RESUMO
Thyroid cancer development is driven by known point mutations or gene fusions found in â¼90% of cases, whereas driver mutations in the remaining tumors are unknown. The insulin-like growth factor 2 mRNA-binding protein 3 (IGF2BP3) plays an important role in cancer, yet the mechanisms of its activation in cancer cells remain poorly understood. Using whole-transcriptome and whole-genome analyses, we identified a recurrent fusion between the thyroid adenoma-associated (THADA) gene on chromosome 2 and the LOC389473 gene on chromosome 7 located 12 kb upstream of the IGF2BP3 gene. We show that THADA fusion to LOC389473 and other regions in the vicinity does not result in the formation of a chimeric protein but instead leads to strong overexpression of the full-length IGF2BP3 mRNA and protein, increased IGF2 translation and IGF1 receptor (IGF1R) signaling via PI3K and MAPK cascades, and promotion of cell proliferation, invasion, and transformation. THADA fusions and IGF2BP3 overexpression are found in â¼5% of thyroid cancers that lack any other driver mutations. We also find that strong IGF2BP3 overexpression via gene fusion, amplification, or other mechanisms occurs in 5 to 15% of several other cancer types. Finally, we provide in vitro and in vivo evidence that growth of IGF2BP3-driven cells and tumors may be blocked by IGF1R inhibition, raising the possibility that IGF2BP3 overexpression in cancer cells may predict an anti-IGF1R benefit.
Assuntos
Regulação Neoplásica da Expressão Gênica , Proteínas de Neoplasias/genética , Proteínas de Fusão Oncogênica/genética , Proteínas de Ligação a RNA/genética , Receptores de Somatomedina/genética , Neoplasias da Glândula Tireoide/genética , Animais , Antineoplásicos/farmacologia , Linhagem Celular Tumoral , Transformação Celular Neoplásica , Feminino , Loci Gênicos , Estudo de Associação Genômica Ampla , Humanos , Imidazóis/farmacologia , Fator de Crescimento Insulin-Like II/genética , Fator de Crescimento Insulin-Like II/metabolismo , Camundongos , Camundongos Nus , Proteínas Quinases Ativadas por Mitógeno/genética , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Proteínas de Neoplasias/metabolismo , Proteínas de Fusão Oncogênica/metabolismo , Fosfatidilinositol 3-Quinases/genética , Fosfatidilinositol 3-Quinases/metabolismo , Biossíntese de Proteínas , Inibidores de Proteínas Quinases/farmacologia , Pirazinas/farmacologia , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Proteínas de Ligação a RNA/metabolismo , Receptor IGF Tipo 1 , Receptores de Somatomedina/antagonistas & inibidores , Receptores de Somatomedina/metabolismo , Transdução de Sinais , Neoplasias da Glândula Tireoide/tratamento farmacológico , Neoplasias da Glândula Tireoide/metabolismo , Neoplasias da Glândula Tireoide/patologia , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Human polyomavirus 7 (HPyV7) is one of 11 HPyVs recently discovered through genomic sequencing technologies. Two lung transplant recipients receiving immunosuppressive therapy developed pruritic, brown plaques on the trunk and extremities showing a distinctive epidermal hyperplasia with virus-laden keratinocytes containing densely packed 36-45-nm icosahedral capsids. Rolling circle amplification and gradient centrifugation testing were positive for encapsidated HPyV7 DNA in skin and peripheral blood specimens from both patients, and HPyV7 early and capsid proteins were abundantly expressed in affected tissues. We describe for the first time that HPyV7 is associated with novel pathogenicity in some immunosuppressed individuals.
Assuntos
Infecções por Polyomavirus/patologia , Infecções por Polyomavirus/virologia , Polyomavirus/isolamento & purificação , Transplantados , Infecções Tumorais por Vírus/patologia , Infecções Tumorais por Vírus/virologia , Idoso , Sangue/virologia , Exantema/patologia , Exantema/virologia , Histocitoquímica , Humanos , Hospedeiro Imunocomprometido , Imuno-Histoquímica , Masculino , Microscopia Eletrônica de Transmissão , Pele/patologia , Pele/virologia , ViremiaRESUMO
BRAF V600E mutation is a driver mutation in the serrated pathway to colorectal cancers. BRAFV600E drives tumorigenesis through constitutive downstream extracellular signal-regulated kinase (ERK) activation, but high-intensity ERK activation can also trigger tumor suppression. Whether and how oncogenic ERK signaling can be intrinsically adjusted to a "just-right" level optimal for tumorigenesis remains undetermined. In this study, we found that FAK (Focal adhesion kinase) expression was reduced in BRAFV600E-mutant adenomas/polyps in mice and patients. In Vill-Cre;BRAFV600E/+;Fakfl/fl mice, Fak deletion maximized BRAFV600E's oncogenic activity and increased cecal tumor incidence to 100%. Mechanistically, our results showed that Fak loss, without jeopardizing BRAFV600E-induced ERK pathway transcriptional output, reduced EGFR (epidermal growth factor receptor)-dependent ERK phosphorylation. Reduction in ERK phosphorylation increased the level of Lgr4, promoting intestinal stemness and cecal tumor formation. Our findings show that a "just-right" ERK signaling optimal for BRAFV600E-induced cecal tumor formation can be achieved via Fak loss-mediated downregulation of ERK phosphorylation.
