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Guillermo Velasco

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DOI: 10.15252/embj.201490784
2015
Cited 993 times
Autophagy in malignant transformation and cancer progression
Review23 February 2015free access Autophagy in malignant transformation and cancer progression Lorenzo Galluzzi Corresponding Author Lorenzo Galluzzi Equipe 11 labellisée pas la Ligue Nationale contre le Cancer, Centre de Recherche des Cordeliers, Paris, France INSERM, U1138, Paris, France Gustave Roussy Cancer Campus, Villejuif, France Université Paris Descartes, Sorbonne Paris Cité, Paris, France Search for more papers by this author Federico Pietrocola Federico Pietrocola Equipe 11 labellisée pas la Ligue Nationale contre le Cancer, Centre de Recherche des Cordeliers, Paris, France INSERM, U1138, Paris, France Gustave Roussy Cancer Campus, Villejuif, France Search for more papers by this author José Manuel Bravo-San Pedro José Manuel Bravo-San Pedro Equipe 11 labellisée pas la Ligue Nationale contre le Cancer, Centre de Recherche des Cordeliers, Paris, France INSERM, U1138, Paris, France Gustave Roussy Cancer Campus, Villejuif, France Search for more papers by this author Ravi K Amaravadi Ravi K Amaravadi Abramson Cancer Center, University of Pennsylvania, Philadelphia, PA, USA Search for more papers by this author Eric H Baehrecke Eric H Baehrecke Department of Molecular, Cell and Cancer Biology, University of Massachusetts Medical School, Worcester, MA, USA Search for more papers by this author Francesco Cecconi Francesco Cecconi Cell Stress and Survival Unit, Danish Cancer Society Research Center, Copenhagen, Denmark IRCCS Fondazione Santa Lucia and Department of Biology, University of Rome Tor Vergata, Rome, Italy Search for more papers by this author Patrice Codogno Patrice Codogno Université Paris Descartes, Sorbonne Paris Cité, Paris, France Institut Necker Enfants-Malades (INEM), Paris, France INSERM, U1151, Paris, France CNRS, UMR8253, Paris, France Search for more papers by this author Jayanta Debnath Jayanta Debnath Department of Pathology and Helen Diller Family Comprehensive Cancer Center, University of California San Francisco, San Francisco, CA, USA Search for more papers by this author David A Gewirtz David A Gewirtz Department of Pharmacology, Toxicology and Medicine, Virginia Commonwealth University, Richmond, Virginia, VA, USA Search for more papers by this author Vassiliki Karantza Vassiliki Karantza Merck Research Laboratories, Rahway, NJ, USA Search for more papers by this author Alec Kimmelman Alec Kimmelman Division of Genomic Stability and DNA Repair, Department of Radiation Oncology, Dana-Farber Cancer Institute, Boston, MA, USA Search for more papers by this author Sharad Kumar Sharad Kumar Centre for Cancer Biology, University of South Australia, Adelaide, SA, Australia Search for more papers by this author Beth Levine Beth Levine Center for Autophagy Research, Department of Internal Medicine, University of Texas Southwestern Medical Center, Dallas, TX, USA Department of Microbiology, University of Texas Southwestern Medical Center, Dallas, TX, USA Howard Hughes Medical Institute, University of Texas Southwestern Medical Center, Dallas, TX, USA Search for more papers by this author Maria Chiara Maiuri Maria Chiara Maiuri Equipe 11 labellisée pas la Ligue Nationale contre le Cancer, Centre de Recherche des Cordeliers, Paris, France INSERM, U1138, Paris, France Gustave Roussy Cancer Campus, Villejuif, France Search for more papers by this author Seamus J Martin Seamus J Martin Department of Genetics, Trinity College, The Smurfit Institute, Dublin, Ireland Search for more papers by this author Josef Penninger Josef Penninger Institute of Molecular Biotechnology of the Austrian Academy of Sciences, Vienna, Austria Search for more papers by this author Mauro Piacentini Mauro Piacentini Department of Biology, University of Rome Tor Vergata, Rome, Italy National Institute for Infectious Diseases IRCCS ‘Lazzaro Spallanzani’, Rome, Italy Search for more papers by this author David C Rubinsztein David C Rubinsztein Department of Medical Genetics, Cambridge Institute for Medical Research, University of Cambridge, Cambridge, UK Search for more papers by this author Hans-Uwe Simon Hans-Uwe Simon Institute of Pharmacology, University of Bern, Bern, Switzerland Search for more papers by this author Anne Simonsen Anne Simonsen Institute of Basic Medical Sciences, University of Oslo, Oslo, Norway Search for more papers by this author Andrew M Thorburn Andrew M Thorburn Department of Pharmacology, University of Colorado School of Medicine, Aurora, CO, USA Search for more papers by this author Guillermo Velasco Guillermo Velasco Department of Biochemistry and Molecular Biology I, School of Biology, Complutense University of Madrid, Madrid, Spain Instituto de Investigaciones Sanitarias San Carlos (IdISSC), Madrid, Spain Search for more papers by this author Kevin M Ryan Kevin M Ryan Cancer Research UK Beatson Institute, Glasgow, UK Search for more papers by this author Guido Kroemer Guido Kroemer Equipe 11 labellisée pas la Ligue Nationale contre le Cancer, Centre de Recherche des Cordeliers, Paris, France INSERM, U1138, Paris, France Université Paris Descartes, Sorbonne Paris Cité, Paris, France Pôle de Biologie, Hôpital Européen Georges Pompidou, AP-HP, Paris, France Metabolomics and Cell Biology Platforms, Gustave Roussy Cancer Campus, Villejuif, France Search for more papers by this author Lorenzo Galluzzi Corresponding Author Lorenzo Galluzzi Equipe 11 labellisée pas la Ligue Nationale contre le Cancer, Centre de Recherche des Cordeliers, Paris, France INSERM, U1138, Paris, France Gustave Roussy Cancer Campus, Villejuif, France Université Paris Descartes, Sorbonne Paris Cité, Paris, France Search for more papers by this author Federico Pietrocola Federico Pietrocola Equipe 11 labellisée pas la Ligue Nationale contre le Cancer, Centre de Recherche des Cordeliers, Paris, France INSERM, U1138, Paris, France Gustave Roussy Cancer Campus, Villejuif, France Search for more papers by this author José Manuel Bravo-San Pedro José Manuel Bravo-San Pedro Equipe 11 labellisée pas la Ligue Nationale contre le Cancer, Centre de Recherche des Cordeliers, Paris, France INSERM, U1138, Paris, France Gustave Roussy Cancer Campus, Villejuif, France Search for more papers by this author Ravi K Amaravadi Ravi K Amaravadi Abramson Cancer Center, University of Pennsylvania, Philadelphia, PA, USA Search for more papers by this author Eric H Baehrecke Eric H Baehrecke Department of Molecular, Cell and Cancer Biology, University of Massachusetts Medical School, Worcester, MA, USA Search for more papers by this author Francesco Cecconi Francesco Cecconi Cell Stress and Survival Unit, Danish Cancer Society Research Center, Copenhagen, Denmark IRCCS Fondazione Santa Lucia and Department of Biology, University of Rome Tor Vergata, Rome, Italy Search for more papers by this author Patrice Codogno Patrice Codogno Université Paris Descartes, Sorbonne Paris Cité, Paris, France Institut Necker Enfants-Malades (INEM), Paris, France INSERM, U1151, Paris, France CNRS, UMR8253, Paris, France Search for more papers by this author Jayanta Debnath Jayanta Debnath Department of Pathology and Helen Diller Family Comprehensive Cancer Center, University of California San Francisco, San Francisco, CA, USA Search for more papers by this author David A Gewirtz David A Gewirtz Department of Pharmacology, Toxicology and Medicine, Virginia Commonwealth University, Richmond, Virginia, VA, USA Search for more papers by this author Vassiliki Karantza Vassiliki Karantza Merck Research Laboratories, Rahway, NJ, USA Search for more papers by this author Alec Kimmelman Alec Kimmelman Division of Genomic Stability and DNA Repair, Department of Radiation Oncology, Dana-Farber Cancer Institute, Boston, MA, USA Search for more papers by this author Sharad Kumar Sharad Kumar Centre for Cancer Biology, University of South Australia, Adelaide, SA, Australia Search for more papers by this author Beth Levine Beth Levine Center for Autophagy Research, Department of Internal Medicine, University of Texas Southwestern Medical Center, Dallas, TX, USA Department of Microbiology, University of Texas Southwestern Medical Center, Dallas, TX, USA Howard Hughes Medical Institute, University of Texas Southwestern Medical Center, Dallas, TX, USA Search for more papers by this author Maria Chiara Maiuri Maria Chiara Maiuri Equipe 11 labellisée pas la Ligue Nationale contre le Cancer, Centre de Recherche des Cordeliers, Paris, France INSERM, U1138, Paris, France Gustave Roussy Cancer Campus, Villejuif, France Search for more papers by this author Seamus J Martin Seamus J Martin Department of Genetics, Trinity College, The Smurfit Institute, Dublin, Ireland Search for more papers by this author Josef Penninger Josef Penninger Institute of Molecular Biotechnology of the Austrian Academy of Sciences, Vienna, Austria Search for more papers by this author Mauro Piacentini Mauro Piacentini Department of Biology, University of Rome Tor Vergata, Rome, Italy National Institute for Infectious Diseases IRCCS ‘Lazzaro Spallanzani’, Rome, Italy Search for more papers by this author David C Rubinsztein David C Rubinsztein Department of Medical Genetics, Cambridge Institute for Medical Research, University of Cambridge, Cambridge, UK Search for more papers by this author Hans-Uwe Simon Hans-Uwe Simon Institute of Pharmacology, University of Bern, Bern, Switzerland Search for more papers by this author Anne Simonsen Anne Simonsen Institute of Basic Medical Sciences, University of Oslo, Oslo, Norway Search for more papers by this author Andrew M Thorburn Andrew M Thorburn Department of Pharmacology, University of Colorado School of Medicine, Aurora, CO, USA Search for more papers by this author Guillermo Velasco Guillermo Velasco Department of Biochemistry and Molecular Biology I, School of Biology, Complutense University of Madrid, Madrid, Spain Instituto de Investigaciones Sanitarias San Carlos (IdISSC), Madrid, Spain Search for more papers by this author Kevin M Ryan Kevin M Ryan Cancer Research UK Beatson Institute, Glasgow, UK Search for more papers by this author Guido Kroemer Guido Kroemer Equipe 11 labellisée pas la Ligue Nationale contre le Cancer, Centre de Recherche des Cordeliers, Paris, France INSERM, U1138, Paris, France Université Paris Descartes, Sorbonne Paris Cité, Paris, France Pôle de Biologie, Hôpital Européen Georges Pompidou, AP-HP, Paris, France Metabolomics and Cell Biology Platforms, Gustave Roussy Cancer Campus, Villejuif, France Search for more papers by this author Author Information Lorenzo Galluzzi 1,2,3,4,‡,‡, Federico Pietrocola1,2,3,‡, José Manuel Bravo-San Pedro1,2,3,‡, Ravi K Amaravadi5, Eric H Baehrecke6, Francesco Cecconi7,8, Patrice Codogno4,9,10,11, Jayanta Debnath12, David A Gewirtz13, Vassiliki Karantza14, Alec Kimmelman15, Sharad Kumar16, Beth Levine17,18,19, Maria Chiara Maiuri1,2,3, Seamus J Martin20, Josef Penninger21, Mauro Piacentini22,23, David C Rubinsztein24, Hans-Uwe Simon25, Anne Simonsen26, Andrew M Thorburn27, Guillermo Velasco28,29, Kevin M Ryan30,‡ and Guido Kroemer1,2,4,31,32,‡ 1Equipe 11 labellisée pas la Ligue Nationale contre le Cancer, Centre de Recherche des Cordeliers, Paris, France 2INSERM, U1138, Paris, France 3Gustave Roussy Cancer Campus, Villejuif, France 4Université Paris Descartes, Sorbonne Paris Cité, Paris, France 5Abramson Cancer Center, University of Pennsylvania, Philadelphia, PA, USA 6Department of Molecular, Cell and Cancer Biology, University of Massachusetts Medical School, Worcester, MA, USA 7Cell Stress and Survival Unit, Danish Cancer Society Research Center, Copenhagen, Denmark 8IRCCS Fondazione Santa Lucia and Department of Biology, University of Rome Tor Vergata, Rome, Italy 9Institut Necker Enfants-Malades (INEM), Paris, France 10INSERM, U1151, Paris, France 11CNRS, UMR8253, Paris, France 12Department of Pathology and Helen Diller Family Comprehensive Cancer Center, University of California San Francisco, San Francisco, CA, USA 13Department of Pharmacology, Toxicology and Medicine, Virginia Commonwealth University, Richmond, Virginia, VA, USA 14Merck Research Laboratories, Rahway, NJ, USA 15Division of Genomic Stability and DNA Repair, Department of Radiation Oncology, Dana-Farber Cancer Institute, Boston, MA, USA 16Centre for Cancer Biology, University of South Australia, Adelaide, SA, Australia 17Center for Autophagy Research, Department of Internal Medicine, University of Texas Southwestern Medical Center, Dallas, TX, USA 18Department of Microbiology, University of Texas Southwestern Medical Center, Dallas, TX, USA 19Howard Hughes Medical Institute, University of Texas Southwestern Medical Center, Dallas, TX, USA 20Department of Genetics, Trinity College, The Smurfit Institute, Dublin, Ireland 21Institute of Molecular Biotechnology of the Austrian Academy of Sciences, Vienna, Austria 22Department of Biology, University of Rome Tor Vergata, Rome, Italy 23National Institute for Infectious Diseases IRCCS ‘Lazzaro Spallanzani’, Rome, Italy 24Department of Medical Genetics, Cambridge Institute for Medical Research, University of Cambridge, Cambridge, UK 25Institute of Pharmacology, University of Bern, Bern, Switzerland 26Institute of Basic Medical Sciences, University of Oslo, Oslo, Norway 27Department of Pharmacology, University of Colorado School of Medicine, Aurora, CO, USA 28Department of Biochemistry and Molecular Biology I, School of Biology, Complutense University of Madrid, Madrid, Spain 29Instituto de Investigaciones Sanitarias San Carlos (IdISSC), Madrid, Spain 30Cancer Research UK Beatson Institute, Glasgow, UK 31Pôle de Biologie, Hôpital Européen Georges Pompidou, AP-HP, Paris, France 32Metabolomics and Cell Biology Platforms, Gustave Roussy Cancer Campus, Villejuif, France ‡These authors contributed equally to this work ‡These authors share senior co-authorship *Corresponding author. Tel: +33 1 44277661; E-mail: [email protected] The EMBO Journal (2015)34:856-880https://doi.org/10.15252/embj.201490784 PDFDownload PDF of article text and main figures. ToolsAdd to favoritesDownload CitationsTrack CitationsPermissions ShareFacebookTwitterLinked InMendeleyWechatReddit Figures & Info Abstract Autophagy plays a key role in the maintenance of cellular homeostasis. In healthy cells, such a homeostatic activity constitutes a robust barrier against malignant transformation. Accordingly, many oncoproteins inhibit, and several oncosuppressor proteins promote, autophagy. Moreover, autophagy is required for optimal anticancer immunosurveillance. In neoplastic cells, however, autophagic responses constitute a means to cope with intracellular and environmental stress, thus favoring tumor progression. This implies that at least in some cases, oncogenesis proceeds along with a temporary inhibition of autophagy or a gain of molecular functions that antagonize its oncosuppressive activity. Here, we discuss the differential impact of autophagy on distinct phases of tumorigenesis and the implications of this concept for the use of autophagy modulators in cancer therapy. Introduction Macroautophagy (herein referred to as autophagy) is a mechanism that mediates the sequestration of intracellular entities within double-membraned vesicles, so-called autophagosomes, and their delivery to lysosomes for bulk degradation (He & Klionsky, 2009). Autophagosomes derive from so-called phagophores, membranous structures also known as ‘isolation membranes’ whose precise origin remains a matter of debate (Lamb et al, 2013). Indeed, the plasma membrane, endoplasmic reticulum (ER), Golgi apparatus, ER-Golgi intermediate compartment (ERGIC), and mitochondria have all been indicated as possible sources for phagophores (Lamb et al, 2013). Upon closure, autophagosomes fuse with lysosomes, forming so-called autolysosomes, and their cargo is exposed to the catalytic activity of lysosomal hydrolases (Mizushima & Komatsu, 2011). The degradation products of the autophagosomal cargo, which includes sugars, nucleosides/nucleotides, amino acids and fatty acids, can be transported back to the cytoplasm and presumably re-enter cellular metabolism (Fig 1) (Rabinowitz & White, 2010; Galluzzi et al, 2013). Of note, the molecular machinery that mediates autophagy is evolutionary conserved, and several components thereof have initially been characterized in yeast (He & Klionsky, 2009). Figure 1. General organization of autophagic responsesAutophagy initiates with the progressive segregation of cytoplasmic material by double-membraned structures commonly known as phagophores or isolation membranes. Phagophores nucleate from the endoplasmic reticulum (ER), but several other membranous organelles have been shown to contribute to their elongation, including the Golgi apparatus, ER-Golgi intermediate compartment (ERGIC), plasma membrane, mitochondria and recycling endosomes. Completely sealed phagophores, which are known as autophagosomes, fuse with lysosomes to form autolysosomes. This promotes the activation of lysosomal hydrolases and hence causes the breakdown of the autophagosomal cargo. The products of these catabolic reactions reach the cytosol via transporters of the lysosomal membrane and are recycled by anabolic or bioenergetic circuitries. Download figure Download PowerPoint In physiological scenarios, autophagy proceeds at basal levels, ensuring the continuous removal of superfluous, ectopic or damaged (and hence potentially dangerous) entities, including organelles and/or portions thereof (Green et al, 2011). Baseline autophagy mediates a key homeostatic function, constantly operating as an intracellular quality control system (Mizushima et al, 2008; Green et al, 2011). Moreover, the autophagic flux can be upregulated in response to a wide panel of stimuli, including (but not limited to) nutritional, metabolic, oxidative, pathogenic, genotoxic and proteotoxic cues (Kroemer et al, 2010). Often, stimulus-induced autophagy underlies and sustains an adaptive response to stress with cytoprotective functions (Kroemer et al, 2010; Mizushima & Komatsu, 2011). Indeed, the pharmacological or genetic inhibition of autophagy generally limits the ability of cells to cope with stress and restore homeostasis (Mizushima et al, 2008; Kroemer et al, 2010). This said, regulated instances of cell death that causally depend on the autophagic machinery have been described (Denton et al, 2009; Denton et al, 2012b; Liu et al, 2013b; Galluzzi et al, 2015). The detailed discussion of such forms of autophagic cell death, however, is beyond the scope of this review. Autophagy is tightly regulated. The best characterized repressor of autophagic responses is mechanistic target of rapamycin (MTOR) complex I (MTORCI) (Laplante & Sabatini, 2012). Thus, several inducers of autophagy operate by triggering signal transduction cascades that result in the inhibition of MTORCI (Inoki et al, 2012). Among other effects, this allows for the activation of several proteins that are crucial for the initiation of autophagic responses, such as unc-51-like autophagy-activating kinase 1 (ULK1, the mammalian ortholog of yeast Atg1) and autophagy-related 13 (ATG13) (Hosokawa et al, 2009; Nazio et al, 2013). A major inhibitor of MTORCI is protein kinase, AMP-activated (PRKA, best known as AMPK), which is sensitive to declining ATP/AMP ratios (Mihaylova & Shaw, 2011). Besides inhibiting the catalytic activity of MTORCI, AMPK directly stimulates autophagy by phosphorylating ULK1 as well as phosphatidylinositol 3-kinase, catalytic subunit type 3 (PIK3C3, best known as VPS34) and Beclin 1 (BECN1, the mammalian ortholog of yeast Atg6), two components of a multiprotein complex that produces a lipid that is essential for the biogenesis of autophagosomes, namely phosphatidylinositol 3-phosphate (Egan et al, 2011; Zhao & Klionsky, 2011; Kim et al, 2013). Autophagy also critically relies on two ubiquitin-like conjugation systems, both of which involve ATG7 (Mizushima, 2007). These systems catalyze the covalent linkage of ATG5 to ATG12 and ATG16-like 1 (ATG16L1), and that of phosphatidylethanolamine to proteins of the microtubule-associated protein 1 light chain 3 (MAP1LC3, best known as LC3) family, including MAP1LC3B (LC3B, the mammalian ortholog of yeast Atg8) (Mizushima, 2007). A detailed discussion of additional factors that are involved in the control and execution of autophagic responses can be found in Boya et al (2013). Importantly, autophagosomes can either take up intracellular material in a relatively non-selective manner or deliver very specific portions of the cytoplasm to degradation, mainly depending on the initiating stimulus (Weidberg et al, 2011; Stolz et al, 2014). Thus, while non-selective forms of autophagy normally develop in response to cell-wide alterations, most often of a metabolic nature, highly targeted autophagic responses follow specific perturbations of intracellular homeostasis, such as the accumulation of permeabilized mitochondria (mitophagy), the formation of protein aggregates (aggrephagy), and pathogen invasion (xenophagy) (Okamoto, 2014; Randow & Youle, 2014). Several receptors participate in the selective recognition and recruitment of autophagosomal cargoes in the course of targeted autophagic responses (Rogov et al, 2014; Stolz et al, 2014). The autophagy receptor best characterized to date, that is, sequestosome 1 (SQSTM1, best known as p62), recruits ubiquitinated proteins to autophagosomes by virtue of an ubiquitin-associated (UBA) and a LC3-binding domain (Pankiv et al, 2007). Owing to its key role in the preservation of intracellular homeostasis, autophagy constitutes a barrier against various degenerative processes that may affect healthy cells, including malignant transformation. Thus, autophagy mediates oncosuppressive effects. Accordingly, proteins with bona fide oncogenic potential inhibit autophagy, while many proteins that prevent malignant transformation stimulate autophagic responses (Morselli et al, 2011). Moreover, autophagy is involved in several aspects of anticancer immunosurveillance, that is, the process whereby the immune system constantly eliminates potentially tumorigenic cells before they establish malignant lesions (Ma et al, 2013). However, autophagy also sustains the survival and proliferation of neoplastic cells exposed to intracellular and environmental stress, hence supporting tumor growth, invasion and metastatic dissemination, at least in some settings (Kroemer et al, 2010; Guo et al, 2013b). Here, we discuss the molecular and cellular mechanisms accounting for the differential impact of autophagy on malignant transformation and tumor progression. Autophagy and malignant transformation In various murine models, defects in the autophagic machinery caused by the whole-body or tissue-specific, heterozygous or homozygous knockout of essential autophagy genes accelerate oncogenesis. For instance, Becn1+/− mice (Becn1−/− animals are not viable) spontaneously develop various malignancies, including lymphomas as well as lung and liver carcinomas (Liang et al, 1999; Qu et al, 2003; Yue et al, 2003; Mortensen et al, 2011), and are more susceptible to parity-associated and Wnt1-driven mammary carcinogenesis than their wild-type counterparts (Cicchini et al, 2014). Similarly, mice lacking one copy of the gene coding for the BECN1 interactor autophagy/beclin-1 regulator 1 (AMBRA1) also exhibit a higher rate of spontaneous tumorigenesis than their wild-type littermates (Cianfanelli et al, 2015). Mice bearing a systemic mosaic deletion of Atg5 or a liver-specific knockout of Atg7 spontaneously develop benign hepatic neoplasms more frequently than their wild-type counterparts (Takamura et al, 2011). Moreover, carcinogen-induced fibrosarcomas appear at an accelerated pace in autophagy-deficient Atg4c−/− mice (Marino et al, 2007), as do KRASG12D-driven and BRAFV600E-driven lung carcinomas in mice bearing lung-restricted Atg5 or Atg7 deletions, respectively (Strohecker et al, 2013; Rao et al, 2014). The pancreas-specific knockout of Atg5 or Atg7 also precipitates the emergence of KRASG12D-driven pre-malignant pancreatic lesions (Rosenfeldt et al, 2013; Yang et al, 2014). Several mechanisms can explain, at least in part, the oncosuppressive functions of autophagy. Proficient autophagic responses may suppress the accumulation of genetic and genomic defects that accompanies malignant transformation, through a variety of mechanisms. Reactive oxygen species (ROS) are highly genotoxic, and autophagy prevents their overproduction by removing dysfunctional mitochondria (Green et al, 2011; Takahashi et al, 2013) as well as redox-active aggregates of ubiquitinated proteins (Komatsu et al, 2007; Mathew et al, 2009). In addition, autophagic responses have been involved in the disposal of micronuclei arising upon perturbation of the cell cycle (Rello-Varona et al, 2012), in the degradation of retrotransposing RNAs (Guo et al, 2014), as well as in the control of the levels of ras homolog family member A (RHOA), a small GTPase involved in cytokinesis (Belaid et al, 2013). Finally, various components of the autophagic machinery appear to be required for cells to mount adequate responses to genotoxic stress (Karantza-Wadsworth et al, 2007; Mathew et al, 2007; Park et al, 2014). This said, the precise mechanisms underlying such genome-stabilizing effects remain elusive, implying that the impact of autophagy on DNA-damage responses may be indirect. Further investigation is required to shed light on this possibility. Autophagy is intimately implicated in the maintenance of physiological metabolic homeostasis (Galluzzi et al, 2014; Kenific & Debnath, 2015). Malignant transformation generally occurs along with a shift from a predominantly catabolic consumption of glycolysis-derived pyruvate by oxidative phosphorylation to a metabolic pattern in which: (1) glucose uptake is significantly augmented to sustain anabolic reactions and antioxidant defenses, (2) mitochondrial respiration remains high to satisfy increased energy demands; and (3) several amino acids, including glutamine and serine, become essential as a means to cope with exacerbated metabolic functions (Hanahan & Weinberg, 2011; Galluzzi et al, 2013). Autophagy preserves optimal bioenergetic functions by ensuring the removal of dysfunctional mitochondria (Green et al, 2011), de facto counteracting the metabolic rewiring that accompanies malignant transformation. Moreover, the autophagic degradation of p62 participates in a feedback circuitry that regulates MTORCI activation in response to nutrient availability (Linares et al, 2013; Valencia et al, 2014). Autophagy appears to ensure the maintenance of normal stem cells. This is particularly relevant for hematological malignancies, which are normally characterized by changes in proliferation or differentiation potential that alter the delicate equilibrium between toti-, pluri- and unipotent precursors in the bone marrow (Greim et al, 2014). The ablation of Atg7 in murine hematopoietic stem cells (HSCs) has been shown to disrupt tissue architecture, eventually resulting in the expansion of a population of bone marrow progenitor cells with neoplastic features (Mortensen et al, 2011). Along similar lines, the tissue-specific deletion of the gene coding for the ULK1 interactor RB1-inducible coiled-coil 1 (RB1CC1, best known as FIP200) alters the fetal HSC compartment in mice, resulting in severe anemia and perinatal lethality (Liu et al, 2010). Interestingly, murine Rb1cc1−/− HSCs do not exhibit increased rates of apoptosis, but an accrued proliferative capacity (Liu et al, 2010). The deletion of Rb1cc1 in murine neuronal stem cells (NSCs) also causes a functional impairment that compromises postnatal neuronal differentiation (Wang et al, 2013). However, this effect appears to stem from the failure of murine Rb1cc1−/− HNCs to control redox homeostasis, resulting in the activation of a tumor protein p53 (TP53)-dependent apoptotic response (Wang et al, 2013). Finally, Becn1+/− mice display an expansion of progenitor-like mammary epithelial cells (Cicchini et al, 2014). Of note, autophagy also appears to be required for the preservation of normal stem cell compartments in the human system. Indeed, human hematopoietic, dermal, and epidermal stem cells transfected with a short-hairpin RNA (shRNA) specific for ATG5 lose their ability to self-renew while differentiating into neutrophils, fibroblasts, and keratinocytes, respectively (Salemi et al, 2012). It has been proposed that autophagy contributes to oncogene-induced cell death or oncogene-induced senescence, two fundamental oncosuppressive mechanisms. The activation of various oncogenes imposes indeed a significant stress on healthy cells, a situation that is normally aborted through the execution of a cell death program (Elgendy et al, 2011), or upon the establishment of permanent proliferative arrest (cell senescence) that engages the innate arm of the immune system (Iannello et al, 2013). The partial depletion of ATG5, ATG7 or BECN1 limited the demise of human ovarian cancer cells pharmacologically stimulated to express HRASG12V from an inducible construct (Elgendy et al, 2011). Similarly, shRNAs specific for ATG5 or ATG7 prevented oncogene-induced senescence in primary human melanocytes or human diploid fibroblasts (HDFs) expressing BRAFV600E or HRASG12V (Young et al, 2009; Liu et al, 2013a). Accordingly, the overexpression of the ULK1 homolog ULK3 was sufficient to limit the proliferative potential of HDFs while promoting autophagy (Young et al, 2009). Moreover, both pharmacological inhibitors of autophagy and small-interfering RNAs targeting ATG5, ATG7 or BECN1 prevented spontaneous senescence in HDFs while preventing the degradation of an endogenous, dominant-neg
DOI: 10.1186/s13059-016-1092-z
2016
Cited 706 times
Tumor immune microenvironment characterization in clear cell renal cell carcinoma identifies prognostic and immunotherapeutically relevant messenger RNA signatures
Tumor-infiltrating immune cells have been linked to prognosis and response to immunotherapy; however, the levels of distinct immune cell subsets and the signals that draw them into a tumor, such as the expression of antigen presenting machinery genes, remain poorly characterized. Here, we employ a gene expression-based computational method to profile the infiltration levels of 24 immune cell populations in 19 cancer types. We compare cancer types using an immune infiltration score and a T cell infiltration score and find that clear cell renal cell carcinoma (ccRCC) is among the highest for both scores. Using immune infiltration profiles as well as transcriptomic and proteomic datasets, we characterize three groups of ccRCC tumors: T cell enriched, heterogeneously infiltrated, and non-infiltrated. We observe that the immunogenicity of ccRCC tumors cannot be explained by mutation load or neo-antigen load, but is highly correlated with MHC class I antigen presenting machinery expression (APM). We explore the prognostic value of distinct T cell subsets and show in two cohorts that Th17 cells and CD8+ T/Treg ratio are associated with improved survival, whereas Th2 cells and Tregs are associated with negative outcomes. Investigation of the association of immune infiltration patterns with the subclonal architecture of tumors shows that both APM and T cell levels are negatively associated with subclone number. Our analysis sheds light on the immune infiltration patterns of 19 human cancers and unravels mRNA signatures with prognostic utility and immunotherapeutic biomarker potential in ccRCC.
