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1.
Ann Gastroenterol ; 34(5): 651-659, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34475735

RESUMO

BACKGROUND: Direct-acting oral anticoagulants (DOACs) are increasingly used, with studies showing a lower risk of gastrointestinal bleeding (GIB), but overall data for GIB risk remains debatable. The objective was to assess non-fatal and fatal GIB risk in patients on DOACs compared with warfarin from randomized clinical trials (RCTs). METHODS: RCTs comparing warfarin and DOACs for various indications (atrial fibrillation, thromboembolism, insertion of mechanical heart valves) were included. The primary endpoint was any GIB event. Other clinical events, such as fatal GIB, and effects of age (≤60 years or older), time in therapeutic range for warfarin, and choice of individual DOACs on GIB risk, were also assessed. RESULTS: 14 RCTs were included, comprising 87,407 participants (DOACs n=46,223, warfarin control n=41,184). The risk of GIB with DOACs was similar to that of warfarin (relative risk [RR] 1.04, 95% confidence interval [CI] 0.85-1.27). Compared with warfarin, rivaroxaban (RR 1.23, 95%CI 1.03-1.48) and dabigatran (RR 1.38, 95%CI 1.12-1.71) had a higher risk of any GIB, whereas fatal GIB risk was lower in the DOACs group (RR 0.36, 95%CI 0.15-0.82). The risk of DOAC-related fatal GIB was lower in patients aged ≤60 years and in those with poor coagulation control (RR 0.39, 95%CI 0.15-0.98). CONCLUSIONS: DOACs compared with warfarin have a lower risk of fatal GIB, especially in those aged <60 years and those with poor coagulation control. However, the risk of GIB was comparable with warfarin and DOACs, except for rivaroxaban and dabigatran.

2.
Int J Colorectal Dis ; 34(9): 1611-1615, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31392404

RESUMO

OBJECTIVES: Study national hospitalization trends for colorectal cancer in patients younger than 50 years of age. METHODS: Patients under age 50 years hospitalized for colorectal cancer were studied using the national inpatient sample databases (2010-2014), using validated ICD-CM-9 codes and hospitalizations represented per 100,000 total inpatient population. RESULTS: Colorectal cancer hospitalizations demonstrated a significant uptrend in the 41-50 years age group, with Caucasians and females most affected, stratifying for age and excluding those with a family history of colorectal cancer (p trend < 0.001). CONCLUSIONS: Younger colorectal cancer patients aged 41-50 years (especially younger Caucasian females) are burdened with increasing hospitalization rates.


Assuntos
Neoplasias Colorretais/epidemiologia , Disparidades em Assistência à Saúde , Hospitalização/tendências , Sexismo , Adolescente , Adulto , Idade de Início , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Grupos Raciais , Estados Unidos/epidemiologia , Adulto Jovem
3.
Gastroenterology ; 149(7): 1872-1883.e9, 2015 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-26344056

RESUMO

BACKGROUND & AIMS: DNA structural lesions are prevalent in sporadic colorectal cancer. Therefore, we proposed that gene variants that predispose to DNA double-strand breaks (DSBs) would be found in patients with familial colorectal carcinomas of an undefined genetic basis (UFCRC). METHODS: We collected primary T cells from 25 patients with UFCRC and matched patients without colorectal cancer (controls) and assayed for DSBs. We performed exome sequence analyses of germline DNA from 20 patients with UFCRC and 5 undiagnosed patients with polyposis. The prevalence of identified variants in genes linked to DNA integrity was compared with that of individuals without a family history of cancer. The effects of representative variants found to be associated with UFCRC was confirmed in functional assays with HCT116 cells. RESULTS: Primary T cells from most patients with UFCRC had increased levels of the DSB marker γ(phosphorylated)histone2AX (γH2AX) after treatment with DNA damaging agents, compared with T cells from controls (P < .001). Exome sequence analysis identified a mean 1.4 rare variants per patient that were predicted to disrupt functions of genes relevant to DSBs. Controls (from public databases) had a much lower frequency of variants in the same genes (P < .001). Knockdown of representative variant genes in HCT116 CRC cells increased γH2AX. A detailed analysis of immortalized patient-derived B cells that contained variants in the Werner syndrome, RecQ helicase-like gene (WRN, encoding T705I), and excision repair cross-complementation group 6 (ERCC6, encoding N180Y) showed reduced levels of these proteins and increased DSBs, compared with B cells from controls. This phenotype was rescued by exogenous expression of WRN or ERCC6. Direct analysis of the recombinant variant proteins confirmed defective enzymatic activities. CONCLUSIONS: These results provide evidence that defects in suppression of DSBs underlie some cases of UFCRC; these can be identified by assays of circulating lymphocytes. We specifically associated UFCRC with variants in WRN and ERCC6 that reduce the capacity for repair of DNA DSBs. These observations could lead to a simple screening strategy for UFCRC, and provide insight into the pathogenic mechanisms of colorectal carcinogenesis.


