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1. A new scaffold-free tumoroid model provides a robust preclinical tool to investigate invasion and drug response in Renal Cell Carcinoma

Author

Irinka Séraudie, Catherine Pillet, Beatrice Cesana, Pauline Bazelle, Florian Jeanneret, Bertrand Evrard, Frédéric Chalmel, Assilah Bouzit, Christophe Battail, Jean-Alexandre Long, Jean Luc Descotes, Claude Cochet, and Odile Filhol

Subjects
Cytology, QH573-671
Abstract

Abstract Clear cell Renal Cell Carcinoma (ccRCC) is one of the most prevalent kidney cancers, which is often asymptomatic and thus discovered at a metastatic state (mRCC). mRCC are highly heterogeneous tumors composed of subclonal populations that lead to poor treatment response rate. Several recent works explored the potential of ccRCC tumoroids culture derived from patients. However, these models were produced following a scaffold-based method using collagen I or Matrigel that exhibit lot variability and whose complexity could induce treatment response modifications and phenotypic alterations. Following the observation that ccRCC tumoroids can create their own niche by secreting extracellular matrix components, we developed the first scaffold-free tumoroid model of ccRCC tumors. Tumoroids from mice as well as from human tumors were generated with high success rate (≥90%) using a magnetic suspension method and standard culture media. Immunofluorescence analysis revealed their self-organization capacities to maintain multiple tumor-resident cell types, including endothelial progenitor cells. Transcriptomic analysis showed the reproducibility of the method highlighting that the majority of gene expression patterns was conserved in tumoroids compared to their matching tumor tissue. Moreover, this model enables to evaluate drug effects and invasiveness of renal cancer cells in a 3D context, providing a robust preclinical tool for drug screening and biomarker assessment in line with alternative ex vivo methods like tumor tissue slice culture or in vivo xenograft models.

Published
2023
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2. Cancer selective cell death induction by a bivalent CK2 inhibitor targeting the ATP site and the allosteric αD pocket

Author

Alexandre Bancet, Rita Frem, Florian Jeanneret, Angélique Mularoni, Pauline Bazelle, Caroline Roelants, Jean-Guy Delcros, Jean-François Guichou, Catherine Pillet, Isabelle Coste, Toufic Renno, Christophe Battail, Claude Cochet, Thierry Lomberget, Odile Filhol, and Isabelle Krimm

Subjects
Molecular biology, Cell biology, Structural biology, Cancer, Science
Abstract

Summary: Although the involvement of protein kinase CK2 in cancer is well-documented, there is a need for selective CK2 inhibitors suitable for investigating CK2 specific roles in cancer-related biological pathways and further exploring its therapeutic potential. Here, we report the discovery of AB668, an outstanding selective inhibitor that binds CK2 through a bivalent mode, interacting both at the ATP site and an allosteric αD pocket unique to CK2. Using caspase activation assay, live-cell imaging, and transcriptomic analysis, we have compared the effects of this bivalent inhibitor to representative ATP-competitive inhibitors, CX-4945, and SGC-CK2-1. Our results show that in contrast to CX-4945 or SGC-CK2-1, AB668, by targeting the CK2 αD pocket, has a distinct mechanism of action regarding its anti-cancer activity, inducing apoptotic cell death in several cancer cell lines and stimulating distinct biological pathways in renal cell carcinoma.

Published
2024
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3. CK2β Regulates Hematopoietic Stem Cell Biology and Erythropoiesis

Author

Laura Quotti Tubi, Sara Canovas Nunes, Elisa Mandato, Marco Pizzi, Nicola Vitulo, Mirco D’Agnolo, Raffaella Colombatti, Maddalena Martella, Maria Paola Boaro, Elena Doriguzzi Breatta, Anna Fregnani, Zaira Spinello, Mitja Nabergoj, Odile Filhol, Brigitte Boldyreff, Mattia Albiero, Gian Paolo Fadini, Carmela Gurrieri, Fabrizio Vianello, Gianpietro Semenzato, Sabrina Manni, Livio Trentin, and Francesco Piazza

Subjects
Diseases of the blood and blood-forming organs, RC633-647.5
Abstract

The Ser-Thr kinase CK2 plays important roles in sustaining cell survival and resistance to stress and these functions are exploited by different types of blood tumors. Yet, the physiological involvement of CK2 in normal blood cell development is poorly known. Here, we discovered that the β regulatory subunit of CK2 is critical for normal hematopoiesis in the mouse. Fetal livers of conditional CK2β knockout embryos showed increased numbers of hematopoietic stem cells associated to a higher proliferation rate compared to control animals. Both hematopoietic stem and progenitor cells (HSPCs) displayed alterations in the expression of transcription factors involved in cell quiescence, self-renewal, and lineage commitment. HSPCs lacking CK2β were functionally impaired in supporting both in vitro and in vivo hematopoiesis as demonstrated by transplantation assays. Furthermore, KO mice developed anemia due to a reduced number of mature erythroid cells. This compartment was characterized by dysplasia, proliferative defects at early precursor stage, and apoptosis at late-stage erythroblasts. Erythroid cells exhibited a marked compromise of signaling cascades downstream of the cKit and erythropoietin receptor, with a defective activation of ERK/JNK, JAK/STAT5, and PI3K/AKT pathways and perturbations of several transcriptional programs as demonstrated by RNA-Seq analysis. Moreover, we unraveled an unforeseen molecular mechanism whereby CK2 sustains GATA1 stability and transcriptional proficiency. Thus, our work demonstrates new and crucial functions of CK2 in HSPC biology and in erythropoiesis.

Published
2023
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4. Intracellular Fate of Sub-Toxic Concentration of Functionalized Selenium Nanoparticles in Aggressive Prostate Cancer Cells

Author

Caroline Bissardon, Olivier Proux, Salvatore Andrea Gazze, Odile Filhol, Benoît Toubhans, Lucie Sauzéat, Sylvain Bouchet, Aled R. Lewis, Thierry Maffeis, Jean-Louis Hazemann, Sam Bayat, Peter Cloetens, R. Steven Conlan, Laurent Charlet, and Sylvain Bohic

Subjects
prostate cancer, selenium, nanoparticle-correlative synchrotron imaging, X-ray fluorescence, speciation, cytotoxicity, Chemistry, QD1-999
Abstract

Selenium 0 (Se0) is a powerful anti-proliferative agent in cancer research. We investigated the impact of sub-toxic concentrations of Se0 functionalized nanoparticles (SeNPs) on prostate cancer PC-3 cells and determined their intracellular localization and fate. An in-depth characterization of functionalized selenium nanoparticles composition is proposed to certify that no chemical bias relative to synthesis issues might have impacted the study. Selenium is an extremely diluted element in the biological environment and therefore requires high-performance techniques with a very low detection limit and high spatial resolution for intracellular imaging. This was explored with state-of-the-art techniques, but also with cryopreparation to preserve the chemical and structural integrity of the cells for spatially resolved and speciation techniques. Monodisperse solutions of SeNPs capped with bovine serum albumin (BSA) were shown to slow down the migration capacity of aggressive prostate cancer cells compared to polydisperse solutions of SeNPs capped with chitosan. BSA coating could prevent interactions between the reactive surface of the nanoparticles and the plasma membrane, mitigating the generation of reactive oxygen species. The intracellular localization showed interaction with mitochondria and also a localization in the lysosome-related organelle. The SeNPs-BSA localization in mitochondria constitute a possible explanation for our result showing a very significant dampening of the PC-3 cell proliferation capabilities. The purpose of the use of sublethal compound concentrations was to limit adverse effects resulting from high cell death to best evaluate some cellular changes and the fate of these SeNPs on PC-3. Our findings provide new insight to further study the various mechanisms of cytotoxicity of SeNPs.

Published
2023
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5. CK2β-regulated signaling controls B cell differentiation and function

Author

Laura Quotti Tubi, Elisa Mandato, Sara Canovas Nunes, Arash Arjomand, Fortunato Zaffino, Sabrina Manni, Alessandro Casellato, Paolo Macaccaro, Nicola Vitulo, Sara Zumerle, Odile Filhol, Brigitte Boldyreff, Christian W. Siebel, Antonella Viola, Giorgio Valle, Federica Mainoldi, Stefano Casola, Valeria Cancila, Alessandro Gulino, Claudio Tripodo, Marco Pizzi, Angelo Paolo Dei Tos, Livio Trentin, Gianpietro Semenzato, and Francesco Piazza

Subjects
B lymphocyte, B cell development, protein kinase CK2, marginal zone, germinal center, Diffuse large B cell lymphoma, Immunologic diseases. Allergy, RC581-607
Abstract

Serine-Threonine kinase CK2 supports malignant B-lymphocyte growth but its role in B-cell development and activation is largely unknown. Here, we describe the first B-cell specific knockout (KO) mouse model of the β regulatory subunit of CK2. CK2βKO mice present an increase in marginal zone (MZ) and a reduction in follicular B cells, suggesting a role for CK2 in the regulation of the B cell receptor (BCR) and NOTCH2 signaling pathways. Biochemical analyses demonstrate an increased activation of the NOTCH2 pathway in CK2βKO animals, which sustains MZ B-cell development. Transcriptomic analyses indicate alterations in biological processes involved in immune response and B-cell activation. Upon sheep red blood cells (SRBC) immunization CK2βKO mice exhibit enlarged germinal centers (GCs) but display a limited capacity to generate class-switched GC B cells and immunoglobulins. In vitro assays highlight that B cells lacking CK2β have an impaired signaling downstream of BCR, Toll-like receptor, CD40, and IL-4R all crucial for B-cell activation and antigen presenting efficiency. Somatic hypermutations analysis upon 4-Hydroxy-3-nitrophenylacetyl hapten conjugated to Chicken Gamma Globulin (NP-CGG) evidences a reduced NP-specific W33L mutation frequency in CK2βKO mice suggesting the importance of the β subunit in sustaining antibody affinity maturation. Lastly, since diffuse large B cell lymphoma (DLBCL) cells derive from GC or post-GC B cells and rely on CK2 for their survival, we sought to investigate the consequences of CK2 inhibition on B cell signaling in DLBCL cells. In line with the observations in our murine model, CK2 inactivation leads to signaling defects in pathways that are essential for malignant B-lymphocyte activation.

Published
2023
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7. CK2β Is a Gatekeeper of Focal Adhesions Regulating Cell Spreading

Author

Odile Filhol, Anne-Marie Hesse, Anne-Pascale Bouin, Corinne Albigès-Rizo, Florian Jeanneret, Christophe Battail, Delphine Pflieger, and Claude Cochet

Subjects
CK2β depletion, EMT, tyrosine-phosphorylated proteins, FAK1-Src-PAX1 signaling pathway, focal adhesions, epithelial cell spreading, Biology (General), QH301-705.5
Abstract

CK2 is a hetero-tetrameric serine/threonine protein kinase made up of two CK2α/αʹ catalytic subunits and two CK2β regulatory subunits. The free CK2α subunit and the tetrameric holoenzyme have distinct substrate specificity profiles, suggesting that the spatiotemporal organization of the individual CK2 subunits observed in living cells is crucial in the control of the many cellular processes that are governed by this pleiotropic kinase. Indeed, previous studies reported that the unbalanced expression of CK2 subunits is sufficient to drive epithelial to mesenchymal transition (EMT), a process involved in cancer invasion and metastasis. Moreover, sub-stoichiometric expression of CK2β compared to CK2α in a subset of breast cancer tumors was correlated with the induction of EMT markers and increased epithelial cell plasticity in breast carcinoma progression. Phenotypic changes of epithelial cells are often associated with the activation of phosphotyrosine signaling. Herein, using phosphotyrosine enrichment coupled with affinity capture and proteomic analysis, we show that decreased expression of CK2β in MCF10A mammary epithelial cells triggers the phosphorylation of a number of proteins on tyrosine residues and promotes the striking activation of the FAK1-Src-PAX1 signaling pathway. Moreover, morphometric analyses also reveal that CK2β loss increases the number and the spatial distribution of focal adhesion signaling complexes that coordinate the adhesive and migratory processes. Together, our findings allow positioning CK2β as a gatekeeper for cell spreading by restraining focal adhesion formation and invasion of mammary epithelial cells.

Published
2022
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8. A RUNX2 stabilization pathway mediates physiologic and pathologic bone formation

Author

Jung-Min Kim, Yeon-Suk Yang, Kwang Hwan Park, Xianpeng Ge, Ren Xu, Na Li, Minkyung Song, Hyunho Chun, Seoyeon Bok, Julia F. Charles, Odile Filhol-Cochet, Brigitte Boldyreff, Teresa Dinter, Paul B. Yu, Ning Kon, Wei Gu, Takeshi Takarada, Matthew B. Greenblatt, and Jae-Hyuck Shim

Subjects
Science
Abstract

Runx2 is essential for tuning the generation of bone from skeletal stem cells (SSCs). Here, the authors demonstrate that the CK2/HAUSP pathway stabilizes RUNX2 protein thereby regulating the commitment of SSCs to osteoprogenitors as well as their subsequent maturation, and that inhibition of this pathway can block heterotopic ossification.

