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Intimal sarcomas (IS) are rare malignant mesenchymal tumors arising in large blood vessels of the systemic and pulmonary circulation and also in the heart. They are morphologically similar to other spindle cell, poorly differentiated sarcomas. The prognosis is poor and depends mainly on surgical options. Three cases of IS were collected from two institutions. Clinical data were retrieved and histological study was performed. A wide immunohistochemical panel was analyzed. FISH of MDM2 gene was performed, and a molecular study with NGS was implemented in all cases. The mean age of our cases was 54 years. Histologically, the tumors presented a diffuse growth pattern with heterogeneous atypical epithelioid or spindle cells and extensive thrombosed areas. All cases presented intense immunoexpression for MDM2, CDK4, CD117, c-myc, PDGFRA, and p16. PDGFRA, HTERT, and pan-TRK gained expression, while p16 lost intensity, being weaker in both the local recurrences and xenografts. The three cases showed amplification of MDM2 by FISH. NGS analysis revealed amplifications in the CDK4, PDGFRA, and KIT genes, together with BRAF mutation and KRAS amplification. P16 was expressed in all cases, losing intensity in local recurrence and xenografts. Two new alterations, a BRAF mutation and a KRAS amplification, were detected by NGS in different tumors, opening up new therapeutic options for these patients.

Supplementary Figures S1-S5. MDM4 modulation by HEY1 occurs downstream of TP53 (S1); Modulation of SIRT1 and TP53 induction upon ectopic expression of Flag-tagged HEY1 in the B-cell malignancy cell lines "697" and NALM6, and in primary keratinocytes (S2); The degree of SIRT1 modulation depends on HEY1 levels (S3); Flow-diagram of samples analysed on tissue micro arrays (TMA1, TMA2, TMA3), and FFPE sections used in this study (S4); Kaplan-Meier survival estimates for patients analysed for SIRT1 expression on TMA3 (S5).

Supplementary Tables S1-S4. Identification of HEY1 regulated genes downstream of EWS-FLI1 silencing in Ewing sarcoma (S1); Combined immunohistochemical staining results for samples from patients with metastases at diagnosis on TMA1 and 2 according to site of metastasis (S2); Clinical characteristics and SIRT1 staining results of primary tumors from 43 ES patients treated at St. Jude Children´s Hospital between 1979 and 1987 using vincristine, doxorubicin, and cyclophosphamide-based regimens, with subsequent addition of ifosfamide and etoposide (S3); Materials used in the study (S4).

Supplementary Figure Legends. Legend for Supplementary Figures S1-S5.

The clinical evolution of solitary fibrous tumors (SFTs) is often uncertain and several risk stratification systems (RSS) have been proposed. The Demicco et al. RSS is the most frequently implemented. In this study we aim to validate two alternative RSS (Sugita et al. and G-Score) using results for the Demicco RSS from a previous study of 97 SFTs. In addition, we aim to determine whether reclassified cases had any distinctive molecular features. As the Sugita et al. system substitutes mitotic count with Ki-67 index we also investigated whether Ki-67 results for tissue microarrays are comparable to those obtained using whole tissue sections. In the present study we detected that many cases classified by Demicco RSS as low-risk were reclassified as intermediate risk using the new system (G-score RSS). Kaplan-Meier survival plots for G-Score RSS showed that the low-risk and intermediate-risk SFTs had a similar evolution that contrasted with the more aggressive high-risk group. Moreover, the similar evolution in both low and intermediate-risk groups occurred despite the G-score system being stricter in classifying low-risk tumors. We observed that Sugita RSS does not provide any better risk stratification in comparison with the Demicco RSS, and testing both RSS in our series produced similar Kaplan-Meier survival data. We found some discordant results when comparing whole sections and the corresponding tissue microarrays samples, finding the hotspot areas easier to locate in whole sections. Forty-one SFTs with initial low-risk assigned by the Demicco RSS were reclassified as intermediate-risk by G-score finding both TP53 and HTER mutations in four cases, only HTER mutation in 11 cases, and only TP53 mutation in 2 cases. All six cases of SFT classified as high-risk by both the Demicco and G-score RSS suffered recurrence/metastasis, and half showed both TP53 and HTER mutations. Five SFTs were categorized as low-risk by both Demicco and G-score, of which 4 cases revealed HTER mutation. Regarding the outcome of these 5 patients, two were lost to follow-up, and one of the remaining three patients suffered recurrence. We believe that although the presence of both TP53 and HTER mutations may confer or be related to poor evolution, the isolated presence of HTER mutation alone would not necessarily be related to poor outcome. The G-score RSS more accurately identified low-risk patients than the other two risk models evaluated in the present series. Late recurrence/metastasis may occasionally be observed even in low-risk SFTs categorized by stricter classification systems such as the G-score RSS. These findings support the possibility that additional, as yet unknown factors may influence the clinical evolution of SFTs. In conclusion, given the possibility of late recurrence, long-term follow-up is recommended for all SFT patients, even in cases classified as low risk by the stricter G-score system. An integration of clinical, radiological, pathological, and molecular findings may improve SFT risk stratification and better predict patient outcome.

Background Osteosarcoma is the most common primary malignant tumour of bone occurring in children and young adolescents and is characterised by complex genetic and epigenetic changes. The miRNA miR-486-5p has been shown to be downregulated in osteosarcoma and in cancer in general. Results To investigate if the mir-486 locus is epigenetically regulated, we integrated DNA methylation and miR-486-5p expression data using cohorts of osteosarcoma cell lines and patient samples. A CpG island in the promoter of the ANK1 host gene of mir-486 was shown to be highly methylated in osteosarcoma cell lines as determined by methylation-specific PCR and direct bisulfite sequencing. High methylation levels were seen for osteosarcoma patient samples, xenografts and cell lines based on quantitative methylation-specific PCR. 5-Aza-2′-deoxycytidine treatment of osteosarcoma cell lines caused induction of miR-486-5p and ANK1, indicating common epigenetic regulation in osteosarcoma cell lines. When overexpressed, miR-486-5p affected cell morphology. Conclusions miR-486-5p represents a highly cancer relevant, epigenetically regulated miRNA in osteosarcoma, and this knowledge contributes to the understanding of osteosarcoma biology.

