scholarly journals Breast tumor-associated metalloproteases restrict reovirus oncolysis by cleaving the σ1 cell-attachment protein, and can be overcome by mutation of σ1

2019 ◽  
Author(s):  
Jason Fernandes ◽  
Francisca Cristi ◽  
Heather Eaton ◽  
Patricia Chen ◽  
Sarah Haeflinger ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Sialic Acid ◽  
Cancer Cells ◽  
Breast Tumor ◽  
Cell Attachment ◽  
T Antigen ◽  
Virus Binding ◽  
Attachment Protein ◽  
Tumor Microenvironments ◽  
Cell Attachment Protein

ABSTRACTReovirus is undergoing clinical testing as an oncolytic therapy for breast cancer. Given that reovirus naturally evolved to thrive in enteric environments, we sought to better understand how breast tumor microenvironments impinge on reovirus infection. Reovirus was treated with extracellular extracts generated from polyoma virus middle T-antigen-derived mouse breast tumors. Unexpectedly, these breast tumor extracellular extracts inactivated reovirus, reducing infectivity of reovirus particles by 100-fold. Mechanistically, inactivation was attributed to proteolytic cleavage of the viral cell attachment protein σ1, which diminished virus binding to sialic acid-low tumor cells. Among various specific protease class inhibitors and metal ions, EDTA and ZnCl2 effectively modulated σ1 cleavage, indicating that breast tumor-associated zinc-dependent metalloproteases are responsible for reovirus inactivation. Moreover, media from MCF7, MB468, MD-MB-231 and HS578T breast cancer cell lines recapitulated σ1 cleavage and reovirus inactivation, suggesting that inactivation of reovirus is shared among mouse and human breast cancers, and that breast cancer cells in by themselves can be a source of reovirus-inactivating proteases. Binding assays and quantification of sialic acid (SA) levels on a panel of cancer cells showed that truncated σ1 reduced virus binding to cells with low surface SA. To overcome this restriction, we generated a reovirus mutant with a mutation (T249I) in σ1 that prevents σ1 cleavage and inactivation by breast tumor-associated proteases. The mutant reovirus showed similar replication kinetics in tumorigenic cells, equivalent toxicity as wild-type reovirus in a severely compromised mouse model, and increased tumor titers. Overall, the data shows that tumor microenvironments have the potential to reduce infectivity of reovirus.SIGNIFICANCEWe demonstrate that metalloproteases in breast tumor microenvironments can inactivate reovirus. Our findings expose that tumor microenvironment proteases could have negative impact on proteinaceous cancer therapies such as reovirus, and that modification of such therapies to circumvent inactivation by tumor metalloproteases merits consideration.

scholarly journals Breast Tumor-Associated Metalloproteases Restrict Reovirus Oncolysis by Cleaving the σ1 Cell Attachment Protein and Can Be Overcome by Mutation of σ1

2019 ◽  
Vol 93 (22) ◽  
Author(s):  
Jason P. Fernandes ◽  
Francisca Cristi ◽  
Heather E. Eaton ◽  
Patricia Chen ◽  
Sarah Haeflinger ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Cancer Cells ◽  
Breast Tumor ◽  
Cell Attachment ◽  
T Antigen ◽  
Virus Binding ◽  
Attachment Protein ◽  
Tumor Microenvironments ◽  
Toxicity Equivalent ◽  
Cell Attachment Protein

ABSTRACT Reovirus is undergoing clinical testing as an oncolytic therapy for breast cancer. Given that reovirus naturally evolved to thrive in enteric environments, we sought to better understand how breast tumor microenvironments impinge on reovirus infection. Reovirus was treated with extracellular extracts generated from polyomavirus middle T-antigen-derived mouse breast tumors. Unexpectedly, these breast tumor extracellular extracts inactivated reovirus, reducing infectivity of reovirus particles by 100-fold. Mechanistically, inactivation was attributed to proteolytic cleavage of the viral cell attachment protein σ1, which diminished virus binding to sialic acid (SA)-low tumor cells. Among various specific protease class inhibitors and metal ions, EDTA and ZnCl2 effectively modulated σ1 cleavage, indicating that breast tumor-associated zinc-dependent metalloproteases are responsible for reovirus inactivation. Moreover, media from MCF7, MB468, MD-MB-231, and HS578T breast cancer cell lines recapitulated σ1 cleavage and reovirus inactivation, suggesting that inactivation of reovirus is shared among mouse and human breast cancers and that breast cancer cells by themselves can be a source of reovirus-inactivating proteases. Binding assays and quantification of SA levels on a panel of cancer cells showed that truncated σ1 reduced virus binding to cells with low surface SA. To overcome this restriction, we generated a reovirus mutant with a mutation (T249I) in σ1 that prevents σ1 cleavage and inactivation by breast tumor-associated proteases. The mutant reovirus showed similar replication kinetics in tumorigenic cells, toxicity equivalent to that of wild-type reovirus in a severely compromised mouse model, and increased tumor titers. Overall, the data show that tumor microenvironments have the potential to reduce infectivity of reovirus. IMPORTANCE We demonstrate that metalloproteases in breast tumor microenvironments can inactivate reovirus. Our findings expose that tumor microenvironment proteases could have a negative impact on proteinaceous cancer therapies, such as reovirus, and that modification of such therapies to circumvent inactivation by tumor metalloproteases merits consideration.

