Genetically engineered mouse models of cancer can be used to filter genome‐wide expression datasets generated from human tumours and to identify gene expression alterations that are functionally ...important to cancer development and progression. In this study, we have generated RNAseq data from tumours arising in two established mouse models of prostate cancer, PB‐Cre/PtenloxP/loxP and p53loxP/loxPRbloxP/loxP, and integrated this with published human prostate cancer expression data to pinpoint cancer‐associated gene expression changes that are conserved between the two species. To identify potential therapeutic targets, we then filtered this information for genes that are either known or predicted to be druggable. Using this approach, we revealed a functional role for the kinase MELK as a driver and potential therapeutic target in prostate cancer. We found that MELK expression was required for cell survival, affected the expression of genes associated with prostate cancer progression and was associated with biochemical recurrence.
Synopsis
Genetically engineered mouse models (GEMM) of human cancers can be used as “biological filters” to identify genes that are important to cancer development and prioritise potential therapeutic targets. The example of MELK in prostate cancer.
Comparisons of high‐throughput sequencing data from two GEMM of prostate cancer with human prostate cancer gene expression data revealed that human and murine tumours share common gene expression alterations.
Integration of human and murine gene expression data was used to identify potential therapeutic targets in prostate cancer, including the protein kinase MELK.
MELK is overexpressed in prostate cancer and associated with poor prognosis.
Silencing of MELK or treatment with an inhibitor targeting MELK resulted in induction of apoptosis in prostate cancer cells in vitro and in vivo.
Genetically engineered mouse models (GEMM) of human cancers can be used as “biological filters” to identify genes that are important to cancer development and prioritise potential therapeutic targets. The example of MELK in prostate cancer.
Osteosarcoma (OSA) is the most common type of primary bone malignancy in people and dogs. Our previous molecular comparisons of canine OSA against healthy bone resulted in the identification of ...differentially expressed protein-expressing genes (forkhead box protein O4 (
), interferon regulatory factor 8 (
), and lymphoid enhancer binding factor 1 (
)). Immunohistochemistry (IHC) and H-scoring provided semi-quantitative assessment of nuclear and cytoplasmic staining alongside qualitative data to contextualise staining (
= 26 patients). FOXO4 was expressed predominantly in the cytoplasm with significantly lower nuclear H-scores. IRF8 H-scores ranged from 0 to 3 throughout the cohort in the nucleus and cytoplasm. LEF1 was expressed in all patients with significantly lower cytoplasmic staining compared to nuclear. No sex or anatomical location differences were observed. While reduced levels of FOXO4 might indicate malignancy, the weak or absent protein expression limits its primary use as diagnostic tumour marker. IRF8 and LEF1 have more potential for prognostic and diagnostic uses and facilitate further understanding of their roles within their respective molecular pathways, including Wnt/beta-catenin/LEF1 signalling and differential regulation of tumour suppressor genes. Deeper understanding of the mechanisms involved in OSA are essential contributions towards the development of novel diagnostic, prognostic, and treatment options in human and veterinary medicine contexts.
The discovery of cytosine hydroxymethylation (5hmC) as a mechanism that potentially controls DNA methylation changes typical of neoplasia prompted us to investigate its behaviour in colon cancer. ...5hmC is globally reduced in proliferating cells such as colon tumours and the gut crypt progenitors, from which tumours can arise.
Here, we show that colorectal tumours and cancer cells express Ten-Eleven-Translocation (TET) transcripts at levels similar to normal tissues. Genome-wide analyses show that promoters marked by 5hmC in normal tissue, and those identified as TET2 targets in colorectal cancer cells, are resistant to methylation gain in cancer. In vitro studies of TET2 in cancer cells confirm that these promoters are resistant to methylation gain independently of sustained TET2 expression. We also find that a considerable number of the methylation gain-resistant promoters marked by 5hmC in normal colon overlap with those that are marked with poised bivalent histone modifications in embryonic stem cells.
Together our results indicate that promoters that acquire 5hmC upon normal colon differentiation are innately resistant to neoplastic hypermethylation by mechanisms that do not require high levels of 5hmC in tumours. Our study highlights the potential of cytosine modifications as biomarkers of cancerous cell proliferation.
