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Objective: To identify the stapedotomy learning curve of two U.K. otolaryngologists. Study Design: A retrospective review of the outcome of first 100 stapedotomy operations by each surgeon. Included in the study was a postal survey of the incidence of stapes surgery by U.K. otolaryngologists. Setting: Two tertiary referral centers. Patients: All ears in which primary stapedotomy was performed for otosclerosis. Nonotosclerotic cases and malleus stapedotomy cases were excluded. Intervention: One surgeon used the technique of small fenestra stapedotomy with either a Teflon-wire or titanium piston but without vein graft interposition, whereas the second used the technique of stapedotomy with vein graft interposition and a Teflon piston. Main Outcome Measures: A moving average with a window of 15 dB was used to plot learning curves for the postoperative air-bone gaps. Using a postoperative air-bone gap of 20 dB or better as a definition of 'success,' the success rates with the increase in surgical experience of both surgeons were plotted on graphs, the learning curves. The end point of the learning curve was defined as the point where the curve reached its peak, and the results were sustainable. Results: There was no clear-cut end point in both learning curves, although it appears that there is a landmark point at 60 to 80 cases for both surgeons. Both surgeons also had one "dead ear" in their first 15 cases. The postal survey showed that some trainers only performed small numbers of stapes surgery, whereas some otolaryngologists who performed stapedotomies regularly were not trainers. Conclusions: The study supports a learning curve in stapes surgery. To maximize the training opportunity of trainee surgeons, it may be advisable for learning centers to form network to provide target training for the trainee who has demonstrated the necessary dexterity and temperament of an otologist. [ABSTRACT FROM AUTHOR]

Epidermal growth factor receptor (EGFR)-tyrosine kinase inhibitors (TKIs) are the standard of care for non-small-cell lung cancer (NSCLC) patients harboring EGFR mutations. However, almost all patients develop resistance after approximately 1 y of treatment, with >50% of cases due to the T790M secondary mutation of the EGFR gene. A large global Phase III study (AURA3) demonstrated that osimertinib significantly prolonged progression-free survival (PFS) over platinum-doublet chemotherapy in patients with T790M-positive NSCLC who had progressed on previous EGFR-TKI therapy. However, it is not clear whether efficacy or safety of osimertinib in Japanese patients is similar to the overall population. We report a pre-planned subgroup analysis of pooled Phase II data from the AURA Extension and AURA2 trials to investigate the efficacy and safety of osimertinib in Japanese patients. This study included 81 Japanese patients. Patients were administered 80 mg osimertinib orally once daily until disease progression. The main endpoints were objective response rate (ORR), PFS, and safety. The ORR was 63.6% and median PFS was 13.8 mo. Overall survival rate at 36 mo was 54.0%. The most common all-cause adverse events (AEs) were rash (grouped term; 65.4%), diarrhea (51.9%), paronychia (grouped term; 49.4%), and dry skin (grouped term; 39.5%). Most AEs were grade 1-2. Five patients (6.2%) developed interstitial lung disease, resulting in two deaths (2.5%). Osimertinib demonstrated favorable ORR and PFS in Japanese patients, similar to the overall population. Additionally, osimertinib has good efficacy and a manageable safety profile in Japanese patients with NSCLC who had acquired resistance due to the T790M mutation., (© 2019 The Authors. Cancer Science published by John Wiley & Sons Australia, Ltd on behalf of Japanese Cancer Association.)

Background: The FLAURA study was a multicenter, double-blind, Phase 3 study in which patients with previously untreated epidermal growth factor receptor mutation-positive advanced non-small-cell lung carcinoma were randomized 1:1 to oral osimertinib 80 mg once daily or standard-of-care (gefitinib 250 mg or erlotinib 150 mg, once daily) to compare safety and efficacy. In the overall FLAURA study, significantly better progression-free survival was shown with osimertinib versus standard-of-care., Methods: Selected endpoints, including progression-free survival (primary endpoint), overall survival, objective response rate, duration of response and safety were evaluated for the Japanese subset of the FLAURA study., Results: In Japan, 120 eligible Japanese patients were randomized to osimertinib (65 patients) or gefitinib (55 patients) treatment from December 2014 to June 2017. Median progression-free survival was 19.1 (95% confidence interval, 12.6, 23.5) and 13.8 (95% confidence interval, 8.3, 16.6) months with osimertinib and gefitinib, respectively (hazard ratio, 0.61; 95% confidence interval, 0.38, 0.99). Median overall survival was not reached in either treatment arm (data were immature). In the osimertinib and gefitinib arms, objective response rate was 75.4% (49/65) and 76.4% (42/55), and median duration of response from onset was 18.4 (95% confidence interval, not calculated) and 9.5 (95% confidence interval, 6.2, 13.9) months, respectively. The incidence of adverse events was similar in the two groups. The frequency of Grade ≥3 interstitial lung disease and pneumonitis in the two groups were the same (one patient)., Conclusions: As the first-line therapy, osimertinib showed significantly improved efficacy versus gefitinib in the Japanese population of the FLAURA study. No new safety concerns were raised., Clinical Trial Registration: NCT02296125 (ClinicalTrials.gov).