Assuntos
Biomarcadores Tumorais/genética , Neoplasias Colorretais/genética , Quebras de DNA de Cadeia Dupla , Variação Genética , Linfócitos T/patologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Biomarcadores Tumorais/metabolismo , Estudos de Casos e Controles , Neoplasias Colorretais/imunologia , Neoplasias Colorretais/metabolismo , Neoplasias Colorretais/patologia , Biologia Computacional , DNA Helicases/genética , DNA Helicases/metabolismo , Reparo do DNA , Enzimas Reparadoras do DNA/genética , Enzimas Reparadoras do DNA/metabolismo , Bases de Dados Genéticas , Exodesoxirribonucleases/genética , Exodesoxirribonucleases/metabolismo , Exoma , Feminino , Frequência do Gene , Técnicas de Silenciamento de Genes , Predisposição Genética para Doença , Instabilidade Genômica , Células HCT116 , Hereditariedade , Histonas/metabolismo , Humanos , Masculino , Pessoa de Meia-Idade , Mutagênicos/farmacologia , Fenótipo , Fosforilação , Proteínas de Ligação a Poli-ADP-Ribose , RecQ Helicases/genética , RecQ Helicases/metabolismo , Análise de Sequência de DNA , Linfócitos T/efeitos dos fármacos , Linfócitos T/imunologia , Linfócitos T/metabolismo , Transfecção , Regulação para Cima , Helicase da Síndrome de Werner
4.
Aging Cell ; 14(1): 139-47, 2015 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-25481981

RESUMO

The cyclin-dependent kinase (Cdk) inhibitor p16(Ink4a) (p16) is a canonical mediator of cellular senescence and accumulates in aging tissues, where it constrains proliferation of some progenitor cells. However, whether p16 induction in tissues is sufficient to inhibit cell proliferation, mediate senescence, and/or impose aging features has remained unclear. To address these issues, we generated transgenic mice that permit conditional p16 expression. Broad induction at weaning inhibited proliferation of intestinal transit-amplifying and Lgr5+ stem cells and rapidly imposed features of aging, including hair loss, skin wrinkling, reduced body weight and subcutaneous fat, an increased myeloid fraction in peripheral blood, poor dentition, and cataracts. Aging features were observed with multiple combinations of p16 transgenes and transactivators and were largely abrogated by a germline Cdk4 R24C mutation, confirming that they reflect Cdk inhibition. Senescence markers were not found, and de-induction of p16, even after weeks of sustained expression, allowed rapid recovery of intestinal cell proliferation and reversal of aging features in most mice. These results suggest that p16-mediated inhibition of Cdk activity is sufficient to inhibit cell proliferation and impose aging features in somatic tissues of mammals and that at least some of these aging features are reversible.


Assuntos
Senilidade Prematura/patologia , Ciclo Celular , Inibidor p16 de Quinase Dependente de Ciclina/metabolismo , Animais , Proliferação de Células , Quinase 4 Dependente de Ciclina/metabolismo , Células Epiteliais/citologia , Humanos , Intestinos/citologia , Camundongos Transgênicos , Reprodutibilidade dos Testes , Células-Tronco/citologia
5.
Cancer Biol Ther ; 13(8): 588-91, 2012 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-22549156

RESUMO

Barrett's esophagus is a metaplasia of the distal esophagus that is the only recognized precursor of esophageal adenocarcinoma. Despite a characteristic histology, the pathogenesis of Barrett's has remained obscure. A recent paper from the laboratories of Wa Xian and Frank McKeon presents evidence for a novel cell of origin of Barrett's. Their work is based on studies of mice engineered to lack the squamous epithelial stem cell survival factor p63. These mice develop a metaplasia of the proximal stomach and esophagus that harbors substantial histological and molecular features of Barrett's. The metaplasia appears to form from embryonic progenitor cells that normally persists post-natally only at the squamo-columnar junction. Moreover, in their model, the metaplasia is initiated not by mutation but by reduced competition between these cells and squamous epithelial cells.


Assuntos
Esôfago de Barrett , Adenocarcinoma , Animais , Esôfago de Barrett/patologia , Neoplasias Esofágicas , Esôfago/patologia , Metaplasia , Camundongos
6.
Genes Cancer ; 3(11-12): 614-8, 2012 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-23634250