DOI: 10.1172/jci37948
2009
Cited 610 times
Cannabinoid action induces autophagy-mediated cell death through stimulation of ER stress in human glioma cells
Autophagy can promote cell survival or cell death, but the molecular basis underlying its dual role in cancer remains obscure. Here we demonstrate that delta(9)-tetrahydrocannabinol (THC), the main active component of marijuana, induces human glioma cell death through stimulation of autophagy. Our data indicate that THC induced ceramide accumulation and eukaryotic translation initiation factor 2alpha (eIF2alpha) phosphorylation and thereby activated an ER stress response that promoted autophagy via tribbles homolog 3-dependent (TRB3-dependent) inhibition of the Akt/mammalian target of rapamycin complex 1 (mTORC1) axis. We also showed that autophagy is upstream of apoptosis in cannabinoid-induced human and mouse cancer cell death and that activation of this pathway was necessary for the antitumor action of cannabinoids in vivo. These findings describe a mechanism by which THC can promote the autophagic death of human and mouse cancer cells and provide evidence that cannabinoid administration may be an effective therapeutic strategy for targeting human cancers.
DOI: 10.1016/j.ccell.2017.04.013
2017
Cited 457 times
A Pan-Cancer Proteogenomic Atlas of PI3K/AKT/mTOR Pathway Alterations
Molecular alterations involving the PI3K/AKT/mTOR pathway (including mutation, copy number, protein, or RNA) were examined across 11,219 human cancers representing 32 major types. Within specific mutated genes, frequency, mutation hotspot residues, in silico predictions, and functional assays were all informative in distinguishing the subset of genetic variants more likely to have functional relevance. Multiple oncogenic pathways including PI3K/AKT/mTOR converged on similar sets of downstream transcriptional targets. In addition to mutation, structural variations and partial copy losses involving PTEN and STK11 showed evidence for having functional relevance. A substantial fraction of cancers showed high mTOR pathway activity without an associated canonical genetic or genomic alteration, including cancers harboring IDH1 or VHL mutations, suggesting multiple mechanisms for pathway activation.
DOI: 10.1016/j.tips.2006.11.001
2007
Cited 330 times
Cannabinoid CB2 receptor: a new target for controlling neural cell survival?
Two types of cannabinoid receptor have been cloned and characterized. Whereas CB1 receptors are ubiquitously expressed in neurons of the CNS, CB2 receptors have been thought to be absent from the CNS. Recent data now question this notion and support the expression of CB2 receptors in microglial cells, astrocytes and even some neuron subpopulations. This discrete distribution makes CB2 receptors interesting targets for treating neurological disorders because CB2-selective agonists lack psychoactivity. Here, we review evidence supporting the idea that CB2 receptors are implicated in the control of fundamental neural cell processes, such as proliferation and survival, and that their pharmacological manipulation might be useful for both delaying the progression of neurodegenerative disorders and inhibiting the growth of glial tumors.
DOI: 10.1016/j.ccr.2006.03.005
2006
Cited 318 times
The stress-regulated protein p8 mediates cannabinoid-induced apoptosis of tumor cells
One of the most exciting areas of current research in the cannabinoid field is the study of the potential application of these compounds as antitumoral drugs. Here, we describe the signaling pathway that mediates cannabinoid-induced apoptosis of tumor cells. By using a wide array of experimental approaches, we identify the stress-regulated protein p8 (also designated as candidate of metastasis 1) as an essential mediator of cannabinoid antitumoral action and show that p8 upregulation is dependent on de novo-synthesized ceramide. We also observe that p8 mediates its apoptotic effect via upregulation of the endoplasmic reticulum stress-related genes ATF-4, CHOP, and TRB3. Activation of this pathway may constitute a potential therapeutic strategy for inhibiting tumor growth.
DOI: 10.1158/0008-5472.can-06-0169
2006
Cited 314 times
Cannabinoids Induce Apoptosis of Pancreatic Tumor Cells via Endoplasmic Reticulum Stress–Related Genes
Abstract Pancreatic adenocarcinomas are among the most malignant forms of cancer and, therefore, it is of especial interest to set new strategies aimed at improving the prognostic of this deadly disease. The present study was undertaken to investigate the action of cannabinoids, a new family of potential antitumoral agents, in pancreatic cancer. We show that cannabinoid receptors are expressed in human pancreatic tumor cell lines and tumor biopsies at much higher levels than in normal pancreatic tissue. Studies conducted with MiaPaCa2 and Panc1 cell lines showed that cannabinoid administration (a) induced apoptosis, (b) increased ceramide levels, and (c) up-regulated mRNA levels of the stress protein p8. These effects were prevented by blockade of the CB2 cannabinoid receptor or by pharmacologic inhibition of ceramide synthesis de novo. Knockdown experiments using selective small interfering RNAs showed the involvement of p8 via its downstream endoplasmic reticulum stress–related targets activating transcription factor 4 (ATF-4) and TRB3 in Δ9-tetrahydrocannabinol–induced apoptosis. Cannabinoids also reduced the growth of tumor cells in two animal models of pancreatic cancer. In addition, cannabinoid treatment inhibited the spreading of pancreatic tumor cells. Moreover, cannabinoid administration selectively increased apoptosis and TRB3 expression in pancreatic tumor cells but not in normal tissue. In conclusion, results presented here show that cannabinoids lead to apoptosis of pancreatic tumor cells via a CB2 receptor and de novo synthesized ceramide-dependent up-regulation of p8 and the endoplasmic reticulum stress–related genes ATF-4 and TRB3. These findings may contribute to set the basis for a new therapeutic approach for the treatment of pancreatic cancer. (Cancer Res 2006; 66(13): 6748-55)
DOI: 10.1038/nrc3247
2012
Cited 307 times
Towards the use of cannabinoids as antitumour agents
DOI: 10.1038/sj.bjc.6603236
2006
Cited 303 times
A pilot clinical study of Δ9-tetrahydrocannabinol in patients with recurrent glioblastoma multiforme
Δ9-Tetrahydrocannabinol (THC) and other cannabinoids inhibit tumour growth and angiogenesis in animal models, so their potential application as antitumoral drugs has been suggested. However, the antitumoral effect of cannabinoids has never been tested in humans. Here we report the first clinical study aimed at assessing cannabinoid antitumoral action, specifically a pilot phase I trial in which nine patients with recurrent glioblastoma multiforme were administered THC intratumoraly. The patients had previously failed standard therapy (surgery and radiotherapy) and had clear evidence of tumour progression. The primary end point of the study was to determine the safety of intracranial THC administration. We also evaluated THC action on the length of survival and various tumour-cell parameters. A dose escalation regimen for THC administration was assessed. Cannabinoid delivery was safe and could be achieved without overt psychoactive effects. Median survival of the cohort from the beginning of cannabinoid administration was 24 weeks (95% confidence interval: 15–33). Δ9-Tetrahydrocannabinol inhibited tumour-cell proliferation in vitro and decreased tumour-cell Ki67 immunostaining when administered to two patients. The fair safety profile of THC, together with its possible antiproliferative action on tumour cells reported here and in other studies, may set the basis for future trials aimed at evaluating the potential antitumoral activity of cannabinoids.
DOI: 10.1155/2010/930509
2010
Cited 298 times
Linking ER Stress to Autophagy: Potential Implications for Cancer Therapy
Different physiological and pathological conditions can perturb protein folding in the endoplasmic reticulum, leading to a condition known as ER stress. ER stress activates a complex intracellular signal transduction pathway, called unfolded protein response (UPR). The UPR is tailored essentially to reestablish ER homeostasis also through adaptive mechanisms involving the stimulation of autophagy. However, when persistent, ER stress can switch the cytoprotective functions of UPR and autophagy into cell death promoting mechanisms. Recently, a variety of anticancer therapies have been linked to the induction of ER stress in cancer cells, suggesting that strategies devised to stimulate its prodeath function or block its prosurvival function, could be envisaged to improve their tumoricidial action. A better understanding of the molecular mechanisms that determine the final outcome of UPR and autophagy activation by chemotherapeutic agents, will offer new opportunities to improve existing cancer therapies as well as unravel novel targets for cancer treatment.
DOI: 10.1158/1535-7163.mct-10-0688
2011
Cited 244 times
A Combined Preclinical Therapy of Cannabinoids and Temozolomide against Glioma
Glioblastoma multiforme (GBM) is highly resistant to current anticancer treatments, which makes it crucial to find new therapeutic strategies aimed at improving the poor prognosis of patients suffering from this disease. Δ(9)-Tetrahydrocannabinol (THC), the major active ingredient of marijuana, and other cannabinoid receptor agonists inhibit tumor growth in animal models of cancer, including glioma, an effect that relies, at least in part, on the stimulation of autophagy-mediated apoptosis in tumor cells. Here, we show that the combined administration of THC and temozolomide (TMZ; the benchmark agent for the management of GBM) exerts a strong antitumoral action in glioma xenografts, an effect that is also observed in tumors that are resistant to TMZ treatment. Combined administration of THC and TMZ enhanced autophagy, whereas pharmacologic or genetic inhibition of this process prevented TMZ + THC-induced cell death, supporting that activation of autophagy plays a crucial role on the mechanism of action of this drug combination. Administration of submaximal doses of THC and cannabidiol (CBD; another plant-derived cannabinoid that also induces glioma cell death through a mechanism of action different from that of THC) remarkably reduces the growth of glioma xenografts. Moreover, treatment with TMZ and submaximal doses of THC and CBD produced a strong antitumoral action in both TMZ-sensitive and TMZ-resistant tumors. Altogether, our findings support that the combined administration of TMZ and cannabinoids could be therapeutically exploited for the management of GBM.
DOI: 10.1038/cdd.2011.32
2011
Cited 230 times
Anti-tumoral action of cannabinoids on hepatocellular carcinoma: role of AMPK-dependent activation of autophagy
Hepatocellular carcinoma (HCC) is the third cause of cancer-related death worldwide. When these tumors are in advanced stages, few therapeutic options are available. Therefore, it is essential to search for new treatments to fight this disease. In this study, we investigated the effects of cannabinoids--a novel family of potential anticancer agents--on the growth of HCC. We found that Δ(9)-tetrahydrocannabinol (Δ(9)-THC, the main active component of Cannabis sativa) and JWH-015 (a cannabinoid receptor 2 (CB(2)) cannabinoid receptor-selective agonist) reduced the viability of the human HCC cell lines HepG2 (human hepatocellular liver carcinoma cell line) and HuH-7 (hepatocellular carcinoma cells), an effect that relied on the stimulation of CB(2) receptor. We also found that Δ(9)-THC- and JWH-015-induced autophagy relies on tribbles homolog 3 (TRB3) upregulation, and subsequent inhibition of the serine-threonine kinase Akt/mammalian target of rapamycin C1 axis and adenosine monophosphate-activated kinase (AMPK) stimulation. Pharmacological and genetic inhibition of AMPK upstream kinases supported that calmodulin-activated kinase kinase β was responsible for cannabinoid-induced AMPK activation and autophagy. In vivo studies revealed that Δ(9)-THC and JWH-015 reduced the growth of HCC subcutaneous xenografts, an effect that was not evident when autophagy was genetically of pharmacologically inhibited in those tumors. Moreover, cannabinoids were also able to inhibit tumor growth and ascites in an orthotopic model of HCC xenograft. Our findings may contribute to the design of new therapeutic strategies for the management of HCC.
DOI: 10.1038/ncb3231
2015
Cited 219 times
AMPK and PFKFB3 mediate glycolysis and survival in response to mitophagy during mitotic arrest
DOI: 10.1038/ncb3072
2014
Cited 209 times
AMBRA1 links autophagy to cell proliferation and tumorigenesis by promoting c-Myc dephosphorylation and degradation
Inhibition of a main regulator of cell metabolism, the protein kinase mTOR, induces autophagy and inhibits cell proliferation. However, the molecular pathways involved in the cross-talk between these two mTOR-dependent cell processes are largely unknown. Here we show that the scaffold protein AMBRA1, a member of the autophagy signalling network and a downstream target of mTOR, regulates cell proliferation by facilitating the dephosphorylation and degradation of the proto-oncogene c-Myc. We found that AMBRA1 favours the interaction between c-Myc and its phosphatase PP2A and that, when mTOR is inhibited, it enhances PP2A activity on this specific target, thereby reducing the cell division rate. As expected, such a de-regulation of c-Myc correlates with increased tumorigenesis in AMBRA1-defective systems, thus supporting a role for AMBRA1 as a haploinsufficient tumour suppressor gene.
DOI: 10.3747/co.23.3080
2016
Cited 188 times
Anticancer Mechanisms of Cannabinoids
In addition to the well-known palliative effects of cannabinoids on some cancer-associated symptoms, a large body of evidence shows that these molecules can decrease tumour growth in animal models of cancer. They do so by modulating key cell signalling pathways involved in the control of cancer cell proliferation and survival. In addition, cannabinoids inhibit angiogenesis and decrease metastasis in various tumour types in laboratory animals. In this review, we discuss the current understanding of cannabinoids as antitumour agents, focusing on recent discoveries about their molecular mechanisms of action, including resistance mechanisms and opportunities for their use in combination therapy. Those observations have already contributed to the foundation for the development of the first clinical studies that will analyze the safety and potential clinical benefit of cannabinoids as anticancer agents.
DOI: 10.1038/ncb3357
2016
Cited 180 times
The metabolic co-regulator PGC1α suppresses prostate cancer metastasis
Cellular transformation and cancer progression is accompanied by changes in the metabolic landscape. Master co-regulators of metabolism orchestrate the modulation of multiple metabolic pathways through transcriptional programs, and hence constitute a probabilistically parsimonious mechanism for general metabolic rewiring. Here we show that the transcriptional co-activator peroxisome proliferator-activated receptor gamma co-activator 1α (PGC1α) suppresses prostate cancer progression and metastasis. A metabolic co-regulator data mining analysis unveiled that PGC1α is downregulated in prostate cancer and associated with disease progression. Using genetically engineered mouse models and xenografts, we demonstrated that PGC1α opposes prostate cancer progression and metastasis. Mechanistically, the use of integrative metabolomics and transcriptomics revealed that PGC1α activates an oestrogen-related receptor alpha (ERRα)-dependent transcriptional program to elicit a catabolic state and metastasis suppression. Importantly, a signature based on the PGC1α–ERRα pathway exhibited prognostic potential in prostate cancer, thus uncovering the relevance of monitoring and manipulating this pathway for prostate cancer stratification and treatment. Torrano et al. use bioinformatics analyses to identify PGC1α as a transcriptional regulator of a metabolic program downstream of ERRα that opposes metastatic dissemination in prostate cancer.
DOI: 10.1200/jco.2016.66.7311
2016
Cited 172 times
Body Mass Index and Metastatic Renal Cell Carcinoma: Clinical and Biological Correlations
Obesity is an established risk factor for clear cell renal cell carcinoma (RCC); however, some reports suggest that RCC developing in obese patients may be more indolent. We investigated the clinical and biologic effect of body mass index (BMI) on treatment outcomes in patients with metastatic RCC.The impact of BMI (high BMI: ≥ 25 kg/m2 v low BMI: < 25 kg/m2) on overall survival (OS) and treatment outcome with targeted therapy was investigated in 1,975 patients from the International Metastatic Renal Cell Carcinoma Database Consortium (IMDC) and in an external validation cohort of 4,657 patients. Gene expression profiling focusing on fatty acid metabolism pathway, in The Cancer Genome Atlas data set, and immunohistochemistry staining for fatty acid synthase (FASN) were also investigated. Cox regression was undertaken to estimate the association of BMI with OS, adjusted for the IMDC prognostic factors.In the IMDC cohort, median OS was 25.6 months (95% CI, 23.2 to 28.6) in patients with high BMI versus 17.1 months (95% CI, 15.5 to 18.5) in patients with low BMI (adjusted hazard ratio, 0.84; 95% CI, 0.73 to 0.95). In the validation cohort, high BMI was associated with improved OS (adjusted hazard ratio, 0.83; 95% CI, 0.74 to 0.93; medians: 23.4 months [95% CI, 21.9 to 25.3 months] v 14.5 months [95% CI, 13.8 to 15.9 months], respectively). In The Cancer Genome Atlas data set (n = 61), FASN gene expression inversely correlated with BMI (P = .034), and OS was longer in the low FASN expression group (medians: 36.8 v 15.0 months; P = .002). FASN immunohistochemistry positivity was more frequently detected in IMDC poor (48%) and intermediate (34%) risk groups than in the favorable risk group (17%; P-trend = .015).High BMI is a prognostic factor for improved survival and progression-free survival in patients with metastatic RCC treated with targeted therapy. Underlying biology suggests a role for the FASN pathway.
DOI: 10.1016/j.cels.2018.08.010
2018
Cited 138 times
A Pan-Cancer Analysis Reveals High-Frequency Genetic Alterations in Mediators of Signaling by the TGF-β Superfamily
We present an integromic analysis of gene alterations that modulate transforming growth factor β (TGF-β)-Smad-mediated signaling in 9,125 tumor samples across 33 cancer types in The Cancer Genome Atlas (TCGA). Focusing on genes that encode mediators and regulators of TGF-β signaling, we found at least one genomic alteration (mutation, homozygous deletion, or amplification) in 39% of samples, with highest frequencies in gastrointestinal cancers. We identified mutation hotspots in genes that encode TGF-β ligands (BMP5), receptors (TGFBR2, AVCR2A, and BMPR2), and Smads (SMAD2 and SMAD4). Alterations in the TGF-β superfamily correlated positively with expression of metastasis-associated genes and with decreased survival. Correlation analyses showed the contributions of mutation, amplification, deletion, DNA methylation, and miRNA expression to transcriptional activity of TGF-β signaling in each cancer type. This study provides a broad molecular perspective relevant for future functional and therapeutic studies of the diverse cancer pathways mediated by the TGF-β superfamily.
DOI: 10.1038/s41591-019-0499-y
2019
Cited 130 times
Inhibition of a G9a/DNMT network triggers immune-mediated bladder cancer regression
DOI: 10.1016/j.annonc.2021.09.014
2021
Cited 125 times
ESMO Clinical Practice Guideline update on the use of immunotherapy in early stage and advanced renal cell carcinoma
•This special article provides key recommendations on the immunotherapy treatments of renal cell carcinoma.•Recommendations are based on available scientific data and the authors' collective expert opinion.•Authorship includes a multidisciplinary group of renal carcinoma experts.
DOI: 10.1001/jamanetworkopen.2020.21869
2021
Cited 112 times
Evaluation of Clear Cell, Papillary, and Chromophobe Renal Cell Carcinoma Metastasis Sites and Association With Survival
There exists considerable biological and clinical variability between histologic variants of metastatic renal cell carcinoma (mRCC). Data reporting on patterns of metastasis in histologic variants of mRCC are sparse.To characterize sites of metastasis and their association with survival across the 3 most common histologic variants of mRCC: clear cell (ccRCC), papillary (pRCC), and chromophobe (chrRCC).In this multicenter, international cohort study, the International mRCC Database Consortium (IMDC) database was used to identify consecutive patients starting systemic therapy for mRCC between 2002 and 2019. Patients with mixed histologic subtype were excluded. Statistical analysis was performed from February to June 2020.Data regarding histologic subtype and sites of metastatic involvement at the time of first systemic therapy initiation were collected.The primary outcomes were prevalence of metastatic site involvement and overall survival (OS) from time of systemic therapy initiation. Patients with multiple sites of metastatic involvement were included in analyses of all groups to which they had metastases.A total of 10 105 patients were eligible for analysis. Median (interquartile range) age at diagnosis was 60 (53-67) years, 7310 (72.4%) were men and 8526 (84.5%) underwent nephrectomy. Of these, 9252 (92%) had ccRCC, 667 (7%) had pRCC, and 186 (2%) had chrRCC. The median number of sites of metastasis was 2 (range, 0-7). In ccRCC, the most common sites of metastasis were lung (70%; 6189 of 8804 patients [448 missing]), lymph nodes (45%; 3874 of 8655 patients [597 missing]), bone (32%; 2847 of 8817 patients [435 missing]), liver (18%; 1560 of 8804 [448 missing]), and adrenal gland (10%; 678 of 6673 patients [2579 missing]). Sites of metastasis varied between subtypes. Lung, adrenal, brain, and pancreatic metastases were more frequent in ccRCC, lymph node involvement was more common in pRCC, and liver metastases were more frequent in chrRCC. Median OS for ccRCC varied by site of metastatic involvement, ranging between 16 months (95% CI, 13.7-18.8 months) for the pleura and 50 months (95% CI, 41.1-55.5 months) for the pancreas. Compared with ccRCC, patients with pRCC tended to have lower OS, regardless of metastatic site.Sites of metastatic involvement differ according to histologic subtype in mRCC and are associated with OS. These data highlight the clinical and biological variability between histologic subtypes of mRCC. Patterns of metastatic spread may reflect differences in underlying disease biology. Further work to investigate differences in immune, molecular, and genetic profiles between metastatic sites and histologic subtypes is encouraged.
