RESUMEN
Germline loss-of-function mutations of the VHL tumor suppressor gene cause von Hippel-Lindau disease, which is associated with an increased risk of hemangioblastomas, clear cell renal cell carcinomas (ccRCCs), and paragangliomas. This Review describes mechanisms involving the VHL gene product in oxygen sensing, protein degradation, and tumor development and current therapeutic strategies targeting these mechanisms. The VHL gene product is the substrate recognition subunit of a ubiquitin ligase that targets the α subunit of the heterodimeric hypoxia-inducible factor (HIF) transcription factor for proteasomal degradation when oxygen is present. This oxygen dependence stems from the requirement that HIFα be prolyl-hydroxylated on one (or both) of two conserved prolyl residues by members of the EglN (also called PHD) prolyl hydroxylase family. Deregulation of HIF, and particularly HIF2, drives the growth of VHL-defective ccRCCs. Drugs that inhibit the HIF-responsive gene product VEGF are now mainstays of ccRCC treatment. An allosteric HIF2 inhibitor was recently approved for the treatment of ccRCCs arising in the setting of VHL disease and has advanced to phase III testing for sporadic ccRCCs based on promising phase I/II data. Orally available EglN inhibitors are being tested for the treatment of anemia and ischemia. Five of these agents have been approved for the treatment of anemia in the setting of chronic kidney disease in various countries around the world.
Asunto(s)
Carcinoma de Células Renales , Neoplasias Renales , Enfermedad de von Hippel-Lindau , Carcinoma de Células Renales/patología , Humanos , Neoplasias Renales/genética , Neoplasias Renales/metabolismo , Oxígeno/metabolismo , Proteolisis , Enfermedad de von Hippel-Lindau/genética , Enfermedad de von Hippel-Lindau/metabolismoRESUMEN
von Hippel-Lindau (VHL) disease is an autosomal dominant hereditary cancer disorder caused by a germline mutation in the VHL tumor suppressor gene. Loss of the wild-type allele results in VHL deficiency and the potential formation of cerebellar hemangioblastomas, which resemble embryonic hemangioblast proliferation and differentiation processes. Multiple, microscopic, VHL-deficient precursors, termed developmentally arrested structural elements (DASEs), consistently involve the cerebellar molecular layer in VHL patients, indicating the tumor site of origin. Unlike hemangioblastomas, however, cerebellar DASEs do not express brachyury, a mesodermal marker for hemangioblasts. In this study, neuronal progenitors occupying the molecular layer were investigated as tumor cells of origin. By immunohistochemistry, cerebellar DASEs and hemangioblastomas lacked immunoreactivity with antibody ZIC1 (Zic family member 1), a granule cell progenitor marker with concordance from oligonucleotide RNA expression array analyses. Rather, cerebellar DASEs and hemangioblastomas were immunoreactive with antibody PAX2 (paired box 2), a marker of basket/stellate cell progenitors. VHL cerebellar cortices also revealed PAX2-positive cells in Purkinje and molecular layers, resembling the histological and molecular development of basket/stellate cells in postnatal non-VHL mouse and human cerebella. These data suggest that VHL deficiency can result in the developmental arrest of basket/stellate cells in the human cerebellum and that these PAX2-positive, initiated cells await another insult or signal to form DASEs and eventually, tumors.
Asunto(s)
Neoplasias Cerebelosas , Hemangioblastoma , Enfermedad de von Hippel-Lindau , Animales , Ratones , Recién Nacido , Humanos , Hemangioblastoma/genética , Hemangioblastoma/metabolismo , Hemangioblastoma/patología , Enfermedad de von Hippel-Lindau/complicaciones , Enfermedad de von Hippel-Lindau/genética , Enfermedad de von Hippel-Lindau/metabolismo , Neoplasias Cerebelosas/genética , Neoplasias Cerebelosas/patología , Cerebelo/patología , Oligonucleótidos/metabolismo , ARN/metabolismo , Proteína Supresora de Tumores del Síndrome de Von Hippel-Lindau/genética , Proteína Supresora de Tumores del Síndrome de Von Hippel-Lindau/metabolismoRESUMEN
Von Hippel-Lindau (VHL) syndrome is a rare inherited cancer disease where the lack of VHL protein triggers the development of multisystemic tumors such us retinal hemangioblastomas (HBs), CNS-HBs, and clear cell renal cell carcinoma (ccRCC). Since standard therapies in VHL have shown limited response, leaving surgery as the only possible treatment, targeting of the ß2-adrenergic receptor (ADRB2) has shown therapeutic antitumor benefits on VHL-retinal HBs (clinical trial), VHL-CNS HBs, and VHL-ccRCC (in vitro and in vivo). In the present study, we wanted to look deep into the effects of the ADRB2 blockers propranolol and ICI-118,551 on two main aspects of cancer progression: (i) the changes on the inflammatory response of ccRCC cells; and (ii) the modulation on the Warburg effect (glycolytic metabolism), concretely, on the expression of genes involved in the cell reactive oxygen species (ROS) balance and levels. Accordingly, in vitro studies with primary VHL-ccRCC and 786-O cells measuring ROS levels, ROS-expression of detoxifying enzymes, and the expression of p65/NF-κB targets by RT-PCR were carried out. Furthermore, histological analyses of ccRCC samples from heterotopic mouse xenografts were performed. The obtained results show that ADRB2 blockade in ccRCC cells reduces the level of oxidative stress and stabilizes the inflammatory response. Thus, these data further support the idea of targeting ADRB2 as a promising strategy for the treatment of VHL and other non-VHL tumors.