Abstract Background Localized prostate cancer is clinically heterogeneous, despite clinical risk groups that represent relative prostate cancer-specific mortality. We previously developed a 100-locus ...DNA classifier capable of substratifying patients at risk of biochemical relapse within clinical risk groups. Objective The 100-locus genomic classifier was refined to 31 functional loci and tested with standard clinical variables for the ability to predict biochemical recurrence (BCR) and metastasis. Design, setting, and participants Four retrospective cohorts of radical prostatectomy specimens from patients with localized disease were pooled, and an additional 102-patient cohort used to measure the 31-locus genomic classifier with the NanoString platform. Outcome measurements and statistical analysis The genomic classifier scores were tested for their ability to predict BCR ( n = 563) and metastasis ( n = 154), and compared with clinical risk stratification schemes. Results and limitations The 31-locus genomic classifier performs similarly to the 100-locus classifier. It identifies patients with elevated BCR rates (hazard ratio = 2.73, p < 0.001) and patients that eventually develop metastasis (hazard ratio = 7.79, p < 0.001). Combining the genomic classifier with standard clinical variables outperforms clinical models. Finally, the 31-locus genomic classifier was implemented using a NanoString assay. The study is limited to retrospective cohorts. Conclusions The 100-locus and 31-locus genomic classifiers reliably identify a cohort of men with localized disease who have an elevated risk of failure. The NanoString assay will be useful for selecting patients for treatment deescalation or escalation in prospective clinical trials based on clinico-genomic scores from pretreatment biopsies. Patient summary It is challenging to determine whether tumors confined to the prostate are aggressive, leading to significant undertreatment and overtreatment. We validated a test based on prostate tumor DNA that improves estimations of relapse risk, and that can help guide treatment planning.
With the exception of classic functional adenomas in dogs and horses, pituitary lesions are infrequently described in the veterinary literature. Approximately 10% of pituitary glands from ...asymptomatic humans contain abnormalities, but the equivalent proportion in small animals is unknown. Pituitary glands from 136 dogs and 65 cats collected during routine necropsies were examined to determine the prevalence of pituitary lesions and their histopathological diagnosis. Lesions were characterized in sections stained with hematoxylin and eosin, periodic acid-Schiff (PAS), Gordon and Sweet’s and reticulin stains, and immunohistochemistry for adrenocorticotropic hormone (ACTH), growth hormone, melanocyte stimulating hormone–α, and prolactin. Pituitary abnormalities were identified in 36 of 136 (26.4%) dogs and 10 of 65 (15.3%) cats. Cystic changes were the most common lesion, occurring in 18 (13.2%) dogs and 8 (12.3%) cats. Pituitary neoplasia was detected in 14.1% (12/85) of middle-aged and old dogs; 1 (1.5%) cat had pituitary nodular hyperplasia. PAS and reticulin stains helped differentiate ACTH-immunoreactive adenomas from hyperplastic nodules: adenomas contained PAS-positive intracytoplasmic granules and loss of the normal reticulin network. One dog had a pituitary carcinoma with infiltration into the thalamus. Other pituitary abnormalities included secondary metastases (2 dogs) and hypophysitis (4 dogs, 1 cat). In most cases, the lesion appeared to be subclinical and could be considered incidental, whereas clinical manifestations were apparent in only 4 dogs (2.9%) and none of the cats with pituitary lesions. Pituitary abnormalities are common in dogs and cats, and their clinical relevance requires further investigation.
