Publications by authors named "Charles-Hugues Lardeau"

12 Publications

  • Page 1 of 1

A widespread role for SLC transmembrane transporters in resistance to cytotoxic drugs.

Nat Chem Biol 2020 04 9;16(4):469-478. Epub 2020 Mar 9.

CeMM Research Center for Molecular Medicine of the Austrian Academy of Sciences, Vienna, Austria.

Solute carriers (SLCs) are the largest family of transmembrane transporters in humans and are major determinants of cellular metabolism. Several SLCs have been shown to be required for the uptake of chemical compounds into cellular systems, but systematic surveys of transporter-drug relationships in human cells are currently lacking. We performed a series of genetic screens in a haploid human cell line against 60 cytotoxic compounds representative of the chemical space populated by approved drugs. By using an SLC-focused CRISPR-Cas9 library, we identified transporters whose absence induced resistance to the drugs tested. This included dependencies involving the transporters SLC11A2/SLC16A1 for artemisinin derivatives and SLC35A2/SLC38A5 for cisplatin. The functional dependence on SLCs observed for a significant proportion of the screened compounds suggests a widespread role for SLCs in the uptake and cellular activity of cytotoxic drugs and provides an experimentally validated set of SLC-drug associations for a number of clinically relevant compounds.
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http://dx.doi.org/10.1038/s41589-020-0483-3DOI Listing
April 2020

Mapping the perturbome network of cellular perturbations.

Nat Commun 2019 11 13;10(1):5140. Epub 2019 Nov 13.

CeMM Research Center for Molecular Medicine of the Austrian Academy of Sciences, Lazarettgasse 14, AKH BT 25.3, A-1090, Vienna, Austria.

Drug combinations provide effective treatments for diverse diseases, but also represent a major cause of adverse reactions. Currently there is no systematic understanding of how the complex cellular perturbations induced by different drugs influence each other. Here, we introduce a mathematical framework for classifying any interaction between perturbations with high-dimensional effects into 12 interaction types. We apply our framework to a large-scale imaging screen of cell morphology changes induced by diverse drugs and their combination, resulting in a perturbome network of 242 drugs and 1832 interactions. Our analysis of the chemical and biological features of the drugs reveals distinct molecular fingerprints for each interaction type. We find a direct link between drug similarities on the cell morphology level and the distance of their respective protein targets within the cellular interactome of molecular interactions. The interactome distance is also predictive for different types of drug interactions.
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http://dx.doi.org/10.1038/s41467-019-13058-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6853941PMC
November 2019

Mapping the Human Kinome in Response to DNA Damage.

Cell Rep 2019 01;26(3):555-563.e6

CeMM Research Center for Molecular Medicine of the Austrian Academy of Sciences, Lazarettgasse 14, AKH BT 25.3, 1090 Vienna, Austria. Electronic address:

We provide a catalog for the effects of the human kinome on cell survival in response to DNA-damaging agents, covering all major DNA repair pathways. By treating 313 kinase-deficient cell lines with ten diverse DNA-damaging agents, including seven commonly used chemotherapeutics, we identified examples of vulnerability and resistance that are kinase specific. To investigate synthetic lethal interactions, we tested the response to carmustine for 25 cell lines by establishing a phenotypic fluorescence-activated cell sorting (FACS) assay designed to validate gene-drug interactions. We show apoptosis, cell cycle changes, and DNA damage and proliferation after alkylation- or crosslink-induced damage. In addition, we reconstitute the cellular sensitivity of DYRK4, EPHB6, MARK3, and PNCK as a proof of principle for our study. Furthermore, using global phosphoproteomics on cells lacking MARK3, we provide evidence for its role in the DNA damage response. Our data suggest that cancers with inactivating mutations in kinases, including MARK3, are particularly vulnerable to alkylating chemotherapeutic agents.
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http://dx.doi.org/10.1016/j.celrep.2018.12.087DOI Listing
January 2019

Pharmacologic inhibition of STAT5 in acute myeloid leukemia.

Leukemia 2018 05 2;32(5):1135-1146. Epub 2018 Feb 2.

Ludwig Boltzmann Institute for Cancer Research, Vienna, Austria.