RESUMO
BRAFV600E mutation is a driver mutation in the serrated pathway to colorectal cancers. BRAFV600E drives tumorigenesis through constitutive downstream extracellular signal-regulated kinase (ERK) activation, but high-intensity ERK activation can also trigger tumor suppression. Whether and how oncogenic ERK signaling can be intrinsically adjusted to a 'just-right' level optimal for tumorigenesis remains undetermined. In this study, we found that FAK (Focal adhesion kinase) expression was reduced in BRAFV600E-mutant adenomas/polyps in mice and patients. In Vil1-Cre;BRAFLSL-V600E/+;Ptk2fl/fl mice, Fak deletion maximized BRAFV600E's oncogenic activity and increased cecal tumor incidence to 100%. Mechanistically, our results showed that Fak loss, without jeopardizing BRAFV600E-induced ERK pathway transcriptional output, reduced EGFR (epidermal growth factor receptor)-dependent ERK phosphorylation. Reduction in ERK phosphorylation increased the level of Lgr4, promoting intestinal stemness and cecal tumor formation. Our findings show that a 'just-right' ERK signaling optimal for BRAFV600E-induced cecal tumor formation can be achieved via Fak loss-mediated downregulation of ERK phosphorylation.
Assuntos
Neoplasias do Ceco , Quinase 1 de Adesão Focal , Proteínas Proto-Oncogênicas B-raf , Animais , Proteínas Proto-Oncogênicas B-raf/metabolismo , Proteínas Proto-Oncogênicas B-raf/genética , Fosforilação , Camundongos , Humanos , Neoplasias do Ceco/metabolismo , Neoplasias do Ceco/genética , Neoplasias do Ceco/patologia , Quinase 1 de Adesão Focal/metabolismo , Quinase 1 de Adesão Focal/genética , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , MAP Quinases Reguladas por Sinal Extracelular/genética , Sistema de Sinalização das MAP Quinases , Receptores ErbB/metabolismo , Receptores ErbB/genética , Carcinogênese/genética , Carcinogênese/metabolismo , Receptores Acoplados a Proteínas G/metabolismo , Receptores Acoplados a Proteínas G/genética , MasculinoRESUMO
Pancreatic lymphoepithelial cysts are rare benign cysts that cannot be reliably differentiated from neoplastic mucinous cysts preoperatively. Although elevated cyst fluid carcinoembryonic antigen (CEA) levels support a diagnosis of a mucinous cyst, the finding of increased CEA levels in lymphoepithelial cysts prompted this study. Nine resected lymphoepithelial cysts were examined for expression of CEA, carbohydrate antigen (CA) 19-9, CK7, p63, PAS-D and a panel of mucins. The pathology data were correlated with clinical information, including serum, cyst fluid and imaging studies. By computed tomography scan, although most lymphoepithelial cysts appeared cystic, 23% were described as masses. The endoscopic ultrasound findings were variable, but the lymphoepithelial cysts tended to be hypoechoic cystic lesions or masses. On cytology, 44% of the cysts had squamous cells, 67% had glandular cells and 56% had atypical cells. The cysts were resected because of size ≥3 cm (89%), symptoms (44%) and/or elevated cyst fluid CEA levels (33%). The cyst fluid CEA levels in the three cysts tested were >450 ng/ml. Histopathologically, all cysts were lined by mature, stratified squamous-type cells and produced keratin. Mucous cells were present in 78% of the cysts. The immunohistochemical profile of the squamous lining was CK7+, p63+, MUC1+, MUC4+, MUC2-, MUC5AC- and MUC6-. Even though lymphoepithelial cysts are lined by squamous-type epithelium, all our resected lymphoepithelial cysts expressed CEA and/or CA19-9, many contained mucous cells, and three exhibited markedly elevated cyst fluid CEA levels. Although cyst fluid CEA levels >200 ng/ml support the diagnosis of mucinous neoplasms, this study emphasizes the need for clinicians and pathologists to recognize that lymphoepithelial cysts can mimic neoplastic mucinous cysts clinically, radiographically and on cyst fluid CEA analysis.
Assuntos
Antígeno Carcinoembrionário/análise , Líquido Cístico/química , Células Epiteliais/química , Tecido Linfoide/química , Mucinas/análise , Neoplasias Císticas, Mucinosas e Serosas/diagnóstico , Cisto Pancreático/diagnóstico , Neoplasias Pancreáticas/diagnóstico , Adulto , Idoso , Biópsia por Agulha Fina , Antígeno CA-19-9/análise , Diagnóstico Diferencial , Endossonografia , Células Epiteliais/patologia , Feminino , Humanos , Imuno-Histoquímica , Queratina-7/análise , Tecido Linfoide/patologia , Masculino , Proteínas de Membrana/análise , Pessoa de Meia-Idade , Neoplasias Císticas, Mucinosas e Serosas/química , Neoplasias Císticas, Mucinosas e Serosas/patologia , Neoplasias Císticas, Mucinosas e Serosas/cirurgia , Cisto Pancreático/química , Cisto Pancreático/patologia , Cisto Pancreático/cirurgia , Neoplasias Pancreáticas/química , Neoplasias Pancreáticas/patologia , Neoplasias Pancreáticas/cirurgia , Pennsylvania , Valor Preditivo dos Testes , Cuidados Pré-Operatórios , Tomografia Computadorizada por Raios XRESUMO
BACKGROUND & AIMS: Identification and validation of new functionally relevant and pharmacologically actionable targets for pancreatic ductal adenocarcinoma (PDAC) remains a great challenge. Premalignant acinar cell reprogramming (acinar-to-ductal metaplasia [ADM]) is a precursor of pancreatic intraepithelial neoplasia (PanIN) lesions that can progress to PDAC. This study investigated the role of proline-rich tyrosine kinase 2 (PYK2) in mutant Kras-induced and pancreatitis-associated ADM and PanIN formation, as well as in PDAC maintenance. METHODS: Genetically engineered mouse models of mutant Kras (glycine 12 to aspartic acid) and Pyk2 deletion were used for investigating the role of PYK2 in PDAC genesis in mice. In vitro ADM assays were conducted using primary pancreatic acinar cells isolated from mice. Immunohistochemistry, immunofluorescence, and a series of biochemical experiments were used to investigate upstream regulators/downstream targets of PYK2 in pancreatic carcinogenesis. PDAC cell line xenograft experiments were performed to study the role of PYK2 and its downstream target in PDAC maintenance. RESULTS: PYK2 was increased substantially in ADM lesions induced by mutant Kras or inflammatory injury. Pyk2 deletion remarkably suppressed ADM and PanIN formation in a mutant Kras-driven and pancreatitis-associated PDAC model, whereas PYK2 knockdown substantially inhibited PDAC cell growth in vitro and in nude mice. This study uncovered a novel yes-associated protein 1/transcriptional co-activator with PDZ binding motif/signal transducer and activator of transcription 3/PYK2/ß-catenin regulation axis in PDAC. Our results suggest that PYK2 contributes to PDAC genesis and maintenance by activating the Wnt/ß-catenin pathway through directly phosphorylating ß-cateninY654. CONCLUSIONS: The current study uncovers PYK2 as a novel downstream effector of mutant KRAS signaling, a previously unrecognized mediator of pancreatitis-induced ADM and a novel intervention target for PDAC.