Purpose: Consensus is lacking regarding the androgen receptor (AR) as a prognostic marker in breast cancer. The objectives of this study were to comprehensively review the literature on AR prognostication and determine optimal criteria for AR as an independent predictor of breast cancer survival. Experimental Design: AR positivity was assessed by immunostaining in two clinically validated primary breast cancer cohorts [training cohort, n = 219; validation cohort, n = 418; 77% and 79% estrogen receptor alpha (ERα) positive, respectively]. The optimal AR cut-point was determined by ROC analysis in the training cohort and applied to both cohorts. Results: AR was an independent prognostic marker of breast cancer outcome in 22 of 46 (48%) previous studies that performed multivariate analyses. Most studies used cut-points of 1% or 10% nuclear positivity. Herein, neither 1% nor 10% cut-points were robustly prognostic. ROC analysis revealed that a higher AR cut-point (78% positivity) provided optimal sensitivity and specificity to predict breast cancer survival in the training (HR, 0.41; P = 0.015) and validation (HR, 0.50; P = 0.014) cohorts. Tenfold cross-validation confirmed the robustness of this AR cut-point. Patients with ERα-positive tumors and AR positivity ≥78% had the best survival in both cohorts ( P < 0.0001). Among the combined ERα-positive cases, those with comparable or higher levels of AR (AR:ERα-positivity ratio >0.87) had the best outcomes ( P < 0.0001). Conclusions: This study defines an optimal AR cut-point to reliably predict breast cancer survival. Testing this cut-point in prospective cohorts is warranted for implementation of AR as a prognostic factor in the clinical management of breast cancer. Clin Cancer Res; 24(10); 2328-41. ©2018 AACR ., (©2018 American Association for Cancer Research.)

Purpose: To develop an image analysis technique that distinguishes pseudoprogression from true progression by analyzing tumour heterogeneity in T2-weighted images using topological descriptors of image heterogeneity called Minkowski functionals (MFs)., Methods: Using a retrospective patient cohort (n = 50), and blinded to treatment response outcome, unsupervised feature estimation was performed to investigate MFs for the presence of outliers, potential confounders, and sensitivity to treatment response. The progression and pseudoprogression groups were then unblinded and supervised feature selection was performed using MFs, size and signal intensity features. A support vector machine model was obtained and evaluated using a prospective test cohort., Results: The model gave a classification accuracy, using a combination of MFs and size features, of more than 85% in both retrospective and prospective datasets. A different feature selection method (Random Forest) and classifier (Lasso) gave the same results. Although not apparent to the reporting radiologist, the T2-weighted hyperintensity phenotype of those patients with progression was heterogeneous, large and frond-like when compared to those with pseudoprogression., Conclusion: Analysis of heterogeneity, in T2-weighted MR images, which are acquired routinely in the clinic, has the potential to detect an earlier treatment response allowing an early change in treatment strategy. Prospective validation of this technique in larger datasets is required.

Due to increased sensitivity, the expression of circulating nucleotides is rapidly gaining popularity in cancer diagnosis. Whole blood mRNA has been used in studies on a number of cancers, most notably two separate studies that used whole blood mRNA to define non-overlapping signatures of prostate cancer that has become castration independent. Prostate cancer is known to rely on androgens for initial growth, and there is increasing evidence on the importance of the androgen axis in advanced disease. Using whole blood mRNA samples from patients with prostate cancer, we have identified the four-gene panel of FAM129A, MME, KRT7 and SOD2 in circulating mRNA that are differentially expressed in a discovery cohort of metastatic samples. Validation of these genes at the mRNA and protein level was undertaken in additional cohorts defined by risk of relapse following surgery and hormone status. All the four genes were downregulated at the mRNA level in the circulation and in primary tissue, but this was not always reflected in tissue protein expression. MME demonstrated significant differences in the hormone cohorts, whereas FAM129A is downregulated at the mRNA level but is raised at the protein level in tumours. Using published ChIP-seq data, we have demonstrated that this may be due to AR binding at the FAM129A and MME loci in multiple cell lines. These data suggest that whole blood mRNA of androgen-regulated genes has the potential to be used for diagnosis and monitoring of prostate cancer., (© 2016 Society for Endocrinology.)

