Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 10 de 10
Filtrar
Mais filtros










Base de dados
Intervalo de ano de publicação
1.
Cancer Metab ; 5: 10, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-29214019

RESUMO

BACKGROUND: Cancer cells express the M2 isoform of the glycolytic enzyme pyruvate kinase (PKM2). PKM2 expression is not required for some cancers, and PKM2 loss can promote cancer progression; however, PKM2 has been reported to be essential in other tumor contexts, including a proposed non-metabolic role in ß-catenin nuclear translocation. PKM2 is expressed in colon cancers where loss of the Apc tumor suppressor results in ß-catenin nuclear translocation and aberrant activation of the canonical Wnt signaling pathway. Whether PKM2 is required in this colon cancer context has not been investigated. RESULTS: Colon tumorigenesis was induced in mice harboring conditional Apc and Pkm2 alleles, and tumor progression was monitored by serial colonoscopy. PKM2 deletion had no effect on overall survival, the number of mice that developed tumors, or the number of tumors that developed per animal. Immunohistochemical analysis demonstrated PKM2 expression in wild-type tumors and the expected loss of PKM2 expression in tumors from Pkm2 conditional mice. Loss of PKM2 resulted in pyruvate kinase M1 expression but had no effect on nuclear ß-catenin staining. These findings are consistent with tumor growth and activated Wnt signaling despite PKM2 loss in this model. We also found a large fraction of human colon cancers had very low or undetectable levels of PKM2 expression. CONCLUSIONS: PKM2 is not required for Apc-deficient colon cancer or for nuclear translocation of ß-catenin in Apc-null tumor cells. These findings suggest that PKM2 expression is not required for colon tumor formation or progression.

2.
Sci Signal ; 7(351): ra107, 2014 Nov 11.
Artigo em Inglês | MEDLINE | ID: mdl-25389372

RESUMO

Targeted blockade of aberrantly activated signaling pathways is an attractive therapeutic strategy for solid tumors, but drug resistance is common. KRAS is a frequently mutated gene in human cancer but remains a challenging clinical target. Inhibitors against KRAS signaling mediators, namely, PI3K (phosphatidylinositol 3-kinase) and mTOR (mechanistic target of rapamycin), have limited clinical efficacy as single agents in KRAS-mutant colorectal cancer (CRC). We investigated potential bypass mechanisms to PI3K/mTOR inhibition in KRAS-mutant CRC. Using genetically engineered mouse model cells that had acquired resistance to the dual PI3K/mTOR small-molecule inhibitor PF-04691502, we determined with chemical library screens that inhibitors of the ERBB [epidermal growth factor receptor (EGFR)] family restored the sensitivity to PF-04691502. Although EGFR inhibitors alone have limited efficacy in reducing KRAS-mutant tumors, we found that PF-04691502 induced the abundance, phosphorylation, and activity of EGFR, ERBB2, and ERBB3 through activation of FOXO3a (forkhead box O 3a), a transcription factor inhibited by the PI3K to AKT pathway. PF-04691502 also induced a stem cell-like gene expression signature. KRAS-mutant patient-derived xenografts from mice treated with PF-04691502 had a similar gene expression signature and exhibited increased EGFR activation, suggesting that this drug-induced resistance mechanism may occur in patients. Combination therapy with dacomitinib (a pan-ERBB inhibitor) restored sensitivity to PF-04691502 in drug-resistant cells in culture and induced tumor regression in drug-resistant allografts in mice. Our findings suggest that combining PI3K/mTOR and EGFR inhibitors may improve therapeutic outcome in patients with KRAS-mutant CRC.


Assuntos
Neoplasias Colorretais/metabolismo , Inibidores Enzimáticos/química , Receptores ErbB/antagonistas & inibidores , Genes ras , Inibidores de Fosfoinositídeo-3 Quinase , Serina-Treonina Quinases TOR/antagonistas & inibidores , Animais , Linhagem Celular Tumoral , Proliferação de Células , Separação Celular , Sobrevivência Celular , Neoplasias Colorretais/genética , Modelos Animais de Doenças , Resistencia a Medicamentos Antineoplásicos , Feminino , Citometria de Fluxo , Engenharia Genética , Humanos , Camundongos , Camundongos SCID , Mutação , Transplante de Neoplasias , Fosforilação , Transdução de Sinais , Transcrição Gênica , Proteína Supressora de Tumor p53/metabolismo , beta Catenina/metabolismo , Proteínas ras/genética
3.
Dis Model Mech ; 7(6): 613-23, 2014 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-24742783

