scholarly journals New monoclonal antibodies that recognize an unglycosylated, conserved, extracellular region of CD44 in vitro and in vivo, and can block tumorigenesis

PLoS ONE ◽  
2021 ◽  
Vol 16 (4) ◽  
pp. e0250175
Author(s):  
Daniel F. Lusche ◽  
Deborah J. Wessels ◽  
Ryan J. Reis ◽  
Cristopher C. Forrest ◽  
Alexis R. Thumann ◽  
...  
Keyword(s):  
Monoclonal Antibodies ◽  
Cancer Cell ◽  
Cell Types ◽  
Western Blots ◽  
Cd44 Isoforms ◽  
Cellular Processes ◽  
Transmembrane Glycoprotein ◽  
Extracellular Region

CD44 is a transmembrane glycoprotein that binds to hyaluronic acid, plays roles in a number of cellular processes and is expressed in a variety of cell types. It is up-regulated in stem cells and cancer. Anti-CD44 monoclonal antibodies affect cell motility and aggregation, and repress tumorigenesis and metastasis. Here we describe four new anti-CD44 monoclonal antibodies originating from B cells of a mouse injected with a plasmid expressing CD44 isoform 12. The four monoclonal antibodies bind to the terminal, extracellular, conserved domain of CD44 isoforms. Based on differences in western blot patterns of cancer cell lysates, the four anti-CD44 mAbs separated into three distinct categories that include P4G9, P3D2, and P3A7, and P3G4. Spot assay analysis with peptides generated inEscherichia colisupport the conclusion that the monoclonal antibodies recognize unglycosylated sequences in the N-terminal conserved region between amino acid 21–220, and analyses with a peptide generated in human embryonic kidney 293 cells, demonstrate that these monoclonal antibodies bind to these peptides only after deglycosylation. Western blots with lysates from three cancer cell lines demonstrate that several CD44 isoforms are unglycosylated in the anti-CD44 target regions. The potential utility of the monoclonal antibodies in blocking tumorigenesis was tested by co-injection of cells of the breast cancer-derived tumorigenic cell line MDA-MB-231 with the anti-CD44 monoclonal antibody P3D2 into the mammary fat pads of mice. All five control mice injected with MDA-MB-231 cells plus anti-IgG formed palpable tumors, while only one of the six test mice injected with MDA-MB-231 cells plus P3D2 formed a tiny tumor, while the remaining five were tumor-free, indicating that the four anti-CD44 mAbs may be useful therapeutically.

scholarly journals Mitogen Kinase Kinase (MKK7) controls cytokine production in vitro and in vivo in mice

2021 ◽  
Author(s):  
Amada D. Caliz ◽  
Hyung-Jin Yoo ◽  
Anastassiia Vertii ◽  
Cathy Tournier ◽  
Roger J. Davis ◽  
...  
Keyword(s):  
Cytokine Production ◽  
Cell Types ◽  
Minor Role ◽  
Cellular Processes ◽  
Mitogen Activated Protein ◽  
And Migration ◽  
Jnk Activation

Mitogen kinase kinase 4 (MKK4) and Mitogen kinase kinase 7 (MKK7) are members of the MAP2K family which can activate downstream mitogen-activated protein kinases (MAPKs). MKK4 has been implicated in the activation of both, c-Jun N-terminal Kinase (JNK) and p38 MAPK, whereas MKK7 only activates JNK in response to different stimuli. The stimuli as well as cell type determine the choice of MAP2K member that mediates the response. In a variety of cell types, the MKK7 contributes to the activation of downstream MAPKs, JNK, which is known to regulate essential cellular processes, such as cell death, differentiation, stress response, and cytokine secretion. Previous studies have implicated the role of MKK7 in stress signaling pathways and cytokine production. However, little is known about the degree to which MKK7 and MKK4 contributes to innate immune response in macrophages as well as during inflammation in vivo. To address this question and elucidate the role of MKK7 and MKK4 in macrophage and in vivo, we developed MKK7- and MKK4-deficient mouse models with tamoxifen-inducible Rosa26 CreERT. This study reports that MKK7 is required for JNK activation both in vitro and in vivo. Additionally, we demonstrated that MKK7 in macrophages is necessary for LPS induced cytokine production and migration which appears to be a major contributor to the inflammatory response in vivo. Whereas MKK4 plays a significant but minor role in cytokine production in vivo.

scholarly journals Variable major proteins of Borrellia hermsii.

