Fusion of the NUP98 gene with the LEDGF/p52 gene defines a recurrent acute myeloid leukemia translocation

BACKGROUND: The NUP98 gene is involved in multiple rearrangements in haematological malignancy. The leukemic cells in an acute myeloid leukemia (AML) patient with a t(9;11)(p22;p15) were recently shown to have a fusion between the NUP98 gene and the LEDGF gene but it was not demonstrated that this fusion was recurrent in other leukaemia patients with the same translocation. RESULTS: We used RT-PCR to analyse the leukemic cells from an AML patient who presented with a cytogenetically identical translocation as the sole chromosomal abnormality. A NUP98-LEDGF fusion transcript was observed and confirmed by sequencing. The reciprocal transcript was also observed. The fusion transcript was not detectable during remission and recurred at relapse. The breakpoints in the NUP98 and LEDGF genes were different to those previously reported. The NUP98 breakpoint occurs in the intron between exons 8 and 9. It is the most 5' breakpoint reported in a translocation involving the NUP98 gene. All of the LEDGF gene is included in the fusion except for exon 1 which codes for the first 24 amino terminal amino acids. CONCLUSIONS: Our results show that fusion of the NUP98 and LEDGF genes is a new recurrent translocation in AML

microRNAs and Esophageal Cancer - Implications for Pathogenesis and Therapy

Author version made available in accordance with the publisher's policy.There are several microRNAs that have been consistently reported to be differentially expressed in esophageal squamous cell carcinoma vs. normal squamous tissue, with prognostic associations for miR-21 (invasion, positive nodes, decreased survival), miR-143 (disease recurrence, invasion depth), and miR-375 (inversely correlated with advanced stage, distant metastasis, poor overall survival, and disease-free survival). There is also evidence that miR-375 regulates gene expression associated with resistance to chemotherapy. Hence, microRNA expression assays have the potential to provide clinically relevant information about prognosis and potential response to chemotherapy in patients with esophageal squamous cell carcinoma. Results are inconsistent, however, for microRNAs across different studies for esophageal adenocarcinoma (EAC) vs. its precursor lesion Barrett’s esophagus. These inconsistencies may partly result from pathological and/or molecular heterogeneity in both Barrett’s esophagus and EAC, but may also result from differences in study designs or different choices of comparator tissues. Despite these inconsistencies, however, several mRNA/protein targets have been identified, the cancer related biology of some of these targets is well understood, and there are clinico-pathological associations for some of these mRNA targets. MicroRNAs also have potential for use in therapy for esophageal cancers. The development of new delivery methods, such as minicells and autologous microvesicles, and molecular modifications such as the addition of aromatic benzene pyridine analogs, have facilitated the exploration of the effects of therapeutic microRNAs in vivo. These approaches are producing encouraging results, with one technology in a phase I/IIa clinical trial

COX-2 mRNA is increased in oesophageal mucosal cells by a proton pump inhibitor

Author version made available in accordance with the publisher's policy.Introduction: Barrett’s oesophagus develops in some individuals with gastro-oesophageal reflux, and is the precursor to oesophageal adenocarcinoma. Proton pump inhibitors (PPIs) suppress gastric acid production and are used to treat reflux. Clinical trials suggest that COX inhibitors might prevent oesophageal cancer, although PPIs could offset this by increasing COX-2 expression in Barrett’s oesophagus. To investigate this, we evaluated the impact of a PPI on COX expression in oesophageal mucosal cells. Methods: The effect of the PPI esomeprazole on COX-1 and COX-2 mRNA levels in oesophageal cells was determined. Oesophageal cell lines OE33 (adenocarcinoma derived) and HET-1A (immortalized squamous cells), and a control intestinal cell line - HT29 (colon carcinoma), were treated for 24 hours with increasing concentrations of the esomeprazole. Results: COX-2, but not COX-1, mRNA levels, dose dependently increased in OE33 and HET-1A cells vs. esomeprazole concentration. COX-2 mRNA levels did not increase in HT29 cells. Conclusions: Exposure to esomeprazole increases COX-2 mRNA in oesophageal cells. This might contribute to the lack of benefit for COX inhibitors for oesophageal cancer prevention in recent clinica

Molecular biomarkers and ablative therapies for Barrett’s esophagus

Author version made available in accordance with the publisher's policy.Barrett’s esophagus is the major risk factor for esophageal adenocarcinoma. Endoscopic interventions which ablate Barrett’s esophagus mucosa lead to replacement with a new squamous (neosquamous) mucosa, but it can be difficult to achieve complete ablation. Knowing whether cancer is less likely to develop in neosquamous mucosa or residual Barrett’s esophagus after ablation is critical for determining the efficacy of treatment. This issue can be informed by assessing biomarkers that are associated with an increased risk of progression to adenocarcinoma. Although there are few post-ablation biomarker studies, evidence suggests that that neosquamous mucosa may have a reduced risk of adenocarcinoma in patients who have been treated for dysplasia or cancer, but some patients who do not have complete eradication of non-dysplastic Barrett’s esophagus may still be at risk. Biomarkers could be used to optimize endoscopic surveillance strategies following ablation, but this needs to be assessed by clinical studies and economic modeling

Ablation of Barrett's oesophagus: towards improved outcomes for oesophageal cancer?