DOI: 10.1038/s41586-021-03422-5
2021
Cited 84 times
AMBRA1 regulates cyclin D to guard S-phase entry and genomic integrity
Mammalian development, adult tissue homeostasis and the avoidance of severe diseases including cancer require a properly orchestrated cell cycle, as well as error-free genome maintenance. The key cell-fate decision to replicate the genome is controlled by two major signalling pathways that act in parallel-the MYC pathway and the cyclin D-cyclin-dependent kinase (CDK)-retinoblastoma protein (RB) pathway1,2. Both MYC and the cyclin D-CDK-RB axis are commonly deregulated in cancer, and this is associated with increased genomic instability. The autophagic tumour-suppressor protein AMBRA1 has been linked to the control of cell proliferation, but the underlying molecular mechanisms remain poorly understood. Here we show that AMBRA1 is an upstream master regulator of the transition from G1 to S phase and thereby prevents replication stress. Using a combination of cell and molecular approaches and in vivo models, we reveal that AMBRA1 regulates the abundance of D-type cyclins by mediating their degradation. Furthermore, by controlling the transition from G1 to S phase, AMBRA1 helps to maintain genomic integrity during DNA replication, which counteracts developmental abnormalities and tumour growth. Finally, we identify the CHK1 kinase as a potential therapeutic target in AMBRA1-deficient tumours. These results advance our understanding of the control of replication-phase entry and genomic integrity, and identify the AMBRA1-cyclin D pathway as a crucial cell-cycle-regulatory mechanism that is deeply interconnected with genomic stability in embryonic development and tumorigenesis.
DOI: 10.1096/fj.06-6638fje
2006
Cited 263 times
Cannabinoid receptors as novel targets for the treatment of melanoma
Melanoma causes the greatest number of skin cancer-related deaths worldwide. Despite intensive research, prevention and early detection are the only effective measures against melanoma, so new therapeutic strategies are necessary for the management of this devastating disease. Here, we evaluated the efficacy of cannabinoid receptor agonists, a new family of potential antitumoral compounds, at skin melanoma. Human melanomas and melanoma cell lines express CB1 and CB2 cannabinoid receptors. Activation of these receptors decreased growth, proliferation, angiogenesis and metastasis, and increased apoptosis, of melanomas in mice. Cannabinoid antimelanoma activity was independent of the immune status of the animal, could be achieved without overt psychoactive effects and was selective for melanoma cells vs. normal melanocytes. Cannabinoid antiproliferative action on melanoma cells was due, at least in part, to cell cycle arrest at the G1-S transition via inhibition of the prosurvival protein Akt and hypophosphorylation of the pRb retinoblastoma protein tumor suppressor. These findings may contribute to the design of new chemotherapeutic strategies for the management of melanoma.
2001
Cited 209 times
Inhibition of Glioma Growth in Vivo by Selective Activation of the CB2 Cannabinoid Receptor
The development of new therapeutic strategies is essential for the management of gliomas, one of the most malignant forms of cancer. We have shown previously that the growth of the rat glioma C6 cell line is inhibited by psychoactive cannabinoids (I. Galve-Roperh et al., Nat. Med., 6: 313-319, 2000). These compounds act on the brain and some other organs through the widely expressed CB(1) receptor. By contrast, the other cannabinoid receptor subtype, the CB(2) receptor, shows a much more restricted distribution and is absent from normal brain. Here we show that local administration of the selective CB(2) agonist JWH-133 at 50 microg/day to Rag-2(-/-) mice induced a considerable regression of malignant tumors generated by inoculation of C6 glioma cells. The selective involvement of the CB(2) receptor in this action was evidenced by: (a) the prevention by the CB(2) antagonist SR144528 but not the CB(1) antagonist SR141716; (b) the down-regulation of the CB(2) receptor but not the CB(1) receptor in the tumors; and (c) the absence of typical CB(1)-mediated psychotropic side effects. Cannabinoid receptor expression was subsequently examined in biopsies from human astrocytomas. A full 70% (26 of 37) of the human astrocytomas analyzed expressed significant levels of cannabinoid receptors. Of interest, the extent of CB(2) receptor expression was directly related with tumor malignancy. In addition, the growth of grade IV human astrocytoma cells in Rag-2(-/-) mice was completely blocked by JWH-133 administration at 50 microg/day. Experiments carried out with C6 glioma cells in culture evidenced the internalization of the CB(2) but not the CB(1) receptor upon JWH-133 challenge and showed that selective activation of the CB(2) receptor signaled apoptosis via enhanced ceramide synthesis de novo. These results support a therapeutic approach for the treatment of malignant gliomas devoid of psychotropic side effects.
DOI: 10.1016/s0014-5793(02)03175-7
2002
Cited 195 times
Phosphorylation of the regulatory subunit of smooth muscle protein phosphatase 1M at Thr850 induces its dissociation from myosin
Rho kinase is known to control smooth muscle contractility by phosphorylating the 110 kDa myosin-targetting subunit (MYPT1) of the myosin-associated form of protein phosphatase 1 (PP1M). Phosphorylation of MYPT1 at Thr695 has previously been reported to inhibit the catalytic activity of PP1. Here, we show that the phosphorylation of Thr850 by Rho kinase dissociates PP1M from myosin, providing a second mechanism by which myosin phosphatase activity is inhibited.
DOI: 10.1124/mol.62.6.1385
2002
Cited 181 times
Mechanism of Extracellular Signal-Regulated Kinase Activation by the CB<sub>1</sub> Cannabinoid Receptor
Cannabinoids, the active components of marijuana and their endogenous counterparts, exert many of their actions in brain through the seven-transmembrane receptor CB<sub>1</sub>. This receptor is coupled to the activation of the extracellular signal-regulated kinase (ERK) cascade. However, the precise molecular mechanism for CB<sub>1</sub>-mediated ERK activation is still unknown. Here, we show that in U373 MG human astrocytoma cells, CB<sub>1</sub> receptor activation with the cannabinoid agonist Δ<sup>8</sup>-tetrahydrocannabinol dimethyl heptyl (HU-210) was coupled to ERK activation and protection from ceramide-induced apoptosis. HU-210-induced ERK activation was inhibited by tyrphostin AG1478 and PP2, widely employed inhibitors of the epidermal growth factor receptor (EGF<sub>R</sub>) and the Src family of cytosolic tyrosine kinases, respectively. However, HU-210 stimulation resulted in neither EGF<sub>R</sub> phosphorylation, Src tyrosine phosphorylation, nor increased Src activity. In addition, dominant-negative forms of both proteins were unable to prevent cannabinoid-induced ERK activation, thus excluding the existence of CB<sub>1</sub>-mediated EGF<sub>R</sub> transactivation or Src activation. Wortmannin and 2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one (LY294,002), inhibitors of the phosphatidylinositol 3-kinase (PI3K) signaling pathway, blocked cannabinoid-induced ERK activation. Likewise, HU-210 stimulated the PI3K downstream targets protein kinase B (PKB), as shown by its phosphorylation in Thr 308 and Ser 473 residues, and Raf-1. Moreover, βγ subunit release mimicked ERK and PI3K/PKB activation, suggesting that activation of class IB PI3K mediates cannabinoid action. Pro-survival HU-210 action also required activation of both PI3K and ERK signaling pathways. In conclusion, CB<sub>1</sub>-induced ERK activation was mediated by PI3K<sub>IB</sub> and this effect may have important consequences in the control of cell death/survival decision.
DOI: 10.1158/0008-5472.can-07-5176
2008
Cited 163 times
Cannabinoids Inhibit Glioma Cell Invasion by Down-regulating Matrix Metalloproteinase-2 Expression
Abstract Cannabinoids, the active components of Cannabis sativa L. and their derivatives, inhibit tumor growth in laboratory animals by inducing apoptosis of tumor cells and impairing tumor angiogenesis. It has also been reported that these compounds inhibit tumor cell spreading, but the molecular targets of this cannabinoid action remain elusive. Here, we evaluated the effect of cannabinoids on matrix metalloproteinase (MMP) expression and its effect on tumor cell invasion. Local administration of Δ9-tetrahydrocannabinol (THC), the major active ingredient of cannabis, down-regulated MMP-2 expression in gliomas generated in mice, as determined by Western blot, immunofluorescence, and real-time quantitative PCR analyses. This cannabinoid-induced inhibition of MMP-2 expression in gliomas (a) was MMP-2–selective, as levels of other MMP family members were unaffected; (b) was mimicked by JWH-133, a CB2 cannabinoid receptor–selective agonist that is devoid of psychoactive side effects; (c) was abrogated by fumonisin B1, a selective inhibitor of ceramide biosynthesis; and (d) was also evident in two patients with recurrent glioblastoma multiforme. THC inhibited MMP-2 expression and cell invasion in cultured glioma cells. Manipulation of MMP-2 expression by RNA interference and cDNA overexpression experiments proved that down-regulation of this MMP plays a critical role in THC-mediated inhibition of cell invasion. Cannabinoid-induced inhibition of MMP-2 expression and cell invasion was prevented by blocking ceramide biosynthesis and by knocking-down the expression of the stress protein p8. As MMP-2 up-regulation is associated with high progression and poor prognosis of gliomas and many other tumors, MMP-2 down-regulation constitutes a new hallmark of cannabinoid antitumoral activity. [Cancer Res 2008;68(6):1945–52]
DOI: 10.1038/onc.2010.402
2010
Cited 161 times
The orphan G protein-coupled receptor GPR55 promotes cancer cell proliferation via ERK
GPR55 is an orphan G protein-coupled receptor that may be engaged by some lipid ligands such as lysophosphatidylinositol and cannabinoid-type compounds. Very little is known about its expression pattern and physio-pathological relevance, and its pharmacology and signaling are still rather controversial. Here we analyzed the expression and function of GPR55 in cancer cells. Our data show that GPR55 expression in human tumors from different origins correlates with their aggressiveness. Moreover, GPR55 promotes cancer cell proliferation, both in cell cultures and in xenografted mice, through the overactivation of the extracellular signal-regulated kinase cascade. These findings reveal the importance of GPR55 in human cancer, and suggest that it could constitute a new biomarker and therapeutic target in oncology.
DOI: 10.1074/jbc.m205797200
2002
Cited 161 times
Cannabinoids Protect Astrocytes from Ceramide-induced Apoptosis through the Phosphatidylinositol 3-Kinase/Protein Kinase B Pathway
Cannabinoids, the active components of marijuana and their endogenous counterparts, exert many of their actions on the central nervous system by binding to the CB1cannabinoid receptor. Different studies have shown that cannabinoids can protect neural cells from different insults. However, those studies have been performed in neurons, whereas no attention has been focused on glial cells. Here we used the pro-apoptotic lipid ceramide to induce apoptosis in astrocytes, and we studied the protective effect exerted by cannabinoids. Results show the following: (i) cannabinoids rescue primary astrocytes from C2-ceramide-induced apoptosis in a dose- and time-dependent manner; (ii) triggering of this anti-apoptotic signal depends on the phosphatidylinositol 3-kinase/protein kinase B pathway; (iii) ERK and its downstream target p90 ribosomal S6 kinase might be also involved in the protective effect of cannabinoids; and (iv) cannabinoids protect astrocytes from the cytotoxic effects of focal C2-ceramide administration in vivo. In summary, results show that cannabinoids protect astrocytes from ceramide-induced apoptosis via stimulation of the phosphatidylinositol 3-kinase/protein kinase B pathway. These findings constitute the first evidence for an “astroprotective” role of cannabinoids. Cannabinoids, the active components of marijuana and their endogenous counterparts, exert many of their actions on the central nervous system by binding to the CB1cannabinoid receptor. Different studies have shown that cannabinoids can protect neural cells from different insults. However, those studies have been performed in neurons, whereas no attention has been focused on glial cells. Here we used the pro-apoptotic lipid ceramide to induce apoptosis in astrocytes, and we studied the protective effect exerted by cannabinoids. Results show the following: (i) cannabinoids rescue primary astrocytes from C2-ceramide-induced apoptosis in a dose- and time-dependent manner; (ii) triggering of this anti-apoptotic signal depends on the phosphatidylinositol 3-kinase/protein kinase B pathway; (iii) ERK and its downstream target p90 ribosomal S6 kinase might be also involved in the protective effect of cannabinoids; and (iv) cannabinoids protect astrocytes from the cytotoxic effects of focal C2-ceramide administration in vivo. In summary, results show that cannabinoids protect astrocytes from ceramide-induced apoptosis via stimulation of the phosphatidylinositol 3-kinase/protein kinase B pathway. These findings constitute the first evidence for an “astroprotective” role of cannabinoids. Δ9-tetrahydrocannabinol extracellular signal-regulated kinase glial-fibrillary acidic protein phosphatidylinositol 3-kinase protein kinase B p90 ribosomal S6 kinase terminal dUTP nick-end labeling phosphate-buffered saline hemagglutinin The effects exerted by marijuana and their derivatives through Δ9-tetrahydrocannabinol (THC)1 and other cannabinoid constituents have been known for many centuries. However, the molecular basis of these actions were not understood until the discovery of an endogenous cannabinoid system comprising two plasma membrane Gi/o-coupled cannabinoid receptors (CB1 (1Matsuda L.A. Lolait S.J. Brownstein M. Young A. Bonner T.I. Nature. 1990; 346: 561-564Crossref PubMed Scopus (4215) Google Scholar) and CB2 (2Munro S. Thomas K.L. Abu-Shaar M. Nature. 1993; 365: 61-65Crossref PubMed Scopus (4150) Google Scholar)) and a family of endogenous ligands for those receptors (3Devane W.A. Hanus L. Breuer A. Pertwee R.G. Stevenson L.A. Griffin G. Gibson D. Mandelbaum A. Etinger A. Mechoulam R. Science. 1992; 258: 1946-1949Crossref PubMed Scopus (4690) Google Scholar, 4Mechoulam R. Ben Shabat S. Hanus L. Ligumsky M. Kaminski N.E. Schatz A.R. Gopher A. Almog S. Martin B.R. Compton D.R. Pertwee R.G. Griffin G. Bayewitch M. Barg J. Vogel Z. Biochem. Pharmacol. 1995; 50: 83-90Crossref PubMed Scopus (2356) Google Scholar). Cannabinoid receptors mediate cannabinoid effects by coupling to different signaling pathways. Both the CB1 and the CB2 receptor signal inhibition of adenylyl cyclase (5Howlett A.C. Annu. Rev. Pharmacol. Toxicol. 1995; 35: 607-634Crossref PubMed Scopus (447) Google Scholar) and stimulation of extracellular signal-regulated kinase (ERK) (6Bouaboula M. Poinot Chazel C. Bourrie B. Canat X. Calandra B. Rinaldi-Carmona M., Le Fur G. Casellas P. Biochem. J. 1995; 312: 637-641Crossref PubMed Scopus (458) Google Scholar), whereas the CB1 receptor is also coupled to modulation of Ca2+ and K+ channels (7Pertwee R.G. Expert. Opin. Investig. Drugs. 2000; 9: 1553-1571Crossref PubMed Scopus (195) Google Scholar), stimulation of the stress-activated p38 and c-Jun N-terminal kinases (8Rueda D. Galve-Roperh I. Haro A. Guzmán M. Mol. Pharmacol. 2000; 58: 814-820Crossref PubMed Scopus (169) Google Scholar), stimulation of the focal adhesion kinase (9Derkinderen P. Toutant M. Kadaré G. Ledent C. Parmentier M. Girault J.-A. J. Biol. Chem. 2001; 276: 38289-38296Abstract Full Text Full Text PDF PubMed Google Scholar), hydrolysis of sphingomyelin (10Sánchez C. Rueda D. Ségui B. Galve-Roperh I. Levade T. Guzmán M. Mol. Pharmacol. 2001; 59: 955-959Crossref PubMed Scopus (83) Google Scholar), and stimulation of phosphatidylinositol 3-kinase/protein kinase B (PI3K/PKB) (11Gómez del Pulgar T. Velasco G. Guzmán M. Biochem. J. 2000; 347: 369-373Crossref PubMed Scopus (218) Google Scholar). The study of the potential therapeutic applications of cannabinoids has become one of the most exciting areas in the field. Ongoing research is determining whether cannabinoid ligands may be effective agents in the treatment of pain, glaucoma, and the wasting and emesis associated with acquired immunodeficiency syndrome and cancer chemotherapy (7Pertwee R.G. Expert. Opin. Investig. Drugs. 2000; 9: 1553-1571Crossref PubMed Scopus (195) Google Scholar, 12Piomelli D. Giuffrida A. Calignano A. Rodrı́guez de Fonseca F. Trends Pharmacol. Sci. 2000; 21: 218-224Abstract Full Text Full Text PDF PubMed Scopus (405) Google Scholar). In addition, cannabinoids are being investigated as potential antitumoral drugs (13Galve-Roperh I. Sánchez C. Cortés M. Gómez del Pulgar T. Izquiedo M. Guzmán M. Nat. Med. 2000; 6: 313-319Crossref PubMed Scopus (570) Google Scholar, 14Sánchez C. de Ceballos M.L. Gómez del Pulgar T. Rueda D. Corbacho C. Velasco G. Galve-Roperh I. Huffman J.W.H. Ramón y Cajal S. Guzmán M. Cancer Res. 2001; 61: 5784-5789PubMed Google Scholar, 15Bifulco M. Laezza C. Portella G. Vitale M. Orlando P., De Petrocellis L. Di Marzo V. FASEB J. 2001; 15: 2745-2747Crossref PubMed Scopus (122) Google Scholar) and therapeutic agents for neurological and neurodegenerative disorders (16Baker D. Pryce G. Croxford J.L. Brown P. Pertwee R.G. Huffman J.W. Layward L. Nature. 2000; 404: 84-87Crossref PubMed Scopus (494) Google Scholar, 17Mechoulam R. Panikashvili D. Sholami E. Trends Mol. Med. 2002; 8: 58-61Abstract Full Text Full Text PDF PubMed Scopus (213) Google Scholar). Neuroprotection by cannabinoids has been related to the CB1-mediated inhibition of voltage-sensitive Ca2+ channels to reduce Ca2+influx, glutamate release and excitotoxicity (12Piomelli D. Giuffrida A. Calignano A. Rodrı́guez de Fonseca F. Trends Pharmacol. Sci. 2000; 21: 218-224Abstract Full Text Full Text PDF PubMed Scopus (405) Google Scholar, 18Shen M. Thayer S.A. Mol. Pharmacol. 1998; 54: 459-462Crossref PubMed Scopus (238) Google Scholar), and to the ability of cannabinoids to act as antioxidants (19Hampson A.J. Grimaldi M. Axelrod J. Wink D. Proc. Natl. Acad. Sci. U. S. A. 1998; 95: 8268-8273Crossref PubMed Scopus (678) Google Scholar, 20Marsicano G. Moosmann H. Lutz B. Bel C. J. Neurochem. 2002; 80: 448-456Crossref PubMed Scopus (249) Google Scholar). However, nothing is known about the possible protective effect of cannabinoids on the major cell population of the central nervous system, namely the astrocytes, despite the pivotal role played by these cells in brain homeostasis. In addition, although the CB1 receptor is coupled to PI3K/PKB (11Gómez del Pulgar T. Velasco G. Guzmán M. Biochem. J. 2000; 347: 369-373Crossref PubMed Scopus (218) Google Scholar) and ERK activation (6Bouaboula M. Poinot Chazel C. Bourrie B. Canat X. Calandra B. Rinaldi-Carmona M., Le Fur G. Casellas P. Biochem. J. 1995; 312: 637-641Crossref PubMed Scopus (458) Google Scholar), and both signaling routes are essential for neural cell survival (21Yuan J. Yankner B.A. Nature. 2000; 407: 802-809Crossref PubMed Scopus (1603) Google Scholar), their possible involvement in the protection of neural cells by cannabinoids is as yet unknown. Ceramide, a sphingosine-based lipid, regulates a variety of cellular processes including differentiation, proliferation, and apoptosis (22Kolesnick R.N. Krönke M. Annu. Rev. Physiol. 1998; 60: 643-665Crossref PubMed Scopus (730) Google Scholar). Interestingly, the pro-apoptotic effect of ceramide may be due, at least partially, to its ability to inhibit PKB (23Salinas M. López-Valdaliso R. Martı́n D. Álvarez A. Cuadrado A. Mol. Cell. Neurosci. 2000; 15: 156-169Crossref PubMed Scopus (175) Google Scholar, 24Schubert K.M. Scheid M.P. Duronio V. J. Biol. Chem. 2000; 275: 13330-13335Abstract Full Text Full Text PDF PubMed Scopus (215) Google Scholar). In addition, it has been shown that accumulation of ceramide in astrocytes leads to apoptosis (25Blázquez C. Galve-Roperh I. Guzmán M. FASEB J. 2000; 14: 2315-2322Crossref PubMed Scopus (141) Google Scholar). Here we employed a cell-permeable analog of ceramide to induce apoptosis in astrocytes, and we studied (i) the protective role of cannabinoids and (ii) the involvement of PI3K/PKB and ERK pathways in such effect. The following materials were kindly donated: HU-210 by Dr. R. Mechoulam (Hebrew University, Jerusalem, Israel); SR 141716 by Sanofi Synthelabo (Montpelier, France); antibodies against total PKB and RSK and the specific PKB/RSK peptide substrate (cross-tide) by Dr. D. Alessi (University of Dundee, Dundee, UK); and wild-type and dominant-negative PKB adenoviral vectors by Dr. W. Ogawa (Kobe University, Kobe, Japan). DNA fragmentation and TUNEL staining kits and biotin-16-dUTP were from Roche Molecular Biochemicals; deoxynucleotidyltransferase was from Invitrogen; streptavidin Alexa Fluor 488 was from Molecular Probes (Leiden, The Netherlands); wortmannin, LY 294002, PD 098059, Ro 318220, and C2-ceramide were from Alexis Biochemicals (San Diego, CA); anti-HA antibody was from Roche Molecular Biochemicals; anti-phospho-ERK antibody was from Santa Cruz Biotechnology (Santa Cruz, CA); anti-phospho-PKB Thr-308 and phospho-PKB Ser-473 were fromCell Signaling Technology (Beverly, MA); anti-glial fibrillary acidic protein (GFAP) polyclonal antibody was from DAKO (Glostrup, Denmark); ABC complex was from Pierce; and WIN 55,212-2 and THC were from Sigma. Cortical astrocytes were prepared from 24- to 48-h Wistar rats as described previously (25Blázquez C. Galve-Roperh I. Guzmán M. FASEB J. 2000; 14: 2315-2322Crossref PubMed Scopus (141) Google Scholar). Briefly, cerebral hemispheres were dissected in PBS supplemented with 0.33% glucose, treated with trypsin (5 mg/ml, 30 min at 37 °C), and after stopping the reaction by addition of 10% serum-containing medium, incubated with DNase I (10 μg/ml, 5 min at 37 °C). Subsequently cells were mechanically dissociated, centrifuged, and seeded (3 × 104 cells/cm2) on plastic plates previously coated with 5 μg/ml poly-l-ornithine and cultured in a mixture of Dulbecco's modified Eagle's medium and Ham's F-12 medium (1:1, v/v) supplemented with 0.5% (w/v) glucose, 5 mg/ml streptomycin, 5 units/ml penicillin, and 10% fetal calf serum. After 10–12 days, cells were trypsinized and reseeded until they reached confluency. Finally, cells were trypsinized, seeded at a density of 3 × 104 cells/cm2, and 24 h before the experiment transferred to a chemically defined serum-free medium consisting of Dulbecco's modified Eagle's medium/Ham's F-12 medium (1:1, v/v). Cell viability was determined by trypan blue exclusion. Oligonucleosomal DNA fragmentation, a characteristic biochemical feature of apoptotic cell death, was measured using a nucleosomal DNA enzyme-linked immunosorbent assay, which quantitatively records histone-associated DNA fragments, according to manufacturer's instructions. TUNEL staining was performed as described previously (26Gómez del Pulgar T. Velasco G. Sánchez C. Haro A. Guzmán M. Biochem. J. 2002; 363: 183-188Crossref PubMed Scopus (156) Google Scholar). PKB and RSK activities were determined as described (11Gómez del Pulgar T. Velasco G. Guzmán M. Biochem. J. 2000; 347: 369-373Crossref PubMed Scopus (218) Google Scholar). Briefly, PKB or RSK was immunoprecipitated from cell lysates with 2 μg of anti-PKBα or anti-RSK antibodies bound to protein G-Sepharose, and kinase activity was determined as the incorporation of [γ-32P]ATP into a specific peptide substrate (GRPRTSSFAEG). Western blot analyses were performed with antibodies that recognize ERK phosphorylated on Thr-202/Tyr-204, PKB-phosphorylated on Thr-308, and PKB-phosphorylated on Ser-473. Adenoviral vectors encoding HA-tagged dominant-negative and wild-type PKB were amplified as described (27Kitamura T. Ogawa W. Sakaue H. Hino Y. Kuroda S. Takata M. Matsumoto M. Maeda T. Konishi H. Kikkawa U. Kasuga M. Mol. Cell. Biol. 1998; 