RESUMO

Cyclin-dependent kinases (Cdks) drive cell cycle progression in all eukaryotes. Yeasts have a single major Cdk that mediates distinct cell cycle transitions via association with different cyclins. The closest homolog in mammals, Cdk1, drives mitosis. Mammals have additional Cdks-Cdk2, Cdk4, and Cdk6-that represent the major Cdks activated during interphase (iCdks). A large body of evidence has accrued that suggests that activation of iCdks dictates progression though interphase. In apparent contradiction, deficiency in each individual iCdk, respectively, in knockout mice proved to be compatible with live birth and in some instances fertility. Moreover, murine embryos could be derived with Cdk1 as the only functional Cdk. Thus, none of the iCdks is strictly essential for mammalian cell cycle progression, raising the possibility that Cdk1 is the dominant regulator in interphase. However, an absence of iCdks has been accompanied by major shifts in cyclin association to Cdk1, suggesting gain in function. After considerable tweaking, a chemical genetic approach has recently been able to examine the impact of acute inhibition of Cdk2 activity without marked distortion of cyclin/Cdk complex formation. The results suggest that, when expressed at its normal levels, Cdk2 performs essential roles in driving human cells into S phase and maintaining genomic stability. These new findings appear to have restored order to the cell cycle field, bringing it full circle to the view that iCdks indeed play important roles. They also underscore the caveat in knockdown and knockout approaches that protein underexpression can significantly perturb a protein interaction network. We discuss the implications of the new synthesis for future cell cycle studies and anti-Cdk-based therapy of cancer and other diseases.

7.
Mol Cell ; 42(1): 36-49, 2011 Apr 08.
Artigo em Inglês | MEDLINE | ID: mdl-21474066

RESUMO

Mutations in both RAS and the PTEN/PIK3CA/AKT signaling module are found in the same human tumors. PIK3CA and AKT are downstream effectors of RAS, and the selective advantage conferred by mutation of two genes in the same pathway is unclear. Based on a comparative molecular analysis, we show that activated PIK3CA/AKT is a weaker inducer of senescence than is activated RAS. Moreover, concurrent activation of RAS and PIK3CA/AKT impairs RAS-induced senescence. In vivo, bypass of RAS-induced senescence by activated PIK3CA/AKT correlates with accelerated tumorigenesis. Thus, not all oncogenes are equally potent inducers of senescence, and, paradoxically, a weak inducer of senescence (PIK3CA/AKT) can be dominant over a strong inducer of senescence (RAS). For tumor growth, one selective advantage of concurrent mutation of RAS and PTEN/PIK3CA/AKT is suppression of RAS-induced senescence. Evidence is presented that this new understanding can be exploited in rational development and targeted application of prosenescence cancer therapies.


Assuntos
Genes ras , Neoplasias/enzimologia , Neoplasias/genética , Fosfatidilinositol 3-Quinases/genética , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/genética , Proteínas Proto-Oncogênicas c-akt/metabolismo , Idoso , Animais , Linhagem Celular Transformada , Proliferação de Células , Senescência Celular/genética , Senescência Celular/fisiologia , Classe I de Fosfatidilinositol 3-Quinases , Modelos Animais de Doenças , Ativação Enzimática , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Masculino , Camundongos , Camundongos Knockout , Camundongos Transgênicos , Pessoa de Meia-Idade , Mutação , Neoplasias/patologia , PTEN Fosfo-Hidrolase/antagonistas & inibidores , PTEN Fosfo-Hidrolase/genética , PTEN Fosfo-Hidrolase/metabolismo , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/metabolismo , Neoplasias Pancreáticas/patologia , Transdução de Sinais
8.
Cell Div ; 5: 16, 2010 Jun 22.
Artigo em Inglês | MEDLINE | ID: mdl-20569479

RESUMO

BACKGROUND: Cellular senescence is a permanent growth arrest that occurs in response to cellular stressors, such as telomere shortening or activation of oncogenes. Although the process of senescence growth arrest is somewhat conserved between mouse and human cells, there are some critical differences in the molecular pathways of senescence between these two species. Recent studies in human fibroblasts have defined a cell signaling pathway that is initiated by repression of a specific Wnt ligand, Wnt2. This, in turn, activates a histone chaperone HIRA, and culminates in formation of specialized punctate domains of facultative heterochromatin, called Senescence-Associated Heterochromatin Foci (SAHF), that are enriched in the histone variant, macroH2A. SAHF are thought to repress expression of proliferation-promoting genes, thereby contributing to senescence-associated proliferation arrest. We asked whether this Wnt2-HIRA-SAHF pathway is conserved in mouse fibroblasts. RESULTS: We show that mouse embryo fibroblasts (MEFs) and mouse skin fibroblasts, do not form robust punctate SAHF in response to an activated Ras oncogene or shortened telomeres. However, senescent MEFs do exhibit elevated levels of macroH2A staining throughout the nucleus as a whole. Consistent with their failure to fully activate the SAHF assembly pathway, the Wnt2-HIRA signaling axis is not overtly regulated between proliferating and senescent mouse cells. CONCLUSIONS: In addition to the previously defined differences between mouse and human cells in the mechanisms and phenotypes associated with senescence, we conclude that senescent mouse and human fibroblasts also differ at the level of chromatin and the signaling pathways used to regulate chromatin. These differences between human and mouse senescence may contribute to the increased propensity of mouse fibroblasts (and perhaps other mouse cell types) to become immortalized and transformed, compared to human cells.

