{'fraction_non_alphanumeric': 0.040586759899673176, 'fraction_numerical': 0.02789389678498138, 'mean_word_length': 4.9809090909090905, 'pattern_counts': {'":': 0, '<': 7, '<?xml version=': 0, '>': 0, 'https://': 0, 'lorem ipsum': 0, 'www.': 0, 'xml': 0}, 'pii_count': 0, 'substrings_counts': 0, 'word_list_counts': {'cursed_substrings.json': 0, 'profanity_word_list.json': 0, 'sexual_word_list.json': 0, 'zh_pornsignals.json': 0}}
PO-498 Spliced RNA panels from tumor-educated platelets (TEP) enable detection of early breast cancer

Introduction Detection of early-stage breast cancer from blood is a highly desirable screening application of liquid biopsies. Tumor-educated platelets (TEP) contain distinct spliced RNA repertoires and can function as a blood-based biosource for the detection of cancer (Best et al. Cancer Cell 2015). Here, we investigated the potential of TEP RNA profiles as an early breast cancer diagnostics algorithm. Material and methods We sequenced platelets of women with stage I-III breast cancer (n=134) and asymptomatic non-cancer females (n=121). Samples were divided over a training, evaluation and validation set. We developed a TEP RNA-based classification algorithm, which was tailored towards two clinically relevant diagnostic applications. The first application is to rule out breast cancer in women with an abnormal mammography. The second application is to rule in breast cancer in asymptomatic women carrying an elevated baseline breast cancer risk. We examined if classifications depended on patient age, tumour stage, subtype, BRCA mutation status, and breast density. In addition, we tested if the algorithm was breast cancer specific by testing our algorithm in a pan-cancer cohort (n=192). Results and discussions In the evaluation cohort (n=48 of which n=24 early breast cancer cases) the TEP RNA-based breast cancer classification had an accuracy of 90% with an AUC of 0.96 (95% confidence interval (CI) 0.92–1.00; p<0.001). In the validation cohort, the TEP RNA-based classification algorithm reached an overall accuracy of 86% and an AUC of 0.93 (95% CI 0.89–0.98, p<0.001). Most important, the accuracy was 91% in stage I/II disease. In a confirmatory rule-out application (positive mammography) sensitivity would be 94% with a specificity of 67%; whereas in a screening setting sensitivity would be 80% with a specificity of 93%. The classifier performance is consistent across subgroups based on patient age, tumour stage, subtype, tissue density, or BRCA1/2-status. The breast cancer TEP profiles were distinct from those of women with other tumour types (n=192, accuracy: 87%, AUC: 0.91, 95% CI 0.87–0.96, p<0.001). Although results look promising, sensitivity and specificity should be improved for clinical implementation. Follow-up studies should, among others, require sample collection in actual screening and confirmatory settings. Conclusion We show that platelet RNA signatures may enable blood-based screening for early breast cancer, and warrant validation in a confirmatory and screening setting.

