Balancing neurotrophin pathway and sortilin function: its role in human disease

Neurotensin receptor-3 or sortilin is a vacuolar protein sorting 10 protein domain (Vps10p) has been firstly discovered in the human brain, it acts as receptor or co-receptor of the cell and traffics different proteins within the cell. Sortilin deregulation contributes to the development of several diseases, including neurological diseases and cancer. On the other hand, neurotrophins which are a family of proteins essential for the nervous system development, function and plasticity. The first discovered member is the nerve growth factor; other members are brain-derived growth factor, neurotrophin-3 and neurotrophin-4. Nerve growth factor and brain-derived growth factor are the common neurotrophins that have a role in cancer. Neurotrophins initiate their signals through interaction with tyrosine receptor kinases TrkA, TrkB, and TrkC; each member has an affinity for a specific receptor to stimulate cell survival, while the interaction with p75NTR initiates cell apoptosis pathway by forming a complex with sortilin and neurotrophin precursors. A number of therapeutic approaches are emerging to target the neurotrophins pathway as well as sortilin

AURKA mRNA expression is an independent predictor of poor prognosis in patients with non-small cell lung cancer

Deregulation of mitotic spindle genes has been reported to contribute to the development and progression of malignant tumours. The aim of the present study was to explore the association between the expression profiles of Aurora kinases (AURKA, AURKB and AURKC), cytoskeleton-associated protein 5 (CKAP5), discs large-associated protein 5 (DLGAP5), kinesin-like protein 11 (KIF11), microtubule nucleation factor (TPX2), monopolar spindle 1 kinase (TTK), and β-tubulins (TUBB) and (TUBB3) genes and clinicopathological characteristics in human non-small cell lung carcinoma (NSCLC). Reverse transcription-quantitative polymerase chain reaction-based RNA gene expression profiles of 132 NSCLC and 44 adjacent wild-type tissues were generated, and Cox's proportional hazard regression was used to examine associations. With the exception of AURKC, all genes exhibited increased expression in NSCLC tissues. Of the 10 genes examined, only AURKA was significantly associated with prognosis in NSCLC. Multivariate Cox's regression analysis demonstrated that AURKA mRNA expression [hazard ratio (HR), 1.81; 95% confidence interval (CI), 1.16-2.84; P=0.009], age (HR, 1.03; 95% CI, 1.00-1.06; P=0.020), pathological tumour stage 2 (HR, 2.43; 95% CI, 1.16-5.10; P=0.019) and involvement of distal nodes (pathological node stage 2) (HR, 3.14; 95% CI, 1.24-7.99; P=0.016) were independent predictors of poor prognosis in patients with NSCLC. Poor prognosis of patients with increased AURKA expression suggests that those patients may benefit from surrogate therapy with AURKA inhibitors

Methylation enrichment pyrosequencing: combining the specificity of MSP with validation by pyrosequencing

It has been suggested that detection of aberrant DNA methylation in clinical specimens such as sputum or saliva may be a valuable tumour biomarker. Any clinically applicable detection technique must combine high sensitivity with high specificity. In this study we describe methylation enrichment pyrosequencing (MEP), which benefits from the high sensitivity and specificity of methylation-specific PCR (MSP) but has a second, confirmatory, pyrosequencing step. The pyrosequencing reaction is rapid, relatively inexpensive and offers significant logistical advantages over previously described validation methods. As proof of principle, we illustrate MEP using assays of p16 and cyclin A1 promoters in a methylated DNA dilution matrix and also in a clinical setting using paired saliva and oral tumour specimens. Our results confirm that mis-priming of MSP, with subsequent false positive results, can occur frequently (perhaps 10%) in assays combining high numbers of PCR cycles and low concentrations of starting DNA. In our clinical example, MEP of saliva-derived DNA was more sensitive than standard non-methylation-specific pyrosequencing as illustrated using p16 and cyclin A1 promoter methylation assays

Detection of hepatitis Β virus DNA and mutations in K-ras and p53 genes in human hepatocellular carcinomas