Ewing sarcoma (EWS) is an aggressive bone and soft tissue tumor of children and young adults in which the principal driver is a fusion gene, EWSR1-FLI1. Although the essential role of EWSR1-FLI1 protein in the regulation of oncogenesis, survival, and tumor progression processes has been described in-depth, little is known about the regulation of chimeric fusion-gene expression. Here, we demonstrate that the active nuclear HDAC6 in EWS modulates the acetylation status of specificity protein 1 (SP1), consequently regulating the SP1/P300 activator complex binding to EWSR1 and EWSR1-FLI1 promoters. Selective inhibition of HDAC6 impairs binding of the activator complex SP1/P300, thereby inducing EWSR1-FLI1 downregulation and significantly reducing its oncogenic functions. In addition, sensitivity of EWS cell lines to HDAC6 inhibition is higher than other tumor or non-tumor cell lines. High expression of HDAC6 in primary EWS tumor samples from patients correlates with a poor prognosis in two independent series accounting 279 patients. Notably, a combination treatment of a selective HDAC6 and doxorubicin (a DNA damage agent used as a standard therapy of EWS patients) dramatically inhibits tumor growth in two EWS murine xenograft models. These results could lead to suitable and promising therapeutic alternatives for patients with EWS., Research in the E.D.A. lab is supported by Asociación Española Contra el Cáncer (AECC), the Ministry of Science of Spain-FEDER (CIBERONC, PI1700464, PI2000003, RD06/0020/0059)S. D.G.D. and L.H.P. are supported by CIBERONC (CB16/12/00361). D.G.D., M.J.R. and L.H.P. are PhD researchers funded by the Consejería de Salud, Junta de Andalucía (PI-0197-2016, ECAI F2-0012-2018 and PI-0013-2018, respectively).

Based on the French Federation Nationale des Centers de Lutte Contre le Cancer (FNCLCC) grading system, this study assesses the accuracy of conventional and modified core biopsy (CB) systems in predicting the final grade (low vs high) assigned to the resected specimen. Substituting Ki-67 immunoexpression for mitotic count, and radiological for histological assessment of necrosis, we used two modified FNCLCC CB grading systems: (1) Ki-67 immunoexpression alone, and (2) Ki-67 plus radiological assessment of necrosis. We graded 199 soft tissue sarcomas (STS) from nine centers, and compared the results for the conventional (obtained from local histopathology reports) and modified CB systems with the final FNCLCC grading of the corresponding resected specimens. Due to insufficient sample quality or lack of available radiologic data, five cases were not evaluated for Ki67 or radiological assessment of necrosis. The conventional FNCLCC CB grading system accurately identified 109 of the 130 high-grade cases (83.8%). The CB grading matched the final FNCLCC grading (low vs high) in 175 (87.9%) of the 199 resected tumors; overestimating the final grade in three cases and underestimating in 21 cases. Modified system 1 (Ki-67) accurately identified 117 of the 130 high-grade cases (90.0%). The CB grading matched the final FNCLCC grading (low vs high) in 175 (89.7%) of the 195 evaluated cases; overestimating seven and underestimating 13 cases. Modified system 2 (Ki-67 plus radiological necrosis) accurately identified 120 of the 130 high-grade cases (92.3%). This last matched the final FNCLCC grading (low vs high) in 177 (91.2%) of the 194 evaluated cases; overestimating seven and underestimating 10 cases. Modified system 2 obtained highest area under ROC curves, although not statistically significant. Underestimated CB grades did not correlate with histological subtypes, although many of the discrepant cases were myxoid tumors (myxofibrosarcomas or myxoid liposarcomas), leiomyosarcomas or undifferentiated pleomorphic/spindle cell sarcomas. Using modified FNCLCC CB grading systems to replace conventional mitotic count and histologic assessment of necrosis may improve the distinction between low and high-grade STS on CB. Our study confirms that classifying grade 1 as low grade and grades 2 and 3 as high grade improves correlation between CB and final grade by up to 21%, irrespective of CB system used. A higher than expected Ki-67 score in a low-grade sarcoma diagnosed on CB should raise concern that a higher-grade component may not have been sampled. Furthermore, correlation of all clinicopathological and radiological findings at multidisciplinary meetings is essential to assess the histological grade on CB as accurately as possible.

Osteosarcoma is the most common primary malignant tumour of bone occurring in children and young adolescents, and is characterised by complex genetic and epigenetic changes. The miRNA miR-486-5p has been shown to be downregulated in osteosarcoma and in cancer in general. To investigate if the miR-486 locus is epigenetically regulated, we integrated DNA methylation and miR-486-5p expression data using cohorts of osteosarcoma cell lines and patient samples. An upstream CpG island of mir-486 was shown to be highly methylated in osteosarcoma cell lines as determined by methylation-specific PCR and direct bisulfite sequencing. High methylation levels were seen for osteosarcoma patient samples, cell xenografts and cell lines based on quantitative methylation-specific PCR. 5-Aza-2’-deoxycytidine treatment of osteosarcoma cell lines caused induction of miR-486-5p in osteosarcoma cell lines. When overexpressed, miR-486-5p affected cell morphology. miR-486-5p represents a highly cancer relevant, epigenetically regulated miRNA in osteosarcoma.

Background Ewing sarcoma (EWS) is the second most common bone cancer in pediatric patients. Angiogenesis is a major factor for tumor growth and metastasis. Our aim was to carry out a histological, immunohistochemical, and molecular characterization of the neovascularization established between xenotransplanted tumors and the host during the initial phases of growth in nude mice in three angiogenesis experiments (ES2, ES3, and ES4). Methods The original human EWS were implanted subcutaneously on the backs of three nude mice. Tumor pieces 3 mm–4 mm in size from early passages of Nu432, Nu495, and Nu471 were also implanted subcutaneously on the backs of three sets (ES2, ES3, and ES4) of athymic Balb-c nude mice (n = 14 each). The animals were sacrificed at 24, 48, and 96 hours and at 7, 14, 21, and 28 days after implantation to perform histological, immunohistochemical, and molecular studies (neovascularization experiments). Results We observed histological, ultrastructural, and immunohistochemical changes in the xenografted tumor at different times after implantation. Chemokine ligand expression peaked twice, once during the first 48 hours and again in the second week. We observed that tumor cells in contact with murine peritumoral stroma presented higher expression of chemokine ligands as well as more tumor cells around the capillary vessels. Mouse serum vascular endothelial growth factor levels peaked twice, once in the first hours and then in the second week after tumor implantation. Conclusion Chemokines and other angiogenic factors may be relevant in the angiogenic mechanism during tumor growth. This model provides information on the early stages of the angiogenic process and could be a useful tool in researching anti-angiogenic drugs for new therapeutic strategies in EWS.

