scholarly journals Direct interaction of metastasis-inducing S100P protein with tubulin causes enhanced cell migration without changes in cell adhesion

2020 ◽  
Vol 477 (6) ◽  
pp. 1159-1178
Author(s):  
Min Du ◽  
Guozheng Wang ◽  
Igor L. Barsukov ◽  
Stephane R. Gross ◽  
Richard Smith ◽  
...  
Keyword(s):  
Cell Migration ◽  
Cell Adhesion ◽  
Hela Cells ◽  
Cancer Metastasis ◽  
Dissociation Constants ◽  
Breast Cancer Metastasis ◽  
Breast Cancers ◽  
Β Tubulin

Overexpression of S100P promotes breast cancer metastasis in animals and elevated levels in primary breast cancers are associated with poor patient outcomes. S100P can differentially interact with nonmuscle myosin (NM) isoforms (IIA > IIC > IIB) leading to the redistribution of actomyosin filaments to enhance cell migration. Using COS-7 cells which do not naturally express NMIIA, S100P is now shown to interact directly with α,β-tubulin in vitro and in vivo with an equilibrium Kd of 2–3 × 10−7 M. The overexpressed S100P is located mainly in nuclei and microtubule organising centres (MTOC) and it significantly reduces their number, slows down tubulin polymerisation and enhances cell migration in S100P-induced COS-7 or HeLa cells. It fails, however, to significantly reduce cell adhesion, in contrast with NMIIA-containing S100P-inducible HeLa cells. When taxol is used to stabilise MTs or colchicine to dissociate MTs, S100P's stimulation of migration is abolished. Affinity-chromatography of tryptic digests of α and β-tubulin on S100P-bound beads identifies multiple S100P-binding sites consistent with S100P binding to all four half molecules in gel-overlay assays. When screened by NMR and ITC for interacting with S100P, four chemically synthesised peptides show interactions with low micromolar dissociation constants. The two highest affinity peptides significantly inhibit binding of S100P to α,β-tubulin and, when tagged for cellular entry, also inhibit S100P-induced reduction in tubulin polymerisation and S100P-enhancement of COS-7 or HeLa cell migration. A third peptide incapable of interacting with S100P also fails in this respect. Thus S100P can interact directly with two different cytoskeletal filaments to independently enhance cell migration, the most important step in the metastatic cascade.

scholarly journals Therapy-induced senescence in normal tissue promotes breast cancer metastasis

2020 ◽  
Author(s):  
Douglas W. Perkins ◽  
Syed Haider ◽  
David Robertson ◽  
Richard Buus ◽  
Lynda O’Leary ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Normal Tissue ◽  
Systemic Chemotherapy ◽  
Cancer Metastasis ◽  
Resistance Mechanisms ◽  
Breast Cancer Metastasis ◽  
Breast Cancers ◽  
Stromal Fibroblasts

SummaryDisseminated tumour cells, particularly in ER+ breast cancers, typically exhibit a period of dormancy that renders them insensitive to targeting by chemotherapy. Additionally, chemotherapy treatment can result in normal tissue damage, including the induction of cellular senescence. Using mouse and human breast cancer models, we demonstrate that systemic chemotherapy administration results in accumulation of long-lived senescent stromal fibroblasts and promotes metastatic outgrowth. Chemotherapy-induced senescent fibroblasts upregulate a senescence associated secretory phenotype (SASP) that accelerates 3D tumour spheroid growth by stimulating mitogenic signalling. Senolytic drugs can effectively eliminate chemotherapy-induced senescent fibroblasts in vitro, but show only modest efficacy in vivo, at least in part due to the upregulation of resistance mechanisms. In conclusion, systemic chemotherapy can establish a productive microenvironment for colonisation and outgrowth of disseminated cancer cells, however, optimisation of senotherapies for effective targeting of senescent fibroblasts is required to establish them as useful additions to standard chemotherapy.

scholarly journals Transcriptional suppression of AMPKα1 promotes breast cancer metastasis upon oncogene activation

