The Information Technology (IT) Infrastructure of the Multicenter Archipelago of Ovarian Cancer Research Biobank: A Potential Blueprint for Other Biobanks.

Objective: Biobanks play a crucial role in fundamental and translational research by storing valuable biomaterials and data for future analyses. However, the design of their information technology (IT) infrastructures is often customized to specific requirements, thereby lacking the ability to be used for biobanks comprising other (types of) diseases. This results in substantial costs, time, and efforts for each new biobank project. The Dutch multicenter Archipelago of Ovarian Cancer Research (AOCR) biobank has developed an innovative, reusable IT infrastructure capable of adaptation to various biobanks, thereby enabling cost-effective and efficient implementation and management of biobank IT systems. Methods and Results: The AOCR IT infrastructure incorporates preexisting biobank software, mainly managed by Health-RI. The web-based registration tool Ldot is used for secure storage and pseudonymization of patient data. Clinicopathological data are retrieved from the Netherlands Cancer Registry and the Dutch nationwide pathology databank (Palga), both established repositories, reducing administrative workload and ensuring high data quality. Metadata of collected biomaterials are stored in the OpenSpecimen system. For digital pathology research, a hematoxylin and eosin-stained slide from each patient's tumor is digitized and uploaded to Slide Score. Furthermore, adhering to the Findable, Accessible, Interoperable, and Reusable (FAIR) principles, genomic data derived from the AOCR samples are stored in cBioPortal. Conclusion: The IT infrastructure of the AOCR biobank represents a new standard for biobanks, offering flexibility to handle diverse diseases and types of biomaterials. This infrastructure bypasses the need for disease-specific, custom-built software, thereby being cost- and time-effective while ensuring data quality and legislative compliance. The adaptability of this infrastructure highlights its potential to serve as a blueprint for the development of IT infrastructures in both new and existing biobanks.

Establishment of the Dutch Nationwide, Interdisciplinary Infrastructure and Biobank for Fundamental and Translational Ovarian Cancer Research: Archipelago of Ovarian Cancer Research.

OBJECTIVES: Ovarian cancer has the worst overall survival rate of all gynecologic malignancies. For the majority of patients, the 5-year overall survival rate of less than 50% has hardly improved over the last decades. To improve the outcome of patients with all subtypes of ovarian cancer, large-scale fundamental and translational research is needed. To accommodate these types of ovarian cancer research, we have established a Dutch nationwide, interdisciplinary infrastructure and biobank: the Archipelago of Ovarian Cancer Research (AOCR). The AOCR will facilitate fundamental and translational ovarian cancer research and enhance interdisciplinary, national, and international collaboration. DESIGN: The AOCR biobank is a prospective ovarian cancer biobank in which biomaterials are collected, processed, and stored in a uniform matter for future (genetic) scientific research. All 19 Dutch hospitals in which ovarian cancer surgery is performed participate and collaborate in the AOCR biobank. PARTICIPANTS/MATERIALS, SETTING, METHODS: Patients of 16 years and older with suspected or diagnosed ovarian, fallopian tube, or primary peritoneal cancer are recruited for participation. Patients who agree to participate give written informed consent for collection, storage, and issue of their biomaterials for future studies. After inclusion, different blood samples are taken at various predefined time points both before and during treatment. In case of a diagnostic paracentesis or biopsy, the residual biomaterials of these procedures are stored in the biobank. During surgery, primary tumor tissue and, if applicable, tissue from metastatic sites are collected and stored. From each patient, a representative histological hematoxylin and eosin stained slide is digitalized for research purposes, including reassessment by a panel of gynecologic pathologists. Clinical and pathological data are obtained on a per-study basis from Dutch registries. Research proposals for the issue of biomaterials and data are evaluated by both the Archipelago Scientific Committee and the Steering Committee. Researchers using the biomaterials from the AOCR biobank are encouraged to enrich the biobank with data and materials resulting from their analyses and experiments. LIMITATIONS: The implementation and first 4 years of collection are financed by an infrastructural grant from the Dutch Cancer Society. Therefore, the main limitation is that the costs for sustaining the biobank after the funding period will have to be covered. This coverage will come from incorporation of budget for biobanking in future grant applications and from fees from external researchers and commercial parties using the biomaterials stored in the AOCR biobank. Moreover, we will apply for grants aimed at sustaining and improving research infrastructures and biobanks. CONCLUSIONS: With the establishment of the Dutch nationwide, interdisciplinary Archipelago of Ovarian Cancer Research infrastructure and biobank, fundamental and translational research on ovarian cancer can be greatly improved. The ultimate aim of this infrastructure is that it will lead to improved diagnostics, treatment, and survival of patients with ovarian cancer.

