Automatic registration of multi-modal microscopy images for integrative analysis of prostate tissue sections

Background Prostate cancer is one of the leading causes of cancer related deaths. For diagnosis, predicting the outcome of the disease, and for assessing potential new biomarkers, pathologists and researchers routinely analyze histological samples. Morphological and molecular information may be integrated by aligning microscopic histological images in a multiplex fashion. This process is usually time-consuming and results in intra- and inter-user variability. The aim of this study is to investigate the feasibility of using modern image analysis methods for automated alignment of microscopic images from differently stained adjacent paraffin sections from prostatic tissue specimens. Methods Tissue samples, obtained from biopsy or radical prostatectomy, were sectioned and stained with either hematoxylin & eosin (H&E), immunohistochemistry for p63 and AMACR or Time Resolved Fluorescence (TRF) for androgen receptor (AR). Image pairs were aligned allowing for translation, rotation and scaling. The registration was performed automatically by first detecting landmarks in both images, using the scale invariant image transform (SIFT), followed by the well-known RANSAC protocol for finding point correspondences and finally aligned by Procrustes fit. The Registration results were evaluated using both visual and quantitative criteria as defined in the text. Results Three experiments were carried out. First, images of consecutive tissue sections stained with H&E and p63/AMACR were successfully aligned in 85 of 88 cases (96.6%). The failures occurred in 3 out of 13 cores with highly aggressive cancer (Gleason score ≥ 8). Second, TRF and H&E image pairs were aligned correctly in 103 out of 106 cases (97%). The third experiment considered the alignment of image pairs with the same staining (H&E) coming from a stack of 4 sections. The success rate for alignment dropped from 93.8% in adjacent sections to 22% for sections furthest away. Conclusions The proposed method is both reliable and fast and therefore well suited for automatic segmentation and analysis of specific areas of interest, combining morphological information with protein expression data from three consecutive tissue sections. Finally, the performance of the algorithm seems to be largely unaffected by the Gleason grade of the prostate tissue samples examined, at least up to Gleason score 7

The expression pattern of matrix-producing tumor stroma is of prognostic importance in breast cancer

Background: There are several indications that the composition of the tumor stroma can contribute to the malignancy of a tumor. Here we utilized expression data sets to identify metagenes that may serve as surrogate marker for the extent of matrix production and vascularization of a tumor and to characterize prognostic molecular components of the stroma. Methods: TCGA data sets from six cancer forms, two breast cancer microarray sets and one mRNA data set of xenografted tumors were downloaded. Using the mean correlation as distance measure compact clusters with genes representing extracellular matrix production (ECM metagene) and vascularization (endothelial metagene) were defined. Explorative Cox modeling was used to identify prognostic stromal gene sets. Results: Clustering of stromal genes in six cancer data sets resulted in metagenes, each containing three genes, representing matrix production and vascularization. The ECM metagene was associated with poor prognosis in renal clear cell carcinoma and in lung adenocarcinoma but not in other cancers investigated. Explorative Cox modeling using gene pairs identified gene sets that in multivariate models were prognostic in breast cancer. This was validated in two microarray sets. Two notable genes are TCF4 and P4HA3 which were included in the sets associated with positive and negative prognosis, respectively. Data from laser-microdissected tumors, a xenografted tumor data set and from correlation analyses demonstrate the stroma specificity of the genes. Conclusions: It is possible to construct ECM and endothelial metagenes common for several cancer forms. The molecular composition of matrix-producing cells, rather than the extent of matrix production seem to be important for breast cancer prognosis

miR-145 suppress the androgen receptor in prostate cancer cells and correlates to prostate cancer prognosis.

Androgen signalling through the androgen receptor (AR) is essential for prostate cancer initiation, progression and transformation to the lethal castration-resistant state. The aim of this study was to characterize the mechanisms by which miR-145 deregulation contribute to prostate cancer progression. The miR-145 levels, measured by quantitative reverse transcription-polymerase chain reaction, were found to inversely correlate with occurrence of metastases, survival and androgen deprivation therapy response in a well-characterized prostate cancer cohort. Introduction of ectopic miR-145 in prostate cancer cells generated an inhibitory effect on the AR at both transcript and protein levels as well as its activity and downstream targets prostate-specific antigen (PSA), kallikrein-related peptidase 2 and TMPRSS2. The regulation was shown to be mediated by direct binding using Ago2-specific immunoprecipitation, but there was also indication of synergetic AR activation. These findings were verified in clinical prostate specimens by demonstrating inverse correlations between miR-145 and AR expression as well as serum PSA levels. In addition, miR-145 was found to regulate androgen-dependent cell growth in vitro. Our findings put forward novel possibilities of therapeutic intervention, as miR-145 potentially could decrease both the stem cells and the AR expressing bulk of the tumour and hence reduce the transformation to the deadly castration-resistant form of prostate cancer