Published
2020
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9. Characterization of RAGE and CK2 Expressions in Human Fetal Membranes

Author

Karen Coste, Shaam Bruet, Caroline Chollat-Namy, Odile Filhol, Claude Cochet, Denis Gallot, Geoffroy Marceau, Loïc Blanchon, Vincent Sapin, and Corinne Belville

Subjects
fetal membranes, amniocyte, RAGE, CK2, rupture, labor, Biology (General), QH301-705.5, Chemistry, QD1-999
Abstract

At the feto-maternal interface, fetal membranes (FM) play a crucial role throughout pregnancy. FM rupture at term implicates different sterile inflammation mechanisms including pathways activated by the transmembrane glycoprotein receptor for advanced glycation end-products (RAGE) belonging to the immunoglobulin superfamily. As the protein kinase CK2 is also implicated in the inflammation process, we aimed to characterize the expressions of RAGE and the protein kinase CK2 as a candidate regulator of RAGE expression. The amnion and choriodecidua were collected from FM explants and/or primary amniotic epithelial cells throughout pregnancy and at term in spontaneous labor (TIL) or term without labor (TNL). The mRNA and protein expressions of RAGE and the CK2α, CK2α′, and CK2β subunits were investigated using reverse transcription quantitative polymerase chain reaction and Western blot assays. Their cellular localizations were determined with microscopic analyses, and the CK2 activity level was measured. RAGE and the CK2α, CK2α′, and CK2β subunits were expressed in both FM layers throughout pregnancy. At term, RAGE was overexpressed in the amnion from the TNL samples, whereas the CK2 subunits were expressed at the same level in the different groups (amnion/choriodecidua/amniocytes, TIL/TNL), without modification of the CK2 activity level and immunolocalization. This work paves the way for future experiments regarding the regulation of RAGE expression by CK2 phosphorylation.

Published
2023
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10. In Search of Small Molecule Inhibitors Targeting the Flexible CK2 Subunit Interface

Author

Benoît Bestgen, Zakia Belaid-Choucair, Thierry Lomberget, Marc Le Borgne, Odile Filhol, and Claude Cochet

Subjects
protein kinase CK2, subunit interface, cyclic peptides, protein–protein interaction, cell death, Medicine, Pharmacy and materia medica, RS1-441
Abstract

Protein kinase CK2 is a tetrameric holoenzyme composed of two catalytic (α and/or α’) subunits and two regulatory (β) subunits. Crystallographic data paired with fluorescence imaging techniques have suggested that the formation of the CK2 holoenzyme complex within cells is a dynamic process. Although the monomeric CK2α subunit is endowed with a constitutive catalytic activity, many of the plethora of CK2 substrates are exclusively phosphorylated by the CK2 holoenzyme. This means that the spatial and high affinity interaction between CK2α and CK2β subunits is critically important and that its disruption may provide a powerful and selective way to block the phosphorylation of substrates requiring the presence of CK2β. In search of compounds inhibiting this critical protein–protein interaction, we previously designed an active cyclic peptide (Pc) derived from the CK2β carboxy-terminal domain that can efficiently antagonize the CK2 subunit interaction. To understand the functional significance of this interaction, we generated cell-permeable versions of Pc, exploring its molecular mechanisms of action and the perturbations of the signaling pathways that it induces in intact cells. The identification of small molecules inhibitors of this critical interaction may represent the first-choice approach to manipulate CK2 in an unconventional way.

Published
2017
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11. Making Hybrids of Two-Hybrid Systems

Author

Marie-Claire Dagher and Odile Filhol-Cochet

Subjects
Biology (General), QH301-705.5
Abstract

Two-hybrid systems are powerful tools to find new partners for a protein of interest. However, exchange of material between two-hybrid users has been handicapped by the various versions of two-hybrid systems available and by the widely accepted idea that they are not compatible. In the present paper, we show that, contrary to the dogma, the most often used two-hybrid systems may be combined by either transformation or mating assays. The protocol to be followed in each case is provided. This will greatly increase the prospects of the growing network of interacting proteins, by reconciling the “two-hybrid systems” and the “interaction trap”.

Published
1997
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12. DSIR: assessing the design of highly potent siRNA by testing a set of cancer-relevant target genes.

Author

Odile Filhol, Delphine Ciais, Christian Lajaunie, Peggy Charbonnier, Nicolas Foveau, Jean-Philippe Vert, and Yves Vandenbrouck

Subjects
Medicine, Science
Abstract

Chemically synthesized small interfering RNA (siRNA) is a widespread molecular tool used to knock down genes in mammalian cells. However, designing potent siRNA remains challenging. Among tools predicting siRNA efficacy, very few have been validated on endogenous targets in realistic experimental conditions. We previously described a tool to assist efficient siRNA design (DSIR, Designer of siRNA), which focuses on intrinsic features of the siRNA sequence. Here, we evaluated DSIR's performance by systematically investigating the potency of the siRNA it designs to target ten cancer-related genes. mRNA knockdown was measured by quantitative RT-PCR in cell-based assays, revealing that over 60% of siRNA sequences designed by DSIR silenced their target genes by at least 70%. Silencing efficacy was sustained even when low siRNA concentrations were used. This systematic analysis revealed in particular that, for a subset of genes, the efficiency of siRNA constructs significantly increases when the sequence is located closer to the 5'-end of the target gene coding sequence, suggesting the distance to the 5'-end as a new feature for siRNA potency prediction. A new version of DSIR incorporating these new findings, as well as the list of validated siRNA against the tested cancer genes, has been made available on the web (http://biodev.extra.cea.fr/DSIR).

Published
2012
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13. A new scaffold-free tumoroid model provides a robust preclinical tool to investigate invasion and drug response in Renal Cell Carcinoma

Author

Odile Filhol, Irinka Séraudie, Catherine Pillet, Beatrice Cesana, Pauline Bazelle, Florian Jeanneret, Bertrand Evrard, Frederic Chalmel, Assilah Bouzit, Jean-Alexandre Long, Jean-Luc Descotes, Claude Cochet, and Christophe Battail

Abstract

Clear cell Renal Cell Carcinoma (ccRCC) is one of the most prevalent kidney cancers, which is often asymptomatic and thus discovered at a metastatic state (mRCC). mRCC are highly heterogeneous tumors composed of subclonal populations that lead to poor treatment response rate. Several recent works explored the potential of ccRCC tumoroids culture derived from patients. However, these models were produced following a scaffold-based method using collagen I or Matrigel that exhibit lot variability and whose complexity could induce treatment response modifications and phenotypic alterations. Following the observation that ccRCC tumoroids can create their own niche by secreting extracellular matrix components, we developed the first scaffold-free tumoroid model of ccRCC tumors. Tumoroids from mice as well as from human tumors were generated with high success rate (≥90 %) using a magnetic suspension method and standard culture media. Immunofluorescence analysis revealed their self-organization capacities to maintain multiple tumor-resident cell types, including endothelial progenitor cells. Transcriptomic analysis showed the reproducibility of the method highlighting that the majority of gene expression patternswas conserved in tumoroids compared to their matching tumor tissue. Moreover, this model enables to evaluate drug effects and invasiveness of renal cancer cells in a 3D context, providing a robust preclinical tool for drug screening and biomarker assessment in line with alternative ex vivo methods like tumor tissue slice culture or in vivoxenograft models.

Published
2023

14. Supplementary Figure 2 from Pharmacological Inhibition of LIM Kinase Stabilizes Microtubules and Inhibits Neoplastic Growth

Author

Laurence Lafanechère, Ora Bernard, Stefan Knapp, Jean-Claude Florent, Diane Braguer, Charles Dumontet, Attilio Di Pietro, Rong Li, Juliana Antonipillai, David Grierson, Samia Aci-Sèche, Stéphane Honoré, Glaucio Valdameri, Anne Beghin, Odile Filhol, Emmanuelle Soleilhac, Caroline Barette, Chloé Prunier, Anne Martinez, Catherine Pillet, Chi-Hung Nguyen, Emilie Vassal-Stermann, and Renaud Prudent

Abstract

PDF file - 486K, Analysis of the consequence of actin filaments depolymerization on Pyr1 -induced formation of Detyr-microtubules

Published
2023

15. Supplementary Figure 1 from Pharmacological Inhibition of LIM Kinase Stabilizes Microtubules and Inhibits Neoplastic Growth

Author

Laurence Lafanechère, Ora Bernard, Stefan Knapp, Jean-Claude Florent, Diane Braguer, Charles Dumontet, Attilio Di Pietro, Rong Li, Juliana Antonipillai, David Grierson, Samia Aci-Sèche, Stéphane Honoré, Glaucio Valdameri, Anne Beghin, Odile Filhol, Emmanuelle Soleilhac, Caroline Barette, Chloé Prunier, Anne Martinez, Catherine Pillet, Chi-Hung Nguyen, Emilie Vassal-Stermann, and Renaud Prudent

Abstract

PDF file - 212K, Effect of Pyr1 treatment on the number and on the size of EB1 comets in HeLa cells

Published
2023

16. Supplementary Figure 3 from Pharmacological Inhibition of LIM Kinase Stabilizes Microtubules and Inhibits Neoplastic Growth

Author

Laurence Lafanechère, Ora Bernard, Stefan Knapp, Jean-Claude Florent, Diane Braguer, Charles Dumontet, Attilio Di Pietro, Rong Li, Juliana Antonipillai, David Grierson, Samia Aci-Sèche, Stéphane Honoré, Glaucio Valdameri, Anne Beghin, Odile Filhol, Emmanuelle Soleilhac, Caroline Barette, Chloé Prunier, Anne Martinez, Catherine Pillet, Chi-Hung Nguyen, Emilie Vassal-Stermann, and Renaud Prudent

Abstract

PDF file - 173K, Characterization of Pyr1 (Pyr1) effects on the inhibition of LIMKs activity in vitro

Published
2023

17. Supplementary Legends for Figures and Tables, Methods from Antitumor Activity of Pyridocarbazole and Benzopyridoindole Derivatives that Inhibit Protein Kinase CK2

Author

Claude Cochet, Jean-Baptiste Reiser, Odile Filhol, Elodie Spreux, Eve Duchemin-Pelletier, Céline F Sautel, Laurence Lafanechère, Jean-Claude Florent, Frédéric Schmidt, Caroline Barette, Chi-Hung Nguyen, Virginie Moucadel, and Renaud Prudent

Abstract

Supplementary Legends for Figures and Tables, Methods from Antitumor Activity of Pyridocarbazole and Benzopyridoindole Derivatives that Inhibit Protein Kinase CK2

Published
2023

18. Supplementary Figure 5 from Pharmacological Inhibition of LIM Kinase Stabilizes Microtubules and Inhibits Neoplastic Growth

Author

Laurence Lafanechère, Ora Bernard, Stefan Knapp, Jean-Claude Florent, Diane Braguer, Charles Dumontet, Attilio Di Pietro, Rong Li, Juliana Antonipillai, David Grierson, Samia Aci-Sèche, Stéphane Honoré, Glaucio Valdameri, Anne Beghin, Odile Filhol, Emmanuelle Soleilhac, Caroline Barette, Chloé Prunier, Anne Martinez, Catherine Pillet, Chi-Hung Nguyen, Emilie Vassal-Stermann, and Renaud Prudent

Abstract

PDF file - 487K, Effects of Pyr1 structural analogues on F-actin organization, on Detyr-microtubules generation, and on in cellulo cofilin phosphorylation

Published
2023

19. Supplementary Figures 1-3 from Antitumor Activity of Pyridocarbazole and Benzopyridoindole Derivatives that Inhibit Protein Kinase CK2

Author

Claude Cochet, Jean-Baptiste Reiser, Odile Filhol, Elodie Spreux, Eve Duchemin-Pelletier, Céline F Sautel, Laurence Lafanechère, Jean-Claude Florent, Frédéric Schmidt, Caroline Barette, Chi-Hung Nguyen, Virginie Moucadel, and Renaud Prudent

Abstract

Supplementary Figures 1-3 from Antitumor Activity of Pyridocarbazole and Benzopyridoindole Derivatives that Inhibit Protein Kinase CK2

Published
2023

20. Supplementary Figure 4 from Pharmacological Inhibition of LIM Kinase Stabilizes Microtubules and Inhibits Neoplastic Growth

Author

Laurence Lafanechère, Ora Bernard, Stefan Knapp, Jean-Claude Florent, Diane Braguer, Charles Dumontet, Attilio Di Pietro, Rong Li, Juliana Antonipillai, David Grierson, Samia Aci-Sèche, Stéphane Honoré, Glaucio Valdameri, Anne Beghin, Odile Filhol, Emmanuelle Soleilhac, Caroline Barette, Chloé Prunier, Anne Martinez, Catherine Pillet, Chi-Hung Nguyen, Emilie Vassal-Stermann, and Renaud Prudent

Abstract

PDF file - 252K, Effect of Pyr1 on NEK11 and MLK1 activity

Published
2023

21. Supplementary Tables 1-9, Movie Legend, Methods from Pharmacological Inhibition of LIM Kinase Stabilizes Microtubules and Inhibits Neoplastic Growth

Author

Laurence Lafanechère, Ora Bernard, Stefan Knapp, Jean-Claude Florent, Diane Braguer, Charles Dumontet, Attilio Di Pietro, Rong Li, Juliana Antonipillai, David Grierson, Samia Aci-Sèche, Stéphane Honoré, Glaucio Valdameri, Anne Beghin, Odile Filhol, Emmanuelle Soleilhac, Caroline Barette, Chloé Prunier, Anne Martinez, Catherine Pillet, Chi-Hung Nguyen, Emilie Vassal-Stermann, and Renaud Prudent

Abstract

PDF file - 253K

Published
2023

22. Data from Antitumor Activity of Pyridocarbazole and Benzopyridoindole Derivatives that Inhibit Protein Kinase CK2

Author

Claude Cochet, Jean-Baptiste Reiser, Odile Filhol, Elodie Spreux, Eve Duchemin-Pelletier, Céline F Sautel, Laurence Lafanechère, Jean-Claude Florent, Frédéric Schmidt, Caroline Barette, Chi-Hung Nguyen, Virginie Moucadel, and Renaud Prudent

Abstract

The alkyloid compound ellipticine derived from the berrywood tree is a topoisomerase II poison that is used in ovarian and breast cancer treatment. In this study, we report the identification of ellipticine derivatives and their tetracyclic angular benzopyridoindole analogues as novel ATP-competitive inhibitors of the protein kinase CK2. In vitro and in vivo assays showed that these compounds have a good pharmacologic profile, causing a marked inhibition of CK2 activity associated with cell cycle arrest and apoptosis in human cancer cells. Further, in vivo assays demonstrate antitumor activity in a mouse xenograft model of human glioblastoma. Finally, crystal structures of CK2–inhibitor complex provide structural insights on the molecular basis of CK2 inhibition. Our work lays the foundation for development of clinically useful CK2 inhibitors derived from a well-studied scaffold with suitable pharmacokinetics parameters. Cancer Res; 70(23); 9865–74. ©2010 AACR.