2020 ◽  
Vol 117 (14) ◽  
pp. 8013-8021 ◽  
Author(s):  
Yong Yi ◽  
Deshi Chen ◽  
Juan Ao ◽  
Wenhua Zhang ◽  
Jianqiao Yi ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Cell Adhesion ◽  
Cancer Metastasis ◽  
Kinase Activity ◽  
Critical Role ◽  
Breast Cancer Metastasis ◽  
Transcription Control ◽  
Cell Cell

AMP-activated protein kinase (AMPK) functions as an energy sensor and is pivotal in maintaining cellular metabolic homeostasis. Numerous studies have shown that down-regulation of AMPK kinase activity or protein stability not only lead to abnormality of metabolism but also contribute to tumor development. However, whether transcription regulation of AMPK plays a critical role in cancer metastasis remains unknown. In this study, we demonstrate that AMPKα1 expression is down-regulated in advanced human breast cancer and is associated with poor clinical outcomes. Transcription of AMPKα1 is inhibited on activation of PI3K and HER2 through ΔNp63α. Ablation of AMPKα1 expression or inhibition of AMPK kinase activity leads to disruption of E-cadherin-mediated cell–cell adhesion in vitro and increased tumor metastasis in vivo. Furthermore, restoration of AMPKα1 expression significantly rescues PI3K/HER2-induced disruption of cell–cell adhesion, cell invasion, and cancer metastasis. Together, these results demonstrate that the transcription control is another layer of AMPK regulation and suggest a critical role for AMPK in regulating cell–cell adhesion and cancer metastasis.

scholarly journals Insight in Adhesion Protein Sialylation and Microgravity Dependent Cell Adhesion—An Omics Network Approach

2020 ◽  
Vol 21 (5) ◽  
pp. 1749 ◽  
Author(s):  
Thomas J. Bauer ◽  
Erich Gombocz ◽  
Markus Wehland ◽  
Johann Bauer ◽  
Manfred Infanger ◽  
...  
Keyword(s):  
Cell Adhesion ◽  
Cancer Metastasis ◽  
Current Knowledge ◽  
Three Dimensional ◽  
Binding Activity ◽  
Omics Data ◽  
Adhesion Proteins ◽  
Adhesion Behavior

The adhesion behavior of human tissue cells changes in vitro, when gravity forces affecting these cells are modified. To understand the mechanisms underlying these changes, proteins involved in cell-cell or cell-extracellular matrix adhesion, their expression, accumulation, localization, and posttranslational modification (PTM) regarding changes during exposure to microgravity were investigated. As the sialylation of adhesion proteins is influencing cell adhesion on Earth in vitro and in vivo, we analyzed the sialylation of cell adhesion molecules detected by omics studies on cells, which change their adhesion behavior when exposed to microgravity. Using a knowledge graph created from experimental omics data and semantic searches across several reference databases, we studied the sialylation of adhesion proteins glycosylated at their extracellular domains with regards to its sensitivity to microgravity. This way, experimental omics data networked with the current knowledge about the binding of sialic acids to cell adhesion proteins, its regulation, and interactions in between those proteins provided insights into the mechanisms behind our experimental findings, suggesting that balancing the sialylation against the de-sialylation of the terminal ends of the adhesion proteins’ glycans influences their binding activity. This sheds light on the transition from two- to three-dimensional growth observed in microgravity, mirroring cell migration and cancer metastasis in vivo.