18: 3708-3717Crossref PubMed Scopus (296) Google Scholar). Astrocytes were transferred to serum-free medium, infected for 3 h with the corresponding adenoviral vector at the multiplicity of infection indicated in the figures, washed with PBS, and transferred to a 10% fetal calf serum medium for 12 h to recover from the infection. Before performing the experiments, infected cells were incubated for 24 h in serum-free medium. Pilot experiments using adenoviruses encoding the green fluorescent protein showed that >95% were infected in our experimental conditions. Expression of HA-tagged wild-type and dominant-negative forms of PKB was confirmed in the infected astrocytes by Western blot analysis with anti-HA antibody. Male Wistar rats (320–350 g) were anaesthetized with equitesin (3.5 ml/kg) and injected stereotactically with C2-ceramide (10 mg/ml in Me2SO) at two sites in the hippocampus. In preliminary experiments the volume and number of sites of C2-ceramide injection were established. Twenty μg were injected into the dorsal dentate gyrus and another 20 μg into the dorsal hippocampus (anteroposterior, bregma −3.8 mm; lateral −3.0 mm, and ventral to the surface of the brain −3.4 and −2.6 mm, respectively). C2-ceramide or vehicle were slowly injected (1 μl/min). The needle was left in place for 2 min before retraction to the more dorsal coordinate, and after injection at the second site left in place for a further 5 min before final retraction. WIN 55,212-2 (2.5 mg/kg, intraperitoneal in 1 ml/kg of 10% Me2SO in saline) was administered 10 min before anesthetic injection and 30 min before focal injection. All procedures were conducted according to the guidelines of the European Community (EC) and were approved by the ethical committee of the Centro Superior de Investigaciones Cientificas (CSIC). Two days post-injection animals were decapitated, the brains removed, and 4-mm coronal slabs around the injected area cut, fixed by immersion in 4% paraformaldehyde in 0.1m phosphate buffer for 3 days, and cryoprotected with 15% sucrose for 24 h and then with 30% sucrose for a further 24 h at 4 °C. Finally, brain slabs were flash-frozen in hexane (−70 °C) and stored at −20 °C until sectioned at 45 μm in a cryostat. TUNEL staining of mounted tissue sections was performed according to the manufacturer's instructions. GFAP immunostaining was performed on free-floating sections. Sections were washed 3 times in PBS, treated with 3% H2O2 for 15 min to block endogenous peroxidase, and rinsed 3 times in PBS. After incubation with 10% normal goat serum (NGS) in PBS containing 0.3% Triton X-100 for 30 min, sections were incubated with anti-GFAP polyclonal antibody (1:1000) in PBS containing 1% NGS and 0.3% Triton X-100 for 6 h at room temperature and overnight at 4 °C. Immunostaining was visualized using the ABC complex and diaminobenzidine oxidation (0.07% plus 0.05% H2O2) and analyzed on a Zeiss microscope by an observer unaware of the different treatments. Results shown represent means ± S.D. Statistical analysis was performed by analysis of variance with apost hoc analysis by the Student-Neuman-Keuls test. We employed the pro-apoptotic lipid C2-ceramide to study the potential protective effect of cannabinoids in primary astrocyte cultures. As shown in Fig.1 A, ceramide-induced astrocyte death was notably reduced by incubation with THC or different synthetic cannabinoids. We employed the cannabinoid agonist WIN 55,212-2 to characterize this effect further. Protection by WIN 55,212-2 was dose-dependent (Fig. 1 B) and reached a maximum at 18 h after the addition of the cannabinoid (Fig.1 C). Next, we investigated the nature of ceramide-induced cell death. Challenge with ceramide induced apoptosis as indicated by TUNEL (Fig. 2 A) and DNA fragmentation enzyme-linked immunosorbent assay (Fig. 2 B), whereas incubation with the cannabinoid agonist HU-210 prevented ceramide-induced apoptosis.Figure 2Cannabinoids prevent ceramide-induced apoptosis. A, astrocytes were incubated in serum-free medium for 24 h and treated with 15 μmC2-ceramide or vehicle (Control) for 90 min. Then, the medium was changed; vehicle (−) or 25 nm HU-210 (HU) was added, and apoptotic DNA fragmentation was determined. Results correspond to four different experiments. *, significantly different (p < 0.01) from the controls.B, cells were treated as in A, and TUNEL staining was performed. Representative micrographs (phase contrast and TUNEL-stained cells) from one experiment are shown. Similar data were obtained in two additional experiments.View Large Image Figure ViewerDownload Hi-res image Download (PPT) We employed pharmacological inhibitors as a first approach to the mechanism of the anti-apoptotic action of cannabinoids in astrocytes. Thus, incubation with SR 141716 (a CB1 receptor antagonist), LY 294002 and wortmannin (two structurally unrelated PI3K inhibitors), PD 098059 (an ERK pathway inhibitor), and Ro 318220 (a protein kinase C inhibitor that has been shown to inhibit equally the ERK-downstream kinase RSK (28Alessi D.R. FEBS Lett. 1997; 402: 121-123Crossref PubMed Scopus (198) Google Scholar)) abrogated the anti-apoptotic effect of cannabinoids (Fig.3), suggesting that this effect is dependent on the CB1 receptor and the PI3K and ERK pathways. It is well established that stimulation of the PI3K pathway leads to activation of the anti-apoptotic kinase PKB (29Vanhaesebroeck B. Alessi D.R. Biochem. J. 2000; 346: 561-576Crossref PubMed Scopus (1399) Google Scholar). As shown in Fig. 4 A, incubation of astrocytes with HU-210 stimulated and incubation with ceramide inhibited PKB activity. Interestingly, incubation with HU-210 also prevented ceramide-induced inhibition of PKB activity (Fig4 A). Because activation of PKB depends on its phosphorylation on residues Thr-308 and Ser-473 (29Vanhaesebroeck B. Alessi D.R. Biochem. J. 2000; 346: 561-576Crossref PubMed Scopus (1399) Google Scholar), we monitored the phosphorylation status of PKB in astrocytes by using specific antibodies raised against the phosphorylated forms of the kinase. Fig.4 B shows that changes in PKB phosphorylation paralleled changes in enzyme activity. Thus, incubation of astrocytes with HU-210 increased and incubation with ceramide decreased PKB phosphorylation on Thr-308 and Ser-473. In addition, after ceramide challenge, incubation with cannabinoids led PKB phosphorylation to the control level. To confirm the involvement of PKB in the anti-apoptotic effect of cannabinoids, we expressed dominant-negative or wild-type forms of PKB (27Kitamura T. Ogawa W. Sakaue H. Hino Y. Kuroda S. Takata M. Matsumoto M. Maeda T. Konishi H. Kikkawa U. Kasuga M. Mol. Cell. Biol. 1998; 18: 3708-3717Crossref PubMed Scopus (296) Google Scholar) in astrocytes. Because primary cells are transfected with very low efficiency, we used adenoviral vectors to ensure that >95% of the cells express the exogenous proteins. As shown in Fig. 4 C, expression of a dominant-negative form of PKB abrogated the protective effect of cannabinoids. In addition, infection with the wild-type form of PKB led to a dose-dependent blockade of the apoptotic effect of ceramide (Fig. 4 D), supporting the notion that the pro-apoptotic effect of this lipid may be mediated, at least partially, by PKB inhibition. As data in Fig. 3 indicated that the protective effect of cannabinoids on astrocytes could also involve the ERK pathway, we determined the extent of ERK activation in the cells by using an antibody raised against the phosphorylated (active) form of this kinase. As shown in Fig.5 A, incubation with HU-210 increased the phosphorylation extent of ERK in the presence and in the absence of ceramide, whereas incubation with ceramide only slightly stimulated ERK. Incubation with SR 141716 or wortmannin partially prevented ERK activation after challenge to ceramide plus HU-210. We also determined the activity of the ERK downstream kinase RSK. As shown in Fig. 5 B, incubation with cannabinoids or ceramide alone induced a 60–80% stimulation of RSK, and treatment with both compounds led to an additive stimulation. The latter effect was prevented by both wortmannin and SR 141716. By contrast, ceramide stimulation of RSK was not affected by incubation with wortmannin or SR141716. We next examined the role of cannabinoids in protecting astrocytes in vivo. As shown in Fig. 6 A, treatment with WIN 55,212-2 prevented the toxic effects of focal administration of C2-ceramide in astrocytes. Thus, whereas administration of ceramide induced an area absolutely devoid of GFAP immunoreactivity coinciding with the site of injection (the ventral dentate gyrus), rats treated with WIN 55,212-2 showed a homogeneous GFAP staining throughout the whole hippocampus and did not present an injured area in the zone of injection. GFAP staining remained increased compared with normal rats or to the contralateral non-injected hemisphere of the brain in both cannabinoid- and vehicle-treated rats. In addition, as shown in Fig. 6 B there was a high number of TUNEL-positive nuclei in ceramide-injected hippocampus that was significantly reduced by cannabinoid administration (number of TUNEL-positive nuclei/mm2: 994 ± 236 after C2-ceramide treatment, 624 ± 193 after WIN 55,212-2 plus C2-ceramide treatment, p < 0.01). No TUNEL-positive nuclei were observed in vehicle-injected controls. During the last few years, a number of reports have indicated that cannabinoids protect nervous cells from different insults (reviewed in Refs. 12Piomelli D. Giuffrida A. Calignano A. Rodrı́guez de Fonseca F. Trends Pharmacol. Sci. 2000; 21: 218-224Abstract Full Text Full Text PDF PubMed Scopus (405) Google Scholar and 17Mechoulam R. Panikashvili D. Sholami E. Trends Mol. Med. 2002; 8: 58-61Abstract Full Text Full Text PDF PubMed Scopus (213) Google Scholar). In line with those observations, data presented here show that cannabinoids, via activation of the CB1 receptor, protect astrocytes from ceramide-induced apoptosis in vitroand in vivo. Astrocytes have been traditionally considered as secondary players in the central nervous system scenario, and therefore all the previous studies on the protective role of cannabinoids on neural cells have involved neurons (see Refs. 18Shen M. Thayer S.A. Mol. Pharmacol. 1998; 54: 459-462Crossref PubMed Scopus (238) Google Scholar and30Nagayama T. Sinor A.D. Simon R.P. Chen J. Graham S.H. Jin K. Greenberg D.A. J. Neurosci. 1999; 19: 2987-2995Crossref PubMed Google Scholar, 31Sinor A.D. Irvin S.M. Greenberg D.A. Neurosci. Lett. 2000; 278: 1257-1260Crossref Scopus (166) Google Scholar, 32Panikashvili D. Simeonidou C. Ben-Shabat S. Hanus L. Breuer A. Mechoulam R. Shohami E. Nature. 2001; 413: 527-531Crossref PubMed Scopus (637) Google Scholar, 33Van der Stelt M. Veldhuis W.B. Bär P.R. Veldink G.A. Vliegenthart J.F.G. Nicolay K. J. Neurosci. 2001; 21: 6475-6479Crossref PubMed Google Scholar, 34Van der Stelt M. Veldhuis W.B. van Haaften G.W. Fezza F. Bisogno T. Bär P.R. Veldink G.A. Vliegenthart J.F.G., Di Marzo V. Nicolay K. J. Neurosci. 2001; 21: 8765-8771Crossref PubMed Google Scholar, for example). However, it is currently well established that astrocytes, the most abundant cells of the mammalian brain, are involved in numerous functions such as supply of nutrients to neurons (35Tsacopoulos M. Magistretti P.J. J. Neurosci. 1996; 16: 877-885Crossref PubMed Google Scholar), establishment of synapses (36Ullian E.M. Sapperstein S.K. Christopherson K.S. Barres B.A. Science. 2001; 291: 657-661Crossref PubMed Scopus (1061) Google Scholar), and generation of neurons (37Doetsch F. Caillé I. Lim D.A. Garcı́a-Verdugo J.M. Álvarez-Buylla A. Cell. 1999; 97: 703-716Abstract Full Text Full Text PDF PubMed Scopus (3242) Google Scholar). In addition, in the context of the present study astrocytes are known to take up (38Di Marzo V. Fontana A. Cadas H. Schinelli S. Cimino G. Schwartz J.C. Piomelli D. Nature. 1994; 372: 686-691Crossref PubMed Scopus (1349) Google Scholar) and produce (39Walter L. Franklin A. Witting A. Moller T. Stella N. J. Biol. Chem. 2002; 277: 20869-20876Abstract Full Text Full Text PDF PubMed Scopus (156) Google Scholar) endocannabinoids. Thus, most likely the complex mechanisms underlying defense against brain injury (and in particular the mechanisms mediated by cannabinoids) also involve protection of astrocytes. Several observations presented in this report indicate that cannabinoids protect primary astrocytes from ceramide-induced apoptosis via CB1 receptor-mediated stimulation of the PI3K/PKB pathway. (i) Blockade of the CB1 receptor or inhibition of PI3K abolishes the protective effect of cannabinoids. (ii) Cannabinoid treatment leads to reactivation of PKB in parallel to prevention of apoptosis. (iii) Overexpression of a dominant-negative form of PKB abrogates the protective effect of cannabinoids. It is well established that challenge with ceramide leads to apoptosis in several experimental models, and this may be at least partially due to dephosphorylation and inactivation of PKB by a ceramide-activated phosphatase (23Salinas M. López-Valdaliso R. Martı́n D. Álvarez A. Cuadrado A. Mol. Cell. Neurosci. 2000; 15: 156-169Crossref PubMed Scopus (175) Google Scholar, 24Schubert K.M. Scheid M.P. Duronio V. J. Biol. Chem. 2000; 275: 13330-13335Abstract Full Text Full Text PDF PubMed Scopus (215) Google Scholar). Our results suggest that cannabinoids (via activation of the PI3K pathway) and ceramide (via phosphatase activation) may compete for the modulation of PKB activity in astrocytes. Supporting this notion, overexpression of ceramide-sensitive wild-type PKB abrogated the apoptotic effect of ceramide. Because activation of PKB triggers the phosphorylation of different targets involved in preventing apoptosis, including Bad, forkhead transcription factors, IκB kinase, and caspase 9 (29Vanhaesebroeck B. Alessi D.R. Biochem. J. 2000; 346: 561-576Crossref PubMed Scopus (1399) Google Scholar), ceramide inhibition of PKB could lead to suppression of the survival signal, whereas cannabinoid-dependent reactivation of the pathway would restore it. Expression of a dominant-negative form of PKB abolishes the protective effect of cannabinoids but does not induce apoptosis by itself, indicating that the apoptotic effect of ceramide and therefore the generation of a survival signal may also depend on the modulation of additional pathways. Thus, several data suggest that the ERK pathway may participate together with PKB activation in the anti-apoptotic effect of cannabinoids as follows: (i) inhibition of the ERK pathway also prevents the protective effect of cannabinoids, and (ii) astrocyte challenge with cannabinoids leads to activation of both ERK and RSK. One of the mechanisms whereby ERK prevents apoptosis in neural cells involves activation of its downstream kinase RSK as this kinase phosphorylates Bad and the transcription factor cAMP-response element-binding protein (21Yuan J. Yankner B.A. Nature. 2000; 407: 802-809Crossref PubMed Scopus (1603) Google Scholar). Thus RSK may act synergistically with PKB to prevent apoptosis (40Nebreda A.R. Gavin A.-C. Science. 1999; 286: 1309-1310Crossref PubMed Scopus (73) Google Scholar). In our model, triggering of the survival signal is accompanied by a consistent activation of ERK and RSK. Nevertheless, incubation with ceramide leads to apoptosis and activation of ERK and RSK, although to a lower extent than with cannabinoid co-treatment. Interestingly, blockade of PI3K prevents the effect of cannabinoids on ERK and RSK but not ceramide-induced activation of these kinases. These data are in line with recent results of our group 2I. Galve-Roperh, D. Rueda, T. Gómez del Pulgar, G. Velasco, and M. Guzmán, submitted for publication. showing that stimulation of ERK by cannabinoids depends on PI3K and suggest that the latter may be involved in the pro-survival effect of cannabinoids also via activation of the ERK/RSK pathway. It is worth noting that RSK activation also depends on phosphorylation by 3-phosphoinositide-dependent kinase 1 on its N-terminal domain (41Williams M.R. Arthur J.S.C. Balendran A. Van der Kaay J. Poli V. Cohen P. Alessi D.R. Curr. Biol. 2000; 10: 439-448Abstract Full Text Full Text PDF PubMed Scopus (396) Google Scholar). Although that phosphorylation site has been suggested to be constitutive (41Williams M.R. Arthur J.S.C. Balendran A. Van der Kaay J. Poli V. Cohen P. Alessi D.R. Curr. Biol. 2000; 10: 439-448Abstract Full Text Full Text PDF PubMed Scopus (396) Google Scholar), it cannot be ruled out that under certain circumstances PI3K activation could lead to 3-phosphoinositide-dependent kinase 1-dependent phosphorylation and activation of RSK (42Park J. Hill M.M. Hess D. Brazil D.P. Hofsteenge J. Hemmings B.A. J. Biol. Chem. 2001; 276: 37459-37471Abstract Full Text Full Text PDF PubMed Scopus (102) Google Scholar). In short, data presented here indicate that cannabinoids protect primary astrocytes from ceramide-induced apoptosis via activation of the PI3K/PKB pathway. Our data also suggest that cannabinoids are involved in protecting astrocytes in vivo. Although the mechanisms of ceramide generation in astrocytes in vivo are still unknown, it is possible that exposure to proinflammatory cytokines (43Singh I. Pahan K. Khan M. Singh A.K. J. Biol. Chem. 1998; 273: 20354-20362Abstract Full Text Full Text PDF PubMed Scopus (182) Google Scholar) or to saturated fatty acids (25Blázquez C. Galve-Roperh I. Guzmán M. FASEB J. 2000; 14: 2315-2322Crossref PubMed Scopus (141) Google Scholar) may increase ceramide production in astrocytes during situations of brain injury. It is curious that, unlike this protective effect on astrocytes, cannabinoids induce apoptosis of glioma cells (13Galve-Roperh I. Sánchez C. Cortés M. Gómez del Pulgar T. Izquiedo M. Guzmán M. Nat. Med. 2000; 6: 313-319Crossref PubMed Scopus (570) Google Scholar, 14Sánchez C. de Ceballos M.L. Gómez del Pulgar T. Rueda D. Corbacho C. Velasco G. Galve-Roperh I. Huffman J.W.H. Ramón y Cajal S. Guzmán M. Cancer Res. 2001; 61: 5784-5789PubMed Google Scholar,26Gómez del Pulgar T. Velasco G. Sánchez C. Haro A. Guzmán M. Biochem. J. 2002; 363: 183-188Crossref PubMed Scopus (156) Google Scholar). This difference between transformed (glioma) and non-transformed cells (astrocytes) could be due to their ability to synthesize ceramide in response to cannabinoids. Thus, cannabinoids induce apoptosis on glioma cells via stimulation of ceramide synthesis de novo(26Gómez del Pulgar T. Velasco G. Sánchez C. Haro A. Guzmán M. Biochem. J. 2002; 363: 183-188Crossref PubMed Scopus (156) Google Scholar), whereas challenge to cannabinoids does not induce ceramide synthesis de novo in astrocytes. 3T. Gómez del Pulgar, G. Velasco, and M. Guzmán, unpublished results. Taken together, these data suggest that cannabinoid receptors are coupled to different pathways and therefore lead to different responses in glioma cells and astrocytes. Accordingly, cannabinoids are being tested as potential antitumoral drugs in the treatment of malignant gliomas and, given the crucial role of astrocytes in brain homeostasis and neuroprotection, our results raise the suggestive although still speculative idea of their usage as therapeutic agents for the management of neurodegenerative disorders. We are grateful to Dr. D. Alessi, Dr. W. Ogawa, Dr. C. Sutherland, Dr. R. Mechoulam, and Sanofi Synthelabo for the kind donation of reagents; Dr. J. Lizcano and Dr. I. Galve-Roperh for helpful suggestions on the signaling experiments; Dr. L. López- Mascaraque and Dr. L. M. Garcı́a-Segura for helpful suggestions on the in vivo experiments; and A. Carracedo, Dr. C. Blázquez, Dr. D. Rueda, and M. E. Fernández de Molina for technical assistance.
DOI: 10.1016/j.pnpbp.2015.05.010
2016
Cited 134 times
The use of cannabinoids as anticancer agents
It is well-established that cannabinoids exert palliative effects on some cancer-associated symptoms. In addition evidences obtained during the last fifteen years support that these compounds can reduce tumor growth in animal models of cancer. Cannabinoids have been shown to activate an ER-stress related pathway that leads to the stimulation of autophagy-mediated cancer cell death. In addition, cannabinoids inhibit tumor angiogenesis and decrease cancer cell migration. The mechanisms of resistance to cannabinoid anticancer action as well as the possible strategies to develop cannabinoid-based combinational therapies to fight cancer have also started to be explored. In this review we will summarize these observations (that have already helped to set the bases for the development of the first clinical studies to investigate the potential clinical benefit of using cannabinoids in anticancer therapies) and will discuss the possible future avenues of research in this area.
DOI: 10.1074/jbc.m608900200
2007
Cited 130 times
Cannabinoids Induce Glioma Stem-like Cell Differentiation and Inhibit Gliomagenesis
Glioma stem-like cells constitute one of the potential origins of gliomas, and therefore, their elimination is an essential factor for the development of efficient therapeutic strategies. Cannabinoids are known to exert an antitumoral action on gliomas that relies on at least two mechanisms: induction of apoptosis of transformed cells and inhibition of tumor angiogenesis. However, whether cannabinoids target human glioma stem cells and their potential impact in gliomagenesis are unknown. Here, we show that glioma stem-like cells derived from glioblastoma multiforme biopsies and the glioma cell lines U87MG and U373MG express cannabinoid type 1 (CB<sub>1</sub>) and type 2 (CB<sub>2</sub>) receptors and other elements of the endocannabinoid system. In gene array experiments, CB receptor activation altered the expression of genes involved in the regulation of stem cell proliferation and differentiation. The cannabinoid agonists HU-210 and JWH-133 promoted glial differentiation in a CB receptor-dependent manner as shown by the increased number of S-100β- and glial fibrillary acidic protein-expressing cells. In parallel, cannabinoids decreased the cell population expressing the neuroepithelial progenitor marker nestin. Moreover, cannabinoid challenge decreased the efficiency of glioma stem-like cells to initiate glioma formation <i>in vivo</i>, a finding that correlated with decreased neurosphere formation and cell proliferation in secondary xenografts. Gliomas derived from cannabinoid-treated cancer stem-like cells were characterized with a panel of neural markers and evidenced a more differentiated phenotype and a concomitant decrease in nestin expression. Overall, our results demonstrate that cannabinoids target glioma stem-like cells, promote their differentiation, and inhibit gliomagenesis, thus giving further support to their potential use in the management of malignant gliomas.
DOI: 10.1038/jid.2015.45
2015
Cited 129 times
Exploiting Cannabinoid-Induced Cytotoxic Autophagy to Drive Melanoma Cell Death
Although the global incidence of cutaneous melanoma is increasing, survival rates for patients with metastatic disease remain <10%. Novel treatment strategies are therefore urgently required, particularly for patients bearing BRAF/NRAS wild-type tumors. Targeting autophagy is a means to promote cancer cell death in chemotherapy-resistant tumors, and the aim of this study was to test the hypothesis that cannabinoids promote autophagy-dependent apoptosis in melanoma. Treatment with Δ<sup>9</sup>-Tetrahydrocannabinol (THC) resulted in the activation of autophagy, loss of cell viability, and activation of apoptosis, whereas cotreatment with chloroquine or knockdown of Atg7, but not Beclin-1 or Ambra1, prevented THC-induced autophagy and cell death <i>in vitro</i>. Administration of Sativex-like (a laboratory preparation comprising equal amounts of THC and cannabidiol (CBD)) to mice bearing BRAF wild-type melanoma xenografts substantially inhibited melanoma viability, proliferation, and tumor growth paralleled by an increase in autophagy and apoptosis compared with standard single-agent temozolomide. Collectively, our findings suggest that THC activates noncanonical autophagy-mediated apoptosis of melanoma cells, suggesting that cytotoxic autophagy induction with Sativex warrants clinical evaluation for metastatic disease.