9.
Cell Div ; 5: 12, 2010 May 13.
Artigo em Inglês | MEDLINE | ID: mdl-20465818

RESUMO

The kinase Wee1 has been recognized for a quarter century as a key inhibitor of Cyclin dependent kinase 1 (Cdk1) and mitotic entry in eukaryotes. Nonetheless, Wee1 regulation is not well understood and its large amino-terminal regulatory domain (NRD) has remained largely uncharted. Evidence has accumulated that cyclin B/Cdk1 complexes reciprocally inhibit Wee1 activity through NRD phosphorylation. Recent studies have identified the first functional NRD elements and suggested that vertebrate cyclin A/Cdk2 complexes also phosphorylate the NRD. A short NRD peptide, termed the Wee box, augments the activity of the Wee1 kinase domain. Cdk1/2-mediated phosphorylation of the Wee box (on T239) antagonizes kinase activity. A nearby region harbors a conserved RxL motif (RxL1) that promotes cyclin A/Cdk2 binding and T239 phosphorylation. Mutation of either T239 or RxL1 bolsters the ability of Wee1 to block mitotic entry, consistent with negative regulation of Wee1 through these sites. The region in human somatic Wee1 that encompasses RxL1 also binds Crm1, directing Wee1 export from the nucleus. These studies have illuminated important aspects of Wee1 regulation and defined a specific molecular pathway through which cyclin A/Cdk2 complexes foster mitotic entry. The complexity, speed, and importance of regulation of mitotic entry suggest that there is more to be learned.

10.
Transl Oncol ; 3(1): 33-42, 2010 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-20165693

RESUMO

Barrett esophagus is an epithelial metaplasia that predisposes to adenocarcinoma. Better markers of cancer risk are urgently needed to identify those patients who are likely to benefit most from emerging methods of endoscopic ablation. Disease progression is associated with genomic DNA changes (segmental gains, losses, or loss of heterozygosity). Although these changes are not easily assayed directly, we hypothesized that the underlying DNA damage should activate a DNA damage response (DDR), detectable by immunohistochemical (IHC) assays of checkpoint proteins and the resulting replicative phase cell cycle delays. Surgical specimens and endoscopic biopsies (N = 28) were subjected to IHC for the cell cycle markers cyclin A and phosphorylated histone H3 (P-H3), the DDR markers gammaH2AX and phosphorylated ATM/ATR substrates (P-ATM/ATRsub), and the DNA damage-responsive tumor suppressors p16 and p53. Correlations were made with histologic diagnoses. The fractions of cells that stained for cyclin A, P-H3, and gammaH2AX increased in parallel in dysplastic tissue, consistent with checkpoint-mediated cell cycle delays. Foci of nuclear gammaH2AX and P-ATM/ATRsub were demonstrated by standard and confocal immunofluorescence. Staining for p16 was more prevalent in early-stage disease with lower staining for gammaH2AX and P-H3. Staining for p53 was moderately increased in some early-stage disease and strongly increased in some advanced disease, consistent with checkpoint-mediated induction and mutational inactivation of p53, respectively. We suggest that IHC for DDR-associated markers may help stratify risk of disease progression in Barrett.

11.
Mol Cell Biol ; 30(1): 116-30, 2010 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-19858290

RESUMO

Sophisticated models for the regulation of mitotic entry are lacking for human cells. Inactivating human cyclin A/Cdk2 complexes through diverse approaches delays mitotic entry and promotes inhibitory phosphorylation of Cdk1 on tyrosine 15, a modification performed by Wee1. We show here that cyclin A/Cdk2 complexes physically associate with Wee1 in U2OS cells. Mutation of four conserved RXL cyclin A/Cdk binding motifs (RXL1 to RXL4) in Wee1 diminished stable binding. RXL1 resides within a large regulatory region of Wee1 that is predicted to be intrinsically disordered (residues 1 to 292). Near RXL1 is T239, a site of inhibitory Cdk phosphorylation in Xenopus Wee1 proteins. We found that T239 is phosphorylated in human Wee1 and that this phosphorylation was reduced in an RXL1 mutant. RXL1 and T239 mutants each mediated greater Cdk phosphorylation and G(2)/M inhibition than the wild type, suggesting that cyclin A/Cdk complexes inhibit human Wee1 through these sites. The RXL1 mutant uniquely also displayed increased nuclear localization. RXL1 is embedded within sequences homologous to Crm1-dependent nuclear export signals (NESs). Coimmunoprecipitation showed that Crm1 associated with Wee1. Moreover, treatment with the Crm1 inhibitor leptomycin B or independent mutation of the potential NES (NESm) abolished Wee1 nuclear export. Export was also reduced by Cdk inhibition or cyclin A RNA interference, suggesting that cyclin A/Cdk complexes contribute to Wee1 export. Somewhat surprisingly, NESm did not display increased G(2)/M inhibition. Thus, nuclear export of Wee1 is not essential for mitotic entry though an important functional role remains likely. These studies identify a novel bifunctional regulatory element in Wee1 that mediates cyclin A/Cdk2 association and nuclear export.


