RESUMO
BACKGROUND: Breast cancer is a heterogenous disease with several histological and molecular subtypes. Models that represent these subtypes are essential for translational research aimed at improving clinical strategy for targeted therapeutics. METHODS: Different combinations of genetic aberrations (Brca1 and Trp53 loss, and inhibition of proteins of the Rb family) were induced in the mammary gland by injection of adenovirus expressing Cre recombinase into the mammary ducts of adult genetically engineered mice. Mammary tumors with different genetic aberrations were classified into molecular subtypes based on expression of molecular markers and RNAseq analysis. In vitro potency assays and Western blots were used to examine their drug sensitivities. RESULTS: Induction of Brca1 and Trp53 loss in mammary ductal epithelium resulted in development of basal-like hormone receptor (HR)-negative mammary tumors. Inhibition of Rb and Trp53 loss or the combination of Rb, Trp53 and Brca1 aberrations resulted in development of luminal ductal carcinoma positive for ER, PR, and Her2 expression. HR positivity in tumors with Rb, Trp53 and Brca1 aberrations indicated that functionality of the Rb pathway rather than Brca1 status affected HR status in these models. Mammary tumor gene expression profiles recapitulated human basal-like or luminal B breast cancer signatures, but HR-positive luminal cancer models were endocrine resistant and exhibited upregulation of PI3K signaling and sensitivity to this pathway inhibition. Furthermore, both tumor subtypes were resistant to CDK4/6 inhibition. CONCLUSIONS: Examination of molecular expression profiles and drug sensitivities of tumors indicate that these breast cancer models can be utilized as a translational platform for evaluation of targeted combinations to improve chemotherapeutic response in patients that no longer respond to hormone therapy or that are resistant to CDK4/6 inhibition.
Assuntos
Neoplasias da Mama , Glândulas Mamárias Humanas , Neoplasias Mamárias Animais , Camundongos , Animais , Humanos , Feminino , Glândulas Mamárias Humanas/metabolismo , Fosfatidilinositol 3-Quinases , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/genética , Neoplasias da Mama/metabolismo , Neoplasias Mamárias Animais/patologia , Epitélio/metabolismo , Hormônios , Proteína BRCA1/genéticaRESUMO
Mice that are homozygous with respect to a mutation (ax(J)) in the ataxia (ax) gene develop severe tremors by 2-3 weeks of age followed by hindlimb paralysis and death by 6-10 weeks of age. Here we show that ax encodes ubiquitin-specific protease 14 (Usp14). Ubiquitin proteases are a large family of cysteine proteases that specifically cleave ubiquitin conjugates. Although Usp14 can cleave a ubiquitin-tagged protein in vitro, it is unable to process polyubiquitin, which is believed to be associated with the protein aggregates seen in Parkinson disease, spinocerebellar ataxia type 1 (SCA1; ref. 4) and gracile axonal dystrophy (GAD). The physiological substrate of Usp14 may therefore contain a mono-ubiquitin side chain, the removal of which would regulate processes such as protein localization and protein activity. Expression of Usp14 is significantly altered in ax(J)/ax(J) mice as a result of the insertion of an intracisternal-A particle (IAP) into intron 5 of Usp14. In contrast to other neurodegenerative disorders such as Parkinson disease and SCA1 in humans and GAD in mice, neither ubiquitin-positive protein aggregates nor neuronal cell loss is detectable in the central nervous system (CNS) of ax(J) mice. Instead, ax(J) mice have defects in synaptic transmission in both the central and peripheral nervous systems. These results suggest that ubiquitin proteases are important in regulating synaptic activity in mammals.
Assuntos
Ataxia/genética , Endopeptidases/genética , Mutação , Alelos , Processamento Alternativo , Animais , Northern Blotting , Southern Blotting , Encéfalo/patologia , Clonagem Molecular , Citosol/metabolismo , Eletrofisiologia , Endopeptidases/fisiologia , Ligação Genética , Immunoblotting , Hibridização In Situ , Meiose , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos CBA , Modelos Genéticos , Neurônios/metabolismo , Fenótipo , Mapeamento Físico do Cromossomo , Proteínas/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Sinapses/metabolismo , Fatores de Tempo , Distribuição Tecidual , Ubiquitina/metabolismo , Ubiquitina TiolesteraseRESUMO
Current therapies for glioblastoma multiforme (GBM), the highest grade malignant brain tumor, are mostly ineffective, and better preclinical model systems are needed to increase the successful translation of drug discovery efforts into the clinic. Previous work describes a genetically engineered mouse (GEM) model that contains perturbations in the most frequently dysregulated networks in GBM (driven by RB, KRAS and/or PI3K signaling and PTEN) that induce development of Grade IV astrocytoma with properties of the human disease. Here, we developed and characterized an orthotopic mouse model derived from the GEM that retains the features of the GEM model in an immunocompetent background; however, this model is also tractable and efficient for preclinical evaluation of candidate therapeutic regimens. Orthotopic brain tumors are highly proliferative, invasive and vascular, and express histology markers characteristic of human GBM. Primary tumor cells were examined for sensitivity to chemotherapeutics and targeted drugs. PI3K and MAPK pathway inhibitors, when used as single agents, inhibited cell proliferation but did not result in significant apoptosis. However, in combination, these inhibitors resulted in a substantial increase in cell death. Moreover, these findings translated into the in vivo orthotopic model: PI3K or MAPK inhibitor treatment regimens resulted in incomplete pathway suppression and feedback loops, whereas dual treatment delayed tumor growth through increased apoptosis and decreased tumor cell proliferation. Analysis of downstream pathway components revealed a cooperative effect on target downregulation. These concordant results, together with the morphologic similarities to the human GBM disease characteristics of the model, validate it as a new platform for the evaluation of GBM treatment.