Road to FAIR genomes: a gap analysis of NGS data generation and sharing in the Netherlands.

Objective: This study investigates current standards and operational gaps in the management and sharing of next generation sequencing (NGS) data within the healthcare and research setting and according to Findable, Accessible, Interoperable and Reusable (FAIR) principles. Methods: The analysis was performed as the basis from which to bridge identified gaps and develop widely accepted working standards that ensure optimal reusability of genomic data in healthcare and research settings in the Netherlands. This work is part of the 'Rational Pharmacotherapy Program' led by ZonMw, The Netherlands Organisation for Health Research and Development, which aims to promote the efficient implementation of NGS and personalised medicine within Dutch healthcare, with an initial focus on oncology and rare diseases. Results: Based on this analysis and as part of this programme, a consortium was formed to develop an instruction manual for FAIR genomic data in clinical care and research based on an inventory of commonly used workflows and standards in the (inter)national field of genome analysis. Conclusions: The gap analysis presented and discussed in this paper represents the starting point for this inventory and is a possible contribution from the Netherlands to the European 1+ Million Genomes Initiative. This paper addresses the topics of data generation, data quality, (meta)data standards, data storage and archiving and data integration and exchange.

Spatial concordance of DNA methylation classification in diffuse glioma.

BACKGROUND: Intratumoral heterogeneity is a hallmark of diffuse gliomas. DNA methylation profiling is an emerging approach in the clinical classification of brain tumors. The goal of this study is to investigate the effects of intratumoral heterogeneity on classification confidence. METHODS: We used neuronavigation to acquire 133 image-guided and spatially separated stereotactic biopsy samples from 16 adult patients with a diffuse glioma (7 IDH-wildtype and 2 IDH-mutant glioblastoma, 6 diffuse astrocytoma, IDH-mutant and 1 oligodendroglioma, IDH-mutant and 1p19q codeleted), which we characterized using DNA methylation arrays. Samples were obtained from regions with and without abnormalities on contrast-enhanced T1-weighted and fluid-attenuated inversion recovery MRI. Methylation profiles were analyzed to devise a 3-dimensional reconstruction of (epi)genetic heterogeneity. Tumor purity was assessed from clonal methylation sites. RESULTS: Molecular aberrations indicated that tumor was found outside imaging abnormalities, underlining the infiltrative nature of this tumor and the limitations of current routine imaging modalities. We demonstrate that tumor purity is highly variable between samples and explains a substantial part of apparent epigenetic spatial heterogeneity. We observed that DNA methylation subtypes are often, but not always, conserved in space taking tumor purity and prediction accuracy into account. CONCLUSION: Our results underscore the infiltrative nature of diffuse gliomas and suggest that DNA methylation subtypes are relatively concordant in this tumor type, although some heterogeneity exists.

Prognostic value of microvessel density in stage II and III colon cancer patients: a retrospective cohort study.

