c-Src signaling in triple negative breast cancer cells: role of Cyr61

Póster presentado al CNIO Frontiers Meeting: "Metastasis Initiation: Mechanistic Insights and Therapeutic Opportunities", celebrado en el Spanish National Cancer Research Centre (CNIO) Auditorium del 28 al 30 de septiembre de 2015.The SFKs (Src Family Kinases) control cellular pathways involved in division, motility, adhesion, angiogenesis, and survival. Therefore, their deregulation is associated with tumorigenesis, and metastasis. c-Src is overexpressed and/or aberrantly activated in epithelial tumors: pancreatic, colorectal, prostatic, ovarian, breast, etc. We previously showed that SFKs catalytic inhibitors (Dasatinib, PP2, and SU6656) reduce proliferation, migration, and invasiveness of MDA-MB-231. Here, we analyzed c-Src contribution to initial steps of metastasis by Tet-On conditional expression of a specific shRNA-c-Src, which suppressed c-Src mRNA and protein levels in MDA-MB-231. c-Src suppression did not alter cell proliferation or survival, but it significantly reduced anchorage-independent growth. Concomitantly with diminished tyrosine-phosphorylation/activation of Fak, caveolin-1, paxillin and p130CAS, c-Src depletion inhibited migration, invasion, transendothelial migration, and reduced MMP2, MMP7 and MMP9 in secretome. Quantitative proteomic analyses of secretome showed that Cyr61 levels, detected in exosomal fraction, were diminished upon shRNA-c-Src expression. However, Cyr61 expression was unaltered inside cells. Cyr61 partially colocalized with cis-Golgi gp74 marker, and with exosomal marker CD63, but c-Src depletion did not alter their distribution. In SUM159PT, transient c-Src suppression also reduced secreted exosomal Cyr61. Furthermore, conditional expression of c-Src dominant negative mutant (c-Src-K295M/Y527F) in MDA-MB-231 and in SUM159PT diminished secreted Cyr61 as well. Cyr61 transient suppression in MDA-MB-231 inhibited invasion and transendothelial migration. Finally, in both MDA-MB-231 and SUM159PT, a neutralizing Cyr61 antibody restrained migration. Collectively, these results suggest that c-Src regulates secreted proteins, including exosomal Cyr61, which are involved in modulating the metastatic potential of triple negative breast cancer cells.Peer Reviewe

Role of c-Src in Human MCF7 Breast Cancer Cell Tumorigenesis

Peer Reviewe

The multidrug transporter P-glycoprotein: A mediator of melanoma invasion?

Malignant melanoma shows high levels of intrinsic drug resistance associated with a highly invasive phenotype. In this study, we investigated the role of the drug transporter P-glycoprotein (Pgp) in the invasion potential of drug-sensitive (M14 WT, Pgp-negative) and drug-resistant (M14 ADR, Pgp-positive) human melanoma cells. Coimmunoprecipitation experiments assessed the association of Pgp with the adhesion molecule CD44 in multidrug resistant (MDR) melanoma cells, compared with parental ones. In MDR cells, the two proteins colocalized in the plasma membrane as visualized by confocal microscopy and immunoelectron microscopy on ultrathin cryosections. MDR melanoma cells displayed a more invasive phenotype compared with parental cells, as demonstrated by quantitative transwell chamber invasion assay. This was accomplished by a different migration strategy adopted by resistant cells ("chain collective") previously described in tumor cells with high metastatic capacity. The Pgp molecule, after stimulation with specific antibodies, appeared to cooperate with CD44, through the activation of ERK1/2 and p38 mitogen-activated protein kinase (MAPK) proteins. This activation led to an increase of metalloproteinase (MMP-2, MMP-3, and MMP-9) mRNAs, and proteolytic activities, which are associated with an increased invasive behavior. RNA interference experiments further demonstrated Pgp involvement in migration and invasion of resistant melanoma cells. A link was identified between MDR transporter Pgp, and MAPK signaling and invasion. © 2007 The Society for Investigative Dermatology

c-Src functionality controls self-renewal and glucose metabolism in MCF7 breast cancer stem cells