RESUMO

Effective treatment options for advanced colorectal cancer (CRC) are limited, survival rates are poor and this disease continues to be a leading cause of cancer-related deaths worldwide. Despite being a highly heterogeneous disease, a large subset of individuals with sporadic CRC typically harbor relatively few established 'driver' lesions. Here, we describe a collection of genetically engineered mouse models (GEMMs) of sporadic CRC that combine lesions frequently altered in human patients, including well-characterized tumor suppressors and activators of MAPK signaling. Primary tumors from these models were profiled, and individual GEMM tumors segregated into groups based on their genotypes. Unique allelic and genotypic expression signatures were generated from these GEMMs and applied to clinically annotated human CRC patient samples. We provide evidence that a Kras signature derived from these GEMMs is capable of distinguishing human tumors harboring KRAS mutation, and tracks with poor prognosis in two independent human patient cohorts. Furthermore, the analysis of a panel of human CRC cell lines suggests that high expression of the GEMM Kras signature correlates with sensitivity to targeted pathway inhibitors. Together, these findings implicate GEMMs as powerful preclinical tools with the capacity to recapitulate relevant human disease biology, and support the use of genetic signatures generated in these models to facilitate future drug discovery and validation efforts.


Assuntos
Neoplasias Colorretais/patologia , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Alelos , Animais , Neoplasias Colorretais/enzimologia , Neoplasias Colorretais/genética , Modelos Animais de Doenças , Genes ras , Humanos , Camundongos , Proteínas Proto-Oncogênicas B-raf/genética , Especificidade da Espécie
4.
Cancer Lett ; 347(2): 204-11, 2014 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-24576621

RESUMO

PI3K inhibition in combination with other agents has not been studied in the context of PIK3CA wild-type, KRAS mutant cancer. In a screen of phospho-kinases, PI3K inhibition of KRAS mutant colorectal cancer cells activated the MAPK pathway. Combination PI3K/MEK inhibition with NVP-BKM120 and PD-0325901 induced tumor regression in a mouse model of PIK3CA wild-type, KRAS mutant colorectal cancer, which was mediated by inhibition of mTORC1, inhibition of MCL-1, and activation of BIM. These findings implicate mitochondrial-dependent apoptotic mechanisms as determinants for the efficacy of PI3K/MEK inhibition in the treatment of PIK3CA wild-type, KRAS mutant cancer.


Assuntos
Apoptose/efeitos dos fármacos , Neoplasias Colorretais/patologia , Genes ras , MAP Quinase Quinase Quinases/antagonistas & inibidores , Mutação , Inibidores de Fosfoinositídeo-3 Quinase , Animais , Classe I de Fosfatidilinositol 3-Quinases , Neoplasias Colorretais/enzimologia , Inibidores Enzimáticos/farmacologia , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Fosfatidilinositol 3-Quinases/genética
5.
Clin Cancer Res ; 19(10): 2688-98, 2013 May 15.
Artigo em Inglês | MEDLINE | ID: mdl-23549875

RESUMO

PURPOSE: BRAF(V600E) mutations are associated with poor clinical prognosis in colorectal cancer (CRC). Although selective BRAF inhibitors are effective for treatment of melanoma, comparable efforts in CRC have been disappointing. Here, we investigated potential mechanisms underlying this resistance to BRAF inhibitors in BRAF(V600E) CRC. EXPERIMENTAL DESIGN: We examined phosphoinositide 3-kinase (PI3K)/mTOR signaling in BRAF(V600E) CRC cell lines after BRAF inhibition and cell viability and apoptosis after combined BRAF and PI3K/mTOR inhibition. We assessed the efficacy of in vivo combination treatment using a novel genetically engineered mouse model (GEMM) for BRAF(V600E) CRC. RESULTS: Western blot analysis revealed sustained PI3K/mTOR signaling upon BRAF inhibition. Our BRAF(V600E) GEMM presented with sessile serrated adenomas/polyps, as seen in humans. Combination treatment in vivo resulted in induction of apoptosis and tumor regression. CONCLUSIONS: We have established a novel GEMM to interrogate BRAF(V600E) CRC biology and identify more efficacious treatment strategies. Combination BRAF and PI3K/mTOR inhibitor treatment should be explored in clinical trials.


