BackgroundRho GTPases are involved in cellular functions relevant to cancer. The roles of RhoA and Rac1 have already been established. However, the role of Rac3 in cancer aggressiveness is less well understood.MethodsThis work was conducted to analyze the implication of Rac3 in the aggressiveness of two breast cancer cell lines, MDA-MB-231 and MCF-7: both express Rac3, but MDA-MB-231 expresses more activated RhoA. The effect of Rac3 in cancer cells was also compared with its effect on the non-tumorigenic mammary epithelial cells MCF-10A. We analyzed the consequences of Rac3 depletion by anti-Rac3 siRNA.ResultsFirstly, we analyzed the effects of Rac3 depletion on the breast cancer cells’ aggressiveness. In the invasive MDA-MB-231 cells, Rac3 inhibition caused a marked reduction of both invasion (40%) and cell adhesion to collagen (84%), accompanied by an increase in TNF-induced apoptosis (72%). This indicates that Rac3 is involved in the cancer cells’ aggressiveness. Secondly, we investigated the effects of Rac3 inhibition on the expression and activation of related signaling molecules, including NF-κB and ERK. Cytokine secretion profiles were also analyzed. In the non-invasive MCF-7 line; Rac3 did not influence any of the parameters of aggressiveness.ConclusionsThis discrepancy between the effects of Rac3 knockdown in the two cell lines could be explained as follows: in the MDA-MB-231 line, the Rac3-dependent aggressiveness of the cancer cells is due to the Rac3/ERK-2/NF-κB signaling pathway, which is responsible for MMP-9, interleukin-6, -8 and GRO secretion, as well as the resistance to TNF-induced apoptosis, whereas in the MCF-7 line, this pathway is not functional because of the low expression of NF-κB subunits in these cells. Rac3 may be a potent target for inhibiting aggressive breast cancer.

BackgroundMultidrug resistance (MDR) is one of the major problems in the treatment of cancer. Overcoming it is therefore expected to improve clinical outcomes for cancer patients. MDR is usually characterized by overexpression of ABC (ATP-binding cassette) protein transporters such as P-gp, MRP1, and ABCG2. Though the importance of ABC transporters for cancer cells is recognized, few studies have looked at its implications for the endothelial cells that are essential to tumor angiogenesis. This study investigated the expression and functions of these ABC transporters in endothelial cells in vitro and their potential contribution to cancer growth in mice.MethodsHuman micro vessel endothelial cells (HMEC-1) and human umbilical vein endothelial cells (HUVEC) were exposed to increasing doses of Doxorubicin (Dox) to induce ABC gene expression. Cell viability was then quantified by 3H-thymidine and MTS assay. Flow cytometry, qPCR, and western blot were used to detect mRNA and the protein expression of P-gp, MRP1, and ABCG2. The intracellular accumulation of Rhodamine 123 (Rho) was used to evaluate drug efflux function and the inhibitors for P-gp, ABCG2, and MRP1 were used to verify their respective roles in vitro. In an attempt to evaluate drug resistance in endothelial cells in vivo, athymic mice were treated with Dox for 15 days before a MDA-MB-435 tumor graft to observe subsequent changes in the inhibition curves of tumor growth in response to Dox treatment. Furthermore, endothelial cells from multiple sites in these mice were also isolated to estimate their P-gp expression by flow cytometry.ResultsDrug resistance in HMEC-1 and HUVEC was successfully induced by the addition of Dox to the culture media. Two stabilized subcell lines of HMEC1 (HMECd1 and HMECd2) showed 15- and 24-fold increases in resistance. Tests also showed that these induced endothelial cells were cross-resistant to the structurally unrelated drugs Daunorubicin, Vinblastine, and Etoposide. P-gp protein levels increased four and six fold in HMECd1 and HMECd2 as revealed by western blot. The qPCR demonstrated 3.4- and 7.2-fold increases in P-gp, and a slight increase in ABCG2, gene expression. The Rho accumulation within these cells was inversely correlated with the expression levels of P-gp. The inhibitors of P-gp, but not of ABCG2 or MRP1, were able to block the induced endothelial cell resistance to Dox. Furthermore, we also showed that injecting Dox into healthy mice induced an increase in P-gp expression in endothelial cells. Using these pretreated mice in a tumor growth experiment, we observed a dramatic diminution in the therapeutic efficiency of Dox treatment, suggesting implications for drug resistance in mice endothelial cells supporting tumor growth.ConclusionsABC transporter expression can be induced in endothelial cells in vitro. This study also indicates that P-gp plays an important role in the acquisition of resistance to Dox in endothelial cells and that this reduces the efficiency of chemotherapy.

Abstract. Voltage-gated Na + channels (VGSCs) are highly expressed in several types of carcinomas including breast, prostate and lung cancers as well as in mesothelioma and cervical cancers. Although the VGSCs activity is considered crucial for the potentiation of cancer cell migration and invasion, the mechanisms responsible for their functional expression and regulation in cancer cells remain unclear. In the present study, the role of the small GTPase RhoA in the regulation of expression and function of the Nav1.5 channel in the breast cancer cell lines MDA-MB 231 and MCF-7 was investigated. RhoA silencing significantly reduced both Nav1.5 channel expression and sodium current indicating that RhoA exerts a stimulatory effect on the synthesis of an active form of Nav1.5 channel in cancer cells. The inhibition of Nav1.5 expression dramatically reduced both cell invasion and proliferation. In addition, a decrease of RhoA protein levels induced by Nav1.5 silencing was observed. Altogether, these findings revealed: i) the key role of the small GTPase RhoA in upregulation of Nav1.5 channel expression and tumor aggressiveness, and ii) the existence of a positive feedback of Nav1.5 channels on RhoA protein levels. IntroductionBreast cancer is the most commonly occurring cancer in women. Each year approximately one million of new cases occurred in the world, 25-40% of patients developing metastasis and dying from cancer (1,2). Many problems remain in its clinical management which is generally related to the malignancy types and drug resistance mechanisms as well as metastasis development. Additional investigations to understand the physiopathology of breast cancer are urgently necessary to develop new therapies based on new targets involved in cancer cell invasion and proliferation.In previous studies we have already demonstrated that the upregulation and activation of the small GTPase RhoA or RhoC contribute to cell invasion leading to breast cancer metastasis (3,4). RhoA signaling pathways are implicated in the activation of FAK, Akt/PI3K, p38MAPK and MLCK, which have been shown to be responsible for cytoskeleton actin reorganization, cell adhesion, motility, migration and invasion (5-9). In many types of cancers, RhoA appears to be overexpressed and/or constitutively activated (10,11), and considered to be a negative clinical prognosis marker (10,12,13).Recently, it has been shown that the voltage-gated Na + channels (VGSCs) could be an accelerating factor in malignant cancers (14-17). VGSCs are membrane panning proteins expressed in a wide variety of excitable and non-excitable cells, as well as in many carcinomas including breast cancer (18,19). VGSCs mainly mediate rapid and transient Na + influx into cells and are classically responsible for generation and propagation of action potential. In cancer cells, overexpression of VGSCs and/or their upregulation has been observed. In particular, the Nav1.5 channel type correlates with cancer cell invasion (20-22). Although it seems that the neonatal isoform of Nav1.5 ch...