Environmental properties of cells improve machine learning-based phenotype recognition accuracy

To answer major questions of cell biology, it is often essential to understand the complex phenotypic composition of cellular systems precisely. Modern automated microscopes produce vast amounts of images routinely, making manual analysis nearly impossible. Due to their efficiency, machine learning-based analysis software have become essential tools to perform single-cell-level phenotypic analysis of large imaging datasets. However, an important limitation of such methods is that they do not use the information gained from the cellular micro- and macroenvironment: the algorithmic decision is based solely on the local properties of the cell of interest. Here, we present how various features from the surrounding environment contribute to identifying a cell and how such additional information can improve single-cell-level phenotypic image analysis. The proposed methodology was tested for different sizes of Euclidean and nearest neighbour-based cellular environments both on tissue sections and cell cultures. Our experimental data verify that the surrounding area of a cell largely determines its entity. This effect was found to be especially strong for established tissues, while it was somewhat weaker in the case of cell cultures. Our analysis shows that combining local cellular features with the properties of the cell's neighbourhood significantly improves the accuracy of machine learning-based phenotyping

Simultaneous detection of BRCA mutations and large genomic rearrangements in germline DNA and FFPE tumor samples

The development of breast and ovarian cancer is strongly connected to the inactivation of the BRCA1 and BRCA2 genes by different germline and somatic alterations, and their diagnosis has great significance in targeted tumor therapy, since recently approved PARP inhibitors show high efficiency in the treatment of BRCA-deficient tumors. This raises the need for new diagnostic methods that are capable of performing an integrative mutation analysis of the BRCA genes not only from germline DNA but also from formalin-fixed and paraffin-embedded (FFPE) tumor samples. Here we describe the development of such a methodology based on next-generation sequencing and a new bioinformatics software for data analysis. The diagnostic method was initially developed on an Illumina MiSeq NGS platform using germline-mutated stem cell lines and then adapted for the Ion Torrent PGM NGS platform as well. We also investigated the usability of NGS coverage data for the detection of copy number variations and exon deletions as a replacement of the conventional MLPA technique. Finally, we tested the developed workflow on FFPE samples from breast and ovarian cancer patients. Our method meets the sensitivity and specificity requirements for the genetic diagnosis of breast and ovarian cancers both from germline and FFPE samples

Prosztatarákban szenvedő betegek definitív ellátásának tanulságai és modern stratégiái = Edifications and modern strategies of localized prostate cancers’ definitive therapy

