Cell death induction by the BH3 mimetic GX15-070 in thyroid carcinoma cells

Background

The evasion of cell death is one of the hallmarks of cancer, contributing to both tumor progression and resistance to therapy. Dedifferentiated and anaplastic thyroid carcinomas that do not take up radioiodine are resistant to conventional anticancer treatments and patients with these tumors are difficult to treat. BH3 mimetics are a new class of drugs that target anti-apoptotic proteins of the BCL-2 family and promote cell death. The purpose of this study was to analyze the molecular effects of the BH3 mimetic GX15-070 on thyroid carcinoma cell lines and to characterize cell death induced by GX15-070.

Methods

A total of 17 cell lines derived from follicular, papillary, and anaplastic thyroid carcinomas were treated with GX15-070. Cell viability was measured with MTT assay while cell cycle phase distribution and subG1 peaks were determined after propidium iodide staining. We assessed cell death via the caspase 3/7 activity, caspase cleavage products, lactate dehydrogenase (LDH) liberation assays, and a LC3 analysis by western blot. Ultrastructural changes were analysed by electron microscopy of GX15-070-treated cells.

Results

After GX15-070 treatment, the number of viable cells was decreased in all cell lines examined, with IC50 values ranging from 48nM to 3.25 μM. We observed biochemical markers of autophagic cell death and necrosis like LC3 conversion and LDH release after the GX15-070 treatment. Electron microscopy revealed several common characteristic ultrastructural changes like swelling of mitochondria, dilatation of rough endoplasmic reticulum, membrane blebbing and formation of vacuoles. GX15-070 treatment induced DNA fragmentation detected by subG1-peak induction and an arrest in G1 phase of the cell cycle. Caspase activation after GX15-070 incubation was detected but had no effect on viability of cells.

Conclusions

With these experiments we demonstrated the efficacy of the BH3 mimetic drug GX15-070 acting against dedifferentiated thyroid carcinoma cells of various histological origins by the induction of cell death. GX15-070-treated cells underwent non-classical cell death with signs of apoptosis, autophagy and necrosis in parallel. GX15-07 and related compounds thus may be a new therapeutic option for dedifferentiated thyroid carcinoma of various histological subtypes.     

Dedifferentiated chondrosarcoma mimicking a giant cell tumor. Is this low grade dedifferentiated chondrosarcoma?

We report a very rare case of a dedifferentiated chondrosarcoma mimicking a benign giant cell tumor. A 22-year-old male was admitted to our hospital with a history of mild left wrist pain after a skiing trauma. Radiology revealed an extensive meta-epiphyseal osteolytic lesion in the distal ulna, which appeared to be a giant cell tumor. Histological examination showed a biphasic tumor comprising chondroid and non-chondroid areas with a giant cell-rich lesion resembling a conventional giant cell tumor of the bone. Immunohistochemistry showed no expression of p16^INK4a, VEGFR1, KDR (VEGFR2), VEGFR3, cKIT, MDM2 or CDK4. However, high expression of the tyrosine kinases PDGFRA and PDGFRB was observed. Molecular analysis showed no amplification of the cMYC gene and no activating mutations in the cKIT (exons 9 and 11) or PDGFRA (exon 18) genes.     

Dedifferentiated gastrointestinal stromal tumor arising de novo from the small intestine

Gastrointestinal stromal tumors (GISTs) are the most common mesenchymal tumors of the gastrointestinal tract and usually display monotonous cytologic features and immunoactivity for CD117. Anaplastic GIST, with pleomorphic cells and loss of CD117, until recently have only been reported in patients with chronic imatinib mesylate treatment. Dedifferentiated GISTs arising de novo is a newly identified entity that may prove to be difficult to diagnose. We present the case of a 52-year-old female found to have a dedifferentiated GIST without prior imatinib mesylate therapy. This case is the first reported dedifferentiated GIST arising de novo from the small bowel, and at 30 cm in greatest diameter, the largest reported to date. Additionally, we demonstrate for the first time the loss of DOG1 in the anaplastic component of the tumor. De novo dedifferentiated GIST is a rare and diagnostically challenging tumor that may be mischaracterized unless considered in the differential diagnosis.     