Introduction In epithelial ovarian cancer (EOC) patients, 15%-20% of the tumours do not respond to first-line chemotherapy treatment, i.e., paclitaxel combined with a platinum-based therapy. The aim of this pilot study was to validate a new approach to measure the sensitivity of ovarian cancer cells to chemotherapeutics. Furthermore, we aimed to relate the in vitro response of tumour cells from ascites to clinical response of ovarian cancer patients. Material and methods For this pilot study, we selected 18 patients with advanced stage EOC and collected ascites. Proliferation assays on tumour cells isolated from the ascites were performed using intracellular ATP content as an indirect measure of cell number. Clinical outcome was determined using status of remission after primary treatment, disease free survival (DFS), and the platinum-sensitivity. Complete remission was defined as a serum-CA125 <35 E/ml, a complete or optimal debulking surgery, and a complete response on CT-scan. We related the sensitivity tests and the clinical outcome using descriptive statistics. Results and discussions The outcome of the proliferation assays, expressed by GI 50 and using dose-response curves, corresponded to the clinical outcome in the patient in most cases. Of the 14 ascites samples eventually tested, seven of the in vitro responses were poor as well as the DFS of the patient. Four samples showed good in vitro response and the patient showed a good DFS as well. Only two of the samples tested did not show similarity in in vitro response and in clinical outcome of the patient. We also determined the in vitro response to second line therapies and new targeted therapies, including PARP-and bromodomain inhibitors. Conclusion This pilot study demonstrates the feasibility of using ascites tumour cells to perform drug sensitivity tests. These are promising results because it seems possible to predict accurately the response on first-line chemotherapy in advanced stages of EOC. However, larger observational studies are required to show a statistical significant relation, to determine the best clinical response measurements and to test second line therapies. Introduction PTEN is a protein that restrains cells from growing unchecked; when it is absent or not functioning (as it often happens in prostate and colorectal cancer -CRC), cancer develops more easily and grows in a more aggressive and therapy-resistant fashion. However, just measuring PTEN mutational status or gene/protein levels in clinical samples may not be sufficient to assess its function, as its regulation is complex. Therefore, we analysed miRNA, transcriptomic, and proteomic profiles of isogenic CRC models differing for PTEN status, in order to identify a set of genes or proteins that are stably modulated by the presence or absence of a functional PTEN. Material and methods To identify biomarkers of PTEN loss of function and functional downstream effectors of PTEN activity, the isogenic CRC cell lines HCT116 (PTEN +/+ ) and HCT116 PTEN -/-(Horizon Discovery Ltd) were subjected to comprehensive miRNA profiling (Human miRNA Microarray Release 21, Agilent), transcriptomic analysis (NextSeq 500, Illumina), and semi-quantitative phosphoproteomic analysis (Antibody Microarray Service, KinexTM).

PO-496
Results and discussions miRNA profiling found 18 miR-NAsdifferentially expressed in HCT116 PTEN +/+ and their HCT116 PTEN -/counterpart: only miR-196a-5p is up-regulated in HCT116 PTEN -/-, whereas other 17 miRNAs were all down-regulated; several of these target miRNAs have been validated by qRT-PCR. RNA-Seq data were analysed via the RAP pipeline available at CINECA that relies on the Tuxedo suite (Tophat-Cufflinks-Cuffdiff) for read mapping, transcript abundance estimation and transcript or gene-based differential expression. RNA-Seq analysis showed 247 differentially expressed genes in HCT116 PTEN +/+ cells as compared to HCT116 PTEN -/-(159 up-regulated and 88 down-regulated in HCT116 PTEN -/-) and clustering into 9 subgroups (including: tissue morphogenesis; negative regulation of phosphatase metabolic process; regulation of sequence-specific DNA binding transcription factor activity). qRT-PCR assay of 10 dysregulated genes was performed to validate the RNA-Seq data set. Proteomic analysis showed that 92 proteins (25 total proteins and 67 phospho-proteins) are differentially expressed in HCT116 PTEN +/+ and their HCT116 PTEN -/counterpart, some of which have been validated by WB. Conclusion Our data identify differentially expressed sets of miRNA/genes/proteins between HCT116 PTEN +/+ and HCT116 PTEN -/-. Bioinformatics analysis to cluster these sets into functional PTEN-loss 'signatures' is ongoing.