Imaging of the metabolism of hyperpolarized [1-(13) C]pyruvate has shown considerable promise in preclinical studies in oncology, particularly for the assessment of early treatment response. The repeatability of measurements of (13) C label exchange between pyruvate and lactate was determined in a murine lymphoma model in fasted and non-fasted animals. The fasted state showed lower intra-individual variability, although the [1-(13) C]lactate/[1-(13) C]pyruvate signal ratio was significantly greater in fasted than in non-fasted mice, which may be explained by the higher tumor lactate concentrations in fasted animals. These results indicate that the fasted state may be preferable for the measurement of (13) C label exchange between pyruvate and lactate, as it reduces the variability and therefore should make it easier to detect the effects of therapy. © 2016 The Authors. NMR in Biomedicine published by John Wiley & Sons Ltd., (© 2016 The Authors. NMR in Biomedicine published by John Wiley & Sons Ltd.)

Background: The androgen receptor (AR) is a major drug target in prostate cancer (PCa). We profiled the AR-regulated kinome to identify clinically relevant and druggable effectors of AR signaling., Methods: Using genome-wide approaches, we interrogated all AR regulated kinases. Among these, choline kinase alpha (CHKA) expression was evaluated in benign (n = 195), prostatic intraepithelial neoplasia (PIN) (n = 153) and prostate cancer (PCa) lesions (n = 359). We interrogated how CHKA regulates AR signaling using biochemical assays and investigated androgen regulation of CHKA expression in men with PCa, both untreated (n = 20) and treated with an androgen biosynthesis inhibitor degarelix (n = 27). We studied the effect of CHKA inhibition on the PCa transcriptome using RNA sequencing and tested the effect of CHKA inhibition on cell growth, clonogenic survival and invasion. Tumor xenografts (n = 6 per group) were generated in mice using genetically engineered prostate cancer cells with inducible CHKA knockdown. Data were analyzed with χ(2) tests, Cox regression analysis, and Kaplan-Meier methods. All statistical tests were two-sided., Results: CHKA expression was shown to be androgen regulated in cell lines, xenografts, and human tissue (log fold change from 6.75 to 6.59, P = .002) and was positively associated with tumor stage. CHKA binds directly to the ligand-binding domain (LBD) of AR, enhancing its stability. As such, CHKA is the first kinase identified as an AR chaperone. Inhibition of CHKA repressed the AR transcriptional program including pathways enriched for regulation of protein folding, decreased AR protein levels, and inhibited the growth of PCa cell lines, human PCa explants, and tumor xenografts., Conclusions: CHKA can act as an AR chaperone, providing, to our knowledge, the first evidence for kinases as molecular chaperones, making CHKA both a marker of tumor progression and a potential therapeutic target for PCa., (© The Author 2015. Published by Oxford University Press.)

Background: Understanding the heterogeneous genotypes and phenotypes of prostate cancer is fundamental to improving the way we treat this disease. As yet, there are no validated descriptions of prostate cancer subgroups derived from integrated genomics linked with clinical outcome., Methods: In a study of 482 tumour, benign and germline samples from 259 men with primary prostate cancer, we used integrative analysis of copy number alterations (CNA) and array transcriptomics to identify genomic loci that affect expression levels of mRNA in an expression quantitative trait loci (eQTL) approach, to stratify patients into subgroups that we then associated with future clinical behaviour, and compared with either CNA or transcriptomics alone., Findings: We identified five separate patient subgroups with distinct genomic alterations and expression profiles based on 100 discriminating genes in our separate discovery and validation sets of 125 and 103 men. These subgroups were able to consistently predict biochemical relapse (p = 0.0017 and p = 0.016 respectively) and were further validated in a third cohort with long-term follow-up (p = 0.027). We show the relative contributions of gene expression and copy number data on phenotype, and demonstrate the improved power gained from integrative analyses. We confirm alterations in six genes previously associated with prostate cancer (MAP3K7, MELK, RCBTB2, ELAC2, TPD52, ZBTB4), and also identify 94 genes not previously linked to prostate cancer progression that would not have been detected using either transcript or copy number data alone. We confirm a number of previously published molecular changes associated with high risk disease, including MYC amplification, and NKX3-1, RB1 and PTEN deletions, as well as over-expression of PCA3 and AMACR, and loss of MSMB in tumour tissue. A subset of the 100 genes outperforms established clinical predictors of poor prognosis (PSA, Gleason score), as well as previously published gene signatures (p = 0.0001). We further show how our molecular profiles can be used for the early detection of aggressive cases in a clinical setting, and inform treatment decisions., Interpretation: For the first time in prostate cancer this study demonstrates the importance of integrated genomic analyses incorporating both benign and tumour tissue data in identifying molecular alterations leading to the generation of robust gene sets that are predictive of clinical outcome in independent patient cohorts.