The transcription factor STAT5 is an essential downstream mediator of many tyrosine kinases (TKs), particularly in hematopoietic cancers. STAT5 is activated by FLT3-ITD, which is a constitutively active TK driving the pathogenesis of acute myeloid leukemia (AML). Since STAT5 is a critical mediator of diverse malignant properties of AML cells, direct targeting of STAT5 is of significant clinical value. Here, we describe the development and preclinical evaluation of a novel, potent STAT5 SH2 domain inhibitor, AC-4-130, which can efficiently block pathological levels of STAT5 activity in AML. AC-4-130 directly binds to STAT5 and disrupts STAT5 activation, dimerization, nuclear translocation, and STAT5-dependent gene transcription. Notably, AC-4-130 substantially impaired the proliferation and clonogenic growth of human AML cell lines and primary FLT3-ITD AML patient cells in vitro and in vivo. Furthermore, AC-4-130 synergistically increased the cytotoxicity of the JAK1/2 inhibitor Ruxolitinib and the p300/pCAF inhibitor Garcinol. Overall, the synergistic effects of AC-4-130 with TK inhibitors (TKIs) as well as emerging treatment strategies provide new therapeutic opportunities for leukemia and potentially other cancers.
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http://dx.doi.org/10.1038/s41375-017-0005-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5940656PMC
May 2018

Repair of UV-Induced DNA Damage Independent of Nucleotide Excision Repair Is Masked by MUTYH.

Mol Cell 2017 Nov;68(4):797-807.e7

CeMM Research Center for Molecular Medicine of the Austrian Academy of Sciences, Lazarettgasse 14, AKH BT 25.3, 1090 Vienna, Austria. Electronic address:

DNA lesions caused by UV damage are thought to be repaired solely by the nucleotide excision repair (NER) pathway in human cells. Patients carrying mutations within genes functioning in this pathway display a range of pathologies, including an increased susceptibility to cancer, premature aging, and neurological defects. There are currently no curative therapies available. Here we performed a high-throughput chemical screen for agents that could alleviate the cellular sensitivity of NER-deficient cells to UV-induced DNA damage. This led to the identification of the clinically approved anti-diabetic drug acetohexamide, which promoted clearance of UV-induced DNA damage without the accumulation of chromosomal aberrations, hence promoting cellular survival. Acetohexamide exerted this protective function by antagonizing expression of the DNA glycosylase, MUTYH. Together, our data reveal the existence of an NER-independent mechanism to remove UV-induced DNA damage and prevent cell death.
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http://dx.doi.org/10.1016/j.molcel.2017.10.021DOI Listing
November 2017

A combinatorial screen of the CLOUD uncovers a synergy targeting the androgen receptor.

Nat Chem Biol 2017 Jul 22;13(7):771-778. Epub 2017 May 22.

CeMM Research Center for Molecular Medicine of the Austrian Academy of Sciences, Vienna, Austria.

Approved drugs are invaluable tools to study biochemical pathways, and further characterization of these compounds may lead to repurposing of single drugs or combinations. Here we describe a collection of 308 small molecules representing the diversity of structures and molecular targets of all FDA-approved chemical entities. The CeMM Library of Unique Drugs (CLOUD) covers prodrugs and active forms at pharmacologically relevant concentrations and is ideally suited for combinatorial studies. We screened pairwise combinations of CLOUD drugs for impairment of cancer cell viability and discovered a synergistic interaction between flutamide and phenprocoumon (PPC). The combination of these drugs modulates the stability of the androgen receptor (AR) and resensitizes AR-mutant prostate cancer cells to flutamide. Mechanistically, we show that the AR is a substrate for γ-carboxylation, a post-translational modification inhibited by PPC. Collectively, our data suggest that PPC could be repurposed to tackle resistance to antiandrogens in prostate cancer patients.
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http://dx.doi.org/10.1038/nchembio.2382DOI Listing
July 2017

Global survey of the immunomodulatory potential of common drugs.

Nat Chem Biol 2017 06 24;13(6):681-690. Epub 2017 Apr 24.

CeMM Research Center for Molecular Medicine of the Austrian Academy of Sciences, Vienna, Austria.