The molecular pathogenesis of dedifferentiated chondrosarcoma

Dedifferentiated chondrosarcomas are cartilaginous tumors that consist of two distinguishable components, a lowgrade chondrosarcoma (chondrogenic) component and a highgrade dedifferentiated (anaplastic) component. The tumor cells in both components seem to originate from a single precursor, but there are a substantial number of genetic alterations in the anaplastic component. The underlying mechanism of dedifferentiation is unknown, but cell cycle regulators p16, p53 and retinoblastoma appear to have important roles in tumor development and dedifferentiation. In this article, molecular pathogenesis of dedifferentiated chondrosarcomas is reviewed.     

Ezrin immunohistochemical expression in cartilaginous tumours: a useful tool for differential diagnosis between chondroblastic osteosarcoma and chondrosarcoma

Ezrin is a cytoskeleton linker protein that is actively involved in the metastatic process of cancer cells. We have recently reported that ezrin expression in conventional osteosarcoma was an independent prognostic factor for event-free survival and overall survival. In this work, ezrin expression was found in all histological subtypes. Especially cartilaginous areas in chondroblastic osteosarcomas were immunopositive for ezrin. We wanted to know if ezrin could be a useful diagnostic marker in bone pathology. We have searched for ezrin expression in 208 cartilaginous tumours by immunohistochemistry and in 16 chondroblastic osteosarcomas. All conventional chondrosarcomas, whatever their grade, were negative, while ten of 16 chondroblastic osteosarcomas were positive. In contrast, dedifferentiated (five of 14) and mesenchymal chondrosarcomas (five of ten) showed ezrin positivity. Some chondroblastomas and more rarely chondromyxoid fibromas also exhibited ezrin expression. These data suggest that ezrin is a useful immunohistochemical marker for differential diagnosis between chondroblastic osteosarcomas and conventional chondrosarcomas with a specificity of 100%. Ezrin expression in dedifferentiated and mesenchymal chondrosarcomas which are aggressive neoplasms resistant to conventional treatment means that ezrin could be a therapeutic target. Ezrin expression in chondroblastomas is more intriguing and requires further study to assess prognostic value.     

Dedifferentiated liposarcoma with peculiar meningothelial-like whorling and metaplastic bone formation

Dedifferentiated liposarcoma with peculiar meningothelial-like whorling pattern and metaplastic bone formation (DDLMB) is an unusual morphologic entity that is characterized by an atypical lipomatous tumor/well-differentiated liposarcoma with epithelioid or spindle cells concentrically arranged into meningothelial-like “whorls,” and mature bone trabeculae rimmed by reactive osteoblasts. We recently experienced 2 cases of DDLMB, one in a 64-year–old male patient with a painless right groin mass and another in a 42-year–old female patient with a painless right abdominal mass. The size of the tumors was 3.5 and 18 cm; and the tumors were located in the right scrotal sac and retroperitoneum in case 1 and case 2, respectively. Under the initial clinical diagnosis of cord lipoma in case 1 and high-grade sarcoma in case 2, the masses were removed. The cut surfaces of the masses were well circumscribed with encapsulation, red-tan, firm, and multinodular. Microscopically, the tumors consisted of atypical lipomatous tumor/well-differentiated liposarcoma with meningothelial-like whorls and metaplastic bone formation in both cases. In addition, the first case showed focal areas of paraganglioma-like pattern; and the second case showed pleomorphic high-grade sarcoma with low-grade myxofibrosarcoma-like areas. Immunohistochemically, the tumor components with meningothelial-like pattern and paraganglioma-like pattern in DDLMB were positive for vimentin and CD56 and negative for pancytokeratin, epithelial membrane antigen (EMA), desmin, and smooth muscle actin. Characteristically, the paraganglioma-like area was immunoreactive for S-100 protein, with a “dot-like” staining pattern. The patient additionally underwent radical orchiectomy in case 1. Review of the literature revealed that only 34 cases of DDLMB have been reported. One case of dedifferentiated liposarcoma with a predominant paraganglioma-like pattern has also been reported in the literature. To our knowledge, case 1 represents the first report of DDLMB with paraganglioma-like pattern. A brief literature review was made with focus on the morphologic variations of DDLMB.     