DOI: 10.1080/15548627.2016.1213927
2016
Cited 121 times
Dihydroceramide accumulation mediates cytotoxic autophagy of cancer cells via autolysosome destabilization
Autophagy is considered primarily a cell survival process, although it can also lead to cell death. However, the factors that dictate the shift between these 2 opposite outcomes remain largely unknown. In this work, we used Δ9-tetrahydrocannabinol (THC, the main active component of marijuana, a compound that triggers autophagy-mediated cancer cell death) and nutrient deprivation (an autophagic stimulus that triggers cytoprotective autophagy) to investigate the precise molecular mechanisms responsible for the activation of cytotoxic autophagy in cancer cells. By using a wide array of experimental approaches we show that THC (but not nutrient deprivation) increases the dihydroceramide:ceramide ratio in the endoplasmic reticulum of glioma cells, and this alteration is directed to autophagosomes and autolysosomes to promote lysosomal membrane permeabilization, cathepsin release and the subsequent activation of apoptotic cell death. These findings pave the way to clarify the regulatory mechanisms that determine the selective activation of autophagy-mediated cancer cell death.
DOI: 10.1158/2326-6066.cir-17-0475
2018
Cited 89 times
The Clinical Activity of PD-1/PD-L1 Inhibitors in Metastatic Non–Clear Cell Renal Cell Carcinoma
Programmed death 1 (PD-1) and PD ligand 1 (PD-L1) inhibitors have shown activity in metastatic clear cell renal cell carcinoma (ccRCC). Data on the activity of these agents in patients with non-clear cell RCC (nccRCC) or patients with sarcomatoid/rhabdoid differentiation are limited. In this multicenter analysis, we explored the efficacy of PD-1/PD-L1 inhibitors in patients with nccRCC or sarcomatoid/rhabdoid differentiation. Baseline and follow-up demographic, clinical, treatment, and radiographic data were collected. The primary endpoint was objective response rate. Secondary endpoints include time-to-treatment failure (TTF), overall survival (OS), and biomarker correlates. Forty-three patients were included: papillary (n = 14; 33%), chromophobe (n = 10; 23%), unclassified (n = 9; 21%), translocation (n = 3; 7%), and ccRCC with sarcomatoid differentiation (n = 7, 16%). Of those 43 patients, 11 patients (26%) had sarcomatoid and/or rhabdoid differentiation (n = 7 with ccRCC; n = 4 nccRCC). Overall, 8 patients (19%) objectively responded, including 4 patients (13%) who received PD-1/PD-L1 monotherapy. Responses were observed in patients with ccRCC with sarcomatoid and/or rhabdoid differentiation (n = 3/7, 43%), translocation RCC (n = 1/3, 33%), and papillary RCC (n = 4/14, 29%). The median TTF was 4.0 months [95% confidence interval (CI), 2.8-5.5] and median OS was 12.9 months (95% CI, 7.4-not reached). No specific genomic alteration was associated with clinical benefit. Modest antitumor activity for PD-1/PD-L1-blocking agents was observed in some patients with nccRCC. Further prospective studies are warranted to investigate the efficacy of PD-1/PD-L1 blockade in this heterogeneous patient population. Cancer Immunol Res; 6(7); 758-65. ©2018 AACR.
DOI: 10.1016/j.bcp.2018.09.007
2018
Cited 77 times
Targeting Glioma Initiating Cells with A combined therapy of cannabinoids and temozolomide
Glioblastoma multiforme (GBM) is the most frequent and aggressive type of brain tumor due, at least in part, to its poor response to current anticancer treatments. These features could be explained, at least partially, by the presence within the tumor mass of a small population of cells termed Glioma Initiating Cells (GICs) that has been proposed to be responsible for the relapses occurring in this disease. Thus, the development of novel therapeutic approaches (and specifically those targeting the population of GICs) is urgently needed to improve the survival of the patients suffering this devastating disease. Previous observations by our group and others have shown that Δ9-Tetrahydrocannabinol (THC, the main active ingredient of marijuana) and other cannabinoids including cannabidiol (CBD) exert antitumoral actions in several animal models of cancer, including gliomas. We also found that the administration of THC (or of THC + CBD at a 1:1 ratio) in combination with temozolomide (TMZ), the benchmark agent for the treatment of GBM, synergistically reduces the growth of glioma xenografts. In this work we investigated the effect of the combination of TMZ and THC:CBD mixtures containing different ratios of the two cannabinoids in preclinical glioma models, including those derived from GICs. Our findings show that TMZ + THC:CBD combinations containing a higher proportion of CDB (but not TMZ + CBD alone) produce a similar antitumoral effect as the administration of TMZ together with THC and CBD at a 1:1 ratio in xenografts generated with glioma cell lines. In addition, we also found that the administration of TMZ + THC:CBD at a 1:1 ratio reduced the growth of orthotopic xenografts generated with GICs derived from GBM patients and enhanced the survival of the animals bearing these intracranial xenografts. Remarkably, the antitumoral effect observed in GICs-derived xenografts was stronger when TMZ was administered together with cannabinoid combinations containing a higher proportion of CBD. These findings support the notion that the administration of TMZ together with THC:CBD combinations - and specifically those containing a higher proportion of CBD - may be therapeutically explored to target the population of GICs in GBM.
DOI: 10.1080/15548627.2020.1761651
2020
Cited 76 times
The anti-cancer drug ABTL0812 induces ER stress-mediated cytotoxic autophagy by increasing dihydroceramide levels in cancer cells
ABTL0812 is a first-in-class small molecule with anti-cancer activity, which is currently in clinical evaluation in a phase 2 trial in patients with advanced endometrial and squamous non-small cell lung carcinoma (NCT03366480). Previously, we showed that ABTL0812 induces TRIB3 pseudokinase expression, resulting in the inhibition of the AKT-MTORC1 axis and macroautophagy/autophagy-mediated cancer cell death. However, the precise molecular determinants involved in the cytotoxic autophagy caused by ABTL0812 remained unclear. Using a wide range of biochemical and lipidomic analyses, we demonstrated that ABTL0812 increases cellular long-chain dihydroceramides by impairing DEGS1 (delta 4-desaturase, sphingolipid 1) activity, which resulted in sustained ER stress and activated unfolded protein response (UPR) via ATF4-DDIT3-TRIB3 that ultimately promotes cytotoxic autophagy in cancer cells. Accordingly, pharmacological manipulation to increase cellular dihydroceramides or incubation with exogenous dihydroceramides resulted in ER stress, UPR and autophagy-mediated cancer cell death. Importantly, we have optimized a method to quantify mRNAs in blood samples from patients enrolled in the ongoing clinical trial, who showed significant increased DDIT3 and TRIB3 mRNAs. This is the first time that UPR markers are reported to change in human blood in response to any drug treatment, supporting their use as pharmacodynamic biomarkers for compounds that activate ER stress in humans. Finally, we found that MTORC1 inhibition and dihydroceramide accumulation synergized to induce autophagy and cytotoxicity, phenocopying the effect of ABTL0812. Given the fact that ABTL0812 is under clinical development, our findings support the hypothesis that manipulation of dihydroceramide levels might represents a new therapeutic strategy to target cancer.Abbreviations: 4-PBA: 4-phenylbutyrate; AKT: AKT serine/threonine kinase; ATG: autophagy related; ATF4: activating transcription factor 4; Cer: ceramide; DDIT3: DNA damage inducible transcript 3; DEGS1: delta 4-desaturase, sphingolipid 1; dhCer: dihydroceramide; EIF2A: eukaryotic translation initiation factor 2 alpha; EIF2AK3: eukaryotic translation initiation factor 2 alpha kinase 3; ER: endoplasmic reticulum; HSPA5: heat shock protein family A (Hsp70) member 5; MAP1LC3B: microtubule associated protein 1 light chain 3 beta; MEF: mouse embryonic fibroblast; MTORC1: mechanistic target of rapamycin kinase complex 1; NSCLC: non-small cell lung cancer; THC: Δ9-tetrahydrocannabinol; TRIB3: tribbles pseudokinase 3; XBP1: X-box binding protein 1; UPR: unfolded protein response.
DOI: 10.1016/j.eururo.2019.07.048
2019
Cited 75 times
First-line Immuno-Oncology Combination Therapies in Metastatic Renal-cell Carcinoma: Results from the International Metastatic Renal-cell Carcinoma Database Consortium
In metastatic renal-cell carcinoma (mRCC), recent data have shown efficacy of first-line ipilimumab and nivolumab (ipi-nivo) as well as immuno-oncology (IO)/vascular endothelial growth factor (VEGF) inhibitor combinations. Comparative data between these strategies are limited. To compare the efficacy of ipi-nivo versus IO-VEGF (IOVE) combinations in mRCC, and describe practice patterns and effectiveness of second-line therapies. Using the International Metastatic Renal-cell Carcinoma Database Consortium (IMDC) dataset, patients treated with any first-line IOVE combination were compared with those treated with ipi-nivo. All patients received first-line IO combination therapies. First- and second-line response rates, time to treatment failure (TTF), time to next treatment (TNT), and overall survival (OS) were analysed. Hazard ratios were adjusted for IMDC risk factors. In total, 113 patients received IOVE combinations and 75 received ipi-nivo. For IOVE combinations versus ipi-nivo, first-line response rates were 33% versus 40% (between-group difference 7%, 95% confidence interval [CI] –8% to 22%, p = 0.4), TTF was 14.3 versus 10.2 mo (p = 0.2), TNT was 19.7 versus 17.9 mo (p = 0.4), and median OS was immature but not statistically different (p = 0.17). Adjusted hazard ratios for TTF, TNT, and OS were 0.71 (95% CI 0.46–1.12, p = 0.14), 0.65 (95% CI 0.38–1.11, p = 0.11), and 1.74 (95% CI 0.82–3.68, p = 0.14), respectively. Sixty-four (34%) patients received second-line treatment. In patients receiving subsequent VEGF-based therapy, second-line response rates were lower in the IOVE cohort than in the ipi-nivo cohort (15% vs 45%; between-group difference 30%, 95% CI 3–57%, p = 0.04; n = 40), though second-line TTF was not significantly different (3.7 vs 5.4 mo; p = 0.4; n = 55). Limitations include the study’s retrospective design and sample size. There were no significant differences in first-line outcomes between IOVE combinations and ipi-nivo. Most patients received VEGF-based therapy in the second line. In this group, second-line response rate was greater in patients who received ipi-nivo initially. There were no significant differences in key first-line outcomes for patients with metastatic renal-cell carcinoma receiving immuno-oncology/vascular endothelial growth factor inhibitor combinations versus ipilimumab and nivolumab.
DOI: 10.18632/oncotarget.24283
2018
Cited 74 times
Analysis of response rate with ANTI PD1/PD-L1 monoclonal antibodies in advanced solid tumors: a meta-analysis of randomized clinical trials
Anti-PD1/PD-L1 monoclonal antibodies (mAbs) increase overall survival compared to standard of care (SOC) in different tumors. However, a proportion of patients (pts) will have progressive disease (PD) as best response. We conducted a meta-analysis to study the rates of response comparing these antibodies with SOC.A search of published trials in MEDLINE and EMBASE analyzing anti-PD1/PD-L1mAbs monotherapy compared to SOC. Relative risk (RR) with 95% confidence interval (CI) of response rates between groups was estimated. Subgroup analyses for location of primary tumor, number of previous treatment lines, selected population by PD-L1 expression and type of radiological assessment were made.Twelve studies accounting for 6,700 pts were included (anti-PD1/PD-L1 mAbs: 3,451 pts; SOC: 3,249 pts [2,823 pts: chemotherapy, 426 pts: targeted therapy]). Adjusted response rates were (N, %): Complete Response (CR) (69/3153, 2.19%), Partial Response (PR) (596/3153, 18.90%), Stable Disease (SD) (632/2463, 25.66%) and PD (1027/2463, 41.70%); and CR (16/2955, 0.54%), PR (263/2955, 8.90%), SD (835/2269, 36.80%) and PD (834/2269, 36.76%) with anti-PD1/PD-L1 mAbs and SOC, respectively. Anti-PD1/PD-L1 mAbs improved CR rate (RR 3.48) and PR rate (RR 2.27). There were no differences in the PD rate between groups (RR 1.10). Subgroup analyses showed an improvement in clinical benefit with anti-PD1/PD-L1 mAbs for melanoma (RR 1.59; 1.37-1.84 95% CI) and those treated in the first line setting (RR 1.57; 1.27-1.95 95% CI).Anti-PD1/PD-L1 mAbs increase overall response rate compared to SOC without an increase in PD rate. Melanoma and pts treated in first line setting seem to have greater benefit with anti-PD1/PD-L1 mAbs.In this systematic meta-analysis, anti-PD1/PD-L1 mAbs were associated with a greater overall response rate. Patients with melanoma and those managed in the first line setting seem to have an additional benefit with anti-PD1/PD-L1 mAbs.
DOI: 10.1136/esmoopen-2020-000828
2020
Cited 69 times
Impact of the COVID-19 outbreak on cancer patient flow and management: experience from a large university hospital in Spain
We report the dramatic shift of the oncology activity at our department during a 5-week period (9 March–13 April 2020) as compared with the same calendar interval in 2019 . Overall, our Medical Oncology Department has experienced remarkable drop in activity. The number of outpatient's visits decreased by 23%. One of the most worrisome concern is that the new oncology referrals were reduced by 37%, and the number of patients enrolled in clinical trials decreased by 43%. These data would mean that nearly 4 out 10 patients with cancer have been missed or their treatment delayed.
DOI: 10.3390/cancers12071945
2020
Cited 57 times
The Value of PD-L1 Expression as Predictive Biomarker in Metastatic Renal Cell Carcinoma Patients: A Meta-Analysis of Randomized Clinical Trials
Immune checkpoint inhibitors (ICIs) are soluble antibodies that have dramatically changed the outcomes including overall survival in a subset of kidney tumors, specifically in renal cell carcinoma (RCC). To date, there is no a single predictive biomarker approved to be used to select the patients that achieve benefit from ICIs targeting. It seems reasonable to analyze whether the programmed death-ligand 1 (PD-L1) expression could be useful. To assess the role of PD-L1 expression as a potential predictive biomarker for benefit of ICIs in RCC patients, we performed a search of randomized clinical trials (RCTs) comparing ICIs (monotherapy or in combination with other therapies) to standard of care in metastatic RCC patients according to PRISMA guidelines. Trials must have included subgroup analyses evaluating the selected outcomes (progression-free survival (PFS) and overall survival (OS)) in different subsets of patients according to PD-L1 expression on tumor samples. Hazard ratios with confidence intervals were used as the measure of efficacy between groups. A total of 4635 patients (six studies) were included (ICIs arm: 2367 patients; standard of care arm: 2268 patients). Globally, PFS and OS results favored ICIs. Differential expression of PD-L1 on tumor samples could select a subset of patients who could benefit more in terms of PFS (those with higher levels; p-value for difference between subgroups: &lt;0.0001) but it did not seem to impact in OS results (p-value for difference: 0.63). As different methods to assess PD-L1 positivity were used among trials, this heterogeneity could have an influence on the results. PD-L1 could represent a biomarker to test PFS in clinical trials but its value for OS is less clear. In this meta-analysis, the usefulness of PD-L1 expression as a predictive biomarker to select treatment in metastatic RCC patients was not clearly shown.
DOI: 10.1001/jamaoncol.2021.4544
2021
Cited 45 times
Clinical Activity and Safety of Cabozantinib for Brain Metastases in Patients With Renal Cell Carcinoma
Patients with brain metastases from renal cell carcinoma (RCC) have been underrepresented in clinical trials, and effective systemic therapy is lacking. Cabozantinib shows robust clinical activity in metastatic RCC, but its effect on brain metastases remains unclear.To assess the clinical activity and toxic effects of cabozantinib to treat brain metastases in patients with metastatic RCC.This retrospective cohort study included patients with metastatic RCC and brain metastases treated in 15 international institutions (US, Belgium, France, and Spain) between January 2014 and October 2020. Cohort A comprised patients with progressing brain metastases without concomitant brain-directed local therapy, and cohort B comprised patients with stable or progressing brain metastases concomitantly treated by brain-directed local therapy.Receipt of cabozantinib monotherapy at any line of treatment.Intracranial radiological response rate by modified Response Evaluation Criteria in Solid Tumors, version 1.1, and toxic effects of cabozantinib.Of the 88 patients with brain metastases from RCC included in the study, 33 (38%) were in cohort A and 55 (62%) were in cohort B; the majority of patients were men (n = 69; 78%), and the median age at cabozantinib initiation was 61 years (range, 34-81 years). Median follow-up was 17 months (range, 2-74 months). The intracranial response rate was 55% (95% CI, 36%-73%) and 47% (95% CI, 33%-61%) in cohorts A and B, respectively. In cohort A, the extracranial response rate was 48% (95% CI, 31%-66%), median time to treatment failure was 8.9 months (95% CI, 5.9-12.3 months), and median overall survival was 15 months (95% CI, 9.0-30.0 months). In cohort B, the extracranial response rate was 38% (95% CI, 25%-52%), time to treatment failure was 9.7 months (95% CI, 6.0-13.2 months), and median overall survival was 16 months (95% CI, 12.0-21.9 months). Cabozantinib was well tolerated, with no unexpected toxic effects or neurological adverse events reported. No treatment-related deaths were observed.In this cohort study, cabozantinib showed considerable intracranial activity and an acceptable safety profile in patients with RCC and brain metastases. Support of prospective studies evaluating the efficacy of cabozantinib for brain metastases in patients with RCC is critical.
DOI: 10.1136/jitc-2022-005977
2023
Cited 12 times
New prognostic model in patients with advanced urothelial carcinoma treated with second-line immune checkpoint inhibitors
Bellmunt Risk Score, based on Eastern Cooperative Oncology Group (ECOG) performance status (PS), hemoglobin levels and presence of liver metastases, is the most established prognostic algorithm for patients with advanced urothelial cancer (aUC) progressing after platinum-based chemotherapy. Nevertheless, existing algorithms may not be sufficient following the introduction of immunotherapy. Our aim was to develop an improved prognostic model in patients receiving second-line atezolizumab for aUC.Patients with aUC progressing after cisplatin/carboplatin-based chemotherapy and enrolled in the prospective, single-arm, phase IIIb SAUL study were included in this analysis. Patients were treated with 3-weekly atezolizumab 1200 mg intravenously. The development and internal validation of a prognostic model for overall survival (OS) was performed using Cox regression analyses, bootstrapping methods and calibration.In 936 patients, ECOG PS, alkaline phosphatase, hemoglobin, neutrophil-to-lymphocyte ratio, liver metastases, bone metastases and time from last chemotherapy were identified as independent prognostic factors. In a 4-tier model, median OS for patients with 0-1, 2, 3-4 and 5-7 risk factors was 18.6, 10.4, 4.8 and 2.1 months, respectively. Compared with Bellmunt Risk Score, this model provided enhanced prognostic separation, with a c-index of 0.725 vs 0.685 and increment in c-statistic of 0.04 (p<0.001). Inclusion of PD-L1 expression did not improve the model.We developed and internally validated a prognostic model for patients with aUC receiving postplatinum immunotherapy. This model represents an improvement over the Bellmunt algorithm and could aid selection of patients with aUC for second-line immunotherapy.NCT02928406.
DOI: 10.1093/jnci/djad155
2023
Cited 11 times
Safety and efficacy of immune checkpoint inhibitors in advanced penile cancer: report from the Global Society of Rare Genitourinary Tumors
Abstract Background Treatment options for penile squamous cell carcinoma are limited. We sought to investigate clinical outcomes and safety profiles of patients with penile squamous cell carcinoma receiving immune checkpoint inhibitors. Methods This retrospective study included patients with locally advanced or metastatic penile squamous cell carcinoma receiving immune checkpoint inhibitors between 2015 and 2022 across 24 centers in the United States, Europe, and Asia. Overall survival and progression-free survival were estimated using the Kaplan-Meier method. Objective response rates were determined per Response Evaluation Criteria in Solid Tumours 1.1 criteria. Treatment-related adverse events were graded per the Common Terminology Criteria for Adverse Events, version 5.0. Two-sided statistical tests were used for comparisons. Results Among 92 patients, 8 (8.7%) were Asian, 6 (6.5%) were Black, and 24 (29%) were Hispanic and/or Latinx. Median (interquartile range) age was 62 (53-70) years. In all, 83 (90%) had metastatic penile squamous cell carcinoma, and 74 (80%) had received at least second-line treatment. Most patients received pembrolizumab monotherapy (n = 26 [28%]), combination nivolumab-ipilimumab with or without multitargeted tyrosine kinase inhibitors (n = 23 [25%]), or nivolumab (n = 16 [17%]) or cemiplimab (n = 15 [16%]) monotherapies. Median overall and progression-free survival were 9.8 months (95% confidence interval = 7.7 to 12.8 months) and 3.2 months (95% confidence interval = 2.5 to 4.2 months), respectively. The objective response rate was 13% (n = 11/85) in the overall cohort and 35% (n = 7/20) in patients with lymph node–only metastases. Visceral metastases, Eastern Cooperative Oncology Group (ECOG) performance status of 1 or higher, and a higher neutrophil/lymphocyte ratio were associated with worse overall survival. Treatment-related adverse events occurred in 27 (29%) patients, and 9.8% (n = 9) of the events were grade 3 or higher. Conclusions Immune checkpoint inhibitors are active in a subset of patients with penile squamous cell carcinoma. Future translational studies are warranted to identify patients more likely to derive clinical benefit from immune checkpoint inhibitors.
DOI: 10.1016/s0014-5793(01)02089-0
2001
Cited 163 times
The AMP‐activated protein kinase prevents ceramide synthesis de novo and apoptosis in astrocytes
Fatty acids induce apoptosis in primary astrocytes by enhancing ceramide synthesis de novo. The possible role of the AMP-activated protein kinase (AMPK) in the control of apoptosis was studied in this model. Long-term stimulation of AMPK with 5-aminoimidazole-4-carboxamide ribonucleoside (AICAR) prevented apoptosis. AICAR blunted fatty acid-mediated induction of serine palmitoyltransferase and ceramide synthesis de novo, without affecting fatty acid synthesis and oxidation. Prevention of ceramide accumulation by AICAR led to a concomitant blockade of the Raf-1/extracellular signal-regulated kinase cascade, which selectively mediates fatty acid-induced apoptosis. Data indicate that AMPK may protect cells from apoptosis induced by stress stimuli.
DOI: 10.1042/0264-6021:3630183
2002
Cited 145 times
De novo-synthesized ceramide is involved in cannabinoid-induced apoptosis
Delta(9)-Tetrahydrocannabinol (THC) and other cannabinoids have been shown to induce apoptosis of glioma cells via ceramide generation. In the present study, we investigated the metabolic origin of the ceramide responsible for this cannabinoid-induced apoptosis by using two subclones of C6 glioma cells: C6.9, which is sensitive to THC-induced apoptosis; and C6.4, which is resistant to THC-induced apoptosis. Pharmacological inhibition of ceramide synthesis de novo, but not of neutral and acid sphingomyelinases, prevented THC-induced apoptosis in C6.9 cells. The activity of serine palmitoyltransferase (SPT), which catalyses the rate-limiting step of ceramide synthesis de novo, was remarkably enhanced by THC in C6.9 cells, but not in C6.4 cells. However, no major changes in SPT mRNA and protein levels were evident. Changes in SPT activity paralleled changes in ceramide levels. Pharmacological inhibition of ceramide synthesis de novo also prevented the stimulation of extracellular-signal-regulated kinase and the inhibition of protein kinase B triggered by cannabinoids. These findings show that de novo-synthesized ceramide is involved in cannabinoid-induced apoptosis of glioma cells.
DOI: 10.1042/0264-6021:3470369
2000
Cited 144 times
The CB1 cannabinoid receptor is coupled to the activation of protein kinase B/Akt
Cannabinoids exert most of their effects in the central nervous system through the CB(1) cannabinoid receptor. This G-protein-coupled receptor has been shown to be functionally coupled to inhibition of adenylate cyclase, modulation of ion channels and activation of extracellular-signal-regulated kinase. Using Chinese hamster ovary cells stably transfected with the CB(1) receptor cDNA we show here that Delta(9)-tetrahydrocannabinol (THC), the major active component of marijuana, induces the activation of protein kinase B/Akt (PKB). This effect of THC was also exerted by the endogenous cannabinoid anandamide and the synthetic cannabinoids CP-55940 and HU-210, and was prevented by the selective CB(1) antagonist SR141716. Pertussis toxin and wortmannin blocked the CB(1) receptor-evoked activation of PKB, pointing to the sequential involvement of a G(i)/G(o) protein and phosphoinositide 3'-kinase. The functionality of the cannabinoid-induced stimulation of PKB was proved by the increased phosphorylation of glycogen synthase kinase-3 serine 21 observed in cannabinoid-treated cells and its prevention by SR141716 and wortmannin. Cannabinoids activated PKB in the human astrocytoma cell line U373 MG, which expresses the CB(1) receptor, but not in the human promyelocytic cell line HL-60, which expresses the CB(2) receptor. Data indicate that activation of PKB may be responsible for some of the effects of cannabinoids in cells expressing the CB(1) receptor.