Background: 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., Results: 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., Conclusions: 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.

Castration-resistant prostate cancer (CRPC) continues to pose a significant clinical challenge with new generation second-line hormonal therapies affording limited improvement in disease outcome. As the androgen receptor (AR) remains a critical driver in CRPC, understanding the determinants of its transcriptional activity is important for developing new AR-targeted therapies. FOXA1 is a key component of the AR transcriptional complex yet its role in prostate cancer progression and the relationship between AR and FOXA1 are not completely resolved. It is well established that FOXA1 levels are elevated in advanced prostate cancer and metastases. We mimicked these conditions by overexpressing FOXA1 in the androgen-responsive LNCaP prostate cancer cell line and observed a significant increase in AR genomic binding at novel regions that possess increased chromatin accessibility. High levels of FOXA1 resulted in increased proliferation at both sub-optimal and high 5α-dihydrotestosterone (DHT) concentrations. Immunohistochemical staining for FOXA1 in a clinical prostate cancer cohort revealed that high FOXA1 expression is associated with shorter time to biochemical recurrence after radical prostatectomy (hazard ratio (HR) 5.0, 95% confidence interval (CI) 1.2-21.1, P=0.028), positive surgical margins and higher stage disease at diagnosis. The gene expression program that results from FOXA1 overexpression is enriched for PTEN, Wnt and other pathways typically represented in CRPC gene signatures. Together, these results suggest that in an androgen-depleted state, elevated levels of FOXA1 enhance AR binding at genomic regions not normally occupied by AR, which in turn facilitates prostate cancer cell growth.

N-acetyl-L-aspartyl-L-glutamate peptidase-like 2 (NAALADL2) is a member of the glutamate carboxypeptidase II family, best characterized by prostate-specific membrane antigen (PSMA/NAALAD1). Using immunohistochemistry (IHC), we have shown overexpression of NAALADL2 in colon and prostate tumours when compared with benign tissue. In prostate cancer, NAALADL2 expression was associated with stage and Grade, as well as circulating mRNA levels of the NAALADL2 gene. Overexpression of NAALADL2 was shown to predict poor survival following radical prostatectomy. In contrast to PSMA/NAALAD1, NAALADL2 was localized at the basal cell surface where it promotes adhesion to extracellular matrix proteins. Using stable knockdown and overexpression cell lines, we have demonstrated NAALADL2-dependent changes in cell migration, invasion and colony-forming potential. Expression arrays of the knockdown and overexpression cell lines have identified nine genes that co-expressed with NAALADL2, which included membrane proteins and genes known to be androgen regulated, including the prostate cancer biomarkers AGR2 and SPON2. Androgen regulation was confirmed in a number of these genes, although NAALADL2 itself was not found to be androgen regulated. NAALADL2 was also found to regulate levels of Ser133 phosphorylated C-AMP-binding protein (CREB), a master regulator of a number of cellular processes involved in cancer development and progression. In combination, these data suggest that changes in expression of NAALADL2 can impact upon a number of pro-oncogenic pathways and processes, making it a useful biomarker for both diagnosis and prognosis.