BACKGROUND: Microvessel density (MVD), as a derived marker for angiogenesis, has been associated with poor outcome in several types of cancer. This study aimed to evaluate the prognostic value of MVD in stage II and III colon cancer and its relation to tumour-stroma-percentage (TSP) and expression of HIF1A and VEGFA. METHODS: Formalin-fixed paraffin-embedded (FFPE) colon cancer tissues were collected from 53 stage II and 54 (5-fluorouracil-treated) stage III patients. MVD was scored by digital morphometric analysis of CD31-stained whole tumour sections. TSP was scored using haematoxylin-eosin stained slides. Protein expression of HIF1A and VEGFA was determined by immunohistochemical evaluation of tissue microarrays. RESULTS: Median MVD was higher in stage III compared to stage II colon cancers (11.1% versus 5.6% CD31-positive tissue area, p < 0.001). High MVD in stage II patients tended to be associated with poor disease free survival (DFS) in univariate analysis (p = 0.056). In contrast, high MVD in 5FU-treated stage III patients was associated with better DFS (p = 0.006). Prognostic value for MVD was observed in multivariate analyses for both cancer stages. CONCLUSIONS: MVD is an independent prognostic factor associated with poor DFS in stage II colon cancer patients, and with better DFS in stage III colon cancer patients treated with adjuvant chemotherapy.

StemBell therapy stabilizes atherosclerotic plaques after myocardial infarction.

BACKGROUND AIMS: After a myocardial infarction (MI) atherosclerosis is accelerated leading to destabilization of the atherosclerotic plaque. mesenchymal stromal cells are a promising therapeutic option for atherosclerosis. Previously, we demonstrated a novel stem cell delivery technique, with adipose stem cells coupled to microbubbles (i.e., StemBells) as therapy after MI. In this study, we aim to investigate the effect of StemBell therapy on atherosclerotic plaques in an atherosclerotic mouse model after MI. METHODS: MI was induced in atherosclerotic Apolipoprotein E-deficient mice that were fed a high-fat Western diet. Six days post-MI, the mice received either 5 × 105/100 µL StemBells or vehicle intravenously. The effects of StemBell treatment on the size and stability of aortic root atherosclerotic plaques and the infarcted heart were determined 28 days post-MI via (immuno)histological analyses. Moreover, monocyte subtypes and lipids in the blood were studied. RESULTS: StemBell treatment resulted in significantly increased cap thickness, decreased intra-plaque macrophage density and increased percentage of intra-plaque anti-inflammatory macrophages and chemokines, without affecting plaque size and serum cholesterol/triglycerides. Furthermore, StemBell treatment significantly increased the percentage of anti-inflammatory macrophages within the infarcted myocardium but did not affect cardiac function nor infarct size. Finally, also the average percentage of anti-inflammatory monocytes in the circulation was increased after StemBell therapy. DISCUSSION: StemBell therapy increased cap thickness and decreased intra-plaque inflammation after MI, indicative of stabilized atherosclerotic plaque. It also induced a shift of circulating monocytes and intra-plaque and intra-cardiac macrophages towards anti-inflammatory phenotypes. Hence, StemBell therapy may be a therapeutic option to prevent atherosclerosis acceleration after MI.

Lumican and versican protein expression are associated with colorectal adenoma-to-carcinoma progression.

BACKGROUND: One prominent event associated with colorectal adenoma-to-carcinoma progression is genomic instability. Approximately 85% of colorectal cancer cases exhibit chromosomal instability characterized by accumulation of chromosome copy number aberrations (CNAs). Adenomas with gain of chromosome 8q, 13q, and/or 20q are at high risk of progression to cancer. Tumor progression is also associated with expansion of the extracellular matrix (ECM) and the activation of non-malignant cells within the tumor stroma. The glycoproteins versican and lumican are overexpressed at the mRNA level in colon carcinomas compared to adenomas, and are associated with the formation of tumor stroma. PURPOSE: The aim of this study was to characterize versican and lumican protein expression in tumor progression and investigate their association with CNAs commonly associated with adenoma-to-carcinoma progression. METHODS: Tissue microarrays were constructed with colon adenomas and carcinomas that were characterized for MSI-status and DNA copy number gains of chromosomes 8q, 13q and 20q. Sections were immunohistochemically stained for lumican and versican. Protein expression levels were evaluated using digitized slides, and scores were finally dichotomized into a positive or negative score per sample. RESULTS: Lumican and versican expression were both observed in neoplastic cells and in the tumor stroma of colon adenomas and carcinomas. Lumican expression was more frequently present in epithelial cells of carcinomas than adenomas (49% versus 18%; P = 0.0001) and in high-risk adenomas and carcinomas combined compared to low-risk adenomas (43% versus 16%; P = 0.005). Versican staining in the tumor stroma was more often present in high-risk adenomas combined with carcinomas compared to low-risk adenomas (57% versus 36%; P = 0.03) and was associated with the presence of gain of 13q (71% versus 44%; P = 0.04). CONCLUSION: Epithelial lumican and stromal versican protein expression are increased during colorectal adenoma-to-carcinoma progression.