DOI: 10.1042/bj3470369
2000
Cited 123 times
The CB1 cannabinoid receptor is coupled to the activation of protein kinase B/Akt
Cannabinoids exert most of their effects in the central nervous system through the CB1 cannabinoid receptor. This G-protein-coupled receptor has been shown to be functionally coupled to inhibition of adenylate cyclase, modulation of ion channels and activation of extracellular-signal-regulated kinase. Using Chinese hamster ovary cells stably transfected with the CB1 receptor cDNA we show here that ∆9-tetrahydrocannabinol (THC), the major active component of marijuana, induces the activation of protein kinase B/Akt (PKB). This effect of THC was also exerted by the endogenous cannabinoid anandamide and the synthetic cannabinoids CP-55940 and HU-210, and was prevented by the selective CB1 antagonist SR141716. Pertussis toxin and wortmannin blocked the CB1 receptor-evoked activation of PKB, pointing to the sequential involvement of a Gi/Go protein and phosphoinositide 3ʹ-kinase. The functionality of the cannabinoid-induced stimulation of PKB was proved by the increased phosphorylation of glycogen synthase kinase-3 serine 21 observed in cannabinoid-treated cells and its prevention by SR141716 and wortmannin. Cannabinoids activated PKB in the human astrocytoma cell line U373 MG, which expresses the CB1 receptor, but not in the human promyelocytic cell line HL-60, which expresses the CB2 receptor. Data indicate that activation of PKB may be responsible for some of the effects of cannabinoids in cells expressing the CB1 receptor.
DOI: 10.1091/mbc.e08-07-0671
2009
Cited 113 times
The TP53INP2 Protein Is Required for Autophagy in Mammalian Cells
Using a bioinformatic approach, we identified a TP53INP1-related gene encoding a protein with 30% identity with tumor protein 53-induced nuclear protein 1 (TP53INP1), which was named TP53INP2. TP53INP1 and TP53INP2 sequences were found in several species ranging from Homo sapiens to Drosophila melanogaster, but orthologues were found neither in earlier eukaryotes nor in prokaryotes. To gain insight into the function of the TP53INP2 protein, we carried out a yeast two-hybrid screening that showed that TP53INP2 binds to the LC3-related proteins GABARAP and GABARAP-like2, and then we demonstrated by coimmunoprecipitation that TP53INP2 interacts with these proteins, as well as with LC3 and with the autophagosome transmembrane protein VMP1. TP53INP2 translocates from the nucleus to the autophagosome structures after activation of autophagy by rapamycin or starvation. Also, we showed that TP53INP2 expression is necessary for autophagosome development because its small interfering RNA-mediated knockdown strongly decreases sensitivity of mammalian cells to autophagy. Finally, we found that interactions between TP53INP2 and LC3 or the LC3-related proteins GABARAP and GABARAP-like2 require autophagy and are modulated by wortmannin as judged by bioluminescence resonance energy transfer assays. We suggest that TP53INP2 is a scaffold protein that recruits LC3 and/or LC3-related proteins to the autophagosome membrane by interacting with the transmembrane protein VMP1. It is concluded that TP53INP2 is a novel gene involved in the autophagy of mammalian cells.
DOI: 10.1371/journal.pone.0054795
2013
Cited 85 times
Local Delivery of Cannabinoid-Loaded Microparticles Inhibits Tumor Growth in a Murine Xenograft Model of Glioblastoma Multiforme
Cannabinoids, the active components of marijuana and their derivatives, are currently investigated due to their potential therapeutic application for the management of many different diseases, including cancer. Specifically, Δ(9)-Tetrahydrocannabinol (THC) and Cannabidiol (CBD) - the two major ingredients of marijuana - have been shown to inhibit tumor growth in a number of animal models of cancer, including glioma. Although there are several pharmaceutical preparations that permit the oral administration of THC or its analogue nabilone or the oromucosal delivery of a THC- and CBD-enriched cannabis extract, the systemic administration of cannabinoids has several limitations in part derived from the high lipophilicity exhibited by these compounds. In this work we analyzed CBD- and THC-loaded poly-ε-caprolactone microparticles as an alternative delivery system for long-term cannabinoid administration in a murine xenograft model of glioma. In vitro characterization of THC- and CBD-loaded microparticles showed that this method of microencapsulation facilitates a sustained release of the two cannabinoids for several days. Local administration of THC-, CBD- or a mixture (1:1 w:w) of THC- and CBD-loaded microparticles every 5 days to mice bearing glioma xenografts reduced tumour growth with the same efficacy than a daily local administration of the equivalent amount of those cannabinoids in solution. Moreover, treatment with cannabinoid-loaded microparticles enhanced apoptosis and decreased cell proliferation and angiogenesis in these tumours. Our findings support that THC- and CBD-loaded microparticles could be used as an alternative method of cannabinoid delivery in anticancer therapies.
DOI: 10.1038/cdd.2010.170
2011
Cited 77 times
Stimulation of the midkine/ALK axis renders glioma cells resistant to cannabinoid antitumoral action
Identifying the molecular mechanisms responsible for the resistance of gliomas to anticancer treatments is an issue of great therapeutic interest. Δ(9)-Tetrahydrocannabinol (THC), the major active ingredient of marijuana, and other cannabinoids inhibit tumor growth in animal models of cancer, including glioma, an effect that relies, at least in part, on the stimulation of autophagy-mediated apoptosis in tumor cells. Here, by analyzing the gene expression profile of a large series of human glioma cells with different sensitivity to cannabinoid action, we have identified a subset of genes specifically associated to THC resistance. One of these genes, namely that encoding the growth factor midkine (Mdk), is directly involved in the resistance of glioma cells to cannabinoid treatment. We also show that Mdk mediates its protective effect via the anaplastic lymphoma kinase (ALK) receptor and that Mdk signaling through ALK interferes with cannabinoid-induced autophagic cell death. Furthermore, in vivo Mdk silencing or ALK pharmacological inhibition sensitizes cannabinod-resistant tumors to THC antitumoral action. Altogether, our findings identify Mdk as a pivotal factor involved in the resistance of glioma cells to THC pro-autophagic and antitumoral action, and suggest that selective targeting of the Mdk/ALK axis could help to improve the efficacy of antitumoral therapies for gliomas.
DOI: 10.1038/cdd.2014.133
2014
Cited 70 times
Loss of Tribbles pseudokinase-3 promotes Akt-driven tumorigenesis via FOXO inactivation
Tribbles pseudokinase-3 (TRIB3) has been proposed to act as an inhibitor of AKT although the precise molecular basis of this activity and whether the loss of TRIB3 contributes to cancer initiation and progression remain to be clarified. In this study, by using a wide array of in vitro and in vivo approaches, including a Trib3 knockout mouse, we demonstrate that TRIB3 has a tumor-suppressing role. We also find that the mechanism by which TRIB3 loss enhances tumorigenesis relies on the dysregulation of the phosphorylation of AKT by the mTORC2 complex, which leads to an enhanced phosphorylation of AKT on Ser473 and the subsequent hyperphosphorylation and inactivation of the transcription factor FOXO3. These observations support the notion that loss of TRIB3 is associated with a more aggressive phenotype in various types of tumors by enhancing the activity of the mTORC2/AKT/FOXO axis.
DOI: 10.1158/1078-0432.ccr-12-0026
2012
Cited 70 times
Nupr1-Aurora Kinase A Pathway Provides Protection against Metabolic Stress-Mediated Autophagic-Associated Cell Death
Abstract Purpose: The limited supply of oxygen and nutrients is thought to result in rigorous selection of cells that will eventually form the tumor. Experimental Design: Nupr1 expression pattern was analyzed in human tissue microarray (TMA) and correlated with survival time of the patient. Microarray analysis was conducted on MiaPaCa2 cells subjected to metabolic stress in Nupr1-silenced conditions. DNA repair and cell cycle–associated gene expression was confirmed by real-time quantitative PCR (qRT-PCR). Nupr1 and AURKA protective role were analyzed using RNA interference (RNAi) silencing or overexpression. DNA damage and autophagy were analyzed by Western blot analysis and immunofluorescence. Results: We showed that both Nupr1 and HIF1α are coexpressed in human pancreatic ductal adenocarcinoma (PDAC) samples and negatively correlate with survival time. PDAC-derived cells submitted to hypoxia and/or glucose starvation induce DNA damage–dependent cell death concomitantly to the overexpression of stress protein Nupr1. Affymetrix-based transcriptoma analysis reveals that Nupr1 knockdown enhances DNA damage and alters the expression of several genes involved in DNA repair and cell-cycle progression. Expression of some of these genes is common to hypoxia and glucose starvation, such as Aurka gene, suggesting that Nupr1 overexpression counteracts the transcriptional changes occurring under metabolic stress. The molecular mechanism by which hypoxia and glucose starvation induce cell death involves autophagy-associated, but not caspase-dependent, cell death. Finally, we have found that AURKA expression is partially regulated by Nupr1 and plays a major role in this response. Conclusions: Our data reveal that Nupr1 is involved in a defense mechanism that promotes pancreatic cancer cell survival when exposed to metabolic stress. Clin Cancer Res; 18(19); 5234–46. ©2012 AACR.
DOI: 10.18632/oncotarget.20028
2017
Cited 69 times
Differential side effects profile in patients with mCRPC treated with abiraterone or enzalutamide: a meta-analysis of randomized controlled trials
Abiraterone and enzalutamide are currently approved for mCRPC patients. Both drugs have distinct mechanisms of action and may have different toxicity profile. There are limited data comparing the side effects of abiraterone and enzalutamide. We performed a meta-analysis of randomized controlled trials (RCT) to better characterize the risk of adverse events associated with both drugs.We performed a literature search on MEDLINE for studies reporting abiraterone and enzalutamide side effects from January 1966 to July 31, 2015. Abstracts presented at ASCO meetings from 2004 to 2015 were selected manually. Phase III RCT were included in analysis. We assessed the risk of adverse events reported in RCT by performing two meta-analyses: abiraterone-prednisone vs. placebo-prednisone (2,283 pts) and enzalutamide vs. placebo (2,914 pts). Summary of incidence, relative-risks (RR), and 95% confidence intervals (CI) were calculated using random-effects or fixed-effects models based on the heterogeneity of included studies.Overall, enzalutamide was not associated with all-grade (RR 1.06 - 95% CI 0.67-1.65) or grade ≥3 (RR 0.81 - 95% CI 0.28-2.33) cardiovascular events, but was associated with increased risk of all-grade fatigue (RR 1.29 - 95% CI 1.15-1.44). On the other hand, abiraterone was associated with increased risk of all-grade (RR 1.28 - 95% CI 1.06-1.55) and grade ≥3 (RR 1.76 - 95% CI 1.12-2.75) cardiovascular events, but was not associated with all-grade (RR 0.85 - 95% CI 0.58-1.23) or grade ≥3 (RR 1.07 - 95% CI 0.97-1.19) fatigue.In this meta-analysis, abiraterone was associated with an increased risk of cardiovascular events, while enzalutamide was associated with an increased risk of fatigue.
DOI: 10.1080/15548627.2016.1240856
2016
Cited 66 times
Human Atg8-cardiolipin interactions in mitophagy: Specific properties of LC3B, GABARAPL2 and GABARAP
The phospholipid cardiolipin (CL) has been proposed to play a role in selective mitochondrial autophagy, or mitophagy. CL externalization to the outer mitochondrial membrane would act as a signal for the human Atg8 ortholog subfamily, MAP1LC3 (LC3). The latter would mediate both mitochondrial recognition and autophagosome formation, ultimately leading to removal of damaged mitochondria. We have applied quantitative biophysical techniques to the study of CL interaction with various Atg8 human orthologs, namely LC3B, GABARAPL2 and GABARAP. We have found that LC3B interacts preferentially with CL over other di-anionic lipids, that CL-LC3B binding occurs with positive cooperativity, and that the CL-LC3B interaction relies only partially on electrostatic forces. CL-induced increased membrane fluidity appears also as an important factor helping LC3B to bind CL. The LC3B C terminus remains exposed to the hydrophilic environment after protein binding to CL-enriched membranes. In intact U87MG human glioblastoma cells rotenone-induced autophagy leads to LC3B translocation to mitochondria and subsequent delivery of mitochondria to lysosomes. We have also observed that GABARAP, but not GABARAPL2, interacts with CL in vitro. However neither GABARAP nor GABARAPL2 were translocated to mitochondria in rotenone-treated U87MG cells. Thus the various human Atg8 orthologs might play specific roles in different autophagic processes.
DOI: 10.1158/2326-6066.cir-16-0110
2016
Cited 63 times
Tumor Mutational Load and Immune Parameters across Metastatic Renal Cell Carcinoma Risk Groups
Patients with metastatic renal cell carcinoma (mRCC) have better overall survival when treated with nivolumab, a cancer immunotherapy that targets the immune checkpoint inhibitor programmed cell death 1 (PD-1), rather than everolimus (a chemical inhibitor of mTOR and immunosuppressant). Poor-risk mRCC patients treated with nivolumab seemed to experience the greatest overall survival benefit, compared with patients with favorable or intermediate risk, in an analysis of the CheckMate-025 trial subgroup of the Memorial Sloan Kettering Cancer Center (MSKCC) prognostic risk groups. Here, we explore whether tumor mutational load and RNA expression of specific immune parameters could be segregated by prognostic MSKCC risk strata and explain the survival seen in the poor-risk group. We queried whole-exome transcriptome data in renal cell carcinoma patients (n = 54) included in The Cancer Genome Atlas who ultimately developed metastatic disease or were diagnosed with metastatic disease at presentation and did not receive immune checkpoint inhibitors. Nonsynonymous mutational load did not differ significantly by the MSKCC risk group, nor was the expression of cytolytic genes-granzyme A and perforin-or selected immune checkpoint molecules different across MSKCC risk groups. In conclusion, this analysis revealed that mutational load and expression of markers of an active tumor microenvironment did not correlate with MSKCC risk prognostic classification in mRCC. Cancer Immunol Res; 4(10); 820-2. ©2016 AACR.
DOI: 10.1158/1078-0432.ccr-15-1808
2016
Cited 59 times
The New Antitumor Drug ABTL0812 Inhibits the Akt/mTORC1 Axis by Upregulating Tribbles-3 Pseudokinase
Abstract Purpose: ABTL0812 is a novel first-in-class, small molecule which showed antiproliferative effect on tumor cells in phenotypic assays. Here we describe the mechanism of action of this antitumor drug, which is currently in clinical development. Experimental Design: We investigated the effect of ABTL0812 on cancer cell death, proliferation, and modulation of intracellular signaling pathways, using human lung (A549) and pancreatic (MiaPaCa-2) cancer cells and tumor xenografts. To identify cellular targets, we performed in silico high-throughput screening comparing ABTL0812 chemical structure against ChEMBL15 database. Results: ABTL0812 inhibited Akt/mTORC1 axis, resulting in impaired cancer cell proliferation and autophagy-mediated cell death. In silico screening led us to identify PPARs, PPARα and PPARγ as the cellular targets of ABTL0812. We showed that ABTL0812 activates both PPAR receptors, resulting in upregulation of Tribbles-3 pseudokinase (TRIB3) gene expression. Upregulated TRIB3 binds cellular Akt, preventing its activation by upstream kinases, resulting in Akt inhibition and suppression of the Akt/mTORC1 axis. Pharmacologic inhibition of PPARα/γ or TRIB3 silencing prevented ABTL0812-induced cell death. ABTL0812 treatment induced Akt inhibition in cancer cells, tumor xenografts, and peripheral blood mononuclear cells from patients enrolled in phase I/Ib first-in-human clinical trial. Conclusions: ABTL0812 has a unique and novel mechanism of action, that defines a new and drugable cellular route that links PPARs to Akt/mTORC1 axis, where TRIB3 pseudokinase plays a central role. Activation of this route (PPARα/γ-TRIB3-Akt-mTORC1) leads to autophagy-mediated cancer cell death. Given the low toxicity and high tolerability of ABTL0812, our results support further development of ABTL0812 as a promising anticancer therapy. Clin Cancer Res; 22(10); 2508–19. ©2015 AACR.
DOI: 10.1158/2326-6066.cir-17-0220
2018
Cited 56 times
Durable Clinical Benefit in Metastatic Renal Cell Carcinoma Patients Who Discontinue PD-1/PD-L1 Therapy for Immune-Related Adverse Events
Abstract The current standard of care for treatment of metastatic renal cell carcinoma (mRCC) patients is PD-1/PD-L1 inhibitors until progression or toxicity. Here, we characterize the clinical outcomes for 19 mRCC patients who experienced an initial clinical response (any degree of tumor shrinkage), but after immune-related adverse events (irAE) discontinued all systemic therapy. Clinical baseline characteristics, outcomes, and survival data were collected. The primary endpoint was time to progression from the date of treatment cessation (TTP). Most patients had clear cell histology and received anti–PD–1/PD-L1 therapy as second-line or later treatment. Median time on PD-1/PD-L1 therapy was 5.5 months (range, 0.7–46.5) and median TTP was 18.4 months (95% CI, 4.7–54.3) per Kaplan–Meier estimation. The irAEs included arthropathies, ophthalmopathies, myositis, pneumonitis, and diarrhea. We demonstrate that 68.4% of patients (n = 13) experienced durable clinical benefit off treatment (TTP of at least 6 months), with 36% (n = 7) of patients remaining off subsequent treatment for over a year after their last dose of anti–PD-1/PD-L1. Three patients with tumor growth found in a follow-up visit, underwent subsequent surgical intervention, and remain off systemic treatment. Nine patients (47.4%) have ongoing irAEs. Our results show that patients who benefitted clinically from anti–PD-1/PD-L1 therapy can experience sustained beneficial responses, not needing further therapies after the initial discontinuation of treatment due to irAEs. Investigation of biomarkers indicating sustained benefit to checkpoint blockers are needed. Cancer Immunol Res; 6(4); 402–8. ©2018 AACR.
DOI: 10.1016/j.euo.2019.06.022
2019
Cited 50 times
Sequencing and Combination of Systemic Therapy in Metastatic Renal Cell Carcinoma
Introduction of additional new agents targeting the vascular endothelial growth factor receptor (VEGFR) and immune checkpoint inhibitors (ICIs) has completely modified the systemic treatment of metastatic renal cell carcinoma (mRCC) during the last years.A comprehensive (nonsystematic) review to determine the suggested sequence or combinations for the systemic treatment of mRCC.PubMed and abstracts from main conferences up to December 2018 were reviewed to retrieve the current evidence for treatment of mRCC. Search terms included renal cell carcinoma, systemic therapy, targeted therapy (TT), and immunotherapy.Marked advances in the treatment of mRCC have been made with novel VEGFR tyrosine kinase inhibitors and multiple ICIs that have been included in the current treatment paradigm of mRCC. Remarkable advance has been made with the combination of double checkpoint blockade. The combination of ipilimumab and nivolumab compared with sunitinib has shown to increase the overall survival in the intermediate- and poor-risk patients based on the International Metastatic Renal Cell Carcinoma Database Consortium (IMDC) model.Double checkpoint blockade with ipilimumab and nivolumab has reported overall survival benefit in IMDC intermediate- and poor-risk patients, providing a durable response for a subset of patients. VEGF inhibitors remain the standard of care for favorable-risk patients in the first line. In the immediate future, more consolidated data on combination of VEGF-TT plus ICIs may show similar robust benefit with different safety profiles.Multiple drugs and sequences are now accepted as effective treatment for metastatic renal cell carcinoma (mRCC). Combination of immune checkpoint inhibitors has shown to increase the overall survival in treatment-naïve mRCC patients. Combinations of immunotherapy and antiangiogenics may be another option in the near future. Outcomes of the first line will determine the sequence, although the best sequence has yet to be defined.
DOI: 10.1016/j.euf.2017.12.007
2019
Cited 45 times
The Development of Brain Metastases in Patients with Renal Cell Carcinoma: Epidemiologic Trends, Survival, and Clinical Risk Factors Using a Population-based Cohort
Background The incidence of brain metastases (BM) in patients with renal cell carcinoma (RCC) is hypothesized to have increased in the last 2 decades. Objective To define incidence trends according to patient and clinical characteristics, to identify risk factors, and to describe outcomes of patients with BM for RCC. Design, setting, and participants Patients diagnosed with RCC between the years 2010 and 2013 within the Surveillance, Epidemiology, and End Results database. An external validation was also considered using patients diagnosed with RCC between 2010 and 2012 within the National Cancer Database. Outcome measurements and statistical analysis Incidence proportions of BM were calculated. Risk factors correlated with BM at diagnosis were identified via a 1000-bootstrap corrected multivariable logistic regression model. A risk model was then developed and evaluated using measures of predictive accuracy. Overall survival was examined using Cox regression analyses. Results and limitations The overall incidence proportions of BM at RCC diagnosis was 1.51% (95% confidence interval: 1.39–1.64%). White/other race, clear cell histology, and sarcomatoid differentiation, T2–4 disease, tumor dimension >10 cm, and N+ disease were significantly associated with BM at RCC diagnosis, and retained within the final prediction model. A risk score was created based on these variables (c-index: 0.803). BM at RCC diagnosis occurred in 0.5%, 3.6%, and 7.7% of patients categorized as low risk, intermediate risk, and high risk. Patients with BM were more likely to succumb to any death than those without BM at diagnosis (median overall survival: 6.4 mo vs not reached, respectively, adjusted hazard ratio: 1.87, 95% confidence interval: 1.67–2.08, p < 0.001). The real incidence of BM at RCC diagnosis is likely underestimated given that the observed rate likely reflects patients who presented with symptoms. Conclusions Patients with BM at RCC have poor oncological outcomes. We have characterized the epidemiology of BM at RCC diagnosis and developed a clinical risk model for the purpose of predicting the development of BMs in patients diagnosed with a cortical renal mass. Patient summary In this report we examined recent proportions of patients with brain metastases at kidney cancer diagnosis in a large community database originating from the US. We developed a model that may be used during routine clinical practice to predict brain metastases. The urologic-oncological community may consider baseline imaging for brain metastases in patients without any symptoms but at high risk of having brain metastases according to the risk model. However, the proposed model certainly needs further testing and validation in the clinical setting. Future studies on brain metastases survival and treatment options are also needed.
DOI: 10.1158/1078-0432.ccr-18-0685
2019
Cited 45 times
CDK4/6 Inhibitor as a Novel Therapeutic Approach for Advanced Bladder Cancer Independently of <i>RB1</i> Status
Abstract Purpose: Bladder cancer is a clinical and social problem due to its high incidence and recurrence rates. It frequently appears in elderly patients showing other medical comorbidities that hamper the use of standard chemotherapy. We evaluated the activity of CDK4/6 inhibitor as a new therapy for patients unfit for cisplatin (CDDP). Experimental Design: Bladder cancer cell lines were tested for in vitro sensitivity to CDK4/6 inhibition. A novel metastatic bladder cancer mouse model was developed and used to test its in vivo activity. Results: Cell lines tested were sensitive to CDK4/6 inhibition, independent on RB1 gene status. Transcriptome analyses and knockdown experiments revealed a major role for FOXM1 in this response. CDK4/6 inhibition resulted in reduced FOXM1 phosphorylation in vitro and in vivo and showed synergy with CDDP, allowing a significant tumor regression. FOXM1 exerted important oncogenic roles in bladder cancer. Conclusions: CDK4/6 inhibitors, alone or in combination, are a novel therapeutic strategy for patients with advanced bladder cancer previously classified as unfit for current treatment options.
DOI: 10.1016/j.ctrv.2020.102142
2021
Cited 29 times
Recent advances in neoadjuvant immunotherapy for urothelial bladder cancer: What to expect in the near future
Urothelial bladder cancer (UC) is the most common malignancy involving the urinary system and represents a significant health problem. Immunotherapy has been used for decades for UC with intravesical bacillus Calmette-Guérin (BCG) set as the standard of care for non-muscle-invasive bladder cancer (NMIBC). The advent of immune checkpoint inhibitors (ICIs) has completely transformed the treatment landscape of bladder cancer enabling to expand the treatment strategies. Novel ICIs have successfully shown improved outcomes on metastatic disease to such an extent that the standard of care paradigm has changed leading to the development of different trials with the aim of determining whether ICIs may have a role in early disease. The localized muscle-invasive bladder cancer (MIBC) scenario remains challenging since the recurrence rate continues to be high despite all therapeutic efforts. This article will review the current experience of ICIs in the neoadjuvant setting of UC, the clinical trials landscape and finally, an insight of what to expect in the immediate and mid-term future.
DOI: 10.1006/abbi.1996.9784
1997
Cited 110 times
Control of Hepatic Fatty Acid Oxidation by 5′-AMP-Activated Protein Kinase Involves a Malonyl-CoA-Dependent and a Malonyl-CoA-Independent Mechanism
Incubation of rat hepatocytes with 5-aminoimidazole-4-carboxamide ribonucleoside (AICAR), an activator of the 5'-AMP-activated protein kinase (AMPK), produced a twofold stimulation of palmitate oxidation and of the activity of carnitine palmitoyltransferase I (CPT-I), together with a profound decrease of the activity of acetyl-CoA carboxylase and of the intracellular level of malonyl-CoA. AICAR-induced CPT-I stimulation progressively blunted with time after cell permeabilization, pointing to reversal of conformational constraints of the enzyme in control cells due to the permeabilization-triggered dilution of intracellular malonyl-CoA. The stimulation stabilized at a steady 20-25%. This 20-25% increase in CPT-I activity survived upon complete removal of malonyl-CoA from the permeabilized cells, indicating that it was not dependent on the malonyl-CoA concentration of the cell. This malonyl-CoA-independent activation of CPT-I was not evident when mitochondria were isolated for assay of enzyme activity or when cells were disrupted by vigorous sonication. In addition, the microtubule stabilizer taxol prevented the malonyl-CoA-independent stimulation of CPT-I induced by AICAR. Hence, stimulation of hepatic fatty acid oxidation by AMPK seems to rely on the activation of CPT-I by two different mechanisms: deinhibition of CPT-I induced by depletion of intracellular malonyl-CoA levels and malonyl-CoA-independent stimulation of CPT-I, which might involve modulation of interactions between CPT-I and cytoskeletal components.