Background: Identification of men harbouring insignificant prostate cancer (PC) is important in selecting patients for active surveillance. Tools have been developed in PSA-screened populations to identify such men based on clinical and biopsy parameters., Methods: Prospectively collected case series of 848 patients was treated with radical prostatectomy between July 2007 and October 2011 at an English tertiary care centre. Tumour volume was assessed by pathological examination. For each tool, receiver operator characteristics were calculated for predicting insignificant disease by three different criteria and the area under each curve compared. Comparison of accuracy in screened and unscreened populations was performed., Results: Of 848 patients, 415 had Gleason 3+3 disease on biopsy. Of these, 32.0% had extra-prostatic extension and 50.2% were upgraded. One had positive lymph nodes. Two hundred and six (24% of cohort) were D'Amico low risk. Of these, 143 had more than two biopsy cores involved. None of the tools evaluated has adequate discriminative power in predicting insignificant tumour burden. Accuracy is low in PSA-screened and -unscreened populations., Conclusions: In our unscreened population, tools designed to identify insignificant PC are inaccurate. Detection of a wider size range of prostate tumours in the unscreened may contribute to relative inaccuracy.

Castrate-resistant prostate cancer (CRPC) is poorly characterized and heterogeneous and while the androgen receptor (AR) is of singular importance, other factors such as c-Myc and the E2F family also play a role in later stage disease. HES6 is a transcription co-factor associated with stem cell characteristics in neural tissue. Here we show that HES6 is up-regulated in aggressive human prostate cancer and drives castration-resistant tumour growth in the absence of ligand binding by enhancing the transcriptional activity of the AR, which is preferentially directed to a regulatory network enriched for transcription factors such as E2F1. In the clinical setting, we have uncovered a HES6-associated signature that predicts poor outcome in prostate cancer, which can be pharmacologically targeted by inhibition of PLK1 with restoration of sensitivity to castration. We have therefore shown for the first time the critical role of HES6 in the development of CRPC and identified its potential in patient-specific therapeutic strategies.

Unlabelled: What's known on the subject? and What does the study add? Ureteric stents cause significant discomfort and this is probably related to ureteric smooth muscle spasm and trigonal irritation. Alpha-adrenoceptor antagonists reduce smooth muscle activity and are already widely used in medical expulsive therapy to aid passage of ureteric calculi. This meta-analysis incorporating five randomized controlled trials provides evidence that alpha-adrenoceptor antagonists reduce stent-related pain and storage symptoms as assessed by the Ureteric Stent Symptoms Questionnaire (USSQ)., Objectives: • To evaluate the efficacy of α-blockers with respect to improving stent-related symptoms. • Ureteric stents remain a source of marked discomfort and their placement is often required after certain ureteroscopic procedures or in the acute setting. This analysis identifies and reviews the several studies that have investigated the role of α-blockers after stent placement., Materials and Methods: • Pubmed/Medline, EMBASE, CINAHL and Cochrane Library databases were scrutinized using standard MeSH headings. • Randomized or controlled trials comparing α-blockers with control or standard therapy were included. • In all studies, patients completed the Ureteral Stent Symptom Questionnaire (USSQ). • The study data were independently reviewed by two assessors., Results: • In total, five studies of varying quality were identified, including 461 patients receiving either tamsulosin or alfuzosin, or control. • On meta-analysis, all five studies showed a reduction in USSQ urinary symptom score and body pain scores. There was mean reduction of 8.4 (95% CI, 5.6-11.1) in the urinary symptom score and 7.2 (95% CI, 2.5-11.8) in the body pain score. • In three studies, the numbers of patients experiencing stent related pain were stated: 45% (51/114) of patients receiving an α-blocker experienced painful episodes within the follow-up period defined for that study compared to 76% (88/116) in the control groups, which is equivalent to a relative risk of pain of 0.59 (95% confidence interval, 0.47-0.71). • There were also reductions in other aspects of the USSQ, such as the general health score and sexual matters score, although these were not statistically significant or uniformly reported., Conclusion: • There is evidence that α-blockers provide an improvement in discomfort after placement of a ureteric stent., (© 2011 THE AUTHORS. BJU INTERNATIONAL © 2011 BJU INTERNATIONAL.)

Objectives: Demand for out-of-hours cranial CT imaging is increasing and some departments have considered addressing this shortfall by allowing non-radiologists to provisionally report imaging studies. The aim of this work was to assess whether it is appropriate for non-radiologists to report head CTs by comparing the misreporting rates of those who regularly report head CTs with two groups of non-radiologists who do not usually report them: neuroradiographers and emergency doctors., Methods: 62 candidates were asked to report 30 head CTs, two-thirds of which were abnormal, and the results were compared by non-parametric statistical analysis., Results: There was no evidence of a difference in the score between neuroradiographers, neuroradiologists and general consultant radiologists. Neuroradiographers scored significantly higher than senior radiology trainees, and the emergency doctors scored least well., Conclusion: The results of this preliminary study show that appropriately trained neuroradiographers are competent at reporting the range of abnormalities assessed with this test and that their misreporting rates are similar to those who already independently report these studies.

