Tissue-based Research Research Articles

Brain tumor epidemiology in Austria and the Austrian Brain Tumor Registry

Cancer registries provide incidence and mortality data on patients with cancer at the population level. Cancer registration is most often restricted to the group of malignant neoplasms, whereas information on benign and intermediate tumors is generally not available. Brain tumors however differ from other sites by 1) the large spectrum of different tumor types, and 2) the exclusive localization in proximity to eloquent areas with considerable neurological comorbidity and mortality irrespective of their biological behavior. In order to obtain a comprehensive overview of the brain tumor burden, specialized brain tumor registries, which provide information on all brain tumor types, have emerged in several countries. This thesis synopsis summarizes the Austrian experience on the establishment of such a specialized brain tumor registry: the Austrian Brain Tumor Registry (ABTR). The initial steps are reported - from consensus and commitment of the Austrian Society of Neuropathology, formation of an interdisciplinary team of experts, setup of the infrastructure including data confidentiality issues, to the sustained support of the Austrian neurooncology community and major cooperation with the Austrian National Cancer Registry. ABTR differs from other registries by its scientific setting and neuropathological background warranting strong expertise in brain tumor typing and tissue-based research. Thereby, ABTR constitutes also a virtual brain tumor biobank. By having achieved these steps, first investigational results demonstrate that ABTR provides valid and accurate population-based incidence and survival data for individual brain tumor types. The exact incidence of rare tumor entities is estimated, key diagnostic criteria of newly proposed tumor entities are refined, common standards for testing of molecular markers are advocated, and medical progress via real-life outcome analyses is assessed. The diverse scientific contributions highlight the enormous scientific potential of ABTR for continued work.

A Texture Based Pattern Recognition Approach to Distinguish Melanoma from Non-Melanoma Cells in Histopathological Tissue Microarray Sections

AimsImmunohistochemistry is a routine practice in clinical cancer diagnostics and also an established technology for tissue-based research regarding biomarker discovery efforts. Tedious manual assessment of immunohistochemically stained tissue needs to be fully automated to take full advantage of the potential for high throughput analyses enabled by tissue microarrays and digital pathology. Such automated tools also need to be reproducible for different experimental conditions and biomarker targets. In this study we present a novel supervised melanoma specific pattern recognition approach that is fully automated and quantitative.Methods and ResultsMelanoma samples were immunostained for the melanocyte specific target, Melan-A. Images representing immunostained melanoma tissue were then digitally processed to segment regions of interest, highlighting Melan-A positive and negative areas. Color deconvolution was applied to each region of interest to separate the channel containing the immunohistochemistry signal from the hematoxylin counterstaining channel. A support vector machine melanoma classification model was learned from a discovery melanoma patient cohort (n = 264) and subsequently validated on an independent cohort of melanoma patient tissue sample images (n = 157).ConclusionHere we propose a novel method that takes advantage of utilizing an immuhistochemical marker highlighting melanocytes to fully automate the learning of a general melanoma cell classification model. The presented method can be applied on any protein of interest and thus provides a tool for quantification of immunohistochemistry-based protein expression in melanoma.

Lyophilized brain tumor specimens can be used for histologic, nucleic acid, and protein analyses after 1 year of room temperature storage

Frozen tissue, a gold standard biospecimen, can yield well preserved nucleic acids and proteins after over a decade but is vulnerable to thawing and has substantial fiscal, spatial, and environmental costs. A long-term room temperature biospecimen storage alternative that preserves broad analytical utility can potentially empower tissue-based research. As there is scant data on the analytical utility of lyophilized brain tumor biospecimens, we evaluated lyophilized (freeze-dried) samples stored for 1 year at room temperature. Lyophilized tumor tissue processed into paraffin sections produced good histology. Yields of extracted DNA, RNA, and protein approximated those of frozen tissue. After 1 year, lyophilized samples yielded high molecular weight DNA that permitted copy number variation analysis, IDH 1 mutation detection, and MGMT promoter methylation PCR. A 27 % decrease in RIN scores over the 1 year suggests that RNA degradation was inhibited though incompletely. Nevertheless, RT-PCR studies on lyophilized tissue performed similarly to frozen tissue. In contrast to FFPE tissues where protein bands were absent or shifted to a lower molecular weight, lyophilized samples showed similar protein bands as frozen tissue on SDS-PAGE analysis. Lyophilized tissue performed similarly to frozen tissue for Western blots and enzyme activity assays. Immunohistochemistry of lyophilized tissue that were processed into FFPE blocks often required longer incubation times for staining than standard FFPE samples but generally provided robust antigen detection. This preliminary study suggests that lyophilization has promise for long-term room temperature storage while permitting varied tests; however, further work is required to better stabilize nucleic acids particularly RNA.

