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The purpose of this study was to investigate rigid ankle-foot orthosis (AFO) prescription practices for adult men among UK orthotists.A cross-sectional study using a survey was distributed online to UK orthotists by the British Association of Prosthetists and Orthotists to its members and through social media and orthotic networks. The survey was completed between November 1, 2020, and November 29, 2020.Descriptive statistics of survey results include information related to the material used, the thickness of the material, positive cast rectification, AFO reinforcement, footplate design, padding, strapping system, and height of AFO.One hundred participants completed the survey, which equates to a response rate of 30.5% of the British Association of Prosthetists and Orthotists members targeted. A clear consensus emerged on the design of a bespoke rigid AFO for the hypothetical patient in this study, which is detailed as follows: 1) 4.5 mm copolymer polypropylene, 2) no additional reinforcement, 3) full-length footplate with mediolateral trimlines terminating behind the metatarsal heads, 4) 3-point correction with parallel sides, 5) padded VELCRO straps with D-rings at the calf and heel, 6) no forefoot or other additional strapping, 7) 3-mm PORON (international Ltd) padding at the malleoli, and 8) AFO height that finishes 2 cm below the fibular head.This study has highlighted a consensus on AFO prescription/design among UK orthotists surveyed, based on the hypothetical patient described in this study.

Objectives To assess whether a commercially available CE-IVD, ELISA-based surrogate neutralisation assay (cPass, Genscript) provides a genuine measure of SARS-CoV-2 neutralisation by human sera, and further to establish whether measuring responses against the RBD of S was a diagnostically useful proxy for responses against the whole S protein.Methods Serum samples from 30 patients were assayed for anti-NP responses, for ‘neutralisation’ by the surrogate neutralisation assay and for neutralisation by SARS-CoV-2 S pseudotyped virus assays utilising two target cell lines. Correlation between assays was measured using linear regression.Results The responses observed within the surrogate neutralisation assay demonstrated an extremely strong, highly significant positive correlation with those observed in both pseudotyped virus assays.Conclusions The tested ELISA-based surrogate assay provides an immunologically useful measure of functional immune responses in a much quicker and highly automatable fashion. It also reinforces that detection of anti-RBD neutralising antibodies alone is a powerful measure of the capacity to neutralise viral infection.

Purpose Retrospective studies indicate that expression of excision repair cross complementing group 1 (ERCC1) protein is associated with platinum resistance and survival in non–small-cell lung cancer (NSCLC). We conducted the first randomized trial, to our knowledge, to evaluate ERCC1 prospectively and to assess the superiority of nonplatinum therapy over platinum doublet therapy for ERCC1-positive NSCLC as well as noninferiority for ERCC1-negative NSCLC. Patients and Methods This trial had a marker-by-treatment interaction phase III design, with ERCC1 (8F1 antibody) status as a randomization stratification factor. Chemonaïve patients with NSCLC (stage IIIB and IV) were eligible. Patients with squamous histology were randomly assigned to cisplatin and gemcitabine or paclitaxel and gemcitabine; nonsquamous patients received cisplatin and pemetrexed or paclitaxel and pemetrexed. Primary end point was overall survival (OS). We also evaluated an antibody specific for XPF (clone 3F2). The target hazard ratio (HR) for patients with ERCC1-positive NSCLC was ≤ 0.78. Results Of patients, 648 were recruited (177 squamous, 471 nonsquamous). ERCC1-positive rates were 54.5% and 76.7% in nonsquamous and squamous patients, respectively, and the corresponding XPF-positive rates were 70.5% and 68.5%. Accrual stopped early in 2012 for squamous patients because OS for nonplatinum therapy was inferior to platinum therapy (median OS, 7.6 months [paclitaxel and gemcitabine] v 10.7 months [cisplatin and gemcitabine]; HR, 1.46; P = .02). Accrual for nonsquamous patients halted in 2013. Median OS was 8.0 (paclitaxel and pemetrexed) versus 9.6 (cisplatin and pemetrexed) months for ERCC1-positive patients (HR, 1.11; 95% CI, 0.85 to 1.44), and 10.3 (paclitaxel and pemetrexed) versus 11.6 (cisplatin and pemetrexed) months for ERCC1-negative patients (HR, 0.99; 95% CI, 0.73 to 1.33; interaction P = .64). OS HR was 1.09 (95% CI, 0.83 to 1.44) for XPF-positive patients, and 1.39 (95% CI, 0.90 to 2.15) for XPF-negative patients (interaction P = .35). Neither ERCC1 nor XPF were prognostic: among nonsquamous patients, OS HRs for positive versus negative were ERCC1, 1.11 ( P = .32), and XPF, 1.08 ( P = .55). Conclusion Superior outcomes were observed for patients with squamous histology who received platinum therapy compared with nonplatinum chemotherapy; however, selecting chemotherapy by using commercially available ERCC1 or XPF antibodies did not confer any extra survival benefit.

