RESUMO
A remarkable feature of regenerative processes is their ability to halt proliferation once an organ's structure has been restored. The Wnt signalling pathway is the major driving force for homeostatic self-renewal and regeneration in the mammalian intestine. However, the mechanisms that counterbalance Wnt-driven proliferation are poorly understood. Here we demonstrate in mice and humans that yes-associated protein 1 (YAP; also known as YAP1)--a protein known for its powerful growth-inducing and oncogenic properties--has an unexpected growth-suppressive function, restricting Wnt signals during intestinal regeneration. Transgenic expression of YAP reduces Wnt target gene expression and results in the rapid loss of intestinal crypts. In addition, loss of YAP results in Wnt hypersensitivity during regeneration, leading to hyperplasia, expansion of intestinal stem cells and niche cells, and formation of ectopic crypts and microadenomas. We find that cytoplasmic YAP restricts elevated Wnt signalling independently of the AXIN-APC-GSK-3ß complex partly by limiting the activity of dishevelled (DVL). DVL signals in the nucleus of intestinal stem cells, and its forced expression leads to enhanced Wnt signalling in crypts. YAP dampens Wnt signals by restricting DVL nuclear translocation during regenerative growth. Finally, we provide evidence that YAP is silenced in a subset of highly aggressive and undifferentiated human colorectal carcinomas, and that its expression can restrict the growth of colorectal carcinoma xenografts. Collectively, our work describes a novel mechanistic paradigm for how proliferative signals are counterbalanced in regenerating tissues. Additionally, our findings have important implications for the targeting of YAP in human malignancies.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Proliferação de Células , Intestinos/citologia , Fosfoproteínas/metabolismo , Regeneração/fisiologia , Células-Tronco/citologia , Células-Tronco/metabolismo , Transporte Ativo do Núcleo Celular , Proteínas Adaptadoras de Transdução de Sinal/deficiência , Proteínas Adaptadoras de Transdução de Sinal/genética , Animais , Proteínas de Ciclo Celular , Neoplasias Colorretais/genética , Neoplasias Colorretais/metabolismo , Neoplasias Colorretais/patologia , Proteínas Desgrenhadas , Genes Supressores de Tumor , Humanos , Intestinos/fisiologia , Camundongos , Camundongos Knockout , Transplante de Neoplasias , Fosfoproteínas/deficiência , Fosfoproteínas/genética , Nicho de Células-Tronco , Trombospondinas/genética , Trombospondinas/metabolismo , Fatores de Transcrição , Proteínas Supressoras de Tumor/genética , Proteínas Supressoras de Tumor/metabolismo , Proteínas Wnt/metabolismo , Via de Sinalização Wnt , Proteínas de Sinalização YAPRESUMO
Ablation of the kinases Mst1 and Mst2, orthologs of the Drosophila antiproliferative kinase Hippo, from mouse intestinal epithelium caused marked expansion of an undifferentiated stem cell compartment and loss of secretory cells throughout the small and large intestine. Although median survival of mice lacking intestinal Mst1/Mst2 is 13 wk, adenomas of the distal colon are common by this age. Diminished phosphorylation, enhanced abundance, and nuclear localization of the transcriptional coactivator Yes-associated protein 1 (Yap1) is evident in Mst1/Mst2-deficient intestinal epithelium, as is strong activation of ß-catenin and Notch signaling. Although biallelic deletion of Yap1 from intestinal epithelium has little effect on intestinal development, inactivation of a single Yap1 allele reduces Yap1 polypeptide abundance to nearly wild-type levels and, despite the continued Yap hypophosphorylation and preferential nuclear localization, normalizes epithelial structure. Thus, supraphysiologic Yap polypeptide levels are necessary to drive intestinal stem cell proliferation. Yap is overexpressed in 68 of 71 human colon cancers and in at least 30 of 36 colon cancer-derived cell lines. In colon-derived cell lines where Yap is overabundant, its depletion strongly reduces ß-catenin and Notch signaling and inhibits proliferation and survival. These findings demonstrate that Mst1 and Mst2 actively suppress Yap1 abundance and action in normal intestinal epithelium, an antiproliferative function that frequently is overcome in colon cancer through Yap1 polypeptide overabundance. The dispensability of Yap1 in normal intestinal homeostasis and its potent proliferative and prosurvival actions when overexpressed in colon cancer make it an attractive therapeutic target.