New tissue aquisition tools for molecular biology.

102 Background: Histological and biomolecular examinations are a prerequisite to understand diseases, determine optimal individualized care, and identify targets for potential novel therapies. Despite this key role of tissue-based research the act of biopsy still remains troublesome. New direct and frontal biopsy technologies are emerging with the aim to alleviate the bottleneck of inappropriate minimal invasive interventions. Recent clinical trials evaluated the use of macrobiopsies in providing the necessary tissues for molecular biology. Methods: The new instruments for minimal-invasive procedures were evaluated in different single and multicenter clinical trials to generate data on providing enough high-quality tissue for biomolecular research in comparison to diagnostic surgery for various soft tissues and bone lesions throughout the body. In addition, patient comfort and costs data were taken up in the survey. Results: Direct and frontal type macrobiopsy systems give tissue samples between 150 mg and 300 mg of highly specified parts of the diseased area in a way very similar to open surgery. In addition, biopsy procedures are increasingly more patient friendly with appropriate comfort and safety. The new macrobiopsies are less expensive, making molecular biology at reach for every oncological patient, company, and health care provider. Conclusions: Direct and frontal macrobiopsies open new avenues for future bio-banking, pharmacogenomics, biomolecular research, and personalized medicine. It is anticipated that drug discovery and clinical implementation of targeted therapies will be highly facilitated, and that clinical research time for multicenter trials will be significantly shortened.

Direct Molecular Tissue Analysis by MALDI Imaging Mass Spectrometry in the Field of Gastrointestinal Disease

*Institute of Pathology, Research Unit Analytical Pathology, Helmholtz Zentrum Munchen, German Research Center for Environmental Health, Neuherberg, Germany; ‡ § Department of Medicine II, Klinikum rechts der Isar, Technische Universitat Munchen, Munich, Germany; Department of Medicine II, Universitatsklinikum Mannheim, University of Heidelberg, Mannheim, Germany; and Institute of Pathology, Technische Universitat Munchen, Munich, Germany

MicroRNA analysis of fine-needle aspirates from pancreatic cancer.

200 Background: Tissue-based research is a major challenge in localized pancreatic cancer (PC) especially those with locally advanced unresectable or borderline resectable disease. Approximately half of the patients are diagnosed by a fine needle aspirate (FNA) /biopsy of the pancreatic primary tumor with limited tissue available for correlative assays. Micro-RNAs (miRNAs) are biomarkers with the potential to be developed for prognostic and predictive purposes. Moreover, these molecules can facilitate the development of targeted therapies in PC. Methods: miRNA expression profiling and quantitative real-time PCR (qRT-PCR) were performed from RNA (25-100 ng per patient) extracted from formalin-embedded cell blocks (FFPE) obtained from diagnostic FNA of the pancreas. Expression profiles of miRNAs were compared to those from patients undergoing pancreatic biopsy for non-malignant conditions. Results: Cell blocks from FNAs from twenty-nine and 15 patients with pancreatic adenocarcinoma and non-malignant conditions were studied, respectively. miRNA profiling demonstrated deregulation of over 228 miRNA. The top 7 were validated by qRT-PCR. The expression of let-7c, let-7f, and miR-200c were significantly reduced in most patients. Conversely, the expression levels of miR-486-5p and miR-451 were significantly elevated. Significant interindividual variations were also noted in expression of certain miRNAs. Conclusions: FFPE obtained from diagnostic FNA of pancreatic tumor can be a valuable resource to study miRNA expression profiles of multiple miRNAs and undertake qRT-PCR in patients with localized PC and further supports the use of diagnostic FNA in RNA-based translational research. miRNA expression can be developed for use as prognostic and predictive biomarkers in future clinical trials in patients with PC.