Asunto(s)
Carcinoma de Células Renales/tratamiento farmacológico , Carcinoma de Células Renales/metabolismo , Inflamación/tratamiento farmacológico , Neoplasias Renales/tratamiento farmacológico , Neoplasias Renales/metabolismo , Estrés Oxidativo/efectos de los fármacos , Receptores Adrenérgicos beta 2/metabolismo , Animales , Línea Celular Tumoral , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Hemangioblastoma/tratamiento farmacológico , Hemangioblastoma/metabolismo , Humanos , Inflamación/metabolismo , Masculino , Ratones , Propanolaminas/farmacología , Propranolol/farmacología , Transducción de Señal/efectos de los fármacos , Proteína Supresora de Tumores del Síndrome de Von Hippel-Lindau/metabolismo , Enfermedad de von Hippel-Lindau/tratamiento farmacológico , Enfermedad de von Hippel-Lindau/metabolismoRESUMEN
Hypoxia-inducible factors (HIFs) activate gene transcription in response to reduced O2 availability and play critical roles in development, physiology, and disease pathogenesis. Mutations that dysregulate HIF activity are the genetic basis for tumor predisposition in the von Hippel-Lindau syndrome and excess red blood cell production in hereditary erythrocytosis.
Asunto(s)
Enfermedades Genéticas Congénitas/genética , Oxígeno/metabolismo , Policitemia/congénito , Enfermedad de von Hippel-Lindau/genética , Translocador Nuclear del Receptor de Aril Hidrocarburo/genética , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/genética , Enfermedades Genéticas Congénitas/metabolismo , Humanos , Subunidad alfa del Factor 1 Inducible por Hipoxia/genética , Mutación/genética , Policitemia/genética , Policitemia/metabolismo , Policitemia/patología , Enfermedad de von Hippel-Lindau/metabolismo , Enfermedad de von Hippel-Lindau/patologíaRESUMEN
Von Hippel-Lindau disease (VHL) is a rare hereditary disease characterized by the predisposal to develop different types of highly vascularized tumors. VHL patients carry a VHL mutation that causes partial lack of functional VHL protein (pVHL) in all cells, and a total lack thereof in cells harboring a second hit mutation. Absence of pVHL generates a prolonged state of pseudo-hypoxia in the cell due to accumulation of hypoxia inducible factor, an important transcription factor regulating pro-tumorigenic genes. The work here presented focuses on characterizing the endothelium of VHL patients, by means of blood outgrowth endothelial cells (BOECs). Transcriptome analysis of VHL-derived BOECs, further supported by in vitro assays, shows that these cells are at a disadvantage, as evidenced by loss of cell adhesion capacity, angiogenesis defects, and immune response and oxidative metabolic gene downregulation, which induce oxidative stress. These results suggest that the endothelium of VHL patients is functionally compromised and more susceptible to tumor development. These findings contribute to shedding light on the vascular landscape of VHL patients preceding the second hit mutation in the VHL gene. This knowledge could be useful in searching for new therapies for these patients and other vascular diseases.