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Proliferação de Células , Colo/metabolismo , Fosfoproteínas/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Células-Tronco/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/genética , Animais , Western Blotting , Proteínas de Ciclo Celular , Linhagem Celular Tumoral , Colo/patologia , Neoplasias do Colo/genética , Neoplasias do Colo/metabolismo , Neoplasias do Colo/patologia , Feminino , Regulação Neoplásica da Expressão Gênica , Células HCT116 , Humanos , Imuno-Histoquímica , Mucosa Intestinal/metabolismo , Intestinos/citologia , Masculino , Camundongos , Camundongos Knockout , Camundongos Transgênicos , Fosfoproteínas/genética , Proteínas Serina-Treonina Quinases/genética , Interferência de RNA , Serina-Treonina Quinase 3 , Células-Tronco/citologia , Análise Serial de Tecidos , Fatores de Transcrição , Proteínas de Sinalização YAPRESUMO
Many tumor types harbor alterations in the Hippo pathway, including mesothelioma, where a high percentage of cases are considered YAP1/TEAD dependent. Identification of autopalmitoylation sites in the hydrophobic palmitate pocket of TEADs, which may be necessary for YAP1 protein interactions, has enabled modern drug discovery platforms to generate compounds that allosterically inhibit YAP1/TEAD complex formation and transcriptional activity. We report the discovery and characterization of a novel YAP1/TEAD inhibitor MRK-A from an aryl ether chemical series demonstrating potent and specific inhibition of YAP1/TEAD activity. In vivo, MRK-A showed a favorable tolerability profile in mice and demonstrated pharmacokinetics suitable for twice daily oral dosing in preclinical efficacy studies. Importantly, monotherapeutic targeting of YAP1/TEAD in preclinical models generated regressions in a mesothelioma CDX model; however, rapid resistance to therapy was observed. RNA-sequencing of resistant tumors revealed mRNA expression changes correlated with the resistance state and a marked increase of hepatocyte growth factor (HGF) expression. In vitro, exogenous HGF was able to fully rescue cytostasis induced by MRK-A in mesothelioma cell lines. In addition, co-administration of small molecule inhibitors of the MET receptor tyrosine kinase suppressed the resistance generating effect of HGF on MRK-A induced growth inhibition. In this work, we report the structure and characterization of MRK-A, demonstrating potent and specific inhibition of YAP1/TAZ-TEAD-mediated transcriptional responses, with potential implications for treating malignancies driven by altered Hippo signaling, including factors resulting in acquired drug resistance.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal , Resistencia a Medicamentos Antineoplásicos , Fator de Crescimento de Hepatócito , Proteínas Proto-Oncogênicas c-met , Transdução de Sinais , Fatores de Transcrição , Proteínas de Sinalização YAP , Humanos , Fator de Crescimento de Hepatócito/metabolismo , Animais , Fatores de Transcrição/metabolismo , Fatores de Transcrição/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-met/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-met/metabolismo , Camundongos , Transdução de Sinais/efeitos dos fármacos , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/antagonistas & inibidores , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Linhagem Celular Tumoral , Ensaios Antitumorais Modelo de Xenoenxerto , Bibliotecas de Moléculas Pequenas/farmacologia , Fatores de Transcrição de Domínio TEA , Proliferação de Células/efeitos dos fármacosRESUMO
The Hippo pathway is an evolutionary conserved signaling network that regulates essential processes such as organ size, cell proliferation, migration, stemness and apoptosis. Alterations in this pathway are commonly found in solid tumors and can lead to hyperproliferation, resistance to chemotherapy, compensation for mKRAS and tumor immune evasion. As the terminal effectors of the Hippo pathway, the transcriptional coactivators YAP1/TAZ and the transcription factors TEAD1-4 present exciting opportunities to pharmacologically modulate the Hippo biology in cancer settings, inflammation and regenerative medicine. This review will provide an overview of the progress and current strategies to directly and indirectly target the YAP1/TAZ protein-protein interaction (PPI) with TEAD1-4 across multiple modalities, with focus on recent small molecules able to selectively bind to TEAD, block its autopalmitoylation and inhibit YAP1/TAZ-TEAD-dependent transcription in cancer.