Early oesophageal adenocarcinoma (OA) and pre-neoplastic dysplasia may be treated with endoscopic resection and ablative techniques such as photodynamic therapy (PDT). Though effective, discrete areas of disease may be missed leading to recurrence. PDT further suffers from the side effects of off-target photosensitivity. A tumour specific and light targeted therapeutic agent with optimised pharmacokinetics could be used to destroy residual cancerous cells left behind after resection. A small molecule antibody-photosensitizer conjugate was developed targeting human epidermal growth factor receptor 2 (HER2). This was tested in an in vivo mouse model of human OA using a xenograft flank model with clinically relevant low level HER2 expression and heterogeneity. In vitro we demonstrate selective binding of the conjugate to tumour versus normal tissue. Light dependent cytotoxicity of the phototherapy agent in vitro was observed. In an in vivo OA mouse xenograft model the phototherapy agent had desirable pharmacokinetic properties for tumour uptake and blood clearance time. PDT treatment caused tumour growth arrest in all the tumours despite the tumours having a clinically defined low/negative HER2 expression level. This new phototherapy agent shows therapeutic potential for treatment of both HER2 positive and borderline/negative OA.

International and national guidelines highlight the importance of accuracy, reproducibility, and quality control of in situ hybridization (ISH) methods for testing breast carcinomas. However, few guidelines cover the reporting of ISH cases with "unusual" signal patterns, including, eg, heterogeneity and loss of chromosome enumeration probe or gene signals. These cases are, in fact, relatively frequent, and there is a need for developing evidence- or consensus-based reporting guidelines to ensure consistency of treatment. Following an audit of cases from a single center (including >1,700 cases) we show that approximately 10% of ISH results reflect unusual signal patterns. We illustrate the most common of these patterns and provide reporting guidelines for diagnosticians and recommendations for future research. Our goal is to ensure that in the future such "rogues" are reported in a consistent manner that, ultimately, will be supported by molecular and biochemical evidence.

Expression of thyroid transcription factor-1 (TTF-1) by primary breast carcinoma and ductal carcinoma in situ (DCIS) is extremely rare and can potentially lead to diagnostic confusion. Here, we present an instructive case, including the first report of mutation analysis on such a tumour, and discuss the ways in which diagnostic pitfalls can be avoided in this situation. A 69-year-old female non-smoker with a past medical history of hypertension presented to the emergency department with cough and a right breast lump. Examination of the right breast revealed a 60 mm mass at the 12 o'clock position, with no skin tethering or fixation to deep structures. Mammography and ultrasound showed a well-defined lobulated mass measuring 30.5 mm in the upper half of the right breast, consistent with malignancy. In addition, chest radiography showed a right paramediastinal swelling of uncertain significance. Core biopsy of the right breast mass showed a grade 3 invasive ductal carcinoma with high-grade DCIS. Immunohistochemistry showed the tumour cells to be negative for oestrogen receptor (ER), progesterone receptor (PR) and human epidermal growth factor receptor 2 (HER2) (figure 1). In view of the unexplained lung lesion on radiology, TTF-1 …

These guidelines supplement existing guidelines on HER2 testing by immunohistochemistry and in-situ hybridisation(ISH) methods in the UK. They provide a specific focus on aspects of guidance relevant to HER2 ISH testing methods, both fluorescent and chromogenic. They are formulated to give advice on methodology, interpretation and quality control for ISH-based testing of HER2 status in common tumour types, including both breast and gastric tumours. The aim is to ensure that all ISH-based testing is accurate, reliable and timely.