Asunto(s)
Células Endoteliales/patología , Neovascularización Patológica , Enfermedad de von Hippel-Lindau/patología , Estudios de Casos y Controles , Adhesión Celular , Movimiento Celular , Proliferación Celular , Células Cultivadas , Células Endoteliales/inmunología , Células Endoteliales/metabolismo , Regulación de la Expresión Génica , Predisposición Genética a la Enfermedad , Humanos , Mutación , Neovascularización Patológica/genética , Estrés Oxidativo , Fenotipo , Transducción de Señal , Transcriptoma , Proteína Supresora de Tumores del Síndrome de Von Hippel-Lindau/genética , Enfermedad de von Hippel-Lindau/genética , Enfermedad de von Hippel-Lindau/inmunología , Enfermedad de von Hippel-Lindau/metabolismoRESUMEN
Von Hippel-Lindau (VHL) disease is characterized by frequent mutation of VHL protein, a tumor suppressor that functions as the substrate recognition subunit of a Cullin2 RING E3 ligase complex (CRL2VHL). CRL2VHL plays important roles in oxygen sensing by targeting hypoxia-inducible factor-alpha (HIF-α) subunits for ubiquitination and degradation. VHL is also commonly hijacked by bifunctional molecules such as proteolysis-targeting chimeras to induce degradation of target molecules. We previously reported the design and characterization of VHL inhibitors VH032 and VH298 that block the VHL:HIF-α interaction, activate the HIF transcription factor, and induce a hypoxic response, which can be beneficial to treat anemia and mitochondrial diseases. How these compounds affect the global cellular proteome remains unknown. Here, we use unbiased quantitative MS to identify the proteomic changes elicited by the VHL inhibitor compared with hypoxia or the broad-spectrum prolyl-hydroxylase domain enzyme inhibitor IOX2. Our results demonstrate that VHL inhibitors selectively activate the HIF response similar to the changes induced in hypoxia and IOX2 treatment. Interestingly, VHL inhibitors were found to specifically upregulate VHL itself. Our analysis revealed that this occurs via protein stabilization of VHL isoforms and not via changes in transcript levels. Increased VHL levels upon VH298 treatment resulted in turn in reduced levels of HIF-1α protein. This work demonstrates the specificity of VHL inhibitors and reveals different antagonistic effects upon their acute versus prolonged treatment in cells. These findings suggest that therapeutic use of VHL inhibitors may not produce overt side effects from HIF stabilization as previously thought.
Asunto(s)
Ciclopropanos/farmacología , Subunidad alfa del Factor 1 Inducible por Hipoxia/antagonistas & inhibidores , Hipoxia/metabolismo , Proteómica/métodos , Pirrolidinas/farmacología , Tiazoles/farmacología , Proteína Supresora de Tumores del Síndrome de Von Hippel-Lindau/antagonistas & inhibidores , Enfermedad de von Hippel-Lindau/patología , Línea Celular , Inhibidores Enzimáticos/farmacología , Humanos , Subunidad alfa del Factor 1 Inducible por Hipoxia/metabolismo , Unión Proteica , Estabilidad Proteica , Proteína Supresora de Tumores del Síndrome de Von Hippel-Lindau/metabolismo , Enfermedad de von Hippel-Lindau/genética , Enfermedad de von Hippel-Lindau/metabolismoRESUMEN
Hypoxia-inducible factors (HIFs) activate gene transcription in response to reduced O2 availability and play critical roles in development, physiology, and disease pathogenesis. Mutations that dysregulate HIF activity are the genetic basis for tumor predisposition in the von Hippel-Lindau syndrome and excess red blood cell production in hereditary erythrocytosis.
Asunto(s)
Estudios de Asociación Genética , Enfermedades Genéticas Congénitas/genética , Enfermedades Genéticas Congénitas/metabolismo , Predisposición Genética a la Enfermedad , Oxígeno/metabolismo , Fenotipo , Biomarcadores , Diagnóstico Diferencial , Enfermedades Genéticas Congénitas/diagnóstico , Humanos , Subunidad alfa del Factor 1 Inducible por Hipoxia/genética , Subunidad alfa del Factor 1 Inducible por Hipoxia/metabolismo , Policitemia/congénito , Policitemia/diagnóstico , Policitemia/metabolismo , Transducción de Señal , Enfermedad de von Hippel-Lindau/diagnóstico , Enfermedad de von Hippel-Lindau/genética , Enfermedad de von Hippel-Lindau/metabolismoRESUMEN
INTRODUCTION: Central nervous system hemangioblastoma is a benign tumor associated with or without von Hippel-Lindau (VHL) disease which is an autosomal dominant hereditary disease that results from a germline mutation in the VHL gene. A main axis of signaling pathways in central nervous system hemangioblastoma is VHL-HIF signaling pathway. Here, we propose an alternative VHL-JAK-STAT signaling pathway in hemangioblastoma and discuss the role. METHODS: Using MACS method, Scl+ hemangioblast-like cells were isolated from multipotent nestin-expressing stem cells. Then, ubiquitination of JAK2 in those cells and immunoprecipitation between JAK2 and VHL were examined. Then, expressions of JAK2 and STAT3 in those cells and expressions of VHL-associated hemangioblastoma tissues were examined. In addition, the VHL genes of patients bearing hemangioblastoma were analyzed. RESULTS: JAK2 and STAT3 in Scl+ hemangioblast-like cells were ubiquitinated after VHL- expression vector was transferred to those cells. Expressions of JAK2 and STAT3 in those cells were well recognized before the transfer, but those disappeared after the transfer. Expressions of both JAK2 and STAT3 in hemangioblastoma tissues were well shown. The VHL gene analysis revealed that patients bearing hemangioblastoma carried missense mutations in 5, small deletions in 2, large deletions in 4, and nonsense mutation in 1 CONCLUSIONS: VHL-JAK-STAT signaling pathway might play an important role in proliferation, angiogenesis, and maintenance of stem-cell-nature in hemangioblastoma as an alternative signaling pathway to supplement VHL-HIF signaling pathway.