This is the accepted version of the following article: Mayne, G. C., Bright, T., Hussey, D. J. and Watson, D. I. (2012), Ablation of Barrett's oesophagus: towards improved outcomes for oesophageal cancer?. ANZ Journal of Surgery, 82: 592–598, which has been published in final form at doi:10.1111/j.1445-2197.2012.06151.xBarrett's oesophagus is the major risk factor for oesophageal adenocarcinoma. The management of Barrett’s oesophagus entails treating reflux symptoms with acid-suppressing medication or surgery (fundoplication). However neither form of anti-reflux therapy produces predictable regression, or prevents cancer development. Patients with Barrett’s oesophagus usually undergo endoscopic surveillance which aims to identify dysplastic changes or cancer at its earliest stage, when treatment outcomes should be better. Alternative endoscopic interventions are now available and are suggested for the treatment of early cancer, and prevention of progression of Barrett’s oesophagus to cancer. Such treatments could minimize the risks associated with oesophagectomy. The current status of these interventions is reviewed. Various endoscopic interventions have been described, but with long term outcomes uncertain, they remain somewhat controversial. Radiofrequency ablation (RFA) of dysplastic Barrett’s oesophagus might reduce the risk of cancer progression, although cancer development has been reported after this treatment. Endoscopic mucosal resection (EMR) allows a 1.5 to 2 cm diameter piece of oesophageal mucosa to be removed. This provides better pathology for diagnosis and staging, and if the lesion is confined to the mucosa and fully excised, EMR can be curative. The combination of EMR and RFA has been used for multifocal lesions, but long term outcomes are unknown. The new endoscopic interventions for Barrett’s oesophagus and early oesophageal cancer have potential to improve clinical outcomes, although evidence which confirms superiority over oesphagectomy is limited. Longer term outcome data and data from larger cohorts is required to confirm the appropriateness of these procedure

A fixed-point algorithm for estimating amplification efficiency from a polymerase chain reaction dilution series

This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly credited. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.Background The polymerase chain reaction amplifies and quantifies small amounts of DNA. It is a cyclic process, during each cycle of which each strand of template DNA is copied with probability approaching one: the amount of DNA approximately doubles and this amount can be estimated fluorimetrically each cycle, producing a set of fluorescence values hereafter referred to as the amplification curve. Commonly the biological question of relevance is one of the ratio of DNA concentrations in two samples: a ratio that is deduced by comparing the two amplification curves, usually by way of a plot of fluorescence against cycle number. Central to this analysis is measuring the extent to which one amplification curve is shifted relative to the other, a measurement often accomplished by defining a threshold or quantification cycle, Cq, for each curve: the fractional cycle number at which fluorescence reaches some threshold or at which some other criterion (maximum slope, maximum rate of change of slope) is satisfied. We propose an alternative where position is measured relative to a reference curve; position equates to the cycle shift which maximizes the correlation between the reference and the observed fluorescence sequence. A key parameter of the reference curve is obtained by fixed-point convergence. Results We consider the analysis of dilution series constructed for the estimation of qPCR amplification efficiency. The estimate of amplification efficiency is based on the slope of the regression line when the Cq is plotted against the logarithm of dilution. We compare the approach to three commonly used methods for determining Cq; each is applied to publicly accessible calibration data sets, and to ten from our own laboratory. As in the established literature we judge their relative merits both from the standard deviation of the slope of the calibration curve, and from the variance in Cq for replicate fluorescence curves. Conclusions The approach does not require modification of experimental protocols, and can be applied retrospectively to existing data. We recommend that it be added to the methodological toolkit with which laboratories interpret their real-time PCR data. Keywords: qPCR; Fixed-point; Amplification efficienc

Circulating microRNAs: emerging biomarkers for diagnosis and prognosis in patients with gastrointestinal cancers