Assuntos
Carcinoma de Células Acinares/metabolismo , Carcinoma Ductal Pancreático/metabolismo , Quinase 2 de Adesão Focal/metabolismo , Neoplasias Pancreáticas/metabolismo , Lesões Pré-Cancerosas/metabolismo , beta Catenina/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Animais , Carcinoma in Situ/genética , Carcinoma in Situ/metabolismo , Carcinoma in Situ/patologia , Carcinoma de Células Acinares/genética , Carcinoma de Células Acinares/patologia , Carcinoma Ductal Pancreático/genética , Carcinoma Ductal Pancreático/patologia , Proteínas de Ciclo Celular/metabolismo , Reprogramação Celular/fisiologia , Modelos Animais de Doenças , Feminino , Quinase 2 de Adesão Focal/genética , Masculino , Metaplasia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Nus , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/patologia , Fosforilação , Lesões Pré-Cancerosas/genética , Lesões Pré-Cancerosas/patologia , Proteínas Proto-Oncogênicas p21(ras)/metabolismo , Via de Sinalização Wnt , Proteínas de Sinalização YAPRESUMO
One of the most encouraging developments in oncology has been the success of BRAF inhibitors in BRAF-mutant melanoma. However, in contrast to its striking efficacy in BRAF-mutant melanomas, BRAF inhibitor monotherapy is ineffective in BRAF-mutant colorectal cancer. Although many studies on BRAF inhibitor resistance in colorectal cancer have focused on mechanisms underlying the reactivation of the EGFR/RAS/RAF/MEK/ERK pathway, the current study focuses on identifying novel adaptive signaling mechanisms, a fresh angle on colorectal cancer resistance to BRAF inhibition. We found that treatment with BRAF inhibitors (both current and next-generation BRAF inhibitors) upregulated the Wnt/ß-catenin pathway in BRAFV600E-mutant colorectal cancer cell lines through activating the cytoplasmic tyrosine kinase focal adhesion kinase (FAK). The results showed that FAK activation upon BRAF inhibitor treatment did not require EGFR or ERK1/2 activation, implying that BRAF inhibitor treatment-induced hyperactivation of Wnt signaling is "pathway reactivation"-independent. BRAF inhibition-induced Wnt pathway activation was further validated in preclinical models of BRAFV600E-mutant colorectal cancer, including cell line xenograft model and a patient-derived xenograft model. Combined inhibition of BRAF/Wnt pathways or BRAF/FAK pathways exerted strong synergistic antitumor effects in cell culture model and mouse xenograft model. Overall, the current study has identified activation of the Wnt/ß-catenin pathway as a novel fundamental cause of colon cancer resistance to BRAF inhibition. Our results suggest that although complete vertical pathway blockade is pivotal for effective and durable control of BRAF-mutant colorectal cancer, cotargeting parallel adaptive signaling-the Wnt/ß-catenin pathway-is also essential. Mol Cancer Ther; 17(4); 806-13. ©2017 AACR.
Assuntos
Neoplasias Colorretais/patologia , Resistencia a Medicamentos Antineoplásicos , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Inibidores de Proteínas Quinases/farmacologia , Proteínas Proto-Oncogênicas B-raf/antagonistas & inibidores , Proteína Wnt1/metabolismo , beta Catenina/metabolismo , Animais , Apoptose , Proliferação de Células , Neoplasias Colorretais/tratamento farmacológico , Neoplasias Colorretais/genética , Humanos , Masculino , Camundongos , Camundongos Nus , Mutação , Proteínas Proto-Oncogênicas B-raf/genética , Células Tumorais Cultivadas , Proteína Wnt1/genética , Ensaios Antitumorais Modelo de Xenoenxerto , beta Catenina/genéticaRESUMO
Universal screening using immunohistochemistry for DNA mismatch-repair proteins (MLH1, MSH2, MSH6, and PMS2) is advocated by major professional medical organizations to identify Lynch syndrome-associated colorectal carcinoma. Loss of MSH6 expression independent of MSH2 expression has been reported in microsatellite-stable (MSS) colorectal carcinoma after neoadjuvant therapy. The mechanism remains unclear. We studied the immunohistochemical expression of MSH2, MSH6, and Ki-67 in MSS colorectal carcinoma with (n=50) or without (n=64) preoperative neoadjuvant therapy and Lynch syndrome-associated colorectal carcinoma with confirmed MSH6 germline mutation (n=3). Twelve of 50 MSS colorectal carcinoma postneoadjuvant resections demonstrated reduced MSH6 expression, with loss of expression ranging from 20% to 100% of tumor cells. Eight of 64 MSS colorectal carcinomas without neoadjuvant therapy also exhibited reduced MSH6 expression but to a lesser degree (10%-50% of tumor cells with loss of expression). In both subgroups, concomitant loss of MSH6 and Ki-67 expressions was demonstrated in the same tumor areas in consecutive tissue sections. However, 3 cases of Lynch syndrome-associated colorectal carcinoma due to germline MSH6 mutation revealed complete loss of MSH6 expression with discordant positive Ki-67 staining in the tumor cells. The MSH2-independent, Ki-67-related expression of MSH6 in colorectal carcinoma helps to explain the heterogeneous MSH6 staining in MSS colorectal carcinoma with or without neoadjuvant therapy.
Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Quimiorradioterapia Adjuvante , Neoplasias Colorretais Hereditárias sem Polipose/terapia , Neoplasias Colorretais/terapia , Proteínas de Ligação a DNA/metabolismo , Antígeno Ki-67/metabolismo , Repetições de Microssatélites , Terapia Neoadjuvante , Adulto , Idoso , Biópsia , Neoplasias Colorretais/química , Neoplasias Colorretais/genética , Neoplasias Colorretais/patologia , Neoplasias Colorretais Hereditárias sem Polipose/química , Neoplasias Colorretais Hereditárias sem Polipose/genética , Neoplasias Colorretais Hereditárias sem Polipose/patologia , Regulação para Baixo , Feminino , Mutação em Linhagem Germinativa , Humanos , Imuno-Histoquímica , Masculino , Pessoa de Meia-Idade , Proteína 2 Homóloga a MutS/análise , Proteína 2 Homóloga a MutS/genética , Resultado do TratamentoRESUMO
Pancreatic cysts >1 cm lined by nonpapillary mucinous epithelium without ovarian-type stroma pose diagnostic challenges. The term "simple mucinous cyst" was recently proposed for this entity. Our goal was to determine the clinicopathologic characteristics of these cysts, as they have not been previously described. Of the 39 patients with pancreatic resections included in this study, the mean age was 65 years and the female-to-male ratio was 4:1. The characteristics of the cysts are as follows: 82% had elevated cyst fluid carcinoembryonic antigen levels, 67% were unilocular, 69% occurred in the body/tail, 92% did not communicate with pancreatic ducts, the mean size was 2.4 cm (range, 1.0 to 5.5 cm), the cyst contents tended to be serous (48%) or viscous (28%), all had a smooth lining (only 1 had focal excrescences) composed of bland columnar mucinous epithelium (low-grade dysplasia) in 92% with focal high-grade dysplasia in 8%, and 65% had degenerative changes (granulation-like tissue, hemorrhage, and myxoid stroma). The cyst lining was CK7+ and 97% had a MUC5AC+ and/or MUC6+ gastric phenotype; overt intestinal features were absent. In total, 55% of cysts tested (fluid and/or resections) harbored KRAS mutations. The term "simple mucinous cyst" is useful to apply to >1 cm mucinous cysts that do not have characteristic features of intraductal papillary mucinous neoplasms or mucinous cystic neoplasms. KRAS mutations can be detected in these typically bland cysts, and in rare instances, focal high-grade dysplasia may be present. Hence, these cysts should be viewed as neoplastic and treated similarly to other mucinous pancreatic cysts.
Assuntos
Cisto Pancreático/diagnóstico , Cisto Pancreático/patologia , Pancreatopatias/diagnóstico , Pancreatopatias/patologia , Adenocarcinoma Papilar/diagnóstico , Adenocarcinoma Papilar/genética , Adenocarcinoma Papilar/patologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Biomarcadores Tumorais/análise , Carcinoma Ductal Pancreático/diagnóstico , Carcinoma Ductal Pancreático/genética , Carcinoma Ductal Pancreático/patologia , Cistadenocarcinoma Papilar/diagnóstico , Cistadenocarcinoma Papilar/genética , Cistadenocarcinoma Papilar/patologia , Análise Mutacional de DNA , Diagnóstico Diferencial , Feminino , Humanos , Imuno-Histoquímica , Masculino , Pessoa de Meia-Idade , Cisto Pancreático/genética , Pancreatopatias/genética , Neoplasias Pancreáticas/diagnóstico , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/patologia , Reação em Cadeia da Polimerase , Proteínas Proto-Oncogênicas p21(ras)/genética , Adulto JovemRESUMO
About 76% of patients with lung adenocarcinoma harbor activating mutations in the receptor tyrosine kinase (RTK)/RAS/RAF pathways, leading to aberrant activation of the mitogen-activated protein kinase (MAPK) pathways particularly the MAPK/ERK pathway. However, many lung adenocarcinomas lacking these genomic mutations also display significant MAPK pathway activation, suggesting that additional MAPK pathway alterations remain undetected. This study has identified serine/threonine kinase mitogen-activated protein 4 kinase 4 (MAP4K4) as a novel positive regulator of MAPK/ERK signaling in lung adenocarcinoma. The results showed that MAP4K4 was drastically elevated in lung adenocarcinoma independently of KRAS or EGFR mutation status. Knockdown of MAP4K4 inhibited proliferation, anchorage-independent growth and migration of lung adenocarcinoma cells, and also inhibited human lung adenocarcinoma xenograft growth and metastasis. Mechanistically, we found that MAP4K4 activated ERK through inhibiting protein phosphatase 2 activity. Our results further showed that downregulation of MAP4K4 prevented ERK reactivation in EGFR inhibitor erlotinib-treated lung adenocarcinoma cells. Together, our findings identify MAP4K4 as a novel MAPK/ERK pathway regulator in lung adenocarcinoma that is required for lung adenocarcinoma maintenance.