Asunto(s)
Neoplasias Cerebelosas/metabolismo , Hemangioblastoma/metabolismo , Transducción de Señal , Enfermedad de von Hippel-Lindau/metabolismo , Adulto , Neoplasias Cerebelosas/complicaciones , Neoplasias Cerebelosas/patología , Femenino , Hemangioblastoma/complicaciones , Hemangioblastoma/patología , Humanos , Janus Quinasa 2/metabolismo , Mutación , Factor de Transcripción STAT3/metabolismo , Proteína Supresora de Tumores del Síndrome de Von Hippel-Lindau/metabolismo , Adulto Joven , Enfermedad de von Hippel-Lindau/complicaciones , Enfermedad de von Hippel-Lindau/patologíaRESUMEN
Genome-wide CRISPR-Cas9 essentiality screening represents a powerful approach to identify genetic vulnerabilities in cancer cells. Here, we applied this technology and designed a strategy to identify target genes that are synthetic lethal (SL) with von Hippel-Lindau (VHL) tumor suppressor gene. Inactivation of VHL has been frequently found in clear cell renal cell carcinoma. Its SL partners serve as potential drug targets for the development of targeted cancer therapies. We performed parallel genome-wide CRISPR screens in two pairs of isogenic clear cell renal cell carcinoma cell lines that differ only in the VHL status. Comparative analyses of screening results not only confirmed a well-known role for mTOR signaling in renal carcinoma, but also identified DNA damage response and selenocysteine biosynthesis pathways as novel SL targets in VHL-inactivated cancer cells. Follow-up studies provided cellular and mechanistic insights into SL interactions of these pathway genes with the VHL gene. Our CRISPR and RNA-seq datasets provide a rich resource for future investigation of the function of the VHL tumor suppressor protein. Our work demonstrates the efficiency of CRISPR-based synthetic lethality screening in human isogenic cell pairs. Similar strategies could be employed to unveil SL partners with other oncogenic drivers.
Asunto(s)
Reparación del ADN , Selenocisteína/biosíntesis , Transducción de Señal , Proteína Supresora de Tumores del Síndrome de Von Hippel-Lindau/metabolismo , Sistemas CRISPR-Cas , Carcinoma de Células Renales/genética , Carcinoma de Células Renales/metabolismo , Línea Celular Tumoral , Daño del ADN , Edición Génica , Humanos , Neoplasias Renales/genética , Neoplasias Renales/metabolismo , Análisis de Secuencia de ARN , Proteína Supresora de Tumores del Síndrome de Von Hippel-Lindau/genética , Enfermedad de von Hippel-Lindau/genética , Enfermedad de von Hippel-Lindau/metabolismoRESUMEN
One of the major consequences of the lack of a functional VHL protein in von Hippel-Lindau disease, a rare cancer, is the constitutive activation of the HIF pathway. This activation ends up in the generation of Central Nervous System (CNS) Hemangioblastomas among other tumours along the lifespan of the patient. Nowadays, only surgery has been proven efficient as therapy since the systemic attempts have failed. Propranolol, a non-specific ß1-and ß2-adrenergic receptor antagonist, was recently designated as the first therapeutic (orphan) drug for VHL disease. Nevertheless, its ß1 affinity provokes the decrease in blood pressure, being not recommended for low or regular blood pressure VHL patients. In order to overcome the ß1-drawback, the properties of a high specific ß2-adrenergic receptor blocker named ICI-118,551 have been studied. ICI-118,551 was able to decrease Hemangioblastomas cell viability in a specific manner, by triggering apoptosis. Moreover, ICI-118,551 also impaired the nuclear internalization of HIF-1α in Hemangioblastomas and hypoxic primary endothelial cells, reducing significantly the activation of HIF-target genes and halting the tumour-related angiogenic processes. In this work, we demonstrate the therapeutical properties of ICI-118,551 in VHL-derived CNS-Hemangioblastoma primary cultures, becoming a promising drug for VHL disease and other HIF-related diseases.