Objective: • To study the outcomes and learning curve of robotic-assisted laparoscopic radical prostatectomy (RALP) in a single centre by two surgeons., Patients and Methods: • In total, 500 consecutive patients underwent RALP between 2005 and 2009 carried out by two surgeons. Using an ethically-approved database, prospective data collection of demographic, surgical, oncological and functional outcomes (patient reported) was performed, with up to 4 years of follow-up. • The learning curves of both surgeons were analyzed and, in addition, the first 100 and last 100 patients were compared to determine the effect of surgeon experience., Results: • The mean age of the patients was 61.5 years and mean preoperative prostate-specific antigen was 7.0 µg/L. Clinical stages were T1 in 63.2%, T2 in 33.8% and T3 in 3.0% of patients. Median (range) operating time was 170 (63-420) min and median (range) blood loss was 200 (20-3000) mL, with significant improvements for both surgeons with increasing experience (P < 0.001 and P= 0.029, respectively). • Pathological stages were pT2 in 53.4%, pT3a in 41.6%, pT3b in 4.0% and pT4 in 0.6% of patients. Overall, the positive margin rate (PMR) was 24.0% and stage-specific rates were 16.1%, 30.4%, 55.0% and 100.0% for pT2, pT3a, pT3b and pT4 disease, respectively. In the last 50 cases performed by each surgeon, the PMRs for pT2 and pT3a disease were 8.0% and 19.1% (surgeon 1) and 12.9% and 23.5% (surgeon 2). • At 12 months of follow-up, 91.3% of patients were continent and, by 48 months of follow-up, 75% of men with preoperative potency who underwent bilateral nerve-sparing RALP were potent., Conclusion: • This is the first report of two surgeons' learning curves in a single centre and shows that key learning curve outcomes continued to improve during the series, suggesting that the learning curve for RALP may be longer than has been previously suggested., (© 2010 THE AUTHORS. BJU INTERNATIONAL © 2010 BJU INTERNATIONAL.)

Background and Purpose: We describe a feasibility study testing the use of gold seeds for the identification of post-operative tumour bed after breast conservation surgery (BCS)., Materials and Methods: Fifty-three patients undergoing BCS for invasive cancer were recruited. Successful use was defined as all six seeds correctly positioned around the tumour bed during BCS, unique identification of all implanted seeds on CT planning scan and ≥ 3 seeds uniquely identified at verification to give couch displacement co-ordinates in 10/15 fractions. Planning target volume (PTV) margin size for four correction strategies were calculated from these data. Variability in tumour bed contouring was investigated with five radiation oncologists outlining five CT datasets., Results: Success in inserting gold seeds, identifying them at CT planning and using them for on-treatment verification was recorded in 45/51 (88%), 37/38 (97%) and 42/43 (98%) of patients, respectively. The clinicians unfamiliar with CT breast planning consistently contoured larger volumes than those already trained. Margin size ranged from 10.1 to 1.4mm depending on correction strategy., Conclusion: It is feasible to implant tumour bed gold seeds during BCS. Whilst taking longer to insert than surgical clips, they have the advantage of visibility for outlining and verification regardless of the ionising radiation beam quality. Appropriate correction strategies enable margins of the order of 5mm as required by the IMPORT trials however, tackling clinician variability in contouring is important., (Copyright © 2011 Elsevier Ireland Ltd. All rights reserved.)