Assuntos
Oncologia , Proteínas Serina-Treonina Quinases/metabolismo , Medicina Regenerativa , Transdução de Sinais , Animais , Ensaios Clínicos como Assunto , Humanos , Inibidores de Proteínas Quinases/química , Inibidores de Proteínas Quinases/farmacologia , Proteínas Serina-Treonina Quinases/antagonistas & inibidoresRESUMO
BACKGROUND AND PURPOSE: Savolitinib (AZD6094, HMPL-504, volitinib) is an oral, potent, and highly MET receptor TK inhibitor. This series of studies aimed to develop a pharmacokinetic-pharmacodynamic (PK/PD) model to link inhibition of MET phosphorylation (pMET) by savolitinib with anti-tumour activity. EXPERIMENTAL APPROACH: Cell line-derived xenograft (CDX) experiments using human lung cancer (EBC-1) and gastric cancer (MKN-45) cells were conducted in athymic nude mice using a variety of doses and schedules of savolitinib. Tumour pMET changes and growth inhibition were calculated after 28 days. Population PK/PD techniques were used to construct a PK/PD model for savolitinib. KEY RESULTS: Savolitinib showed dose- and dose frequency-dependent anti-tumour activity in the CDX models, with more frequent, lower dosing schedules (e.g., twice daily) being more effective than intermittent, higher dosing schedules (e.g., 4 days on/3 days off or 2 days on/5 days off). There was a clear exposure-response relationship, with maximal suppression of pMET of >90%. Data from additional CDX and patient-derived xenograft (PDX) models overlapped, allowing calculation of a single EC50 of 0.38 ng·ml-1 . Tumour growth modelling demonstrated that prolonged, high levels of pMET inhibition (>90%) were required for tumour stasis and regression in the models. CONCLUSION AND IMPLICATIONS: High and persistent levels of MET inhibition by savolitinib were needed for optimal monotherapy anti-tumour activity in preclinical models. The modelling framework developed here can be used to translate tumour growth inhibition from the mouse to human and thus guide choice of clinical dose and schedule.
Assuntos
Antineoplásicos , Proteínas Proto-Oncogênicas c-met , Animais , Antineoplásicos/farmacologia , Linhagem Celular Tumoral , Camundongos , Camundongos Nus , Inibidores de Proteínas Quinases/farmacologia , Pirazinas , Triazinas , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Mouse embryonic stem cells (ESCs) proliferate with rapid cell cycle kinetics but without loss of pluripotency. The histone methyltransferase Dot1L is responsible for methylation of histone H3 at lysine 79 (H3K79me). We investigated whether ESCs require Dot1L for proper stem cell behavior. ESCs deficient in Dot1L tolerate a nearly complete loss of H3K79 methylation without a substantial impact on proliferation or morphology. However, shortly after differentiation is induced, Dot1L-deficient cells cease proliferating and arrest in G2/M-phase of the cell cycle, with increased levels of aneuploidy. In addition, many aberrant mitotic spindles occur in Dot1L-deficient cells. Surprisingly, these mitotic and cell cycle defects fail to trigger apoptosis, indicating that mouse ESCs lack stringent cell cycle checkpoint control during initial stages of differentiation. Transcriptome analysis indicates that Dot1L deficiency causes the misregulation of a select set of genes, including many with known roles in cell cycle control and cellular proliferation as well as markers of endoderm differentiation. The data indicate a requirement for Dot1L function for early stages of ESC differentiation where Dot1L is necessary for faithful execution of mitosis and proper transcription of many genes throughout the genome.