Published
2023

23. Supplementary Video 1 from Pharmacological Inhibition of LIM Kinase Stabilizes Microtubules and Inhibits Neoplastic Growth

Author

Laurence Lafanechère, Ora Bernard, Stefan Knapp, Jean-Claude Florent, Diane Braguer, Charles Dumontet, Attilio Di Pietro, Rong Li, Juliana Antonipillai, David Grierson, Samia Aci-Sèche, Stéphane Honoré, Glaucio Valdameri, Anne Beghin, Odile Filhol, Emmanuelle Soleilhac, Caroline Barette, Chloé Prunier, Anne Martinez, Catherine Pillet, Chi-Hung Nguyen, Emilie Vassal-Stermann, and Renaud Prudent

Abstract

MOV file - 6MB, Effect of Pyr1 on microtubule dynamic instability - Time-lapse microscopy after 2 hours of incubation, on GFP-EB3 HeLa cells treated with DMSO (control), 5 muM, 10 muM or 25 muM Pyr1, as indicated. 4.Supplementary information

Published
2023

24. Supplementary Figure 6 from Pharmacological Inhibition of LIM Kinase Stabilizes Microtubules and Inhibits Neoplastic Growth

Author

Laurence Lafanechère, Ora Bernard, Stefan Knapp, Jean-Claude Florent, Diane Braguer, Charles Dumontet, Attilio Di Pietro, Rong Li, Juliana Antonipillai, David Grierson, Samia Aci-Sèche, Stéphane Honoré, Glaucio Valdameri, Anne Beghin, Odile Filhol, Emmanuelle Soleilhac, Caroline Barette, Chloé Prunier, Anne Martinez, Catherine Pillet, Chi-Hung Nguyen, Emilie Vassal-Stermann, and Renaud Prudent

Abstract

PDF file - 173K, Effect of LIMK down regulation on Detyr-tubulin levels

Published
2023

25. Regulation of sclerostin by the SIRT1 stabilization pathway in osteocytes

Author

Jung-Min Kim, Yeon-Suk Yang, Jun Xie, Oksun Lee, JiHea Kim, Jaehyoung Hong, Brigitte Boldyreff, Odile Filhol, Hyonho Chun, Matthew B. Greenblatt, Guangping Gao, and Jae-Hyuck Shim

Subjects
Mice, Osteoblasts, Sirtuin 1, Osteogenesis, Animals, Cell Biology, Osteocytes, Molecular Biology, Adaptor Proteins, Signal Transducing
Abstract

Osteocytes play a critical role in bone remodeling through the secretion of paracrine factors regulating the differentiation and activity of osteoblasts and osteoclasts. Sclerostin is a key osteocyte-derived factor that suppresses bone formation and promotes bone resorption, therefore regulators of sclerostin secretion are a likely source of new therapeutic strategies for treatment of skeletal disorders. Here, we demonstrate that protein kinase CK2 (casein kinase 2) controls sclerostin expression in osteocytes via the deubiquitinase ubiquitin-specific peptidase 4 (USP4)-mediated stabilization of Sirtuin1 (SIRT1). Deletion of CK2 regulatory subunit, Csnk2b, in osteocytes (Csnk2b

Published
2022

26. AB668, a novel highly selective protein kinase CK2 inhibitor with a distinct anti-tumor mechanism as compared to CX-4945 and SGC-CK2-1

Author

Alexandre Bancet, Rita Frem, Florian Jeanneret, Angélique Mularoni, Pauline Bazelle, Caroline Roelants, Jean-Guy Delcros, Jean-François Guichou, Catherine Pillet, Isabelle Coste, Toufic Renno, Christophe Battail, Claude Cochet, Thierry Lomberget, Odile Filhol, and Isabelle Krimm

Abstract

Although the involvement of protein kinase CK2 in cancer is well-documented, there is a need for selective CK2 inhibitors suitable for investigating CK2 specific roles in cancer-related biological pathways and further explore its therapeutic potential. Here we have discovered AB668, a new bivalent inhibitor that binds both at the ATP site and an allosteric αD pocket unique to CK2. The molecule inhibits CK2 activity with an outstanding selectivity over other kinases. Using caspase activation assay, live-cell imaging and transcriptomic analysis, we have compared the effects of this bivalent inhibitor to the non-selective ATP-competitive inhibitor CX-4945 that reached clinic and to the selective ATP-competitive SGC-CK2-1 molecule. Our results show that in contrast to CX-4945 or SGC-CK2-1, AB668 has a distinct mechanism of action regarding its anti-cancer activity, inducing apoptotic cell death and stimulating distinct biological pathways in several cancer cell lines while sparing healthy cells. Our data suggest that targeting a cryptic CK2 αD pocket validates an allosteric approach to targeting CK2 and provides a starting point for creating drug-like CK2 inhibitors for aggressive cancers.

Published
2022

28. Editorial

Author

Valérian, Dormoy, Odile, Filhol, Carole, Sourbier, and Thierry, Massfelder

Published
2022

29. NLRP7 Promotes Choriocarcinoma Growth and Progression through the Establishment of an Immunosuppressive Microenvironment

Author

Padma Murthi, Roland Abi Nahed, Deborah Reynaud, Vincent Sapin, Nadia Alfaidy, Christophe Battail, Mohamed Benharouga, Nicolas Lemaitre, Touria Aboussaouira, Pierre-Adrien Bolze, Wael Traboulsi, Rima Slim, Odile Filhol, Pascale Hoffmann, Houssine Boufettal, Frederic Sergent, BioSanté (UMR BioSanté), Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes (UGA), CEA- Saclay (CEA), Commissariat à l'énergie atomique et aux énergies alternatives (CEA), Physiologie cellulaire et végétale (LPCV), Centre National de la Recherche Scientifique (CNRS)-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE)-Université Grenoble Alpes (UGA), Centre Hospitalier Lyon Sud [CHU - HCL] (CHLS), Hospices Civils de Lyon (HCL), Université Claude Bernard Lyon 1 (UCBL), Université de Lyon, Georgetown University Medical Center, Génétique, Reproduction et Développement (GReD), Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Université Clermont Auvergne (UCA), CHU Clermont-Ferrand, Ibn Rochd University Hospital of Casablanca, University Hassan II of Casablanca, Morocco., Centre Hospitalier Universitaire [Grenoble] (CHU), Monash Biomedicine Discovery Institute, Monash University [Clayton], University of Melbourne, McGill University = Université McGill [Montréal, Canada], Mécanisme de l’Angiogenèseet des BarrièresBiologiques (MAB2), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes (UGA)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Alfaidy, Nadia, and ANR-17-EURE-0003,CBH-EUR-GS,CBH-EUR-GS(2017)

Subjects
0301 basic medicine, Cancer Research, orthotopic model of choriocarcinoma, [SDV.BDLR.RA]Life Sciences [q-bio]/Reproductive Biology/Asexual reproduction, Article, Gestational choriocarcinoma, Immune tolerance, Proinflammatory cytokine, NLRP7, 03 medical and health sciences, 0302 clinical medicine, Immune system, Downregulation and upregulation, medicine, choriocarcinoma, tumor microenvironment, [SDV.BDLR.RA] Life Sciences [q-bio]/Reproductive Biology/Asexual reproduction, reproductive and urinary physiology, RC254-282, Tumor microenvironment, Gene knockdown, business.industry, Choriocarcinoma, Neoplasms. Tumors. Oncology. Including cancer and carcinogens, medicine.disease, 3. Good health, hydatidiform mole, 030104 developmental biology, Oncology, 030220 oncology & carcinogenesis, embryonic structures, Cancer research, business, maternal immune tolerance
Abstract

The inflammatory gene NLRP7 is the major gene responsible for recurrent complete hydatidiform moles (CHM), an abnormal pregnancy that can develop into gestational choriocarcinoma (CC). However, the role of NLRP7 in the development and immune tolerance of CC has not been investigated. Three approaches were employed to define the role of NLRP7 in CC development: (i) a clinical study that analyzed human placenta and sera collected from women with normal pregnancies, CHM or CC, (ii) an in vitro study that investigated the impact of NLRP7 knockdown on tumor growth and organization, and (iii) an in vivo study that used two CC mouse models, including an orthotopic model. NLRP7 and circulating inflammatory cytokines were upregulated in tumor cells and in CHM and CC. In tumor cells, NLRP7 functions in an inflammasome-independent manner and promoted their proliferation and 3D organization. Gravid mice placentas injected with CC cells invalidated for NLRP7, exhibited higher maternal immune response, developed smaller tumors, and displayed less metastases. Our data characterized the critical role of NLRP7 in CC and provided evidence of its contribution to the development of an immunosuppressive maternal microenvironment that not only downregulates the maternal immune response but also fosters the growth and progression of CC.

Published
2021

30. Cibler la dégradation des ARNms médiée par les éléments riches en A et U à l'aide d'une forme active tronquée de la protéine à doigts de zincs TIS11b/BRF1 affecte les principales propriétés de la tumorigenèse mammaire

Author

Nadia Cherradi, Josiane Denis, Laurent Guyon, Felicitas Rataj, Séverine Planel, Odile Filhol, Jean-Jacques Feige, Caroline Roelants, Biologie du Cancer et de l'Infection (BCI ), Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut National de la Santé et de la Recherche Médicale (INSERM), Invasion mechanisms in angiogenesis and cancer (IMAC), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Famille BMP dans l'angiogenèse et la lymphangiogenèse (BAL ), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA), and Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG)

Subjects
0301 basic medicine, Untranslated region, Cancer Research, Carcinogenesis, RNA Stability, Recombinant Fusion Proteins, [SDV]Life Sciences [q-bio], Tristetraprolin, Breast Neoplasms, Biology, Mice, 03 medical and health sciences, chemistry.chemical_compound, 0302 clinical medicine, Chlorocebus aethiops, Human Umbilical Vein Endothelial Cells, Genetics, medicine, Animals, Humans, Protein Isoforms, Molecular Biology, Cells, Cultured, AU Rich Elements, AU-rich element, Mice, Inbred BALB C, TATA-Binding Protein Associated Factors, Tumor microenvironment, Tumor hypoxia, Cancer, Zinc Fingers, medicine.disease, 3. Good health, Cell biology, Gene Expression Regulation, Neoplastic, Vascular endothelial growth factor, 030104 developmental biology, chemistry, Gain of Function Mutation, 030220 oncology & carcinogenesis, COS Cells, Cancer cell, Female
Abstract

International audience; Altered expression of regulatory RNA-binding proteins (RBPs) in cancer leads to abnormal expression of mRNAs encoding many factors involved in cancer hallmarks. While conventional anticancer therapies usually target one pathway at a time, targeting key RBP would affect multiple genes and thus overcome drug resistance. Among the Tristetraprolin family of RBP, TIS11b/BRF1/ZFP36L1 mediates mRNA decay through binding to Adenylate/Uridylate (AU-rich elements) in mRNA 3'-untranslated region and recruitment of mRNA degradation enzymes. Here, we show that TIS11b is markedly underexpressed in three breast cancer cell lines, as well as in breast tumor samples. We hypothesized that restoring intracellular TIS11b levels could impair cancer cell phenotypic traits. We thus generated a derivative of TIS11b called R9-ZnCS334D, by combining N-terminal domain deletion, serine-to-aspartate substitution at position 334 to enhance the function of the protein and fusion to the cell-penetrating peptide polyarginine R9. R9-ZnCS334D not only blunted secretion of vascular endothelial growth factor (VEGF) but also inhibited proliferation, migration, invasion, and anchorage-independent growth of murine 4T1 or human MDA-MB-231 breast cancer cells. Moreover, R9-ZnCS334D prevented endothelial cell organization into vessel-like structures, suggesting that it could potentially target various cell types within the tumor microenvironment. In vivo, injection of R9-ZnCS334D in 4T1 tumors impaired tumor growth, decreased tumor hypoxia, and expression of the epithelial-to-mesenchymal transition (EMT) markers Snail, Vimentin, and N-cadherin. R9-ZnCS334D also hindered the expression of chemokines and proteins involved in cancer-related inflammation and invasion including Fractalkine (CX3CL1), SDF-1 (CXCL12), MCP-1 (CCL2), NOV (CCN3), and Pentraxin-3 (PTX3). Collectively, our data indicate that R9-ZnCS334D counteracts multiple traits of breast cancer cell aggressiveness and suggest that this novel protein could serve as the basis for innovative multi-target therapies in cancer.