Guidance for laboratories performing molecular pathology for cancer patients

Molecular testing is becoming an important part of the diagnosis of any patient with cancer. The challenge to laboratories is to meet this need, using reliable methods and processes to ensure that patients receive a timely and accurate report on which their treatment will be based. The aim of this paper is to provide minimum requirements for the management of molecular pathology laboratories. This general guidance should be augmented by the specific guidance available for different tumour types and tests. Preanalytical considerations are important, and careful consideration of the way in which specimens are obtained and reach the laboratory is necessary. Sample receipt and handling follow standard operating procedures, but some alterations may be necessary if molecular testing is to be performed, for instance to control tissue fixation. DNA and RNA extraction can be standardised and should be checked for quality and quantity of output on a regular basis. The choice of analytical method(s) depends on clinical requirements, desired turnaround time, and expertise available. Internal quality control, regular internal audit of the whole testing process, laboratory accreditation, and continual participation in external quality assessment schemes are prerequisites for delivery of a reliable service. A molecular pathology report should accurately convey the information the clinician needs to treat the patient with sufficient information to allow for correct interpretation of the result. Molecular pathology is developing rapidly, and further detailed evidence-based recommendations are required for many of the topics covered here

Positional and functional mapping of a neuroblastoma differentiation gene on chromosome 11

BACKGROUND: Loss of chromosome 11q defines a subset of high-stage aggressive neuroblastomas. Deletions are typically large and mapping efforts have thus far not lead to a well defined consensus region, which hampers the identification of positional candidate tumour suppressor genes. In a previous study, functional evidence for a neuroblastoma suppressor gene on chromosome 11 was obtained through microcell mediated chromosome transfer, indicated by differentiation of neuroblastoma cells with loss of distal 11q upon introduction of chromosome 11. Interestingly, some of these microcell hybrid clones were shown to harbour deletions in the transferred chromosome 11. We decided to further exploit this model system as a means to identify candidate tumour suppressor or differentiation genes located on chromosome 11. RESULTS: In a first step, we performed high-resolution arrayCGH DNA copy-number analysis in order to evaluate the chromosome 11 status in the hybrids. Several deletions in both parental and transferred chromosomes in the investigated microcell hybrids were observed. Subsequent correlation of these deletion events with the observed morphological changes lead to the delineation of three putative regions on chromosome 11: 11q25, 11p13->11p15.1 and 11p15.3, that may harbour the responsible differentiation gene. CONCLUSION: Using an available model system, we were able to put forward some candidate regions that may be involved in neuroblastoma. Additional studies will be required to clarify the putative role of the genes located in these chromosomal segments in the observed differentiation phenotype specifically or in neuroblastoma pathogenesis in general

Dark stars at the Galactic centre - the main sequence

In regions of very high dark matter density such as the Galactic centre, the capture and annihilation of WIMP dark matter by stars has the potential to significantly alter their evolution. We describe the dark stellar evolution code DarkStars, and present a series of grids of WIMP-influenced stellar models for main sequence stars. We describe changes in which occur as a function of the rate of energy injection by WIMPs, for stars of 0.3-2.0 solar masses and metallicities Z = 0.0003-0.02. We show what rates of energy injection can be obtained using realistic orbital parameters for stars at the Galactic centre, including detailed consideration of the velocity and density profiles of dark matter. Capture and annihilation rates are strongly boosted when stars follow elliptical rather than circular orbits. If there is a spike of dark matter induced by the supermassive black hole at the Galactic centre, single solar-mass stars following orbits with periods as long as 50 years and eccentricities as low as 0.9 could be significantly affected. Binary systems with similar periods about the Galactic centre could be affected on even less eccentric orbits. The most striking evidence of this scenario would be the existence of a binary consisting of a low-mass protostar and a higher-mass evolved star. The observation of low-mass stars and/or binaries on such orbits would either provide a detection of WIMP dark matter, or place stringent limits on the combination of the WIMP mass, spin-dependent nuclear-scattering cross-section, halo density and velocity distribution near the Galactic centre. In some cases, the limits on the WIMP mass and spin-dependent nuclear-scattering cross-section would be of comparable sensitivity to current direct-detection experiments.Comment: v2: Accepted version, added discussion on binaries. v3: Updated to be consistent with erratum submitted to MNRAS. Main results and all conclusions unchange