Assuntos
Neoplasias Colorretais/tratamento farmacológico , Inibidores de Fosfoinositídeo-3 Quinase , Inibidores de Proteínas Quinases/farmacologia , Proteínas Proto-Oncogênicas B-raf/antagonistas & inibidores , Serina-Treonina Quinases TOR/antagonistas & inibidores , Animais , Apoptose/efeitos dos fármacos , Western Blotting , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Neoplasias Colorretais/genética , Neoplasias Colorretais/patologia , Relação Dose-Resposta a Droga , Células HCT116 , Humanos , Indenos/farmacologia , Camundongos , Camundongos Knockout , Mutação , Neoplasias Experimentais/tratamento farmacológico , Neoplasias Experimentais/genética , Neoplasias Experimentais/patologia , Fosfatidilinositol 3-Quinases/genética , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas B-raf/genética , Proteínas Proto-Oncogênicas B-raf/metabolismo , Pirazóis/farmacologia , Transdução de Sinais/efeitos dos fármacos , Serina-Treonina Quinases TOR/metabolismo , Carga Tumoral/efeitos dos fármacos
6.
Clin Cancer Res ; 19(11): 2929-40, 2013 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-23403635

RESUMO

PURPOSE: Effective therapies for KRAS-mutant colorectal cancer (CRC) are a critical unmet clinical need. Previously, we described genetically engineered mouse models (GEMM) for sporadic Kras-mutant and non-mutant CRC suitable for preclinical evaluation of experimental therapeutics. To accelerate drug discovery and validation, we sought to derive low-passage cell lines from GEMM Kras-mutant and wild-type tumors for in vitro screening and transplantation into the native colonic environment of immunocompetent mice for in vivo validation. EXPERIMENTAL DESIGN: Cell lines were derived from Kras-mutant and non-mutant GEMM tumors under defined media conditions. Growth kinetics, phosphoproteomes, transcriptomes, drug sensitivity, and metabolism were examined. Cell lines were implanted in mice and monitored for in vivo tumor analysis. RESULTS: Kras-mutant cell lines displayed increased proliferation, mitogen-activated protein kinase signaling, and phosphoinositide-3 kinase signaling. Microarray analysis identified significant overlap with human CRC-related gene signatures, including KRAS-mutant and metastatic CRC. Further analyses revealed enrichment for numerous disease-relevant biologic pathways, including glucose metabolism. Functional assessment in vitro and in vivo validated this finding and highlighted the dependence of Kras-mutant CRC on oncogenic signaling and on aerobic glycolysis. CONCLUSIONS: We have successfully characterized a novel GEMM-derived orthotopic transplant model of human KRAS-mutant CRC. This approach combines in vitro screening capability using low-passage cell lines that recapitulate human CRC and potential for rapid in vivo validation using cell line-derived tumors that develop in the colonic microenvironment of immunocompetent animals. Taken together, this platform is a clear advancement in preclinical CRC models for comprehensive drug discovery and validation efforts.


Assuntos
Neoplasias do Colo/genética , Mutação , Proteínas ras/genética , Animais , Carcinogênese/genética , Carcinogênese/metabolismo , Linhagem Celular Tumoral , Análise por Conglomerados , Neoplasias do Colo/tratamento farmacológico , Neoplasias do Colo/metabolismo , Modelos Animais de Doenças , Perfilação da Expressão Gênica , Genes APC , Genes p53 , Genótipo , Glucose/metabolismo , Humanos , Ácido Láctico/biossíntese , Camundongos , Camundongos Transgênicos , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Transdução de Sinais , Serina-Treonina Quinases TOR/metabolismo , Via de Sinalização Wnt , Ensaios Antitumorais Modelo de Xenoenxerto , Proteínas ras/metabolismo
7.
PLoS One ; 6(9): e25132, 2011.
Artigo em Inglês | MEDLINE | ID: mdl-21966435