Sinonasal tract (SNT) alveolar rhabdomyosarcoma (ARMS) are frequently misdiagnosed, especially in adults. Fifty-two adult (≥18 years) patients with SNT ARMS were reviewed and characterized by immunohistochemistry and molecular studies. Twenty-six females and 26 males (18–72 years; mean 43.2 years), presented after a short duration (mean 2.6 months) with a large (mean 5.5 cm) destructive nasal cavity mass, involving multiple contiguous paranasal sites (n = 46) and with cervical adenopathy (n = 41). The tumors showed an alveolar, nested to solid growth pattern below an intact, but often involved (n = 9) epithelium with frequent necrosis (n = 37), destructive bone invasion (n = 30), and lymphovascular invasion (n = 25). The neoplastic cells were dyshesive and dilapidated, with crush artifacts. Rhabdoid features (n = 36) and tumor cell multinucleation (n = 28) were common. Mitotic counts were high (mean 17/10 HPFs). The neoplastic cells showed the following immunohistochemical positive findings: desmin (100%), myogenin (100%), MYOD1 (100%), MSA (96%), SMA (52%), CAM5.2 (50%), AE1/AE3 (36%); other positive markers included S100 protein (27%), CD56 (100%), synaptophysin (35%), and chromogranin (13%). Overall, 54% show epithelial marker reactivity. Molecular studies showed FOXO1 translocations (81%) with PCR demonstrating PAX3 in 72.7% tested. Patients presented with high stage (IV 24; III 26) and metastatic disease (lymph nodes n = 41; distant metastases n = 25) (IRSG grouping). Surgery (n = 16), radiation (n = 41) and chemotherapy (n = 45) yielded an overall survival of 36.1 months (mean; range 2.4–286); 18 alive without disease (mean 69.6 months); 7 alive with disease (mean 11.0 months); 1 dead without disease (63.7 months); and 26 dead with disease (mean 18.5 months). SNT ARMS frequently present in adults as a large, destructive midline mass of short symptom duration, with high stage disease. The alveolar to solid pattern of growth of cells with rhabdoid-plasmacytoid features suggests the diagnosis, but epithelial immunohistochemistry markers are present in 54% of cases, leading to misdiagnosis as carcinomas if muscle markers are not also performed. Overall survival of 36.1 months is achieved with multimodality therapy, but 64% have incurable disease (16.9 months). Mixed anatomic site (p = 0.02) was a significant adverse prognostic indicator, while stage (0.06) and tumor size >5 cm (0.06) approached marginal significance.

Background: Gastrointestinal stromal tumour (GIST) is the most common primary mesenchymal tumour of the gastrointestinal tract. Spindle cell monophasic synovial sarcoma (SS) can be morphologically similar. Angiogenesis is a major factor for tumour growth and metastasis. Our aim was to compare the angiogenic expression profiles of high-risk GIST and spindle cell monophasic SS by histological, immunohistochemical and molecular characterisation of the neovascularisation established between xenotransplanted tumours and the host during the initial phases of growth in nude mice. Methods: The angiogenic profile of two xenotransplanted human soft-tissue tumours were evaluated in 15 passages in nude mice using tissue microarrays (TMA). Tumour pieces were also implanted subcutaneously on the backs of 14 athymic Balb-c nude mice. The animals were sacrificed at 24, 48, and 96 h; and 7, 14, 21, and 28 days after implantation to perform histological, immunohistochemical, and molecular studies (neovascularisation experiments). Results: Morphological similarities were apparent in the early stages of neoplastic growth of these two soft-tissue tumours throughout the passages in nude mice and in the two neovascularisation experiments. Immunohistochemistry demonstrated overexpression of proangiogenic factors between 24 h and 96 h after xenotransplantation in both tumours. Additionally, neoplastic cells coexpressed chemokines (CXCL9, CXCL10, GRO, and CXCL12) and their receptors in both tumours. Molecular studies showed two expression profiles, revealing an early and a late phase in the angiogenic process. Conclusion: This model could provide information on the early stages of the angiogenic process in monophasic spindle cell SS and high-risk GIST and offers an excellent way to study possible tumour response to antiangiogenic drugs.

The developmental receptor NOTCH plays an important role in various human cancers as a consequence of oncogenic mutations. Here we describe a novel mechanism of NOTCH-induced tumor suppression involving modulation of the deacetylase SIRT1, providing a rationale for the use of SIRT1 inhibitors to treat cancers where this mechanism is inactivated because of SIRT1 overexpression. In Ewing sarcoma cells, NOTCH signaling is abrogated by the driver oncogene EWS-FLI1. Restoration of NOTCH signaling caused growth arrest due to activation of the NOTCH effector HEY1, directly suppressing SIRT1 and thereby activating p53. This mechanism of tumor suppression was validated in Ewing sarcoma cells, B-cell tumors, and human keratinocytes where NOTCH dysregulation has been implicated pathogenically. Notably, the SIRT1/2 inhibitor Tenovin-6 killed Ewing sarcoma cells in vitro and prohibited tumor growth and spread in an established xenograft model in zebrafish. Using immunohistochemistry to analyze primary tissue specimens, we found that high SIRT1 expression was associated with Ewing sarcoma metastasis and poor prognosis. Our findings suggest a mechanistic rationale for the use of SIRT1 inhibitors being developed to treat metastatic disease in patients with Ewing sarcoma. Cancer Res; 74(22); 6578–88. ©2014 AACR.

[Background] To report on long-term results of a phase 3 trial comparing three versus five cycles of adjuvant chemotherapy (CT) with full-dose epirubicin+ifosfamide in high-risk soft tissue sarcomas (STS)., [Methods] Patients (pts) were randomized to receive three preoperative cycles of epirubicin 120 mg/m2 and ifosfamide 9 g/m2 (Arm A) or to receive the same three preoperative cycles plus two postoperative cycles (Arm B). Radiotherapy could be either delivered in the preoperative or in the postoperative setting. Non-inferiority of the primary end point, OS, was assessed by the confidence interval of the hazard ratio (HR; Arm A/Arm B) derived from Cox model., [Results] Between January 2002 and April 2007, 164 pts were assigned to arm A and 164 to arm B. At a median follow-up (FU) of 117 months (IQ range 103–135 months), 123 deaths were recorded: 58 in Arm A and 65 in Arm B. Ten-year OS was 61% for the entire group of patients: 64% in Arm A and 59% in Arm B. The intention-to-treat analysis confirmed that three cycles were not inferior to five cycles (one-sided 95% upper confidence limit was 1.24). A per protocol analysis was consistent with these results. Pts with leiomyosarcoma and undifferentiated pleomorphic sarcoma (UPS) had the lowest, and the highest response rates, respectively. Consistently, Leiomyosarcoma and UPS had the worse and the best prognosis, respectively., [Conclusions] At a longer FU, the non-inferiority of three cycles of a full-dose conventional CT in comparison to five is confirmed. Response to therapy is also confirmed to be associated with better survival. This regimen is currently tested within an ongoing international trial against three cycles of a neoadjuvant histology-tailored CT (ClinicalTrials.gov Identifier: NCT01710176).

В останні роки в науковій пресі з'явилисm численні дані про молекулярну біологію раку молочної залози. У зв'язку з цим, гістологічна класифікація ВООЗ (2004 ) виявилась застарілою. Сьогодні більшість відомостей про прогноз, лікування та клінічний результат захворювання грунтується на результатах молекулярних і біологічних досліджень останніх років. Розглянуто сучасні підходи до питань гістопатологічної діагностики раку молочної залози на основі нових технологій. Обговорено рекомендації з імуногістохімічного тестування рецепторів естрогенів і прогестерону , а також рецепторів до епідермального фактора росту 2 (HERB2) у зразках пухлинної тканини. Запропоновано алгоритм оцінки отриманих даних, включаючи позитивні, спірні й негативні результати, що стосуються експресії білків HERB2 і ампліфікації гена. На основі характеру експресії генів, вивченого за технологією мікрочіпів з подальшою ієрархічної кластеризацією РНК нативної тканини, виділено нові генетичні профілі пухлини. Показано можливість застосування методів імуногістохімії в тканинах, залитих у парафін. Рекомендовано імуногістохімічну панель для діагностики наступних типів раку молочної залози: люмінальної А (Luminal А) , люмінальної В (Luminal В), базальноподібний і HER2/neu.Заключна частина роботи присвячена варіантам раку молочної залози з генетичною сприйнятливістю до BRCA1 і 2. Такі пухлини вкрай агресивні, характеризуються високим ступенем злоякісності і мають несприятливий прогноз. Проаналізовано нові підходи до терапії захворювання.