Recombinant Human Erythropoietin (rHuEPO) In Combination with Chemotherapy Increases Breast Cancer Metastasis In Pre-Clinical Mouse Models

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 3345-3345
Author(s):  
Anargyros Xenocostas ◽  
Benjamin D Hedley ◽  
Jenny E Chu ◽  
D. George Ormond ◽  
Michel Beausoleil ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Cell Lines ◽  
Research Funding ◽  
Breast Cancer Cells ◽  
Cancer Metastasis ◽  
Breast Cancer Metastasis ◽  
In Vivo Studies ◽  
Metastatic Process

Abstract Abstract 3345 Background: Erythropoietin (EPO) is a key regulator of erythropoiesis, and has been shown to stimulate growth, maintain viability, and promote differentiation of red blood cell precursors. The EPO receptor (EPO-R) is expressed by erythroid cells and by several non-hematopoietic cell types including various neoplastic cells. Erythropoiesis-stimulating agents (ESAs) are used clinically for the treatment of chemotherapy-induced anemia. The results of some recent randomized clinical trials have reported an increased incidence in adverse events and reduced survival in ESA-treated metastatic breast cancer patients receiving chemotherapy, potentially related to EPO-induced cancer progression. These results have raised concerns over ESA treatment in metastatic cancer patients. However, very little pre-clinical data is available regarding the impact of EPO on breast cancer metastasis. The goal of the current study was therefore to determine if EPO can influence the malignant behavior of breast cancer cells and/or influence the metastatic process. Methods: MDA-MB-468, MDA-MB-231, MDA-MB-435, and 4T-1 breast cancer cell lines were treated with recombinant human EPO (rHuEPO; 10 U/ml) or control media and screened for EPO-R mRNA expression levels by RT-PCR, and for EPO-R protein expression by Western blot and flow cytometry. MDA-MB-231 (231) and MDA-MB-435 (435) cell lines were used for functional assays in vitro and in vivo. Untreated or rHuEPO treated cells were grown in 2D and 3D in vitro systems (standard tissue culture plates and 0.6% soft agar, respectively) to determine if rHuEPO influenced growth. In vitro cell survival was also assessed in response to treatment with rHuEPO in the presence or absence of paclitaxel chemotherapy (10mg/ml), radiation (10G), or hypoxic conditions (1% O2). Following mammary fat pad injection, in vivo effects of rHuEPO (300U/kg) alone or in combination with paclitaxel treatment (10mg/kg) were assessed in mouse models of tumorigenicity and spontaneous metastasis. Results: Expression analysis of EPO-R mRNA and protein revealed a large variation in levels across different cell lines. The majority of cell lines did not express cell surface EPO-R by flow cytometry, although two cell lines (231 and 435) did show weak expression of EPO-R mRNA, with only the 231 cell line showing EPO-R expression by Western blot. In vitro, a small protective effect from rHuEPO on radiation-treated 435 cells was seen (p<0.05); however, rHuEPO treatment alone or combined with chemotherapy or hypoxia did not cause a significant increase in cell survival relative to untreated controls cells. In contrast, in vivo studies demonstrated that rHuEPO increased the incidence and burden of lung metastases in immunocompromised mice injected with 231 or 435 cells and treated with paclitaxel relative to mice treated with paclitaxel alone (p<0.05). Conclusions: The lack of an in vitro effect of rHuEPO highlights the importance of in vivo studies to delineate the effects of EPO on the metastatic process. Our novel findings demonstrate that rHuEPO can reduce the efficacy of chemotherapy in the metastatic setting in vivo, and in some cases enhance the inherent metastatic growth potential of human breast cancer cells. This work was supported by funding from the London Regional Cancer Program and Janssen Ortho Canada Disclosures: Xenocostas: Janssen Ortho: Consultancy, Honoraria, Research Funding. Allan:Janssen Ortho: Research Funding.

scholarly journals The quinoxaline di-N-oxide DCQ blocks breast cancer metastasis in vitro and in vivo by targeting the hypoxia inducible factor-1 pathway

2014 ◽  
Vol 13 (1) ◽  
pp. 12 ◽  
Author(s):  
Khaled Ghattass ◽  
Sally El-Sitt ◽  
Kazem Zibara ◽  
Saide Rayes ◽  
Makhluf J Haddadin ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Cancer Metastasis ◽  
Hypoxia Inducible Factor ◽  
Breast Cancer Metastasis ◽  
Hypoxia Inducible Factor 1

scholarly journals Mammalian Adenylyl Cyclase-associated Protein 1 (CAP1) Regulates Cofilin Function, the Actin Cytoskeleton, and Cell Adhesion