1982 ◽  
Vol 156 (5) ◽  
pp. 1312-1324 ◽  
Author(s):  
A G Barbour ◽  
S L Tessier ◽  
H G Stoenner
Keyword(s):  
Molecular Weight ◽  
Monoclonal Antibodies ◽  
Antigenic Variation ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Major Protein ◽  
Relapsing Fever ◽  
Western Blots

Borrelia hermsii, a relapsing fever agent, manifests antigenic variation in vivo and in vitro. We studied three mouse-passaged serotypes of strain HS1 (7, 14, and 21) and a HS1 derivative obtained after multiple in vitro passages (C serotype). All four serotypes had two major proteins in whole cell lysates fractionated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. One major protein species (pII) had the same apparent subunit molecular weight (or approximately 3.9 X 10(4) in all the serotypes. In contrast, the other abundant protein in lysates, pI, had a different apparent molecular weight in each serotype. In one gel the molecular weights of pIc, pI7, pI14, and pI21 were 1.9, 4.2, 4.1, and 4.0 X 10(4), respectively. Serotype-specific mouse antisera bound to both hemologous and heterologous pIIs, to homologous pI, but not to heterologous pI in Western blots. Hybridomas were raised from spleens of mice infected with B. hermsii. Monoclonal antibodies were identified by immunofluorescence assays using whole organisms. Monoclonal antibodies specific for serotype 7 (H1826) or for serotype 21 (H3326) bound only to pI7 or pI21, respectively, in Western blots. The surface location of the pI was suggested not only by the immunofluorescence studies but also by the labeling of pI7 and pI21 when whole cells of serotypes 7 and 21 were incubated with 125I in the presence of Iodogen. Under the same circumstances, pII was relatively poorly labeled. These studies have identified the variable pI proteins of B. hermsii as serotype-specific antigens. A change from one pI to another may be the basis of antigenic variation of Borrelia species during relapsing fever.

scholarly journals Anticancer Effect of Fucoidan in Combination with Tyrosine Kinase Inhibitor Lapatinib

2014 ◽  
Vol 2014 ◽  
pp. 1-6 ◽  
Author(s):  
Byeongsang Oh ◽  
Jihun Kim ◽  
Weidong Lu ◽  
David Rosenthal
Keyword(s):  
Cancer Cells ◽  
Cancer Cell ◽  
Kinase Inhibitor ◽  
Cell Types ◽  
Anticancer Effect ◽  
Inhibit Cell Growth ◽  
Mechanism Of Interaction ◽  
Synergistic Antitumor Activity

Background. Despite a number ofin vitroandin vivostudies reporting the efficacy of fucoidan in treating various cancers, few studies have measured the efficacy of dietary fucoidan (DF) in combination with cancer drugs. Thus, we examined the sensitivity of DF in combination with the EGFR/ERBB2-targeting reagent lapatinib on cancer cells.Method. We selected six EGFR/ERBB2-amplified cancer cell lines (OE19, NCI-N87, OE33, ESO26, MKN7, and BT474) as anin vitromodel and tested their sensitivity to DF alone and to DF in combination with the well-known EGFR/ERBB2-targeting reagent lapatinib.Result. Overall, in drug independent sensitivity test, DF alone did not significantly inhibit the growth of EGFR/ERBB2-amplified cancer cellsin vitro. When DF was given in combination with lapatinib, however, it tended to synergistically inhibit cell growth in OE33 but antagonized the action of lapatinib in ESO26, NCI-N87, and OE19.Conclusion. This study suggests that DF has the potential to increase or decrease the effects of certain anticancer drugs on certain cancer cell types. Further study is needed to explore the mechanism of interaction and synergistic antitumor activity of DF in combination with chemotherapy and targeted therapy.

scholarly journals FiloQuant reveals increased filopodia density during DCIS progression

2017 ◽  
Author(s):  
Guillaume Jacquemet ◽  
Ilkka Paatero ◽  
Alexandre F. Carisey ◽  
Artur Padzik ◽  
Jordan S. Orange ◽  
...  
Keyword(s):  
Cancer Cell ◽  
Ductal Carcinoma ◽  
Biological Significance ◽  
Cell Types ◽  
Neuronal Cultures ◽  
Cellular Models ◽  
Tumour Spheroids ◽  
Microscopy Data