Asunto(s)
Antagonistas Adrenérgicos beta/farmacología , Núcleo Celular/metabolismo , Neoplasias del Sistema Nervioso Central/metabolismo , Hemangioblastoma/metabolismo , Subunidad alfa del Factor 1 Inducible por Hipoxia/metabolismo , Propanolaminas/farmacología , Enfermedad de von Hippel-Lindau/metabolismo , Apoptosis , Neoplasias del Sistema Nervioso Central/complicaciones , Hemangioblastoma/complicaciones , Humanos , Terapia Molecular Dirigida , Mutación/genética , Neovascularización Patológica , Transducción de Señal , Células Tumorales Cultivadas , Proteína Supresora de Tumores del Síndrome de Von Hippel-Lindau/genética , Enfermedad de von Hippel-Lindau/complicacionesRESUMEN
Von Hippel-Lindau (VHL) syndrome is a familial cancer syndrome caused by mutations in the tumor suppressor gene VHL. We generated human iPSC lines from primary dermal fibroblasts of three VHL syndrome patients carrying distinct VHL germ line mutations (c.194C>G, c.194C>T and nt440delTCT, respectively). Characterization of the iPSC lines confirmed expression of pluripotency markers, trilineage differentiation potential and absence of exogenous vector expression. The three hiPSC lines were genetically stable and retained the VHL mutation of each donor. These iPSC lines, the first derived from VHL syndrome patients, offer a useful resource to study disease pathophysiology and for anti-cancer drug development.
Asunto(s)
Mutación de Línea Germinal , Células Madre Pluripotentes Inducidas , Proteína Supresora de Tumores del Síndrome de Von Hippel-Lindau , Enfermedad de von Hippel-Lindau , Línea Celular , Femenino , Humanos , Células Madre Pluripotentes Inducidas/metabolismo , Células Madre Pluripotentes Inducidas/patología , Proteína Supresora de Tumores del Síndrome de Von Hippel-Lindau/genética , Proteína Supresora de Tumores del Síndrome de Von Hippel-Lindau/metabolismo , Enfermedad de von Hippel-Lindau/genética , Enfermedad de von Hippel-Lindau/metabolismo , Enfermedad de von Hippel-Lindau/patologíaRESUMEN
Intracellular proteins continuously turn over by degradation and synthesis in all organ tissues. Owing to its irreversible nature, protein degradation is a highly selective process to avoid irreparable breakdown of cellular constituents, thereby disrupting cellular stability, integrity and signalling. The majority of intracellular proteins are degraded by the ubiquitin-proteasome system (UPS), a multi-enzyme process that involves the covalent conjugation of ubiquitin to a substrate protein and its recognition and degradation by the core multicomponent proteolytic complex of the UPS, the proteasome. In addition to labelling misfolded, damaged, aggregation-prone and intact but unneeded proteins for proteasomal degradation, ubiquitylation regulates a multitude of cellular processes, such as transcription, translation, endocytosis, and receptor activity and subcellular localization. In addition, the proteasome generates peptides for antigen presentation in the immune system and for further degradation by peptidases to provide amino acids for protein biosynthesis and gluconeogenesis. Alterations of the UPS or of protein substrates that render them more or less susceptible to degradation are responsible for disorders associated with renal cell dysfunction. In this Review, we provide insight into the elegant and complex nature of UPS-mediated proteostasis and focus on its established and potential roles in renal cell physiology and pathophysiology.