2013 ◽  
Vol 288 (29) ◽  
pp. 20966-20977 ◽  
Author(s):  
Haitao Zhang ◽  
Pooja Ghai ◽  
Huhehasi Wu ◽  
Changhui Wang ◽  
Jeffrey Field ◽  
...  
Keyword(s):  
Cell Adhesion ◽  
Actin Cytoskeleton ◽  
Hela Cells ◽  
Actin Polymerization ◽  
Actin Dynamics ◽  
Ras Signaling ◽  
Filamentous Actin ◽  
Camp Pathway

CAP (adenylyl cyclase-associated protein) was first identified in yeast as a protein that regulates both the actin cytoskeleton and the Ras/cAMP pathway. Although the role in Ras signaling does not extend beyond yeast, evidence supports that CAP regulates the actin cytoskeleton in all eukaryotes including mammals. In vitro actin polymerization assays show that both mammalian and yeast CAP homologues facilitate cofilin-driven actin filament turnover. We generated HeLa cells with stable CAP1 knockdown using RNA interference. Depletion of CAP1 led to larger cell size and remarkably developed lamellipodia as well as accumulation of filamentous actin (F-actin). Moreover, we found that CAP1 depletion also led to changes in cofilin phosphorylation and localization as well as activation of focal adhesion kinase (FAK) and enhanced cell spreading. CAP1 forms complexes with the adhesion molecules FAK and Talin, which likely underlie the cell adhesion phenotypes through inside-out activation of integrin signaling. CAP1-depleted HeLa cells also had substantially elevated cell motility as well as invasion through Matrigel. In summary, in addition to generating in vitro and in vivo evidence further establishing the role of mammalian CAP1 in actin dynamics, we identified a novel cellular function for CAP1 in regulating cell adhesion.

scholarly journals Perturbation of BRMS1 interactome reveals pathways that impact cell migration

2021 ◽  
Author(s):  
Rosalyn Zimmermann ◽  
Mihaela E. Sardiu ◽  
Christa A. Manton ◽  
Md. Sayem Miah ◽  
Charles A.S. Banks ◽  
...  
Keyword(s):  
Cell Migration ◽  
Protein Interactions ◽  
Cancer Metastasis ◽  
Breast Cancer Metastasis ◽  
Metastasis Suppressor ◽  
Protein Protein Interactions ◽  
Multiple Cancer ◽  
C Terminus ◽  
Metastasis Suppression

AbstractBreast Cancer Metastasis Suppressor 1 (BRMS1) expression has been associated with longer patient survival in multiple cancer types. Understanding BRMS1 at the protein level will provide insights into both mechanism of action and enhance potential therapeutic development. We previously mapped the C-terminus of BRMS1 as critical for metastasis suppression and hypothesized that critical protein interactions in this region will explain function. These studies indicate that phosphorylation status at S237 regulates BRMS1 interactions related to a variety of biological processes, phenotypes [cell cycle (e.g., CDKN2A), DNA repair (e.g., BRCA1)], and metastasis [(e.g., TCF2 and POLE2)]. Presence of the C-terminal site appears to be critical for BRMS1 directed metastasis suppression, as demonstrated by in vitro migration assays. These assays demonstrated that presence of S237 directly decreased MDA-MB-231 migration. This study furthers our understanding of BRMS1’s molecular role, as it demonstrates that BRMS1 C-terminus is involved in direct protein-protein interactions. Several of the interacting proteins are associated with cancer and metastasis, which may result in metastasis suppression as suggested by in vitro findings.Abstract FigureGraphical AbstractUtilizing BRMS1 mutants to mimic-phosphorylation, this study demonstrates that S237-phosphorylation disrupts BRMS1 protein-protein interactions. The disruption includes both known Sin3/HDAC interactors as well as additionally previously unidentified Sin3-indepedent binding partners (indicated by increased opacity). It is revealed that BRMS1-phosphorylation status also more greatly inhibits cell migration (indicated by +) compared to the unphosphorylated state, suggesting that phosphorylation plays a role in BRMS1 metastatsis suppresion function, potentially though altered protein interactions.

