RÉSUMÉ
The histone 3 lysine 79 (H3K79) methyltransferase (HMT) DOT1L is known to play a critical role for growth and survival of MLL-rearranged leukemia. Serendipitous observations during high-throughput drug screens indicated that the use of DOT1L inhibitors might be expandable to multiple myeloma (MM). Through pharmacologic and genetic experiments, we could validate that DOT1L is essential for growth and viability of a subset of MM cell lines, in line with a recent report from another team. In vivo activity against established MM xenografts was observed with a novel DOT1L inhibitor. In order to understand the molecular mechanism of the dependency in MM, we examined gene expression changes upon DOT1L inhibition in sensitive and insensitive cell lines and discovered that genes belonging to the endoplasmic reticulum (ER) stress pathway and protein synthesis machinery were specifically suppressed in sensitive cells. Whole-genome CRISPR screens in the presence or absence of a DOT1L inhibitor revealed that concomitant targeting of the H3K4me3 methyltransferase SETD1B increases the effect of DOT1L inhibition. Our results provide a strong basis for further investigating DOT1L and SETD1B as targets in MM.
RÉSUMÉ
Aberrant activation of the JAK/STAT pathway is thought to be the critical event in the pathogenesis of the chronic myeloproliferative neoplasms, polycythemia vera, essential thrombocythemia and primary myelofibrosis. The most frequent genetic alteration in these pathologies is the activating JAK2V617F mutation, and expression of the mutant gene in mouse models was shown to cause a phenotype resembling the human diseases. Given the body of genetic evidence, it has come as a sobering finding that JAK inhibitor therapy only modestly suppresses the JAK2V617F allele burden, despite showing clear benefits in terms of reducing splenomegaly and constitutional symptoms in patients. To gain a better understanding if JAK2V617F is required for maintenance of myeloproliferative disease once it has evolved, we generated a conditional inducible transgenic JAK2V617F mouse model using the SCL-tTA-2S tet-off system. Our model corroborates that expression of JAK2V617F in hematopoietic stem and progenitor cells recapitulates key hallmarks of human myeloproliferative neoplasms, and exhibits gender differences in disease manifestation. The disease was found to be transplantable, and importantly, reversible when transgenic JAK2V617F expression was switched off. Our results indicate that mutant JAK2V617F-specific inhibitors should result in profound disease modification by disabling the myeloproliferative clone bearing mutant JAK2.
Sujet(s)
Régulation de l'expression des gènes , Cellules souches hématopoïétiques , Kinase Janus-2 , Syndromes myéloprolifératifs , Transgènes , Substitution d'acide aminé , Animaux , Modèles animaux de maladie humaine , Cellules souches hématopoïétiques/métabolisme , Cellules souches hématopoïétiques/anatomopathologie , Humains , Kinase Janus-2/biosynthèse , Kinase Janus-2/génétique , Souris , Souris transgéniques , Mutation faux-sens , Syndromes myéloprolifératifs/génétique , Syndromes myéloprolifératifs/métabolisme , Syndromes myéloprolifératifs/anatomopathologieRÉSUMÉ
More than 90% of drugs with preclinical activity fail in human trials, largely due to insufficient efficacy. We hypothesized that adequately powered trials of patient-derived xenografts (PDX) in mice could efficiently define therapeutic activity across heterogeneous tumors. To address this hypothesis, we established a large, publicly available repository of well-characterized leukemia and lymphoma PDXs that undergo orthotopic engraftment, called the Public Repository of Xenografts (PRoXe). PRoXe includes all de-identified information relevant to the primary specimens and the PDXs derived from them. Using this repository, we demonstrate that large studies of acute leukemia PDXs that mimic human randomized clinical trials can characterize drug efficacy and generate transcriptional, functional, and proteomic biomarkers in both treatment-naive and relapsed/refractory disease.