Published
2019

31. Cooperative Blockade of CK2 and ATM Kinases Drives Apoptosis in VHL-Deficient Renal Carcinoma Cells through ROS Overproduction

Author

Yann Wallez, Laurent Guyon, Caroline Roelants, Christophe Battail, Q. Franquet, Frédéric Chalmel, Irinka Séraudie, Claude Cochet, Caroline Barette, Odile Filhol, Sofia Giacosa, Nicolas Peilleron, Marie-Odile Fauvarque, Catherine Pillet, Emmanuelle Soleilhac, Clément Sarrazin, Bertrand Evrard, Gaelle Fiard, Jean-Luc Descotes, Jean-Alexandre Long, BioSanté (UMR BioSanté), Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes (UGA), Invasion mechanisms in angiogenesis and cancer (IMAC), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes (UGA)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Inovarion, Institut de recherche en santé, environnement et travail (Irset), Université d'Angers (UA)-Université de Rennes (UR)-École des Hautes Études en Santé Publique [EHESP] (EHESP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Structure Fédérative de Recherche en Biologie et Santé de Rennes ( Biosit : Biologie - Santé - Innovation Technologique ), Genetics and Chemogenomics (GenChem), CHU Grenoble, Gestes Medico-chirurgicaux Assistés par Ordinateur (TIMC-GMCAO), Translational Innovation in Medicine and Complexity / Recherche Translationnelle et Innovation en Médecine et Complexité - UMR 5525 (TIMC ), VetAgro Sup - Institut national d'enseignement supérieur et de recherche en alimentation, santé animale, sciences agronomiques et de l'environnement (VAS)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes (UGA)-Institut polytechnique de Grenoble - Grenoble Institute of Technology (Grenoble INP ), Université Grenoble Alpes (UGA)-VetAgro Sup - Institut national d'enseignement supérieur et de recherche en alimentation, santé animale, sciences agronomiques et de l'environnement (VAS)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes (UGA)-Institut polytechnique de Grenoble - Grenoble Institute of Technology (Grenoble INP ), Université Grenoble Alpes (UGA), UMR1036, Institut National de la Santé et de la Recherche Médicale, 2018, Groupement des Entreprises Françaises dans la lutte contre le Cancer, Isère, Ligue Contre le Cancer, 2017, Association Française d'Urologie, 2019, Centre Hospitalier Universitaire de Grenoble Alpes, ANR-17-EURE-0003,CBH-EUR-GS,CBH-EUR-GS(2017), Université d'Angers (UA)-Université de Rennes 1 (UR1), Université de Rennes (UNIV-RENNES)-Université de Rennes (UNIV-RENNES)-École des Hautes Études en Santé Publique [EHESP] (EHESP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Structure Fédérative de Recherche en Biologie et Santé de Rennes ( Biosit : Biologie - Santé - Innovation Technologique ), and Chard-Hutchinson, Xavier

Subjects
Cancer Research, kinase inhibitor, CK2, [SDV.CAN]Life Sciences [q-bio]/Cancer, Biology, urologic and male genital diseases, lcsh:RC254-282, [SDV.MHEP.UN]Life Sciences [q-bio]/Human health and pathology/Urology and Nephrology, Article, NOX4, 03 medical and health sciences, 0302 clinical medicine, Mediator, [SDV.CAN] Life Sciences [q-bio]/Cancer, medicine, ROS pathway, tumor tissue slices, 030304 developmental biology, 0303 health sciences, Kinase, ccRCC, apoptosis, Cancer, HIF-2α, medicine.disease, lcsh:Neoplasms. Tumors. Oncology. Including cancer and carcinogens, [SDV.MHEP.UN] Life Sciences [q-bio]/Human health and pathology/Urology and Nephrology, 3. Good health, Clear cell renal cell carcinoma, Oncology, Apoptosis, Cell culture, 030220 oncology & carcinogenesis, [SDV.SP.PHARMA] Life Sciences [q-bio]/Pharmaceutical sciences/Pharmacology, ATM, Cancer research, [SDV.SP.PHARMA]Life Sciences [q-bio]/Pharmaceutical sciences/Pharmacology, Ex vivo
Abstract

Simple Summary Renal cell carcinoma (RCC) is the eighth leading malignancy in the world, accounting for 4% of all cancers with poor outcome when metastatic. Protein kinases are highly druggable proteins, which are often aberrantly activated in cancers. The aim of our study was to identify candidate targets for metastatic clear cell renal cell carcinoma therapy, using chemo-genomic-based high-throughput screening. We found that the combined inhibition of the CK2 and ATM kinases in renal tumor cells and patient-derived tumor samples induces synthetic lethality. Mechanistic investigations unveil that this drug combination triggers apoptosis through HIF-2α-(Hypoxic inducible factor HIF-2α) dependent reactive oxygen species (ROS) overproduction, giving a new option for patient care in metastatic RCC. Abstract Kinase-targeted agents demonstrate antitumor activity in advanced metastatic clear cell renal cell carcinoma (ccRCC), which remains largely incurable. Integration of genomic approaches through small-molecules and genetically based high-throughput screening holds the promise of improved discovery of candidate targets for cancer therapy. The 786-O cell line represents a model for most ccRCC that have a loss of functional pVHL (von Hippel-Lindau). A multiplexed assay was used to study the cellular fitness of a panel of engineered ccRCC isogenic 786-O VHL− cell lines in response to a collection of targeted cancer therapeutics including kinase inhibitors, allowing the interrogation of over 2880 drug–gene pairs. Among diverse patterns of drug sensitivities, investigation of the mechanistic effect of one selected drug combination on tumor spheroids and ex vivo renal tumor slice cultures showed that VHL-defective ccRCC cells were more vulnerable to the combined inhibition of the CK2 and ATM kinases than wild-type VHL cells. Importantly, we found that HIF-2α acts as a key mediator that potentiates the response to combined CK2/ATM inhibition by triggering ROS-dependent apoptosis. Importantly, our findings reveal a selective killing of VHL-deficient renal carcinoma cells and provide a rationale for a mechanism-based use of combined CK2/ATM inhibitors for improved patient care in metastatic VHL-ccRCC.

Published
2021

32. A RUNX2 stabilization pathway mediates physiologic and pathologic bone formation

Author

Jae-Hyuck Shim, Julia F. Charles, Matthew B. Greenblatt, Yeon Suk Yang, Paul B. Yu, Takeshi Takarada, Xianpeng Ge, Hyunho Chun, Wei Gu, Jung-Min Kim, Minkyung Song, Teresa Dinter, Na Li, Odile Filhol-Cochet, Seoyeon Bok, Kwang Hwan Park, Ning Kon, Ren Xu, Brigitte Boldyreff, University of Massachusetts Medical School [Worcester] (UMASS), University of Massachusetts System (UMASS), Yonsei University College of Medicine [Séoul, Corée du Sud], Xiamen University, Sungkyunkwan University [Suwon] (SKKU), Korea Advanced Institute of Science and Technology (KAIST), Weill Medical College of Cornell University [New York], Harvard Medical School [Boston] (HMS), Biologie du Cancer et de l'Infection (BCI ), Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes (UGA), KinaseDetect ApS [Krusa, Danemark], Columbia University College of Physicians and Surgeons, Okayama University Graduate School of Medicine [Okayama, Japon], This research was supported by Basic Science Research Program through the National Research Foundation of Korea (NRF) funded by the Ministry of Education (2014R1A6A3A03055719, J.M.K.). M.B.G. holds a Career Award for Medical Scientists from the Burroughs Wellcome Fund, a Basil O’Connor Award from the March of Dimes and is supported by an NIH Director’s Early Independence Award (1DP5OD021351), and a Pershing Square Sohn Cancer Research Prize for Young Investigators. J.H.S holds support from NIAMS of the NIH under R01AR068983, R21AR072836, and R21AR073331, an UMCCTS pilot project program award., Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes (UGA)-Institut National de la Santé et de la Recherche Médicale (INSERM), and Bodescot, Myriam

Subjects
0301 basic medicine, Male, Cellular differentiation, Regulator, General Physics and Astronomy, Core Binding Factor Alpha 1 Subunit, Haploinsufficiency, Ubiquitin-Specific Peptidase 7, 0302 clinical medicine, Osteogenesis, Phosphorylation, lcsh:Science, Casein Kinase II, [SDV.MHEP.RSOA] Life Sciences [q-bio]/Human health and pathology/Rhumatology and musculoskeletal system, Multidisciplinary, Chemistry, Protein Stability, musculoskeletal, neural, and ocular physiology, Bone development, Osteoblast, Cell Differentiation, [SDV.BBM.MN]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Molecular Networks [q-bio.MN], Middle Aged, Cell biology, Hindlimb, RUNX2, medicine.anatomical_structure, [SDV.MHEP.RSOA]Life Sciences [q-bio]/Human health and pathology/Rhumatology and musculoskeletal system, 030220 oncology & carcinogenesis, embryonic structures, Female, Stem cell, Casein kinase 2, Cleidocranial Dysplasia, Deubiquitination, Adult, musculoskeletal diseases, endocrine system, Science, Trauma, General Biochemistry, Genetics and Molecular Biology, Article, 03 medical and health sciences, stomatognathic system, medicine, Animals, Humans, RNA, Messenger, Bone, Aged, Osteoblasts, Ossification, Heterotopic, fungi, General Chemistry, Mice, Inbred C57BL, 030104 developmental biology, [SDV.BBM.MN] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Molecular Networks [q-bio.MN], lcsh:Q, Gene Deletion
Abstract

The osteoblast differentiation capacity of skeletal stem cells (SSCs) must be tightly regulated, as inadequate bone formation results in low bone mass and skeletal fragility, and over-exuberant osteogenesis results in heterotopic ossification (HO) of soft tissues. RUNX2 is essential for tuning this balance, but the mechanisms of posttranslational control of RUNX2 remain to be fully elucidated. Here, we identify that a CK2/HAUSP pathway is a key regulator of RUNX2 stability, as Casein kinase 2 (CK2) phosphorylates RUNX2, recruiting the deubiquitinase herpesvirus-associated ubiquitin-specific protease (HAUSP), which stabilizes RUNX2 by diverting it away from ubiquitin-dependent proteasomal degradation. This pathway is important for both the commitment of SSCs to osteoprogenitors and their subsequent maturation. This CK2/HAUSP/RUNX2 pathway is also necessary for HO, as its inhibition blocked HO in multiple models. Collectively, active deubiquitination of RUNX2 is required for bone formation and this CK2/HAUSP deubiquitination pathway offers therapeutic opportunities for disorders of inappropriate mineralization., Runx2 is essential for tuning the generation of bone from skeletal stem cells (SSCs). Here, the authors demonstrate that the CK2/HAUSP pathway stabilizes RUNX2 protein thereby regulating the commitment of SSCs to osteoprogenitors as well as their subsequent maturation, and that inhibition of this pathway can block heterotopic ossification.

Published
2020

33. Protein kinase CK2 contributes to placental development: physiological and pathological implications

Author

Nicolas Lemaitre, Claude Cochet, Daniel Vaiman, Déborah Reynaud, Odile Filhol, Mohamed Benharouga, Nadia Alfaidy, Roland Abi Nahed, Solene Lartigue, Caroline Roelants, Invasion mechanisms in angiogenesis and cancer (IMAC), Biologie du Cancer et de l'Infection (BCI ), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Grenoble Alpes (UGA)-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Grenoble Alpes (UGA)-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA), Inovarion, Inovarion [Paris], Institut Cochin (IC UM3 (UMR 8104 / U1016)), Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Biologie des Métaux (BioMet), Laboratoire de Chimie et Biologie des Métaux (LCBM - UMR 5249), Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes (UGA)-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes (UGA)-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes (UGA)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes (UGA), Biologie des Métaux (BioMet ), Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes (UGA)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), and Centre National de la Recherche Scientifique (CNRS)-Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM)

Subjects
Adult, Invasion de trophoblastes, Modèle animal, Prééclampsie, Adolescent, Protein subunit, Placenta, Biology, Preeclampsia, Andrology, Mice, Young Adult, 03 medical and health sciences, 0302 clinical medicine, Pre-Eclampsia, Pregnancy, In vivo, Cell Line, Tumor, Drug Discovery, medicine, [SDV.MHEP.PHY]Life Sciences [q-bio]/Human health and pathology/Tissues and Organs [q-bio.TO], Animals, Humans, Casein Kinase II, Genetics (clinical), reproductive and urinary physiology, Trophoblast, medicine.disease, Molecular medicine, Placentation, In vitro, Trophoblasts, Mice, Inbred C57BL, Disease Models, Animal, Pregnancy Trimester, First, Protéine kinase CK2, medicine.anatomical_structure, [SDV.BDD.EO]Life Sciences [q-bio]/Development Biology/Embryology and Organogenesis, embryonic structures, Molecular Medicine, Female, Ex vivo, 030215 immunology
Abstract

International audience; Preeclampsia (PE) is the most threatening pathology of human pregnancy. Its development is thought to be due to a failure in the invasion of trophoblast cells that establish the feto-maternal circulation. Protein kinase CK2 is a ubiquitous enzyme reported to be involved in the control of cell invasion. CK2 consists of two subunits, a catalytic subunit, CK2α, and a regulatory subunit, CK2β. To date, no data exist regarding the expression and role of this enzyme in normal and PE pregnancies. We performed studies, at the clinical level using distinctive cohorts from early pregnancy (n = 24) and from PE (n = 23) and age-matched controls (n = 28); in vitro, using trophoblast cell lines; ex vivo, using placental explants; and in vivo, using PE mouse models. We demonstrated that (i) CK2 is more expressed during the late first trimester of pregnancy and is mainly localized in differentiated trophoblast cells, (ii) the inhibition of its enzymatic activity decreased the proliferation, migration, invasion, and syncytialization of trophoblast cells, both in 2D and 3D culture systems, and (iii) CK2 activity and the CK2α/CK2β protein ratio were increased in PE human placentas. The pattern and profile of CK2 expression were confirmed in gravid mice along with an increase in the PE mouse models. Altogether, our results demonstrate that CK2 plays an essential role in the establishment of the feto-maternal circulation and that its deregulation is associated with PE development. The increase in CK2 activity in PE might constitute a compensatory mechanism to ensure proper pregnancy progress.