AbstractFilopodia are commonly observed cellular protrusions in vitro and in vivo. Defective filopodia formation is linked to several pathologies including cancer, wherein actively protruding filopodia, at the invasive front, and filopodia-mediated probing of the microenvironment accompanies cancer cell dissemination. Despite wide biological significance, delineating the function of these finger-like protrusions in more complex systems remains technically challenging, particularly hindered by lack of compatible methods to quantify filopodia properties. Here, we present FiloQuant, a freely available ImageJ plugin, to detect filopodia and filopodia-like protrusions in both fixed and live-cell microscopy data. We demonstrate that FiloQuant can extract quantifiable information including protrusion dynamics, density and length from multiple cell types and in a range of microenvironments, such as during collective or single cancer cell migration in 2D and 3D, in fixed neuronal cultures, in activated natural killer cells and in sprouting endothelial cells in vivo. In cellular models of breast ductal carcinoma in situ (DCIS) we reveal a link between filopodia formation at the cell-matrix interface, during collective invasion and in 3D tumour spheroids, with the previously reported local invasive potential of these breast cancer models in vivo. Finally, using intravital microscopy, we observed that tumour spheroids display prominent filopodia in vivo, supporting a potential role for these protrusions during tumorigenesis.

Local adipocyte cancer cell paracrine loop: can “sick fat” be more detrimental?

2015 ◽  
Vol 21 (1) ◽  
Author(s):  
Marie-Christine Rio ◽  
Nassim Dali-Youcef ◽  
Catherine Tomasetto
Keyword(s):  
Cancer Cell ◽  
Molecular Mechanisms ◽  
Tumor Stroma ◽  
Cell Types ◽  
Migration And Invasion ◽  
Molecular Alterations ◽  
Increased Risk ◽  
Human Carcinomas

AbstractThis review article focuses on the emerging role of tumor resident adipocytes. It provides in vitro and in vivo evidence that they are essential for cancer development/progression. In addition to systemic effects, their tumor-promoting impact is dependent on local functions, notably via a complex adipocyte cancer cell paracrine loop (ACCPL). Indeed, this event leads to dramatic phenotypic and/or functional modifications of both cell types as well as of the extracellular matrix. Adipocytes undergo delipidation leading to adipocytes/cancer-associated adipocytes/cancer-associated fibroblasts de-differentiation processes. In turn, cancer cell aggressiveness is exacerbated through increased proliferation, migration, and invasion properties. This is accompanied by intense tissue remodeling, conducting to the occurrence of the tumor stroma. The molecular pathways involved in ACCPL remain largely unknown. Nevertheless, several clues are starting to emerge. Moreover, obesity is currently a sign of increased risk and poor prognosis in human carcinomas. How adiposopathy might impact tumors and specifically the ACCPL is still under investigation. However, available experimental, epidemiological, and clinical data allow to draw some directions. Interestingly, there are numerous similarities between the ACCPL-induced and obesity-related molecular alterations. It might, therefore, be hypothesized that obesity provides a “constitutively active” local permissive environment for cancer cells. Improving our knowledge about ACCPL in both lean and obese patients remains a challenging task. Indeed, deciphering the cellular and molecular mechanisms behind ACCPL might provide new targets for improving diagnosis/prognosis and the design of innovative therapeutic strategies, and even, in case of obesity, for preventing cancer.

scholarly journals miR-375-NAT10 Axis Dysfunction Promotes Oral Cancer Development and Mediates Cdk7 Stabilizing and Cell Cycle Progression

2021 ◽  
Author(s):  
Chen ZOU ◽  
Xia LI ◽  
Haigang WEI ◽  
Siyuan WU ◽  
Jing SONG ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Cell Proliferation ◽  
Oral Cancer ◽  
Cancer Cell ◽  
In Vivo Studies ◽  
Cancer Tissue ◽  
Cancer Cell Proliferation ◽  
Western Blots