Glucose Transporter 1 (SLC2A1) and Vascular Endothelial Growth Factor A (VEGFA) Predict Survival After Resection of Colorectal Cancer Liver Metastasis.

OBJECTIVE: To investigate the individual and combined prognostic value of HIF1α, SLC2A1, and vascular endothelial growth factor A (VEGFA) in a multi-institutional cohort of patients with resected colorectal cancer liver metastasis (CRCLM). BACKGROUND: In the majority of patients with CRCLM, resection seems not to be curative, despite its curative intent. Overexpression of hypoxia-inducible factor 1α (HIF1α), glucose transporter 1 (SLC2A1; also known as GLUT1), and VEGFA has been associated with tumor progression and poor prognosis of patients with colorectal cancer (CRC). METHODS: Tissue microarrays were generated using CRCLM and patient-matched primary CRC from patients who underwent CRCLM resection between 1990 and 2010. Prognostic value of HIF1α, SLC2A1, and VEGFA was determined by immunohistochemistry. A 500-fold cross-validated hazard rate ratio (HRRav) for overall survival was calculated. RESULTS: HIF1α, SLC2A1, and VEGFA expression could be evaluated in 328, 350, and 335 patients, respectively. High SLC2A1 expression was associated with good prognosis (HRRav, 0.67; P (HRR >1)  < 0.01) and high VEGFA expression to poor prognosis (HRRav, 1.84; P (HRR < 1)  = 0.02), also after multivariate analysis including established clinicopathological prognostic variables (HRRav, 0.67; P (HRR > 1)  < 0.01 and HRRav, 1.50; P (HRR < 1)  = 0.02, respectively). SLC2A1 showed prognostic value particularly in patients treated with systemic therapy (P < 0.01), whereas the prognostic value of VEGFA expression was mainly observed in patients not treated with systemic therapy (P < 0.01). Prognosis was especially poor in patients with both low SLC2A1 and high VEGFA expression (P < 0.01). HIF1α expression was not associated with survival. CONCLUSIONS: SLC2A1 and VEGFA expression are prognostic molecular biomarkers for patients with CRCLM with added value to established clinicopathological variables.

Extent and Location of Tumor-Infiltrating Lymphocytes in Microsatellite-Stable Colon Cancer Predict Outcome to Adjuvant Active Specific Immunotherapy.

PURPOSE: To determine the prognostic and predictive value of tumor-infiltrating lymphocytes (TIL) in colon cancer in a cohort of patients who previously took part in a trial on adjuvant active specific immunotherapy (ASI). EXPERIMENTAL DESIGN: We determined the number and location of CD3 and CD8 positive T cells in archival tumor samples of 106 colon cancers. We correlated stromal and epithelial TIL numbers with tumor stage and treatment and determined the effects on disease-specific survival (DSS) and recurrence-free interval (RFI). RESULTS: On the basis of the data presented, we concluded that (i) high numbers of stromal CD3 T cells have positive prognostic value measured as DSS for patients with stage II microsatellite-stable tumors and (ii) high numbers of epithelial CD8-positive T cells have positive prognostic value measured as RFI for the group of patients with stage II microsatellite-stable tumors as well as for the whole group (so stage II plus stage III together). Furthermore, we concluded that high numbers of pre-existing stromal CD3-positive T cells are of positive predictive value in adjuvant ASI treatment measured as DSS as well as RFI. CONCLUSIONS: ASI therapy may contribute to an improved DSS and RFI in patients with microsatellite-stable colon tumors harboring high numbers of pre-existing stromal CD3(+) TIL. Validation in future clinical trials is awaited.