Published
2020

34. Discovery of holoenzyme-disrupting chemicals as substrate-selective CK2 inhibitors

Author

Ruben Abagyan, Renaud Prudent, Thierry Lomberget, Céline F. Sautel, Béatrice Laudet, Matthias Engel, Isabelle Krimm, Odile Filhol, Marc Le Borgne, Irina Kufareva, Mohamed Ettaoussi, Claude Cochet, Paul Brear, Benoit Bestgen, Virginie Moucadel, Skaggs School of Pharmacy and Pharmaceutical Sciences [San Diego], University of California [San Diego] (UC San Diego), University of California-University of California, Molécules bioactives et chimie médicinale (B2MC), Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon, University of Cambridge [UK] (CAM), Invasion mechanisms in angiogenesis and cancer (IMAC), Biologie du Cancer et de l'Infection (BCI ), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA), Laboratoire Adaptation et pathogénie des micro-organismes [Grenoble] (LAPM), Université Joseph Fourier - Grenoble 1 (UJF)-Centre National de la Recherche Scientifique (CNRS), Université de Lyon, Institute for Advanced Biosciences / Institut pour l'Avancée des Biosciences (Grenoble) (IAB), Centre Hospitalier Universitaire [Grenoble] (CHU)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Etablissement français du sang - Auvergne-Rhône-Alpes (EFS)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019]), Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut National de la Santé et de la Recherche Médicale (INSERM), Centre de Recherche Jean-Pierre AUBERT Neurosciences et Cancer - U1172 Inserm - U837 (JPArc), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Lille Nord de France (COMUE)-Université de Lille, Centre National de la Recherche Scientifique (CNRS)-Université Joseph Fourier - Grenoble 1 (UJF), Centre de Résonance Magnetique Nucleaire (CRMN), Institut de Chimie et Biochimie Moléculaires et Supramoléculaires (ICBMS), Université de Lyon-Université de Lyon-Institut National des Sciences Appliquées de Lyon (INSA Lyon), Université de Lyon-Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-Institut de Chimie du CNRS (INC)-École Supérieure Chimie Physique Électronique de Lyon-Centre National de la Recherche Scientifique (CNRS)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-Institut de Chimie du CNRS (INC)-École Supérieure Chimie Physique Électronique de Lyon-Centre National de la Recherche Scientifique (CNRS), Saarland University [Saarbrücken], Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), University of California (UC)-University of California (UC), Centre de Recherche Jean-Pierre AUBERT Neurosciences et Cancer - U837 (JPArc), Université Lille Nord de France (COMUE)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Lille, Université de Lyon-Université de Lyon-École Supérieure de Chimie Physique Électronique de Lyon (CPE)-Institut National des Sciences Appliquées de Lyon (INSA Lyon), Université de Lyon-Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS), ANR-17-EURE-0003,CBH-EUR-GS,CBH-EUR-GS(2017), Kufareva, Irina [0000-0001-9083-7039], Filhol, Odile [0000-0003-1964-7958], Borgne, Marc Le [0000-0003-1398-075X], Apollo - University of Cambridge Repository, and Le Borgne, Marc

Subjects
0301 basic medicine, genetic structures, lcsh:Medicine, [CHIM.THER]Chemical Sciences/Medicinal Chemistry, Crystallography, X-Ray, 01 natural sciences, Substrate Specificity, Adenosine Triphosphate, Cell Movement, Catalytic Domain, Phosphorylation, lcsh:Science, Casein Kinase II, ComputingMilieux_MISCELLANEOUS, Cancer, Tumor, Crystallography, Multidisciplinary, Molecular medicine, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], Chemistry, Small molecule, 3. Good health, Molecular Docking Simulation, 5.1 Pharmaceuticals, [SDV.SP.PHARMA] Life Sciences [q-bio]/Pharmaceutical sciences/Pharmacology, Development of treatments and therapeutic interventions, [SDV.SP.MED] Life Sciences [q-bio]/Pharmaceutical sciences/Medication, [SDV.BBM.BS] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], [CHIM.THER] Chemical Sciences/Medicinal Chemistry, Protein subunit, Kinases, [SDV.BC]Life Sciences [q-bio]/Cellular Biology, Article, Target validation, Cell Line, 03 medical and health sciences, Targeted therapies, [SDV.SP.MED]Life Sciences [q-bio]/Pharmaceutical sciences/Medication, Target identification, Cell Line, Tumor, Humans, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, Binding site, Protein kinase A, Protein Kinase Inhibitors, Cell Proliferation, Binding Sites, 010405 organic chemistry, Cell growth, lcsh:R, Surface Plasmon Resonance, In vitro, 0104 chemical sciences, Protein Subunits, Kinetics, 030104 developmental biology, Cell culture, X-Ray, [SDV.SP.PHARMA]Life Sciences [q-bio]/Pharmaceutical sciences/Pharmacology, Biophysics, lcsh:Q, Holoenzymes, Peptides
Abstract

CK2 is a constitutively active protein kinase overexpressed in numerous malignancies. Interaction between CK2α and CK2β subunits is essential for substrate selectivity. The CK2α/CK2β interface has been previously targeted by peptides to achieve functional effects; however, no small molecules modulators were identified due to pocket flexibility and open shape. Here we generated numerous plausible conformations of the interface using the fumigation modeling protocol, and virtually screened a compound library to discover compound 1 that suppressed CK2α/CK2β interaction in vitro and inhibited CK2 in a substrate-selective manner. Orthogonal SPR, crystallography, and NMR experiments demonstrated that 4 and 6, improved analogs of 1, bind to CK2α as predicted. Both inhibitors alter CK2 activity in cells through inhibition of CK2 holoenzyme formation. Treatment with 6 suppressed MDA-MB231 triple negative breast cancer cell growth and induced apoptosis. Altogether, our findings exemplify an innovative computational-experimental approach and identify novel non-peptidic inhibitors of CK2 subunit interface disclosing substrate-selective functional effects.

Published
2019

35. The size-speed-force relationship governs migratory cell response to tumorigenic factors

Author

Andreas Christ, Aldo Leal-Egaña, Timothée Vignaud, Caroline Roelants, Gaëlle Letort, Manuel Théry, Odile Filhol, Jean Louis Martiel, Physiologie cellulaire et végétale (LPCV), Institut National de la Recherche Agronomique (INRA)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes (UGA)-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA), Biologie du Cancer et de l'Infection (BCI ), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Grenoble Alpes (UGA)-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Invasion mechanisms in angiogenesis and cancer (IMAC), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Grenoble Alpes (UGA)-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Alloimmunité-Autoimmunité-Transplantation (A2T), Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut Universitaire d'Hématologie (IUH), Université Paris Diderot - Paris 7 (UPD7)-Université Paris Diderot - Paris 7 (UPD7), Institut National du Cancer (INCA PLBIO2011), Institut Universitaire d'Hématologie (IUH), Université Paris Diderot - Paris 7 (UPD7)-Université Paris Diderot - Paris 7 (UPD7)-Institut National de la Santé et de la Recherche Médicale (INSERM), Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Recherche Agronomique (INRA)-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut National de la Santé et de la Recherche Médicale (INSERM), Institut National du Cancer [INCA PLBIO2011], Institut National de la Recherche Agronomique (INRA)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), and Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG)

Subjects
0301 basic medicine, Cell contractility, Carcinogenesis, Receptor, ErbB-2, Cell, Tumorigenic factors, [SDV.BC.BC]Life Sciences [q-bio]/Cellular Biology/Subcellular Processes [q-bio.SC], Contractile Proteins, Cell Movement, Transforming Growth Factor beta, Tumor development, ComputingMilieux_MISCELLANEOUS, Cancer, education.field_of_study, Human breast cell, Cell migration, Anatomy, Cell mechanics, Articles, Cell biology, Biomechanical Phenomena, Cell Motility, medicine.anatomical_structure, Cell Transformation, Neoplastic, Female, Population, [SDV.CAN]Life Sciences [q-bio]/Cancer, Breast Neoplasms, [SDV.BC]Life Sciences [q-bio]/Cellular Biology, Biology, Collagen Type I, Contractility, 03 medical and health sciences, Cell Line, Tumor, medicine, Cell Adhesion, Humans, education, Molecular Biology, Cell Proliferation, Cell Size, Cell growth, Cell Biology, Traction forces, Fibronectin, 030104 developmental biology, Cell culture, biology.protein, Transforming growth factor
Abstract

Normal and transformed motile cells follow a common trend in which size and contractile forces are negatively correlated with cell speed. However, tumorigenic factors amplify the preexisting population heterogeneity and lead some cells to exhibit biomechanical properties that are more extreme than those observed with normal cells., Tumor development progresses through a complex path of biomechanical changes leading first to cell growth and contraction and then cell deadhesion, scattering, and invasion. Tumorigenic factors may act specifically on one of these steps or have a wider spectrum of actions, leading to a variety of effects and thus sometimes to apparent contradictory outcomes. Here we used micropatterned lines of collagen type I/fibronectin on deformable surfaces to standardize cell behavior and measure simultaneously cell size, speed of motion and magnitude of the associated traction forces at the level of a single cell. We analyzed and compared the normal human breast cell line MCF10A in control conditions and in response to various tumorigenic factors. In all conditions, a wide range of biomechanical properties was identified. Despite this heterogeneity, normal and transformed motile cells followed a common trend whereby size and contractile forces were negatively correlated with cell speed. Some tumorigenic factors, such as activation of ErbB2 or loss of the βsubunit of casein kinase 2, shifted the whole population toward a faster speed and lower contractility state. Treatment with transforming growth factor β induced some cells to adopt opposing behaviors such as extremely high versus extremely low contractility. Thus tumor transformation amplified preexisting population heterogeneity and led some cells to exhibit biomechanical properties that were more extreme than those observed with normal cells.

Published
2017

36. 2-Aminothiazole Derivatives as Selective Allosteric Modulators of the Protein Kinase CK2. 2. Structure-Based Optimization and Investigation of Effects Specific to the Allosteric Mode of Action

Author

Thierry Lomberget, Odile Filhol, Claude Cochet, Rolf W. Hartmann, Irina Kufareva, Ruben Abagyan, Matthias Engel, Chris Abell, Wei-Guang Seetoh, Marc Le Borgne, Benoit Bestgen, Molécules bioactives et chimie médicinale (B2MC), Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon, Skaggs School of Pharmacy and Pharmaceutical Sciences [San Diego], University of California [San Diego] (UC San Diego), University of California (UC)-University of California (UC), Univ Cambridge, Dept Chem, Cambridge CB2 1EW, England, affiliation inconnue, Jena Optronik GmbH (Jena Optronik), entreprise, Invasion mechanisms in angiogenesis and cancer (IMAC), Biologie du Cancer et de l'Infection (BCI ), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA), University of California-University of California, Saarland University [Saarbrücken], U873, Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Institut National de la Santé et de la Recherche Médicale (INSERM), Transduction du signal : signalisation calcium, phosphorylation et inflammation, Université Joseph Fourier - Grenoble 1 (UJF)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Institut National de la Santé et de la Recherche Médicale (INSERM), Filhol, Odile, and HIPS, Helmholtz-Institut für Pharmazeutische Forschung Saarland, Universitätscampus E8.1 66123 Saarbrücken, Germany.

Subjects
Druggability, Apoptosis, [CHIM.THER]Chemical Sciences/Medicinal Chemistry, Plasma protein binding, Ligands, Benzoates, 01 natural sciences, chemistry.chemical_compound, Drug Discovery, kinase inhibitors, Prodrugs, Casein Kinase II, [SDV.BBM.BC] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biochemistry [q-bio.BM], anti-cancer, ComputingMilieux_MISCELLANEOUS, Benzoic acid, 0303 health sciences, Tumor, Molecular Structure, Pharmacology and Pharmaceutical Sciences, Small molecule, kinase selectivity, 3. Good health, [SDV.BBM.BC]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biomolecules [q-bio.BM], Molecular Docking Simulation, 5.1 Pharmaceuticals, Molecular Medicine, Development of treatments and therapeutic interventions, Lead compound, [CHIM.CHEM]Chemical Sciences/Cheminformatics, Allosteric Site, Protein Binding, Stereochemistry, CK2, Medicinal & Biomolecular Chemistry, Allosteric regulation, Antineoplastic Agents, allosteric inhibition, Article, Cell Line, Structure-Activity Relationship, Medicinal and Biomolecular Chemistry, 03 medical and health sciences, [SDV.SP.MED]Life Sciences [q-bio]/Pharmaceutical sciences/Medication, Allosteric Regulation, Cell Line, Tumor, Humans, Structure–activity relationship, Naphthyridines, [SDV.BBM.BC]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biochemistry [q-bio.BM], Mode of action, Protein Kinase Inhibitors, 030304 developmental biology, organic chemicals, Organic Chemistry, 0104 chemical sciences, Thiazoles, 010404 medicinal & biomolecular chemistry, chemistry, [SDV.SP.PHARMA]Life Sciences [q-bio]/Pharmaceutical sciences/Pharmacology, Phenazines
Abstract

Protein CK2 has gained much interest as an anti-cancer drug target in the last decade. We had previously described the identification of a new allosteric site on the catalytic α-subunit, along with first small molecule ligands based on the 4-(4-phenylthiazol-2-ylamino) benzoic acid scaffold. In the present work, structure optimizations guided by a binding model led to the identification of the lead compound 2-hydroxy-4-((4-(naphthalen-2-yl)thiazol-2-yl)amino)benzoic acid (27), showing a submicromolar potency against purified CK2α (IC(50) = 0.6 μM). Furthermore, 27 induced apoptosis and cell death in 786-O renal cell carcinoma cells (EC(50) = 5 μM) and inhibited STAT3 activation even more potently than the ATP-competitive drug candidate CX-4945 (EC(50)s: 1.6 μM vs. 5.3 μM). Notably, the potencies of our allosteric ligands to inhibit CK2 varied depending on the individual substrate. Altogether, the novel allosteric pocket was proved a druggable site, offering an excellent perspective to develop efficient and selective allosteric CK2 inhibitors.