Sujet(s)
Hétérogreffes , Leucémies/anatomopathologie , Lymphomes/anatomopathologie , Banques de tissus , Tests d'activité antitumorale sur modèle de xénogreffe , Animaux , Antinéoplasiques/pharmacologie , Antinéoplasiques/usage thérapeutique , Marqueurs biologiques tumoraux , Lignage cellulaire , Femelle , Analyse de profil d'expression de gènes , Gènes p53 , Humains , Internet , Isoquinoléines/pharmacologie , Isoquinoléines/usage thérapeutique , Leucémies/métabolisme , Leucémie expérimentale/traitement médicamenteux , Lymphomes/métabolisme , Mâle , Souris , Souris de lignée NOD , Thérapie moléculaire ciblée , Protéines tumorales/antagonistes et inhibiteurs , Transplantation tumorale , Phénotype , Pipérazines/pharmacologie , Pipérazines/usage thérapeutique , Leucémie-lymphome lymphoblastique à précurseurs B/traitement médicamenteux , Leucémie-lymphome lymphoblastique à précurseurs B/génétique , Leucémie-lymphome lymphoblastique à précurseurs B/anatomopathologie , Protéome , Protéines proto-oncogènes c-mdm2/antagonistes et inhibiteurs , Répartition aléatoire , Essais contrôlés randomisés comme sujet/méthodes , Plan de recherche , TranscriptomeRÉSUMÉ
The tumor suppressor p53 is a key regulator of apoptosis and functions upstream in the apoptotic cascade by both indirectly and directly regulating Bcl-2 family proteins. In cells expressing wild-type (WT) p53, the HDM2 protein binds to p53 and blocks its activity. Inhibition of HDM2:p53 interaction activates p53 and causes apoptosis or cell-cycle arrest. Here, we investigated the ability of the novel HDM2 inhibitor CGM097 to potently and selectively kill WT p53-expressing AML cells. The antileukemic effects of CGM097 were studied using cell-based proliferation assays (human AML cell lines, primary AML patient cells, and normal bone marrow samples), apoptosis, and cell-cycle assays, ELISA, immunoblotting, and an AML patient-derived in vivo mouse model. CGM097 potently and selectively inhibited the proliferation of human AML cell lines and the majority of primary AML cells expressing WT p53, but not mutant p53, in a target-specific manner. Several patient samples that harbored mutant p53 were comparatively unresponsive to CGM097. Synergy was observed when CGM097 was combined with FLT3 inhibition against oncogenic FLT3-expressing cells cultured both in the absence as well as the presence of cytoprotective stromal-secreted cytokines, as well as when combined with MEK inhibition in cells with activated MAPK signaling. Finally, CGM097 was effective in reducing leukemia burden in vivo. These data suggest that CGM097 is a promising treatment for AML characterized as harboring WT p53 as a single agent, as well as in combination with other therapies targeting oncogene-activated pathways that drive AML.
Sujet(s)
Antinéoplasiques/pharmacologie , Isoquinoléines/pharmacologie , Leucémie aigüe myéloïde/traitement médicamenteux , Pipérazines/pharmacologie , Protéines proto-oncogènes c-mdm2/antagonistes et inhibiteurs , Protéine p53 suppresseur de tumeur/métabolisme , Animaux , Apoptose/effets des médicaments et des substances chimiques , Benzothiazoles/pharmacologie , Lignée cellulaire tumorale , Prolifération cellulaire/effets des médicaments et des substances chimiques , Synergie des médicaments , Femelle , Expression des gènes , Humains , Concentration inhibitrice 50 , Souris de lignée NOD , Souris SCID , Phénylurées/pharmacologie , Staurosporine/analogues et dérivés , Staurosporine/pharmacologie , Charge tumorale/effets des médicaments et des substances chimiques , Cellules cancéreuses en culture , Protéine p53 suppresseur de tumeur/génétique , Tests d'activité antitumorale sur modèle de xénogreffeRÉSUMÉ
Biomarkers for patient selection are essential for the successful and rapid development of emerging targeted anti-cancer therapeutics. In this study, we report the discovery of a novel patient selection strategy for the p53-HDM2 inhibitor NVP-CGM097, currently under evaluation in clinical trials. By intersecting high-throughput cell line sensitivity data with genomic data, we have identified a gene expression signature consisting of 13 up-regulated genes that predicts for sensitivity to NVP-CGM097 in both cell lines and in patient-derived tumor xenograft models. Interestingly, these 13 genes are known p53 downstream target genes, suggesting that the identified gene signature reflects the presence of at least a partially activated p53 pathway in NVP-CGM097-sensitive tumors. Together, our findings provide evidence for the use of this newly identified predictive gene signature to refine the selection of patients with wild-type p53 tumors and increase the likelihood of response to treatment with p53-HDM2 inhibitors, such as NVP-CGM097.