Introduction Stomach cancer is the 3rd commonest cause of death by cancer worldwide. As many as 60% of these gastric tumours express the Human Epidermal Growth Factor receptor (HER 1–4), which play a role in cell proliferation and survival. Endoscopic submucosal dissection (ESD) is a minimally invasive surgical technique used to resect early gastric neoplasias. The purpose of this study was to establish a relationship between HER family expression in early gastric cancers, their clinical utility as a diagnostic or prognostic adjunct and how over-expression of the HER receptor affects the curative rate of ESD. Method Formalin fixed paraffin embedded tissue from 22 early gastric ESD resection specimens were sectioned. Normal gastric resections were used as a control. Immunohistochemical staining for the HER family receptors were optimised and staining performed against EGFR external (ED) and internal domain (ID), HER2 (ED and ID), HER3 (ID) and HER4 (ID). Scoring was performed by two independent observers and confirmed by diagnostic software. Results 2/22 (9%), 2/22 (9%) and 3/22 (13%) of cases were positive for EGFR ID, HER2 ID and HER2 ED respectively. 1/22 (4.5%) of cases showed co-expression of EGFR ID, HER2 ID and HER2 ED. 1 showed HER2 ED and ID co-expression. EGFR ED, HER3 and HER4 were all negative for overexpression. Over-expression of HER2 ID (p = 0.047) and HER2 ED (p = 0.0045) correlated positively with cancer aggressiveness. 10/20 (50%) cases showing no over-expression treated with ESD had reoccurrence of dysplasia, compared to 3/3 (100%) in the over-expressed group (p = 0.21). Conclusion Though small, our study showed a positive relationship between EGFR and HER2 expression, a potential target for therapeutic treatment. HER3 and HER4 expression did not correlate with any other antibody, in contrast to reported relationships. Repeats with different antibody clones would confirm the reliability of these results. The prognostic utility EGFR and HER2 co-expression to predict an aggressive early gastric cancer phenotype warrants further investigation. Although insignificant, the negative correlation between HER over-expression and ESD curative rates should be followed up with a larger cohort. Disclosure of interest None Declared.

Introduction Gastric (GC) and oesophageal adenocarcinoma’s (OA) are a major global health problem. This study analysed the relationship between demographic, clinical and pathological variables on and all-cause mortality in a cohort of GC and OA patients. Method Data was captured from 227 patients for age, sex, highest histological grade in tissue, tumour type, tumour location, TNM stage, HER2 status, R0 resection, nodal status, presence of linitis plastica, inflammation, atrophic gastritis (for GC) or Barrett’s epithelium (BE) (for EC). All patients’ received standard treatment through a multidisciplinary team and tissue specimens were examined by expert pathologists. Mortality was the primary outcome and was measured from date of diagnosis to date of death from any cause to calculate median survival (MS) of assessed variables. Kaplan-Meir curves were then compared between related variables (log-rank test, Mantel-Cox) and (log-rank test for trend). Results Using Mantel-Cox analysis, MS (months=m) was significantly worse in GC vs EA (12.5 m, n = 126; 23.8 m; n = 51; p = 0.027); diffuse vs intestinal type tumours (17.4 m, n = 69; 27.9 m, n = 97; p = 0.008); poorly vs moderately differentiated tumours (20.78 m, n = 51; 32.68 m, n = 125; p = 0.08); metastatic vs localised cancers (6.5 m, n = 28; 26.4 m, n = 64; p= Conclusion Survival was worse in tumours of gastric origin, diffuse grade, poor differentiation, higher TNM stage, and for OA, in those arising from Barrett’s epithelium. Older age was a worse prognostic indicator. This data helps understand the prognostic implications of GC and OA histology when counselling patients. Disclosure of interest None Declared.