Small-molecule drugs may complement antibody-based therapies in an immune-oncology setting, yet systematic methods for the identification and characterization of the immunomodulatory properties of these entities are lacking. We surveyed the immumomodulatory potential of 1,402 small chemical molecules, as defined by their ability to alter the cell-cell interactions among peripheral mononuclear leukocytes ex vivo, using automated microscopy and population-wide single-cell image analysis. Unexpectedly, ∼10% of the agents tested affected these cell-cell interactions differentially. The results accurately recapitulated known immunomodulatory drug classes and revealed several clinically approved drugs that unexpectedly harbor the ability to modulate the immune system, which could potentially contribute to their physiological mechanism of action. For instance, the kinase inhibitor crizotinib promoted T cell interactions with monocytes, as well as with cancer cells, through inhibition of the receptor tyrosine kinase MSTR1 and subsequent upregulation of the expression of major histocompatibility complex molecules. The approach offers an attractive platform for the personalized identification and characterization of immunomodulatory therapeutics.
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http://dx.doi.org/10.1038/nchembio.2360DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5438060PMC
June 2017

Artemisinins Target GABA Receptor Signaling and Impair α Cell Identity.

Cell 2017 Jan 1;168(1-2):86-100.e15. Epub 2016 Dec 1.

CeMM Research Center for Molecular Medicine of the Austrian Academy of Sciences. Lazarettgasse 14, 1090 Vienna, Austria.

Type 1 diabetes is characterized by the destruction of pancreatic β cells, and generating new insulin-producing cells from other cell types is a major aim of regenerative medicine. One promising approach is transdifferentiation of developmentally related pancreatic cell types, including glucagon-producing α cells. In a genetic model, loss of the master regulatory transcription factor Arx is sufficient to induce the conversion of α cells to functional β-like cells. Here, we identify artemisinins as small molecules that functionally repress Arx by causing its translocation to the cytoplasm. We show that the protein gephyrin is the mammalian target of these antimalarial drugs and that the mechanism of action of these molecules depends on the enhancement of GABA receptor signaling. Our results in zebrafish, rodents, and primary human pancreatic islets identify gephyrin as a druggable target for the regeneration of pancreatic β cell mass from α cells.
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http://dx.doi.org/10.1016/j.cell.2016.11.010DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5236063PMC
January 2017

Heme drives hemolysis-induced susceptibility to infection via disruption of phagocyte functions.

Nat Immunol 2016 Dec 31;17(12):1361-1372. Epub 2016 Oct 31.

CeMM Research Center for Molecular Medicine of the Austrian Academy of Sciences, Vienna, Austria.

Hemolysis drives susceptibility to bacterial infections and predicts poor outcome from sepsis. These detrimental effects are commonly considered to be a consequence of heme-iron serving as a nutrient for bacteria. We employed a Gram-negative sepsis model and found that elevated heme levels impaired the control of bacterial proliferation independently of heme-iron acquisition by pathogens. Heme strongly inhibited phagocytosis and the migration of human and mouse phagocytes by disrupting actin cytoskeletal dynamics via activation of the GTP-binding Rho family protein Cdc42 by the guanine nucleotide exchange factor DOCK8. A chemical screening approach revealed that quinine effectively prevented heme effects on the cytoskeleton, restored phagocytosis and improved survival in sepsis. These mechanistic insights provide potential therapeutic targets for patients with sepsis or hemolytic disorders.
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http://dx.doi.org/10.1038/ni.3590DOI Listing
December 2016

Acquired nintedanib resistance in FGFR1-driven small cell lung cancer: role of endothelin-A receptor-activated ABCB1 expression.

Oncotarget 2016 Aug;7(31):50161-50179

Institute of Cancer Research, Department of Medicine I, Medical University of Vienna, Austria.