Integration of next-generation sequencing in clinical diagnostic molecular pathology laboratories for analysis of solid tumours; an expert opinion on behalf of IQN Path ASBL

Contains fulltext : 169719.pdf (publisher's version ) (Open Access)The clinical demand for mutation detection within multiple genes from a single tumour sample requires molecular diagnostic laboratories to develop rapid, high-throughput, highly sensitive, accurate and parallel testing within tight budget constraints. To meet this demand, many laboratories employ next-generation sequencing (NGS) based on small amplicons. Building on existing publications and general guidance for the clinical use of NGS and learnings from germline testing, the following guidelines establish consensus standards for somatic diagnostic testing, specifically for identifying and reporting mutations in solid tumours. These guidelines cover the testing strategy, implementation of testing within clinical service, sample requirements, data analysis and reporting of results. In conjunction with appropriate staff training and international standards for laboratory testing, these consensus standards for the use of NGS in molecular pathology of solid tumours will assist laboratories in implementing NGS in clinical services

A global fit of the MSSM with GAMBIT

We study the seven-dimensional Minimal Supersymmetric Standard Model (MSSM7) with the new GAMBIT software framework, with all parameters defined at the weak scale. Our analysis significantly extends previous weak-scale, phenomenological MSSM fits, by adding more and newer experimental analyses, improving the accuracy and detail of theoretical predictions, including dominant uncertainties from the Standard Model, the Galactic dark matter halo and the quark content of the nucleon, and employing novel and highly-efficient statistical sampling methods to scan the parameter space. We find regions of the MSSM7 that exhibit co-annihilation of neutralinos with charginos, stops and sbottoms, as well as models that undergo resonant annihilation via both light and heavy Higgs funnels. We find high-likelihood models with light charginos, stops and sbottoms that have the potential to be within the future reach of the LHC. Large parts of our preferred parameter regions will also be accessible to the next generation of direct and indirect dark matter searches, making prospects for discovery in the near future rather good

Neuroblastoma Cell Differentiation: The Role of Neurotrophin Receptor Signaling and N-myc Expression

Neuroblastoma is a tumor of sympathetic nervous system derivation, mostly afflicting young children. This thesis is focused on a group of receptor proteins for neurotrophic factors, the Trk family, and on N-Myc, a transcription factor, all important in the formation of the sympathetic nervous system as well as in determining neuroblastoma patient outcome. Expression of trkA and trkC is linked to favorable prognosis, while amplification of the N-myc gene is strongly correlated to poor outcome. The role of trkA and trkC in neuroblastoma cell differentiation was studied using neuroblastoma cell lines constitutively expressing trkA or trkC. Stimulation of the trkA- and trkC-transfected cells with their cognate ligands resulted in differentiation of both cell clones, the differentiated trkC-transfected cells lacking important neuronal features present in the differentiated trkA-transfected cells. Signaling elicited by the two receptors was studied and differences described. The possible connection between expression of N-myc and trkB, another trk family member, was tested by examination of expression of these genes in a set of neuroblastomas and neuroblastoma cell lines. Results suggested high expression of N-myc per se to be insufficient to induce trkB expression. In other experiments, retinoic acid, an agent known to induce trkB expression was added to neuroblastoma cells in combination with the ligand of TrkB, brain-derived neurotrophic factor. Data from characterization of the resulting phenotype indicated that a sympathetic neuronal differentiation did not take place in these cells and alternative explanations were suggested. N-myc expression in the developing human sympathetic nervous system was studied using in situ hybridization and the role of N-myc in neuroblastoma cell was examined, utilizing non-N-myc-amplified neuroblastoma cells with constitutive N-myc overexpression as a model system. Overall, the capacity of these cells to differentitate morphologically in response to various differentiation protocols was retained, thus offering an explanation to the lack of correlation between N-myc expression and patient outcome in non-N-myc-amplified tumors