A prognostic classifier for patients with colorectal cancer liver metastasis, based on AURKA, PTGS2 and MMP9.

BACKGROUND: Prognosis of patients with colorectal cancer liver metastasis (CRCLM) is estimated based on clinicopathological models. Stratifying patients based on tumor biology may have additional value. METHODS: Tissue micro-arrays (TMAs), containing resected CRCLM and corresponding primary tumors from a multi-institutional cohort of 507 patients, were immunohistochemically stained for 18 candidate biomarkers. Cross-validated hazard rate ratios (HRRs) for overall survival (OS) and the proportion of HRRs with opposite effect (P(HRR < 1) or P(HRR > 1)) were calculated. A classifier was constructed by classification and regression tree (CART) analysis and its prognostic value determined by permutation analysis. Correlations between protein expression in primary tumor-CRCLM pairs were calculated. RESULTS: Based on their putative prognostic value, EGFR (P(HRR < 1) = .02), AURKA (P(HRR < 1) = .02), VEGFA (P(HRR < 1) = .02), PTGS2 (P(HRR < 1) = .01), SLC2A1 (P(HRR > 1) < 01), HIF1α (P(HRR > 1) = .06), KCNQ1 (P(HRR > 1) = .09), CEA (P (HRR > 1) = .05) and MMP9 (P(HRR < 1) = .07) were included in the CART analysis (n = 201). The resulting classifier was based on AURKA, PTGS2 and MMP9 expression and was associated with OS (HRR 2.79, p < .001), also after multivariate analysis (HRR 3.57, p < .001). The prognostic value of the biomarker-based classifier was superior to the clinicopathological model (p = .001). Prognostic value was highest for colon cancer patients (HRR 5.71, p < .001) and patients not treated with systemic therapy (HRR 3.48, p < .01). Classification based on protein expression in primary tumors could be based on AURKA expression only (HRR 2.59, p = .04). CONCLUSION: A classifier was generated for patients with CRCLM with improved prognostic value compared to the standard clinicopathological prognostic parameters, which may aid selection of patients who may benefit from adjuvant systemic therapy.

Expression of apoptosis regulating proteins identifies stage II and III colon cancer patients with high risk of recurrence.

BACKGROUND AND OBJECTIVES: Deregulation of apoptosis related genes may be associated with poor outcome in cancer. Aim of the present study was to investigate the prognostic role of expression levels of apoptosis related proteins in stage II and III colon cancer. METHODS: From tumor samples of 386 stage II and III colon cancer patients, DNA was isolated and tissue microarrays were constructed. Expression of Bcl-2, Bcl-X, BAX, XIAP, Fas, FasL and c-FLIP was evaluated and PCR-based microsatellite instability analysis was performed. RESULTS: High FasL expressing tumors were associated with high disease recurrence rates in stage II colon cancer patients overall, as was low Bcl-X expression in microsatellite stable stage II patients. In stage II patients, a multivariable model based on FasL and Bcl-XL expression revealed a significant association with disease free survival (DFS). In stage III colon cancer patients, low Bcl-2, low BAX and low Fas expression levels were associated with worse outcome. In these patients a multivariable model based on angioinvasion and Bcl-2, Fas and FasL expression was significantly associated with DFS. CONCLUSIONS: Stage II patients with low Bcl-X and high FasL protein expression levels and stage III patients with low Fas, high FasL and low Bcl-2 expression could be considered as high risk for disease recurrence.

Changes in tissue composition of the vaginal wall of premenopausal women with prolapse.