Asunto(s)
Enfermedades Renales/fisiopatología , Complejo de la Endopetidasa Proteasomal/metabolismo , Proteínas Ubiquitinadas/metabolismo , Ubiquitinación/fisiología , Autofagia/fisiología , Transporte Biológico/fisiología , Eritropoyetina/metabolismo , Glucosa/metabolismo , Homeostasis/fisiología , Humanos , Riñón/fisiología , Enfermedades Renales/metabolismo , Lisosomas/metabolismo , Atrofia Muscular/fisiopatología , Podocitos/metabolismo , Inhibidores de Proteasoma/farmacología , Sodio/metabolismo , Equilibrio Hidroelectrolítico/fisiología , Enfermedad de von Hippel-Lindau/metabolismo , Enfermedad de von Hippel-Lindau/fisiopatologíaRESUMEN
Familiar cancers represent a privileged point of view for studying the complex cellular events inducing tumor transformation. Von Hippel-Lindau syndrome, a familiar predisposition to develop cancer is a clear example. Here, we present our efforts to decipher the role of von Hippel-Lindau tumor suppressor protein (pVHL) in cancer insurgence. We collected high quality information about both pVHL mutations and interactors to investigate the association between patient phenotypes, mutated protein surface and impaired interactions. Our data suggest that different phenotypes correlate with localized perturbations of the pVHL structure, with specific cell functions associated to different protein surfaces. We propose five different pVHL interfaces to be selectively involved in modulating proteins regulating gene expression, protein homeostasis as well as to address extracellular matrix (ECM) and ciliogenesis associated functions. These data were used to drive molecular docking of pVHL with its interactors and guide Petri net simulations of the most promising alterations. We predict that disruption of pVHL association with certain interactors can trigger tumor transformation, inducing metabolism imbalance and ECM remodeling. Collectively taken, our findings provide novel insights into VHL-associated tumorigenesis. This highly integrated in silico approach may help elucidate novel treatment paradigms for VHL disease.
Asunto(s)
Mutación , Proteína Supresora de Tumores del Síndrome de Von Hippel-Lindau/genética , Carcinoma de Células Renales/genética , Biología Computacional , Genes Supresores de Tumor , Estudios de Asociación Genética , Predisposición Genética a la Enfermedad , Humanos , Neoplasias Renales/genética , Simulación del Acoplamiento Molecular , Policitemia/genética , Dominios y Motivos de Interacción de Proteínas , Mapas de Interacción de Proteínas , Procesamiento Proteico-Postraduccional , Proteína Supresora de Tumores del Síndrome de Von Hippel-Lindau/química , Proteína Supresora de Tumores del Síndrome de Von Hippel-Lindau/metabolismo , Enfermedad de von Hippel-Lindau/genética , Enfermedad de von Hippel-Lindau/metabolismoRESUMEN
Renal cell carcinoma (RCC) is the major cause of death among patients with von Hippel-Lindau (VHL) disease. Resistance to therapies targeting tumor angiogenesis opens the question about the underlying mechanisms. Previously we have described that RWDD3 or RSUME (RWD domain-containing protein SUMO Enhancer) sumoylates and binds VHL protein and negatively regulates HIF degradation, leading to xenograft RCC tumor growth in mice. In this study, we performed a bioinformatics analysis in a ccRCC dataset showing an association of RSUME levels with VHL mutations and tumor progression, and we demonstrate the molecular mechanism by which RSUME regulates the pathologic angiogenic phenotype of VHL missense mutations. We report that VHL mutants fail to downregulate RSUME protein levels accounting for the increased RSUME expression found in RCC tumors. Furthermore, we prove that targeting RSUME in RCC cell line clones carrying missense VHL mutants results in decreased early tumor angiogenesis. The mechanism we describe is that RSUME sumoylates VHL mutants and beyond its sumoylation capacity, interacts with Type 2 VHL mutants, reduces HIF-2α-VHL mutants binding, and negatively regulates the assembly of the Type 2 VHL, Elongins and Cullins (ECV) complex. Altogether these results show RSUME involvement in VHL mutants deregulation that leads to the angiogenic phenotype of RCC tumors.