Assuntos
Apoptose/fisiologia , Ciclo Celular/fisiologia , Diferenciação Celular/fisiologia , Células-Tronco Embrionárias/citologia , Células-Tronco Embrionárias/metabolismo , Metiltransferases/fisiologia , Animais , Apoptose/genética , Ciclo Celular/genética , Diferenciação Celular/genética , Proliferação de Células , Imunoprecipitação da Cromatina , Histona-Lisina N-Metiltransferase , Cariotipagem , Metiltransferases/genética , Camundongos , Reação em Cadeia da Polimerase , Interferência de RNARESUMO
PURPOSE: The emergence of secondary mutations is a cause of resistance to current KIT inhibitors used in the treatment of patients with gastrointestinal stromal tumors (GIST). AZD3229 is a selective inhibitor of wild-type KIT and a wide spectrum of primary and secondary mutations seen in patients with GIST. The objective of this analysis is to establish the pharmacokinetic-pharmacodynamic (PKPD) relationship of AZD3229 in a range of mouse GIST tumor models harboring primary and secondary KIT mutations, and to benchmark AZD3229 against other KIT inhibitors. EXPERIMENTAL DESIGN: A PKPD model was developed for AZD3229 linking plasma concentrations to inhibition of phosphorylated KIT using data generated from several in vivo preclinical tumor models, and in vitro data generated in a panel of Ba/F3 cell lines. RESULTS: AZD3229 drives inhibition of phosphorylated KIT in an exposure-dependent manner, and optimal efficacy is observed when >90% inhibition of KIT phosphorylation is sustained over the dosing interval. Integrating the predicted human pharmacokinetics into the mouse PKPD model predicts that an oral twice daily human dose greater than 34 mg is required to ensure adequate coverage across the mutations investigated. Benchmarking shows that compared with standard-of-care KIT inhibitors, AZD3229 has the potential to deliver the required target coverage across a wider spectrum of primary or secondary mutations. CONCLUSIONS: We demonstrate that AZD3229 warrants clinical investigation as a new treatment for patients with GIST based on its ability to inhibit both ATP-binding and A-loop mutations of KIT at clinically relevant exposures.
Assuntos
Tumores do Estroma Gastrointestinal/tratamento farmacológico , Inibidores de Proteínas Quinases/farmacologia , Proteínas Proto-Oncogênicas c-kit/antagonistas & inibidores , Quinazolinas/farmacologia , Triazóis/farmacologia , Animais , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Tumores do Estroma Gastrointestinal/patologia , Humanos , Camundongos , Modelos Biológicos , Mutação , Fosforilação/efeitos dos fármacos , Inibidores de Proteínas Quinases/uso terapêutico , Proteínas Proto-Oncogênicas c-kit/genética , Proteínas Proto-Oncogênicas c-kit/metabolismo , Quinazolinas/uso terapêutico , Receptor alfa de Fator de Crescimento Derivado de Plaquetas/genética , Triazóis/uso terapêutico , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Gastrointestinal stromal tumor (GIST) is the most common human sarcoma driven by mutations in KIT or platelet-derived growth factor α (PDGFRα). Although first-line treatment, imatinib, has revolutionized GIST treatment, drug resistance due to acquisition of secondary KIT/PDGFRα mutations develops in a majority of patients. Second- and third-line treatments, sunitinib and regorafenib, lack activity against a plethora of mutations in KIT/PDGFRα in GIST, with median time to disease progression of 4 to 6 months and inhibition of vascular endothelial growth factor receptor 2 (VEGFR2) causing high-grade hypertension. Patients with GIST have an unmet need for a well-tolerated drug that robustly inhibits a range of KIT/PDGFRα mutations. Here, we report the discovery and pharmacological characterization of AZD3229, a potent and selective small-molecule inhibitor of KIT and PDGFRα designed to inhibit a broad range of primary and imatinib-resistant secondary mutations seen in GIST. In engineered and GIST-derived cell lines, AZD3229 is 15 to 60 times more potent than imatinib in inhibiting KIT primary mutations and has low nanomolar activity against a wide spectrum of secondary mutations. AZD3229 causes durable inhibition of KIT signaling in patient-derived xenograft (PDX) models of GIST, leading to tumor regressions at doses that showed no changes in arterial blood pressure (BP) in rat telemetry studies. AZD3229 has a superior potency and selectivity profile to standard of care (SoC) agents-imatinib, sunitinib, and regorafenib, as well as investigational agents, avapritinib (BLU-285) and ripretinib (DCC-2618). AZD3229 has the potential to be a best-in-class inhibitor for clinically relevant KIT/PDGFRα mutations in GIST.