Abstract Background: Oral cancer is the most common cancer with poor prognosis and outcome for the patients due to the challenging diagnosis and limited treatment possibilities. However, the molecular underpinnings behind the malignant progression of oral cancer remain incompletely understood. Methods: The expression profiling of NAT10 and CDK7 in oral cancer patients were assessed by IHC, qPCR and western blots. ShRNA was used to silence gene expression. The biological function of NAT10 and CDK7 in cholangiocarcinoma was investigated using in vitro and in vivo studies including, transwell cell migration, plate cloning, CCK8, shRNA interference, western blots, flow cytometry and xenograft mouse model. The underlying molecular mechanism was determined by western blots and immunoprecipitation.Results: In this study, we demonstrated that deregulation of miR-375-NAT10 axis is among the most causes in inducing the acquisition of a tumorigenesis phenotype in oral cancer cells. NAT10 is abundant in oral cancer tissue. and its protein level is positively correlated with poor overall survival. Increased the level of NAT10 promotes oral cancer cell proliferation in vitro as well as xenograft tumorigenicity in vivo. Most importantly, NAT10 regulates cancer cell proliferation through stabilizing CDK7 thus regulating the cell cycle. NAT10 as an acetyltransferase is responsible for CDK7 acetylation at lysine 328 (K328Ac). Moreover, it was found that the expression of miR-375 is abnormally alleviated in oral cancer tissues. Bioinformatics analysis revealed a targeted complementary binding site between miR-375 and NAT10. Decreased expression of miR-375 promotes expression of NAT10.Conclusion: Our study showed that NAT10 plays a strong carcinogenic role in oral cancer tumorigenesis by acetylating CDK7 at K382 thus promotion stability. Moreover, NAT10 may serve as a target for miR-375. Therefore, targeting NAT10 may provide a new and effective therapeutic strategy to inhibit the tumorigenicity of oral cancer.

scholarly journals Hypoxia induces transcriptional and translational downregulation of the type I interferon (IFN) pathway in multiple cancer cell types

2019 ◽  
Author(s):  
Ana Miar ◽  
Esther Arnaiz ◽  
Esther Bridges ◽  
Shaunna Beedie ◽  
Adam P Cribbs ◽  
...  
Keyword(s):  
Cancer Cell ◽  
Type I Interferon ◽  
Regulation Of Gene Expression ◽  
Cell Types ◽  
Type I ◽  
Type I Ifn ◽  
Multiple Cancer ◽  
Independent Manner

AbstractHypoxia is a common phenomenon in solid tumours and is considered a hallmark of cancer. Increasing evidence shows that hypoxia promotes local immune suppression. Type I IFN is involved in supporting cytotoxic T lymphocytes by stimulating the maturation of dendritic cells (DCs) and enhancing their capacity to process and present antigens. However, there is little information about the relationship between hypoxia and the type I interferon (IFN) pathway, which comprises the sensing of double-stranded RNA and DNA (dsRNA/dsDNA), followed by IFNα/β secretion and transcription activation of IFN-stimulated genes (ISGs). The aims of this study were to determine both the effect and mechanisms of hypoxia on the I IFN pathway in breast cancer.There was a downregulation of the type I IFN pathway expression at mRNA and protein level in cancer cell lines under hypoxia in vitro and in vivo in xenografts. This pathway was suppressed at each level of signalling, from the dsRNA sensors (RIG-I, MDA5), the adaptor (MAVS), transcription factors (IRF3, IRF7, STAT1) and several ISGs (RIG-I, IRF7, STAT1, ADAR-p150). There was also lower IFN secretion under hypoxic conditions. HIF1 and HIF2 regulation of gene expression did not explain most of the effects. However, ATAC-Seq data revealed that in hypoxia peaks with STAT1 and IRF3 motifs had decreased accessibility.Thus hypoxia leads to an overall 50% downregulation of the type I IFN pathway due to repressed transcription and lower chromatin accessibility in a HIF1/2α-independent manner, which could contribute to immunosuppression in hypoxic tumours.

scholarly journals Mitogen Kinase Kinase (MKK7) Controls Cytokine Production In Vitro and In Vivo in Mice

2021 ◽  
Vol 22 (17) ◽  
pp. 9364
Author(s):  
Amada D. Caliz ◽  
Hyung-Jin Yoo ◽  
Anastassiia Vertii ◽  
Ana C. Dolan ◽  
Cathy Tournier ◽  
...  
Keyword(s):  
Cytokine Production ◽  
Cell Types ◽  
Minor Role ◽  
Cellular Processes ◽  
Mitogen Activated Protein ◽  
And Migration ◽  
Jnk Activation

