CXCL12 mRNA expression. CXCL12 treatment enhanced extracellular signal-regulated kinase (ERK) pathway activation and the motility/invasiveness of LHSCC cell lines, which were blocked by treatment with a CXCR4 antagonist (AMD3100) and a specific MEK inhibitor (U0126). Results show that the mRNA and protein levels of matrix metalloproteinase (MMP)-13, but not MMP-2 or MMP-9, were elevated in HEp-2 cells in response to CXCL12. Again, U0126 almost inhibited the induction of MMP-13 in HEp-2 cells by stimulating CXCL12. The transcriptional factor, c-Jun, a downstream factor of ERK pathway, was found to be readily phosphorylated and translocated to the nucleus after 10 min of exposure to CXCL12. Blockage of c-Jun activity by transfection with c-jun antisense oligodeoxynucleotide significantly decreased CXCL12-induced MMP-13 expression and cell invasion. CXCL12 seems to enhance LHSCC cell invasion through paracrine-activated CXCR4, which triggers ERK/c-Jun-dependent MMP-13 upregulation. [ABSTRACT FROM AUTHOR]