RESUMO

PURPOSE: To examine the in vitro and in vivo efficacy of the dual PI3K/mTOR inhibitor NVP-BEZ235 in treatment of PIK3CA wild-type colorectal cancer (CRC). EXPERIMENTAL DESIGN: PIK3CA mutant and wild-type human CRC cell lines were treated in vitro with NVP-BEZ235, and the resulting effects on proliferation, apoptosis, and signaling were assessed. Colonic tumors from a genetically engineered mouse (GEM) model for sporadic wild-type PIK3CA CRC were treated in vivo with NVP-BEZ235. The resulting effects on macroscopic tumor growth/regression, proliferation, apoptosis, angiogenesis, and signaling were examined. RESULTS: In vitro treatment of CRC cell lines with NVP-BEZ235 resulted in transient PI3K blockade, sustained decreases in mTORC1/mTORC2 signaling, and a corresponding decrease in cell viability (median IC(50) = 9.0-14.3 nM). Similar effects were seen in paired isogenic CRC cell lines that differed only in the presence or absence of an activating PIK3CA mutant allele. In vivo treatment of colonic tumor-bearing mice with NVP-BEZ235 resulted in transient PI3K inhibition and sustained blockade of mTORC1/mTORC2 signaling. Longitudinal tumor surveillance by optical colonoscopy demonstrated a 97% increase in tumor size in control mice (p = 0.01) vs. a 43% decrease (p = 0.008) in treated mice. Ex vivo analysis of the NVP-BEZ235-treated tumors demonstrated a 56% decrease in proliferation (p = 0.003), no effects on apoptosis, and a 75% reduction in angiogenesis (p = 0.013). CONCLUSIONS: These studies provide the preclinical rationale for studies examining the efficacy of the dual PI3K/mTOR inhibitor NVP-BEZ235 in treatment of PIK3CA wild-type CRC.


Assuntos
Neoplasias Colorretais/tratamento farmacológico , Imidazóis/uso terapêutico , Inibidores de Fosfoinositídeo-3 Quinase , Inibidores de Proteínas Quinases/uso terapêutico , Quinolinas/uso terapêutico , Animais , Apoptose/efeitos dos fármacos , Western Blotting , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Classe I de Fosfatidilinositol 3-Quinases , Neoplasias Colorretais/genética , Neoplasias Colorretais/metabolismo , Células HCT116 , Humanos , Imuno-Histoquímica , Camundongos , Camundongos Knockout , Fosfatidilinositol 3-Quinases/genética , Transdução de Sinais/efeitos dos fármacos
8.
Proc Natl Acad Sci U S A ; 108(1): 290-5, 2011 Jan 04.
Artigo em Inglês | MEDLINE | ID: mdl-21173247

RESUMO

Patients with inflammatory bowel disease (IBD) have increased numbers of human tryptase-ß (hTryptase-ß)-positive mast cells (MCs) in the gastrointestinal tract. The amino acid sequence of mouse mast cell protease (mMCP)-6 is most similar to that of hTryptase-ß. We therefore hypothesized that this mMCP, or the related tryptase mMCP-7, might have a prominent proinflammatory role in experimental colitis. The dextran sodium sulfate (DSS) and trinitrobenzene sulfonic acid (TNBS) colitis models were used to evaluate the differences between C57BL/6 (B6) mouse lines that differ in their expression of mMCP-6 and mMCP-7 with regard to weight loss, colon histopathology, and endoscopy scores. Microarray analyses were performed, and confirmatory real-time PCR, ELISA, and/or immunohistochemical analyses were carried out on a number of differentially expressed cytokines, chemokines, and matrix metalloproteinases (MMPs). The mMCP-6-null mice that had been exposed to DSS had significantly less weight loss as well as significantly lower pathology and endoscopy scores than similarly treated mMCP-6-expressing mice. This difference in colitis severity was confirmed endoscopically in the TNBS-treated mice. Evaluation of the distal colon segments revealed that numerous proinflammatory cytokines, chemokines that preferentially attract neutrophils, and MMPs that participate in the remodeling of the ECM were all markedly increased in the colons of DSS-treated WT mice relative to untreated WT mice and DSS-treated mMCP-6-null mice. Collectively, our data show that mMCP-6 (but not mMCP-7) is an essential MC-restricted mediator in chemically induced colitis and that this tryptase acts upstream of many of the factors implicated in IBD.