Sujet(s)
Marqueurs biologiques/métabolisme , Régulation de l'expression des gènes tumoraux/effets des médicaments et des substances chimiques , Isoquinoléines/pharmacologie , Tumeurs/traitement médicamenteux , Sélection de patients , Pipérazines/pharmacologie , Protéines proto-oncogènes c-mdm2/antagonistes et inhibiteurs , Protéine p53 suppresseur de tumeur/génétique , Lignée cellulaire tumorale , Transfert d'énergie par résonance de fluorescence , Analyse de profil d'expression de gènes , Humains , Séquençage par oligonucléotides en batterie , Protéines proto-oncogènes c-mdm2/métabolisme , Protéine p53 suppresseur de tumeur/métabolismeRÉSUMÉ
Malignant rhabdoid tumors (MRTs) are aggressive pediatric cancers arising in brain, kidney and soft tissues, which are characterized by loss of the tumor suppressor SNF5/SMARCB1. MRTs are poorly responsive to chemotherapy and thus a high unmet clinical need exists for novel therapies for MRT patients. SNF5 is a core subunit of the SWI/SNF chromatin remodeling complex which affects gene expression by nucleosome remodeling. Here, we report that loss of SNF5 function correlates with increased expression of fibroblast growth factor receptors (FGFRs) in MRT cell lines and primary tumors and that re-expression of SNF5 in MRT cells causes a marked repression of FGFR expression. Conversely, siRNA-mediated impairment of SWI/SNF function leads to elevated levels of FGFR2 in human fibroblasts. In vivo, treatment with NVP-BGJ398, a selective FGFR inhibitor, blocks progression of a murine MRT model. Hence, we identify FGFR signaling as an aberrantly activated oncogenic pathway in MRTs and propose pharmacological inhibition of FGFRs as a potential novel clinical therapy for MRTs.
Sujet(s)
Protéines chromosomiques nonhistones/métabolisme , Protéines de liaison à l'ADN/métabolisme , Récepteur facteur croissance fibroblaste/métabolisme , Tumeur rhabdoïde/métabolisme , Facteurs de transcription/métabolisme , Animaux , Carcinogenèse/génétique , Carcinogenèse/métabolisme , Lignée cellulaire tumorale , Prolifération cellulaire , Assemblage et désassemblage de la chromatine/génétique , Protéines chromosomiques nonhistones/génétique , Protéines de liaison à l'ADN/génétique , Fibroblastes/métabolisme , Cellules HEK293 , Humains , Souris , Régions promotrices (génétique)/génétique , Récepteur facteur croissance fibroblaste/génétique , Tumeur rhabdoïde/génétique , Protéine SMARCB1 , Transduction du signal/génétique , Facteurs de transcription/génétiqueRÉSUMÉ
The Hippo (Hpo) pathway is a novel signaling pathway that controls organ size in Drosophila and mammals and is deregulated in a variety of human cancers. It consists of a set of kinases that, through a number of phosphorylation events, inactivate YAP, a transcriptional co-activator that controls cellular proliferation and apoptosis. We have identified PTPN14 as a YAP-binding protein that negatively regulates YAP activity by controlling its localization. Mechanistically, we find that the interaction of ectopic YAP with PTPN14 can be mediated by the respective WW and PPxY motifs. However, the PTPN14 PPxY motif and phosphatase activity appear to be dispensable for the negative regulation of endogenous YAP, likely suggesting more complex mechanisms of interaction and modulation. Finally, we demonstrate that PTPN14 downregulation can phenocopy YAP activation in mammary epithelial cells and synergize with YAP to induce oncogenic transformation.
Sujet(s)
Phosphoprotéines/métabolisme , Protein Tyrosine Phosphatases, Non-Receptor/métabolisme , Lignée cellulaire , Cellules épithéliales/cytologie , Cellules épithéliales/métabolisme , Humains , Phosphoprotéines/génétique , Liaison aux protéines , Protein Tyrosine Phosphatases, Non-Receptor/génétique , Transduction du signal/physiologieRÉSUMÉ
UNLABELLED: Patient stratification biomarkers that enable the translation of cancer genetic knowledge into clinical use are essential for the successful and rapid development of emerging targeted anticancer therapeutics. Here, we describe the identification of patient stratification biomarkers for NVP-BGJ398, a novel and selective fibroblast growth factor receptor (FGFR) inhibitor. By intersecting genome-wide gene expression and genomic alteration data with cell line-sensitivity data across an annotated collection of cancer cell lines called the Cancer Cell Line Encyclopedia, we show that genetic alterations for FGFR family members predict for sensitivity to NVP-BGJ398. For the first time, we report oncogenic FGFR1 amplification in osteosarcoma as a potential patient selection biomarker. Furthermore, we show that cancer cell lines harboring FGF19 copy number gain at the 11q13 amplicon are sensitive to NVP-BGJ398 only when concomitant expression of ß-klotho occurs. Thus, our findings provide the rationale for the clinical development of FGFR inhibitors in selected patients with cancer harboring tumors with the identified predictors of sensitivity. SIGNIFICANCE: The success of a personalized medicine approach using targeted therapies ultimately depends on being able to identify the patients who will benefit the most from any given drug. To this end, we have integrated the molecular profiles for more than 500 cancer cell lines with sensitivity data for the novel anticancer drug NVP-BGJ398 and showed that FGFR genetic alterations are the most significant predictors for sensitivity. This work has ultimately endorsed the incorporation of specific patient selection biomakers in the clinical trials for NVP-BGJ398.