Assuntos
Adenocarcinoma/enzimologia , Adenocarcinoma/patologia , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Neoplasias Pulmonares/enzimologia , Neoplasias Pulmonares/patologia , Quinases de Proteína Quinase Ativadas por Mitógeno/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Adenocarcinoma de Pulmão , Animais , Movimento Celular/efeitos dos fármacos , Movimento Celular/genética , Proliferação de Células/efeitos dos fármacos , Proliferação de Células/genética , Regulação para Baixo , Receptores ErbB/genética , Receptores ErbB/metabolismo , Cloridrato de Erlotinib/farmacologia , Técnicas de Silenciamento de Genes , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/genética , Camundongos , Camundongos Nus , Quinases de Proteína Quinase Ativadas por Mitógeno/antagonistas & inibidores , Inibidores de Proteínas Quinases/farmacologia , Proteína Fosfatase 2/antagonistas & inibidores , Proteína Fosfatase 2/genética , Proteína Fosfatase 2/metabolismo , Proteínas Serina-Treonina Quinases/genética , Proteínas Proto-Oncogênicas p21(ras)/genética , Proteínas Proto-Oncogênicas p21(ras)/metabolismo , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Most gastrointestinal stromal tumors (GISTs) are caused by activating mutations of the KIT receptor tyrosine kinase. The small molecule inhibitor imatinib mesylate was initially developed to target the ABL1 kinase, which is constitutively activated through chromosomal translocation in BCR-ABL1-positive chronic myeloid leukemia. Because of cross-reactivity of imatinib against the KIT kinase, the drug is also successfully used for the treatment of GIST. Although inhibition of KIT clearly has a major role in the therapeutic response of GIST to imatinib, the contribution of concomitant inhibition of ABL in this context has never been explored. We show here that ABL1 is expressed in the majority of GISTs, including human GIST cell lines. Using siRNA-mediated knockdown, we demonstrate that depletion of KIT in conjunction with ABL1 - hence mimicking imatinib treatment - leads to reduced apoptosis induction and attenuated inhibition of cellular proliferation when compared to depletion of KIT alone. These results are explained by an increased activity of the AKT survival kinase, which is mediated by the cyclin-dependent kinase CDK2, likely through direct phosphorylation. Our results highlight that distinct inhibitory properties of targeted agents can impede antitumor effects and hence provide insights for rational drug development. Novel KIT-targeted agents to treat GIST should therefore comprise an increased specificity for KIT while at the same time displaying a reduced ability to inhibit ABL1.
Assuntos
Neoplasias Gastrointestinais/metabolismo , Tumores do Estroma Gastrointestinal/metabolismo , Mesilato de Imatinib/farmacologia , Inibidores de Proteínas Quinases/farmacologia , Proteínas Proto-Oncogênicas c-abl/metabolismo , Proteínas Proto-Oncogênicas c-kit/metabolismo , Adulto , Idoso , Idoso de 80 Anos ou mais , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Feminino , Neoplasias Gastrointestinais/tratamento farmacológico , Tumores do Estroma Gastrointestinal/tratamento farmacológico , Regulação Neoplásica da Expressão Gênica , Humanos , Masculino , Pessoa de Meia-Idade , Proteínas Proto-Oncogênicas c-abl/genética , Proteínas Proto-Oncogênicas c-kit/genética , RNA Interferente Pequeno/farmacologia , Análise Serial de TecidosRESUMO
Between 10% and 15% of colorectal carcinomas demonstrate sporadic DNA mismatch-repair protein deficiency as a result of MLH1 promoter methylation and are thought to arise from sessile serrated adenomas, termed the serrated neoplasia pathway. Although the presence of the BRAF V600E mutation is indicative of a sporadic cancer, up to 30% to 50% of colorectal carcinomas with MLH1 promoter hypermethylation will lack a BRAF mutation. We report the clinicopathologic and molecular features of MLH1-deficient colorectal carcinoma with wild-type BRAF and MLH1 promoter hypermethylation (referred to as MLH1-hypermethylated BRAF wild-type colorectal carcinoma, n=36) in comparison with MLH1-deficient BRAF-mutated colorectal carcinoma (n=113) and Lynch syndrome-associated colorectal carcinoma (n=36). KRAS mutations were identified in 31% of MLH1-hypermethylated BRAF wild-type colorectal carcinomas compared with 0% of MLH1-deficient BRAF-mutated colorectal carcinomas and 37% of Lynch syndrome-associated colorectal carcinomas. When a precursor polyp was identified, MLH1-hypermethylated BRAF wild-type colorectal carcinomas arose from precursor polyps resembling conventional tubular/tubulovillous adenomas in contrast to MLH1-deficient BRAF-mutated colorectal carcinomas, which arose from precursor sessile serrated adenomas (P<0.001). Both MLH1-hypermethylated BRAF wild-type colorectal carcinoma and MLH1-deficient BRAF-mutated colorectal carcinoma had a predilection for the right colon compared with Lynch syndrome-associated colorectal carcinoma (86% vs. 92% vs. 49%, P<0.001). There was no significant difference in mucinous differentiation, tumor-infiltrating lymphocytes, Crohn-like reaction, and medullary differentiation between the 3 tumor groups. Using Kaplan-Meier survival functions, there was no significant difference in disease-specific survival between the 3 patient groups (P>0.05). In conclusion, our results indicate that MLH1-hypermethylated BRAF wild-type colorectal carcinomas can harbor KRAS mutations and arise from precursor polyps resembling conventional tubular/tubulovillous adenomas.