scholarly journals Reduction of Virion-Associated σ1 Fibers on Oncolytic Reovirus Variants Promotes Adaptation toward Tumorigenic Cells

2015 ◽  
Vol 89 (8) ◽  
pp. 4319-4334 ◽  
Author(s):  
Adil Mohamed ◽  
Carmit Teicher ◽  
Sarah Haefliger ◽  
Maya Shmulevitz
Keyword(s):  
Cancer Cells ◽  
Cell Attachment ◽  
Forward Genetics ◽  
Productive Infection ◽  
Enteric Infection ◽  
Wild Type ◽  
Attachment Protein ◽  
Oncolytic Therapy ◽  
Cell Attachment Protein

ABSTRACTWild-type mammalian orthoreovirus serotype 3 Dearing (T3wt) is nonpathogenic in humans but preferentially infects and kills cancer cells in culture and demonstrates promising antitumor activityin vivo. Using forward genetics, we previously isolated two variants of reovirus, T3v1 and T3v2, with increased infectivity toward a panel of cancer cell lines and improvedin vivooncolysis in a murine melanoma model relative to that of T3wt. Our current study explored how mutations in T3v1 and T3v2 promote infectivity. Reovirions contain trimers of σ1, the reovirus cell attachment protein, at icosahedral capsid vertices. Quantitative Western blot analysis showed that purified T3v1 and T3v2 virions had ∼2- and 4-fold-lower levels of σ1 fiber than did T3wt virions. Importantly, using RNA interference to reduce σ1 levels during T3wt production, we were able to generate wild-type reovirus with reduced levels of σ1 per virion. As σ1 levels were reduced, virion infectivity increased by 2- to 5-fold per cell-bound particle, demonstrating a causal relationship between virion σ1 levels and the infectivity of incoming virions. During infection of tumorigenic L929 cells, T3wt, T3v1, and T3v2 uncoated the outer capsid proteins σ3 and μ1C at similar rates. However, having started with fewer σ1 molecules, a complete loss of σ1 was achieved sooner for T3v1 and T3v2. Distinct from intracellular uncoating, chymotrypsin digestion, as a mimic of natural enteric infection, resulted in more rapid σ3 and μ1C removal, unique disassembly intermediates, and a rapid loss of infectivity for T3v1 and T3v2 compared to T3wt. Optimal infectivity toward natural versus therapeutic niches may therefore require distinct reovirus structures and σ1 levels.IMPORTANCEWild-type reovirus is currently in clinical trials as a potential cancer therapy. Our molecular studies on variants of reovirus with enhanced oncolytic activityin vitroandin vivonow show that distinct reovirus structures promote adaptation toward cancer cells and away from conditions that mimic natural routes of infection. Specifically, we found that reovirus particles with fewer molecules of the cell attachment protein σ1 became more infectious toward transformed cells. Reduced σ1 levels conferred a benefit to incoming particles only, resulting in an earlier depletion of σ1 and a higher probability of establishing productive infection. Conversely, reovirus variants with fewer σ1 molecules showed reduced stability and infectivity and distinct disassembly when exposed to conditions that mimic natural intestinal proteolysis. These findings support a model where the mode of infection dictates the precise optimum of reovirus structure and provide a molecular rationale for considering alternative reovirus structures during oncolytic therapy.