Objective: To determine whether the density of CD4(+) and CD8(+) T-lymphocytes in a transrectal ultrasonography (TRUS) biopsy of the prostate can be used to predict the progression of lower urinary tract symptoms (LUTS) in benign prostatic hyperplasia (BPH)., Patients and Methods: In total, 100 patients were randomly selected from a pool of patients with histologically proven, benign TRUS biopsy specimens. There were seven full years of follow-up available. Clinical data were recorded, including prostate volume, International Prostate Symptom Score (IPSS), prostate-specific antigen, urine flow rate, postvoid residual urine volume and previous prostate surgery. Markers of disease progression included the subsequent development of acute urinary retention (AUR), ≥4 point rise in IPSS, prescription of medical therapy (α-blocker or 5-α-reductase inhibitor) and bladder outlet surgery. Four patients' specimens were unsuitable for analysis. Biopsy sections from 96 patients were immunohistochemically stained for the presence of CD4(+) and CD8(+) T-lymphocytes and the density of infiltrate was assessed using random field sampling and point counting., Results: Some 29% of patients (28/96) did not have BPH at the time of biopsy. Of all patients, 41% (39/96) progressed, 10% of whom (4/39) did not have BPH at the time of biopsy. A further 10% (10/96) developed AUR, 7% (7/96) had a ≥4 point rise in IPSS, 33% (32/96) required medical therapy for BPH and 11% (11/96) required bladder outlet surgery. There was low correlation between CD4(+) and CD8(+) densities in paired sections. CD4(+) and CD8(+) densities did not provide any significant predictive function in the progression of BPH, nor was their any predictive association noted between CD4(+) and CD8(+) scores and the development of prostate cancer. Sub-analysis did show that a threshold mean of ≥1.35 CD8(+) cells per field predicted progression to AUR with a sensitivity of 60% (95% confidence interval, CI, 26.2-87.8), specificity of 73.3% (95% CI 62.6-82.2) but a positive predictive value of 20.6% (95% CI 8.0-39.7). CD4(+) infiltrate density suggested a trend to general progression but without statistical significance., Conclusion: The present study, despite certain trends, shows no evidence for an association between CD4(+) and CD8(+) T-lymphocytes and the progression of LUTS in BPH., (© 2010 THE AUTHORS. BJU INTERNATIONAL © 2010 BJU INTERNATIONAL.)

Current standards of data presentation and analysis in biological journals often fall short of ideal. This is the first of a planned series of short articles, to be published in a number of journals, aiming to highlight the principles of clear data presentation and appropriate statistical analysis. This article considers the methods used to show data, in particular the value of the dot plot, and methods to summarise the distribution of values. The uses of measures such as standard deviation, standard error of the mean, and confidence intervals are contrasted.

Background and Aims: Although aberrant methylation of key genes in the progression of colorectal neoplasia has been reported, no model-based analysis of the incremental changes through the intermediate adenoma stage has been described. In addition, the biological drivers for these methylation changes have yet to be defined. Linear mixed-effects modelling was used in this study to understand the onset and patterns of the methylation changes of SFRP2, IGF2 DMR0, H19, LINE-1 and a CpG island methylator phenotype (CIMP) marker panel, and they were correlated with DNA methyltransferase 3B (DNMT3B) levels of expression in a sample set representative of colorectal neoplastic progression., Methods: Methylation of the above CpG islands was measured using quantitative pyrosequencing assays in 261 tissue samples. This included a prospective collection of 44 colectomy specimens with concurrent normal mucosa, adenoma and invasive cancer tissues. Tissue microarrays from a subset of 64 cases were used for immunohistochemical analysis of DNMT3B expression., Results: It is shown that the onset and pattern of methylation changes during colorectal neoplastic progression are locus dependent. The CIMP marker RUNX3 was the earliest CpG island showing significant change, followed by the CIMP markers NEUROG1 and CACNA1G at the hyperplastic polyp stage. SFRP2 and IGF2 DMR0 showed significant methylation changes at the adenomatous polyp stage, followed by the CIMP markers CDKN2A and hMLH1 at the adenocarcinoma stage. DNMT3B levels of immunohistochemical expression increased significantly (p < 0.001) from normal to hyperplastic and from adenomatous polyps to carcinoma samples. DNMT3B expression correlated positively with SFRP2 methylation (r = 0.42, p < 0.001, 95% CI 0.25 to 0.56), but correlated negatively with IGF2 DMR0 methylation (r = 0.26, p = 0.01, 95% CI -0.45 to -0.05). A subset of the CIMP panel (NEUROG1, CACNA1G and CDKN2A) positively correlated with DNMT3B levels of expression (p < 0.05)., Conclusion: Hierarchical epigenetic alterations occur at transition points during colorectal neoplastic progression. These cumulative changes are closely correlated with a gain of DNMT3B expression, suggesting a causal relationship.