Introduction Detection of early-stage breast cancer from blood is a highly desirable screening application of liquid biopsies. Tumor-educated platelets (TEP) contain distinct spliced RNA repertoires and can function as a blood-based biosource for the detection of cancer (Best et al. Cancer Cell 2015). Here, we investigated the potential of TEP RNA profiles as an early breast cancer diagnostics algorithm. Material and methods We sequenced platelets of women with stage I-III breast cancer (n=134) and asymptomatic non-cancer females (n=121). Samples were divided over a training, evaluation and validation set. We developed a TEP RNA-based classification algorithm, which was tailored towards two clinically relevant diagnostic applications. The first application is to rule out breast cancer in women with an abnormal mammography. The second application is to rule in breast cancer in asymptomatic women carrying an elevated baseline breast cancer risk. We examined if classifications depended on patient age, tumour stage, subtype, BRCA mutation status, and breast density. In addition, we tested if the algorithm was breast cancer specific by testing our algorithm in a pan-cancer cohort (n=192). Results and discussions In the evaluation cohort (n=48 of which n=24 early breast cancer cases) the TEP RNA-based breast cancer classification had an accuracy of 90% with an AUC of 0.96 (95% confidence interval (CI) 0.92-1.00; p<0.001). In the validation cohort, the TEP RNA-based classification algorithm reached an overall accuracy of 86% and an AUC of 0.93 (95% CI 0.89-0.98, p<0.001). Most important, the accuracy was 91% in stage I/II disease. In a confirmatory rule-out application (positive mammography) sensitivity would be 94% with a specificity of 67%; whereas in a screening setting sensitivity would be 80% with a specificity of 93%. The classifier performance is consistent across subgroups based on patient age, tumour stage, subtype, tissue density, or BRCA1/2status. The breast cancer TEP profiles were distinct from those of women with other tumour types (n=192, accuracy: 87%, AUC: 0.91, 95% CI 0.87-0.96, p<0.001). Although results look promising, sensitivity and specificity should be improved for clinical implementation. Follow-up studies should, among others, require sample collection in actual screening and confirmatory settings. Conclusion We show that platelet RNA signatures may enable blood-based screening for early breast cancer, and warrant validation in a confirmatory and screening setting. Introduction miR-21 is an oncomir that is overexpressed in many types of solid tumours. According to some studies miR-21 expression is higher in breast tumours than normal breast tissue and it is significantly correlated with shorter survival of patients. Many studies have focused on miR-21-5 p and its correlation with clinic pathological features and survival of breast tumours. In some of the studies miR-21-5 p expression has been shown to be associated with ER status, PR status, HER2 status, clinical stage, lymph node, tumour grade and size. The purpose of this study was to compare the expression of miR-21-3 p and miR-21-5 p in progression of breast tumours and their association with corresponding clinic pathological data from the breast cancer patients. Material and methods Six non-neoplastic breast tissues and 144 tumours, from breast cancer patients diagnosed 1987-2003, were obtained from the Department of Pathology at Landspitali University Hospital. The expression levels of miR-21-3 p and miR-21-5 p were measured with StepOnePlus Real-Time PCR System by applying EXIQON primer sets and SYBR Green master mix using miR-16-5 p as endogenous control.
Results and discussions miR-21-5 p was expressed at significantly higher levels than miR-21-3 p in both normal (p=0.0003) and tumour tissues (p=6.378e-12). A significant difference was not observed between miR-21-5 p expression in normal breast tissue and tumours. High miR-21-5 p expression correlated significantly with HER2 expression (p=0.001) but it did not correlate with other clinical and pathological factors or survival. miR-21-3 p was expressed at significantly higher levels in tumour tissues than normal breast tissue (p=0.003). Its expression was significantly higher in tumours from patients with metastasis than without metastasis (p=0.02). Furthermore, higher levels of miR-21-3 p correlated with shorter disease free survival (p=0.03, HR=1.72, 95% CI=1.03-2.9). Conclusion Although miR-21 is one of the most studied and upregulated miRNAs in human solid tumours the comparison of miR-21-3 p expression with miR-21-5 p expression in breast tumours has not been well studied. Our results suggest that miR-21-3 p could be a better indicator of breast cancer progression than miR-21-5 p. More studies in larger breast cohorts and functional cell based assays will be needed to shed light on the potential role of miR-21-3 p in breast tumours progression.

A NOVEL ROLE FOR JUNCTIONAL ADHESION MOLECULE-A (JAM-A) IN HER2-POSITIVE GASTRO-OESOPHAGEAL CANCERS
Introduction Junctional Adhesion Molecule-A (JAM-A) is a cell-cell adhesion protein that regulates physiological adhesion in epithelial and endothelial cells. Our previous work has shown that increased JAM-A expression on breast tumour epithelial tissue associates with poor prognosis in patients with invasive breast cancer, and identified a correlation between high JAM-A expression and levels of the important oncogene human epidermal growth factor receptor-2 (HER2). An important role for HER2 in gastro-oesophageal cancer is emerging, however the specific role of JAM-A in this setting is unclear. Material and methods In a pilot study, gastric cancer patient tissue sections (n=11) were stained for JAM-A expression. A