RESUMO
BACKGROUND: Recent publications have shown patients with defects in the DNA mismatch repair (MMR) pathway driven by either MSH2 or MSH6 loss experience a significant increase in the incidence of prostate cancer. Moreover, this increased incidence of prostate cancer is accompanied by rapid disease progression and poor clinical outcomes. METHODS AND RESULTS: We show that androgen-receptor activation, a key driver of prostate carcinogenesis, can disrupt the MSH2 gene in prostate cancer. We screened tumours from two cohorts (recurrent/non-recurrent) of prostate cancer patients to confirm the loss of MSH2 protein expression and identified decreased MSH2 expression in recurrent cases. Stratifying the independent TCGA prostate cancer cohort for MSH2/6 expression revealed that patients with lower levels of MSH2/6 had significant worse outcomes, in contrast, endometrial and colorectal cancer patients with lower MSH2/6 levels. MMRd endometrial and colorectal tumours showed the expected increase in mutational burden, microsatellite instability and enhanced immune cell mobilisation but this was not evident in prostate tumours. CONCLUSIONS: We have shown that loss or reduced levels of MSH2/MSH6 protein in prostate cancer is associated with poor outcome. However, our data indicate that this is not associated with a statistically significant increase in mutational burden, microsatellite instability or immune cell mobilisation in a cohort of primary prostate cancers.
Assuntos
Neoplasias Colorretais/genética , Neoplasias do Endométrio/genética , Proteína 2 Homóloga a MutS/genética , Neoplasias da Próstata/genética , Neoplasias Colorretais/imunologia , Reparo de Erro de Pareamento de DNA , Neoplasias do Endométrio/imunologia , Feminino , Rearranjo Gênico , Mutação em Linhagem Germinativa , Humanos , Masculino , Instabilidade de Microssatélites , Neoplasias da Próstata/imunologia , Transcriptoma , Células Tumorais Cultivadas , Sequenciamento Completo do GenomaRESUMO
DNA-fluorescence in situ hybridisation (DNA-FISH) allows visualisation of chromosome organisation and rearrangement. FISH probes are pools of short fluorescently labelled DNA fragments that are often produced from template plasmids that contain large genomic inserts. For effective sample penetration and target hybridisation it is critical that probe fragments are between 200 and 500bp. Production of these short probes requires significant optimisation and can be confounded access to expensive sonication equipment or inherent sequence features that influence enzymatic fragmentation or amplification. Here we demonstrate that effective FISH probes can be prepared without the need for optimisation of fragmentation using a cocktail of two the 4bp recognition sequence restriction enzymes CviQI and AluI.
RESUMO
Evidence suggests that altered adipose tissue homeostasis may be an important contributor to the development and/or progression of prostate cancer. In this study, we investigated the adipose transcriptional profiles of low- and high-risk disease to determine both prognostic potential and possible biological drivers of aggressive disease. RNA was extracted from periprostatic adipose tissue from patients categorised as having prostate cancer with either a low or high risk of progression based on tumour characteristics at prostatectomy and profiled by RNA sequencing. The expression of selected genes was then quantified by qRT-PCR in a cross-validation cohort. In the first phase, a total of 677 differentially transcribed genes were identified, from which a subset of 14 genes was shortlisted. In the second phase, a 3 gene (IGHA1, OLFM4, RERGL) signature was refined and evaluated using recursive feature selection and cross-validation, obtaining a promising discriminatory utility (area under curve 0.72) at predicting the presence of high-risk disease. Genes implicated in immune and/or inflammatory responses predominated. Periprostatic adipose tissue from patients with high-risk prostate cancer has a distinct transcriptional signature that may be useful for detecting its occult presence. Differential expression appears to be driven by a local immune/inflammatory reaction to more advanced tumours, than any specific adipose tissue-specific tumour-promoting mechanism. This signature is transferable into a clinically usable PCR-based assay, which in a cross-validation cohort shows diagnostic potential.
Assuntos
Tecido Adiposo/fisiopatologia , Neoplasias da Próstata/diagnóstico , Transcriptoma/genética , Adulto , Idoso , Humanos , Masculino , Pessoa de Meia-Idade , Prognóstico , Neoplasias da Próstata/metabolismo , Fatores de RiscoRESUMO
The maintenance of a pristine genome, free from errors, is necessary to prevent cellular transformation and degeneration. When errors in DNA are detected, DNA damage repair (DDR) genes and their regulators are activated to effect repair. When these DDR pathways are themselves mutated or aberrantly downregulated, cancer and neurodegenerative disorders can ensue. Multiple lines of evidence now indicate, however, that defects in key regulators of DNA repair pathways are highly enriched in human metastasis specimens and hence may be a key step in the acquisition of metastasis and the ability of localized disease to disseminate. Some of the key regulators of checkpoints in the DNA damage response are the TP53 protein and the PARP enzyme family. Targeting of these pathways, especially through PARP inhibition, is now being exploited therapeutically to effect significant clinical responses in subsets of individuals, particularly in patients with ovarian cancer or prostate cancer, including cancers with a marked metastatic burden. Targeting DNA repair-deficient tumors with drugs that take advantage of the fundamental differences between normal repair-proficient cells and repair-deficient tumors offers new avenues for treating advanced disease in the future. Clin Cancer Res; 22(13); 3132-7. ©2016 AACR.