DOI: 10.1016/j.yexcr.2006.03.009
2006
Cited 87 times
The CB2 cannabinoid receptor signals apoptosis via ceramide-dependent activation of the mitochondrial intrinsic pathway
Δ9-Tetrahydrocannabinol and other cannabinoids exert pro-apoptotic actions in tumor cells via the CB2 cannabinoid receptor. However, the molecular mechanism involved in this effect has remained elusive. Here we used the human leukemia cell line Jurkat—that expresses CB2 as the unique CB receptor—to investigate this mechanism. Our results show that incubation with the selective CB2 antagonist SR144528 abrogated the pro-apoptotic effect of Δ9-tetrahydrocannabinol. Cannabinoid treatment led to a CB2 receptor-dependent stimulation of ceramide biosynthesis and inhibition of this pathway prevented Δ9-tetrahydrocannabinol-induced mitochondrial hypopolarization and cytochrome c release, indicating that ceramide acts at a pre-mitochondrial level. Inhibition of ceramide synthesis de novo also prevented caspase activation and apoptosis. Caspase 8 activation—an event typically related with the extrinsic apoptotic pathway—was also evident in this model. However, activation of this protease was post-mitochondrial since (i) a pan-caspase inhibitor as well as a selective caspase 8 inhibitor were unable to prevent Δ9-tetrahydrocannabinol-induced loss of mitochondrial-membrane transmembrane potential, and (ii) cannabinoid-induced caspase 8 activation was not observed in Bcl-xL over-expressing cells. In summary, results presented here show that CB2 receptor activation signals apoptosis via a ceramide-dependent stimulation of the mitochondrial intrinsic pathway.
DOI: 10.1007/s12035-007-0002-5
2007
Cited 86 times
Cannabinoids and Gliomas
DOI: 10.2174/138161206777585139
2006
Cited 86 times
Endocannabinoids: A New Family of Lipid Mediators Involved in the Regulation of Neural Cell Development
The endocannabinoids (eCBs) anandamide and 2-arachidonoylglycerol are important retrograde messengers that inhibit neurotransmitter release via presynaptic CB1 receptors. In addition, cannabinoids are known to modulate the cell death/survival decision of different neural cell types, leading to different outcomes that depend on the nature of the target cell and its proliferative/differentiation status. Thus, cannabinoids protect primary neurons, astrocytes and oligodendrocytes from apoptosis, whereas transformed glial cells are prone to apoptosis by cannabinoid challenge. Moreover, a potential role of the eCB system in neurogenesis and neural differentiation has been proposed. Recent research shows that eCBs stimulate neural progenitor proliferation and inhibit hippocampal neurogenesis in normal adult brain. Cannabinoids inhibit cortical neuron differentiation and promote glial differentiation. On the other hand, experiments with differentiated neurons have shown that cannabinoids also regulate neuritogenesis, axonal growth and synaptogenesis. These new observations support that eCBs constitute a new family of lipid signaling cues responsible for the regulation of neural progenitor proliferation and differentiation, acting as instructive proliferative signals through the CB1 receptor. Keywords: neurogenesis, glioprotective action, synaptogenesis, Cannabinoid receptors, ERK pathway, neural progenitor differentiation
DOI: 10.4161/auto.5.7.9508
2009
Cited 69 times
TRB3 links ER stress to autophagy in cannabinoid antitumoral action
AbstractΔ9-tetrahydrocannabinol (THC), the main active component of marijuana, is being investigated as a potential anti-tumoral agent. We find that THC stimulates an endoplasmic reticulum (ER) stress-related signaling pathway, which activates autophagy via inhibition of the Akt/mTORC1 axis. We also show that autophagy is upstream of apoptosis in cannabinoid-induced cancer cell death and that activation of this pathway is necessary for the anti-tumoral action of cannabinoids in vivo.This article refers to:
DOI: 10.1016/j.fertnstert.2009.01.153
2010
Cited 65 times
The CB2 cannabinoid receptor regulates human sperm cell motility
To analyze the expression and distribution of cannabinoid receptors in human sperm cells and evaluate the effects of activation of receptors by specific agonists and antagonists, with a special emphasis on the CB(2) receptor.We performed expression assays for CB(1) and CB(2) by reverse transcriptase PCR, Western blot, and immunofluorescence techniques in spermatozoa and performed motility analysis after incubation of semen samples with cannabinoid agonists and CB(2) antagonist SR144528.Academic research laboratory.Semen from 50 normozoospermic, healthy human donors.Spermatozoa isolated from semen by two consecutive swim-ups were used for all techniques.Reverse transcriptase PCR amplification gels, immunoblots, indirect immunofluorescence antibody assays, and percentage of motile sperm.We have verified the presence of CB(1) and CB(2) receptors in human spermatozoa. The distribution of both of these receptors was distinct. Incubation with selective cannabinoid receptor agonists induced a significant reduction in the proportion of rapidly progressive motile spermatozoa, and whereas the CB(1) agonist increased the proportion of immobile sperm cells, the CB(2) receptor agonist increased the slow/sluggish progressive sperm cell population. The effect of the CB(2) agonist was antagonized by the CB(2)-specific antagonist.The functional CB(2) cannabinoid receptor is present in human spermatozoa and regulates the sperm motility in a more distinct manner than CB(1).
DOI: 10.1007/978-3-319-20825-1_16
2015
Cited 48 times
Endocannabinoids and Cancer
A large body of evidence shows that cannabinoids, in addition to their well-known palliative effects on some cancer-associated symptoms, can reduce tumour growth in animal models of cancer. They do so by modulating key cell signalling pathways involved in the control of cancer cell proliferation and survival. In addition, cannabinoids inhibit angiogenesis and cell proliferation in different types of tumours in laboratory animals. By contrast, little is known about the biological role of the endocannabinoid system in cancer physio-pathology, and several studies suggest that it may be over-activated in cancer. In this review, we discuss our current understanding of cannabinoids as antitumour agents, focusing on recent advances in the molecular mechanisms of action, including resistance mechanisms and opportunities for combination therapy approaches.
DOI: 10.1016/j.bbalip.2013.03.014
2013
Cited 47 times
The pseudokinase tribbles homologue-3 plays a crucial role in cannabinoid anticancer action
Δ9-Tetrahydrocannabinol (THC), the major active ingredient of marijuana, and other cannabinoids inhibit tumor growth in animal models of cancer. This effect relies, at least in part, on the up-regulation of several endoplasmic reticulum stress-related proteins including the pseudokinase tribbles homologue-3 (TRIB3), which leads in turn to the inhibition of the AKT/mTORC1 axis and the subsequent stimulation of autophagy-mediated apoptosis in tumor cells. Here, we took advantage of the use of cells derived from Trib3-deficient mice to investigate the precise mechanisms by which TRIB3 regulates the anti-cancer action of THC. Our data show that RasV12/E1A-transformed embryonic fibroblasts derived from Trib3-deficient mice are resistant to THC-induced cell death. We also show that genetic inactivation of this protein abolishes the ability of THC to inhibit the phosphorylation of AKT and several of its downstream targets, including those involved in the regulation of the AKT/mammalian target of rapamycin complex 1 (mTORC1) axis. Our data support the idea that THC-induced TRIB3 up-regulation inhibits AKT phosphorylation by regulating the accessibility of AKT to its upstream activatory kinase (the mammalian target of rapamycin complex 2; mTORC2). Finally, we found that tumors generated by inoculation of Trib3-deficient cells in nude mice are resistant to THC anticancer action. Altogether, the observations presented here strongly support that TRIB3 plays a crucial role on THC anti-neoplastic activity. This article is part of a Special Issue entitled Lipid Metabolism in Cancer.
DOI: 10.1016/j.bcp.2018.08.023
2018
Cited 45 times
Optimization of a preclinical therapy of cannabinoids in combination with temozolomide against glioma
Glioblastoma multiforme (GBM) is the most frequent and aggressive form of brain cancer. These features are explained at least in part by the high resistance exhibited by these tumors to current anticancer therapies. Thus, the development of novel therapeutic approaches is urgently needed to improve the survival of the patients suffering this devastating disease. Δ9-Tetrahydrocannabinol (THC, the major active ingredient of marijuana), and other cannabinoids have been shown to exert antitumoral actions in animal models of cancer, including glioma. The mechanism of these anticancer actions relies, at least in part, on the ability of these compounds to stimulate autophagy-mediated apoptosis in tumor cells. Previous observations from our group demonstrated that local administration of THC (or of THC + CBD at a 1:1 ratio, a mixture that resembles the composition of the cannabinoid-based medicine Sativex®) in combination with Temozolomide, the benchmark agent for the treatment of GBM, synergistically reduces the growth of glioma xenografts. With the aim of optimizing the possible clinical utilization of cannabinoids in anti-GBM therapies, in this work we explored the anticancer efficacy of the systemic administration of cannabinoids in combination with TMZ in preclinical models of glioma. Our results show that oral administration of Sativex-like extracts (containing THC and CBD at a 1:1 ratio) in combination with TMZ produces a strong antitumoral effect in both subcutaneous and intracranial glioma cell-derived tumor xenografts. In contrast, combined administration of Sativex-like and BCNU (another alkylating agent used for the treatment of GBM which share structural similarities with the TMZ) did not show a stronger effect than individual treatments. Altogether, our findings support the notion that the combined administration of TMZ and oral cannabinoids could be therapeutically exploited for the management of GBM.
DOI: 10.1200/jco.2020.38.6_suppl.542
2020
Cited 32 times
Impact of the combination of durvalumab (MEDI4736) plus olaparib (AZD2281) administered prior to surgery in the molecular profile of resectable urothelial bladder cancer: NEODURVARIB Trial.
542 Background: Cisplatin-based chemotherapy remains the perioperative treatment in muscle-invasive bladder carcinoma (MIBC). Recent evidence suggests that immune checkpoint inhibitors could be incorporated in this setting. Olaparib is a PARP inhibitor with well-established activity in HRD tumor. Results from trials assessing the combination of durvalumab and olaparib suggest a synergistic effect. However, a molecular characterization is crucial to warrant a rational development. Methods: A phase II clinical trial was designed to assess the impact of neoadjuvant treatment with the combination of durvalumab plus olaparib in the molecular profile of MIBC (NCT03534492; SOGUG-2017-A-IEC(VEJ)-2). Efficacy and safety were secondary objectives. Subjects with cT2-T4a MIBC aimed for cystectomy were treated during 6 to 8 weeks pre-cystectomy. Diagnostic and surgical samples, pre and postreatment blood samples have been collected for the molecular analysis. We present results regarding efficacy and safety. Results: From November 2018 to October 2019 28 patients have been enrolled. 52%/48% of patients had PS 0/1. Median age was 70. TNM stage was: pT2 in 73,6% patients, pT3 in 10.6%, pT4 in 15.8% and 10.6% presented nodal spread. 13 patients have completed neoadjuvant treatment so far and 12 have undergone cystectomy. A wound dehiscence and one death related to surgical procedures were postoperative complications. Pathological complete response rate is 44,5%. Radiological evaluation is ongoing. 10 serious adverse events non-treatment related have been communicated. Any grade of toxicity has been reported in 91% of patients but adverse events grade 3-4 was detected in only 8.3% of cases. Grade 1 pruritus was the unique IR adverse event described in one patient. PARP inhibitors-related adverse events were grade 1 nausea and vomiting (25%), and grade 1 anemia (25%). Conclusions: Preliminary clinical data suggest that Durvalumab in combination with Olaparib could be active and well-tolerated neoadjuvant treatment of MIBC. Molecular characterization and biomarker discovery will be presented separately. Clinical trial information: NCT03534492.
DOI: 10.1038/s41379-020-0607-z
2020
Cited 31 times
Molecular characterization of chromophobe renal cell carcinoma reveals mTOR pathway alterations in patients with poor outcome
DOI: 10.1200/jco.2021.39.15_suppl.4576
2021
Cited 25 times
Efficacy of nivolumab/ipilimumab in patients with initial or late progression with nivolumab: Updated analysis of a tailored approach in advanced renal cell carcinoma (TITAN-RCC).
4576 Background: TITAN-RCC uses a tailored immunotherapy approach in renal cell carcinoma (RCC), starting with nivolumab (nivo) induction followed by nivo + ipilimumab (ipi) as immunotherapeutic “boost” in non-responders. Patients with initial partial or complete response (PR/CR) continued with nivo maintenance but received later “boosts” for progressive disease (PD). Here we report updated results focusing on the efficacy of nivo+ipi in patients with initial PD vs. initial responders with later PD. Methods: Patients with IMDC intermediate and poor risk advanced clear cell RCC were recruited between OCT 2016 and DEC 2018. Patients started with nivo 240 mg Q2W induction. Patients with early significant PD (week 8) or non-responders at week 16 received 2-4 nivo+ipi “boost” cycles. Responders (PR/CR) to nivo monotherapy continued with maintenance but could receive nivo+ipi for later PD. The primary endpoint is confirmed investigator assessed objective response rate (ORR) per RECIST in first line (1L) and second line (2L). Secondary endpoints included activity of nivo monotherapy, remission rate with nivo+ipi “boost”, safety and overall survival (OS). Results: 109 1L and 98 2L (after TKI) patients were analyzed for efficacy. Median age was 65 years (range 20-87). 71 % were intermediate and 25 % poor risk. Confirmed ORR with nivo monotherapy was 28 % for 1L and 17 % for 2L. After a median follow-up of 12.8 months best overall response after nivo induction ± nivo+ipi was 36 % in 1L and 30 % in 2L. Of all patients, 38 received nivo+ipi for stable disease (SD) up to week 16, with 1 (3 %), 4 (11 %) and 26 (68 %) achieving CR, PR and SD, respectively. 28 patients in 1L and 43 in 2L were boosted with nivo+ipi for initial PD. Of these, 3 (11 %) and 8 (29 %) achieved PR and SD, respectively, in 1L, whereas 3 (7.0 %) achieved CR, 6 (14 %) PR and 13 (30 %) SD in 2L. 16 and 10 patients received “boosts” later than week 16 for PD during nivo maintenance in 1L and 2L, respectively. Thereof, 3 (19 %) achieved PR and 5 (31 %) SD in 1L, whereas 2 (20 %) achieved PR and 3 (30 %) SD in 2L. Progression-free survival was 6.3 months (95 % CI 3.7 – 10.1) and 3.7 months (95 % CI 2.0 - 4.5) in 1L and 2L, respectively. OS was 27.2 months (95 % CI 19.9 – not estimable (NE)) in 1L and 20.2 months (95 % CI 15.6 – NE) in 2L. Treatment-related adverse events will be presented. Conclusions: Our tailored approach with nivo+ipi “boosts” results in improved response rates compared to nivo monotherapy. Our updated analysis suggests that almost half of the patients receiving “boosts” for PD improve to either PR/CR (18 %) or SD (30 %), irrespective of initial or later progression with nivo. Clinical trial information: NCT02917772. [Table: see text]
DOI: 10.1002/cam4.3717
2021
Cited 24 times
Cabozantinib real‐world effectiveness in the first‐through fourth‐line settings for the treatment of metastatic renal cell carcinoma: Results from the International Metastatic Renal Cell Carcinoma Database Consortium
Abstract Background Cabozantinib is approved for metastatic renal cell carcinoma (mRCC) based on the METEOR and CABOSUN trials. However, real‐world effectiveness and dosing patterns of cabozantinib are not well characterized. Methods Patients with mRCC treated with cabozantinib between 2011 and 2019 were identified and stratified using the International mRCC Database Consortium (IMDC) risk groups. First‐ (1L), second‐ (2L), third‐ (3L), and fourth‐line (4L) overall response rate (ORR), time to treatment failure (TTF), and overall survival (OS) were analyzed. Dose reduction rates and their association with TTF and OS were determined. Results A total of 413 patients were identified. The ORRs across 1L to 4L were 32%, 26%, 25%, and 29%, respectively, and the median TTF rates were 8.3, 7.3, 7.0, and 8.0 months, respectively. The median OS (mOS) rates in 1L to 4L were 30.7, 17.8, 12.6, and 14.9 months, respectively. For patients treated with 1L PD(L)1 combination agent ( n = 31), 2L cabozantinib had ORR of 22%, median TTF of 5.4 months, and mOS of 17.4 months. About 50% (129/258) of patients required dose reductions. The TTF and mOS were significantly longer for patients who required dose reduction vs. patients who did not, with an adjusted hazard ratio of 0.37 (95% CI 0.202–0.672, p &lt; 0.01) and 0.46 (95% CI 0.215–0.980, p = 0.04), respectively. Limitations include the retrospective study design and the lack of central radiology review. Conclusion The ORR and TTF of cabozantinib were maintained from the 1L to 4L settings. Dose reductions due to toxicity were associated with improved TTF and OS. Cabozantinib has clinical activity after 1L Immuno‐oncology combination agents.
DOI: 10.1016/j.ctrv.2023.102652
2024
Vascular endothelial growth factor-targeted therapy in patients with renal cell carcinoma pretreated with immune checkpoint inhibitors: A systematic literature review
IntroductionWe conducted a systematic literature review to identify evidence for use of vascular endothelial growth factor (VEGF)-targeted (anti-VEGF) treatment in patients with renal cell carcinoma (RCC) following prior checkpoint inhibitor (CPI)-based therapy.MethodsThis was a PRISMA-standard systematic literature review; registered with PROSPERO (CRD42021255568). Literature searches were conducted in MEDLINE®, Embase, and the Cochrane Library (January 28, 2021; updated September 13, 2022) to identify publications reporting efficacy/effectiveness and safety/tolerability evidence for anti-VEGF treatment in patients with RCC who had received prior CPI therapy.ResultsOf 2,639 publications screened, 48 were eligible and featured 2,759 patients treated in trials and 2,209 in real-world studies (RWS). Most patients with available data were treated with anti-VEGF tyrosine kinase inhibitor-based regimens (trials: 93%; RWS: 100%), most commonly cabozantinib, which accounted for 46% of trial and 62% of RWS patients in publications with available data. Collectively, there was consistent evidence of anti-VEGF treatment activity after prior CPI therapy. Activity was reported for all anti-VEGF regimens and regardless of prior CPI-based regimen. No new safety signals were detected for subsequent anti-VEGF therapy; no studies suggested increased immune-related adverse events associated with prior CPI therapy. The results were limited by data quality; study heterogeneity prohibited meta-analyses.ConclusionBased on the available data (most commonly for cabozantinib), anti-VEGF therapy appears to be a rational treatment choice in patients with RCC who have progressed despite prior CPI-based therapy. Results from ongoing trials of combination anti-VEGF plus CPI regimen post prior CPI therapy trials will contribute more definitive evidence.Plain language summaryAnticancer treatments that work by reducing levels of a substance in the body called Vascular Endothelial Growth Factor are known as anti-VEGF drugs. Reducing VEGF levels helps to reduce blood supply to tumors, which can slow the speed at which the cancer grows. Some other types of anticancer drugs that help the immune system to fight cancer cells are called checkpoint inhibitors. Here, we looked at published studies that investigated how anti-VEGF drugs work, and what side effects they cause, in people who have already been treated with checkpoint inhibitors for a type of kidney cancer called renal cell carcinoma. We aimed to summarize the available evidence to help doctors decide how best to use anti-VEGF drugs in these patients. We found 48 studies that included almost 5,000 patients. The results of the studies showed that anti-VEGF drugs have anticancer effects in people with renal cell carcinoma who had already been treated with checkpoint inhibitors. All of the VEGF-targeting drugs had anticancer effects, irrespective of what checkpoint inhibitor treatment people had received before. There were different amounts of evidence available for the different anti-VEGF drugs. The anti-VEGF cabozantinib has the largest amount of evidence. Importantly, previous checkpoint inhibitor treatment did not seem to affect the number or type of side-effects associated with anti-VEGF drugs. Results from ongoing, well-designed studies will be helpful to confirm these results. Our findings may be useful for doctors considering using anti-VEGF drugs in patients with renal cell carcinoma who have received checkpoint inhibitor treatment.
DOI: 10.1186/s12964-023-01462-0
2024
Transcending frontiers in prostate cancer: the role of oncometabolites on epigenetic regulation, CSCs, and tumor microenvironment to identify new therapeutic strategies
Prostate cancer, as one of the most prevalent malignancies in males, exhibits an approximate 5-year survival rate of 95% in advanced stages. A myriad of molecular events and mutations, including the accumulation of oncometabolites, underpin the genesis and progression of this cancer type. Despite growing research demonstrating the pivotal role of oncometabolites in supporting various cancers, including prostate cancer, the root causes of their accumulation, especially in the absence of enzymatic mutations, remain elusive. Consequently, identifying a tangible therapeutic target poses a formidable challenge. In this review, we aim to delve deeper into the implications of oncometabolite accumulation in prostate cancer. We center our focus on the consequential epigenetic alterations and impacts on cancer stem cells, with the ultimate goal of outlining novel therapeutic strategies.
DOI: 10.1016/j.canlet.2024.216661
2024
Immune checkpoints between epithelial-mesenchymal transition and autophagy: A conflicting triangle
Inhibitory immune checkpoint (ICP) molecules are pivotal in inhibiting innate and acquired antitumor immune responses, a mechanism frequently exploited by cancer cells to evade host immunity. These evasion strategies contribute to the complexity of cancer progression and therapeutic resistance. For this reason, ICP molecules have become targets for antitumor drugs, particularly monoclonal antibodies, collectively referred to as immune checkpoint inhibitors (ICI), that counteract such cancer-associated immune suppression and restore antitumor immune responses. Over the last decade, however, it has become clear that tumor cell-associated ICPs can also induce tumor cell-intrinsic effects, in particular epithelial-mesenchymal transition (EMT) and macroautophagy (hereafter autophagy). Both of these processes have profound implications for cancer metastasis and drug responsiveness. This article reviews the positive or negative cross-talk that tumor cell-associated ICPs undergo with autophagy and EMT. We discuss that tumor cell-associated ICPs are upregulated in response to the same stimuli that induce EMT. Moreover, ICPs themselves, when overexpressed, become an EMT-inducing stimulus. As regards the cross-talk with autophagy, ICPs have been shown to either stimulate or inhibit autophagy, while autophagy itself can either up- or downregulate the expression of ICPs. This dynamic equilibrium also extends to the autophagy-apoptosis axis, further emphasizing the complexities of cellular responses. Eventually, we delve into the intricate balance between autophagy and apoptosis, elucidating its role in the broader interplay of cellular dynamics influenced by ICPs. In the final part of this article, we speculate about the driving forces underlying the contradictory outcomes of the reciprocal, inhibitory, or stimulatory effects between ICPs, EMT, and autophagy. A conclusive identification of these driving forces may allow to achieve improved antitumor effects when using combinations of ICIs and compounds acting on EMT and/or autophagy. Prospectively, this may translate into increased and/or broadened therapeutic efficacy compared to what is currently achieved with ICI-based clinical protocols.
DOI: 10.1046/j.1471-4159.1998.71041597.x
1998
Cited 84 times
Role of Carnitine Palmitoyltransferase I in the Control of Ketogenesis in Primary Cultures of Rat Astrocytes
Abstract: The role of carnitine palmitoyltransferase I (CPT‐I) in the control of ketogenesis was studied in primary cultures of rat astrocytes. Ketone bodies were the major product of [ 14 C]palmitate oxidation by cultured astrocytes, whereas CO 2 made a minor contribution to the total oxidation products. Using tetradecylglycidate as a specific, cell‐permeable inhibitor of CPT‐I, a flux control coefficient of 0.77 ± 0.07 was calculated for CPT‐I over the flux of [ 14 C]palmitate to ketone bodies. CPT‐I from astrocytes was sensitive to malonyl‐CoA (IC 50 = 3.4 ± 0.8 µ M ) and cross‐reacted on western blots with an antibody raised against liver CPT‐I. On the other hand, astrocytes expressed significant acetyl‐CoA carboxylase (ACC) activity, and consequently they contained considerable amounts of malonyl‐CoA. Western blot analysis of ACC isoforms showed that ACC in astrocytes—like in neurons, liver, and white adipose tissue—mostly comprised the 265‐kDa isoform, whereas the 280‐kDa isoform—which was highly expressed in skeletal muscle—showed much lower abundance. Forskolin was used as a tool to study the modulation of the ketogenic pathway in astrocytes. Thus, forskolin decreased in parallel ACC activity and intracellular malonyl‐CoA levels, whereas it stimulated CPT‐I activity and [ 14 C]palmitate oxidation to both ketone bodies and CO 2 . Results show that in cultured astrocytes (a) CPT‐I exerts a very high degree of control over ketogenesis from palmitate, (b) the ACC/malonyl‐CoA/CPT‐I system is similar to that of liver, and (c) the ACC/malonyl‐CoA/CPT‐I system is subject to regulation by cyclic AMP.
DOI: 10.1016/j.febslet.2005.08.021
2005
Cited 76 times
p38 MAPK is involved in CB<sub>2</sub> receptor‐induced apoptosis of human leukaemia cells
Cannabinoids have been shown to inhibit the growth of a broad spectrum of tumour cells. However, the molecular mechanisms involved in that effect have not been completely elucidated. Here, we investigated the possible involvement of mitogen‐activated protein kinases (MAPKs) in CB 2 receptor‐induced apoptosis of human leukaemia cells. Results show that stimulation of the CB 2 receptor leads to p38 MAPK activation and that inhibition of this kinase attenuates CB 2 receptor‐induced caspase activation and apoptosis. These findings support a role for p38 MAPK in CB 2 receptor‐induced apoptosis of human leukaemia cells.