Absztrakt: Bevezetés: A korszerű diagnosztikus és kezelési lehetőségek igénybevételével a prosztatarák mortalitása jelentősen csökkenthető. A korai stádiumú daganatos betegek radikális műtéttel vagy sugárkezeléssel is meggyógyíthatók. Célkitűzés: A jelenlegi kivizsgálási és a terápiás módszerek áttekintése és összehasonlítása a korábbi gyakorlattal a diagnózis pontossága, a hatékonyság javítása és a mellékhatások csökkentése szempontjából, valamint általános összefüggések megvilágítása súlyos szövődményes esetek kapcsán. Módszer: Két prosztatadaganatos eset mentén ismertetjük a pontos szövettani diagnózis jelentőségének és változásának, a megfelelő képalkotó vizsgálatok alkalmazásának, a besugárzás hagyományos és modern paramétereinek, valamint a sugárkezelés akut és késői szövődményeinek részleteit. Elemzésre kerülnek a korábbi és a napjainkban alkalmazott módszerek lényegi különbségei és ezek konzekvenciái is. Eredmények: Betegeink diagnózisában napjainkra módosult a szövettani vélemény. Mindkét esetben 2009–2011-ben hagyományos, 3 dimenziós konformális technikával definitív sugárkezelés történt, melynek hatására meggyógyultak prosztatatumorukból. Az egyik beteg besugárzást kapott a hólyagra is, mivel a daganat beszűrte azt. A másiknál a biztonsági zóna kiterjesztése miatt, a kor színvonalán álló besugárzás a hólyagalapot is kiterjedtebben érintette. Bár mindkét esetben hetek alatt szanálódott a hevenyen jelentkező kettes fokozatú cystitis, mégis késői szövődményként zsugorhólyag alakult ki. Ezek elhárítására – az első esetben a vérzés megállítására, a második esetben az incontinentia megszüntetésére – a hagyományos eljárások kudarca miatt radikális cystoprostatectomiára volt szükség. Következtetések: A prosztatadaganatok diagnosztikai és terápiás fejlődésének összetett lépései együttesen vezetnek a súlyos szövődmények elkerüléséhez. Az adatok szintézise azonban akkor lehet még sikeresebb, ha azokat a korábbi tapasztalatok birtokában is elemezzük. Orv Hetil. 2018; 159(32): 1317–1325. | Abstract: Introduction: Mortality of prostate carcinoma can be significantly decreased by the use of modern diagnostic and therapeutic options. Patients in early stages can be cured by radical surgery or radiotherapy. Aim: Overview and comparison of previous and present diagnostic and therapeutic methods regarding accuracy of diagnosis, improvement of efficiency and decrease of toxicities. We also aimed to explore general correlations in case of serious complications. Method: By the help of two prostate cancer patients we demonstrate the importance of accuracy and change of histological diagnosis, significance of proper imaging techniques, and also show parameters of conventional and modern radiotherapy and their acute and chronic complications. Differences of previous and present methods and their consequences were analyzed. Results: By now, histological findings in the patients’ diagnosis have changed. Both patients received conventional three-dimensional definitive radiotherapy in 2009–2011, and their prostate cancer was cured. In one case, urinary bladder also received radiotherapy because prostate carcinoma had infiltrated it. In the other case, the contemporary radiotherapy involved urinary bladder’s fundus due to safety margins. Although acute grade 2 cystitis developed in both cases and recovered in several weeks, as late complication bladder shrinkage developed, which after the ineffectiveness of conventional therapies had to be cured by radical cystoprostatectomy – in order to cease bleeding and to cure incontinence. Conclusions: In case of prostate carcinomas, serious complications can be avoided by the improvement of diagnostic and therapeutic options. Synthesis of data could be more successful if they were analyzed in the light of previous experiences. Orv Hetil. 2018; 159(32): 1317–1325

Novel Diagnostic Value of Driver Gene Transcription Signatures to Characterise Clear Cell Renal Cell Carcinoma, ccRCC

Routine molecular tumour diagnostics are augmented by DNA-based qualitative and quantitative molecular techniques detecting mutations of DNA. However, in the past decade, it has been unravelled that the phenotype of cancer, as it’s an extremely complex disease, cannot be fully described and explained by single or multiple genetic variants affecting only the coding regions of the genes. Moreover, studying the manifestation of these somatic mutations and the altered transcription programming—driven by genomic rearrangements, dysregulation of DNA methylation and epigenetic landscape—standing behind the tumorigenesis and detecting these changes could provide a more detailed characterisation of the tumour phenotype. Consequently, novel comparative cancer diagnostic pipelines, including DNA- and RNA-based approaches, are needed for a global assessment of cancer patients. Here we report, that by monitoring the expression patterns of key tumour driver genes by qPCR, the normal and the tumorous samples can be separated into distinct categories. Furthermore, we also prove that by examining the transcription signatures of frequently affected genes at 3p25, 3p21 and 9p21.3 genomic regions, the ccRCC (clear cell renal cell carcinoma) and non-tumorous kidney tissues can be distinguished based on the mRNA level of the selected genes. Our results open new diagnostics possibilities where the mRNA signatures of tumour drivers can supplement the DNA-based approaches providing a more precise diagnostics opportunity leading to determine more precise therapeutic protocols