Genomically amplified fibroblast growth factor receptor 1 (FGFR1) is an oncogenic driver in defined lung cancer subgroups and predicts sensibility against FGFR1 inhibitors in this patient cohort. The FGFR inhibitor nintedanib has recently been approved for treatment of lung adenocarcinoma and is currently evaluated for small cell lung cancer (SCLC). However, tumor recurrence due to development of nintedanib resistance might occur. Hence, we aimed at characterizing the molecular mechanisms underlying acquired nintedanib resistance in FGFR1-driven lung cancer. Chronic nintedanib exposure of the FGFR1-driven SCLC cell line DMS114 (DMS114/NIN) but not of two NSCLC cell lines induced massive overexpression of the multidrug-resistance transporter ABCB1. Indeed, we proved nintedanib to be both substrate and modulator of ABCB1-mediated efflux. Importantly, the oncogenic FGFR1 signaling axis remained active in DMS114/NIN cells while bioinformatic analyses suggested hyperactivation of the endothelin-A receptor (ETAR) signaling axis. Indeed, ETAR inhibition resensitized DMS114/NIN cells against nintedanib by downregulation of ABCB1 expression. PKC and downstream NFκB were identified as major downstream players in ETAR-mediated ABCB1 hyperactivation. Summarizing, ABCB1 needs to be considered as a factor underlying nintedanib resistance. Combination approaches with ETAR antagonists or switching to non-ABCB1 substrate FGFR inhibitors represent innovative strategies to manage nintedanib resistance in lung cancer.
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http://dx.doi.org/10.18632/oncotarget.10324DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5226575PMC
August 2016

Mapping the chemical chromatin reactivation landscape identifies BRD4-TAF1 cross-talk.

Nat Chem Biol 2016 07 9;12(7):504-10. Epub 2016 May 9.

CeMM Research Center for Molecular Medicine of the Austrian Academy of Sciences, Vienna, Austria.

Bromodomain-containing proteins of the BET family recognize histone lysine acetylation and mediate transcriptional activation of target genes such as the MYC oncogene. Pharmacological inhibitors of BET domains promise therapeutic benefits in a variety of cancers. We performed a high-diversity chemical compound screen for agents capable of modulating BRD4-dependent heterochromatization of a generic reporter in human cells. In addition to known and new compounds targeting BRD4, we identified small molecules that mimic BRD4 inhibition without direct engagement. One such compound was a potent inhibitor of the second bromodomain of TAF1. Using this inhibitor, we discovered that TAF1 synergizes with BRD4 to control proliferation of cancer cells, making TAF1 an attractive epigenetic target in cancers driven by MYC.
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http://dx.doi.org/10.1038/nchembio.2080DOI Listing
July 2016

Quantitative proteomic analysis of A549 cells infected with human respiratory syncytial virus.

Mol Cell Proteomics 2010 Nov 20;9(11):2438-59. Epub 2010 Jul 20.

Institute of Molecular and Cellular Biology, Faculty of Biological Sciences, University of Leeds, Leeds, United Kingdom.

Human respiratory syncytial virus (HRSV) is a major cause of pediatric lower respiratory tract disease to which there is no vaccine or efficacious chemotherapeutic strategy. Although RNA synthesis and virus assembly occur in the cytoplasm, HRSV is known to induce nuclear responses in the host cell as replication alters global gene expression. Quantitative proteomics was used to take an unbiased overview of the protein changes in transformed human alveolar basal epithelial cells infected with HRSV. Underpinning this was the use of stable isotope labeling with amino acids in cell culture coupled to LC-MS/MS, which allowed the direct and simultaneous identification and quantification of both cellular and viral proteins. To reduce sample complexity and increase data return on potential protein localization, cells were fractionated into nuclear and cytoplasmic extracts. This resulted in the identification of 1,140 cellular proteins and six viral proteins. The proteomics data were analyzed using Ingenuity Pathways Analysis to identify defined canonical pathways and functional groupings. Selected data were validated using Western blot, direct and indirect immunofluorescence confocal microscopy, and functional assays. The study served to validate and expand upon known HRSV-host cell interactions, including those associated with the antiviral response and alterations in subnuclear structures such as the nucleolus and ND10 (promyelocytic leukemia bodies). In addition, novel changes were observed in mitochondrial proteins and functions, cell cycle regulatory molecules, nuclear pore complex proteins and nucleocytoplasmic trafficking proteins. These data shed light into how the cell is potentially altered to create conditions more favorable for infection. Additionally, the study highlights the application and advantage of stable isotope labeling with amino acids in cell culture coupled to LC-MS/MS for the analysis of virus-host interactions.
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http://dx.doi.org/10.1074/mcp.M110.001859DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2984239PMC
November 2010