Assuntos
Antineoplásicos , Tumores do Estroma Gastrointestinal , Animais , Antineoplásicos/farmacologia , Antineoplásicos/uso terapêutico , Resistencia a Medicamentos Antineoplásicos , Tumores do Estroma Gastrointestinal/tratamento farmacológico , Tumores do Estroma Gastrointestinal/genética , Humanos , Mutação , Naftiridinas , Inibidores de Proteínas Quinases/farmacologia , Inibidores de Proteínas Quinases/uso terapêutico , Proteínas Proto-Oncogênicas c-kit/genética , Pirazóis , Pirróis , Ratos , Receptor alfa de Fator de Crescimento Derivado de Plaquetas/genética , Triazinas , Ureia/análogos & derivados , Fator A de Crescimento do Endotélio VascularRESUMO
Following fertilization, the newly formed zygote faces several critical decisions regarding cell fate and lineage commitment. First, the parental genomes must be reprogrammed and reset for the zygotic genome to assume responsibility for gene expression. Second, blastomeres must be committed to form either the inner cell mass or trophectoderm before implantation. A variety of epigenetic mechanisms underlies each of these steps, allowing for proper activation of transcriptional circuits which function to specify a cell's identity and maintain or adjust that state as developmental and environmental conditions dictate. These epigenetic mechanisms encompass DNA methylation, post-translational histone modification, chromatin remodeling, and alterations in nuclear architecture. In recent years, stem cells derived from the inner cell mass have been used to examine the epigenetic pathways that regulate pluripotency, differentiation, and lineage commitment. From a technical standpoint, embryonic stem cells provide an easier system to work with compared to preimplantation embryos; however, it is currently unknown how closely the epigenetic mechanisms of cultured stem cells resemble their counterparts in the intact embryo. Furthermore, it remains unclear how similar the reprogramming pathways in artificially created systems, such as nuclear transfer-derived embryos and induced pluripotent stem cells, are to those in naturally created embryos. In this review, we summarize the current knowledge of epigenetic influences during preimplantation development and shed light on the extent to which these pathways are conserved in cultured pluripotent cells in vitro. In doing so, we demonstrate the critical role that epigenetic mechanisms play in the establishment of cell fate during the earliest stages of mammalian development.
Assuntos
Desenvolvimento Embrionário/genética , Epigênese Genética , Animais , Montagem e Desmontagem da Cromatina , Metilação de DNA , Células-Tronco Embrionárias/metabolismo , Feminino , Regulação da Expressão Gênica no Desenvolvimento , Histonas/metabolismo , Humanos , Modelos Genéticos , Oócitos/metabolismo , Células-Tronco Pluripotentes/metabolismo , Gravidez , RNA não Traduzido/genética , Inativação do Cromossomo X , Zigoto/metabolismoRESUMO
While the treatment of gastrointestinal stromal tumors (GISTs) has been revolutionized by the application of targeted tyrosine kinase inhibitors capable of inhibiting KIT-driven proliferation, diverse mutations to this kinase drive resistance to established therapies. Here we describe the identification of potent pan-KIT mutant kinase inhibitors that can be dosed without being limited by the tolerability issues seen with multitargeted agents. This effort focused on identification and optimization of an existing kinase scaffold through the use of structure-based design. Starting from a series of previously reported phenoxyquinazoline and quinoline based inhibitors of the tyrosine kinase PDGFRα, potency against a diverse panel of mutant KIT driven Ba/F3 cell lines was optimized, with a particular focus on reducing activity against a KDR driven cell model in order to limit the potential for hypertension commonly seen in second and third line GIST therapies. AZD3229 demonstrates potent single digit nM growth inhibition across a broad cell panel, with good margin to KDR-driven effects. Selectivity over KDR can be rationalized predominantly by the interaction of water molecules with the protein and ligand in the active site, and its kinome selectivity is similar to the best of the approved GIST agents. This compound demonstrates excellent cross-species pharmacokinetics, shows strong pharmacodynamic inhibition of target, and is active in several in vivo models of GIST.