OBJECTIVE: The objective of this study was to compare histological and biochemical features of the (normal) precervical anterior vaginal wall and the prolapsed anterior vaginal wall of women with pelvic organ prolapse (POP). These data were compared to tissue of the precervical anterior vaginal wall of age-matched controls without POP to identify possible intrinsic and acquired effects. STUDY DESIGN: Biopsies were collected from the apex of the anterior vaginal cuff after hysterectomy from a control group of 13 premenopausal women undergoing hysterectomy for benign gynecological diseases, and a case group of 13 premenopausal women undergoing prolapse surgery (cystocele POP-Quantification stage ≥2). In women with POP an additional full-thickness vaginal wall sample was taken from the POP site during anterior colporrhaphy. Histomorphometric and biochemical analysis were performed for different components of the extracellular matrix. RESULTS: There were no differences between case and control group in the precervical vaginal wall tissue with respect to the different components of the extracellular matrix and the biochemical parameters. However, there was a tendency toward a higher amount of collagen III and elastin, and a significant increase of smooth muscle cells and pyridinoline collagen cross-links in the POP site compared to the non-POP site of the same POP patient. CONCLUSION: Our findings suggest that the changes seen in connective tissue in the anterior vaginal wall of women with POP are the effect, rather than the cause, of POP.

In compressed lung tissue microscopic sections of adenocarcinoma in situ may mimic papillary adenocarcinoma.

CONTEXT: Surgical removal and pathologic handling of lung tissue has a compressive effect upon its architecture. The effect of surgical atelectasis on morphology has not been examined in depth, especially with respect to lung adenocarcinomas. OBJECTIVE: To examine the influence of surgical atelectasis on morphologic lepidic growth pattern, mimicking papillary adenocarcinoma pattern. DESIGN: In 2 cases serial sections of resected pulmonary adenocarcinoma were used, as was a 3-dimensional reconstruction. Elastin stains were performed on primary and metastatic adenocarcinomas. RESULTS: Perfusion fixation of another case showed marked morphologic differences of less compressed peripheral lung tissue, emphasizing the preexisting alveolar structure. An elastic stain may help identify true lesional architecture. CONCLUSIONS: We demonstrate that microscopic sections of adenocarcinoma in situ in compressed/collapsed tissue may give rise to a pseudopapillary pattern mimicking invasive adenocarcinoma. Accurate appreciation of different tumor architecture in lung adenocarcinoma has important biologic and clinical implications. Pathologists should be aware of the possibility of misclassification of adenocarcinoma pattern due to tissue artifacts caused by lung tissue handling.

Lumican and versican are associated with good outcome in stage II and III colon cancer.

BACKGROUND: Tumor stroma plays an important role in the progression and metastasis of colon cancer. The glycoproteins versican and lumican are overexpressed in colon carcinomas and are associated with the formation of tumor stroma. The aim of the present study was to investigate the potential prognostic value of versican and lumican expression in the epithelial and stromal compartment of Union for International Cancer Control (UICC) stage II and III colon cancer. METHODS: Clinicopathological data and tissue samples were collected from stage II (n = 226) and stage III (n = 160) colon cancer patients. Tissue microarrays were constructed with cores taken from both the center and the periphery of the tumor. These were immunohistochemically stained for lumican and versican. Expression levels were scored on digitized slides. Statistical evaluation was performed. RESULTS: Versican expression by epithelial cells in the periphery of the tumor, i.e., near the invasive front, was correlated to a longer disease-free survival for the whole cohort (P = 0.01), stage III patients only (P = 0.01), stage III patients with microsatellite-instable tumors (P = 0.04), and stage III patients with microsatellite-stable tumors who did not receive adjuvant chemotherapy (P = 0.006). Lumican expression in epithelial cells overall in the tumor was correlated to a longer disease-specific survival in stage II patients (P = 0.05) and to a longer disease-free survival and disease-specific survival in microsatellite-stable stage II patients (P = 0.02 and P = 0.004). CONCLUSIONS: Protein expression of versican and lumican predicted good clinical outcome for stage III and II colon cancer patients, respectively.

Targeting JNK-interacting-protein-1 (JIP1) sensitises osteosarcoma to doxorubicin.

