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Extracellular matrix (ECM)-adhesion proteins and actin cytoskeleton are pivotal in cancer cell invasion. Ras Suppressor-1 (RSU-1), a cell-ECM adhesion protein that interacts with PINCH-1, thus being connected to Integrin Linked Kinase (ILK), alpha-parvin (PARVA), and actin cytoskeleton, is up-regulated in metastatic breast cancer (BC) samples. Apart from the originally-identified gene (RSU-1L), an alternatively-spliced isoform (RSU-1-X1) has been reported. We used non-invasive MCF-7 cells, expressing only RSU-1L, and highly invasive MDA-MB-231-LM2 expressing both isoforms and generated stable shRNA-transduced cells lacking RSU-1L, while the truncated RSU-1-X1 isoform was depleted by siRNA-mediated silencing. RSU-1L depletion in MCF-7 cells resulted in complete abrogation of tumor spheroid invasion in three-dimensional collagen gels, whereas it promoted MDA-MB-231-LM2 invasion, through a compensatory upregulation of RSU-1-X1. When RSU-1-X1 was also eliminated, RSU-1L-depletion-induced migration and invasion were drastically reduced being accompanied by reduced urokinase plasminogen activator expression. Protein expression analysis in 23 human BC samples corroborated our findings showing RSU-1L to be upregulated and RSU-1-X1 downregulated in metastatic samples. We demonstrate for the first time, that both RSU-1 isoforms promote invasion in vitro while RSU-1L elimination induces RSU-1-X1 upregulation to compensate for the loss. Hence, we propose that both isoforms should be blocked to effectively eliminate metastasis.

Dexamethasone is a glucocorticoid steroid with anti-inflammatory properties used to treat many diseases, including cancer, in which it helps manage various side effects of chemo-, radio-, and immunotherapies. Here, we investigate the tumor microenvironment (TME)-normalizing effects of dexamethasone in metastatic murine breast cancer (BC). Dexamethasone normalizes vessels and the extracellular matrix, thereby reducing interstitial fluid pressure, tissue stiffness, and solid stress. In turn, the penetration of 13 and 32 nm dextrans, which represent nanocarriers (NCs), is increased. A mechanistic model of fluid and macromolecule transport in tumors predicts that dexamethasone increases NC penetration by increasing interstitial hydraulic conductivity without significantly reducing the effective pore diameter of the vessel wall. Also, dexamethasone increases the tumor accumulation and efficacy of ∼30 nm polymeric micelles containing cisplatin (CDDP/m) against murine models of primary BC and spontaneous BC lung metastasis, which also feature a TME with abnormal mechanical properties. These results suggest that pretreatment with dexamethasone before NC administration could increase efficacy against primary tumors and metastases. 13 6 6396 6408

Metastatic dissemination of cancer cells to distal organs is the major cause of death for patients suffering from the aggressive basal-like breast cancer (BLBC) subtype. Recently, we have shown that interleukin 13 receptor alpha 2 (IL13Rα2) is a critical gene that is overexpressed in a subset of BLBC primary tumors associated with poor distant metastasis-free survival (DMFS) and can promote extravasation and metastasis of breast cancer cells to the lungs. However, the upstream signaling mechanisms that promote aberrant IL13Rα2 expression during tumor progression remain unknown. Driven by our previously published gene expression microarray data derived from a well-characterized cell line model for BLBC progression, we show that both Inhibin βA (INHBA) and IL13Rα2 genes exhibit similarly higher expression levels in metastatic compared to non-metastatic cells and that overexpression of both genes predicts worse metastasis-free survival of patients with high grade tumors. Activin A, a member of the TGFβ superfamily comprising two INHBA subunits, has been shown to play context-depended roles in cancer progression. Here, we demonstrate that INHBA depletion downregulates IL13Rα2 expression in metastatic breast cancer cells, whereas treatment with Activin A in non-metastatic cells increases its expression levels. We also find that Activin A predominantly induces Smad2 phosphorylation and to a lesser extent activates Smad3 and Akt. Interestingly, we also show that Activin A-mediated upregulation of IL13Rα2 is Smad2-dependent since knocking down Smad2 or using the ALK4/ALK5 inhibitors EW-7197 and SB-505124 abolishes this effect. Most importantly, our data indicate that knocking down INHBA levels in breast cancer cells delays primary tumor growth, suppresses migration in vitro and inhibits the formation of lung metastases in vivo. Conclusively, our findings presented here suggest that the development of therapeutic interventions employing small molecule inhibitors against Activin receptors or neutralizing antibodies targeting Activin A ligand, could serve as alternative approaches against breast tumors overexpressing INHBA and/or IL13Rα2. 9

Simple Summary Despite recent therapeutic advances against cancer, many patients do not respond well or respond poorly, to treatment and develop resistance to more than one anti-cancer drug, a term called multi-drug resistance (MDR). One of the main factors that contribute to MDR is the deregulation of apoptosis or programmed cell death. Herein, we describe the major apoptotic pathways and discuss how pro-apoptotic and anti-apoptotic proteins are modified in cancer cells to convey drug resistance. We also focus on our current understanding related to the interactions between survival and cell death pathways, as well as on mechanisms underlying the balance shift towards cancer cell growth and drug resistance. Moreover, we highlight the role of the tumor microenvironment components in blocking apoptosis in MDR tumors, and we discuss the significance and potential exploitation of epigenetic modifications for cancer treatment. Finally, we summarize the current and future therapeutic approaches for overcoming MDR. The ability of tumor cells to evade apoptosis is established as one of the hallmarks of cancer. The deregulation of apoptotic pathways conveys a survival advantage enabling cancer cells to develop multi-drug resistance (MDR), a complex tumor phenotype referring to concurrent resistance toward agents with different function and/or structure. Proteins implicated in the intrinsic pathway of apoptosis, including the Bcl-2 superfamily and Inhibitors of Apoptosis (IAP) family members, as well as their regulator, tumor suppressor p53, have been implicated in the development of MDR in many cancer types. The PI3K/AKT pathway is pivotal in promoting survival and proliferation and is often overactive in MDR tumors. In addition, the tumor microenvironment, particularly factors secreted by cancer-associated fibroblasts, can inhibit apoptosis in cancer cells and reduce the effectiveness of different anti-cancer drugs. In this review, we describe the main alterations that occur in apoptosis-and related pathways to promote MDR. We also summarize the main therapeutic approaches against resistant tumors, including agents targeting Bcl-2 family members, small molecule inhibitors against IAPs or AKT and agents of natural origin that may be used as monotherapy or in combination with conventional therapeutics. Finally, we highlight the potential of therapeutic exploitation of epigenetic modifications to reverse the MDR phenotype.