Chondrosarcomas are malignant cartilage-forming tumors that represent the third most common malignant solid tumor of bone. In patients with grades II and III, local recurrence, increasing tumor size and dedifferentiation have been associated with lower survival rates. These biologically poorly-understood neoplasms vary considerably in clinical presentation and biological behavior. Cytogenetic studies have shown that heterogeneity is related to karyotypic complexity; moreover, alterations in the 9p21 locus and TP53 gene are related to disease progression. Despite the relatively high frequency of chondrosarcoma only a limited number of cell lines exist in the scientific community, limiting the possibility to study hypothesis-derived research or primary drug interaction necessary for pre-clinical studies. We report a chondrosarcoma cell line, CH-3573, derived from a primary tumor that may serve as a useful tool for both in vitro and in vivo models to study the molecular pathogenesis. In addition, xenograft passages in nude mice were studied to characterize the genetic stability over the course of tumor progression. In contrary to other reported cell lines, an important feature of our established cell line was the retained matrix production, a characteristic feature of a conventional grade II chondrosarcoma. The cell line (CH-3573) was characterized by pathological, immunohistochemical and molecular genetic methods.

BACKGROUND: Human tumors transplanted into immunodeficient mice (xenografts) are good preclinical models, and it is important to identify possible systematic changes during establishment and passaging in mice. METHODS: High-resolution microarray-based comparative genomic hybridization (array CGH) was used to investigate how well a series of sarcoma xenografts, including 9 patient/xenograft pairs and 8 early versus late xenograft passage pairs, represented the patient tumor from which they originated. RESULTS: In all analyses, the xenografts were more similar to their tumor of origin than other xenografts of the same type. Most changes in aberration patterns were toward a more normal genome complement, and the increased aberrations observed were mostly toward more loss. In general, the changes were scattered over the genome, but some changes were significant in osteosarcomas. These were rather focused and consistent with amplifications frequent in patient samples, involving the genes platelet-derived growth factor receptor A (PDGFRA), cysteine-rich hydrophobic domain 2 (CHIC2), FIP-like 1 (FIP1L1), ligand of numb-protein X1 (LNX1), RAS-like family 11 member B (RASL11B), and sec1 family domain containing 2 (SCFD2), probably a sign of continued tumor progression. Some changes that disappeared may have been involved in host-stroma interactions or chemotherapy resistance, possibly because of the absence of selection in the mouse. CONCLUSIONS: Direct xenografts reflected well the genomic patterns of their tumors of origin. The few significant aberrations that were lost during passaging in immune-defective mice may have been caused by the lack of selection in the new host, whereas aberrations that were gained appeared to be the result of general tumor progression rather than model-specific artifacts. Cancer 2011;. © 2011 American Cancer Society.

Artículo publicado en: International Journal of Surgical Pathology published online 18 July 2010 The purpose of this study was to examine the immunoexpression of cell adhesion molecules (CAMs) E-cadherin, CD44s, and CD44v3 in cervical cancer and compare it with that in benign exo-endocervical tissue. In all, 81 cervical cancer biopsy specimens and 22 benign controls were included. Primary monoclonal antibodies NHC-38, F10-44-2, and 3G5 for E-cadherin, CD44s, and CD44v3 were used, respectively. Statistical significance was evaluated by the ?2 test. Antigen expression was significantly different in cervical cancer specimens compared with controls, showing marked decrease in membrane expression: E-cadherin, 6.5% and 77.3% (P < .000); CD44s, 3.9% and 81.8% (P < .000); and CD44v3, 0% and 81.8% (P < .000), respectively. The immunoexpression was significantly heterogeneous in carcinomas (P < .034) and adenocarcinomas (P < .000) for E-cadherin and CD44s. For CD44v3, no case of cancer showed immunostaining in membranes. These findings reaffirm that cell adhesion is markedly altered in cervical cancer. The authors suggest that these proteins could serve as markers for invasive cervical neoplasia. tmorelva@ula.ve antonio.llombart@uv.es

Background: To assess whether deletions involving codons 557 and/or 558 (critical deletions) of exon 11 of KIT are relevant in the prognosis of relapse-free survival (RFS) in gastrointestinal stromal tumor (GIST) patients with a long follow-up. Patients and methods: A univariate and multivariate analysis for RFS were carried out on 162 localized GIST patients over the entire follow-up period and over the intervals 0-4 years and >4 years. Factors assessed among others were Fletcher/National Institutes of Health and Miettinen-Lasota/Armed Forces Institute of Pathology (M-L/AFIP) risk categories, critical deletions and non-deletion-type mutation (NDTM) within exon 11 of KIT. Results: Multivariate analyses revealed that M-L/AFIP [relative risk (RR) 11.45, confidence interval (CI) 4.40-29.76, for the high-risk subgroup and RR 5.97, CI 2.09-17.06, for the intermediate subgroup] and critical deletions (RR 3.05, CI 1.59-5.85) were independent prognostic factors for RFS for the first 4 years and for the entire follow-up period. Beyond 4 years, the high-risk M-L/AFIP subgroup (RR 8.12, CI 1.48-44.4) and NDTM (RR 6.42, CI 1.17-35.12) were independent prognostic factors for RFS. The median follow-up was 84 months. Conclusion: Critical deletions represent a time-dependent prognostic factor limited to the first 4 years after surgery, which could help identify a subset with higher and earlier risk for relapse in GIST patients. © The Author 2010. Published by Oxford University Press.

Starting from an experimental model that accounts for the 2 most important adverse processes to successful therapy of Ewing's sarcoma (EWS), chemoresistance and the presence of metastasis at the time of diagnosis, we defined a molecular signature of potential prognostic value. Functional annotation of differentially regulated genes revealed 3 major networks related to cell cycle, cell-to-cell interactions and cellular development. The prognostic impact of 8 genes, representative of these 3 networks, was validated in 56 EWS patients. High mRNA expression levels of HINT1, IFITM2, LGALS3BP, STOML2 and c-MYC were associated with reduced risk to death and lower risk to develop metastasis. At multivariate analysis, LGALS3BP, a matricellular protein with a role in tumor progression and metastasis, was the most important predictor of event-free survival and overall survival. The association between LGALS3BP and prognosis was confirmed at protein level, when expression of the molecule was determined in tumor tissues but not in serum, indicating a role for the protein at local tumor microenvironment. Engineered enhancement of LGALS3BP expression in EWS cells resulted in inhibition of anchorage independent cell growth and reduction of cell migration and metastasis. Silencing of LGALS3BP expression reverted cell behavior with respect to in vitro parameters, thus providing further functional validation of genetic data obtained in clinical samples. Thus, we propose LGALS3BP as a novel reliable indicator of prognosis, and we offer genetic signatures to the scientific communities for cross-validation and meta-analysis, which are indispensable tools for a rare tumor such as EWS.

