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Abstract Background Chondrosarcoma (Chs) is the third most frequent primary malignant tumour of bone and can be primary or secondary, the latter results mainly from the malignant transformation of a benign pre-existing tumour. Methods All the cases diagnosed as Chs (primary tumours, recurrences and/or metastasis and xenotransplanted Chs) from the files of our Department were collected. Only cases with paraffin blocks available were selected (Total 32 cases). Six Tissue Microarrays (TMAs) were performed and all the cases and biopsies were distributed into the following groups: a) only paraffin block available from primary and/or metastatic tumours (3 TMAs), b) paraffin block available from primary and/or metastatic tumours as well as from the corresponding Nude mice xenotransplant (2 TMAs), c) only paraffin block available from xenotransplanted Chs (1 TMA). A reclassification of all the cases was performed; in addition, conventional hematoxylin-eosin as well as immunohistochemistry staining (S100, SOX-9, Ki-67, BCL-2, p53, p16, CK, CD99, Survivin and Caveolin) was analyzed in all the TMA. Results The distribution of the cases according to the histopathological pattern and the location of tumours were as follows: fourteen Grade I Chs (all primaries), two primary Grade II Chs, ten Grade III Chs (all primaries), five dedifferentiated Chs (four primaries and one primary with metastasis), and two Chs from cell cultures (Ch grade III). One recurrent extraskeletal myxoid Chs was included as a control in the TMA. Although there was heterogeneity in immunohistochemistry results of the different material analyzed, S100, SOX-9, Caveolin and Survivin were more expressed. The number of passages in xenotransplants fluctuated between 1 and 13. Curiously, in Grade I Chs, these implanted tumours hardly grew, and the number of passages did not exceed one. Conclusion The study of Chs by means of TMA techniques is very important because it will improve the assessment of different antibodies applied in the immunohistochemical assays. Xenotransplanted tumours in TMA improve knowledge concerning the variability in the morphological pattern shown by these tumours during the evolution in nudes.

Abstract Primary sarcomas of the major blood vessels can be classified based on location in relationship to the wall or by histologic type. Angiosarcomas are malignant neoplasms that arise from the endothelial lining of the blood vessels; those arising in the intimal compartment of pulmonary artery are rare. We report a case of pulmonary artery angiosarcoma in a 36-year old female with pulmonary masses. The patient had no other primary malignant neoplasm, thus excluding a metastatic lesion. Gross examination revealed a thickened right pulmonary artery and a necrotic and hemorrhagic tumor, filling and occluding the vascular lumen. The mass extended distally, within the pulmonary vasculature of the right lung. Microscopically, an intravascular undifferentiated tumor was identified. The tumor cells showed expression for vascular markers VEGFR, VEGFR3, PDGFRa, FGF, Ulex europaeus, FVIII, FLI-1, CD31 and CD34; p53 was overexpressed and Ki67 proliferative rate was increased. Intravascular angiosarcomas are aggressive neoplasms, often associated with poor outcome. Virtual slide The virtual slide(s) for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/2315906377648045.

Abstract Background Chondrosarcoma is the second most common primary sarcoma of bone. High-grade conventional chondrosarcoma and dedifferentiated chondrosarcoma have a poor outcome. In pre-clinical research aiming at the identification of novel treatment targets, the need for representative cell lines and model systems is high, but availability is scarce. Methods We developed and characterized three cell lines, derived from conventional grade III chondrosarcoma (L835), and dedifferentiated chondrosarcoma (L2975 and L3252) of bone. Proliferation and migration were studied and we used COBRA-FISH and array-CGH for karyotyping and genotyping. Immunohistochemistry for p16 and p53 was performed as well as TP53 and IDH mutation analysis. Cells were injected into nude mice to establish their tumorigenic potential. Results We show that the three cell lines have distinct migrative properties, L2975 had the highest migration rate and showed tumorigenic potential in mice. All cell lines showed chromosomal rearrangements with complex karyotypes and genotypic aberrations were conserved throughout late passaging of the cell lines. All cell lines showed loss of CDKN2A, while TP53 was wild type for exons 5–8. L835 has an IDH1 R132C mutation, L2975 an IDH2 R172W mutation and L3252 is IDH wild type. Conclusions Based on the stable culturing properties of these cell lines and their genotypic profile resembling the original tumors, these cell lines should provide useful functional models to further characterize chondrosarcoma and to evaluate new treatment strategies.