Published
2019

37. The tyrosine-kinase inhibitor sunitinib targets vascular endothelial (VE)-cadherin: a marker of response to antitumoural treatment in metastatic renal cell carcinoma

Author

Helena Polena, Ellen Blanc, Abir Khalil-Mgharbel, Delphine Borchiellini, Gilles Pagès, Jean-Marc Ferrero, Maeva Dufies, Isabelle Vilgrain, Céline Ferlay-Segura, Aude Salomon, Alban Deroux, Odile Filhol, Adama Sidibé, Jean-Louis Quesada, Sylvie Negrier, Julie Creuzet, Bernard Escudier, Caroline Roelants, Claude Cochet, Biologie du Cancer et de l'Infection (BCI ), Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut National de la Santé et de la Recherche Médicale (INSERM), Centre méditerranéen de médecine moléculaire (C3M), Université Nice Sophia Antipolis (... - 2019) (UNS), COMUE Université Côte d'Azur (2015-2019) (COMUE UCA)-COMUE Université Côte d'Azur (2015-2019) (COMUE UCA)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Côte d'Azur (UCA), Université Grenoble Alpes - Institut d'Administration des Entreprises (UGA IAE), Université Grenoble Alpes [2016-2019] (UGA [2016-2019]), Invasion mechanisms in angiogenesis and cancer (IMAC), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Université Grenoble Alpes - UFR Médecine (UGA UFRM), CIC - Grenoble, Université Joseph Fourier - Grenoble 1 (UJF)-Institut National de la Santé et de la Recherche Médicale (INSERM), Délégation à la Recherche Clinique et à l'Innovation [Lyon] (DRCI), UNICANCER - Centre Léon Bérard Lyon (Rhône)-Centre de Lutte Contre le Cancer de Lyon et Rhône-Alpes, Centre de Lutte contre le Cancer Antoine Lacassagne [Nice] (UNICANCER/CAL), UNICANCER-Université Côte d'Azur (UCA), Oncologie génito-urinaire, Département de médecine oncologique [Gustave Roussy], Institut Gustave Roussy (IGR)-Institut Gustave Roussy (IGR), Université de Lyon, Institut de signalisation, biologie du développement et cancer (ISBDC), Centre National de la Recherche Scientifique (CNRS)-Université Nice Sophia Antipolis (... - 2019) (UNS), COMUE Université Côte d'Azur (2015-2019) (COMUE UCA)-COMUE Université Côte d'Azur (2015-2019) (COMUE UCA)-Université Côte d'Azur (UCA), COMUE Université Côte d'Azur (2015 - 2019) (COMUE UCA), Angiogenèse hormono-regulée et angiogenèse tumorale (LAPV), Université Joseph Fourier - Grenoble 1 (UJF)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Institut National de la Santé et de la Recherche Médicale (INSERM), Transduction du signal : signalisation calcium, phosphorylation et inflammation, Centre Léon Bérard [Lyon], Laboratoire de Probabilités, Statistiques et Modélisations (LPSM (UMR_8001)), Université Paris Diderot - Paris 7 (UPD7)-Sorbonne Université (SU)-Centre National de la Recherche Scientifique (CNRS), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA), Université Nice Sophia Antipolis (1965 - 2019) (UNS), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Centre Léon Bérard [Lyon]-UNICANCER - Centre Léon Bérard Lyon (Rhône), and COMUE Université Côte d'Azur (2015-2019) (COMUE UCA)-COMUE Université Côte d'Azur (2015-2019) (COMUE UCA)-Centre National de la Recherche Scientifique (CNRS)-Université Côte d'Azur (UCA)

Subjects
0301 basic medicine, Cancer Research, Bevacizumab, Endothelium, medicine.drug_class, Antineoplastic Agents, [SDV.CAN]Life Sciences [q-bio]/Cancer, [SDV.BC]Life Sciences [q-bio]/Cellular Biology, Tyrosine-kinase inhibitor, Biomarkers, Pharmacological, Article, 03 medical and health sciences, 0302 clinical medicine, medicine, Biomarkers, Tumor, Human Umbilical Vein Endothelial Cells, Sunitinib, Humans, Molecular Targeted Therapy, Neoplasm Metastasis, [SDV.BBM.BC]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biochemistry [q-bio.BM], Carcinoma, Renal Cell, Cells, Cultured, ComputingMilieux_MISCELLANEOUS, Retrospective Studies, Clinical Trials as Topic, business.industry, medicine.disease, Cadherins, Metastatic breast cancer, Kidney Neoplasms, 3. Good health, Endothelial stem cell, 030104 developmental biology, medicine.anatomical_structure, Treatment Outcome, Oncology, 030220 oncology & carcinogenesis, Cancer research, Endothelium, Vascular, VE-cadherin, business, Kidney cancer, medicine.drug
Abstract

Background Vascular endothelial (VE)-cadherin is an endothelial cell-specific protein responsible for endothelium integrity. Its adhesive properties are regulated by post-translational processing, such as tyrosine phosphorylation at site Y685 in its cytoplasmic domain, and cleavage of its extracellular domain (sVE). In hormone-refractory metastatic breast cancer, we recently demonstrated that sVE levels correlate to poor survival. In the present study, we determine whether kidney cancer therapies had an effect on VE-cadherin structural modifications and their clinical interest to monitor patient outcome. Methods The effects of kidney cancer biotherapies were tested on an endothelial monolayer model mimicking the endothelium lining blood vessels and on a homotypic and heterotypic 3D cell model mimicking tumour growth. sVE was quantified by ELISA in renal cell carcinoma patients initiating sunitinib (48 patients) or bevacizumab (83 patients) in the first-line metastatic setting (SUVEGIL and TORAVA trials). Results Human VE-cadherin is a direct target for sunitinib which inhibits its VEGF-induced phosphorylation and cleavage on endothelial monolayer and endothelial cell migration in the 3D model. The tumour cell environment modulates VE-cadherin functions through MMPs and VEGF. We demonstrate the presence of soluble VE-cadherin in the sera of mRCC patients (n = 131) which level at baseline, is higher than in a healthy donor group (n = 96). Analysis of sVE level after 4 weeks of treatment showed that a decrease in sVE level discriminates the responders vs. non-responders to sunitinib, but not bevacizumab. Conclusions These data highlight the interest for the sVE bioassay in future follow-up of cancer patients treated with targeted therapies such as tyrosine-kinase inhibitors.

Published
2018

38. Ex-vivo mice mammary glands characterization using energy-dispersive x-ray diffraction and spacially resolved CdZnTe detectors

Author

Joachim Tabary, Vera Feldman, Odile Filhol, C. Paulus, Loick Verger, Isabelle Vilgrain, Jean-Louis Hazemann, Commissariat à l'énergie atomique et aux énergies alternatives - Laboratoire d'Electronique et de Technologie de l'Information (CEA-LETI), Direction de Recherche Technologique (CEA) (DRT (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA), Matériaux, Rayonnements, Structure (MRS), Institut Néel (NEEL), Institut polytechnique de Grenoble - Grenoble Institute of Technology (Grenoble INP )-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut polytechnique de Grenoble - Grenoble Institute of Technology (Grenoble INP )-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019]), and Matériaux, Rayonnements, Structure (NEEL - MRS)

Subjects
Diffraction, [PHYS]Physics [physics], medicine.medical_specialty, Materials science, Hardware_MEMORYSTRUCTURES, medicine.diagnostic_test, 010308 nuclear & particles physics, Breast imaging, Detector, [PHYS.NEXP]Physics [physics]/Nuclear Experiment [nucl-ex], medicine.disease, 01 natural sciences, GeneralLiterature_MISCELLANEOUS, 3. Good health, Spectral imaging, 03 medical and health sciences, 0302 clinical medicine, Breast cancer, 030220 oncology & carcinogenesis, 0103 physical sciences, medicine, Mammography, Energy-dispersive X-ray diffraction, Image resolution, Biomedical engineering
Abstract

False-positive mammography results in breast cancer diagnosis can lead to unnecessary biopsies which are invasive and time-consuming. X-Ray Diffraction (XRD) has the potential of providing diagnosis-relevant information and thus can be used after a mammography to verify its results and possibly avoid needless biopsy. We present an energy-dispersive X-Ray Diffraction (EDXRD) system and data analysis method which allowed us to characterize healthy and cancerous mice mammary glands ex-vivo. Our technique showed decent glad localization along the z-axis as well as scatter signatures coherent with ones previously described in literature. We used an in-house spatially resolved CdZnTe detector, and a subpixelation technique which enhances spatial resolution. Acquisition time and dose delivered are to be optimized yet, however our results demonstrate the potential of EDXRD systems for depth-resolved breast imaging. Different geometries and processing algorithms are currently being investigated in the development of a future EDXRD clinical system for breast cancer diagnosis.

Published
2018

39. 3D polyelectrolyte scaffolds to mimic exocrine glands: a step towards a prostate-on-chip platform

Author

Odile Filhol-Cochet, Frédérique Kermarrec, Maxime Legues, Xavier Gidrol, Guillaume Laffitte, Christophe A. Marquette, Jean-Pierre Alcaraz, Olivier Nicoud, Nathalie Picollet-D'hahan, Florence Rivera, Isabelle Vilgrain, Lavinia Liguori, Jacques Thélu, Patricia Obeid, Donald K. Martin, Ali Bouamrani, Sophie Gerbaud, Benoit Gilquin, Thibaud Moufle-Milot, Adrien Mombrun, Cristina Belda Marin, Axel Tollance, Biomicrotechnologie et génomique fonctionnelle (BIOMICS), Laboratoire de Biologie à Grande Échelle (BGE - UMR S1038), Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019]), Institut d'oncologie/développement Albert Bonniot de Grenoble (INSERM U823), Université Joseph Fourier - Grenoble 1 (UJF)-CHU Grenoble-EFS-Institut National de la Santé et de la Recherche Médicale (INSERM), Systèmes Nanobiotechnologiques et Biomimétiques (TIMC-IMAG-SyNaBi), Techniques de l'Ingénierie Médicale et de la Complexité - Informatique, Mathématiques et Applications, Grenoble - UMR 5525 (TIMC-IMAG), Institut polytechnique de Grenoble - Grenoble Institute of Technology (Grenoble INP )-VetAgro Sup - Institut national d'enseignement supérieur et de recherche en alimentation, santé animale, sciences agronomiques et de l'environnement (VAS)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut polytechnique de Grenoble - Grenoble Institute of Technology (Grenoble INP )-VetAgro Sup - Institut national d'enseignement supérieur et de recherche en alimentation, santé animale, sciences agronomiques et de l'environnement (VAS)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019]), Génie Enzymatique, Membrane Biomimétique et Assemblages Supramoléculaires (GEMBAS), Institut de Chimie et Biochimie Moléculaires et Supramoléculaires (ICBMS), Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Institut National des Sciences Appliquées de Lyon (INSA Lyon), Université de Lyon-Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-Institut de Chimie du CNRS (INC)-École Supérieure Chimie Physique Électronique de Lyon-Centre National de la Recherche Scientifique (CNRS)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-Institut de Chimie du CNRS (INC)-École Supérieure Chimie Physique Électronique de Lyon-Centre National de la Recherche Scientifique (CNRS), Invasion mechanisms in angiogenesis and cancer (IMAC), Biologie du Cancer et de l'Infection (BCI ), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut National de la Santé et de la Recherche Médicale (INSERM), Commissariat à l'énergie atomique et aux énergies alternatives - Laboratoire d'Electronique et de Technologie de l'Information (CEA-LETI), Direction de Recherche Technologique (CEA) (DRT (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA), Gestes Medico-chirurgicaux Assistés par Ordinateur (TIMC-IMAG-GMCAO), Clinatec - Centre de recherche biomédicale Edmond J.Safra (SCLIN), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Direction de Recherche Technologique (CEA) (DRT (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Centre Hospitalier Universitaire [Grenoble] (CHU)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Grenoble Alpes (UGA), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Université de Lyon-Université de Lyon-École Supérieure de Chimie Physique Électronique de Lyon (CPE)-Institut National des Sciences Appliquées de Lyon (INSA Lyon), Université de Lyon-Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS), and Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Centre Hospitalier Universitaire [Grenoble] (CHU)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])

Subjects
Exocrine gland, 3D scaffolds, Biomedical Engineering, Nanotechnology, polyelectrolytes, 03 medical and health sciences, 0302 clinical medicine, Prostate, Genetics, medicine, Liquid biopsy, [SDV.IB.BIO]Life Sciences [q-bio]/Bioengineering/Biomaterials, Molecular Biology, ComputingMilieux_MISCELLANEOUS, 030304 developmental biology, 0303 health sciences, liquid biopsy, Polyelectrolyte, secretome, medicine.anatomical_structure, 030220 oncology & carcinogenesis, Molecular Medicine, organ-on-chip, TP248.13-248.65, Food Science, Biotechnology
Abstract

We report our approach to creating a microfluidic chip (namely UroLOC) that mimics the acinar/tubular structure and the luminal microenvironment of exocrine glands. The chip utilises a nanostructured membrane that is designed to provide a 3-dimensional supporting scaffold for the growth of exocrine acinus epithelial cells. The nanostructured membrane was produced using layer-by-layer assembly of polyelectrolytes, and formed into 3-dimensional hemispherical cavities and “finger-like” structures in order to mimic the natural architecture of acini found in exocrine glands. We utilised normal (PNT2) and cancerous (PC3, LNCaP) prostate epithelial cells to demonstrate the proof-of-concept of using MALDI (Matrix Assisted Laser Desorption Ionisation) profiling of secretions collected after 48 hours of cell growth, with no concentration or purification steps and without any a priori on the knowledge of targeted proteins. This MALDI profiling analysis of the crude supernatants from 3 different cell lines (PNT2, PC3 and LNCaP) demonstrated the capacity of the MALDI profiling approach to discriminate between the different secretome signatures. The UroLOC concept and secretome profiling that we describe opens new opportunities in terms of liquid-biopsy based diagnosis, particularly for the early stages of carcinogenesis.