Background Neuroblastic tumors (NT) are pediatric neoplasms with a heterogeneous genetic profile. They present genotypic alterations of prognostic value, the study of which is mandatory in designing therapeutic management. Tissue microarrays (TMA) from paraffin material allow the analysis of a large number of cases with minimal costs. The main purpose of the present study is to analyze specific genetic markers of neuroblastic tumors included in TMAs and determine their prognostic value. We compare the results obtained by different molecular techniques at different substrates to evaluate the feasibility of these assays. Procedure One hundred thirty-nine samples were included in four different TMAs. We performed FISH assays to determine the status of MYCN gene, 1p36 region and 17q23 arm. The prognostic value of the genetic markers as well as the statistical correlation among clinical variables and outcome were analyzed by SPSS. Results MYCN amplification was detected in 35.3% of the cases, whereas 1p36 deletion and 17q23 gain was observed in 46.8% and 58.3% of the cases, respectively. An adverse prognosis was noted among these patients. Other adverse factors were age (>18 months) as well as high stage of disease (stage 4). Phenotypic signs of differentiation correlated with good outcome. Conclusion Retrospective studies using paraffin-embedded tissues assembled in TMA are a useful tool for the analysis of prognostic factors in NT. Pediatr Blood Cancer 2009;52:209–214. © 2008 Wiley-Liss, Inc.

We report two cases of malignant peripheral nerve sheath tumor (MPNST) in an uncommon location (heart and retropharynx) both with divergent osseous heterologous differentiation. We present the pathological and immunohistochemical studies that confirmed the neurogenic origin. The histopathology of the tumor arising in the retropharynx showed a transition from a neurofibroma to MPNST, making this a new report of an MPNST arising from a plexiform neurofibroma without neurofibromatosis. Primary cardiac MPNST with osseous differentiation has never been reported before. In conclusion, the histology of MPNSTs is very heterogeneous, showing no specific diagnostic immunoprofile or genetic alteration. Thus, it is important to rule out other histologically similar tumors, particularly in cases arising in uncommon locations or tumors with divergent heterologous differentiation.

Introducción: El sarcoma de estroma endometrial uterino es un tumor muy poco frecuente con una incidencia de 0,4 -3,4 por 100.000 mujeres, comprende menos del 1% de tumores malignos ginecológicos y el 2-5% de tumores malignos uterinos, siendo el tercer sarcoma en frecuencia después del carcinosarcoma y el leiomiosarcoma. Material y métodos: Presentamos cinco casos de sarcoma de estroma endometrial diagnosticados en un periodo de siete años con la evolución clínica de las pacientes. Resultados: Immunohistoquímicamente, estos tumores expresaban vimentina, receptores hormonales, CD10 y p53 (sarcomas de alto grado), siendo los marcadores musculares, la CK, el c-kit y ALK negativos. De estos cinco sarcomas dos de ellos eran de alto grado y tres de bajo grado (uno con diferenciación miogénica y otro combinado con áreas tipo «cordones sexuales», áreas miogénicas y áreas estromales puras). Discusión: Asimismo se plantean una serie de diagnósticos diferenciales con metodología práctica para poder caracterizar estos tumores. Introduction: Endometrial stromal sarcoma (ESS) is an infrequent neoplasm with an incidence of 0,4-3,4 cases/100.000 woman. This tumor represents less than 1% of the gynecologic malignancies as well as the 2,5% of the uterine malignant tumors being the third malignancy after carcinosarcoma and leiomyosarcoma. Patients and methods: We present five cases of endometrial sarcoma, diagnosed in a period of seven years. This study includes a complete immunohistochemical analysis as well as the follow-up of the patients. Results: Immunohistochemically, these tumors expressed vimentin, hormone receptors, CD10 and p53 (high grade sarcomas), whereas muscle markers, CK, c-Kit and ALK resulted negative. Two of the ESS were high grade tumors whereas the other three were considered low grade tumors (one with myogenic differentiation and the other combining sex cord areas, myogenic foci and pure stromal areas). Discussion: We discuss their differential diagnosis with other uterine malignant tumors. Martinez Rodriguez, Miguel, Miguel.Martinez-Rodriguez@uv.es ; Llombart Bosch, Antonio, Antonio.Llombart@uv.es ; Navarro Fos, Samuel, Samuel.Navarro@uv.es

Resumen El dermatofibrosarcoma protuberans (DFSP) es un tumor poco frecuente, de malignidad intermedia, con poca tendencia a desarrollar metastasis, pero con una alta frecuencia de recidiva local. Citogeneticamente, el DFSP se caracteriza por presentar la translocacion reciproca t(17;22) (q22;q13) que condiciona en la fusion del gen del colageno tipo Iα (COL1A1), en el cromosoma 17q, con el gen de la cadena β del factor de crecimiento derivado de las plaquetas (PDGFB), en el cromosoma 22q. La fusion de estos genes es variable, implicando a alguno de los 51 exones del gen COL1A1 con el exon 2 del gen PDGFB. Presentamos el caso de una mujer de 37 anos con una tumoracion en el brazo cuya histologia muestra una infiltracion neoplasica del tejido celular subcutaneo constituido por celulas fusiformes de nucleo elongado con un patron estoriforme y otras celulas mas pleomorficas con un patron en espina de pescado siendo compatible con DFSP con componente de fibrosarcoma. El estudio de biologia molecular con material incluido en parafina mediante transcripcion inversa y reaccion en cadena de la polimerasa (RT-PCR) y posterior secuenciacion muestra una nueva fusion del exon 19 del gen COL1A1 con el exon 2 de PDGFB, apoyando un diagnostico de DFSP. El estudio de los productos de fusion COL1A1-PDGFB es util en casos donde la histologia y la inmunohistoquimica son insuficientes para el diagnostico diferencial del DFSP con otros sarcomas y ademas, justifica la aplicacion de nuevas vias de tratamiento farmacologico con los inhibidores de la tirosincinasa.