Protein σ1 is the reovirus cell attachment protein

Virology ◽  
1981 ◽  
Vol 108 (1) ◽  
pp. 156-163 ◽  
Author(s):  
Patrick W.K. Lee ◽  
Edward C. Hayes ◽  
Wolfgang K. Joklik
Keyword(s):  
Cell Attachment ◽  
Attachment Protein ◽  
Cell Attachment Protein

scholarly journals Evaluation of the Impact of Imprinted Polymer Particles on Morphology and Motility of Breast Cancer Cells by Using Digital Holographic Cytometry

2020 ◽  
Vol 10 (3) ◽  
pp. 750 ◽  
Author(s):  
Megha Patel ◽  
Marek Feith ◽  
Birgit Janicke ◽  
Kersti Alm ◽  
Zahra El-Schich
Keyword(s):  
Breast Cancer ◽  
Sialic Acid ◽  
Cancer Cells ◽  
Cell Lines ◽  
Cell Morphology ◽  
Molecularly Imprinted Polymers ◽  
Breast Cancer Cells ◽  
Molecularly Imprinted ◽  
Imprinted Polymers ◽  
Mcf 7

Breast cancer is the second most common cancer type worldwide and breast cancer metastasis accounts for the majority of breast cancer-related deaths. Tumour cells produce increased levels of sialic acid (SA) that terminates the monosaccharide on glycan chains of the glycosylated proteins. SA can contribute to cellular recognition, cancer invasiveness and increase the metastatic potential of cancer cells. SA-templated molecularly imprinted polymers (MIPs) have been proposed as promising reporters for specific targeting of cancer cells when deployed in nanoparticle format. The sialic acid-molecularly imprinted polymers (SA-MIPs), which use SA for the generation of binding sites through which the nanoparticles can target and stain breast cancer cells, opens new strategies for efficient diagnostic tools. This study aims at monitoring the effects of SA-MIPs on morphology and motility of the epithelial type MCF-7 and the highly metastatic MDAMB231 breast cancer cell lines, using digital holographic cytometry (DHC). DHC is a label-free technique that is used in cell morphology studies of e.g., cell volume, area and thickness as well as in motility studies. Here, we show that MCF-7 cells move slower than MDAMB231 cells. We also show that SA-MIPs have an effect on cell morphology, motility and viability of both cell lines. In conclusion, by using DH microscopy, we could detect SA-MIPs impact on different breast cancer cells regarding morphology and motility.

scholarly journals Estrogen receptor signaling regulates the expression of the breast tumor kinase in breast cancer cells

BMC Cancer ◽  
2019 ◽  
Vol 19 (1) ◽  
Author(s):  
Sayem Miah ◽  
Edward Bagu ◽  
Raghuveera Goel ◽  
Yetunde Ogunbolude ◽  
Chenlu Dai ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Estrogen Receptor ◽  
Cancer Cells ◽  
Breast Tumor ◽  
Breast Cancer Cells ◽  
Receptor Signaling ◽  
Estrogen Receptor Signaling ◽  
Breast Tumor Kinase

TOPOGRAPHY MEDIATED REGULATION OF HER-2 EXPRESSION IN BREAST CANCER CELLS

Nano LIFE ◽  
2012 ◽  
Vol 02 (03) ◽  
pp. 1241009 ◽  
Author(s):  
AMITA DAVEREY ◽  
AUSTIN C. MYTTY ◽  
SRIVATSAN KIDAMBI
Keyword(s):  
Breast Cancer ◽  
Cell Adhesion ◽  
Cancer Cells ◽  
Cancer Progression ◽  
Breast Tumor ◽  
Cancer Biology ◽  
Breast Cancer Cells ◽  
Molecular Mechanisms ◽  
Her 2 ◽  
Micro Topography

This article demonstrates that the surface micro-topography regulates the biology of breast cancer cells, including the expression of HER-2 gene and protein. The breast tumor microenvironment is made up of heterogenous mixture of pores, ridges and collagen fibers with well defined topographical features. Although, significant progress has been achieved towards elucidating the biochemical and molecular mechanisms that underlie breast cancer progression, quantitative characterization of the associated mechanical/topographical properties and their role in breast tumor progression remains largely unexplored. Therefore, the aim of this study is to investigate the effect of topography on the adhesion and biology of breast cancer cells in in vitro cultures. Polydimethylsiloxane (PDMS) surfaces containing different topographies were coated with polyelectrolyte multilayers (PEMs) to improve cell adhesion and maintain cell culture. HER-2 expressing breast cancer cells, BT-474 and SKBr3, were cultured on these PDMS surfaces. We demonstrate that micro-topography affects the cell adhesion and distribution depending on the topography on the PDMS surfaces. We also report for the first time that surface topography down-regulates the HER-2 gene transcription and protein expression in breast cancer cells when cultured on PDMS surfaces with micro-topographies compared to the tissue culture polystyrene surface (TCPS) control. Results from this study indicate that micro-topography modulates morphology of cells, their distribution and expression of HER-2 gene and protein in breast cancer cells. This study provides a novel platform for studying the role of native topography in the progression of breast cancer and has immense potential for understanding the breast cancer biology.