DOI: 10.1016/j.lfs.2005.05.015
2005
Cited 75 times
Cannabinoids and ceramide: Two lipids acting hand-by-hand
Cannabinoids, the active components of Cannabis sativa (marijuana) and their endogenous counterparts, exert their effects by binding to specific G-protein-coupled receptors that modulate adenylyl cyclase and ion channels. Recent research has shown that the CB1 cannabinoid receptor is also coupled to the generation of the lipid second messenger ceramide via two different pathways: sphingomyelin hydrolysis and ceramide synthesis de novo. Sustained ceramide accumulation in tumor cells mediates cannabinoid-induced apoptosis, as evidenced by in vitro and in vivo studies. This effect seems to be due to the impact of ceramide on key cell signalling systems such as the extracellular signal-regulated kinase cascade and the Akt pathway. These findings provide a new conceptual view on how cannabinoids act, and raise interesting physiological and therapeutic questions.
DOI: 10.1016/j.neuropharm.2004.04.016
2004
Cited 66 times
Hypothesis: cannabinoid therapy for the treatment of gliomas?
Gliomas, in particular glioblastoma multiforme or grade IV astrocytoma, are the most frequent class of malignant primary brain tumours and one of the most aggressive forms of cancer. Current therapeutic strategies for the treatment of glioblastoma multiforme are usually ineffective or just palliative. During the last few years, several studies have shown that cannabinoids-the active components of the plant Cannabis sativa and their derivatives--slow the growth of different types of tumours, including gliomas, in laboratory animals. Cannabinoids induce apoptosis of glioma cells in culture via sustained ceramide accumulation, extracellular signal-regulated kinase activation and Akt inhibition. In addition, cannabinoid treatment inhibits angiogenesis of gliomas in vivo. Remarkably, cannabinoids kill glioma cells selectively and can protect non-transformed glial cells from death. These and other findings reviewed here might set the basis for a potential use of cannabinoids in the management of gliomas.
DOI: 10.1111/j.1755-148x.2008.00481.x
2008
Cited 56 times
The antidepressant sertraline downregulates Akt and has activity against melanoma cells
Summary Melanoma is a common malignancy which is poorly responsive to chemotherapy and radiation. One of the major reasons melanoma responds poorly to these modalities is constitutive expression of Akt, which protects against apoptosis. The antidepressant sertraline was found to be a potent cytotoxic agent against A375 human melanoma. To determine the mechanism by which sertraline kills melanoma cells, Western blot analysis of signaling molecules, including phosphorylated Akt, caspase 9 and phospho‐p70 S6 kinase was performed. Finally, the effects of sertraline on A375 xenografts in mice were assessed. Sertaline potently inhibited the phosphorylation of Akt, and caused cell death through induction of endoplasmic reticulum in vitro. Sertraline monotherapy demonstrated activity against A375 xenografts in vivo. Akt is a major cause of resistance of melanoma to current therapy. Antidepressants are commonly used to prevent interferon‐induced depression. Use of antidepressants that decrease Akt may improve the efficacy of interferon and other therapies against melanoma. Further studies are needed to elucidate whether sertraline acts as an Akt inhibitor in melanoma.
DOI: 10.1038/ncb3171
2015
Cited 41 times
Erratum: Corrigendum: AMBRA1 links autophagy to cell proliferation and tumorigenesis by promoting c-Myc dephosphorylation and degradation
Nat. Cell Biol. 17, 20–30 (2015); published online 1 December 2014; corrected after print 1 April 2015 In the version of this Article originally published, incorrect western blot scans were provided for the actin panels in Figure 4h,i. These panels have been corrected online and are shown above. Allsamples in 4i were collected and processed simultaneously, on the same or on parallel gels/blots.
DOI: 10.1007/s12307-018-0211-7
2018
Cited 35 times
Exosomes: Definition, Role in Tumor Development and Clinical Implications
Exosomes are microvesicles released by cells in both physiological and pathological situations. They are surrounded by a lipid bilayer with proteins derived from the origin cell, and contain a variety of molecules, such as nucleic acids. They represent an emerging mechanism of intercellular communication, and they play an important role in the pathogenesis of cancer, stimulating proliferation and aggressiveness of cancer cells, inducing a microenvironment favorable to tumor development and controlling immune responses. Because of the growing understanding of the potential implications of extracellular vesicles in the development of malignancies, research on exosomes, and its role as a diagnostic and therapeutic tool, constitutes nowadays a very exciting and promising field.
DOI: 10.1038/s41416-018-0064-3
2018
Cited 35 times
Targeted genomic landscape of metastases compared to primary tumours in clear cell metastatic renal cell carcinoma
The genomic landscape of primary clear cell renal cell carcinoma (ccRCC) has been well described. However, little is known about cohort genomic alterations (GA) landscape in ccRCC metastases, or how it compares to primary tumours in aggregate. The genomic landscape of metastases may have biological, clinical, and therapeutic implications.We collected targeted next-generation sequencing mutation calls from two independent cohorts and described the metastases GA landscape and descriptively compared it to the GA landscape in primary tumours.The cohort 1 (n = 578) consisted of 349 primary tumours and 229 metastases. Overall, the most common mutations in the metastases were VHL (66.8%), PBRM1 (41.87%), and SETD2 (24.7%). TP53 was more frequently mutated in metastases compared to primary tumours (14.85% versus 8.9%; p = 0.031). No other gene had significant difference in the cohort frequency of mutations between the metastases and primary tumours. Mutation burden was not significantly different between the metastases and primary tumours or between metastatic sites. The second cohort (n = 257) consisted of 177 primary tumours and 80 metastases. No differences in frequency of mutations or mutational burden were observed between primaries and metastases.These data support the theory that ccRCC primary tumours and metastases encompass a uniform distribution of common genomic alterations tested by next-generation sequencing targeted panels. This study does not address variability between matched primary tumours and metastases or the change in genomic alterations over time and after sequential systemic therapies.
DOI: 10.1016/j.ejso.2018.05.026
2018
Cited 31 times
Predictors, utilization patterns, and overall survival of patients undergoing metastasectomy for metastatic renal cell carcinoma in the era of targeted therapy
Introduction Metastasectomy (MSX) is considered a treatment option in patients with metastatic renal cell carcinoma (mRCC) at diagnosis, but its role in the targeted therapy era is unclear. We sought to describe the utilization trends of MSX and survival outcomes in a large US cohort. Materials and methods Using the National Cancer Database, we identified patients undergoing MSX for mRCC at diagnosis between 2006 and 2013. Linear regression methods estimated utilization trends, and hierarchical models identified independent predictors of MSX after adjusting for hospital clustering. Kaplan-Meier survival estimates and Cox proportional hazard models were used to evaluate overall survival according to treatment after propensity-score matching. Results Of 6994 mRCC patients, 1976 underwent MSX (28.3%). Those treated at academic facilities were more likely to undergo a MSX (OR: 1.57, 95% CI 1.20–2.06, p = 0.001). In contrast, older patients (OR: 0.99, 95% CI: 0.98–1.00), black race (OR: 0.65, 95% CI: 0.51–0.82), higher pT stage (OR: 0.76, 95% CI: 0.65–0.89), and who received targeted therapy (OR: 0.72, 95% CI: 0.63–0.82, all p ≤ 0.008) were less likely to undergo MSX. Overall, MSX patients had an improved survival compared to non-MSX patients (HR: 0.83, 95% CI: 0.77–0.90, p < 0.001), as well as among patients treated with targeted therapy (HR: 0.77, 95% CI: 0.77–0.96, p = 0.008). Conclusions Our findings indicate that MSX-treated patients may benefit from an improved overall survival compared to non-MSX treated patients. Good patient selection and a proper risk stratification strategy are still very important considerations.
DOI: 10.1242/jcs.234120
2019
Cited 31 times
Inhibiting SUMO1-mediated SUMOylation induces autophagy-mediated cancer cell death and reduces tumour cell invasion via RAC1
Post-translational modifications directly control protein activity and, thus, they represent an important means to regulate the responses of cells to different stimuli. Protein SUMOylation has recently been recognised as one such modification, and it has been associated with various diseases, including different types of cancer. However, the precise way that changes in SUMOylation influence the tumorigenic properties of cells remains to be fully clarified. Here, we show that blocking the SUMO pathway by depleting SUMO1 and UBC9, or by exposure to ginkgolic acid C15:1 or 2-D08 (two different SUMOylation inhibitors), induces cell death, also inhibiting the invasiveness of tumour cells. Indeed, diminishing the formation of SUMO1 complexes induces autophagy-mediated cancer cell death through increasing the expression of Tribbles pseudokinase 3 (TRIB3). Moreover, we found that blocking the SUMO pathway inhibits tumour cell invasion by decreasing RAC1 SUMOylation. These findings shed new light on the mechanisms by which SUMO1 modifications regulate the survival, and the migratory and invasive capacity of tumour cells, potentially establishing the bases to develop novel anti-cancer treatments based on the inhibition of SUMOylation.
DOI: 10.7150/thno.41450
2020
Cited 28 times
Midkine signaling maintains the self-renewal and tumorigenic capacity of glioma initiating cells
Glioblastoma (GBM) is one of the most aggressive forms of cancer.It has been proposed that the presence within these tumors of a population of cells with stem-like features termed Glioma Initiating Cells (GICs) is responsible for the relapses that take place in the patients with this disease.Targeting this cell population is therefore an issue of great therapeutic interest in neuro-oncology.We had previously found that the neurotrophic factor MIDKINE (MDK) promotes resistance to glioma cell death.The main objective of this work is therefore investigating the role of MDK in the regulation of GICs.Methods: Assays of gene and protein expression, self-renewal capacity, autophagy and apoptosis in cultures of GICs derived from GBM samples subjected to different treatments.Analysis of the growth of GICs-derived xenografts generated in mice upon blockade of the MDK and its receptor the ALK receptor tyrosine kinase (ALK) upon exposure to different treatments.Results: Genetic or pharmacological inhibition of MDK or ALK decreases the self-renewal and tumorigenic capacity of GICs via the autophagic degradation of the transcription factor SOX9. Blockade of the MDK/ALK axis in combination with temozolomide depletes the population of GICs in vitro and has a potent anticancer activity in xenografts derived from GICs.Conclusions: The MDK/ALK axis regulates the self-renewal capacity of GICs by controlling the autophagic degradation of the transcription factor SOX9. Inhibition of the MDK/ALK axis may be a therapeutic strategy to target GICs in GBM patients.
DOI: 10.1016/j.ejca.2021.04.009
2021
Cited 19 times
Response to systemic therapy in fumarate hydratase–deficient renal cell carcinoma
Fumarate hydratase-deficient (FHdef) renal cell carcinoma (RCC) is a rare entity associated with the hereditary leiomyomatosis and RCC syndrome with no standard therapy approved. The aim of this retrospective study was to evaluate the efficacy of different systemic treatments in this population.We performed a multicentre retrospective analysis of Fhdef RCC patients to determine the response to systemic treatments. The endpoints were objective response rate (ORR), time-to-treatment failure (TTF), and overall survival (OS). The two latter were estimated using the Kaplan-Meier method.Twenty-four Fhdef RCC patients were identified, and 21 under systemic therapy were included in the analysis: ten received cabozantinib, 14 received sunitinib, nine received "other antiangiogenics" (sorafenib, pazopanib, and axitinib), three received erlotinib-bevacizumab (E-B), three received mTOR inhibitors, and 11 received immune checkpoint blockers (ICBs). ORR for treatments were 50% for cabozantinib, 43% for sunitinib, 63% for "other antiangiogenics," and 30% for E-B, whereas ORR was 0% for mTOR inhibitors and 18% for ICBs. The median TTF (mTTF) was significantly higher with antiangiogenics (11.6 months) than with mTOR inhibitors (4.4 months) or ICBs (2.7 months). In the first-line setting, antiangiogenics presented a higher ORR compared with nivolumab-ipilimumab (64% versus 25%) and a significantly superior mTTF (11.0 months vs 2.5 months; p = 0.0027). The median OS from the start of the first systemic treatment was 44.0 months (95% confidence interval: 13.0-95.0).We report the first European retrospective study of Fhdef RCC patients treated with systemic therapy with a remarkably long median OS of 44.0 months. Our results suggest that antiangiogenics may be superior to ICB/mTOR inhibitors in this population.
DOI: 10.1080/2162402x.2022.2067944
2022
Cited 12 times
Characterization of plasma circulating small extracellular vesicles in patients with metastatic solid tumors and newly diagnosed brain metastasis
Nearly 40% of the advanced cancer patients will present brain metastases during the course of their disease, with a 2-year life expectancy of less than 10%. Immune system impairment, including the modulation of both STAT3 and PD-L1, is one of the hallmarks of brain metastases. Liquid biopsy could offer several advantages in brain metastases management, such as the possibility of noninvasive dynamic monitoring. Extracellular vesicles (EVs) have been recently proposed as novel biomarkers especially useful in liquid biopsy due to their secretion in biofluids and their role in cell communication during tumor progression. The main aim of this work was to characterize the size and protein cargo of plasma circulating EVs in patients with solid tumors and their correlation with newly diagnosed brain metastases, in addition to their association with other relevant clinical variables. We analyzed circulating EVs in the plasma of 123 patients: 42 patients with brain metastases, 50 without brain metastases and 31 healthy controls. Patients with newly diagnosed brain metastases had a lower number of circulating EVs in the plasma and a higher protein concentration in small EVs (sEVs) compared to patients without brain metastases and healthy controls. Interestingly, melanoma patients with brain metastases presented decreased STAT3 activation and increased PD-L1 levels in circulating sEVs compared to patients without central nervous system metastases. Decreased STAT3 activation and increased PD-L1 in plasma circulating sEVs identify melanoma patients with brain metastasis.
DOI: 10.1016/j.xcrm.2023.101101
2023
Cited 4 times
Single-cell dissection of Merkel cell carcinoma heterogeneity unveils transcriptomic plasticity and therapeutic vulnerabilities
Merkel cell carcinoma (MCC), a rare but aggressive skin cancer, remains a challenge in the era of precision medicine. Immune checkpoint inhibitors (ICIs), the only approved therapy for advanced MCC, are impeded by high primary and acquired resistance. Hence, we dissect transcriptomic heterogeneity at single-cell resolution in a panel of patient tumors, revealing phenotypic plasticity in a subset of treatment-naive MCC. The tumor cells in a "mesenchymal-like" state are endowed with an inflamed phenotype that portends a better ICI response. This observation is also validated in the largest whole transcriptomic dataset available from MCC patient tumors. In contrast, ICI-resistant tumors predominantly express neuroepithelial markers in a well-differentiated state with "immune-cold" landscape. Importantly, a subtle shift to "mesenchymal-like" state reverts copanlisib resistance in primary MCC cells, highlighting potential strategies in patient stratification for therapeutics to harness tumor cell plasticity, augment treatment efficacy, and avert resistance.
DOI: 10.1021/jm0407714
2004
Cited 64 times
4,5-Dihydro-1<i>H-</i>pyrazole Derivatives with Inhibitory nNOS Activity in Rat Brain: Synthesis and Structure−Activity Relationships
In an attempt to find new compounds with neuroprotective activity, we have designed, synthesized and characterized 19 new nNOS inhibitors with a 4,5-dihydro-1H-pyrazole structure. Compounds 11r [1-cyclopropanecarbonyl-3-(2-amino-5-chlorophenyl)-4,5-dihydro-1H-pyrazole] and 11e [1-cyclopropanecarbonyl-3-(2-amino-5-methoxyphenyl)- 4,5-dihydro-1H-pyrazole] show the highest activities with inhibition percentages of 70% and 62%, respectively. A structure−activity relationship for the nNOS inhibition can be established from the structural comparison of these new pyrazole derivatives and the described synthetic kynurenines 10.
DOI: 10.1042/bj20031714
2004
Cited 59 times
Ceramide sensitizes astrocytes to oxidative stress: protective role of cannabinoids
Cannabinoids induce apoptosis on glioma cells via stimulation of ceramide synthesis de novo, whereas they do not affect viability of primary astrocytes. In the present study, we show that incubation with Δ9-tetrahydrocannabinol did not induce accumulation of ceramide on astrocytes, although incubation of these cells in a serum-free medium (with or without cannabinoids) led to stimulation of ceramide synthesis de novo and sensitization to oxidative stress. Thus treatment with H2O2 induced apoptosis of 5-day-serum-deprived astrocytes and this effect was abrogated by pharmacological blockade of ceramide synthesis de novo. The sensitizing effect of ceramide accumulation may depend on p38 mitogen-activated protein kinase activation rather than on other ceramide targets. Finally, a protective role of cannabinoids on astrocytes is shown as a long-term incubation with cannabinoids prevented H2O2-induced loss of viability in a CB1 receptor-dependent manner. In summary, our results show that whereas challenge of glioma cells with cannabinoids induces accumulation of de novo-synthesized ceramide and apoptosis, long-term treatment of astrocytes with these compounds does not stimulate this pathway and also abrogates the sensitizing effects of ceramide accumulation.
DOI: 10.1016/j.neuropharm.2007.06.021
2008
Cited 46 times
Down-regulation of tissue inhibitor of metalloproteinases-1 in gliomas: a new marker of cannabinoid antitumoral activity?
Cannabinoids, the active components of Cannabis sativa L. and their derivatives, inhibit tumor growth in laboratory animals by inducing apoptosis of tumor cells and inhibiting tumor angiogenesis. It has also been reported that cannabinoids inhibit tumor cell invasiveness, but the molecular targets of this cannabinoid action remain elusive. Here we evaluated the effects of cannabinoids on the expression of tissue inhibitors of metalloproteinases (TIMPs), which play critical roles in the acquisition of migrating and invasive capacities by tumor cells. Local administration of Δ9-tetrahydrocannabinol (THC), the major active ingredient of cannabis, down-regulated TIMP-1 expression in mice bearing subcutaneous gliomas, as determined by Western blot and immunofluorescence analyses. This cannabinoid-induced inhibition of TIMP-1 expression in gliomas (i) was mimicked by JWH-133, a selective CB2 cannabinoid receptor agonist that is devoid of psychoactive side effects, (ii) was abrogated by fumonisin B1, a selective inhibitor of ceramide synthesis de novo, and (iii) was also evident in two patients with recurrent glioblastoma multiforme (grade IV astrocytoma). THC also depressed TIMP-1 expression in cultures of various human glioma cell lines as well as in primary tumor cells obtained from a glioblastoma multiforme patient. This action was prevented by pharmacological blockade of ceramide biosynthesis and by knocking-down the expression of the stress protein p8. As TIMP-1 up-regulation is associated with high malignancy and negative prognosis of numerous cancers, TIMP-1 down-regulation may be a hallmark of cannabinoid-induced inhibition of glioma progression.
DOI: 10.1158/1535-7163.mct-08-1147
2009
Cited 45 times
Cannabinoid receptor 1 is a potential drug target for treatment of translocation-positive rhabdomyosarcoma
Abstract Gene expression profiling has revealed that the gene coding for cannabinoid receptor 1 (CB1) is highly up-regulated in rhabdomyosarcoma biopsies bearing the typical chromosomal translocations PAX3/FKHR or PAX7/FKHR. Because cannabinoid receptor agonists are capable of reducing proliferation and inducing apoptosis in diverse cancer cells such as glioma, breast cancer, and melanoma, we evaluated whether CB1 is a potential drug target in rhabdomyosarcoma. Our study shows that treatment with the cannabinoid receptor agonists HU210 and Δ9-tetrahydrocannabinol lowers the viability of translocation-positive rhabdomyosarcoma cells through the induction of apoptosis. This effect relies on inhibition of AKT signaling and induction of the stress-associated transcription factor p8 because small interfering RNA–mediated down-regulation of p8 rescued cell viability upon cannabinoid treatment. Finally, treatment of xenografts with HU210 led to a significant suppression of tumor growth in vivo. These results support the notion that cannabinoid receptor agonists could represent a novel targeted approach for treatment of translocation-positive rhabdomyosarcoma. [Mol Cancer Ther 2009;8(7):1838–45]
DOI: 10.1016/j.ejca.2013.01.002
2013
Cited 36 times
Gene expression changes associated with erlotinib response in glioma cell lines
Erlotinib (ERL), a tyrosine kinase inhibitor that acts on the epidermal growth factor receptor (EGFR), is used as a second line treatment for glioma therapy, with controversial findings regarding its response. Here, we analysed the gene expression profiles of a series of human glioma cell lines with differing sensitivities to ERL to identify the gene expression changes associated with ERL response. The varying responses to ERL were associated with different expression levels of specific genes (HRAS, CTFG, ERCC5 and HDAC3) and genes associated with specific pathways (apoptosis and cell death). PI3K pathway genes were primarily affected by ERL, as we found that PIK3R3 was repressed by ERL treatment in sensitive glioma cell lines. The cell cycle and ubiquitin pathways were also affected by EGFR inhibition, as GAS5, PLK1 and BIRC5 were the most significantly affected genes. In this study we have identified several genes such as PIK3R3 and GAS5, that can be targeted in order to enhance the response to ERL therapy.
DOI: 10.3390/cancers12061550
2020
Cited 23 times
Clinical Applications of Molecular Biomarkers in Prostate Cancer
There is clinically relevant molecular heterogeneity in prostate cancer (PCa), but this biological diversity has had only a minimal impact on clinical practice. Treatment outcomes in patients with localised PCa are often highly variable, even among patients stratified to the same risk group or disease state based on standard clinical and pathological parameters. In recent years, the development of gene panels has provided valuable data on the differential expression of genes in patients with PCa. Nevertheless, there is an urgent need to identify and validate prognostic and predictive biomarkers that can be applied across clinical scenarios, ranging from localised disease to metastatic castration-resistant PCa. The availability of such tools would allow for precision medicine to finally reach PCa patients. In this review, we evaluate current data on molecular biomarkers for PCa, with an emphasis on the biomarkers and gene panels with the most robust evidence to support their application in routine clinical practice.
DOI: 10.3389/fimmu.2021.786069
2022
Cited 10 times
Transcriptomic Mapping of Non-Small Cell Lung Cancer K-RAS p.G12C Mutated Tumors: Identification of Surfaceome Targets and Immunologic Correlates
Targeting K-RAS-mutant non-small cell lung cancer (NSCLC) with novel inhibitors has shown promising results with the recent approval of sotorasib in this indication. However, progression to this agent is expected, as it has previously been observed with other inhibitors. Recently, new immune therapeutics, including vectorized compounds with antibodies or modulators of the host immune response, have demonstrated clinical activity. By interrogating massive datasets, including TCGA, we identified genes that code for surface membrane proteins that are selectively expressed in K-RAS mutated NSCLC and that could be used to vectorize novel therapies. Two genes, CLDN10 and TMPRSS6, were selected for their clear differentiation. In addition, we discovered immunologic correlates of outcome that were clearly de-regulated in this particular tumor type and we matched them with immune cell populations. In conclusion, our article describes membrane proteins and immunologic correlates that could be used to better select and optimize current therapies.
DOI: 10.1042/bj3630183
2002
Cited 48 times
De novo-synthesized ceramide is involved in cannabinoid-induced apoptosis
Δ9-Tetrahydrocannabinol (THC) and other cannabinoids have been shown to induce apoptosis of glioma cells via ceramide generation. In the present study, we investigated the metabolic origin of the ceramide responsible for this cannabinoid-induced apoptosis by using two subclones of C6 glioma cells: C6.9, which is sensitive to THC-induced apoptosis; and C6.4, which is resistant to THC-induced apoptosis. Pharmacological inhibition of ceramide synthesis de novo, but not of neutral and acid sphingomyelinases, prevented THC-induced apoptosis in C6.9 cells. The activity of serine palmitoyltransferase (SPT), which catalyses the rate-limiting step of ceramide synthesis de novo, was remarkably enhanced by THC in C6.9 cells, but not in C6.4 cells. However, no major changes in SPT mRNA and protein levels were evident. Changes in SPT activity paralleled changes in ceramide levels. Pharmacological inhibition of ceramide synthesis de novo also prevented the stimulation of extracellular-signal-regulated kinase and the inhibition of protein kinase B triggered by cannabinoids. These findings show that de novo-synthesized ceramide is involved in cannabinoid-induced apoptosis of glioma cells.
DOI: 10.1021/jm050740o
2005
Cited 47 times
Kynurenamines as Neural Nitric Oxide Synthase Inhibitors
To find new compounds with potential neuroprotective activity, we have designed, synthesized, and characterized a series of neural nitric oxide synthase (nNOS) inhibitors with a kynurenamine structure. Among them, N-[3-(2-amino-5-methoxyphenyl)-3-oxopropyl]acetamide is the main melatonin metabolite in the brain and shows the highest activity in the series, with an inhibition percentage of 65% at a 1 mM concentration. The structure-activity relationship of the new series partially reflects that of the previously reported 2-acylamido-4-(2-amino-5-methoxyphenyl)-4-oxobutyric acids, endowed with a kynurenine-like structure. Structural comparisons between these new kinurenamine derivatives, kynurenines, and 1-acyl-3-(2-amino-5-methoxyphenyl)-4,5-dihydro-1H-pyrazole derivatives also reported confirm our previous model for the nNOS inhibition.