Osteosarcoma (OS) is the most common primary malignant bone tumour in children and adolescents. Despite aggressive therapy, survival outcomes remain unsatisfactory, especially for patients with metastatic disease or patients with a poor chemotherapy response. Chemoresistance contributes to treatment failure. To increase the efficacy of conventional chemotherapy, essential survival pathways should be targeted concomitantly. Here, we performed a loss-of-function siRNA screen of the human kinome in SaOS-2 cells to identify critical survival kinases after doxorubicin treatment. Gene silencing of JNK-interacting-protein-1 (JIP1) elicited the most potent sensitisation to doxorubicin. This candidate was further explored as potential target for chemosensitisation in OS. A panel of OS cell lines and human primary osteoblasts was examined for sensitisation to doxorubicin using small molecule JIP1-inhibitor BI-78D3. JIP1 expression and JIP1-inhibitor effects on JNK-signalling were investigated by Western blot analysis. JIP1 expression in human OS tumours was assessed by immunohistochemistry on tissue micro arrays. BI-78D3 blocked JNK-signalling and sensitised three out of four tested OS cell lines, but not healthy osteoblasts, to treatment with doxorubicin. Combination treatment increased the induction of apoptosis. JIP1 was found to be expressed in two-thirds of human primary OS tissue samples. Patients with JIP1 positive tumours showed a trend to inferior overall survival. Collectively, JIP1 appears a clinically relevant novel target in OS to enhance the efficacy of doxorubicin treatment by means of RNA interference or pharmacological inhibition.

CSE1L, DIDO1 and RBM39 in colorectal adenoma to carcinoma progression.

BACKGROUND: Gain of chromosome 20q is an important factor in the progression from colorectal adenomas to carcinomas. Genes that drive 20q gain are expected to show correlation of mRNA and protein expression levels with 20q DNA copy number status while functionally influencing cancer processes. CSE1L, DIDO1 and RBM39 are located on the 20q amplicon and affect processes such as cell viability and anchorage-independent growth in colorectal cancer. This study aimed to investigate whether CSE1L, DIDO1 and RBM39 may drive 20q amplification. METHODS: Protein expression levels were examined by immunohistochemical evaluation of tissue microarrays containing a series of colorectal adenoma and carcinoma samples, which were characterized by genome-wide (microarray-based) DNA and mRNA profiling. RESULTS: CSE1L, DIDO1 and RBM39 mRNA expression levels correlated with chromosome 20q DNA copy number status. CSE1L protein expression was not associated with 20q gain, although its expression was increased in carcinomas compared to adenomas. DIDO1 and RBM39 protein expression was quite strong in the majority of tumors irrespective of 20q DNA copy number status. CONCLUSION: The lack of correlation between protein expression levels and 20q DNA copy number status implies that CSE1L, DIDO1 and RBM39 are merely passengers rather than drivers of chromosome 20q gain in colorectal adenoma-to-carcinoma progression.

Cell cycle proteins predict recurrence in stage II and III colon cancer.

PURPOSE: To investigate the prognostic value of multiple cell cycle-associated proteins in a large series of stage II and III colon cancers. METHODS: From formalin-fixed, paraffin-embedded tumor samples of 386 patients with stage II and III colon cancer, DNA was isolated and tissue microarrays were constructed. Tissue microarray slides were immunohistochemically stained for p21, p27, p53, epidermal growth factor receptor, Her2/Neu, ß-catenin, cyclin D1, Ki-67, thymidylate synthase, and Aurora kinase A (AURKA). Polymerase chain reaction-based microsatellite instability analysis was performed to allow for stratification of protein expression by microsatellite instability status. RESULTS: Overall, low p21, high p53, low cyclin D1, and high AURKA expression were significantly associated with recurrence (P = 0.01, P < 0.01, P = 0.04, and P < 0.01, respectively). In stage II patients who did not receive adjuvant chemotherapy (n = 190), significantly more recurrences were observed in case of low-p21 and high-p53-expressing tumors (P < 0.01 and P = 0.03, respectively). In stage III patients who did not receive chemotherapy, high p53 expression was associated with recurrence (P = 0.02), and in patients who received chemotherapy, high AURKA expression was associated with relapse (P < 0.01). In patients with microsatellite stable tumors, high levels of p53 and AURKA were associated with recurrence (P = 0.01 and P < 0.01, respectively). Multivariate analysis showed p21 (odds ratio 1.6, 95% confidence interval 0.9-2.8) and AURKA (odds ratio 2.7, 95% confidence interval 1.3-5.6) to be independently associated with disease recurrence. CONCLUSIONS: p21, p53, cyclin D1, and AURKA could possibly be used as prognostic markers to identify colon cancer patients with high risk of disease recurrence.