Abstract Background Xenografts have been shown to provide a suitable source of tumor tissue for molecular analysis in the absence of primary tumor material. We utilized ES xenograft series for integrated microarray analyses to identify novel biomarkers. Method Microarray technology (array comparative genomic hybridization (aCGH) and micro RNA arrays) was used to screen and identify copy number changes and differentially expressed miRNAs of 34 and 14 passages, respectively. Incubated cells used for xenografting (Passage 0) were considered to represent the primary tumor. Four important differentially expressed miRNAs (miR-31, miR-31*, miR-145, miR-106) were selected for further validation by real time polymerase chain reaction (RT-PCR). Integrated analysis of aCGH and miRNA data was performed on 14 xenograft passages by bioinformatic methods. Results The most frequent losses and gains of DNA copy number were detected at 9p21.3, 16q and at 8, 15, 17q21.32-qter, 1q21.1-qter, respectively. The presence of these alterations was consistent in all tumor passages. aCGH profiles of xenograft passages of each series resembled their corresponding primary tumors (passage 0). MiR-21, miR-31, miR-31*, miR-106b, miR-145, miR-150*, miR-371-5p, miR-557 and miR-598 showed recurrently altered expression. These miRNAS were predicted to regulate many ES-associated genes, such as genes of the IGF1 pathway, EWSR1, FLI1 and their fusion gene (EWS-FLI1). Twenty differentially expressed miRNAs were pinpointed in regions carrying altered copy numbers. Conclusion In the present study, ES xenografts were successfully applied for integrated microarray analyses. Our findings showed expression changes of miRNAs that were predicted to regulate many ES associated genes, such as IGF1 pathway genes, FLI1, EWSR1, and the EWS-FLI1 fusion genes.

Abstract Background Conventional high-grade osteosarcoma is a primary malignant bone tumor, which is most prevalent in adolescence. Survival rates of osteosarcoma patients have not improved significantly in the last 25 years. Aiming to increase this survival rate, a variety of model systems are used to study osteosarcomagenesis and to test new therapeutic agents. Such model systems are typically generated from an osteosarcoma primary tumor, but undergo many changes due to culturing or interactions with a different host species, which may result in differences in gene expression between primary tumor cells, and tumor cells from the model system. We aimed to investigate whether gene expression profiles of osteosarcoma cell lines and xenografts are still comparable to those of the primary tumor. Methods We performed genome-wide mRNA expression profiling on osteosarcoma biopsies (n = 76), cell lines (n = 13), and xenografts (n = 18). Osteosarcoma can be subdivided into several histological subtypes, of which osteoblastic, chondroblastic, and fibroblastic osteosarcoma are the most frequent ones. Using nearest shrunken centroids classification, we generated an expression signature that can predict the histological subtype of osteosarcoma biopsies. Results The expression signature, which consisted of 24 probes encoding for 22 genes, predicted the histological subtype of osteosarcoma biopsies with a misclassification error of 15%. Histological subtypes of the two osteosarcoma model systems, i.e. osteosarcoma cell lines and xenografts, were predicted with similar misclassification error rates (15% and 11%, respectively). Conclusions Based on the preservation of mRNA expression profiles that are characteristic for the histological subtype we propose that these model systems are representative for the primary tumor from which they are derived.