Journal URL: http://www.spandidos-publications.com/ijo/Hepatitis Β virus (HBV) infection is considered as one of the major risk factors in the development of human hepatocellular carcinoma (HCC). Recent studies have also suggested the implication of oncogene and onco-suppressor genes in liver carcinogenesis. We studied 41 cases of HCC for the presence of HBV DNA and point mutations in codon 12 of K-ras and codon 249 of p53. We used 'nested' PCR for the amplification of HBV because of the expected low incidence of the virus DNA in the samples. PCR was also used for the amplification of K-ras and p53 regions that contain the codons of interest, followed by RFLP analysis for the detection of point mutations. HBV DNA was amplified in 22 cases (53.7%), while 5 cases (12.2%) appeared to carry mutations in codon 12 of K-ras and 7 cases (17.1%) had mutations in codon 249 of the p53 gene. These results further support the correlation between HBV infection and HCC and also indicate an implication of K-ras and p53 genes in hepatocarcinogenesis

Superfast Broadband and Rural Community Resilience : Examining the Rural Need for Speed

Acknowledgements The authors would also like to thank colleagues at the University of Aberdeen and Scotland's Rural College for feedback and input throughout the early research process. Funding: This work is supported by the University of Aberdeen College of Physical Sciences as well as the dot.rural Digital Economy Research Hub at the University of Aberdeen, award reference EP/G066051/1.Peer reviewedPublisher PD

p53 gene aberrations in non-small-cell lung carcinomas from a smoking population.

We examined 46 non-small-cell lung carcinomas (NSCLCs) for the presence of p53 mutations in exons 4-9, positive p53 immunostaining and loss of heterozygosity (LOH) in the TP53 locus. p53 mutations were detected in 13 tumour samples (28.3%), whereas overexpression of the p53 protein was found in 30 of 45 (67%) samples. Allelic loss was found in 9 of 38 (23.6%) informative cases. The statistical analysis revealed no significant correlation between p53 mutations and clinicopathological data, although mutations appear to occur more frequently in squamous cell carcinomas (7 of 18) than in adenocarcinomas (2 of 15). All but three individuals in this study group smoked. In contrast to previous reports, we found a higher prevalence of GC-->AT transitions than of GC-->TA transversions, as expected in a smoking population. A trend was found between p53-positive immunostaining and a history of heavy smoking (76-126 pack-years) and was inversely correlated with allelic deletion (LOH) at the TP53 locus. Eight of the 12 NSCLCs containing p53 mutations also had concomitant p53 overexpression, and it is of specific note that three of the four tumours containing p53 'mutations' with no overexpression of the p53 protein had either insertions or deletions in the p53 gene. No correlation was found between p53 mutations and fractional allele loss or ras mutations. p53 mutations in this Merseyside population in the UK do not appear to be as common as in other reports for NSCLC and exhibit predominance of GC-->AT transitions preferentially at non-CpG sites, suggesting that other carcinogens in addition to those in tobacco smoke may be involved in NSCLC in the Merseyside area of the UK

METH-2 silencing and promoter hypermethylation in NSCLC

The antiangiogenic factor METH-2 (ADAMTS-8) was identified in a previous dual-channel cDNA microarray analysis to be at least two-fold under-represented in 85% (28 out of 33) of primary non-small-cell lung carcinomas (NSCLCs). This observation has been validated in an independent series of NSCLCs and adjacent normal tissues by comparative multiplex RT—PCR, and METH-2 mRNA expression was dramatically reduced in all 23 tumour samples analysed. Immunohistochemical analysis of the same sample set demonstrated that METH-2 was strongly expressed in 14 out of 19 normal epithelial sites examined but only one out of 20 NSCLCs. DNA methylation analysis of the proximal promoter region of this gene revealed abnormal hypermethylation in 67% of the adenocarcinomas and 50% of squamous cell carcinomas, indicating that epigenetic mechanisms are involved in silencing this gene in NSCLC. No homozygous deletions of METH-2 were found in lung cancer cell lines. Allelic imbalance in METH-2 was assessed by an intronic single nucleotide polymorphism (SNP) assay and observed in 44% of informative primary samples. In conclusion, the downregulation of METH-2 expression in primary NSCLC, often associated with promoter hypermethylation, is a frequent event, which may be related to the development of the disease

Activating transcription factor-2 (ATF2) is a key determinant of resistance to endocrine treatment in an in vitro model of breast cancer.