Assuntos
Adenocarcinoma/genética , Adenoma/genética , Neoplasias Colorretais/genética , Metilação de DNA , Proteína 1 Homóloga a MutL/deficiência , Proteínas Proto-Oncogênicas B-raf/genética , Proteínas Proto-Oncogênicas p21(ras)/genética , Adenocarcinoma/mortalidade , Adenocarcinoma/patologia , Adenoma/mortalidade , Adenoma/patologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Biomarcadores Tumorais/deficiência , Biomarcadores Tumorais/genética , Estudos de Casos e Controles , Neoplasias Colorretais/mortalidade , Neoplasias Colorretais/patologia , Neoplasias Colorretais Hereditárias sem Polipose/genética , Bases de Dados Factuais , Feminino , Seguimentos , Humanos , Imuno-Histoquímica , Estimativa de Kaplan-Meier , Masculino , Instabilidade de Microssatélites , Pessoa de Meia-Idade , Proteína 1 Homóloga a MutL/genética , Mutação , Regiões Promotoras GenéticasRESUMO
The molecular alterations in colorectal poorly differentiated neuroendocrine carcinoma remain incompletely characterized, particularly with respect to mutations in BRAF and KRAS. We analyzed 32 colorectal poorly differentiated neuroendocrine carcinomas and 40 colorectal poorly differentiated conventional adenocarcinomas for mutations in KRAS and BRAF and for DNA mismatch repair protein abnormalities to correlate histopathology with molecular alterations and survival. Compared with poorly differentiated conventional adenocarcinoma, poorly differentiated neuroendocrine carcinoma frequently harbored BRAF mutations (59% versus 5%; P < .001) and less frequently demonstrated KRAS codon 12 or 13 mutations (17% versus 43%; P = .03). BRAF mutations were identified in both pure poorly differentiated neuroendocrine carcinoma (60%) and poorly differentiated neuroendocrine carcinoma associated with a signet ring cell adenocarcinoma component (82%). Most (93%) poorly differentiated neuroendocrine carcinomas demonstrated proficient DNA mismatch repair by either microsatellite instability polymerase chain reaction or DNA mismatch repair immunohistochemistry. Patients with poorly differentiated neuroendocrine carcinoma had a significantly worse overall survival compared with patients with poorly differentiated conventional adenocarcinoma (P < .001). There was no significant difference in overall survival between patients with pure poorly differentiated neuroendocrine carcinoma and patients with both poorly differentiated neuroendocrine carcinoma and adenocarcinoma components (P = .5). In conclusion, colorectal poorly differentiated neuroendocrine carcinomas frequently harbor BRAF mutations and are associated with poor overall survival.
Assuntos
Adenocarcinoma/genética , Biomarcadores Tumorais/genética , Carcinoma Neuroendócrino/genética , Diferenciação Celular , Neoplasias Colorretais/genética , Mutação , Proteínas Proto-Oncogênicas B-raf/genética , Adenocarcinoma/enzimologia , Adenocarcinoma/mortalidade , Adenocarcinoma/patologia , Adenocarcinoma/terapia , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Biomarcadores Tumorais/análise , Biópsia , Carcinoma Neuroendócrino/enzimologia , Carcinoma Neuroendócrino/mortalidade , Carcinoma Neuroendócrino/patologia , Carcinoma Neuroendócrino/terapia , Neoplasias Colorretais/enzimologia , Neoplasias Colorretais/mortalidade , Neoplasias Colorretais/patologia , Neoplasias Colorretais/terapia , Reparo de Erro de Pareamento de DNA , Análise Mutacional de DNA , Feminino , Predisposição Genética para Doença , Humanos , Imuno-Histoquímica , Estimativa de Kaplan-Meier , Masculino , Instabilidade de Microssatélites , Pessoa de Meia-Idade , Fenótipo , Modelos de Riscos Proporcionais , Proteínas Proto-Oncogênicas p21(ras)/genética , Fatores de Tempo , Resultado do Tratamento , Adulto JovemRESUMO
Aberrant activation of Wnt/ß-catenin signaling plays an unequivocal role in colorectal cancer, but identification of effective Wnt inhibitors for use in cancer remains a tremendous challenge. New insights into the regulation of this pathway could reveal new therapeutic point of intervention, therefore are greatly needed. Here we report a novel FAK/PYK2/GSK3ß(Y216)/ß-catenin regulation axis: FAK and PYK2, elevated in adenomas in APC(min/+) mice and in human colorectal cancer tissues, functioned redundantly to promote the Wnt/ß-catenin pathway by phosphorylating GSK3ß(Y216) to reinforce pathway output-ß-catenin accumulation and intestinal tumorigenesis. We previously showed that Wnt-induced ß-catenin accumulation requires Wnt-induced GSK3ß/ß-TrCP interaction; the current study revealed that phosphorylation of GSK3ß(Y216) was a molecular determinant of GSK3ß recruitment of ß-TrCP. Pharmacological inhibition of FAK/PYK2 suppressed adenoma formation in APC(min/+) mice accompanied with reduced intestinal levels of phospho-GSK3ß(Y216) and ß-catenin, indicating that FAK/PYK2/GSK3ß(Y216) axis is critical for the activation of Wnt/ß-catenin signaling in APC driven intestinal tumorigenesis.
Assuntos
Carcinogênese , Quinase 1 de Adesão Focal/metabolismo , Quinase 2 de Adesão Focal/metabolismo , Quinase 3 da Glicogênio Sintase/metabolismo , Via de Sinalização Wnt , Animais , Neoplasias Colorretais/patologia , Feminino , Regulação da Expressão Gênica , Glicogênio Sintase Quinase 3 beta , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Camundongos NusRESUMO
Anomalous expression of certain types of mucins occurs in pancreatic tumors, but little is known about the causes. MUC2 and MUC5AC are not expressed in normal pancreas. In the present study an immunohistochemical screening showed that MUC2 antigen was expressed in 6% of invasive tumors. Of the pancreatic cell lines, 2.4% expressed MUC2 message and antigen. In contrast, MUC5AC antigen was expressed in 86-100% of invasive adenocarcinoma (depending on the antibody). MUC5AC message and antigen were expressed in 66.7% of the cell lines tested. A polymerase chain reaction based assay was used to determine if methylation of CpG sites immediately 5' of the transcription initiation sites of the MUC2 and MUC5AC genes could be related to mucin expression in the cell lines. Digestibility by the methylation sensitive restriction enzyme HpaII correlated with the presence or absence of mRNA in 100% and 77.8% of the cell lines for MUC2 and MUC5AC, respectively. Treatment of a cell line that did not express MUC2 or MUC5AC gene products with the DNA methyltransferase inhibitor 5-aza-2'-deoxycytidine resulted in an increase in MUC2 message but no change in MUC5AC message. In summary, the expression of MUC2 gene products correlated well with methylation of the proximal region of the promoter whereas expression of MUC5AC may involve additional regions or other mechanisms.