Introduction Genetic factors play a crucial role in the predisposition to Barrett’s epithelium (BE) and progression to oesophageal adenocarcinoma (OA). Using next generation sequencing (NGS) methods on frozen tissue, TP53 mutations have been identified in a stage specific manor in BE (1). The standard method of preserving BE is formalin fixing and paraffin embedding (FFPE). NGS on tiny FFPE specimens is challenging. We examined the prevalence of TP53 genomic aberrations and p53 expression in tiny FFPE BE biopsies using an adapted NGS platform. Method 37 cases including non-dysplastic BE (NDBE; n = 7), low grade dysplasia (LGD; n = 7), high grade dysplasia (HGD; n = 9), intramucosal cancer (IMC; n = 6) and invasive OA (IOA; n = 8) were sequenced and immunostained. We used an inhouse, high throughput multiplexing method for targeted mutational screening using NGS on DNA extracted from FFPE tissue and analysed the files with our unique hidden Markov analytical method. Automated p53 IHC using clone D07 was used to correlate detected TP53 mutations with tissue expression. Results DNA extracted varied from 2–50 ng, including some highly degraded DNA material. TP53 was mutated in 40.5% (15/37) of cases; 28.6% of NDBE (2/7), 71.4% of LGD (5/7), 44.4% of HGD (4/9), 33.3% of IMC (2/6) and 25% of IOA (2/8). Interestingly, only 40% (6/15) of cases showed both TP53 mutations and p53 aberrations (over- or absence of expression), 60% (9/15) had mutations but no protein aberration, and 45% (5/11) had p53 aberration but no mutational abnormality. The data quality was excellent with all cases having a depth of coverage of at least x800 (mean coverage x1200 (Figure 1A). The level of p53 staining did not however correlate with mutation data (Figure 1B). Conclusion These results show discordance between detection of TP53 using mutation analysis and current p53 IHC methods. Furthermore, we identified patients with TP53 mutations in NDBE, which may pre-empt progression to OA. Identifying early genomic aberrations driving the development of OA from BE holds the key to accurately risk stratifying BE patients into more aggressive treatment strategies or frequent surveillance regimes. Disclosure of interest None Declared. Reference Weaver et al . Nat Genet . 2014;46(8):837-43

Introduction We have previously shown how the Allred scoring system may be used to semi-quantify expression of nuclear biomarkers in the Barrett’s (BE) to oesophageal adenocarcinoma (OA) sequence. Recently, a number of digital image analysis (DIA) platforms have been clinically validated for quantification of immunohistochemistry (IHC) in breast tissue. This study aims to compare pathologists scores (PS) with DIA for the quantification of nuclear biomarkers in BE to OA sequence. Methods Paraffin embedded specimens were selected from 34 patients. Pathology grade (PG) was scored as 1 (non-dysplastic BE; n = 5), 2 (low grade dysplasia; n = 5), 3 (high grade dysplasia; n = 11) and 4 (OA; n = 14). Sections were immunostained with antibodies PLK1-M, PLK1-L and Geminin. Intensity (I-PS) (0 to 3+) and extent (E-PS) (0; 66% = 5) of staining were scored by 2 GI pathologists, and mean PS calculated. Intensity and proportions of +ve staining were digitally quantified using Ariol® software. Analysis classifiers were trained to identify thresholds of positive (brown/DAB) and negative (blue/Hx) nuclei (Figure A) in the areas of interest. Background tissue was digitally excluded. Mean intensity (I-DIA) and mean counts for DIA positive (red) and negative (green) nuclei (Figure B) were quantified to calculate percentage positivity (E-DIA) and compared with I-PS, E-PS and total Allred score (A-PS) using Pearson correlation coefficient. Results Significant correlation was seen between E-DIA and A-PS (r = 0.76, p = 0.006; r = 0.73, p = 0.008; r = 0.94, p = 0.0004) with all biomarkers (PLK1-M; PLK1-L; Geminin). PLK1-L showed additional correlation between DIA and PS for intensity (r = 0.985, p = 0.02) and extent (r = 0.95, p = 0.90). Geminin showed additional correlations between DIA and PS for extent (r = 0.99, p = 0.0008) and PG (r = 0.97, p = 0.03). Following training, Ariol® analysis took a mean of 4mins (Range 3–5) per tissue region highlighted. Conclusion This study has demonstrated how DIA may be used to quantify expression of nuclear biomarkers. Significant correlation between DIA with Allred score was seen with all biomarkers, but only PLK1-L correlated with intensity and Geminin with PG. Background staining ignored by pathologists was found to be a confounder for DIA, particularly with PLK1-M. Nonetheless, DIA has great potential to enhance current grading and risk stratification systems for BE, and help select patients for targeted therapies dependent on biomarker expression. Disclosure of Interest None Declared