Digital Image Analysis in Pathology: Benefits and Obligation

Pathology has recently entered the era of personalized medicine. This brings new expectations for the accuracy and precision of tissue-based diagnosis, in particular, when quantification of histologic features and biomarker expression is required. While for many years traditional pathologic diagnosis has been regarded as ground truth, this concept is no longer sufficient in contemporary tissue-based biomarker research and clinical use. Another major change in pathology is brought by the advancement of virtual microscopy technology enabling digitization of microscopy slides and presenting new opportunities for digital image analysis. Computerized vision provides an immediate benefit of increased capacity (automation) and precision (reproducibility), but not necessarily the accuracy of the analysis. To achieve the benefit of accuracy, pathologists will have to assume an obligation of validation and quality assurance of the image analysis algorithms. Reference values are needed to measure and control the accuracy. Although pathologists' consensus values are commonly used to validate these tools, we argue that the ground truth can be best achieved by stereology methods, estimating the same variable as an algorithm is intended to do. Proper adoption of the new technology will require a new quantitative mentality in pathology. In order to see a complete and sharp picture of a disease, pathologists will need to learn to use both their analogue and digital eyes.

Digital image analysis in pathology: benefits and obligation.

Pathology has recently entered the era of personalized medicine. This brings new expectations for the accuracy and precision of tissue-based diagnosis, in particular, when quantification of histologic features and biomarker expression is required. While for many years traditional pathologic diagnosis has been regarded as ground truth, this concept is no longer sufficient in contemporary tissue-based biomarker research and clinical use. Another major change in pathology is brought by the advancement of virtual microscopy technology enabling digitization of microscopy slides and presenting new opportunities for digital image analysis. Computerized vision provides an immediate benefit of increased capacity (automation) and precision (reproducibility), but not necessarily the accuracy of the analysis. To achieve the benefit of accuracy, pathologists will have to assume an obligation of validation and quality assurance of the image analysis algorithms. Reference values are needed to measure and control the accuracy. Although pathologists' consensus values are commonly used to validate these tools, we argue that the ground truth can be best achieved by stereology methods, estimating the same variable as an algorithm is intended to do. Proper adoption of the new technology will require a new quantitative mentality in pathology. In order to see a complete and sharp picture of a disease, pathologists will need to learn to use both their analogue and digital eyes.

MALDI imaging mass spectrometry for direct tissue analysis: technological advancements and recent applications

Matrix assisted laser desorption/ionization (MALDI) imaging mass spectrometry (IMS) is a method that allows the investigation of the molecular content of tissues within its morphological context. Since it is able to measure the distribution of hundreds of analytes at once, while being label free, this method has great potential which has been increasingly recognized in the field of tissue-based research. In the last few years, MALDI-IMS has been successfully used for the molecular assessment of tissue samples mainly in biomedical research and also in other scientific fields. The present article will give an update on the application of MALDI-IMS in clinical and preclinical research. It will also give an overview of the multitude of technical advancements of this method in recent years. This includes developments in instrumentation, sample preparation, computational data analysis and protein identification. It will also highlight a number of emerging fields for application of MALDI-IMS like drug imaging where MALDI-IMS is used for studying the spatial distribution of drugs in tissues.