Published
2018

41. Protein kinase CK2 in breast cancer: the CK2β regulatory subunit takes center stage in epithelial plasticity

Author

Sofia Giacosa, Claude Cochet, Odile Filhol, and Yann Wallez

Subjects
Epithelial-Mesenchymal Transition, Breast Neoplasms, MAP2K7, Cellular and Molecular Neuroscience, Humans, ASK1, Casein Kinase II, Molecular Biology, Pharmacology, biology, Cyclin-dependent kinase 4, Akt/PKB signaling pathway, Cyclin-dependent kinase 2, Forkhead Transcription Factors, Cell Biology, Gene Expression Regulation, Neoplastic, embryonic structures, Cyclin-dependent kinase complex, biology.protein, Cancer research, Molecular Medicine, Female, Snail Family Transcription Factors, Casein kinase 2, FOXC2, Signal Transduction, Transcription Factors
Abstract

Structurally, protein kinase CK2 consists of two catalytic subunits (α and α') and two regulatory subunits (β), which play a critical role in targeting specific CK2 substrates. Compelling evidence shows the complexity of the CK2 cellular signaling network and supports the view that this enzyme is a key component of regulatory protein kinase networks that are involved in several aspects of cancer. CK2 both activates and suppresses the expression of a number of essential oncogenes and tumor suppressors, and its expression and activity are upregulated in blood tumors and virtually all solid tumors. The prognostic significance of CK2α expression in association with various clinicopathological parameters highlighted this kinase as an adverse prognostic marker in breast cancer. In addition, several recent studies reported its implication in the regulation of the epithelial-to-mesenchymal transition (EMT), an early step in cancer invasion and metastasis. In this review, we briefly overview the contribution of CK2 to several aspects of cancer and discuss how in mammary epithelial cells, the expression of its CK2β regulatory subunit plays a critical role in maintaining an epithelial phenotype through CK2-mediated control of key EMT-related transcription factors. Importantly, decreased CK2β expression in breast tumors is correlated with inefficient phosphorylation and nuclear translocation of Snail1 and Foxc2, ultimately leading to EMT induction. This review highlights the pivotal role played by CK2β in the mammary epithelial phenotype and discusses how a modest alteration in its expression may be sufficient to induce dramatic effects facilitating the early steps in tumor cell dissemination through the coordinated regulation of two key transcription factors.

Published
2015

42. Protein kinase CK2 enables regulatory T cells to suppress excessive TH2 responses in vivo

Author

Susanne Hahn, Toszka Bohn, Hansjörg Schild, Alexander Ulges, Frauke Zipp, Nadine Grebe, Tobias Bopp, Ari Waisman, Sonja Reißig, Andreas Beilhack, Sabine Muth, Natascha Stergiou, Sebastian Reuter, Till-Julius Brühl, Matthias Klein, Markus Hoffmann, Odile Filhol-Cochet, Irma Haben, Edgar Schmitt, Hajime Yurugi, Hans Christian Probst, Krishnaraj Rajalingam, Brigitte Boldyreff, Thierry Buchou, Iris Bellinghausen, Minka Breloer, Bastian Gerlitzki, Helmut Jonuleit, Andrea Tuettenberg, and Valérie Staudt

Subjects
CD4-Positive T-Lymphocytes, Male, T cell, Immunology, Mice, Transgenic, Receptors, Cell Surface, chemical and pharmacologic phenomena, Cell Growth Processes, T-Lymphocytes, Regulatory, Cell Line, Mice, Th2 Cells, Immune system, Hypersensitivity, medicine, Animals, Humans, Immunology and Allergy, IL-2 receptor, Casein Kinase II, Mice, Inbred BALB C, Chemistry, Peripheral tolerance, FOXP3, Cell Differentiation, Forkhead Transcription Factors, Dendritic Cells, Acquired immune system, Cell biology, Mice, Inbred C57BL, medicine.anatomical_structure, Cell culture, Interferon Regulatory Factors, Leukocytes, Mononuclear, IRF4
Abstract

The quality of the adaptive immune response depends on the differentiation of distinct CD4(+) helper T cell subsets, and the magnitude of an immune response is controlled by CD4(+)Foxp3(+) regulatory T cells (Treg cells). However, how a tissue- and cell type-specific suppressor program of Treg cells is mechanistically orchestrated has remained largely unexplored. Through the use of Treg cell-specific gene targeting, we found that the suppression of allergic immune responses in the lungs mediated by T helper type 2 (TH2) cells was dependent on the activity of the protein kinase CK2. Genetic ablation of the β-subunit of CK2 specifically in Treg cells resulted in the proliferation of a hitherto-unexplored ILT3(+) Treg cell subpopulation that was unable to control the maturation of IRF4(+)PD-L2(+) dendritic cells required for the development of TH2 responses in vivo.

Published
2015

43. Protein kinase CK2 controls T-cell polarization through dendritic cell activation in response to contact sensitizers

Author

Philippe Rousseau, Yann Gallais, Zeina El Ali, Odile Filhol, Claude Cochet, Marie-Hélène Damiens, Marie de Bourayne, Marc Pallardy, Sylvie Chollet-Martin, Saadia Kerdine-Römer, Filhol, Odile, Laboratoire Matériaux et Phénomènes Quantiques (MPQ (UMR_7162)), Université Paris Diderot - Paris 7 (UPD7)-Centre National de la Recherche Scientifique (CNRS), Cytokines, chimiokines et immunopathologie, Université Paris-Sud - Paris 11 (UP11)-Institut National de la Santé et de la Recherche Médicale (INSERM), Inflammation, Chimiokines et Immunopathologie [Châtenay-Malabry], Université Paris-Sud - Paris 11 (UP11)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Paris-Saclay, Invasion mechanisms in angiogenesis and cancer (IMAC), Biologie du Cancer et de l'Infection (BCI ), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA), Laboratoire d'Immunologie 'Autoimmunité et Hypersensibilités' [AP-HP Hôpital Bichat, Paris], AP-HP - Hôpital Bichat - Claude Bernard [Paris], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP), Faculté de Pharmacie, Université Paris-Sud - Paris 11 (UP11), Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), and Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut National de la Santé et de la Recherche Médicale (INSERM)

Subjects
CD4-Positive T-Lymphocytes, 0301 basic medicine, Proteome, dendritic cell, T-Lymphocytes, T cell, Immunology, Down-Regulation, [SDV.BC]Life Sciences [q-bio]/Cellular Biology, Monocytes, Proinflammatory cytokine, 03 medical and health sciences, Nickel, Dinitrochlorobenzene, medicine, cytokine, Humans, Immunology and Allergy, RNA, Messenger, Naphthyridines, Casein Kinase II, Protein kinase A, [SDV.BC] Life Sciences [q-bio]/Cellular Biology, Cell Proliferation, CD86, CD40, biology, Cell Polarity, Cell Differentiation, Dextrans, protein kinase, Dendritic Cells, Cell Biology, Dendritic cell, 3. Good health, Cell biology, 030104 developmental biology, medicine.anatomical_structure, biology.protein, Cytokines, Phenazines, Cytokine secretion, allergic contact dermatitis, Th1/Th2/Th17 cell, Biomarkers, Fluorescein-5-isothiocyanate, CD8, Transcription Factors
Abstract

Allergic contact dermatitis (ACD) represents a severe health problem with increasing worldwide prevalence. It is a T-cell-mediated inflammatory skin disease caused by chemicals present in the daily or professional environment. NiSO4 and 2,4-dinitrochlorobenzene (DNCB) are 2 chemicals involved in ACD. These contact sensitizers are known to induce an up-regulation of phenotypic markers and cytokine secretion in dendritic cells (DCs; professional APCs), leading to the generation of CD8+ Tc1/Tc17 and CD4+ Th1/Th17 effector T cells. In the present study, using a peptide array approach, we identified protein kinase CK2 as a novel kinase involved in the activation of human monocyte-derived DCs (MoDCs) in response to NiSO4 and DNCB. Inhibition of CK2 activity in MoDCs led to an altered mature phenotype with lower expression of CD54, PDL-1, CD86, and CD40 in response to NiSO4 or DNCB. CK2 activity also regulated proinflammatory cytokine production, such as TNF-α, IL-1β, and IL-23 in MoDCs. Moreover, in a DC/T cell coculture model in an allogeneic setup, CK2 activity in MoDCs played a major role in Th1 polarization in response to NiSO4 and DNCB. CK2 inhibition in MoDCs led to an enhanced Th2 polarization in the absence of contact sensitizer stimulation.

Published
2017

44. Polarity Reversal by Centrosome Repositioning Primes Cell Scattering during Epithelial-to-Mesenchymal Transition

Author

Joanne Young, Gaëlle Letort, Mithila Burute, Thomas Bessy, Sandrine Truchet, Odile Filhol, Qingzong Tseng, Guillaume Blin, Sally Lowell, Manuel Théry, Magali Prioux, Alloimmunité-Autoimmunité-Transplantation (A2T), Institut Universitaire d'Hématologie (IUH), Université Paris Diderot - Paris 7 (UPD7)-Université Paris Diderot - Paris 7 (UPD7)-Institut National de la Santé et de la Recherche Médicale (INSERM), Physiologie cellulaire et végétale (LPCV), Institut National de la Recherche Agronomique (INRA)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA), Génétique Animale et Biologie Intégrative (GABI), Institut National de la Recherche Agronomique (INRA)-AgroParisTech, CytoMorphoLab, Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Institut National de la Recherche Agronomique (INRA)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), MRC Centre for Regenerative Medicine [Edinburgh, UK] (Institute for Stem Cell Research), University of Edinburgh-Institute for Stem Cell Research [Edinburgh, UK] (School of Biological Sciences), CYTOO SA [Grenoble, France], Invasion mechanisms in angiogenesis and cancer (IMAC), Biologie du Cancer et de l'Infection (BCI ), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), BPI France (ETICS, I1107018W), Ligue Nationale et Regionale contre le Cancer and Espoir Foundation, European Project: 310472,EC:FP7:ERC,ERC-2012-StG_20111109,SPICY(2012), Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Recherche Agronomique (INRA)-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Recherche Agronomique (INRA)-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Institute for Stem Cell Research [Edinburgh, UK] (School of Biological Sciences)-University of Edinburgh, Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut National de la Santé et de la Recherche Médicale (INSERM), Laboratoire de physiologie cellulaire végétale (LPCV), Université Joseph Fourier - Grenoble 1 (UJF)-Institut National de la Recherche Agronomique (INRA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Centre National de la Recherche Scientifique (CNRS)-Université Joseph Fourier - Grenoble 1 (UJF)-Institut National de la Recherche Agronomique (INRA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Centre National de la Recherche Scientifique (CNRS)-Laboratoire de physiologie cellulaire végétale (LPCV), Université Joseph Fourier - Grenoble 1 (UJF)-Institut National de la Recherche Agronomique (INRA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Recherche Agronomique (INRA)-Physiologie cellulaire et végétale (LPCV), Institut National de la Recherche Agronomique (INRA)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes (UGA)-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Institut National de la Recherche Agronomique (INRA)-Université Grenoble Alpes (UGA)-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), AgroParisTech-Institut National de la Recherche Agronomique (INRA), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Grenoble Alpes (UGA)-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Grenoble Alpes (UGA)-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), This work was supported by grants from the ERC (SpiCy, StG-310472) and the BPI France (ETICS, I1107018W). M.B. received a fellowship from the ANRT (CIFRE 2011/1730) and M.P. a fellowship from the Laboratory of Excellence (Grenoble Alliance for Integrated Structural Cell Biology). GB and SL are supported by fellowships from the Wellcome Trust. OFC is supported by La Ligue Nationale et Regionale contre le Cancer and Espoir Foundation., CIC - Biotherapie - Saint Louis ((CIC-BT 301)), Assistance publique - Hôpitaux de Paris (AP-HP) (APHP)-Université Paris Diderot - Paris 7 (UPD7)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Groupe Hospitalier Saint Louis - Lariboisière - Fernand Widal [Paris], Assistance publique - Hôpitaux de Paris (AP-HP) (APHP), CYTOO SA, Cytoo SA, Centre Development in Stem Cell Biology, University of Edinburgh-Institute for Stem Cell Research, Ligue Nationale et Regionale contre le Cancer and Espoir Foundation., Filhol, Odile, Spatial Integration in Cell Cytoskeleton - SPICY - - EC:FP7:ERC2012-12-01 - 2017-11-30 - 310472 - VALID, Physiologie cellulaire et végétale [2016-2019] (LPCV [2016-2019]), Institut National de la Santé et de la Recherche Médicale (INSERM)-Assistance publique - Hôpitaux de Paris (AP-HP) (APHP)-Université Paris Diderot - Paris 7 (UPD7)-Groupe Hospitalier Saint Louis - Lariboisière - Fernand Widal [Paris], and Invasion mechanisms in angiogenesis and cancer [?-2019] (IMAC [?-2019])

Subjects
0301 basic medicine, [SDV]Life Sciences [q-bio], Cell Cycle Proteins, migration, Cell junction, Mouse embryo, Microtubules, Mice, Cell polarity, polarity, Cytoskeleton, ComputingMilieux_MISCELLANEOUS, Cells, Cultured, Centrosome repositioning, biology, EMT, Cell Polarity, cytoskeleton, Epithelial to Mesenchymal Transition EMT, Cell biology, Biomechanical Phenomena, Mouse mammary gland, intercellular junctions, Mammalia, Female, Intracellular, microtubule, Epithelial-Mesenchymal Transition, stathmin, Microtubule network dynamics, Embryonic Development, Stathmin, [SDV.BC]Life Sciences [q-bio]/Cellular Biology, General Biochemistry, Genetics and Molecular Biology, Article, 03 medical and health sciences, Mammary Glands, Animal, Microtubule, [SDV.EE.SANT] Life Sciences [q-bio]/Ecology, environment/Health, Animals, Humans, Epithelial–mesenchymal transition, Molecular Biology, Adaptor Proteins, Signal Transducing, [SDV.EE.SANT]Life Sciences [q-bio]/Ecology, environment/Health, Cell Biology, micropatterning, Polarity reversals, 030104 developmental biology, centrosome, Centrosome, biology.protein, Cell Adhesion Molecules, Developmental Biology, Micropatterned cells
Abstract

International audience; During epithelial-to-mesenchymal transition (EMT), cells lining the tissue periphery break up their cohesion to migrate within the tissue. This dramatic reorganization involves a poorly characterized reorientation of the apicobasal polarity of static epithelial cells into the front-rear polarity of migrating mesenchymal cells. To investigate the spatial coordination of intra-cellular reorganization with morphological changes, we monitored centrosome positioning during EMT in vivo, in developing mouse embryos and mammary gland, and in vitro, in cultured 3D cell aggregates and micropatterned cell doublets. In all conditions, centrosomes moved from their off-centered position next to intercellular junctions toward extracellular matrix adhesions on the opposite side of the nucleus, resulting in an effective internal polarity reversal. This move appeared to be supported by controlled micro- tubule network disassembly. Sequential release of cell confinement using dynamic micropatterns, and modulation of microtubule dynamics, confirmed that centrosome repositioning was responsible for further cell disengagement and scattering.

