The c-myc and c-fos proto-oncogenes have several putative functions, including regulation of cell growth. In many neoplasms c-myc overexpression has been linked to poor prognosis. In order to study the role of c-myc and c-fos expression on the tumorigenesis, and the metastatic spread of osteosarcoma, frozen and paraffin-embedded tissue 38 primary osteosarcoma and 10 lung metastases were analyzed. The mRNA analysis was performed by quantitative reverse transcription-polymerase chain reaction and in situ hybridization. The protein expression was studied by Western blot analysis and immunohistochemistry. C-myc and c-fos were found overexpressed in a high percentage of the relapsed tumors and of the metastases, and overexpression of both oncogenes in the same tumor was strongly correlated to the development of metastases (p < 0.05), as 6 of the 7 primary tumors overexpressing both the oncogenes gave metastases. In conclusion, both c-myc and c-fos are involved in the growth and spread of osteosarcoma and a synchronous overexpression of both oncogenes is highly significant for a metastatic potential of a primary tumor.

No abstract

Our series confirms that CIC-DUX4 fusion-positive sarcomas are aggressive tumours with an adverse prognosis, and with clinical, histological and genetic differences from Ewing sarcoma. The best therapeutic approach needs to be investigated.

These prognostic markers that influence growth and spread of tumor cells might be useful to define tumor aggressiveness and risk of the metastasic event.

Cell‐cycle regulation depends on a fine balance between cyclin‐cyclin‐dependent kinase complexes and a family of kinase inhibitors that bind cyclin‐cdk complexes and block their activity. To investigate the role of mechanisms regulating cell‐cycle progression in human osteosarcomas (OS), pRb/p16/cdk4 expression was analyzed in 39 high‐grade OS; 19 of these developed metastasis during follow‐up. Positive reaction for functional pRB was shown by 18/39 (46%) OS, while 21/39 (54%) were negative. A higher probability of metastasis was seen in patients with negative pRb expression (p < 0.05). Furthermore, while functional pRb and D1 expression are inversely associated to metastasis occurrence, the presence of D1/cdk4 complex in our study was related to poor prognosis. We found that 10/18 pRb‐positive and 14/21 pRb‐negative tumors were p16‐positive. No significant correlation was found between pRb and p16 expression. On the other hand, high cdk4 levels in p16‐positive tumors as compared with p16‐negative tumors resulted in a positive association between p16 and cdk4 expression (Chi squared = 5.98; p = 0.01). No extensive p16INK4A genomic alterations were found in tumors lacking p16‐protein expression. To determine which mechanisms are involved in the down‐regulation of p16 protein, the methylation status of the p16INK4 gene was evaluated on the 15 p16‐negative tumors: 8 samples showed 5′ CpG‐island methylation; 4/8 had a complete methylation status, while in the remaining 4 the gene was only partially methylated. These data confirm the role of the pRb/p16/cdk4 pathway in OS development. Int. J. Cancer (Pred. Oncol.) 84:489–493, 1999. © 1999 Wiley‐Liss, Inc.

Musculoskeletal sarcomas are aggressive malignancies often characterized by an adverse prognosis despite the use of intense multiagent chemotherapy or molecular targeted therapy in combination to surgery and radiotherapy. Stem-like cells identified within solid tumors have been recently implicated in drug resistance, metastasis and local relapse. Here, we report the identification of putative cancer stem cells (CSCs) in sarcomas using a sphere culture system. These sarcospheres, able to grow in anchorage-independent and serum-starved conditions, express the pluripotent embryonic stem cell marker genes OCT3/4, Nanog and SOX2. Expression levels of these genes were greater in sarcospheres than in the parental tumor cultures. Importantly, the isolated tumor spheres transplanted into mice were tumorigenic and capable of recapitulating the human disease. Finally, we demonstrated that low (1%) O2 conditions, reproducing those found within the tumor microenvironment, significantly increase the number and the size of sarcospheres. The sphere formation assay is, therefore, a valuable method for the isolation of putative CSCs from human sarcomas and its efficiency is improved by controlling oxygen availability. This method provides a reliable preclinical model that can be used for future studies aimed at investigating crucial aspects of sarcoma biology, such as resistance to treatments and relapse.

No abstract

The Ewing's family of tumors (EFTs) are characterized by chimeric transcripts generated by specific chromosomal rearrangements. The most common fusions are between the EWSR1 gene on chromosome 22 and the ETS family of transcription factors; rarely, FUS (on chromosome 16) substitutes for EWSR1. The detection of specific translocations using molecular analysis is now a routine part of the pathological examination of EFT. Here, we report our experience with molecular diagnosis of EFT during the 4 years (2006 -2009) at the Rizzoli Institute. We analyzed 222 consecutive tumors with a presumptive diagnosis of EFT using molecular techniques and IHC. We found five distinct types of EWSR1-FLI1 fusion transcripts resulting from translocation t(11;22), three types of EWSR1-ERG transcripts resulting from t(21;22), and one type of t(2;22) resulting in EWSR1-FEV fusion. Molecular investigation validated 92% of cases ultimately diagnosed as EFT; IHC validated 76% of the cases. Thus, despite the difficulties and limitations associated with both molecular and IHC analysis on fresh and formalin-fixed, paraffin-embedded tissue, a combination of these techniques is the best approach to enhancing the accuracy of EFT diagnosis. We also present our method for choosing which molecular techniques to apply. Finally, we collected the most prevalent breakpoints reported in the literature, indicating which exons are involved, the sequence breakpoints, and the NCBI reference sequences. Ewing's sarcoma (ES) is a highly malignant small round cell tumor (SRCT) that can arise in both bone and soft tissues of children and young adults. Rarely, it occurs also in adults.1,2 It is the second most common pediatric bone tumor after osteosarcoma, accounting for ϳ30% of all primary bone tumors in this age group. 1,3,4 Based on their shared immunophenotypes and molecular parameters, several diagnostic entities that were previously considered distinct are now assembled as a single entity, the Ewing's sarcoma family of tumors (EFTs). These include bone and extraskeletal Ewing's sarcoma, primitive neuroectodermal tumor (PNET), and Askin tumor of the thoracopulmonary region. 1,5-7The histological evaluation of EFT is based on morphological, immunophenotypic, and molecular features. The morphology of ES is variable. These tumors are frequently composed of uniform small round cells with round nuclei containing fine chromatin, scanty clear or eosinophilic cytoplasm, and indistinct cytoplasmic membranes. In less differentiated cases, tumor cells are larger, with prominent nucleoli and irregular contours. The cytoplasm frequently contains PAS-positive diastase-resistant glycogen. PNET is characterized by HomerWright rosettes, which are typical of neuroectodermic differentiation. 6,8,9 Immunohistochemical analysis (IHC), the first ancillary technique used in pathology to confirm the diagnosis of EFT, can distinguish among different variants. The immunohistochemical features of ES are positivity for CD99, FLI1, and caveolin 1 antigens, whereas PNET also shows immun...