Assuntos
Descoberta de Drogas , Tumores do Estroma Gastrointestinal/tratamento farmacológico , Proteínas Mutantes/antagonistas & inibidores , Mutação , Inibidores de Proteínas Quinases/química , Inibidores de Proteínas Quinases/farmacologia , Proteínas Proto-Oncogênicas c-kit/antagonistas & inibidores , Quinazolinas/química , Quinazolinas/farmacologia , Triazóis/química , Triazóis/farmacologia , Neoplasias Gastrointestinais/tratamento farmacológico , Neoplasias Gastrointestinais/metabolismo , Neoplasias Gastrointestinais/patologia , Tumores do Estroma Gastrointestinal/metabolismo , Tumores do Estroma Gastrointestinal/patologia , Humanos , Modelos Moleculares , Proteínas Mutantes/genética , Proteínas Mutantes/metabolismo , Conformação Proteica , Inibidores de Proteínas Quinases/farmacocinética , Proteínas Proto-Oncogênicas c-kit/genética , Proteínas Proto-Oncogênicas c-kit/metabolismo , Quinazolinas/farmacocinética , Distribuição Tecidual , Triazóis/farmacocinética , Células Tumorais CultivadasRESUMO
Cancer genome sequencing projects have identified hundreds of genetic alterations, often at low frequencies, raising questions as to their functional relevance. One exemplar gene is HUWE1, which has been found to be mutated in numerous studies. However, due to the large size of this gene and a lack of functional analysis of identified mutations, their significance to carcinogenesis is unclear. To determine the importance of HUWE1, we chose to examine its function in colorectal cancer, where it is mutated in up to 15 per cent of tumours. Modelling of identified mutations showed that they inactivate the E3 ubiquitin ligase activity of HUWE1. Genetic deletion of Huwe1 rapidly accelerated tumourigenic in mice carrying loss of the intestinal tumour suppressor gene Apc, with a dramatic increase in tumour initiation. Mechanistically, this phenotype was driven by increased MYC and rapid DNA damage accumulation leading to loss of the second copy of Apc The increased levels of DNA damage sensitised Huwe1-deficient tumours to DNA-damaging agents and to deletion of the anti-apoptotic protein MCL1. Taken together, these data identify HUWE1 as a bona fide tumour suppressor gene in the intestinal epithelium and suggest a potential vulnerability of HUWE1-mutated tumours to DNA-damaging agents and inhibitors of anti-apoptotic proteins.
Assuntos
Carcinogênese , Neoplasias Colorretais/patologia , Dano ao DNA , Genes Supressores de Tumor , Proteínas Proto-Oncogênicas c-myc/metabolismo , Transdução de Sinais , Ubiquitina-Proteína Ligases/metabolismo , Animais , Deleção de Genes , Camundongos , Proteínas Mutantes/genética , Proteínas Mutantes/metabolismo , Proteínas Supressoras de Tumor , Ubiquitina-Proteína Ligases/genéticaRESUMO
Lung cancer is the most common cause of cancer death globally with a significant, unmet need for more efficacious treatments. The receptor tyrosine kinase MET has been implicated as an oncogene in numerous cancer subtypes, including non-small cell lung cancer (NSCLC). Here we explore the therapeutic potential of savolitinib (volitinib, AZD6094, HMPL-504), a potent and selective MET inhibitor, in NSCLC. In vitro, savolitinib inhibits MET phosphorylation with nanomolar potency, which correlates with blockade of PI3K/AKT and MAPK signaling as well as MYC down-regulation. In vivo, savolitinib causes inhibition of these pathways and significantly decreases growth of MET-dependent xenografts. To understand resistance mechanisms, we generated savolitinib resistance in MET-amplified NSCLC cell lines and analyzed individual clones. We found that upregulation of MYC and constitutive mTOR pathway activation is a conserved feature of resistant clones that can be overcome by knockdown of MYC or dual mTORC1/2 inhibition. Lastly, we demonstrate that mechanisms of resistance are heterogeneous, arising via a switch to EGFR dependence or by a requirement for PIM signaling. This work demonstrates the efficacy of savolitinib in NSCLC and characterizes acquired resistance, identifying both known and novel mechanisms that may inform combination strategies in the clinic.