Assuntos
Neoplasias Encefálicas/metabolismo , Glioblastoma/metabolismo , Proteínas Quinases Ativadas por Mitógeno/antagonistas & inibidores , Inibidores de Fosfoinositídeo-3 Quinase , Transdução de Sinais , Animais , Antineoplásicos/química , Apoptose , Biomarcadores Tumorais/metabolismo , Neoplasias Encefálicas/tratamento farmacológico , Linhagem Celular Tumoral , Proliferação de Células , Sobrevivência Celular , Relação Dose-Resposta a Droga , Regulação Neoplásica da Expressão Gênica , Proteína Glial Fibrilar Ácida , Glioblastoma/tratamento farmacológico , Humanos , Sistema de Sinalização das MAP Quinases , Camundongos , Invasividade Neoplásica , Transplante de Neoplasias , Fosfatidilinositol 3-Quinases/metabolismoRESUMO
The high mortality rate from ovarian cancers can be attributed to late-stage diagnosis and lack of effective treatment. Despite enormous effort to develop better targeted therapies, platinum-based chemotherapy still remains the standard of care for ovarian cancer patients, and resistance occurs at a high rate. One of the rate limiting factors for translation of new drug discoveries into clinical treatments has been the lack of suitable preclinical cancer models with high predictive value. We previously generated genetically engineered mouse (GEM) models based on perturbation of Tp53 and Rb with or without Brca1 or Brca2 that develop serous epithelial ovarian cancer (SEOC) closely resembling the human disease on histologic and molecular levels. Here, we describe an adaptation of these GEM models to orthotopic allografts that uniformly develop tumors with short latency and are ideally suited for routine preclinical studies. Ovarian tumors deficient in Brca1 respond to treatment with cisplatin and olaparib, a PARP inhibitor, whereas Brca1-wild type tumors are non-responsive to treatment, recapitulating the relative sensitivities observed in patients. These mouse models provide the opportunity for evaluation of effective therapeutics, including prediction of differential responses in Brca1-wild type and Brca1-deficient tumors and development of relevant biomarkers.
Assuntos
Cistadenocarcinoma Seroso/genética , Cistadenocarcinoma Seroso/metabolismo , Neoplasias Epiteliais e Glandulares/genética , Neoplasias Epiteliais e Glandulares/metabolismo , Neoplasias Ovarianas/genética , Neoplasias Ovarianas/metabolismo , Aloenxertos , Animais , Antineoplásicos/administração & dosagem , Antineoplásicos/farmacologia , Proteína BRCA1/genética , Carcinoma Epitelial do Ovário , Linhagem Celular Tumoral , Análise por Conglomerados , Cistadenocarcinoma Seroso/tratamento farmacológico , Cistadenocarcinoma Seroso/mortalidade , Cistadenocarcinoma Seroso/patologia , Modelos Animais de Doenças , Progressão da Doença , Relação Dose-Resposta a Droga , Resistencia a Medicamentos Antineoplásicos/genética , Feminino , Perfilação da Expressão Gênica , Humanos , Camundongos , Mutação , Neoplasias Epiteliais e Glandulares/tratamento farmacológico , Neoplasias Epiteliais e Glandulares/mortalidade , Neoplasias Epiteliais e Glandulares/patologia , Neoplasias Ovarianas/tratamento farmacológico , Neoplasias Ovarianas/mortalidade , Neoplasias Ovarianas/patologia , Carga Tumoral/efeitos dos fármacosRESUMO
The majority of human high-grade serous epithelial ovarian cancer (SEOC) is characterized by frequent mutations in p53 and alterations in the RB and FOXM1 pathways. A subset of human SEOC harbors a combination of germline and somatic mutations as well as epigenetic dysfunction for BRCA1/2. Using Cre-conditional alleles and intrabursal induction by Cre-expressing adenovirus in genetically engineered mice, we analyzed the roles of pathway perturbations in epithelial ovarian cancer initiation and progression. Inactivation of RB-mediated tumor suppression induced surface epithelial proliferation with progression to stage I carcinoma. Additional biallelic inactivation and/or missense p53 mutation in the presence or absence of Brca1/2 caused progression to stage IV disease. As in human SEOC, mice developed peritoneal carcinomatosis, ascites, and distant metastases. Unbiased gene expression and metabolomic profiling confirmed that Rb, p53, and Brca1/2-triple mutant tumors aligned with human SEOC, and not with other intraperitoneal cancers. Together, our findings provide a novel resource for evaluating disease etiology and biomarkers, therapeutic evaluation, and improved imaging strategies in epithelial ovarian cancer.