Assuntos
Proteínas de Ciclo Celular/fisiologia , Núcleo Celular/metabolismo , Ciclina A/metabolismo , Quinase 2 Dependente de Ciclina/metabolismo , Carioferinas/fisiologia , Proteínas Nucleares/fisiologia , Proteínas Tirosina Quinases/fisiologia , Receptores Citoplasmáticos e Nucleares/fisiologia , Transporte Ativo do Núcleo Celular , Motivos de Aminoácidos , Sequência de Aminoácidos , Linhagem Celular , Sequência Conservada , Ciclina A/genética , Ciclina E/metabolismo , Fase G2 , Humanos , Carioferinas/antagonistas & inibidores , Mitose , Dados de Sequência Molecular , Fosforilação , Ligação Proteica , Estrutura Terciária de Proteína , Interferência de RNA , Receptores Citoplasmáticos e Nucleares/antagonistas & inibidores , Fase S , Proteína Exportina 1
12.
Cancer Prev Res (Phila) ; 2(9): 800-6, 2009 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-19723896

RESUMO

Despite advances in screening and treatment, colorectal cancer remains the second leading cause of cancer-related death in the United States. Cyclin-dependent kinases (Cdk) are deregulated in colorectal cancer by silencing of the Cdk inhibitor p16(Ink4a) and other mechanisms. We tested whether the small molecule Cdk inhibitor SNS-032 (formerly BMS-387032), which targets Cdk2, Cdk7, and Cdk9, can prevent intestinal tumorigenesis in mouse models. We generated mice with high intestinal tumor loads by combining the multiple intestinal neoplasia (Min) mutation with Ink4a/Arf mutations and inducing colitis with dextran sulfate sodium. p16-null Min mice (n = 17) began dextran sulfate sodium treatment at week 5 and i.p. injection of carrier or SNS-032 at week 6. Mice were sacrificed at week 12. SNS-032 was well tolerated and reduced colon tumor burden to 36% of that in carrier-treated mice (P < 0.001). We then extended the study to Ink4/Arf-null Min mice (n = 14) and increased the drug dose frequency. SNS-032 treatment reduced the intestinal tumor number to 25% and intestinal tumor burden to 16% of carrier-treated mice (P < 0.0001). DNA synthesis in non-neoplastic and tumor epithelial cells, detected by bromodeoxyuridine incorporation, was modestly reduced by acute SNS-032 treatment. The mitotic index, detected by histone H3 phosphorylation, was distinctly decreased (P < 0.03), and apoptosis, detected by caspase 3 activation, was increased (P < 0.005). These results show the chemoprevention of intestinal tumorigenesis by SNS-032. Our findings support further study of Cdk inhibitors for chemoprevention and therapy of colon cancer.


Assuntos
Quinases Ciclina-Dependentes/antagonistas & inibidores , Neoplasias Intestinais/prevenção & controle , Oxazóis/uso terapêutico , Inibidores de Proteínas Quinases/uso terapêutico , Tiazóis/uso terapêutico , Fator 1 de Ribosilação do ADP/fisiologia , Animais , Apoptose/efeitos dos fármacos , Ciclo Celular/efeitos dos fármacos , Colite/enzimologia , Colite/patologia , Colite/prevenção & controle , Inibidor p16 de Quinase Dependente de Ciclina/fisiologia , Feminino , Neoplasias Intestinais/enzimologia , Neoplasias Intestinais/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Fosforilação/efeitos dos fármacos
13.
J Clin Invest ; 119(4): 758-60, 2009 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-19348043

RESUMO

The Wnt pathway has been found to play a role in the development of many tissues and to spur growth and differentiation of adult osteoblasts, sparking interest in its potential clinical application for bone growth. However, when deregulated, this pathway can be oncogenic in some tissues. In this issue of the JCI, Kansara and colleagues reveal that Wnt inhibitory factor 1 is epigenetically silenced in human osteosarcomas and that its absence augments osteosarcoma formation in mice (see the related article beginning on page 837). These observations suggest the need for caution in stimulating the Wnt pathway for therapeutic bone growth.