Deregulation of Src kinases is associated with cancer. We previously showed that SrcDN conditional expression in MCF7 cells reduces tumorigenesis and causes tumor regression in mice. However, it remained unclear whether SrcDN affected breast cancer stem cell functionality or it reduced tumor mass. Here, we address this question by isolating an enriched population of Breast Cancer Stem Cells (BCSCs) from MCF7 cells with inducible expression of SrcDN. Induction of SrcDN inhibited self-renewal, and stem-cell marker expression (Nanog, Oct3-4, ALDH1, CD44). Quantitative proteomic analyses of mammospheres from MCF7-Tet-On-SrcDN cells (data are available via ProteomeXchange with identifier PXD017789, project DOI: 10.6019/PXD017789) and subsequent GSEA showed that SrcDN expression inhibited glycolysis. Indeed, induction of SrcDN inhibited expression and activity of hexokinase, pyruvate kinase and lactate dehydrogenase, resulting in diminished glucose consumption and lactate production, which restricted Warburg effect. Thus, c-Src functionality is important for breast cancer stem cell maintenance and renewal, and stem cell transcription factor expression, effects linked to glucose metabolism reduction.This work has been supported by grand SAF2016–75991-R (MINECO, AEI/FEDER, UE) to Jorge Martín-Pérez and ISCIII [grand PI 16/00789] to Miguel Ángel Fernández-Moreno. Víctor Mayoral-Varo was supported by the grand SAF2016–75991-R (MINECO, AEI/FEDER, UE). We acknowledge support for publication fee by the CSIC Open Access Publication Support Initiative through its Unit for Information Resources for Research (URICI)

Drug Delivery Systems of Natural Products in Oncology

In recent decades, increasing interest in the use of natural products in anticancer therapy field has been observed, mainly due to unsolved drug-resistance problems. The antitumoral effect of natural compounds involving different signaling pathways and cellular mechanisms has been largely demonstrated in in vitro and in vivo studies. The encapsulation of natural products into different delivery systems may lead to a significant enhancement of their anticancer efficacy by increasing in vivo stability and bioavailability, reducing side adverse effects and improving target-specific activity. This review will focus on research studies related to nanostructured systems containing natural compounds for new drug delivery tools in anticancer therapies

L'Auto-vélo : automobilisme, cyclisme, athlétisme, yachting, aérostation, escrime, hippisme / dir. Henri Desgranges

04 août 19101910/08/04 (A11,N3580)

Design of new nanocarriers for biomedical applications

One of the main prerequisite of a therapeutic drug is to overcome a series of physiological barriers and it to be less toxic for the human. To this aim, it is necessary to find a carrying vector able to enhance drug internalization and to improve its therapeutic efficacy. Presently, biocompatible and biodegradable nanoparticles (NPs) as some effective drug delivery devices, are widely studied. In order to increase the antimicrobial and antitumor efficacy of different natural products numerous studies are in progress to evaluate different drug delivery systems (cationic liposomes, lysozyme-shelled hollow nano/microbubbles and nano/microcapsules). This delivery system would have a synergistic and additive effect decreasing the drug resistance reactions too. The present article has tried to review some of the latest research on nanocarriers in clinical care

Activation of polyamine catabolic enzymes involved in diverse responses against epibrassinolide-induced apoptosis in LNCaP and DU145 prostate cancer cell lines

Epibrassinolide (EBR) is a biologically active compound of the brassinosteroids, steroid-derived plant growth regulator family. Generally, brassinosteroids are known for their cell expansion and cell division-promoting roles. Recently, EBR was shown as a potential apoptotic inducer in various cancer cells without affecting the non-tumor cell growth. Androgen signaling controls cell proliferation through the interaction with the androgen receptor (AR) in the prostate gland. Initially, the development of prostate cancer is driven by androgens. However, in later stages, a progress to the androgen-independent stage is observed, resulting in metastatic prostate cancer. The androgen-responsive or -irresponsive cells are responsible for tumor heterogeneity, which is an obstacle to effective anti-cancer therapy. Polyamines are amine-derived organic compounds, known for their role in abnormal cell proliferation as well as during malignant transformation. Polyamine catabolism-targeting agents are being investigated against human cancers. Many chemotherapeutic agents including polyamine analogs have been demonstrated to induce polyamine catabolism that depletes polyamine levels and causes apoptosis in tumor models. In our study, we aimed to investigate the mechanism of apoptotic cell death induced by EBR, related with polyamine biosynthetic and catabolic pathways in LNCaP (AR+), DU145 (AR-) prostate cancer cell lines and PNT1a normal prostate epithelial cell line. Induction of apoptotic cell death was observed in prostate cancer cell lines after EBR treatment. In addition, EBR induced the decrease of intracellular polyamine levels, accompanied by a significant ornithine decarboxylase (ODC) down-regulation in each prostate cancer cell and also modulated ODC antizyme and antizyme inhibitor expression levels only in LNCaP cells. Catabolic enzymes SSAT and PAO expression levels were up-regulated in both cell lines; however, the specific SSAT and PAO siRNA treatments prevented the EBR-induced apoptosis only in LNCaP (AR+) cells. In a similar way, MDL 72,527, the specific PAO and SMO inhibitor, co-treatment with EBR during 24 h, reduced the formation of cleaved fragments of PARP in LNCaP (AR+) cells. © 2013 Springer-Verlag Wien