Biomarker Expression in Pelvic High-grade Serous Carcinoma

Neoadjuvant therapy has an emerging role in the treatment of high-stage ovarian carcinoma. Some ovarian carcinoma subtypes do not respond well to standard chemotherapy, making accurate subtype diagnosis before starting therapy important. This diagnosis is frequently based on omental biopsy specimens. In particular, with very small biopsies, immunostaining for diagnostic biomarkers may be needed. To assess intratumoral heterogeneity of biomarker expression in pelvic high-grade serous carcinoma, we compared the expression of a set of 10 biomarkers between ovarian and omental sites. Tissue microarrays were constructed from 123 high-grade serous carcinomas with paired ovarian and omental tumor samples. These samples were stained with biomarkers that have been used in ovarian carcinoma subtype diagnosis (WT1, TP53/p53, MUC16/CA125, CDKN2A/p16), and with biomarkers of the tumor microenvironment (CD8, CD163, SPARC, PDGFRB), cell adhesion (CDH1/E-Cadherin), and proliferation (Ki67) as well. Expression frequencies in samples from the 2 sites were compared, as was concordance at the 2 sites for individual tumors. The 2 markers of desmoplastic stromal response (PDGFRB, SPARC) were more frequently expressed in the omentum compared with the ovary (P<0.001; McNemar test). The other 8 markers did not show a significant difference in the frequency of expression between sites. Within individual cases, some markers such as Ki67 and CDKN2A showed variability, indicating that these markers are affected by intratumoral heterogeneity. The intratumoral variability for MUC16, TP53, and WT1 was modest. Commonly used diagnostic markers, such as TP53 and WT1, show little variability between ovarian and omental sites, suggesting that they can be successfully used in small biopsy specimens from extraovarian sites. In contrast, markers of host stromal response do vary between sites, suggesting a biologic difference of the microenvironment at different sites that should be taken into account when tissue-based research is carried out.

Maintaining Clinical Tissue Archives and Supporting Human Research: Challenges and Solutions

The increasing number of requests for use of clinically archived tissue in translational research poses unique challenges. Conflicts may arise between pathologists who are responsible for overseeing and preserving the tissues and investigators who need these materials for research purposes. To evaluate the status of our institution's Tissue Registry Archive and to develop updated written policies and procedures to support a new modern and robust tracking system with features of a library loan system. An observational study was performed. We found the existing process for managing loans of tissue (slides and paraffin blocks) to be insufficient for the complexity and volume of this task. After extensive customization, a new tracking system was implemented in January 2008. Analysis of the first year of the system's use (2008) showed that of the 206,330 slides and 51,416 blocks loaned out in 2008, 92% and 94%, respectively, were returned by the due date. These rates were markedly improved from those before the new system: 61% and 47%, respectively, in 2005. Material permanently "lost" in 2008 represented only 0.02% of slides and 0.05% of blocks, none of which was the only diagnostic material for the case. With expanding needs for archived tissues for clinical care and growing demands for translational research, it is essential that pathology departments at institutions with large tissue-based research endeavors have a tracking and management system in place to meet clinical, educational, and research needs, as well as legal requirements.

How much will the biobanking industry come to rely on private companies?

Biopreservation and BiobankingVol. 8, No. 4 The Experts SpeakHow Much Will the Biobanking Industry Come to Rely on Private Companies?Brian J. Clark, John M. Baust, and Glyn StaceyBrian J. ClarkSearch for more papers by this author, John M. BaustSearch for more papers by this author, and Glyn StaceySearch for more papers by this authorPublished Online:18 Dec 2010https://doi.org/10.1089/bio.2010.8415AboutSectionsView articleView Full TextPDF/EPUB ToolsPermissionsDownload CitationsTrack CitationsAdd to favorites Back To Publication ShareShare onFacebookTwitterLinked InRedditEmail View article"How Much Will the Biobanking Industry Come to Rely on Private Companies?." , 8(4), pp. 179–180"How Much Will the Biobanking Industry Come to Rely on Private Companies?." , 8(4), pp. 179–180"How Much Will the Biobanking Industry Come to Rely on Private Companies?." , 8(4), pp. 179–180"How Much Will the Biobanking Industry Come to Rely on Private Companies?." , 8(4), pp. 179–180FiguresReferencesRelatedDetailsCited ByBeyond and within public engagement: a broadened approach to engagement in biobanking27 October 2015 | New Genetics and Society, Vol. 34, No. 4Commercialization of Tissue-Based Research: Time to Move Beyond the Gift/Market Dichotomy16 December 2013 | Biopreservation and Biobanking, Vol. 11, No. 6Comprehensive Outsourcing Biobanking Facility to Serve the International Research Community30 June 2011 | Biopreservation and Biobanking, Vol. 9, No. 2 Volume 8Issue 4Dec 2010 InformationCopyright 2010, Mary Ann Liebert, Inc.To cite this article:Brian J. Clark, John M. Baust, and Glyn Stacey.How Much Will the Biobanking Industry Come to Rely on Private Companies?.Biopreservation and Biobanking.Dec 2010.179-180.http://doi.org/10.1089/bio.2010.8415Published in Volume: 8 Issue 4: December 18, 2010PDF download