Author version made available in accordance with the publisher's policy. Under embargo for 6 months from time of publication. The final version of record is available at http://www.clinsci.org/cs/128/0001/cs1280001.htmTo identify novel non-invasive biomarkers for improved detection, risk assessment and prognostic evaluation of cancer, expression profiles of circulating microRNAs are currently under evaluation. Circulating microRNAs are highly promising candidates in this context, as they present some key characteristics for cancer biomarkers: they are tissue-specific with reproducible expression and consistency among individuals from the same species, they are potentially derived directly from the tumor and therefore might correlate with tumor progression and recurrence, and they are bound to proteins or contained in sub-cellular particles such as microvesicles or exosomes, making them highly stable and resistant to degradation. This review highlights the origin of circulating microRNAs, their stability in blood samples, and techniques to isolate exosomal microRNAs, and then addresses the current evidence supporting potential clinical applications for circulating miRNAs for diagnostic and prognostic purposes

Simplified Algorithms for Determining Cycle Shift between qPCR Fluorescence Curves

This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.The polymerase chain reaction is a central component of current molecular biology. It is a cyclic process, in each early cycle of which the template DNA approximately doubles. An indicator which fluoresces when bound to DNA quantifies the DNA present at the end of each cycle, giving rise to a fluorescence curve which is characteristically sigmoid in shape. The fluorescence curve quantifies the amount of DNA initially present; the more the initial DNA, the earlier the rise in the fluorescence. Accordingly the amount of DNA initially present in two samples can be compared: the sample with the less DNA gives rise to a relatively delayed fluorescence curve and the ratio of the DNAs can be deduced from the separation of the curves. There is, however, a second determinant of this separation, the fold increase in DNA per cycle: ideally a twofold increase but frequently less. Current guidelines recommend that this be determined experimentally by carrying out PCR on a series of dilutions. If the value of the fold increase is known, then the algorithm for determining the separation can be reduced to a relatively simple computation, rather than employing a multidimensional nonlinear optimization such as the Marquardt-Levenberg as currently employed

MicroRNAs and Their Impact on Radiotherapy for Cancer

Published version made available in accordance with publisher copyright policy.Resistance to radiation is considered to be an important reason for local failure after radiotherapy and tumor recurrence. However, the exact mechanisms of tumor resistance remain poorly understood. Current investigations of microRNAs as potential diagnostic and therapeutic tools for cancer treatment have shown promising results. With respect to radiotherapy resistance and response, there is now emerging evidence that microRNAs modulate key cellular pathways that mediate response to radiation. These data suggest that microRNAs might have significant potential as targets for the development of new therapeutic strategies to overcome radioresistance in cancer. This review summarizes the current literature pertinent to the influence of microRNAs in the response to radiotherapy for cancer treatment, with an emphasis on microRNAs as novel diagnostic and prognostic markers, as well as their potential to alter radiosensitivity

Proton pump inhibitors (PPIs) impact on tumour cell survival, metastatic potential and chemotherapy resistance, and affect expression of resistance-relevant miRNAs in esophageal cancer

This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly credited. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.Background Neoadjuvant treatment plays a crucial role in the therapy of advanced esophageal cancer. However, response to radiochemotherapy varies widely. Proton pump inhibitors (PPIs) have been demonstrated to impact on chemotherapy in a variety of other cancers. We analyzed the impact of PPI treatment on esophageal cancer cell lines, and investigated mechanisms that mediate the effect of PPI treatment in this tumour. Methods We investigated the effect of esomeprazole treatment on cancer cell survival, adhesion, migration and chemotherapy in human adeno-(OE19) and squamous-cell-carcinoma (KYSE410) cell lines. Furthermore, we investigated the effect of PPI treatment on intra-/extracellular pH and on expression of resistance-relevant miRNAs. Results Esomeprazole significantly inhibited tumour cell survival (in a dose-dependent manner), adhesion and migration in both tumour subtypes. Furthermore, esomeprazole augmented the cytotoxic effect of cisplatin and 5-FU in both tumour subtypes. Surprisingly, PPI treatment led to a significant increase of intracellular pH and a decrease of the extracellular pH. Finally, we found esomeprazole affected expression of resistance-relevant miRNAs. Specifically, miR-141 and miR-200b were upregulated, whereas miR-376a was downregulated after PPI treatment in both tumour types. Conclusion Our study demonstrates for the first time that PPIs impact on tumour cell survival, metastatic potential and sensitivity towards chemotherapy in esophageal cancer cell lines. Furthermore, we observed that in this tumour entity, PPIs do not lead to intracellular acidification, but affect the expression of resistance-relevant miRNAs. Keywords: Proton pump inhibitor; PPI; Esophageal cancer; Metastasis; Chemotherapy; Resistance; microRN