Abstract The androgen receptor (AR) is the dominant growth factor in prostate cancer (PCa). Therefore, understanding how ARs regulate the human transcriptome is of paramount importance. The early ...effects of castration on human PCa have not previously been studied 27 patients medically castrated with degarelix 7 d before radical prostatectomy. We used mass spectrometry, immunohistochemistry, and gene expression array (validated by reverse transcription-polymerase chain reaction) to compare resected tumour with matched, controlled, untreated PCa tissue. All patients had levels of serum androgen, with reduced levels of intraprostatic androgen at prostatectomy. We observed differential expression of known androgen-regulated genes ( TMPRSS2, KLK3, CAMKK2, FKBP5 ). We identified 749 genes downregulated and 908 genes upregulated following castration. AR regulation of α-methylacyl-CoA racemase expression and three other genes ( FAM129A, RAB27A , and KIAA0101 ) was confirmed. Upregulation of oestrogen receptor 1 (ESR1) expression was observed in malignant epithelia and was associated with differential expression of ESR1-regulated genes and correlated with proliferation (Ki-67 expression). Patient summary This first-in-man study defines the rapid gene expression changes taking place in prostate cancer (PCa) following castration. Expression levels of the genes that the androgen receptor regulates are predictive of treatment outcome. Upregulation of oestrogen receptor 1 is a mechanism by which PCa cells may survive despite castration.
Objective-To compare long-term survival and quality of life data in dogs with clinical signs associated with a congenital portosystemic shunt (CPSS) that underwent medical or surgical treatment. ...Design-Prospective cohort study. Animals-124 client-owned dogs with CPSS. Procedures-Dogs received medical or surgical treatment without regard to signalment, clinical signs, or clinicopathologic results. Survival data were analyzed with a Cox regression model. Quality of life information, obtained from owner questionnaires, included frequency of CPSS-associated clinical signs (from which a clinical score was derived), whether owners considered their dog normal, and (for surgically treated dogs) any ongoing medical treatment for CPSS. A Mann-Whitney U test was used to compare mean clinical score data between surgically and medically managed dogs during predetermined follow-up intervals. Results-97 dogs underwent surgical treatment; 27 were managed medically. Median follow-up time for all dogs was 1,936 days. Forty-five dogs (24 medically managed and 21 surgically managed) died or were euthanized during the follow-up period. Survival rate was significantly improved in dogs that underwent surgical treatment (hazard ratio, 8.11; 95% CI, 4.20 to 15.66) than in those treated medically for CPSS. Neither age at diagnosis nor shunt type affected survival rate. Frequency of clinical signs was lower in surgically versus medically managed dogs for all follow-up intervals, with a significant difference between groups at 4 to 7 years after study entry. Conclusions and Clinical Relevance-Surgical treatment of CPSS in dogs resulted in significantly improved survival rate and lower frequency of ongoing clinical signs, compared with medical management. Age at diagnosis did not affect survival rate and should not influence treatment choice.
Cardiac disease is a leading cause of morbidity and mortality in dogs and humans, with dilated cardiomyopathy being a large contributor to this. The Irish Wolfhound (IWH) is one of the most commonly ...affected breeds and one of the few breeds with genetic loci associated with the disease. Mutations in more than 50 genes are associated with human dilated cardiomyopathy (DCM), yet very few are also associated with canine DCM. Furthermore, none of the identified canine loci explain many cases of the disease and previous work has indicated that genotypes at multiple loci may act together to influence disease development. In this study, loci previously associated with DCM in IWH were tested for associations in a new cohort both individually and in combination. We have identified loci significantly associated with the disease individually, but no genotypes individually or in pairs conferred a significantly greater risk of developing DCM than the population risk. However combining three loci together did result in the identification of a genotype which conferred a greater risk of disease than the overall population risk. This study suggests multiple rather than individual genetic factors, cooperating to influence DCM risk in IWH.
Methods that do not require animal sacrifice to detect botulinum neurotoxins (BoNTs) are critical for BoNT antagonist discovery and the advancement of quantitative assays for biodefense and ...pharmaceutical applications. Here we describe the development and optimization of fluorogenic reporters that detect the proteolytic activity of BoNT/A, B, D, E, F, and G serotypes in real time with femtomolar to picomolar sensitivity. Notably, the reporters can detect femtomolar concentrations of BoNT/A in 4h and BoNT/E in 20h, sensitivity that equals that of animal-based methods. The reporters can be used to determine the specific activity of BoNT preparations with intra- and inter-assay coefficients of variation of approximately 10%. Finally, we find that the greater sensitivity of our reporters compared with those used in other commercially available assays makes the former attractive candidates for high-throughput screening of BoNT antagonists.
PDF
