Beyond germline genetic testing - heterozygous pathogenic variants in PMS2 in two children with Osteosarcoma and Ependymoma.

BACKGROUND: Lynch syndrome (LS) is not considered part of childhood cancer predisposition syndromes. CASE PRESENTATION: Analysis of a pediatric osteosarcoma (OS) displayed hypermutation (16.8), alternative lengthening of telomeres (ALT), loss of PMS2 expression in tumor tissue (retained in non-neoplastic cells), PMS2 loss of heterozygosity (LOH), and high-degree of microsatellite instability (MSI) tested by PCR. A heterozygous duplication c.1076dup p.(Leu359Phefs*6) in exon 10 of NM_000535.6:PMS2 was detected by SNV analysis in peripheral blood, confirming diagnosis of LS in the patient. The tumor molecular features suggest LS-associated development of OS. In a second case, whole-genome sequencing identified a heterozygous SNV c.1 A > T p.? in exon 1 of PMS2 in tumor and germline material of a girl with ependymoma. Tumor analysis displayed evidence for ALT and low mutational burden (0.6), PMS2 expression was retained, MSI was low. Multiplex ligation-dependent probe amplification identified no additional PMS2 variant and germline MSI testing did not reveal increased gMSI ratios in the patient´s lymphocytes. Thus, CMMRD was most closely excluded and our data do not suggest that ependymoma was related to LS in the child. CONCLUSIONS: Our data suggest that the LS cancer spectrum may include childhood cancer. The importance of LS in pediatric cancers necessitates prospective data collection. Comprehensive molecular workup of tumor samples is necessary to explore the causal role of germline genetic variants.

Evolutionary patterns of chromosomal instability and mismatch repair deficiency in proximal and distal colorectal cancer.

AIM: Colorectal carcinomas (CRCs) progress through heterogeneous pathways. The aim of this study was to analyse whether or not the cytogenetic evolution of CRC is linked to tumour site, level of chromosomal imbalance and metastasis. METHOD: A set of therapy-naïve pT3 CRCs comprising 26 proximal and 49 distal pT3 CRCs was studied by combining immunohistochemistry of mismatch repair (MMR) proteins, microsatellite analyses and molecular karyotyping as well as clinical parameters. RESULTS: A MMR deficient/microsatellite-unstable (dMMR/MSI-H) status was associated with location of the primary tumour proximal to the splenic flexure, and dMMR/MSI-H tumours presented with significantly lower levels of chromosomal imbalances compared with MMR proficient/microsatellite-stable (pMMR/MSS) tumours. Oncogenetic tree modelling suggested two evolutionary clusters characterized by dMMR/MSI-H and chromosomal instability (CIN), respectively, for both proximal and distal CRCs. In CIN cases, +13q, -18q and +20q were predicted as preferentially early events, and -1p, -4 -and -5q as late events. Separate oncogenetic tree models of proximal and distal cases indicated similar early events independent of tumour site. However, in cases with high CIN defined by more than 10 copy number aberrations, loss of 17p occurred earlier in cytogenetic evolution than in cases showing low to moderate CIN. Differences in the oncogenetic trees were observed for CRCs with lymph node and distant metastasis. Loss of 8p was modelled as an early event in node-positive CRC, while +7p and +8q comprised early events in CRC with distant metastasis. CONCLUSION: CRCs characterized by CIN follow multiple, interconnected genetic pathways in line with the basic 'Vogelgram' concept proposed for the progression of CRC that places the accumulation of genetic changes at centre of tumour evolution. However, the timing of specific genetic events may favour metastatic potential.

Drug loading of polymeric micelles.

PURPOSE: To gain mechanistic insights into drug loading and lyophilization of polymeric micelles. METHODS: PEGylated poly-4-(vinylpyridine) micelles were loaded with dexamethasone. Three different methods were applied and compared: O/W emulsion, direct dialysis, cosolvent evaporation. Micellar dispersions with the highest drug load were lyophilized with varying lyoprotectors: sucrose, trehalose, maltose, a polyvinylpyrrolidine derivative, and ß-cyclodextrin derivatives. For comparison, other PEGylated block copolymer micelles (PEGylated polylactic acid, polylactic acid-co-glycolic acid, polycaprolactone) were freeze-dried. RESULTS: Drug loading via direct dialysis from acetone was a less effective loading method which led to dexamethasone loads <2% w/w. O/W emulsion technique from dichlormethane increased drug load up to ~13% w/w; optimized cosolvent evaporation increased load up to ~19% w/w. An important step for cosolvent evaporation was solubility screen of the drug prior to preparation. Loading was maintained upon lyophilization with ß-cyclodextrins which proved to be versatile stabilizers for other block copolymer micelles. CONCLUSION: Careful solvent selection prior to cosolvent evaporation was a beneficial approach to load hydrophobic drugs into polymeric micelles. Moreover, ß-cyclodextrins could be used as versatile lyoprotectors for these micelles.