Experience with high-risk human papillomavirus testing on vaginal brush-based self-samples of non-attendees of the cervical screening program.

We evaluated the effect of offering brush-based vaginal self-sampling for high-risk human papillomavirus (hrHPV) testing to non-attendees of the cervical screening program on response rate, compliance to follow-up and cervical intraepithelial neoplasia grade 2 or 3 (CIN2+/CIN3+) yield. In addition, concordance of hrHPV test results between physician-taken cervical scrapes and vaginal self-samples was determined. A total of 26,409 nonattending women were randomly assigned to receive a vaginal brush device for hrHPV testing by Hybrid Capture-2 method (i.e., self-sampling group, n = 26,145) or a reinvitation for regular cytology-based screening (i.e., recall control group, n = 264). hrHPV-positive self-sampling responders were invited for a physician-taken scrape for cytology and blinded hrHPV testing. If cytology was abnormal, women were referred for colposcopy. Response rate in the self-sampling group was significantly increased compared to the recall control group (30.8% versus 6.5%; p < 0.001). The concordance rate between hrHPV detection in self-samples and corresponding physician-taken cervical scrape samples was 68.8%. Amongst women with CIN3+ and CIN2+, the concordance rates in hrHPV positivity between both samples were 95.5% and 93.8%, respectively. Adherence at baseline to cytology triage of hrHPV-positive self-sampling women (89.1%) and colposcopy referral of those with abnormal cytology (95.8%) was high. The CIN2+/CIN3+/carcinoma yields were 1.5%, 1.0% and 0.1%, respectively, in self-sampling responders. In conclusion, offering hrHPV testing on self-sampled vaginal material with a brush device to non-attendees significantly increases the attendance to the regular screening program, yields hrHPV test results that are in very good concordance with those of physician-taken scrapes in women with CIN2+/CIN3+, and is effective in detecting CIN2+/CIN3+.

Cell surface proteomics identifies glucose transporter type 1 and prion protein as candidate biomarkers for colorectal adenoma-to-carcinoma progression.

BACKGROUND AND OBJECTIVE: Early detection of colon adenomas at high risk of progression and early-stage colorectal cancer (CRC) is an effective approach to reduce CRC death rates. Current screening methods lack specificity as they detect many adenomas that will never progress to CRC. The authors aimed to identify cell surface protein biomarkers with extracellular domains that could be targeted for molecular imaging and discriminate low-risk adenomas and normal colon from high-risk adenomas and CRC. DESIGN: Cell surface proteins of five CRC cell lines were biotinylated, isolated and analysed by in-depth proteomics using gel electrophoresis and nanoliquid chromatography coupled to tandem mass spectrometry. Differential expression in adenomas and CRCs was based on mRNA expression and verified by immunohistochemical staining of tissue microarrays. RESULTS: In total, 2609 proteins were identified in the cell surface fractions. Of these, 44 proteins were selected as promising cell surface candidate biomarkers for adenoma-to-carcinoma progression based on the following criteria: protein identification in at least four out of five cell lines, a predicted (trans)membrane location and increased mRNA expression in CRCs compared to adenomas. Increased protein expression in high-risk adenomas and CRCs compared to low-risk adenomas was confirmed by immunohistochemistry for glucose transporter type 1 (gene symbol SLC2A1; p<0.00001) and prion protein (gene symbol PRNP; p<0.005). CONCLUSION: This study revealed glucose transporter type 1, prion protein and 42 other cell surface candidate biomarkers for adenoma-to-carcinoma progression that could potentially serve as targets for emerging molecular imaging modalities like optical imaging, ¹9F-MRI and positron emission tomography.