Published
2017

45. In Search of Small Molecule Inhibitors Targeting the Flexible CK2 Subunit Interface

Author

Odile Filhol, Claude Cochet, Benoit Bestgen, Thierry Lomberget, Marc Le Borgne, Zakia Belaid-Choucair, Le Borgne, Marc, Biologie du Cancer et de l'Infection (BCI ), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA), Universität des Saarlandes [Saarbrücken], Molécules bioactives et chimie médicinale (B2MC), Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon, Imagine - Institut des maladies génétiques (IMAGINE - U1163), Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM), Invasion mechanisms in angiogenesis and cancer (IMAC), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut National de la Santé et de la Recherche Médicale (INSERM), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Grenoble Alpes (UGA)-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Centre National de la Recherche Scientifique (CNRS)-Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM), and Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Grenoble Alpes (UGA)-Institut de Recherche Interdisciplinaire de Grenoble (IRIG)

Subjects
0301 basic medicine, Fluorescence-lifetime imaging microscopy, Protein subunit, Specificity factor, [CHIM.THER] Chemical Sciences/Medicinal Chemistry, subunit interface, Pharmaceutical Science, lcsh:Medicine, lcsh:RS1-441, [SDV.CAN]Life Sciences [q-bio]/Cancer, [SDV.BC]Life Sciences [q-bio]/Cellular Biology, [CHIM.THER]Chemical Sciences/Medicinal Chemistry, Biology, Article, Protein–protein interaction, lcsh:Pharmacy and materia medica, 03 medical and health sciences, [SDV.CAN] Life Sciences [q-bio]/Cancer, Drug Discovery, ComputingMilieux_MISCELLANEOUS, chemistry.chemical_classification, protein kinase CK2, cyclic peptides, protein–protein interaction, cell death, lcsh:R, fungi, [SDV.SP]Life Sciences [q-bio]/Pharmaceutical sciences, Small molecule, Cyclic peptide, [SDV.SP] Life Sciences [q-bio]/Pharmaceutical sciences, 030104 developmental biology, chemistry, Biochemistry, Biophysics, Molecular Medicine, Phosphorylation, Signal transduction
Abstract

International audience; Protein kinase CK2 is a tetrameric holoenzyme composed of two catalytic (α and/or α') subunits and two regulatory (β) subunits. Crystallographic data paired with fluorescence imaging techniques have suggested that the formation of the CK2 holoenzyme complex within cells is a dynamic process. Although the monomeric CK2α subunit is endowed with a constitutive catalytic activity, many of the plethora of CK2 substrates are exclusively phosphorylated by the CK2 holoenzyme. This means that the spatial and high affinity interaction between CK2α and CK2β subunits is critically important and that its disruption may provide a powerful and selective way to block the phosphorylation of substrates requiring the presence of CK2β. In search of compounds inhibiting this critical protein–protein interaction, we previously designed an active cyclic peptide (Pc) derived from the CK2β carboxy-terminal domain that can efficiently antagonize the CK2 subunit interaction. To understand the functional significance of this interaction, we generated cell-permeable versions of Pc, exploring its molecular mechanisms of action and the perturbations of the signaling pathways that it induces in intact cells. The identification of small molecules inhibitors of this critical interaction may represent the first-choice approach to manipulate CK2 in an unconventional way.

Published
2017

46. Heterogeneity of single-cell mechanical responses to tumorigenic factors

Author

Manuel Théry, Jean-Louis Martiel, Aldo Leal-Egaña, Odile Filhol, Andreas Christ, Timothée Vignaud, Caroline Roelants, and Gaëlle Letort

Subjects
0303 health sciences, education.field_of_study, biology, Cell growth, Population, Cell, Transforming growth factor beta, Anatomy, Cell biology, Fibronectin, Contractility, 03 medical and health sciences, 0302 clinical medicine, medicine.anatomical_structure, Cell culture, 030220 oncology & carcinogenesis, medicine, biology.protein, Casein kinase 2, education, 030304 developmental biology
Abstract

Tumor development progresses through a complex path of biomechanical changes leading first to cell growth and contraction followed by cell de-adhesion, scattering and invasion. Tumorigenic factors may act specifically on one of these steps or have wider spectrum of actions, leading to a variety of effects and thus sometimes to apparent contradictory outcomes. Here we used micropatterned lines of collagen type-I/fibronectin on deformable surfaces to standardize cell behavior and to measure simultaneously cell size, speed of motion and the magnitude of the associated contractile forces at the level of a single cell. We analyzed and compared normal human breast cell line MCF10A in control conditions and in response to various tumorigenic factors. In all conditions, distinct populations of cells with a wide range of biomechanical properties were identified. Despite this heterogeneity, normal and transformed motile cells followed a common trend whereby size and contractile forces were negatively correlated with cell speed. Some tumorigenic factors, such as activation of ErbB2 or the loss of the beta subunit of casein kinase 2 (CK2), shifted the whole population towards a faster speed and lower contractility state. Treatment with transforming growth factor beta (TGF-β), induced some cells to adopt opposing behaviors such as extreme high contractility versus extreme low contractility. Thus, tumor transformation amplified the pre-existing population heterogeneity and led some cells to exhibit biomechanical properties that were more extreme than that observed with normal cells.

Published
2016

47. A 'DropChip' cell array for DNA and siRNA transfection combined with drug screening

Author

Francois Chatelain, Sandra Boccard, Julien Reboud, Béatrice Schaack, François Berger, Brigitte Fouque, Stéphanie Combe, and Odile Filhol-Cochet

Subjects
Cell type, Microarray, Cell, Biomedical Engineering, Bioengineering, Transfection, Biology, Cell biology, medicine.anatomical_structure, Cell culture, RNA interference, High-content screening, medicine, Nucleic acid, Molecular Biology
Abstract

We have developed a "DropChip" microarray for multiplexed cell-based assays to perform information-rich high-throughput screening. This microarray features cell culture nanoliter droplets on a glass slide; the culture's size, shape, and location are accurately controlled by surface tension. We report parallel gene expression and gene silencing analyses on the same array, combining nucleic acid transfection, anti-cancer drug screening, and cell cultures. In particular, we analyzed the synergic effects of siRNA and cisplatin on cancerous cells. With up to 100 cells per drop, we could carry out the detection and analysis of cell behavior at the individual cell level, using high-resolution fluorescence microscopy and automated image analysis. Nucleic and chemical compounds require early multiplexed testing using nanomolar quantities. The NanoDrop technology is a unique solution to perform cell-based assay in nanoliter drops, providing a major decrease of cost and time, as well as a precise description of the behavior of several cell types in a single test at the single cell level. This novel cell array format could enable highly informative functional genomic studies and large scale in vitro toxicity testing. Copyright © 2005 Humana Press Inc. All rights of any nature whatsoever are reserved.

Published
2016

48. Structure–function analysis of the beta regulatory subunit of protein kinase CK2 by targeting embryonic stem cell

Author

Béatrice Laudet, Claude Cochet, Odile Filhol, Léa Ziercher, Thierry Buchou, and Renaud Prudent

Subjects
Homeobox protein NANOG, Cell Survival, Cellular differentiation, Rex1, Clinical Biochemistry, Biology, Mice, Structure-Activity Relationship, Neural Stem Cells, Catalytic Domain, Neurosphere, Animals, Cell Lineage, Progenitor cell, Casein Kinase II, Molecular Biology, Cell potency, Embryonic Stem Cells, Cell Proliferation, Teratoma, Cell Differentiation, Cell Biology, General Medicine, Embryonic stem cell, Cell biology, Oligodendroglia, Mutation, Mutant Proteins, Stem cell
Abstract

Programs that govern stem cell maintenance and pluripotency are dependent on extracellular factors and of intrinsic cell modulators. Embryonic stem (ES) cells with a specific depletion of the gene encoding the regulatory subunit of protein kinase CK2 (CK2β) revealed a viability defect. However, analysis of CK2β functions along the neural lineage established CK2β as a positive regulator for neural stem/progenitor cell (NSC) proliferation and multipotency. By using an in vitro genetic conditional approach, we demonstrate in this work that specific domains of CK2β involved in the regulatory function towards CK2 catalytic subunits are crucial structural determinants for ES cell homeostasis.

Published
2011

49. Regulation of epithelial to mesenchymal transition: CK2β on stage

Author

Alexandre Deshiere, Claude Cochet, Eve Duchemin-Pelletier, Odile Filhol, Elodie Spreux, Christelle Forcet, and Delphine Ciais

Subjects
Epithelial-Mesenchymal Transition, Clinical Biochemistry, Biology, Cell morphology, medicine.disease_cause, Cell Line, Mice, Transforming Growth Factor beta2, Cell polarity, Cell Adhesion, medicine, Animals, Humans, Gene silencing, Epithelial–mesenchymal transition, RNA, Small Interfering, Casein Kinase II, Cell adhesion, Cell Shape, Molecular Biology, Cell Polarity, Epithelial Cells, Cell Biology, General Medicine, Phenotype, Up-Regulation, Cell biology, Gene Knockdown Techniques, embryonic structures, NIH 3T3 Cells, Signal transduction, Carcinogenesis
Abstract

Protein kinase CK2 participates in the regulation of fundamental cellular processes. Among these processes, cell polarity and cell morphology are controlled by this enzyme probably through the phosphorylation of key proteins. To further study the involvement of CK2 in these processes, we showed that in epithelial cells, the regulatory CK2β subunit was required for LKB1-dependent polarization and cell adhesion. Moreover, CK2β silencing in MCF10A mammary epithelial cells triggered changes in their morphology correlated with the acquisition of mesenchymal phenotype, which were reminiscent to TGFβ-induced epithelial-to-mesenchymal-transition (EMT). TGFβ has emerged as a major inducer of EMT both in vitro and in vivo. We found that among the TGFβ isoforms, TGFβ2 expression was strongly induced in CK2β-knockdown cells. However, the EMT phenotype induced in response to CK2β silencing was not abolished by blocking the TGFβ signaling pathway at TGFβ receptor level, suggesting that alternative pathways might be involved. Given the importance of CK2 in tumorigenesis, a dysregulation of CK2β expression might contribute to EMT induction during cancer progression.

Published
2011

50. Protein kinase CK2α subunit over-expression correlates with metastatic risk in breast carcinomas: Quantitative immunohistochemistry in tissue microarrays

Author

Stéphane Garcia, Xavier Carcopino, Juan L. Iovanna, Colette Charpin, Odile Filhol, Véronique Secq, Daniel Birnbaum, Sophie Giusiano, Pascal Bonnier, Eve Duchemin-Pelletier, Claude Cochet, and Léon Boubli

Subjects
Cancer Research, Univariate analysis, Tissue microarray, Cancer, Breast Neoplasms, Biology, Prognosis, medicine.disease, Immunohistochemistry, Metastasis, Breast cancer, ROC Curve, Oncology, Risk Factors, Tissue Array Analysis, Biomarkers, Tumor, medicine, Cancer research, Humans, Female, Breast disease, Neoplasm Metastasis, Casein Kinase II, Breast carcinoma
Abstract

CK2α is a signalling molecule that participates in major events in solid tumour progression. The aim of this study was to evaluate the prognostic significance of the immunohistochemical expression of CK2α in breast carcinomas.Quantitative measurements of immunohistochemical expression of 33 biomarkers using high-throughput densitometry, assessed on digitised microscopic tissue micro-array images were correlated with clinical outcome in 1000 breast carcinomas using univariate and multivariate analyses.In univariate analysis, CK2α was a significant prognostic indicator (p0.001). Moreover, a multivariable model allowed the selection of the best combination of the 33 biomarkers to predict patients' outcome through logistic regression. A nine-marker signature highly predictive of metastatic risk, associating SHARP-2, STAT1, eIF4E, pmapKAPk-2, pAKT, caveolin, VEGF, FGF-1 and CK2α permitted to classify well 82.32% of patients (specificity 81.59%, sensitivity 92.55%, area under ROC curve 0.939). Importantly, in a node negative subset of patients an even more (86%) clinically relevant association of eleven markers was found predictive of poor outcome.A strong quantitative CK2α immunohistochemical expression in breast carcinomas is individually a significant indicator of poor prognosis. Moreover, an immunohistochemical signature of 11 markers including CK2α accurately (86%) well classifies node negative patients in good and poor outcome subsets. Our results suggest that CK2α evaluation together with key downstream CK2 targets might be a useful tool to identify patients at high risk of distant metastases and that CK2 can be considered as a relevant target for potential specific therapy.

Published
2011

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