HistoViewer: an interactive e-learning platform facilitating group and peer group learning.

Understanding tissue architecture and the morphological characteristics of cells is a central prerequisite to comprehending the basis of physiological tissue function in healthy individuals and relating this to disease states. Traditionally, medical curricula include courses where students examine glass slides of cytological or tissue samples under a light microscope. However, it is challenging to implement group and peer group learning in these courses and to give students sufficient time to study specimens. An increasing number of medical schools have thus started to implement digital slide viewers, so-called virtual microscopes, in histology and histopathology. These websites are mostly based on standard commercial software and offer limited adaptation to the special needs of first-year students. An e-learning platform has therefore been developed for use in cytology and histology courses. This virtual microscopy tool is coupled to a central database in which students can label and store the positions of individual structures for later repetition. As learning in pairs and peer groups has been shown to provide a high learning outcome, identified structures can be shared and discussed with students' peers or faculty via a built-in communication module. This website has the possibility of opening an arbitrary number of frames which all can actively be moved and changed in magnification to enable the comparison of specimens and thus encourage a more global understanding of related tissues. HistoViewer is thus suggested as an e-learning tool combining several modern teaching concepts.

Follicular dendritic cell sarcoma of the spleen.

Diagnosis of primary spindle cell tumors of the spleen is challenging because of the limited immunologic and cytogenetic characterization of this rare entity. We report a case of primary follicular dendritic cell (FDC) sarcoma of the spleen in a 44-year-old woman. Indications for FDC included positive staining for CD21, Ki-M4P, CD14, and fascin. Expression of both standard FDC markers CD23 and CD35 was detected immunohistochemically using tyramide signal amplification. Cytogenetic analysis revealed multiple clonal chromosomal aberrations involving unbalanced translocations of chromosomes X, 3, 5, 7, 8, 9, and 10, leading to net gains at 3q, 7p, 8q, and 9q and net losses at Xp, 8p, 9p, and 10p. Loss at Xp has been described previously in another tumor with FDC features, suggesting that this aberration might play a common role in this malignancy.

Comparative genomic hybridization analysis on male breast cancer.

The spectrum of genetic alterations in primary male breast cancer is not well established. We analyzed chromosomal imbalances in 39 tumor samples from primary male breast cancer by comparative genomic hybridization (CGH) and correlated CGH findings with clinicopathological factors. Chromosomal gains were most frequent at 1q (46%), 8q (46%), 16p (36%), 17q (36%), Xq (28%), 20q (26%) and Xp (18%). Losses were most commonly observed at 8p (36%), 16q (28%), 13q (28%), 6q (18%), 11q (18%) and 22q (18%). Gains at 16p, 20q and Xq and losses at 13q correlated significantly with higher degree of cytogenetic complexity. Significant associations with clinicopathological factors were observed for +8q and -16q with larger tumor size and -16q with lower proliferative activity and lower grade of malignancy. A comparison with reported CGH data from female breast cancer showed a similar pattern of chromosomal imbalances, including +1q, -8p, +8q, -13q, +16p, -16q, +17q and +20q. Our results indicate that male breast cancer shares a common pattern of imbalances with female breast cancer, suggesting that similar genetic events may underlie the development and progression of male and female breast cancer.

Differential diagnosis of gastrointestinal leiomyoma versus gastrointestinal stromal tumor.

INTRODUCTION: Strategies for the diagnosis of tumors arising in the intestinal muscular wall are rapidly evolving. Immunoreactivity for CD117 (KIT) usually supports the diagnosis of gastrointestinal stromal tumor (GIST), but a small subset of GISTs lacks KIT expression. In these cases the differential diagnosis of KIT-negative GIST versus one of their morphological mimics is difficult and bears critical implications for therapeutic management. CASE REPORT: Here, we report a case of a KIT-negative smooth muscle cell tumor of the colon in a 21-year-old man with the clinical appearance of GIST. Mutations of the KIT and platelet-derived growth factor receptor alpha (PDGFRA) gene could be ruled out. No chromosomal imbalances characteristic of GIST were found. However, cytogenetic analysis revealed losses at 7q, which has previously been reported in cases of uterine leiomyoma. DISCUSSION: We discuss current approaches to the differential diagnosis of true gastrointestinal smooth muscle cell tumor versus GIST.