Saudi views on consenting for research on medical records and leftover tissue samples

BackgroundConsenting for retrospective medical records-based research (MR) and leftover tissue-based research (TR) continues to be controversial. Our objective was to survey Saudis attending outpatient clinics at a tertiary care hospital on their personal preference and perceptions of norm and current practice in relation to consenting for MR and TR.MethodsWe surveyed 528 Saudis attending clinics at a tertiary care hospital in Saudi Arabia to explore their preferences and perceptions of norm and current practice. The respondents selected one of 7 options from each of 6 questionnaires.ResultsRespondents' mean (SD) age was 33 (11) years, 42% were males, 56% were patients, 84% had ≥ secondary school education, and 10% had previously volunteered for research. Respectively, 40% and 49% perceived that the norm is to conduct MR and TR without consent and 38% and 37% with general or proposal-specific consent; the rest objected to such research. There was significant difference in the distribution of choices according to health status (patients vs. companions) for MR (adjusted Kruskal-Wallis test P = 0.03) but not to age group, gender, education level, or previous participation in research (unadjusted P = 0.02 - 0.59). The distributions of perceptions of current practice and norm were similar (unadjusted Marginal Homogeneity test P = 0.44 for MR and P = 0.89 for TR), whereas the distributions of preferences and perceptions of norm were different (adjusted P = 0.09 for MR and P = 0.02 for TR). The distributions of perceptions of norm, preferences, and perceptions of current practice for MR were significantly different from those of TR (adjusted P < 0.009 for all).ConclusionsWe conclude that: 1) there is a considerable diversity among Saudi views regarding consenting for retrospective research which may be related to health status, 2) the distribution of perceptions of norm was similar to the distribution of perceptions of current practice but different from that of preferences, and 3) MR and TR are perceived differently in regard to consenting.

Pathology: coming in from the cold

Following the UK's organ retention scandals that occurred a decade ago, politicians unleashed a deluge of well-intentioned but naïve and unnecessarily burdensome regulations that have progressively stymied human tissue-based research, stifling development of improved diagnostic and prognostic tests as well as discovery of new treatments based on sound knowledge of human-specific biology. For the UK to maintain a leading role in medical research, more sensible levels of regulation need to be introduced that recognise differences between tissue from donors who have passed-away and surgical tissue that is surplus to diagnostic requirements and that otherwise will be incinerated. While it is important to reassure the public that research using their tissues will be conducted within an approved ethical framework, it is equally important to ensure that, as hospital staff and academic researchers, we are able to fulfil our unwritten covenant with patients to do our utmost to seek better diagnostic assays and more predictive prognostic indicators, while collaborating with our colleagues in academia and industry and hence bring hope to patients with illnesses for which no effective treatments are yet available. There is a clear case for introducing an opt-out system as the default to allow all surplus surgical tissues to be immediately available for research, concomitant with an education campaign. This is the optimal and most ethical approach to ensure the wishes of the vast majority of patients are respected by allowing their residual surgical tissues and relevant clinical information to be made available for research without the current levels of obstruction and hindrance.

Efficacy and safety of new medicines: a human focus

The introduction of safe and effective new medicines is proving ever more difficult, a problem arguably due at least in part to over-reliance on experimental animal-based test systems. In light of the increasing awareness of the lack of predictiveness of such non-human approaches, the necessity to focus on human-based test methods is clear. There has been considerable progress in human in vivo (microdosing) and in silico approaches, primarily to identify ADMET issues, however, in vitro functional studies using human tissues are receiving inadequate attention. The potential scope of human tissue-based research is considerable, but much methodological development is required, which necessitates an increased willingness on the part of the Pharma industry to support it. This approach also requires considerably improved access to the cells and tissues themselves. While current acquisition is almost exclusively from surgery and post mortem, the range of tissue types, the quantity, quality and frequency of supply will remain inadequate to support human tissue as a key component of pre-clinical efficacy and safety testing. Additional routine access to non-transplantable tissues from organ donors for research purposes would be of inestimable value, but in order to realise this, true collaboration will be required between NHS, the Pharma and biotech industries, and the general public.