Assuntos
Antineoplásicos/farmacologia , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Resistencia a Medicamentos Antineoplásicos , Neoplasias Pulmonares/tratamento farmacológico , Proteínas Proto-Oncogênicas c-myc/metabolismo , Pirazinas/química , Serina-Treonina Quinases TOR/metabolismo , Triazinas/química , Animais , Carcinoma Pulmonar de Células não Pequenas/metabolismo , Linhagem Celular Tumoral , Sobrevivência Celular , Regulação para Baixo , Receptores ErbB/metabolismo , Feminino , Humanos , Neoplasias Pulmonares/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Transplante de Neoplasias , Fosfatidilinositol 3-Quinases/metabolismo , Fosforilação , Proteínas Proto-Oncogênicas c-met/metabolismoRESUMO
Although endocrine therapy is successfully used to treat patients with estrogen receptor (ER) positive breast cancer, a substantial proportion of this population will relapse. Several mechanisms of acquired resistance have been described including activation of the mTOR pathway, increased activity of CDK4 and activating mutations in ER. Using a patient derived xenograft model harboring a common activating ER ligand binding domain mutation (D538G), we evaluated several combinatorial strategies using the selective estrogen receptor degrader (SERD) fulvestrant in combination with chromatin modifying agents, and CDK4/6 and mTOR inhibitors. In this model, fulvestrant binds WT and MT ER, reduces ER protein levels, and downregulated ER target gene expression. Addition of JQ1 or vorinostat to fulvestrant resulted in tumor regression (41% and 22% regression, respectively) though no efficacy was seen when either agent was given alone. Interestingly, although the CDK4/6 inhibitor palbociclib and mTOR inhibitor everolimus were efficacious as monotherapies, long-term delayed tumor growth was only observed when co-administered with fulvestrant. This observation was consistent with a greater inhibition of compensatory signaling when palbociclib and everolimus were co-dosed with fulvestrant. The addition of fulvestrant to JQ1, vorinostat, everolimus and palbociclib also significantly reduced lung metastatic burden as compared to monotherapy. The combination potential of fulvestrant with palbociclib or everolimus were confirmed in an MCF7 CRISPR model harboring the Y537S ER activating mutation. Taken together, these data suggest that fulvestrant may have an important role in the treatment of ER positive breast cancer with acquired ER mutations.
Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Neoplasias da Mama/tratamento farmacológico , Quinase 4 Dependente de Ciclina/antagonistas & inibidores , Quinase 6 Dependente de Ciclina/antagonistas & inibidores , Mutação , Receptores de Estrogênio/genética , Serina-Treonina Quinases TOR/antagonistas & inibidores , Animais , Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Resistencia a Medicamentos Antineoplásicos , Estradiol/administração & dosagem , Estradiol/análogos & derivados , Everolimo/administração & dosagem , Feminino , Fulvestranto , Humanos , Células MCF-7 , Camundongos , Piperazinas/administração & dosagem , Piridinas/administração & dosagem , Receptores de Estrogênio/análise , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
PURPOSE: Papillary renal cell carcinoma (PRCC) is the second most common cancer of the kidney and carries a poor prognosis for patients with nonlocalized disease. The HGF receptor MET plays a central role in PRCC and aberrations, either through mutation, copy number gain, or trisomy of chromosome 7 occurring in the majority of cases. The development of effective therapies in PRCC has been hampered in part by a lack of available preclinical models. We determined the pharmacodynamic and antitumor response of the selective MET inhibitor AZD6094 in two PRCC patient-derived xenograft (PDX) models. EXPERIMENTAL DESIGN: Two PRCC PDX models were identified and MET mutation status and copy number determined. Pharmacodynamic and antitumor activity of AZD6094 was tested using a dose response up to 25 mg/kg daily, representing clinically achievable exposures, and compared with the activity of the RCC standard-of-care sunitinib (in RCC43b) or the multikinase inhibitor crizotinib (in RCC47). RESULTS: AZD6094 treatment resulted in tumor regressions, whereas sunitinib or crizotinib resulted in unsustained growth inhibition. Pharmacodynamic analysis of tumors revealed that AZD6094 could robustly suppress pMET and the duration of target inhibition was dose related. AZD6094 inhibited multiple signaling nodes, including MAPK, PI3K, and EGFR. Finally, at doses that induced tumor regression, AZD6094 resulted in a dose- and time-dependent induction of cleaved PARP, a marker of cell death. CONCLUSIONS: Data presented provide the first report testing therapeutics in preclinical in vivo models of PRCC and support the clinical development of AZD6094 in this indication.