Asunto(s)
Carcinoma de Células Renales/genética , Neoplasias Renales/genética , Factores de Transcripción/metabolismo , Proteína Supresora de Tumores del Síndrome de Von Hippel-Lindau/genética , Enfermedad de von Hippel-Lindau/genética , Animales , Células COS , Carcinoma de Células Renales/metabolismo , Carcinoma de Células Renales/mortalidad , Línea Celular Tumoral , Chlorocebus aethiops , Medios de Cultivo Condicionados , Elonguina/genética , Elonguina/metabolismo , Regulación Neoplásica de la Expresión Génica , Células Endoteliales de la Vena Umbilical Humana , Humanos , Neoplasias Renales/metabolismo , Neoplasias Renales/mortalidad , Masculino , Ratones , Ratones Endogámicos NOD , Ratones SCID , Mutación Missense , Neovascularización Patológica/genética , Neovascularización Patológica/metabolismo , Sumoilación , Factores de Transcripción/genética , Proteína Supresora de Tumores del Síndrome de Von Hippel-Lindau/metabolismo , Enfermedad de von Hippel-Lindau/complicaciones , Enfermedad de von Hippel-Lindau/metabolismoRESUMEN
Autophagy is a highly conserved, homeostatic process by which cytosolic components reach lysosomes for degradation. The roles played by different autophagic processes in cancer are complex and remain cancer type and stage dependent. Renal cell carcinoma (RCC) is the most common subtype of kidney cancer and is characterized by the inactivation of the von Hippel-Lindau (VHL) tumor suppressor. Our previous study identified a small compound, STF-62247, as an autophagy-modulating molecule causing selective cytotoxicity for VHL-inactivated cells. This present study investigates the effects of STF-62247 specifically on the macroautophagic flux to better characterize its mechanism of action in RCC. Our results clearly demonstrate that this compound is a potent blocker of late stages of autophagy. We show that inhibiting autophagy by CRISPR knockouts of autophagy-related genes rendered VHL-deficient cells insensitive to STF-62247, uncovering the importance of the autophagic pathway in STF-selective cell death. By exploiting the autofluorescence of STF-62247, we pinpointed its cellular localization to lysosomes. Finally, in response to prolonged STF treatments, we show that VHL-proficient cells are able to surmount the block in late stages of autophagy by restoring their lysosome numbers. Conversely, an increase in autophagic vesicles accompanied by a time-dependent decrease in lysosomes was observed in VHL-deficient cells. This is the first mechanistic study investigating STF-62447's effects on the autophagic flux in RCC. Importantly, our study reclassifies STF-62247 as a blocker of later stages of autophagy and highlights the possibility of blocking this process through lysosome disruption in VHL-mutated RCCs.
Asunto(s)
Autofagia/fisiología , Carcinoma de Células Renales/metabolismo , Neoplasias Renales/metabolismo , Lisosomas/metabolismo , Piridinas/metabolismo , Tiazoles/metabolismo , Enfermedad de von Hippel-Lindau/metabolismo , Autofagia/efectos de los fármacos , Carcinoma de Células Renales/tratamiento farmacológico , Relación Dosis-Respuesta a Droga , Sistemas de Liberación de Medicamentos/métodos , Células HeLa , Humanos , Neoplasias Renales/tratamiento farmacológico , Lisosomas/efectos de los fármacos , Piridinas/administración & dosificación , Tiazoles/administración & dosificación , Enfermedad de von Hippel-Lindau/tratamiento farmacológicoRESUMEN
We present a rare case of a 42-year-old man with a long history of von Hippel-Lindau disease that was scanned with Ga-DOTA-TOC PET/CT for suspicion of disease relapse. Ga-DOTA-TOC imaging demonstrated increased DOTA-TOC uptake in pancreas tail and intramedullary and extramedullary spinal hemangioblastomas, only some of which have already been highlighted at MRI examination. This case illustrates the significant role the Ga-labeled somatostatin receptor analogs PET/CT in the management of the von Hippel-Lindau disease.
Asunto(s)
Octreótido/análogos & derivados , Compuestos Organometálicos , Tomografía Computarizada por Tomografía de Emisión de Positrones , Enfermedad de von Hippel-Lindau/diagnóstico por imagen , Adulto , Humanos , Imagen por Resonancia Magnética , Masculino , Receptores de Somatostatina/metabolismo , Enfermedad de von Hippel-Lindau/metabolismo , Enfermedad de von Hippel-Lindau/patología , Enfermedad de von Hippel-Lindau/terapiaRESUMEN
The von Hippel-Lindau (VHL) syndrome is a rare inherited cancer, caused by mutations in the VHL gene, many of which render the VHL protein (pVHL) unstable. pVHL is a tumor-suppressor protein implicated in a variety of cellular processes, most notably in response to changes in oxygen availability, due to its role as part of an E3-ligase complex which targets the hypoxia-inducible factor (HIF) for degradation. Previously we reported, using in silico and in vitro analyses, that common oncogenic VHL mutations render pVHL less stable than the wild-type protein, distort its core domain and as a result reduce the ability of the protein to bind its target HIF-1α. Among various chemical chaperones tested, arginine was the most effective in refolding mutant of pVHL. Here we examined the consequences of administering L- or D-arginine to a Drosophila VHL model and to human renal carcinoma cells, both expressing misfolded versions of human pVHL. Arginine treatment increased pVHL solubility in both models and increased the half-life of the mutant pVHL proteins in the cell culture. In both models, L- as well as D-arginine enhanced the ability of wild-type pVHL and certain misfolded mutant versions of pVHL to bind ODD, the HIF-derived target peptide, reflecting restoration of pVHL function. Moreover, continuous feeding of Drosophila expressing misfolded versions of pVHL either L- or D-arginine rich diet rescued their lethal phenotype. Collectively, these in vivo results suggest that arginine supplementation should be examined as a potential novel treatment for VHL cancer syndrome.