Background & Aims: The incidence of esophageal and junctional adenocarcinoma has increased 6-fold in the past 30 years and 5-year survival remains approximately 20%. Current staging is limited in its ability to predict survival which has ramifications for treatment choices. The aim of this study was to generate and validate a molecular prognostic signature for esophageal adenocarcinoma., Methods: Gene expression profiling was performed and the resulting 42,000 gene signatures correlated with clinical and pathologic features for 75 snap-frozen esophageal and junctional resection specimens. External validation of selected targets was performed on 371 independent cases using immunohistochemistry to maximize clinical applicability., Results: A total of 119 genes were associated significantly with survival and 270 genes with the number of involved lymph nodes. Filtering of these lists resulted in a shortlist of 10 genes taken forward to validation. Four genes proved to be prognostic at the protein level (deoxycytidine kinase [DCK], 3'-phosphoadenosine 5'-phosphosulfate synthase 2 [PAPSS2], sirtuin 2 [SIRT2], and tripartite motif-containing 44 [TRIM44]) and were combined to create a molecular prognostic signature. This 4-gene signature was highly predictive of survival in the independent external validation cohort (0/4 genes dysregulated 5-year survival, 58%; 95% confidence interval [CI], 36%-80%; 1-2/4 genes dysregulated 5-year survival, 26%; 95% CI, 20%-32%; and 3-4/4 genes dysregulated 5-year survival, 14%; 95% CI, 4%-24% (P = .001). Furthermore, this 4-gene signature was independently prognostic in a multivariable model together with the existing clinical TNM staging system (P = .013)., Conclusions: This study has generated a clinically applicable prognostic gene signature that independently predicts survival in an external validation cohort and may inform management decisions., (Copyright © 2010 AGA Institute. Published by Elsevier Inc. All rights reserved.)

Introduction: Survival in small cell lung cancer (SCLC) is limited by the development of chemoresistance. Factors associated with chemoresistance in vitro have been difficult to validate in vivo. Both Bcl-2 and β(1)-integrin have been identified as in vitro chemoresistance factors in SCLC but their importance in patients remains uncertain. Tissue microarrays (TMAs) are useful to validate biomarkers but no large TMA exists for SCLC. We designed an SCLC TMA to study potential biomarkers of prognosis and then used it to clarify the role of both Bcl-2 and β(1)-integrin in SCLC., Methods: A TMA was constructed consisting of 184 cases of SCLC and stained for expression of Bcl-2 and β(1)-integrin. The slides were scored and the role of the proteins in survival was determined using Cox regression analysis. A meta-analysis of the role of Bcl-2 expression in SCLC prognosis was performed based on published results., Results: Both proteins were expressed at high levels in the SCLC cases. For Bcl-2 (n=140), the hazard ratio for death if the staining was weak in intensity was 0.55 (0.33-0.94, P=0.03) and for β(1)-integrin (n=151) was 0.60 (0.39-0.92, P=0.02). The meta-analysis showed an overall hazard ratio for low expression of Bcl-2 of 0.91(0.74-1.09)., Conclusions: Both Bcl-2 and β(1)-integrin are independent prognostic factors in SCLC in this cohort although further validation is required to confirm their importance. A TMA of SCLC cases is feasible but challenging and an important tool for biomarker validation.

Background & Aims: Increasing age is associated with impaired immune function and in chronic HCV infection specifically, with progressive fibrosis, liver failure, HCC and impaired responses to antiviral therapy. T-lymphocyte telomere length declines with age. We hypothesised that shorter T-lymphocyte telomere length would be associated with poor clinical outcome in HCV infection., Methods: Circulating T-lymphocyte telomere length, an objective measure of immune senescence, was measured by flow-FISH in 135 HCV-RNA-positive, treatment-naïve patients and 41 healthy controls in relation to clinical outcome., Results: Shorter CD4+CD45RO+ T-lymphocyte telomeres were associated with severe fibrosis (p=0.003), independent of male sex (p=0.04), CMV positivity (p=0.003), previous HBV infection (p=0.007), and age (p=ns) in viraemic patients compared to controls. There were inverse correlations between CD4+CD45RO+ telomere length and fibrosis stage (p<0.001), portal tract inflammatory grade (p=0.035), prothrombin time (p<0.001) and bilirubin (p=0.001). One hundred and twenty-four viraemic individuals were followed prospectively to a composite endpoint of death, hepatic decompensation or HCC. Independent of age, those with shorter CD4+CD45RO+ telomeres were less likely to be complication free after 2-years than those with longer telomeres (86% versus 96%, p=0.009) with an age-adjusted hazard ratio of 0.93 (0.90-0.96). In addition, CD4+CD45RO+ telomere length predicted successful antiviral therapy (p=0.001) independent of other factors., Conclusions: CD4+ T-lymphocyte telomere length, independent of age, was related to inflammatory grade, fibrosis stage, laboratory indices of severity, subsequent hepatic decompensation and treatment outcome in patients with chronic HCV infection., (Copyright 2010 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.)