El neuroblastoma presenta alteraciones genéticas que predicen su evolución clínica. Ganancias cromosómicas completas están asociadas a estadios clínicos no avanzados y evolución favorable, mientras que pérdidas de 1p, ganancia de 17q y amplificación del gen MYCN (MNA) son indicativas de estadios clínicos avanzados y pronóstico desfavorable. Son neuroblastomas de alto riesgo (NB-HR) los presentes en niños mayores de un año: estadio 4 o MNA en cualquier estadio de enfermedad, excluyendo estadio 1. El pronóstico de estos enfermos es malo, incluso con tratamientos agresivos. Sólo MNA confiere valor pronóstico negativo. Se remitieron al Centro de Referencia Nacional del neuroblastoma 60 casos de NB-HR. En todos analizamos MYCN y 1p36 con la técnica de hibridación in situ fluorescente (FISH) y en 24 de ellos el perfil genómico con la técnica de hibridación genómica comparada (CGH). Mediante FISH detectamos 31 casos MNA presentando 29 pérdida de 1p36; 21 casos no amplificados (MNNA) con pérdida de 1p36 en 7 casos; y 8 casos con ganancia del gen MYCN (MNG), seis con pérdida de 1p36. Mediante CGH detectamos otros reordenamientos cromosómicos. De 11 casos MNA ninguno presentó pérdida en 11q, todos presentaban di-somía del 17 o ganancia 17q. De 7 casos MNNA, cuatro mostraron pérdida de 11q, 2 con pérdida de 3p. Todos mostraron disomía del 17 o ganancia 17q. De 6 casos MNG, cuatro mostraron pérdida de 11q y cinco disomía del 17 o ganancia 17q. El estudio del perfil genómico en NB-HR revela la interacción de alteraciones genéticas cuyo significado pronóstico debe ser evaluado para establecer nuevos criterios terapéuticos. : Different subtypes of neuroblastoma (NB) carry associated genetic aberrations that predict their clinical course. Whole chromosome gains are usually associated with early clinical stages and good prognosis, while 1p deletion, 17q gain and MYCN amplification (MNA) are related to advanced stages and poor prognosis. High-risk neuroblastomas (NB-HR) include NB in children aged more than 1 year old, either stage 4 or any stages showing MNA except stage 1. The prognosis of NB-HR patients remains poor, despite aggressive therapy. Only MNA confers poor prognosis. Between January 2000 and February 2005, tumoral specimens from 60 patients with NB-HR were sent to the Spanish Reference Center for NB biological studies. In all cases, MYCN together with 1p36 status was analyzed by fluorescence in situ hybridization (FISH). Comparative genomic hybridization (CGH) was performed in 24 cases. Using FISH we detected 31 MNA cases including 29 with 1p36 deletion; there were 21 cases without MYCN amplification (MNNA) but 7 of these had 1p36 deletion; 8 cases showed MYCN gain (MNG) but 6 of these had 1p36 deletion. CGH showed other chromosomal alterations. Of 11 MNA cases, none had 11q loss and all of them showed 17q gain or 17 disomy. Of the 7 MNNA cases, there were 4 with 11q loss including 2 with 3p loss and all presented 17q gain or 17 disomy. The 6 MNG cases included 4 cases with 11q loss and 5 cases with 17q gain or 17 disomy. Genomic profiling by CGH in NB-HR confirms the interaction among genetic alterations, the prognostic significance of which should be evaluated to establish new treatment criteria.

When designing infrastructure for a networked virtual tumour bank (samples remain at the collector institutes and sample data are collected in a searchable central database), it is apparent that this can only function properly after developing an adequate set of rules for use and access. These rules must include sufficient incentives for the tissue sample collectors to remain active within the network and maintain sufficient sample levels in the local bank. These requirements resulted in a key TuBaFrost rule, stating that the custodianship of the samples remains under the authority of the local collector. As a consequence, the samples and the decision to issue the samples to a requestor are not transferred to a large organisation but instead remain with the collector, thus allowing autonomous negotiation between collector and requestor, potential co-authorship in publications or compensation for collection and processing costs. Furthermore, it realises a streamlined cost effective network, ensuring tissue visibility and accessibility thereby improving the availability of large amounts of samples of highly specific or rare tumour types as well as providing contact opportunities for collaboration between scientists with cutting edge technology and tissue collectors. With this general purpose in mind, the rules and responsibilities for collectors, requestors and central office were generated.

We report, for the first time, mutations in the Alu repeat regions in the genome of kidney tumors induced by diethylstilbestrol in Syrian hamsters. Among the 66 loci amplified by 11 random primers, 28 loci exhibited insertions, deletions or losses or gains in intensity in the genome of kidney tumor tissues compared with normal kidney tissues from age-matched hamsters. Higher numbers of mutated Alu loci were observed in the tumors of old hamsters compared with young hamsters. In N-ethyl-N-nitrosourea- and diethylstilbestrol-treated hamsters deletion of a 0.59 kb locus amplified with primer OPC03 was observed in most of the female hamsters, but not in male hamsters. An insertion mutation of a 0.498 kb locus amplified with primer OPC03 was observed in 12 of 36 diethylstilbestrol-induced kidney tumors. The cloning and sequencing of the 0.498 kb locus amplified with primer OPC03 revealed that it had significant sequence similarity to the mouse RIKEN cDNA clone. These findings indicate that age, sex and co-exposure to N-ethyl-N-nitrosourea influence mutations in the Alu repeat sequences in the genome of diethylstilbestrol-induced kidney tumors in Syrian hamsters. Structural alterations in Alu repeats in critical target genes may be involved in diethylstilbestrol-induced carcinogenesis.

Introduccion . La invasion de vasos linfaticos por melanocitos de nevos melanociticos es un hallazgo infrecuente. Por otro lado, la presencia de melanocitos en los ganglios linfaticos se detecta de forma casual en linfadenectomias regionales por carcinomas y melanomas. Metodos . Hemos estudiado 8 casos de nevos melanociticos cutaneos con invasion de vasos linfaticos, analizando sus caracteristicas clinicas e histologicas, asi como la inmunotincion para Ki-67. Resultados . Siete de los 8 casos correspondian a mujeres; cinco fueron diagnosticados como nevos congenitos, observandose en estos ultimos un crecimiento solido melanocitico intravascular con integridad de la superficie endotelial. En los 3 casos restantes, los vasos linfaticos con melanocitos en su luz se encontraban distantes al nevo, y los melanocitos adoptaban un patron mas irregular en agregados ocasionalmente adheridos a la superficie interna endotelial. Uno de los casos mostro los dos patrones. En ningun caso los melanocitos intravasculares fueron positivos para Ki-67. Conclusiones . La presencia de vasos linfaticos con melanocitos intraluminales es mas frecuente en nevos congenitos y en particular en mujeres. Se distinguen dos patrones bien definidos de invasion: crecimiento polipoide intravascular y agregados melanociticos irregulares ocasionalmente adheridos a la superficie endotelial. El primero es el habitual en los nevos congenitos y el segundo en los adquiridos. La ausencia de inmunotincion para Ki-67 apoya el caracter no agresivo de la poblacion intravascular.