Assuntos
Adenocarcinoma Mucinoso/patologia , Adenocarcinoma/patologia , Azacitidina/análogos & derivados , Metilação de DNA , DNA de Neoplasias/química , Regulação Neoplásica da Expressão Gênica , Mucinas/genética , Proteínas de Neoplasias/genética , Neoplasias Pancreáticas/patologia , Regiões Promotoras Genéticas , Adenocarcinoma/genética , Adenocarcinoma Mucinoso/genética , Azacitidina/farmacologia , Sequência de Bases , DNA (Citosina-5-)-Metiltransferases/antagonistas & inibidores , Metilação de DNA/efeitos dos fármacos , DNA de Neoplasias/genética , Decitabina , Inibidores Enzimáticos/farmacologia , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Dados de Sequência Molecular , Mucina-5AC , Mucina-2 , Mucinas/biossíntese , Invasividade Neoplásica , Proteínas de Neoplasias/antagonistas & inibidores , Proteínas de Neoplasias/biossíntese , Pâncreas/metabolismo , Neoplasias Pancreáticas/genética , Regiões Promotoras Genéticas/genética , RNA Mensageiro/biossíntese , RNA Neoplásico/biossínteseRESUMO
Syndromic and sporadic fundic gland polyps are morphologically indistinguishable but may arise via different pathogenetic mechanisms involving mutations of the adenomatous polyposis coli (APC) and its downstream target beta-catenin genes. Although a higher frequency of dysplasia has been reported in syndromic forms, the risk of developing invasive carcinoma is exceedingly low. The current study was designed to investigate whether syndromic and sporadic fundic gland polyps differ in protein expression of a number of genes that are thought to be important in the control of neoplastic transformation. A total of 262 fundic gland polyps, including 155 syndromic polyps obtained from 35 patients with familial adenomatous polyposis or Gardner's syndrome and 107 sporadic polyps randomly selected from 45 patients with gastroesophageal reflux disease or Barrett's esophagus, were included in this study. Immunohistochemical evaluation showed that loss of immunoreactivity to the antibody against the carboxyl terminus of the APC protein, presumably resulting from APC gene mutations, was more frequent in syndromic than in sporadic cases (40% versus 6.7%, P<0.001). However, immunostaining failed to show aberrant nuclear localization of beta-catenin, a protein regulated by APC, in any of the polyps, irrespective of syndromic or sporadic types. Instead, positive membranous staining for beta-catenin was observed in all the cases. In addition, the expression characteristics of 2 other proteins, c-Myc and cyclin D1, whose genes have been reported to be transcriptionally regulated by the APC/beta-catenin pathway, were similar in these two types of polyps. Furthermore, all cases, including those harboring dysplasia, showed negative nuclear staining for p53 and positive nuclear staining for retinoblastoma (RB). Taken together, these data show a lack of dysregulation in the APC/beta-catenin signaling pathway and in the expression of p53 and RB in fundic gland polyps despite a high frequency of somatic mutations of the APC and beta-catenin genes reported in these polyps. These findings may explain at least in part why fundic gland polyps show a negligible malignant potential even in the presence of dysplasia.
Assuntos
Biomarcadores Tumorais/metabolismo , Fundo Gástrico/metabolismo , Proteínas de Neoplasias/metabolismo , Pólipos/metabolismo , Neoplasias Gástricas/metabolismo , Proteína da Polipose Adenomatosa do Colo/metabolismo , Adolescente , Adulto , Idoso , Núcleo Celular/metabolismo , Núcleo Celular/patologia , Ciclina D1/metabolismo , Proteínas do Citoesqueleto/metabolismo , Feminino , Fundo Gástrico/patologia , Humanos , Técnicas Imunoenzimáticas , Masculino , Pessoa de Meia-Idade , Pólipos/patologia , Proteínas Proto-Oncogênicas c-myc/metabolismo , Proteína do Retinoblastoma/metabolismo , Neoplasias Gástricas/patologia , Síndrome , Transativadores/metabolismo , Proteína Supressora de Tumor p53/metabolismo , beta CateninaRESUMO
The BRAF V600E mutation occurs in 15% of colorectal carcinomas (CRCs) and has important genetic, prognostic, and therapeutic implications. A monoclonal antibody (VE1) targeting the BRAF V600E mutant protein has become available with variable efficacy in literature reports. We investigated the utility of the VE1 antibody in detecting BRAF V600E mutant protein in two cohorts: (1) a retrospectively accrued series of 103 resected CRCs with (N = 57) and without (N = 46) known BRAF V600E mutation status by PCR and (2) a prospective series of 25 CRCs requiring BRAF analysis during routine screening for Lynch syndrome. All 74 cases with positive BRAF V600E mutation demonstrated cytoplasmic positivity with the VE1 antibody with most tumors (70/74, 95%) demonstrating moderate to strong staining. Of the 54 BRAF V600E-negative cases, 51/54 CRCs (94%) were negative with the VE1 antibody while 3 CRCs (6%) demonstrated weak cytoplasmic staining. The sensitivity and specificity of VE1 was 100% and 94%, respectively. Ten BRAF V600E-mutated CRCs had adjacent precursor lesions including 7 sessile serrated adenomas associated with CRCs with high-level microsatellite instability (MSI-H). All 10 precursor adenomas were positive for VE1 staining with the 7 sessile serrated adenomas maintaining preserved MLH1 expression. Our results indicate that VE1 immunohistochemistry is a useful surrogate for the detection of the BRAF V600E mutation in CRC, although weak staining must be evaluated by BRAF PCR analysis to exclude a false positive result. In addition, the BRAF V600E mutation appears to be an early event before the divergent development into MSS and MSI-H pathways.