Assuntos
Osteoporose/terapia , Proteínas Wnt/fisiologia , Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Adaptadoras de Transdução de Sinal/fisiologia , Animais , Proteínas da Matriz Extracelular/deficiência , Proteínas da Matriz Extracelular/genética , Inativação Gênica , Genes Supressores de Tumor , Humanos , Peptídeos e Proteínas de Sinalização Intercelular/deficiência , Peptídeos e Proteínas de Sinalização Intercelular/genética , Camundongos , Modelos Biológicos , Osteossarcoma/etiologia , Osteossarcoma/genética , Proteínas Repressoras/genética , Proteínas Repressoras/fisiologia , Transdução de Sinais , Proteínas Wnt/genética
14.
Clin Cancer Res ; 15(9): 3156-62, 2009 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-19383812

RESUMO

PURPOSE: Immunohistochemistry for mismatch repair proteins has shown utility in the identification of Lynch syndrome, but majority of tumors with loss of MLH1 expression are due to sporadic hypermethylation of the MLH1 promoter. These tumors can also show epigenetic silencing of other genes, such as p16. The aim of our study is to evaluate the utility of p16 immunohistochemistry in the prediction of MLH1 germline mutations. EXPERIMENTAL DESIGN: p16 immunohistochemistry was appropriately evaluated in 79 colorectal cancers with loss of MLH1 expression. Methylation of MLH1 and p16 were quantitatively studied using real-time PCR assay Methylight. BRAF V600E mutation in tumor tissue was also investigated. Genetic testing for germline mutation of MLH1 was made on 52 patients. RESULTS: Loss of p16 expression was seen in 21 of 79 samples (26.6%). There was found statistically significant association between p16 expression and p16 methylation (P < 0.001), MLH1 methylation (P < 0.001), and BRAF mutation (P < 0.005). All tumors with loss of p16 expression showed hypermethylation of p16 (21 of 21), 95.2% (20 of 21) showed MLH1 methylation, and 71.4% (15 of 21) were mutated for BRAF V600E. Mutational analysis showed pathogenic germline mutations in 8 of the patients, harboring 10 tumors. All 10 of these tumors showed normal staining of p16 in the immunochemical analysis. CONCLUSIONS: p16 immunohistochemistry is a good surrogate marker for p16 and MLH1 epigenetic silencing due to hypermethylation, and is useful as screening tool in the selection of patients for genetic testing in Lynch syndrome.


Assuntos
Neoplasias Colorretais Hereditárias sem Polipose/metabolismo , Proteínas de Neoplasias/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/genética , Neoplasias Colorretais Hereditárias sem Polipose/genética , Inibidor p16 de Quinase Dependente de Ciclina , Metilação de DNA , Epigênese Genética , Feminino , Mutação em Linhagem Germinativa/genética , Humanos , Técnicas Imunoenzimáticas , Masculino , Pessoa de Meia-Idade , Proteína 1 Homóloga a MutL , Proteínas de Neoplasias/genética , Proteínas Nucleares/genética , Prognóstico , Proteínas Proto-Oncogênicas B-raf/genética
15.
Gastroenterology ; 136(2): 596-606.e4, 2009 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-19056388

RESUMO

BACKGROUND & AIMS: Vascular endothelial growth factor (VEGF) is expressed robustly in human colon neoplasia and is a major new "rational" target of therapy for cancers of the colon and other organs. Nonetheless, the mechanism(s) of action of VEGF-targeted therapies and the biologic roles of VEGF in tumorigenesis have not been well defined. We used a transgenic approach to directly test the hypothesis that augmented VEGF expression can drive progression of intestinal neoplasia. METHODS: Transgenic mouse lines were generated with moderate (vilVEGF1) and high (vilVEGF2) VEGF expression from the intestinal epithelium. vilVEGF1 mice were bred to Min mice (adenomatous polyposis coli [APC] +/-). Colon epithelial cells from an APC patient were cocultured with endothelial cells and fibroblasts. RESULTS: vilVEGF mice were generally healthy but displayed red small intestines. Vessels were larger and more numerous in the submucosa but not the mucosa. The mucosa showed striking stromal and epithelial hypercellularity, with increased epithelial proliferation. Many crypts formed cysts composed of relatively undifferentiated epithelial cells surrounded by cells with endothelial and myofibroblast markers. Compared with Min controls, vilVEGF1-Min mice developed 6-fold more intestinal adenomas of all sizes, with more advanced histologic features. Polycystic masses were also observed. Coculture of human colonocytes with endothelial cells and fibroblasts directly stimulated colonocyte proliferation. CONCLUSIONS: Augmented VEGF expression from intestinal epithelium potently stimulated cross talk with mesenchymal cells and proliferation of normal and neoplastic epithelium. These effects of VEGF, largely occurring prior to the canonical angiogenic switch in tumors, may be in part independent of angiogenesis.