Asunto(s)
Proteínas de Drosophila/metabolismo , Neoplasias Experimentales/metabolismo , Enfermedad de von Hippel-Lindau/metabolismo , Animales , Carcinoma de Células Renales/genética , Carcinoma de Células Renales/metabolismo , Carcinoma de Células Renales/patología , Proteínas de Drosophila/genética , Drosophila melanogaster , Humanos , Neoplasias Renales/genética , Neoplasias Renales/metabolismo , Neoplasias Renales/patología , Neoplasias Experimentales/genética , Neoplasias Experimentales/patología , Proteína Supresora de Tumores del Síndrome de Von Hippel-Lindau/genética , Proteína Supresora de Tumores del Síndrome de Von Hippel-Lindau/metabolismo , Enfermedad de von Hippel-Lindau/genética , Enfermedad de von Hippel-Lindau/patologíaRESUMEN
The inactivation of the von Hippel-Lindau (VHL) tumor suppressor gene plays a crucial role in the development of hemangioblastomas (HBs) within the human central nervous system (CNS). However, both the cytological origin and the evolutionary process of HBs (including neovascularization) remain controversial, and anti-angiogenesis for VHL-HBs, based on classic HB angiogenesis, have produced disappointing results in clinical trials. One major obstacle to the successful clinical translation of anti-vascular treatment is the lack of a thorough understanding of neovascularization in this vascular tumor. In this article, we present a comprehensive procedure to evaluate in vitro whether classic tumor angiogenesis exists in HBs, as well as its role in HBs. With this procedure, researchers can accurately understand the complexity of HB neovascularization and identify the function of this common form of angiogenesis in HBs. These protocols can be used to evaluate the most promising anti-vascular therapy for tumors, which has high translational potential either for tumors treatment or for aiding in the optimization of the anti-angiogenic treatment for HBs in future translations. The results highlight the complexity of HB neovascularization and suggest that this common form angiogenesis is only a complementary mechanism in HB neovascularization.
Asunto(s)
Hemangioblastoma/genética , Neovascularización Patológica/genética , Esferoides Celulares/metabolismo , Proteína Supresora de Tumores del Síndrome de Von Hippel-Lindau/genética , Enfermedad de von Hippel-Lindau/genética , Femenino , Hemangioblastoma/metabolismo , Hemangioblastoma/patología , Humanos , Masculino , Neovascularización Patológica/metabolismo , Enfermedad de von Hippel-Lindau/metabolismoAsunto(s)
Sistema Nervioso Central/metabolismo , Fibronectinas/metabolismo , Hemangioblastoma/metabolismo , Proteína Supresora de Tumores del Síndrome de Von Hippel-Lindau/metabolismo , Enfermedad de von Hippel-Lindau/metabolismo , Matriz Extracelular/metabolismo , Fibronectinas/genética , Humanos , Riñón/metabolismo , Mutación , Proteína Supresora de Tumores del Síndrome de Von Hippel-Lindau/genéticaRESUMEN
Von Hippel-Lindau disease (vHL, familial cerebello-retinal angiomatosis) is a rare genetic autosomal dominant disorder associated with predisposition to vascular tumors. Mutations of VHL tumor suppressor gene, located on chromosome 3p25-26, are responsible for clinical manifestation of the disease. The VHL gene product encodes VHL protein, which is responsible for HIF-1 (hypoxia-inducible factor-1) dependent cell cycle regulation and cellular pathways mediated by VEGF, PDGF, TGF-α, EPO. The mechanism substantiates the hypoxia dependent vascular tumor growth caused by loss of wild-type VHL protein. The clinical spectrum of vHL syndrome includes multiple tumors of various localization and low histologic grade, often bilateral. The most typical for the syndrome are: hemangioblastoma of central nervous system (typically posterior fossa or medulla), retinal hemangioblastoma, renal cell carcinoma and pheochromocytoma. The aim of the case report is to remind the typical clinical manifestation of von Hippel- Lindau syndrome, update the diagnostic criteria, recommended diagnostic and follow up methods.