scholarly journals Daratumumab, a Novel Anti-CD38 Monoclonal Antibody Shows Anti-Tumor Activity in CLL and hampers Leukemia-Microenvironment Interactions

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 4680-4680 ◽  
Author(s):  
Alba Matas-Céspedes ◽  
Anna Vidal-Crespo ◽  
Vanina Rodriguez ◽  
Julio Delgado ◽  
Neus Villamor ◽  
...  
Keyword(s):  
Extracellular Matrix ◽  
Cell Migration ◽  
Cell Adhesion ◽  
Tumor Microenvironment ◽  
Primary Cells ◽  
Isotype Control ◽  
Mmp9 Expression ◽  
Anti Tumor Activity

Abstract Daratumumab (DARA) is a anti-human CD38 antibody with Fc-mediated cell killing activity. DARA induces killing of tumor cells, mainly via complement-dependent cytotoxicity (CDC), antibody-dependent cell-mediated cytotoxicity (ADCC) (de Weers M. J Immunol 2011), and antibody-dependent cellular phagocytosis (ADCP) by macrophages (mΦ), both murine and human in multiple myeloma (MM) and Burkitt lymphoma cells. DARA is currently being evaluated in phase III clinical trials in patients with MM. We have previously reported that DARA induces cytotoxic activity in vitro via ADCC in primary cells and cell lines from Chronic Lymphoctic Leukemia (CLL), and significantly prolongs overall survival of animals in a systemic CLL mouse model. Here, we present additional data on in vivo mechanism of DARA and its effect on tumor-microenvironment interactions in CLL. We first evaluated whether ADCP contributes to DARA activity both in vitro and in vivo. For in vitro ADCP, mΦ were generated from monocytes of normal PBMCs and stimulated with GM-CSF (10ng/mL, 7 days). CLL cell lines and primary cells were labeled with calcein and incubated for 4h with mΦ at an effector:target ratio of 2:1 in the presence of a fixed mAb concentration of 1 μg/mL, followed by flow cytometric analysis. The amount of remaining CLL target cells (CD19+, CD11b-) was reduced by 3-16%. ADCP defined as percentage of mφ which had phagocytosed, referred to as double positive mΦ (CD11b+, calcein+, CD19-), ranged from 3-10%. To analyze ADCP in vivo, SCID beige mice, devoid of NK cells but with active macrophages, were inoculated intraperitoneally with CLL cells (20×106) and simultaneously treated with a single dose of DARA or isotype control (20mg/kg, n=3-5 per group). Forty-eight hours later, CLL cells were recovered from the intraperitoneal cavity and counted in a flow cytometer (identified as human CD45+/CD19+/CD5+cells). In DARA-treated mice the number of CLL cells recovered was reduced by 42% (n=2, p<0.05) compared to the isotype control group. Remarkably, the decrease in cell number was already detectable 2h after DARA administration. CLL pathogenesis relies on supportive tumor-microenvironment interactions both in the bone marrow (BM) and in the lymph node (LN), and CD38 constitutes a molecular hub integrating proliferative and migratory signals for CLL (Malavasi, F. Blood 2011). We evaluated the effect of DARA on migration and adhesion. In in vitro migrations assays, we have demonstrated that DARA (10-30 μg/mL) inhibited CXCL12/SDF1α-mediated migration up to 70% (n=5). In addition, DARA reduced up to 55% (n=2) of downstream pERK activation, that peaked after 5min of CXCL12/SDF1α stimulation. We analyzed the effect of DARA on primary CLL cell migration from Peripheral Blood (PB) to BM and spleen in vivo, using NOD/SCID/gamma (NSG) null mice (lacking NK cells and effective macrophages). In this system, NSG mice were pretreated (day 0) with DARA, control IgG or anti-CXCR4 as positive control for inhibition of cell homing, prior to injection of fresh primary CLL cells (50×106 cells/per mice) on day 1. PB, BM and spleen cells were isolated on day 2 and CLL cells were identified by staining for human CD45/CD19/CD5 and counted using a flow cytometer. Cell counting showed that CLL cells mainly migrate to the spleen, and that DARA significantly reduced this migration (55% inhibition on average, p<0.05). In addition to migration, CD38 also plays a key role in cell adhesion through interaction with integrins (CD49d/CD29) and with extracellular matrix proteins. We analyzed the effect of DARA on the adhesion of CLL cells to the extracellular matrix vascular-cell adhesion molecule-1 (VCAM-1) mediated by CD49d/CD29. DARA reduced adhesion of CLL cells (n=4), to VCAM-1 by 46±13% (range 27-57) compared to isotype control. By RT-PCR we observed an up-regulation of MMP9 transcripts (average 2 fold, n=2), and DARA abrogated both constitutive MMP9 expression (90% reduction) and VCAM-derived (94% reduction) MMP9 expression. In summary, DARA shows a positive effect on ADCP-mediated anti-tumor activity on CLL cells both in vitro and in vivo. In addition DARA exhibits a strong effect on CLL cell migration and adhesion. Based on these data, we hypothesize that DARA may exert unique and substantial effects on CLL tumor cell growth and contributes to potent therapeutic efficacy in a clinical setting. Disclosures Doshi: Janssen R&D: Employment. Parren:Genmab: Employment, Equity Ownership. Lammerts van Bueren:Genmab : Employment. Pérez-Galán:Genmab: Research Funding.