Assuntos
Comunicação Celular/fisiologia , Mucosa Intestinal/metabolismo , Neoplasias Intestinais/metabolismo , Mesoderma/metabolismo , Neoplasias Epiteliais e Glandulares/metabolismo , Fator A de Crescimento do Endotélio Vascular/metabolismo , Polipose Adenomatosa do Colo/metabolismo , Polipose Adenomatosa do Colo/patologia , Animais , Proliferação de Células , Células Cultivadas , Técnicas de Cocultura , Colo/citologia , Colo/metabolismo , Colo/patologia , Modelos Animais de Doenças , Endotélio Vascular/citologia , Endotélio Vascular/metabolismo , Endotélio Vascular/patologia , Humanos , Mucosa Intestinal/citologia , Mucosa Intestinal/patologia , Neoplasias Intestinais/patologia , Mesoderma/citologia , Camundongos , Camundongos Transgênicos , Neoplasias Epiteliais e Glandulares/patologia
16.
Cancer Biol Ther ; 7(11): 1706-11, 2008 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-18836285

RESUMO

Studies of early neoplasia have revealed fundamental molecular pathways that drive tumorigenesis. Despite this progress, synthesis of principles of tumorigenesis that span tissue types has lagged. Such forays into the 'comparative anatomy' of cancer can stimulate new models and refine key questions. We envision commonality of pathways important in formation of two early benign neoplasms that are found in different tissues and which are not generally thought to be similar: dysplastic nevi of the skin and intestinal aberrant crypt foci. We propose that these neoplasms result from an ongoing 'tug of war' between the tumor suppression barrier posed by cellular senescence and the tumor-promoting activity of Wnt-signaling. Whether or not such neoplasms progress to malignancy or persist in a benign state for many years might be largely determined by the outcome of this tug of war and its modulation by other genetic and epigenetic alterations, such as inactivation of p16(INK4a).


Assuntos
Regulação Neoplásica da Expressão Gênica , Neoplasias/metabolismo , Proteínas Wnt/metabolismo , Animais , Senescência Celular , Células Epiteliais/metabolismo , Humanos , Modelos Biológicos , Modelos Genéticos , Mutação , Nevo/metabolismo , Oncogenes , Fosforilação , Transdução de Sinais
17.
J Biol Chem ; 283(26): 17749-52, 2008 Jun 27.
Artigo em Inglês | MEDLINE | ID: mdl-18424430

RESUMO

Chk1 is a conserved kinase that imposes cell cycle delays in response to impediments to DNA replication. Recent experiments have further defined effects of Chk1 on the activity of mammalian origins of DNA replication and progression of replication forks. Moreover, Chk1 now appears to help defend genomic integrity through effects on several other pathways, including Fanconi anemia proteins, the mitotic spindle, and transcription of cell cycle-related genes. These findings can account for the requirement for Chk1 in normal proliferating cells of the early embryo and suggest the potential for diverse effects of Chk1 inhibition in cancer therapy.


Assuntos
Replicação do DNA , Regulação da Expressão Gênica , Genoma , Proteínas Quinases/fisiologia , Animais , Ciclo Celular , Linhagem Celular Tumoral , Proliferação de Células , Centrossomo/metabolismo , Quinase 1 do Ponto de Checagem , Galinhas , DNA/metabolismo , Humanos , Modelos Biológicos , Neoplasias/genética , Neoplasias/terapia , Proteínas Quinases/metabolismo , Fuso Acromático
18.
Nat Genet ; 38(9): 1060-5, 2006 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-16878133

RESUMO

Human adenocarcinomas commonly harbor mutations in the KRAS and MYC proto-oncogenes and the TP53 tumor suppressor gene. All three genetic lesions are potentially pro-angiogenic, as they sustain production of vascular endothelial growth factor (VEGF). Yet Kras-transformed mouse colonocytes lacking p53 formed indolent, poorly vascularized tumors, whereas additional transduction with a Myc-encoding retrovirus promoted vigorous vascularization and growth. In addition, VEGF levels were unaffected by Myc, but enhanced neovascularization correlated with downregulation of anti-angiogenic thrombospondin-1 (Tsp1) and related proteins, such as connective tissue growth factor (CTGF). Both Tsp1 and CTGF are predicted targets for repression by the miR-17-92 microRNA cluster, which was upregulated in colonocytes coexpressing K-Ras and c-Myc. Indeed, miR-17-92 knockdown with antisense 2'-O-methyl oligoribonucleotides partly restored Tsp1 and CTGF expression; in addition, transduction of Ras-only cells with a miR-17-92-encoding retrovirus reduced Tsp1 and CTGF levels. Notably, miR-17-92-transduced cells formed larger, better-perfused tumors. These findings establish a role for microRNAs in non-cell-autonomous Myc-induced tumor phenotypes.