Chondrosarcomas are malignant cartilage-forming tumors that represent the second most common malignant solid tumor of bone. These biologically poorly understood neoplasms vary considerably in clinical presentation and biologic behavior. Chemotherapy and radiation therapy are generally ineffective. Here we describe the establishment and characterization of a new human chondrosarcoma cell line named ch-2879, and we compare the cell line with its tumor of origin. The cell line was established from a recurrent grade 3 chondrosarcoma of the chest wall and characterized by growth kinetics and morphologic studies. Immunocytochemistry and RT-PCR were performed to examine the expression of cartilage-specific phenotypes. Genetic characterization was performed using cytogenetics, fluorescence in situ hybridization, flow cytometry, and molecular techniques for analysis of the genes implicated in cell cycle control, amplification of MDM2, CDK4, and Cyclin D1, and mutations in the p53 gene. ch-2879 cells were subcultured for more than 80 passages. They expressed vimentin, HNK-1, HBA-71, Ki-67, cyclin D1, Fli-1, S-100, p21, p27, and p53 and were negative for cytokeratin, EMA, p14, p16, MDM2, Rb, and c-erb-b2 antigens. Cytogenetically the recurrent tumor showed a hyperhaploid karyotype with clonal numerical and structural abnormalities. The sole structural abnormality was a chromosome derivative of a t(1;21) translocation. The cell line at passage 3 showed two populations: the hyperhaploid and an exactly duplicated, hypotriploid population. After the 18th passage, only the hypotriploid population was present. The cells expressed collagen 2. Molecular comparison of the primary and recurrent tumor evidenced an in vivo molecular change consisting of a deletion of 9p21 genes in the recurrence, probably caused by a selection process. Because of its gene expression profile, including expression of genes implicated in chondrogenesis in uncoated plastic dishes, this cell line may prove useful for cellular and molecular studies as well as studies of chondrosarcoma characterization and treatment.

Increased angiogenesis is related to boosted growth and malignancy in carcinomas. "Chronic Persistent Low-Dose Ionizing Radiation" (CPLDIR) exposure increases incidence and aggressive behavior of clear-cell renal-cell carcinoma (CCRCC). The aim was to study the biology of angiogenesis, including microvessel density (MVD), in human clear-cell renal-cell carcinomas (CCRCC) originating from a radio-contaminated geographical area (Ukraine) and to compare with similar tumors diagnosed in non-contaminated regions of Europe (Spain, Valencia) and Latin America (Colombia, Barranquilla). MVD was comparatively examined in 124 patients diagnosed with CCRCC from three geographical areas by means of digital micro-imaging and computerized analysis. Additionally, 50 adult normal kidneys were used for controls (autopsy kidneys from Valencia and Barranquilla). Furthermore, an immunohistochemical study of several vascular related growth factors was undertaken using a similar methodology. MVD as well as VEFG are the most discriminating factors associated with an aggressive behavior of CCRCC. Their expression increased in proportion to the level of exposure to chronic low-dose ionizing radiation in Ukrainian patients in the 25 years since the Chernobyl accident substantiated by comparison with the two control groups of renal carcinomas present in non-irradiated areas (Spain and Colombia). No major biological differences relating to angiogenesis appear to exist between the CCRCC diagnosed in two distant geographical areas of the world. HIF-1α expression was similar in all groups, with no statistical significance. Present findings demonstrate the existence of a significant relationship between MVD and VEGF in CCRCC: an increased expression of VEGF is associated with a high level of angiogenesis.

Rhabdomyosarcomas (RMS) may resemble other non-myogenic sarcomas and malignant rhabdoid tumor (MRT). Alveolar rhabdomyosarcoma (ARMS) often harbors a typical translocation, but embryonal rhabdomyosarcoma (ERMS) lacks any specific rearrangement. Histopathology is not always sufficient for an unequivocal diagnosis, necessitating ancillary studies, including immunohistochemistry (IHC). Sixteen genetically tested RMS and two MRT were xenografted and followed in successive passages. Tissue microarrays were constructed including samples from original and xenograft tumors. Desmin, myogenin, CK, EMA, INI1, LSD1, AP2 beta, fibrillin-2, HMGA2, nestin, and SIRT1 were tested using immunohistochemical staining. Desmin and myogenin were positive in all RMS, and the epithelial markers were negative in almost all RMS. New markers (LSD1, AP2 beta, HMGA2, Nestin, and SIRT1) were positive in all RMS and MRT. There were no differences in IHC expression between the three RMS subtypes tested except fibrillin-2, which was negative in ARMS. Applying new IHC markers can contribute to RMS diagnosis. Nevertheless, most markers are also expressed in MRT, and further studies are needed to confirm their value against this and other small round cell tumors.

Ductal carcinoma in situ (DCIS) is a heterogeneous group of lesions that has been subdivided into three types: well differentiated (grade I), moderately differentiated (grade II), and poorly differentiated (grade III). Forty-five cases of DCIS were analyzed for image analysis: nuclear area, DNA ploidy, and vascularization in order to establish a more precise correlation between the histologic grade and these morphometric parameters. Our results confirm that the mean nuclear area, DNA ploidy, and microvessel density (MVD) progressively increased from DCIS grade I to DCIS grade III. The analysis of the nuclear area in relationship to DCIS grading demonstrated a progressive increase of values between grades I/II to grade III, but these data have no statistical significance. An analysis of DNA ploidy demonstrated significant differences between grades I/III (p0.05), but there was no statistical significance between grades I/II, grades II/III, or both (p0.005). The analysis of MVD was extremely significant between grades I/III (p0.001) and grades II/III (p0.001), but between grades I/II, these values showed no significant differences (p0.05). Based on this study, it can be concluded that image analysis techniques confirm how DCIS presents morphometric values that increase from DCIS grade I to DCIS grade III and that within this spectrum, DCIS grade III can be identified as a group of tumors presenting a large nuclear area, aneuploid DNA, and abundant vascular neogenesis, confirming that this neoplasm displays more aggressive patterns than the other two types. These criteria should justify a higher rate of tumor progression to DCIS grade III.

Atypical fibroxanthoma (AFX), a pleomorphic superficial cutaneous tumor of low-grade malignancy, shares many morphologic features with malignant melanoma (MM) and squamous cell carcinoma (SCC). Absence of S-100, keratin, and desmin immunoreactivity is the clue for this diagnosis. In a search for positive markers, we tested 26 cases of AFX with 2 antibodies: O13 (CD99) and protein gene product 9.5 (PGP9.5). We also included 10 cases of poorly differentiated SCC and 10 cases of MM in the study. In AFX, CD99 immunoreactivity was present in 19 cases (73%), whereas focal PGP9.5 immunoreactivity was found in only 9 cases (35%). None of the SCC cases showed CD99 immunostaining. No CD99 immunoreactivity was found in 9 of 10 cases of MM. To our knowledge, this is the first report of CD99 and PGP9.5 immunostaining in AFX. We believe, based on our results, that CD99 may be a helpful "positive" feature in the histopathologic diagnosis of AFX.