Mitogen kinase kinase 4 (MKK4) and mitogen kinase kinase 7 (MKK7) are members of the MAP2K family that can activate downstream mitogen-activated protein kinases (MAPKs). MKK4 has been implicated in the activation of both c-Jun N-terminal kinase (JNK) and p38 MAPK, while MKK7 has been reported to activate only JNK in response to different stimuli. The stimuli, as well as the cell type determine which MAP2K member will mediate a given response. In various cell types, MKK7 contributes to the activation of downstream MAPKs, JNK, which is known to regulate essential cellular processes, such as cell death, differentiation, stress response, and cytokine secretion. Previous studies have also implicated the role of MKK7 in stress signaling pathways and cytokine production. However, little is known about the degree to which MKK4 and MKK7 contribute to innate immune responses in macrophages or during inflammation in vivo. To address this question and to elucidate the role of MKK4 and MKK7 in macrophage and in vivo, we developed MKK4- and MKK7-deficient mouse models with tamoxifen-inducible Rosa26 CreERT. This study reports that MKK7 is required for JNK activation both in vitro and in vivo. Additionally, we demonstrated that MKK7 in macrophages is necessary for lipopolysaccharide (LPS)-induced cytokine production, M1 polarization, and migration, which appear to be a major contributor to the inflammatory response in vivo. Conversely, MKK4 plays a significant, but minor role in cytokine production in vivo.

scholarly journals Ras Suppressor-1 (RSU1) in Cancer Cell Metastasis: A Tale of a Tumor Suppressor

2020 ◽  
Vol 21 (11) ◽  
pp. 4076
Author(s):  
Maria Louca ◽  
Triantafyllos Stylianopoulos ◽  
Vasiliki Gkretsi
Keyword(s):  
Tumor Suppressor ◽  
Cancer Cell ◽  
Tumor Suppressor Genes ◽  
Ras Oncogene ◽  
Cellular Processes ◽  
Cell Metastasis ◽  
Cancer Types

Cancer is a multifactorial disease responsible for millions of deaths worldwide. It has a strong genetic background, as mutations in oncogenes or tumor suppressor genes contribute to the initiation of cancer development. Integrin signaling as well as the signaling pathway of Ras oncogene, have been long implicated both in carcinogenesis and disease progression. Moreover, they have been involved in the promotion of metastasis, which accounts for the majority of cancer-related deaths. Ras Suppressor-1 (RSU1) was identified as a suppressor of Ras-induced transformation and was shown to localize to cell-extracellular matrix adhesions. Recent findings indicate that its expression is elevated in various cancer types, while its role in regulating metastasis-related cellular processes remains largely unknown. Interestingly, there is no in vivo work in the field to date, and thus, all relevant knowledge stems from in vitro studies. In this review, we summarize recent studies using breast, liver and brain cancer cell lines and highlight the role of RSU1 in regulating cancer cell invasion.

scholarly journals Hic-5 expression is a major indicator of cancer cell morphology, migration, and plasticity in three-dimensional matrices

2018 ◽  
Vol 29 (14) ◽  
pp. 1704-1717 ◽  
Author(s):  
Anushree C. Gulvady ◽  
Fatemeh Dubois ◽  
Nicholas O. Deakin ◽  
Gregory J. Goreczny ◽  
Christopher E. Turner
Keyword(s):  
Tumor Cell ◽  
Cell Lines ◽  
Cancer Cell ◽  
Ectopic Expression ◽  
Three Dimensional ◽  
Cell Types ◽  
Cell Phenotype ◽  
In Vitro Invasiveness

The focal adhesion proteins Hic-5 and paxillin have been previously identified as key regulators of MDA-MB-231 breast cancer cell migration and morphologic mesenchymal-amoeboid plasticity in three-dimensional (3D) extracellular matrices (ECMs). However, their respective roles in other cancer cell types have not been evaluated. Herein, utilizing 3D cell–derived matrices and fibronectin-coated one-dimensional substrates, we show that across a variety of cancer cell lines, the level of Hic-5 expression serves as the major indicator of the cells primary morphology, plasticity, and in vitro invasiveness. Domain mapping studies reveal sites critical to the functions of both Hic-5 and paxillin in regulating phenotype, while ectopic expression of Hic-5 in cell lines with low endogenous levels of the protein is sufficient to induce a Rac1-dependent mesenchymal phenotype and, in turn, increase amoeboid-mesenchymal plasticity and invasion. We show that the activity of vinculin, when coupled to the expression of Hic-5 is required for the mesenchymal morphology in the 3D ECM. Taken together, our results identify Hic-5 as a critical modulator of tumor cell phenotype that could be utilized in predicting tumor cell migratory and invasive behavior in vivo.

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