BACKGROUND: Activating transcription factor-2 (ATF2), a member of the leucine zipper family of DNA binding proteins, has been implicated as a tumour suppressor in breast cancer. However, its exact role in breast cancer endocrine resistance is still unclear. We have previously shown that silencing of ATF2 leads to a loss in the growth-inhibitory effects of tamoxifen in the oestrogen receptor (ER)-positive, tamoxifen-sensitive MCF7 cell line and highlighted that this multi-faceted transcription factor is key to the effects of tamoxifen in an endocrine sensitive model. In this work, we explored further the in vitro role of ATF2 in defining the resistance to endocrine treatment. MATERIALS AND METHODS: We knocked down ATF2 in TAMR, LCC2 and LCC9 tamoxifen-resistant breast cancer cell lines as well as the parental tamoxifen sensitive MCF7 cell line and investigated the effects on growth, colony formation and cell migration. We also performed a microarray gene expression profiling (Illumina Human HT12_v4) to explore alterations in gene expression between MCF7 and TAMRs after ATF2 silencing and confirmed gene expression changes by quantitative RT-PCR. RESULTS: By silencing ATF2, we observed a significant growth reduction of TAMR, LCC2 and LCC9 with no such effect observed with the parental MCF7 cells. ATF2 silencing was also associated with a significant inhibition of TAMR, LCC2 and LCC9 cell migration and colony formation. Interestingly, knockdown of ATF2 enhanced the levels of ER and ER-regulated genes, TFF1, GREB1, NCOA3 and PGR, in TAMR cells both at RNA and protein levels. Microarray gene expression identified a number of genes known to mediate tamoxifen resistance, to be differentially regulated by ATF2 in TAMR in relation to the parental MCF7 cells. Moreover, differential pathway analysis confirmed enhanced ER activity after ATF2 knockdown in TAMR cells. CONCLUSION: These data demonstrate that ATF2 silencing may overcome endocrine resistance and highlights further the dual role of this transcription factor that can mediate endocrine sensitivity and resistance by modulating ER expression and activity

CpG island methylation phenotype (CIMP) in oral cancer: associated with a marked inflammatory response and less aggressive tumour biology.

Studies in several tumour sites highlight the significance of the CpG island methylation phenotype (CIMP), with distinct features of histology, biological aggression and outcome. We utilise pyrosequencing techniques of quantitative methylation analysis to investigate the presence of CIMP in oral squamous cell carcinoma (OSCC) for the first time, and evaluate its correlation with allelic imbalance, pathology and clinical behaviour. Tumour tissue, control tissue and PBLs were obtained from 74 patients with oral squamous cell carcinoma. Pyrosequencing was used to analyse methylation patterns in 75-200 bp regions of the CpG rich gene promoters of 10 genes with a broad range of cellular functions. Allelic imbalance was investigated using a multiplexed panel of 11 microsatellite markers. Corresponding variables, histopathological staging and grading were correlated with these genetic and epigenetic aberrations. A cluster of tumours with a greater degree of promoter methylation than would be predicted by chance alone (P=0.001) were designated CIMP+ve. This group had less aggressive tumour biology in terms of tumour thickness (p=0.015) and nodal metastasis (P=0.012), this being apparently independent of tumour diameter. Further, it seems that these CIMP+ve tumours excited a greater host inflammatory response (P=0.019). The exact mechanisms underlying CIMP remain obscure but the association with a greater inflammatory host response supports existing theories relating these features in other tumour sites. As CIMP has significant associations with other well documented prognostic indicators, it may prove beneficial to include methylation analyses in molecular risk modelling of tumours