Research involving storage and use of human tissue: how did the Human Tissue Act 2004 affect decisions by research ethics committees?

Background:Little is known about the types of issues research ethics committees (RECs) raise in their letters about research involving the storage and use of human tissue.Aims:To classify the issues that...

Tissue biomarkers in renal cell carcinoma: Issues and solutions

Renal cell carcinoma (RCC) is an aggressive malignancy that is associated with a high rate of metastasis. Although several promising therapeutic strategies are now available for the treatment of patients with metastatic kidney cancer, the prognosis of these patients remains poor. Research is ongoing to identify RCC-specific biomarkers that can improve early diagnosis, surveillance of tumor progression, and prediction of patient prognosis. The identification of biomarkers that may predict response to specific therapies also will be useful in stratifying patients with RCC for treatment selection. Unfortunately, biomarker detection and measurement in kidney tumor tissues can be biased significantly by the lack of standardization in tissue sample acquisition, storage, and analysis. Consequently, the establishment of standardized operating procedures is necessary to maximize the accuracy of tissue-based biomarker assays. Herein, the authors discuss current issues in tissue-based translational research aimed at identifying clinically useful biomarkers for kidney cancer.

Do we see what we think we see? The complexities of morphological assessment

Reliable pathological interpretation is vital to so many aspects of tissue-based research as well as being central to patient care. Understanding the complex processes involved in decision-making is the starting point to improve both diagnostic reproducibility and the definition of diagnostic groups that underpin our experiments. Unfortunately, there is a paucity of research in this field and it is encouraging to see The Journal of Pathology publishing work in this area. This review attempts to highlight the opportunities that exist in this field and the technologies that are now available to support this type of research. Key amongst these are the use of decision analysis tools such as inference networks, and virtual microscopy that allows us to simulate diagnostic decision-making. These tools have roles, not only in studying the subtleties of diagnostic decision-making, but also in delivering new methods of training and proficiency testing. Research which helps us to better understand what we see, why we see it, and standardizing interpretative reasoning in pathological classification is essential for improving the wide range of activities that pathologists support, including clinical diagnosis, teaching, training, and experimental research.

The Human Protein Atlas—a tool for pathology

Tissue-based diagnostics and research is incessantly evolving with the development of new molecular tools. It has long been realized that immunohistochemistry can add an important new level of information on top of morphology and that protein expression patterns in a cancer may yield crucial diagnostic and prognostic information. We have generated an immunohistochemistry-based map of protein expression profiles in normal tissues, cancer and cell lines. For each antibody, altogether 708 spots of tissues and cells are analysed and the resulting images and data are presented as freely available in the Human Protein Atlas (www.proteinatlas.org). The new version 4 of the atlas, including more than 5 million images of immunohistochemically stained tissues and cells, is based on 6122 antibodies, representing 5011 human proteins encoded by approximately 25% of the human genome. The gene-centric database includes a putative classification of proteins in various protein classes, both functional classes, such as kinases or transcription factors and project-related classes, such as candidate genes for cancer or cardiovascular diseases. For each of the internally generated antibodies, the exact antigen sequence is presented, together with a visualization of application-specific validation data, including a protein array assay, western blot analysis, immunohistochemistry and, in most cases, immunofluorescent-based confocal microscopy. The updated version also includes new search algorithms to allow complex queries regarding expression profiles, protein classes and chromosome location. Thus, the presented Human Protein Atlas provides a resource for pathology-based biomedical research, including protein science and biomarker discovery.

Letter to the Editor: Towards a new era of tissue-based diagnosis and research

Letter to the Editor: Towards a new era of tissue-based diagnosis and research