Assuntos
Adenoma/diagnóstico , Neoplasias Colorretais Hereditárias sem Polipose/diagnóstico , Neoplasias Colorretais/diagnóstico , Imuno-Histoquímica/métodos , Proteínas Mutantes/metabolismo , Proteínas Proto-Oncogênicas B-raf/metabolismo , Adenoma/metabolismo , Adenoma/patologia , Idoso , Idoso de 80 Anos ou mais , Anticorpos Monoclonais , Biomarcadores Tumorais/metabolismo , Neoplasias Colorretais/metabolismo , Neoplasias Colorretais/patologia , Neoplasias Colorretais Hereditárias sem Polipose/metabolismo , Neoplasias Colorretais Hereditárias sem Polipose/patologia , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Biologia MolecularRESUMO
Reduced CDX2 and cytokeratin 20 (CK20) expression in colorectal carcinoma with BRAF mutation and high-level microsatellite instability (MSI-H) has been well documented. The immunophenotype of BRAF-mutated microsatellite stable (MSS) colorectal carcinoma has not been reported. We analyzed 205 colorectal carcinomas including 28 BRAF-mutated MSS, 53 BRAF-mutated MSI-H, and 124 BRAF wild-type MSS tumors for CDX2, cytokeratin 7 (CK7), and CK20 immunohistochemical expression. CDX2 was scored semiquantitatively for both staining intensity and percent of tumor cells staining and a modified CDX2 H-score was calculated. Patients with BRAF-mutated MSS colorectal carcinomas were more frequently stage IV at presentation compared to patients with BRAF-mutated MSI-H colorectal carcinomas and BRAF wild-type MSS colorectal carcinomas (32% versus 8% versus 15%, P < .001). BRAF-mutated MSS colorectal carcinoma displayed reduced CDX2 expression compared to BRAF wild-type MSS colorectal carcinoma (75% versus 94%; mean CDX2 H-score 98 versus 150, P < .001). CK7 expression was more often identified in BRAF-mutated MSS colorectal carcinoma compared to both BRAF-mutated MSI-H colorectal carcinoma and BRAF wild-type MSS colorectal carcinoma (39% versus 6% versus 6%, P = .0001). BRAF-mutated MSI-H colorectal carcinomas were less often CK20 positive compared to BRAF-mutated MSS and BRAF wild-type MSS tumors (70% versus 93% versus 90%, P = 0.001). In summary, BRAF-mutated MSS colorectal carcinoma often displays reduced CDX2 and increased CK7 expression. Knowledge of this altered immunophenotype is important as patients with BRAF-mutated MSS colorectal carcinoma often present with metastatic disease and the altered tumor immunophenotype may lead to the erroneous assumption that origin from the colon/rectum is unlikely.
Assuntos
Adenocarcinoma/metabolismo , Neoplasias Colorretais/metabolismo , Proteínas de Homeodomínio/metabolismo , Queratina-7/metabolismo , Proteínas Proto-Oncogênicas B-raf/metabolismo , Adenocarcinoma/patologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Fator de Transcrição CDX2 , Neoplasias Colorretais/patologia , Feminino , Humanos , Imuno-Histoquímica , Queratina-20/metabolismo , Masculino , Instabilidade de Microssatélites , Repetições de Microssatélites , Pessoa de Meia-Idade , Mutação , PrognósticoRESUMO
We present a case of a 61-year-old woman with end-stage renal disease (ESRD) who developed painless hematochezia following initiation of anticoagulation. Work-up revealed a large ulceration in the sigmoid colon, and histologic images revealed sevelamer crystals embedded in the colonic mucosa, consistent with sevelamer crystal-mediated injury. This is a novel cause of gastrointestinal hemorrhage that has not previously been described in the literature. Physicians should be aware of the potential for sevelamer-induced injury.
RESUMO
Mucin core proteins (MUCs) are expressed in tissue-specific patterns in the gastrointestinal tract and expression is deregulated in Barrett's metaplasia. Based on differential expression, MUCs have been used to classify adenocarcinomas into distinct phenotypes (eg, intestinal, gastric, pancreaticobiliary, etc). Because MUC expression patterns carry prognostic significance in other tumors, we evaluated MUC expression in superficial adenocarcinomas of the gastroesophageal junction and esophagus (EAC) to determine whether there are differences in outcome associated with MUC subtype in this potentially curable subset of EAC. We classified 142 resected, superficial (T1) EAC based on their pattern of expression of MUC2, MUC5AC, MUC6 and MUC1. The association between survival and MUC expression pattern was determined in univariate and multivariate analyses. The MUC2 positive "intestinal" phenotype was associated with significantly worse prognosis in submucosal EAC (hazard ratio 2.2, 95% confidence interval 1.2-4.2), independent of node stage and other prognostic factors. MUC2 expression in submucosal EAC also showed significantly accelerated time to recurrence (hazard ratio 2.8, 95% confidence interval 1.2-6.8) after adjusting for node stage. The classification of superficial EAC by MUC protein expression has prognostic significance. MUC2 expression is an adverse prognostic indicator in submucosal EAC, independent of node stage and other prognostic factors.