Assuntos
Doenças Inflamatórias Intestinais/imunologia , Mastócitos/imunologia , Triptases/imunologia , Animais , Quimiocinas/imunologia , Colo/metabolismo , Colo/patologia , Citocinas/imunologia , Sulfato de Dextrana/toxicidade , Ensaio de Imunoadsorção Enzimática , Imuno-Histoquímica , Doenças Inflamatórias Intestinais/induzido quimicamente , Doenças Inflamatórias Intestinais/enzimologia , Doenças Inflamatórias Intestinais/patologia , Mastócitos/metabolismo , Metaloproteinases da Matriz/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Análise em Microsséries , Reação em Cadeia da Polimerase , Ácido Trinitrobenzenossulfônico/toxicidade , Triptases/metabolismo
9.
Carcinogenesis ; 29(4): 880-6, 2008 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-18258601

RESUMO

Mast cells have been observed in numerous types of tumors; however, their role in carcinogenesis remains poorly understood. The majority of epidemiological evidence suggests a negative association between the presence of mast cells and tumor progression in breast, lung and colonic neoplasms. Intestinal adenomas in the multiple intestinal neoplasia (Min, APC(Min/+)) mouse displayed increased numbers of mast cells and increased abundance of mast cell-associated proteinases as determined by transcriptional profiling with the Hu/Mu ProtIn microarray. To examine the role of mast cells in intestinal tumorigenesis, a mutant mouse line deficient in mast cells, Sash mice (c-kit(W-sh/W-sh)), was crossed with the Min mouse, a genetic model of intestinal neoplasia. The resulting mast cell-deficient Min-Sash mice developed 50% more adenomas than littermate controls and the tumors were 33% larger in Min-Sash mice. Mast cell deficiency did not affect tumor cell proliferation; however, apoptosis was significantly inhibited in mast cell-deficient mice. Mast cells have been shown to act as critical upstream regulators of numerous inflammatory cells. Neutrophil, macrophage and T cell populations were similar between Min and Min-Sash mice; however, eosinophils were significantly less abundant in tumors obtained from Min-Sash animals. These results indicate a protective, antitumor role of mast cells in a genetic model of early-stage intestinal tumorigenesis.


Assuntos
Neoplasias Intestinais/prevenção & controle , Mastócitos/fisiologia , Animais , Endopeptidases/genética , Feminino , Neoplasias Intestinais/enzimologia , Neoplasias Intestinais/genética , Neoplasias Intestinais/patologia , Masculino , Mastócitos/enzimologia , Camundongos , Camundongos Endogâmicos C57BL , Reação em Cadeia da Polimerase
10.
Int J Exp Pathol ; 89(6): 466-75, 2008 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-19134056

RESUMO

Matrix metalloproteinases (MMPs) are a family of 23 extracellular proteases that are best known for their collective ability to degrade all components of the extracellular matrix. We previously demonstrated that genetic ablation of MMP-7 reduced tumour multiplicity in multiple intestinal neoplasia (Min) mice possessing a genetic alteration in the adenomatous polyposis coli gene (APC). These mice, commonly referred to as APC-Min mice, are a frequently used model of early intestinal tumourigenesis. To examine further the role of MMPs in intestinal tumour development, we generated APC-Min mice genetically deficient in MMP-2, -9, -12 or -19. Genetic ablation of MMP-2, -12 or -19 did not affect multiplicity or size of intestinal tumours when crossed into the APC-Min system. However, MMP-9 deficient animals developed 40% fewer tumours than littermate controls, although tumour size distribution remained unaffected. Intestinal adenomas from MMP-9 deficient mice demonstrated a 50% decrease in proliferating cells compared with control tissues, with no difference in apoptosis. To determine the cellular origin of MMP-9 in these tumours, immunofluorescent co-staining with markers for different leucocyte lineages was used to demonstrate that intratumoural MMP-9 is largely a product of neutrophils. These studies extend the potential targets for chemoprevention of intestinal adenomas to MMP-9 in addition to MMP-7 and exclude MMP-2,-12,-19 as attractive targets for intervention.


Assuntos
Polipose Adenomatosa do Colo/enzimologia , Neoplasias do Colo/enzimologia , Mucosa Intestinal/enzimologia , Metaloproteinase 9 da Matriz/análise , Polipose Adenomatosa do Colo/patologia , Animais , Cruzamento , Neoplasias do Colo/patologia , Genes APC , Imuno-Histoquímica , Metaloproteinase 12 da Matriz/análise , Metaloproteinase 2 da Matriz/análise , Metaloproteinase 2 da Matriz/genética , Metaloproteinase 9 da Matriz/genética , Metaloproteinase 9 da Matriz/metabolismo , Metaloproteinases da Matriz Secretadas/análise , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Microscopia de Fluorescência , Neutrófilos/enzimologia
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA
...