scholarly journals Prostate tumor-induced stromal reprogramming generates Tenascin C that promotes prostate cancer metastasis through YAP/TAZ inhibition

2021 ◽  
Author(s):  
Sue-Hwa Lin ◽  
Yu-Chen Lee ◽  
Song-Chang Lin ◽  
Guoyu Yu ◽  
Ming Zhu ◽  
...  
Keyword(s):  
Prostate Cancer ◽  
Cell Migration ◽  
Cancer Metastasis ◽  
Neutralizing Antibody ◽  
Metastatic Potential ◽  
In Vivo Studies ◽  
Hybrid Cells ◽  
Tenascin C

Metastatic prostate cancer (PCa) in bone induces bone-forming lesions that enhance PCa progression. How tumor-induced bone formation enhances PCa progression is not known. We have previously shown that PCa-induced bone originates from endothelial cells (EC) that have undergone endothelial-to-osteoblast (EC-to-OSB) transition by tumor-secreted BMP4. Here, we show that EC-to-OSB transition leads to changes in the tumor microenvironment that increases the metastatic potential of PCa cells. We found that conditioned medium (CM) from EC-OSB hybrid cells increases the migration, invasion and survival of PC3-mm2 and C4-2B4 PCa cells. Quantitative mass spectrometry (iTRAQ) identified Tenascin C (TNC) as one of the major proteins secreted from EC-OSB hybrid cells. TNC expression in tumor-induced osteoblasts was confirmed by immunohistochemistry of MDA-PCa118b xenograft and human bone metastasis specimens. Mechanistically, BMP4 increases TNC expression in EC-OSB cells through the Smad1-Notch/Hey1 pathway. How TNC promotes PCa metastasis was next interrogated by in vitro and in vivo studies. In vitro studies showed that a TNC neutralizing antibody inhibits EC-OSB-CM-mediated PCa cell migration and survival. TNC knockdown decreased, while addition of recombinant TNC or TNC overexpression increased migration and anchorage-independent growth of PC3 or C4-2b cells. When injected orthotopically, PC3-mm2-shTNC clones decreased metastasis to bone, while C4-2b-TNC overexpressing cells increased metastasis to lymph nodes. TNC enhances PCa cell migration through α5β1 integrin-mediated YAP/TAZ inhibition. These studies elucidate that tumor-induced stromal reprogramming generates TNC that enhances PCa metastasis and suggest that TNC may be a target for PCa therapy.

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