Abstract Background Aromatase (CYP19A1) regulates estrogen biosynthesis. Polymorphisms in CYP19A1 have been related to the pathogenesis of breast cancer (BC). Inhibition of aromatase with letrozole constitutes the best option for treating estrogen-dependent BC in postmenopausal women. We evaluate a series of polymorphisms of CYP19A1 and their effect on response to neoadjuvant letrozole in early BC. Methods We analyzed 95 consecutive postmenopausal women with stage II-III ER/PgR [+] BC treated with neoadjuvant letrozole. Response to treatment was measured by radiology at 4th month by World Health Organization (WHO) criteria. Three polymorphisms of CYP19A1, one in exon 7 (rs700519) and two in the 3'-UTR region (rs10046 and rs4646) were evaluated on DNA obtained from peripheral blood. Results Thirty-five women (36.8%) achieved a radiological response to letrozole. The histopathological and immunohistochemical parameters, including hormonal receptor status, were not associated with the response to letrozole. Only the genetic variants (AC/AA) of the rs4646 polymorphism were associated with poor response to letrozole (p = 0.03). Eighteen patients (18.9%) reported a progression of the disease. Those patients carrying the genetic variants (AC/AA) of rs4646 presented a lower progression-free survival than the patients homozygous for the reference variant (p = 0.0686). This effect was especially significant in the group of elderly patients not operated after letrozole induction (p = 0.009). Conclusions Our study reveals that the rs4646 polymorphism identifies a subgroup of stage II-III ER/PgR [+] BC patients with poor response to neoadjuvant letrozole and poor prognosis. Testing for the rs4646 polymorphism could be a useful tool in order to orientate the treatment in elderly BC patients.

Simple Summary: Solitary fibrous tumors (SFTs) are tumors that can vary greatly in how they behave, with some being harmless while others become aggressive and life-threatening. This study evaluated 52 fusion-confirmed SFTs with clinical follow-up. The aim was to analyze the immune intratumoral infiltrate and correlate the presence of specific immune cells and risk stratification systems to see if the presence or absence of certain cells could help predict how the tumor will behave or correlate with other clinicopathologic characteristics. The research found that, while certain immune cells were associated with specific tumor features with prognostic significance, these immune cells did not reliably predict the tumor's overall risk level. This suggests that the immune environment within SFTs is complex and may not be useful on its own for determining the tumor progression risk. Nonetheless, the presented results give an insight into the relationship between immune tumor cells and histologic characteristics such as necrosis, proliferation index, and neoplastic cell morphology. Background and objectives: Solitary fibrous tumors (SFTs) are morphologically heterogeneous tumors characterized by the NAB2::STAT6 gene fusion. Clinical outcomes may vary widely, and while most cases have favorable outcomes, some can progress to aggressive disease, manifesting as recurrence and metastasis, and ultimately resulting in patient death. Herein, we analyze the immunological tumor microenvironment (ITME) of SFTs, aiming to determine its prognostic value and correlation with established risk stratification systems (RSSs). Methods: A retrospective observational multicenter study of 52 fusion-confirmed SFTs with clinical follow-up data. Immunohistochemical analysis including CD163, CD68, CD3, CD8, CD20, PDL-1, PD-1, and LAG1 were evaluated in tissue microarrays, using an analog scale with scores ranging from 0 to 3 (0 = ≤9, 1 = 10–49, 2 = 50–99, and 3 = >100 positive cells per 10 high-power fields). The expression of these markers was correlated with clinical outcomes, morphological characteristics previously evaluated in whole slide tissue sections (hypercellularity/hypocellularity, round–oval or spindle dominant constituent cell (DCC) morphology, and necrosis), Ki67, overall survival, and RSS. Results: Only one of the fifty-two cases studied showed progression. In the multivariate analysis, neither the presence nor absence of immune cells (B-lymphocytes, T-lymphocytes, and macrophages) showed any association with the assessed RSSs (Demicco, Sugita, G-score, and Huang). Interestingly, the case that showed progression had high immune infiltrate with expression of CD68, CD163, CD8, and CD20 markers (score of 3). Round–oval cell morphology was associated with the presence of higher levels of CD163 macrophages. Lastly, the scant presence of CD20+ lymphocytes correlated with less necrosis, and cases with higher PDL-1 expression correlated with increased Ki67 values. All cases were negative for LAG-1 and PD-1. Conclusions: SFT ITME components correlated with independent variables with prognostic significance. Nevertheless, ITME did not correlate with RSS scores. [ABSTRACT FROM AUTHOR]