Introduction Evaluation of HER2 status is standard practise for people with inoperable locally advanced or recurrent and/or metastatic adenocarcinoma of the stomach or gastroesophageal junction (GOJ). HER2 positivity is defined as a histology score of 3+, or 2+ with HER2 gene amplification. This study analysed gastroesophageal HER2 testing in UCL-AD over the past 2.5 years. Methods All referrals for HER2 testing (n = 844) were reviewed. Data was captured for age (n = 367), sex (n = 320), tumour origin (n = 104), referring lab (n = 768), sample adequacy, HER2 grading (0–3+) and tumour type. Sections were immunostained with the Roche Tissue Diagnostics Pathway HER2 assay. For cases scoring 2+, F/DDISH testing was performed and the turn-around-time (TAT) from receipt to result analysed. Results Cases were received from 61 labs, mean age 63yo (26–93yo, SD = 13) and the majority were men (71.5%). Specimens originated from oesophageal (OE) (33.7%), GOJ (13.5%), stomach (48%) and UGI metastases (4.8%). The majority (98.7%, n = 833) were HER2-graded. The tumour types were intestinal (INT) (49.6%, n = 257), diffuse (D) (29%, n = 150), mixed D/INT (14.3%, n = 74), carcinoma-in-situ (HGD/IMC) (6.4%, n = 33), squamous (Sq) and adenosquamous (ASq) (0.4%, n = 2 each). HER2 scores were 0 (44%, n = 366), 1+ (14.5%, n = 121), 2+ (18.8%, n = 156) and 3+ (22.7%, n = 189).Gene amplification of 2+ cases identified 124 (79.5%) negative, 23 (14.7%) positive and 9 (1.1%) unrecordable results. HER2 positivity (3+/2+&F/DDISH+) was noted in 10% (15/150) of D, 30.4% (78/257) of INT, 13.5% (10/74) of mixed D/INT, 54.5% (18/33) of HGD/IMC and in 50% (1/2) of Sq tumours. A TAT of Conclusion Overall HER2-positivity rate was 25.8% in a cohort twice the size of the UK cohort reported in the benchmark ToGA trial. When compared with ToGA, subgroup analysis showed comparable but higher rates in D cancers (10% vs 6.1%), lower in both INT/mixed cancers (30.4/13.5% vs 32.2/20.4%), showed opposite proportions of GOJ/stomach cancers (28.6/31.3% vs 33.1/20.9%), and grouping proximal (OE+GOJ) cancers together, a higher ratio compared to stomach cancers was seen (0.51 vs 0.33). Positivity in Sq cancer is of uncertain significance and warrants further research into new potential applications for HER2 targeting. Disclosure of Interest None Declared