Sujet(s)
Tumeurs/traitement médicamenteux , Tumeurs/enzymologie , Phénylurées/pharmacologie , Pyrimidines/pharmacologie , Récepteur facteur croissance fibroblaste/génétique , Animaux , Lignée cellulaire tumorale , Amplification de gène/effets des médicaments et des substances chimiques , Cellules HEK293 , Humains , Tumeurs du foie/traitement médicamenteux , Tumeurs du foie/enzymologie , Tumeurs du foie/génétique , Souris , Modèles moléculaires , Tumeurs/génétique , Tumeurs/anatomopathologie , Phénylurées/composition chimique , Inhibiteurs de protéines kinases/composition chimique , Inhibiteurs de protéines kinases/pharmacologie , Pyrimidines/composition chimique , Récepteur FGFR1/antagonistes et inhibiteurs , Récepteur FGFR1/génétique , Récepteur FGFR1/métabolisme , Récepteur FGFR2/antagonistes et inhibiteurs , Récepteur FGFR2/génétique , Récepteur FGFR2/métabolisme , Récepteur facteur croissance fibroblaste/antagonistes et inhibiteurs , Récepteur facteur croissance fibroblaste/métabolisme , Tests d'activité antitumorale sur modèle de xénogreffeRÉSUMÉ
PURPOSE: Reactivation of p53 tumor suppressor activity in diseases such as soft-tissue sarcoma is considered an attractive means of targeted therapy. By systematically assessing alterations affecting the p53 pathway, we aimed to (a) classify sarcoma subtypes, (b) define a potential role in malignancy, and (c) identify potential patient biomarkers in this heterogeneous disease. EXPERIMENTAL DESIGN: We have mapped mutational events in a panel of 192 benign or malignant bone and soft-tissue sarcomas. Analyses included TP53 and CDKN2A mutational and SNP status, MDM2 and MDM4 amplification and MDM2 SNP309 status. RESULTS: We found an inverse relationship between MDM2 amplification and TP53 mutations, with a predominantly wild-type CDKN2A background. A high rate of point mutations in TP53 was observed uniquely in leiomyosarcoma, osteosarcoma, and MFH. Both MDM2 and MDM4 were also amplified in a subtype-specific manner, which was frequently seen as a coamplification event. We have also analyzed the risk allele frequencies for MDM2 SNP309, and show that the G allele was strongly associated with both liposarcomas and MDM2 amplification. CONCLUSIONS: Our data emphasize the critical role of p53 inactivation in sarcomagenesis, whereby different pathway alterations may be related to the heterogeneity of the disease. Moreover, we observed a strong association of malignancy with TP53 mutation, or MDM2 amplification and the presence of a G allele in SNP309, especially in lipoma versus liposarcoma. We propose, therefore, that MDM2 markers along with TP53 sequencing should be considered as patient biomarkers in clinical trials of sarcomas using MDM2 antagonists.
Sujet(s)
Tumeurs osseuses/génétique , Amplification de gène , Gènes p53 , Protéines proto-oncogènes c-mdm2/génétique , Sarcomes/génétique , Tumeurs des tissus mous/génétique , Marqueurs biologiques tumoraux/analyse , Cartographie chromosomique , Dosage génique , Humains , Mutation , Polymorphisme de nucléotide simpleRÉSUMÉ
BACKGROUND: The serotonin type 4 (5-HT4) receptor has been associated with functions of the gastrointestinal tract such as modulation of the peristaltic reflex, smooth muscle tone, intestinal secretion and visceral sensitivity. The activation of peripheral 5-HT4 receptors with agonists such as tegaserod has been shown to accelerate gastric emptying and improve symptoms of constipation in animals and humans. However, detailed data on the expression profile and on the localization of this receptor subtype are lacking so far. OBJECTIVE: To study the pattern and expression levels of 5-HT4 receptor messenger RNA expression in the gut. METHOD: Normal tissue samples were collected from the whole gastrointestinal tract of patients undergoing abdominal surgery and, in addition, of monkeys. We performed a comprehensive analysis of 5-HT4 receptor expression by quantitative reverse transcription-polymerase chain reaction, using human and non-human primate tissues from the oesophagus to the rectum. In addition, the brain and heart of non-human primates were analysed. RESULTS: Significantly higher levels of 5-HT4 receptor mRNA were measured in the human stomach, duodenum, jejunum, ileum and caecum and also in the corresponding non-human primate gut segments, ranging from 2- to 12-fold compared with the liver. No differences were found between females and males of both human and non-human primates. CONCLUSIONS: These results show 5-HT4 receptor mRNA expression throughout the gastrointestinal tract in humans and primates, and also support the preclinical and clinical findings of 5-HT4 receptors ligands exhibiting multiple effects throughout the gastrointestinal tract.