Garcinol sensitizes breast cancer cells to Taxol through the suppression of caspase-3/iPLA2and NF-κB/Twist1 signaling pathways in a mouse 4T1 breast tumor model

2017 ◽  
Vol 8 (3) ◽  
pp. 1067-1079 ◽  
Author(s):  
Shih-Hsin Tu ◽  
Yi-Shiou Chiou ◽  
Nagabhushanam Kalyanam ◽  
Chi-Tang Ho ◽  
Li-Ching Chen ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Cancer Cells ◽  
Signaling Pathways ◽  
Breast Tumor ◽  
Breast Cancer Cells ◽  
Caspase 3 ◽  
Tumor Model ◽  
Breast Tumor Model

Garcinol sensitizes breast cancer cells to Taxolviasynergistically inducing antimitotic effects and effectively repressing the activation of caspase-3/iPLA2and Taxol-promoted NF-κB/Twist1 signaling pathways.

BINDING OF CELL ATTACHMENT PROTEIN TO COLLAGEN: EFFECT OF CHEMICAL MODIFICATIONS

1978 ◽  
Vol 312 (1 Fibroblast Su) ◽  
pp. 436-438 ◽  
Author(s):  
Hynda K. Kleinman ◽  
Ermona B McGoodwin ◽  
George R. Martin ◽  
Robert J. Klebe
Keyword(s):  
Cell Attachment ◽  
Chemical Modifications ◽  
Attachment Protein ◽  
Cell Attachment Protein

Estrogen receptor β represses breast tumor growth and angiogenesis in vivo

2006 ◽  
Vol 24 (18_suppl) ◽  
pp. 10101-10101
Author(s):  
J. Hartman ◽  
K. Lindberg ◽  
J. Inzunza ◽  
J. Wan ◽  
A. Ström ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Tumor Growth ◽  
Cancer Cells ◽  
Breast Tumor ◽  
Breast Cancer Cells ◽  
Cancer Cell Growth ◽  
Breast Cancer Cell Growth ◽  
Angiogenic Effect

10101 Background: Estrogens are well known stimulators of breast cancer cell growth in vitro as well as in vivo. Two different estrogen receptors exist, namely estrogen receptor (ER) α and β. ERα mediates the proliferative effect of estrogen in breast cancer cells and we have earlier shown that ERβ inhibits cell-cycle progression in vitro. Estrogens are well known stimulators of in vivo breast cancer cell growth as well as angiogenesis, and the effect is mediated through ERα. The function of ERβ in this context is not well understood. Methods: We have used ERα-positive T47D breast cancer cells stably transfected with a Tet/Off regulated ERβ expression vector system. The ERβ-inducible tumor cells are studied in vitro as well as in vivo. Results: By transplanting ERβ-inducible breast cancer cells into SCID-mice, we show that ERβ inhibits tumor growth and reduces the volume of established tumors. Furthermore, we show by immunohistochemistry, that the number of blood microvessels in the tumor periphery is decreased by ERβ expression, counteracting the well-known pro-angiogenic effect of ERα. By Western blot analysis on tumor extracts, we show that the concentration of the important pro-angiogenic growth factors VEGF and bFGF, normally expressed by breast tumor cells, is decreased in the ERβ-expressing tumors compared to the normal tumors. To exclude that the observed anti-angiogenic effect is just a result of reduced tumor growth, we incubated Tet/Off regulated ERβ expressing cells in vitro, during non-hypoxic conditions. We found that the expression of ERβ leads to decreased expression of VEGF and PDGFβ at the mRNA and protein-levels. In transient transfection assays, we found estrogen-ERα mediated up regulation of VEGF, PDGFβ and bFGF-promoter activities in T47D cells, and these activities were all suppressed following co-transfection with an ERβ-expression vector. Conclusions: We conclude that ERβ inhibits growth factor expression at transcriptional level in breast cancer cells; taken together, our data indicates that ERβ inhibits growth and angiogenesis of tumors formed by T47D breast cancer cells. This makes ERβ an interesting therapeutic target in breast cancer and perhaps treatment with the newly designed ERβ-selective ligands might work as a new anti-proliferative and anti-angiogenic therapy. No significant financial relationships to disclose.

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