Assuntos
Reparo do DNA/efeitos dos fármacos , Neoplasias Ovarianas/genética , Inibidores de Poli(ADP-Ribose) Polimerases/uso terapêutico , Neoplasias da Próstata/genética , Proteína Supressora de Tumor p53/antagonistas & inibidores , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Dano ao DNA/genética , Reparo do DNA/fisiologia , Feminino , Humanos , Masculino , Neoplasias Ovarianas/tratamento farmacológico , Neoplasias Ovarianas/patologia , Poli(ADP-Ribose) Polimerases/genética , Neoplasias da Próstata/tratamento farmacológico , Neoplasias da Próstata/patologia , Transdução de Sinais/efeitos dos fármacos , Proteína Supressora de Tumor p53/genéticaRESUMO
Assessments of animal-based outcomes form the core of routine veterinary clinical examinations and are being increasingly used as indicators of animal welfare. A method of group observation that did not require gathering and handling of individual sheep, was used to assess eight animal-based indicators of sheep welfare: demeanour, skin irritation, wool loss, excessive panting, coughing, lameness, and cleanliness of the ventral abdominal and 'breech' (perineum/gluteal/caudal hindlimb) areas. The inter-observer reliability of two or three observers who independently assessed these indicators was tested on 2406 adult sheep and growing lambs across 36 farms and the intra-observer reliability of an experienced, veterinary assessor--the 'test standard observer'--was assessed on 88 adult sheep during four on-farm assessments. Observer reliability was evaluated using Cronbach's alpha (α) comparison of the recorded proportion (%) of sheep affected by each welfare condition and by binomial logistic regression modelling. High levels of inter-observer reliability were identified for the assessment of group lameness (α 0.76-1.00) and cleanliness of the breech area (α 0.97-1.00). Excellent intra-observer reliability was determined for lameness (α 0.99), cleanliness of the breech area (α 0.97), demeanour (α 1.00) and wool loss (α 1.00). In addition, proportion data and logistic regression models identified few between-observer differences. The results suggest that welfare outcomes based on observations of the behaviour and physical appearance of individual animals within a group may offer a reliable and feasible measurement tool for the on-farm assessment of sheep welfare.
Assuntos
Criação de Animais Domésticos/normas , Bem-Estar do Animal/estatística & dados numéricos , Bem-Estar do Animal/normas , Doenças dos Ovinos/epidemiologia , Ovinos , Animais , Comportamento Animal , Modelos Logísticos , Variações Dependentes do Observador , Reprodutibilidade dos Testes , Reino Unido/epidemiologia , LãRESUMO
Despite 12 yr since the discovery of SRY, little is known at the molecular level about how SRY and the SRY-related protein, SOX9 [SRY-related high-mobility group (HMG) box 9], initiate the program of gene expression required to commit the bipotential embryonic gonad to develop into a testis rather than an ovary. Analysis of SRY and SOX9 clinical mutant proteins and XX mice transgenic for testis-determining genes have provided some insight into their normal functions. SRY and SOX9 contain an HMG domain, a DNA-binding motif. The HMG domain plays a central role, being highly conserved between species and the site of nearly all missense mutations causing XY gonadal dysgenesis. SRY and SOX9 are architectural transcription factors; their HMG domain is capable of directing nuclear import and DNA bending. Whether SRY and SOX9 activate testis-forming genes, repress ovary-forming genes, or both remains speculative until downstream DNA target genes are identified. However, factors that control SRY and SOX9 gene expression have been identified, as have a dozen sex-determining genes, allowing some of the pieces in this molecular genetic puzzle to be connected. Many genes, however, remain unidentified, because in the majority of cases of XY females and in all cases of XX males lacking SRY, the mutated gene is unknown.
Assuntos
Proteínas de Ligação a DNA/fisiologia , Proteínas de Grupo de Alta Mobilidade/fisiologia , Proteínas Nucleares , Diferenciação Sexual , Fatores de Transcrição/fisiologia , Sequência de Aminoácidos , Animais , Proteínas de Ligação a DNA/genética , Transtornos do Desenvolvimento Sexual , Feminino , Regulação da Expressão Gênica , Domínios HMG-Box/genética , Domínios HMG-Box/fisiologia , Proteínas de Grupo de Alta Mobilidade/genética , Masculino , Camundongos , Dados de Sequência Molecular , Mutação de Sentido Incorreto , Organogênese/genética , Organogênese/fisiologia , Fatores de Transcrição SOX9 , Alinhamento de Sequência , Proteína da Região Y Determinante do Sexo , Fatores de Transcrição/genética , Cromossomo Y/genética , Cromossomo Y/fisiologiaRESUMO
Although gonads are not required for development or survival, defects in gonadal development undoubtedly have a profound influence on affected individuals. Recent complementary studies in the fields of cytology, biochemistry and molecular genetics have revealed that normal gonad development involves an exquisitely regulated network of gene expression and protein-protein interactions. The initial event of gonadogenesis, in both males and females, involves the formation of a bipotential primordium. A Y chromosome then activates the male-specific pathway. The demonstration that mutations in the SOX proteins, SRY and SOX9, are responsible for disorders associated with male-to-female sex reversal showed dramatically that SRY and SOX9 have an essential role in male sex differentiation. This was emphasized when it was shown that female mice carrying transgenes that encode these proteins developed as males. SRY and SOX9 proteins have been characterized extensively and aspects of their function and regulation are now known.