Synovial sarcoma (SS) is a rare soft-tissue tumor characterized by a monomorphic blue spindle cell histology and variable epithelial differentiation. Morphologically, SSs may be confused with other sarcomas. Systemic treatment is more effective for patients with high-risk SSs, patients with advanced disease, and younger patients. However, further studies are required to find new prognostic biomarkers. Herein, we describe the morphological, molecular, and clinical findings, using a wide immunohistochemical panel, of a series of SS cases. We studied 52 cases confirmed as SSs by morphological diagnosis and/or molecular studies. Clinical data (gender, age, tumor size, tumor location, resection margins, adjuvant treatment, recurrences, metastasis, and survival) were also retrieved for each patient. All the available H&E slides were examined by four pathologists. Three tissue microarrays (TMAs) were constructed for each of the tumors, and a wide immunohistochemical panel was performed. For time-to-event variables, survival analysis was performed using Kaplan–Meier curves and log-rank testing, or Cox regression. Statistical significance was considered at p < 0.05. The mean age of our patients was 40.33, and the median was 40.5 years. We found a predominance of males versus females (1.7:1). The most frequent morphological subtype was monophasic. TRPS1, SS18-SSX, and SSX-C-terminus were positive in 96% of cases. GLI1 expression was strong in six and focal (cytoplasmic) in twenty patients. Moreover, BCOR was expressed in more than half of SSs. Positive expression of both proteins, BCOR and GLI1, was correlated with a worse prognosis. Multivariate analysis was also performed, but only BCOR expression appeared to be significant. The combination of GLI1 and BCOR antibodies can be used to group SSs into three risk groups (low, intermediate, and high risk). We hypothesize that these findings could identify which patients would benefit from receiving adjuvant treatment and which would not. Moreover, these markers could represent therapeutic targets in advanced stages. However, further, larger series of SSs and molecular studies are necessary to corroborate our present findings. [ABSTRACT FROM AUTHOR]

The Ewing sarcoma family of tumors (EFT) is a group of highly malignant small round blue cell tumors occurring in children and young adults. We report here the largest genomic survey to date of 101 EFT (65 tumors and 36 cell lines). Using a combination of whole genome sequencing and targeted sequencing approaches, we discover that EFT has a very low mutational burden (0.15 mutations/Mb) but frequent deleterious mutations in the cohesin complex subunit STAG2 (21.5% tumors, 44.4% cell lines), homozygous deletion of CDKN2A (13.8% and 50%) and mutations of TP53 (6.2% and 71.9%). We additionally note an increased prevalence of the BRCA2 K3326X polymorphism in EFT patient samples (7.3%) compared to population data (OR 7.1, p = 0.006). Using whole transcriptome sequencing, we find that 11% of tumors pathologically diagnosed as EFT lack a typical EWSR1 fusion oncogene and that these tumors do not have a characteristic Ewing sarcoma gene expression signature. We identify samples harboring novel fusion genes including FUS-NCATc2 and CIC-FOXO4 that may represent distinct small round blue cell tumor variants. In an independent EFT tissue microarray cohort, we show that STAG2 loss as detected by immunohistochemistry may be associated with more advanced disease (p = 0.15) and a modest decrease in overall survival (p = 0.10). These results significantly advance our understanding of the genomic and molecular underpinnings of Ewing sarcoma and provide a foundation towards further efforts to improve diagnosis, prognosis, and precision therapeutics testing.