Prostate-specific antigen: gene structure and regulation of expression

As a start of our study of prostate-specific and androgen-dependent expression of PA, various PA cDNAs (Chapter II) and the PA gene (Chapter Ill) were isolated and characterized. The PA gene turned out to be a member of a small kaHikrein family, encompassing two other closely related genes Ithe human Glandular Kal!ikrein-1 gene (hGK-1), and the tissue kallikrein gene {KLK1)]. Genetic characterization of the human kallikrein genes showed clustering in a 60 kb segment on chromosome 19q13.2-13.4 (Chapters IV and V). The hGK-1 gene shows a strong homology to PA and is, similarly to PA, exclusively expressed in prostate tissue. KLK1 is mainly expressed in kidney, pancreas and salivary glands. In addition to PA eDNA, hGK-1 cDNAs were isolated and characterized. This allowed the comparison of PA and hGK-1 mRNA expression. Using hGK-1 and ?A-specific eDNA probes, androgen-stimulated mRNA expression of PA and hGK-1 could be determined (Chapter VI). Further, an androgen responsive element in the promoter region of the PA gene was defined and tested for its functional activity (Chapter Vl!). The homology of the PA, hGK-1 and KLK1 genes does not only include the open reading frame, but extends into the promoter regions, although the genes are at least partially expressed in different tissues and at different levels. The above mentioned aspects, resulted in the development of a model system for the study of tissue-specific and hormone-responsive gene expression in the human prostate

How standardization of the pre-analytical phase of both research and diagnostic biomaterials can increase reproducibility of biomedical research and diagnostics.

Comparison of published biomedical studies shows that a large proportion are irreproducible, causing severe damage to society and creating an image of wasted investments. These observations are of course damaging to the biomedical research field, which is currently full of future promise. Precision medicine and disease prevention are successful, but are progressing slowly due to irreproducible study results. Although standardization is mentioned as a possible solution, it is not always clear how this could decrease or prevent irreproducible results in biomedical studies. In this article more insight is given into what quality, norms, standardization, certification, accreditation and optimized infrastructure can accomplish to reveal causes of irreproducibility and increase reproducibility when collecting biomaterials. CEN and ISO standards for the sample pre-analytical phase are currently being developed with the support of the SPIDIA4P project, and their role in increasing reproducibility in both biomedical research and diagnostics is demonstrated. In particular, it is described how standardized methods and quality assurance documentation can be exploited as tools for: 1) recognition and rejection of 'not fit for purpose' samples on the basis of detailed sample metadata, and 2) identification of methods that contribute to irreproducibility which can be adapted or replaced

Buccal swab as a reliable predictor for X inactivation ratio in inaccessible tissues

Background As a result of the epigenetic phenomenon of X chromosome inactivation (XCI) every woman is a mosaic of cells with either an inactive paternal X chromosome or an inactive maternal X chromosome. The ratio between inactive paternal and maternal X chromosomes is different for every female individual, and can influence an X-encoded trait or disease. A multitude of X linked conditions is known, and for many of them it is recognised that the phenotype in affected female carriers of the causative mutation is modulated by the XCI ratio. To predict disease severity an XCI ratio is usually determined in peripheral blood samples. However, the correlation between XCI ratios in peripheral blood and disease affected tissues, that are often inaccessible, is poorly understood. Here, we tested several tissues obtained from autopsies of 12 female individuals for patch size and XCI ratio. Methods XCI ratios were analysed using methylsensitive PCR-based assays for the AR, PCSK1N and SLITRK4 loci. XCI patch size was analysed by testing the XCI ratio of tissue samples with decreasing size. Results XCI patch size was analysed for liver, muscle, ovary and brain samples and was found too small to confound testing for XCI ratio in these tissues. XCI ratios were determined in the easily accessible tissues, blood, buccal epithelium and hair follicle, and compared with ratios in several inaccessible tissues. Conclusions Buccal epithelium is preferable over peripheral blood for predicting XCI ratios of inaccessible tissues. Ovary is the only inaccessible tissue showing a poor correlation to blood and buccal epithelium, but has a good correlation to hair follicle instead

Improvement of European translational cancer research. Collaboration between comprehensive cancer centers

Even though the increasing incidence of cancer is mainly a consequence of a population with a longer life span, part of this augmentation is related to the increasing prevalence of patients living with a chronic cancer disease. To fight the problem, improved preventive strategies are mandatory in combination with an innovative health care provision that is driven by research. To overcome the weakness of translational research the OECI is proposing a practical approach as part of a strategy foreseen by the EUROCAN+PLUS feasibility study, which was launched by the EC in order to identify mechanisms for the coordination of cancer research in Europe

Factors that drive the increasing use of FFPE tissue in basic and translational cancer research

The decision to use 10% neutral buffered formalin fixed, paraffin embedded (FFPE) archival pathology material may be dictated by the cancer research question or analytical technique, or may be governed by national ethical, legal and social implications (ELSI), biobank, and sample availability and access policy. Biobanked samples of common tumors are likely to be available, but not all samples will be annotated with treatment and outcomes data and this may limit their application. Tumors that are rare or very small exist mostly in FFPE pathology archives. Pathology departments worldwide contain millions of FFPE archival samples, but there are challenges to availability. Pathology departments lack resources for retrieving materials for research or for having pathologists select precise areas in paraffin blocks, a critical quality control step. When samples must be sourced from several pathology departments, different fixation and tissue processing approaches create variability in quality. Researchers must de