Driven by genomic somatic variation, tumour tissues are typically heterogeneous, yet unbiased quantitative methods are rarely used to analyse heterogeneity at the protein level. Motivated by this problem, we developed automated image segmentation of images of multiple biomarkers in Ewing sarcoma to generate distributions of biomarkers between and within tumour cells. We further integrate high dimensional data with patient clinical outcomes utilising random survival forest (RSF) machine learning. Using material from cohorts of genetically diagnosed Ewing sarcoma with EWSR1 chromosomal translocations, confocal images of tissue microarrays were segmented with level sets and watershed algorithms. Each cell nucleus and cytoplasm were identified in relation to DAPI and CD99, respectively, and protein biomarkers (e.g. Ki67, pS6, Foxo3a, EGR1, MAPK) localised relative to nuclear and cytoplasmic regions of each cell in order to generate image feature distributions. The image distribution features were analysed with RSF in relation to known overall patient survival from three separate cohorts (185 informative cases). Variation in pre-analytical processing resulted in elimination of a high number of non-informative images that had poor DAPI localisation or biomarker preservation (67 cases, 36%). The distribution of image features for biomarkers in the remaining high quality material (118 cases, 104 features per case) were analysed by RSF with feature selection, and performance assessed using internal cross-validation, rather than a separate validation cohort. A prognostic classifier for Ewing sarcoma with low cross-validation error rates (0.36) was comprised of multiple features, including the Ki67 proliferative marker and a sub-population of cells with low cytoplasmic/nuclear ratio of CD99. Through elimination of bias, the evaluation of high-dimensionality biomarker distribution within cell populations of a tumour using random forest analysis in quality controlled tumour material could be achieved. Such an automated and integrated methodology has potential application in the identification of prognostic classifiers based on tumour cell heterogeneity.

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., Author Summary The Ewing sarcoma family of tumors is a group of aggressive cancers that primarily affects the pediatric and young adult population. Increasingly, genomics are being used to better define the disease biology and to identify targets for therapy in many cancer types. Here, we report one of the first and largest genomic studies to date in the Ewing sarcoma family of tumors. Using a combination of modern sequencing techniques in >100 samples, we discover that Ewing sarcomas have a genome that is less complex compared to most cancer types previously surveyed. We find that this cancer is frequently affected by mutations in STAG2, a gene that has recently gained attention due to its importance in the biology of several cancer types. We show that Ewing sarcoma patients whose tumors are affected by STAG2 loss may have a worse prognosis. Additionally, we identify a subset of tumors that were diagnosed as Ewing sarcoma that appear to be distinct from the majority based on genetic and molecular characteristics. Our findings help to define the genetic landscape of Ewing sarcoma and provide a starting point for improving individualization of diagnosis, prognosis and treatment in this cancer.

During the 13-year period subsequent to the Chernobyl accident, the morbidity of malignant renal tumors in Ukraine has increased from 4.7 to 7.5 per 100,000 of total population. Cesium 137 ( 137 Cs) accounts for 90% of the incorporated radioactivity in the Ukrainian population, which has been exposed to long-term, low dose ionizing radiation and 90% of the more labile pool of ( 137 Cs) is excreted via kidneys. The present study was performed to evaluate the histopathological features and the immunohistochemical status of proliferating cell nuclear antigen (PCNA) and K-ras in renal cell carcinomas (RCCs) of 236 Ukrainian patients (groups I to V), which represents a varying degrees of internal exposure to radiation and were operated in 2 different periods of time after the Chernobyl accident. The control group VI of 112 analog patients with RCCs was selected in Spain. The strong significant differences between the Ukrainian and Spanish groups were found in tumoral nuclear grade, in the percentage of sarcomatoid changes, the level of the peritumoral inflammatory response as well as in the peritumoral lesions. The dramatic increase of aggressivity and proliferative activity supported by strong PCNA and K-ras expression of RCCs from Ukrainian groups, associated with chronic radiation nephropathy of peritumoral kidney tissue, showed good correlation with the duration of radiation exposure and confirmed the influence of chronic but regular and sustained low dose of ionizing radiation on renal carcinogenesis of the Ukrainian population. Int. J. Cancer 87:880 ‐ 883, 2000. © 2000 Wiley-Liss, Inc.

The aim of the present study was to explore ERG immunoreactivity in a series of sarcomas, GIST and malignant rhabdoid tumor (MRT), considering the not fully elucidated specificity and sensitivity of this antibody. Paraffin-embedded tissue microarrays from those tumors were stained with anti-ERG against the C-terminus [(EPR3864(2)] and N-terminus (Clone 9FY). EPR3864(2) was positive in almost all angiosarcomas, and MRT.GIST were positive in a large proportion of cases (38.4%), and more than half the synovial sarcomas (52.7%) revealed EPR3864(2) staining. Several chondrosarcomas, osteosarcomas, rhabdomyosarcoma and Ewing's sarcoma family of tumors (ESFT) presented EPR3864(2) expression in a lower number of cases. 9FY was positive in most of the angiosarcomas; however, only sporadic ESFT and synovial sarcoma were positive and the other tumors tested were negative. Fourteen ESFT with EWSR1/Fli-1 gene fusion presented positive nuclear staining for EPR3864(2). Similarly, 5 ESFT with EWSR1/Fli-1 gene fusion presented positive staining for 9FY. We must stress that the difference between the present and previous studies may be due to the source of the anti-ERG employed, anti-ERG against C or N-terminus, protein cross-reactivity and dilution. In conclusion, specificity for ERG staining in sarcomas should be considered with caution and the immunoexpression is undoubtedly influenced by clone and antibody selection. (C) 2014 Elsevier GmbH. All rights reserved.

Contribution over time of human papillomavirus (HPV) types in human cancers has been poorly documented. Such data is fundamental to measure current HPV vaccines impact in the years to come. We estimated the HPV type-specific distribution in a large international series of invasive cervical cancer (ICC) over 70 years prior to vaccination. Paraffin embedded ICC cases diagnosed between 1940 and 2007 were retrieved from eleven countries in Central-South America, Asia and Europe. Included countries reported to have low-medium cervical cancer screening uptake. Information on age at and year of diagnosis was collected from medical records. After histological confirmation, HPV DNA detection was performed by SPF-10/DEIA/LiPA25 (version1). Logistic regression models were used for estimating the adjusted relative contributions (RC) of HPV16 and of HPV18 over time. Among 4,771 HPV DNA positive ICC cases, HPV16 and HPV18 were the two most common HPVs in all the decades with no statistically significant variations of their adjusted-RC from 1940–59 to 2000–07 (HPV16—from 61.5 to 62.1%, and HPV18—from 6.9 to 7.2%). As well, the RC of other HPV types did not varied over time. In the stratified analysis by histology, HPV16 adjusted-RC significantly increased across decades in adenocarcinomas. Regarding age, cases associated to either HPV16, 18 or 45 were younger than those with other HPV types in all the evaluated decades. The observed stability on the HPV type distribution predicts a high and stable impact of HPV vaccination in reducing the cervical cancer burden in future vaccinated generations.

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.

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.