Assuntos
MicroRNAs/metabolismo , Neoplasias/irrigação sanguínea , Neovascularização Patológica/metabolismo , Proteínas Proto-Oncogênicas c-myc/fisiologia , Animais , Linhagem Celular , Linhagem Celular Transformada , Transformação Celular Viral , Células Cultivadas , Fator de Crescimento do Tecido Conjuntivo , Meios de Cultivo Condicionados/análise , Regulação Neoplásica da Expressão Gênica , Vetores Genéticos , Humanos , Proteínas Imediatamente Precoces/genética , Proteínas Imediatamente Precoces/metabolismo , Técnicas In Vitro , Peptídeos e Proteínas de Sinalização Intercelular/genética , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Neoplasias/patologia , Neovascularização Patológica/genética , Oligonucleotídeos Antissenso/farmacologia , Proteínas Proto-Oncogênicas c-myc/genética , RNA Neoplásico/metabolismo , Retroviridae/genética , Células-Tronco/citologia , Trombospondina 1/genética , Trombospondina 1/metabolismo , Transplante Homólogo , Fator A de Crescimento do Endotélio Vascular/análise , Fator A de Crescimento do Endotélio Vascular/metabolismo
19.
Neoplasia ; 8(6): 429-36, 2006 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-16820088

RESUMO

p16(INK4a) is a major tumor-suppressor protein, but its regulation and settings of fuction remain poorly understood. To explore the notion that p16 is induced in vivo in response to replicative stress, we examined p16 expression in tissues from human ulcerative colitis (UC; n = 25) and normal controls (n = 20). p16 was expressed strongly in UC-associated neoplasms (n = 17), as seen previously in sporadic colonic neoplasms. In non-neoplastic UC epithelium, p16 was expressed in 33% of crypts (the proliferative compartment) compared to < 1% of normal controls. p16 expression did not correlate with degree of inflammation but did correlate with the degree of crypt architecture distortion (P = .002)-a reflection of epithelial regeneration. In coimmunofluorescence studies with Ki67, p16 expression was associated with cell cycle arrest (P < .001). Both UC and normal crypts displayed evidence for the activation of the DNA damage checkpoint pathway, and p16 was induced in primary cultures of normal epithelial cells by ionizing irradiation (IR). However, induction by IR displayed delayed kinetics, implying that p16 is not an immediate target of the checkpoint pathway. These findings support a model in which p16 is induced as an "emergency brake" in cells experiencing sustained replicative stress.


Assuntos
Colite Ulcerativa/metabolismo , Inibidor p16 de Quinase Dependente de Ciclina/biossíntese , Regulação Neoplásica da Expressão Gênica , Regulação da Expressão Gênica , Dano ao DNA , Células Epiteliais/citologia , Epitélio/patologia , Humanos , Inflamação , Mucosa Intestinal/patologia , Antígeno Ki-67/biossíntese , Cinética , Microscopia de Fluorescência , Regeneração
20.
Cell Cycle ; 5(6): 661-6, 2006 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-16582612

RESUMO

In mammalian somatic cells, the Anaphase Promoting Complex (APC) is inactivated during S phase by active cyclin A-Cyclin dependent kinase (Cdk) 2 complexes promoting accumulation of mitotic regulators, such as cyclin B and Polo like kinase 1 (Plk1). However, mitotic entry does not appear to be perturbed in some human cancer cells or in normal mouse cells following Cdk2 RNA interference (i) or deletion of the Cdk2 gene. These results suggest functional complementation of APC regulation by a compensatory kinase. Using Plk1 protein level as readout of APC activity, we show that APC is inactivated during S phase in human cells by both cyclin A-Cdk2 and cyclin A-Cdk1 complexes. Expression of a dominant negative mutant of Cdk2 or Cdk2 RNAi in early S phase destabilizes Plk1 as it begins to accumulate. However, this effect wanes in late S phase, where destabilization of Plk1 also requires Cdk1 RNAi. Although Cdk2 is the dominant partner of cyclin A in these settings, cyclin A also binds Cdk1. Both complexes bind the APC targeting factor Cdh1, but Cdk1 complexes are inactive in early S phase, accounting for the stronger regulation of APC function by Cdk2. These results provide further evidence that cyclin A-Cdk2 and -Cdk1 complexes display overlapping and partially redundant roles in preparing cells for mitosis, through regulation of the APC.


Assuntos
Proteína Quinase CDC2/metabolismo , Ciclina A/metabolismo , Quinase 2 Dependente de Ciclina/metabolismo , Mitose/genética , Complexos Ubiquitina-Proteína Ligase/metabolismo , Ciclossomo-Complexo Promotor de Anáfase , Proteína Quinase CDC2/genética , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Linhagem Celular , Linhagem Celular Tumoral , Ciclina A/genética , Quinase 2 Dependente de Ciclina/genética , Regulação para Baixo/genética , Humanos , Substâncias Macromoleculares/metabolismo , Mutação/genética , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismo , Proteínas Proto-Oncogênicas/genética , Proteínas Proto-Oncogênicas/metabolismo , Interferência de RNA/fisiologia , Fase S/fisiologia , Complexos Ubiquitina-Proteína Ligase/genética , Quinase 1 Polo-Like
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