Dedifferentiated liposarcoma of the cheek

Liposarcomas of the head and neck region are rare; only a few cases have been reported to arise in the cheek or buccal mucosa. Dedifferentiated liposarcoma has rarely been reported in the head and neck region and, to the best of our knowledge, this is the first reported case of dedifferentiated liposarcoma of the cheek. Dedifferentiated liposarcoma is a mixed histologic subtype defined by the association of an atypical lipomatous tumor or well-differentiated liposarcoma and a nonlipogenic sarcoma. The patient was a 61-year-old man who presented with a soft-tissue mass of the left cheek and a presumptive diagnosis of salivary neoplasm based on a fine needle aspiration. The tumor was excised and consisted histologically of an atypical lipomatous tumor/well-differentiated liposarcoma composed a well-differentiated lipomatous neoplasm with atypical cells and rare lipoblasts. The tumor recurred in the same location 5 months after surgery. The recurrent tumor was primarily composed of a nonlipogenic spindle sarcoma with focal rhabdomyoblastic differentiation associated with areas of an atypical lipomatous tumor/well-differentiated liposarcoma.     

Fluorescence in Situ Hybridization (FISH) for Differential Diagnosis of Soft Tissue Sarcomas

Introduction:Soft tissue sarcomas are rare tumors comprising 1 percent of solid malignancies. The latest editionof WHO soft tissue pathology lists 94 benign and malignant soft tissue tumors. Many of these show a large degreeof morphological overlap. Immunohistochemistry has been shown to be reliable in many cases for differential diagnosisof lesions, although cytogenetic tests are considered the gold standard for many entities.Fluorescence in-situ hybridization(FISH) is a cytogenetic technique that uses fluorescent probes that bind to only those parts of the chromosome whichhave a high degree of sequence complementarity. Many soft tissue tumors show recurrent genetic mutations that arenow being used as diagnostic markers. Knowledge of the molecular identity allows prediction of behavior, prognosisand treatment response. Objective:The aim of this study was to identify genetic mutations in soft tissue sarcomas usingFISH testing and to assess correlations with histological diagnosis. Material and methods:A total of 25 cases of differentsoft tissue sarcomas diagnosed on histology with the help of immunohistochemical staining and for which FISH studieswere requested were included in this study. Three pathologists with a special interest in soft tissue sarcomas reviewedthe cases. FISH tests for EWS, the X:18 translocation, FOXO1 and MDM2 were respectively applied for 8 cases ofEwing sarcoma, 8 cases of synovial sarcoma, 2 cases of rhabdomyosarcoma and 7 cases of dedifferentiated liposarcomaand atypical lipomatous tumors/well differentiated liposarcomas. Results:EWS gene fusion was detected in 7 out of8 cases of Ewing sarcoma and the X:18 translocation was positive in 3 of the 8 cases of synovial sarcoma. FOXO1was not detected in either of the two rhabdomyosarcomas. MDM2 by FISH was detected in only one out of 5 cases ofatypical lipomatous tumors and 1 out of 2 dedifferentiated liposarcomas. Conclusion: FISH is a useful adjunct in thediagnostic assessment of different types of soft tissue sarcomas. It is easy to set up, is relatively inexpensive and hasthe ability to diagnose sarcomas with great accuracy, especially in cases which can not be accurately classified evenafter thorough histological and immunohistochemical evaluation. It may play a very important role in the accuratediagnosis and correct management of patients.