Assuntos
Antineoplásicos/farmacologia , Carcinoma Papilar/metabolismo , Carcinoma Papilar/patologia , Carcinoma de Células Renais/metabolismo , Carcinoma de Células Renais/patologia , Inibidores de Proteínas Quinases/farmacologia , Proteínas Proto-Oncogênicas c-met/antagonistas & inibidores , Pirazinas/farmacologia , Triazinas/farmacologia , Animais , Antineoplásicos/administração & dosagem , Carcinoma Papilar/tratamento farmacológico , Carcinoma Papilar/genética , Carcinoma de Células Renais/tratamento farmacológico , Carcinoma de Células Renais/genética , Linhagem Celular Tumoral , Crizotinibe , Modelos Animais de Doenças , Relação Dose-Resposta a Droga , Feminino , Humanos , Indóis/farmacologia , Inibidores de Proteínas Quinases/administração & dosagem , Proteínas Proto-Oncogênicas c-met/genética , Pirazinas/administração & dosagem , Pirazóis/farmacologia , Piridinas/farmacologia , Pirróis/farmacologia , Sunitinibe , Triazinas/administração & dosagem , Carga Tumoral/efeitos dos fármacos , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Tissue regeneration is vital to the form and function of an organ. At the core of an organs' ability to self-renew is the stem cell, which maintains homeostasis, and repopulates injured or aged tissue. Tissue damage can dramatically change the dimensions of an organ, and during regeneration, an organ must halt growth once the original tissue dimensions have been restored. Therefore, stem cells must give rise to the appropriate number of differentiated progeny to achieve homeostasis. How this tissue-size checkpoint is regulated and how tissue size information relayed to stem cell compartments is unclear, however, it is likely that these mechanisms are altered during the course of tumorigenesis. An emerging signaling cascade, the Hippo Signaling Pathway, is a broadly conserved potent organ size regulator [1]. However, this pathway does not act alone. A number of examples demonstrate crosstalk between Hippo and other signaling pathways including Wnt, Tgfß and Notch, with implications for stem cell biology. Here, we focus on these interactions primarily in the context of well characterized stem cell populations.
Assuntos
Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Transdução de Sinais , Células-Tronco/metabolismo , Animais , Proteínas de Drosophila/metabolismo , Desenvolvimento Embrionário , Células Epidérmicas , Humanos , Intestinos/citologia , Células-Tronco Neoplásicas/metabolismo , Proteínas Nucleares/metabolismo , Tamanho do Órgão , Células-Tronco/citologia , Células-Tronco/patologia , Transativadores/metabolismo , Proteínas Wnt/metabolismo , Proteínas de Sinalização YAPRESUMO
IMPORTANCE OF THE FIELD: The establishment and maintenance of specialized chromatin is crucial for correct gene expression and chromosome stability in mammalian cells. Therefore, epigenetic insults are frequently observed in cancer. Several chromatin modifying enzymes have been implicated in leukemia, and are attractive candidates for the development of therapeutic agents. AREAS COVERED IN THIS REVIEW: The histone methyltransferase DOT1L is responsible for methylation of histone H3 at lysine 79 and is involved in the pathobiology of several leukemias, the majority of which are characterized by chromosomal translocations involving the mixed lineage leukemia (MLL) gene. Leukemic translocations yield fusion proteins involving MLL and other proteins that physically interact with DOT1L. These oncogenic fusion proteins recruit DOT1L to ectopic loci (including HOX gene clusters), whose mis-expression contributes to the transformed phenotype. Studies from stem cells and certain leukemias suggest a second mechanism of leukemogenesis, in which reduced or mistargeted DOT1L activity yields altered centromeric chromatin and consequent chromosomal instability. Targeting DOT1L enzymatic activity as well as interactions with leukemogenic fusion proteins is discussed as possible leads in therapeutic interventions. WHAT THE READER WILL GAIN: In this review, we discuss the normal functions of DOT1L, its mechanistic roles in leukemogenesis, and possible strategies for targeting DOT1L in leukemia. DOT1L is an atypical histone lysine methyltransferase in that it does not contain an enzymatic domain common to all other lysine methyltranferases. This attribute makes DOT1L a unique and specifically targetable enzyme. An emerging role for DOT1L under normal cellular conditions as well as transformed conditions is emerging and shedding light on the biology and mechanisms of some translocation-induced leukemias. TAKE HOME MESSAGE: DOT1L is critical in development, as shown in studies in mouse embryos and embryonic stem cells. DOT1L enzymatic activity is also required for the leukemic transformation capabilities of a number of oncogenic fusion proteins. In addition, interactions between DOT1L and oncogenic fusion proteins are necessary for the transformation process. Therefore, it may be possible to specifically target DOT1L enzymatic activity or DOT1L interactions with leukemogenic fusion proteins.