Eribulin induces tumor vascular remodeling through intussusceptive angiogenesis in a sarcoma xenograft model.

Eribulin is a novel microtubule inhibitor that, similar to other types of microtubule inhibitors, induces apoptosis by inhibiting the mitotic division of cells. Besides this direct effect on tumor cells, previous studies have shown that eribulin has the potential to induce tumor vascular remodeling in several different cancers; however, the mechanisms underlying this phenomenon remain unclear. In the present study, we aimed to elucidate whether eribulin is effective against synovial sarcoma, a relatively rare sarcoma that often affects adolescents and young adults, and to histologically investigate the microstructure of tumor vessels after the administration of eribulin. We found that eribulin exhibits potent antitumor activity against synovial sarcoma in a tumor xenograft model and that tumor vessels frequently have intervascular pillars, a hallmark of intussusceptive angiogenesis (IA), after the administration of eribulin. IA is a distinct form of angiogenesis that is involved in normal developmental processes as well as pathological conditions. Our data indicate that IA is potentially involved in eribulin-induced vascular remodeling and thereby suggest previously unacknowledged role of IA in regulating the tumor vasculature after eribulin administration.

Fenretinide induces a new form of dynamin-dependent cell death in pediatric sarcoma.

Alveolar rhabdomyosarcoma (aRMS) is a highly malicious childhood malignancy characterized by specific chromosomal translocations mostly encoding the oncogenic transcription factor PAX3-FOXO1 and therefore also referred to as fusion-positive RMS (FP-RMS). Previously, we have identified fenretinide (retinoic acid p-hydroxyanilide) to affect PAX3-FOXO1 expression levels as well as FP-RMS cell viability. Here, we characterize the mode of action of fenretinide in more detail. First, we demonstrate that fenretinide-induced generation of reactive oxygen species (ROS) depends on complex II of the mitochondrial respiratory chain, since ROS scavenging as well as complexing of iron completely abolished cell death. Second, we co-treated cells with a range of pharmacological inhibitors of specific cell death pathways including z-vad (apoptosis), necrostatin-1 (necroptosis), 3-methyladenine (3-MA) (autophagy), and ferrostatin-1 (ferroptosis) together with fenretinide. Surprisingly, none of these inhibitors was able to prevent cell death. Also genetic depletion of key players in the apoptotic and necroptotic pathway (BAK, BAX, and RIPK1) confirmed the pharmacological data. Interestingly however, electron microscopy of fenretinide-treated cells revealed an excessive accumulation of cytoplasmic vacuoles, which were distinct from autophagosomes. Further flow cytometry and fluorescence microscopy experiments suggested a hyperstimulation of macropinocytosis, leading to an accumulation of enlarged early and late endosomes. Surprisingly, pharmacological inhibition as well as genetic depletion of large dynamin GTPases completely abolished fenretinide-induced vesicle formation and subsequent cell death, suggesting a new form of dynamin-dependent programmed cell death. Taken together, our data identify a new form of cell death mediated through the production of ROS by fenretinide treatment, highlighting the value of this compound for treatment of sarcoma patients including FP-RMS.

Intracranial Low-Grade Fibromyxoid Sarcoma: Findings on Electron Microscopy and Histologic Analysis.

BACKGROUND: Low-grade fibromyxoid sarcoma is a rare tumor that manifests as a deep soft tissue mass characterized by a benign histology, but with potentially aggressive clinical behavior and a high rate of recurrence; primary intracranial sarcomas are even rarer. We present a case of primary intracranial low-grade fibromyxoid sarcoma, emphasizing its clinical, radiologic, and histologic features. CASE DESCRIPTION: A 39-year-old woman presented with a recent history of headache and seizures. A right parietal mass was detected on computed tomography and magnetic resonance imaging, and she was subsequently operated on to remove the intracranial mass. Histologic examination of the resected tumor revealed mildly atypical fibroblastic cells embedded within a myxoid matrix. The diagnosis was confirmed by electron microscopy and cytogenetic analyses. CONCLUSIONS: This report describes electron microscopic evaluation of intracranial low-grade fibromyxoid sarcoma, which has an extremely rare occurrence.

Primary cutaneous biphasic sarcomatoid basal cell carcinoma with myoepithelial carcinoma differentiation: A new variant.

Isolated cases of basal cell carcinoma (BCC) with partial myoepithelial component have been described. However, myoepithelial differentiation has not been described in sarcomatoid basal cell carcinomas, which usually show features resembling osteosarcoma, chondrosarcoma, or leiomyosarcoma. We report a case of an 87-year-old man with a forehead lesion that histologically showed a minor component of conventional nodular BCC in transition with a major biphasic sarcomatoid growth composed of invasive spindle-cell and epithelial-like components, the latter with a reticular pattern and scattered ductal structures. Both components showed cytological atypia and high mitotic rate (26/10HPF), with atypical mitotic figures. BER-EP4 immunostaining was exclusively found in the nodular BCC component whereas the sarcomatoid component revealed immunostaining for α-smooth muscle actin (SMA), muscle-specific actin (MSA), calponin, and p63 in both epithelial-like and spindle-cell populations. Focal immunoreactivity was observed in the epithelial component for S100 and glial fibrillary acidic protein (GFAP). Furthermore, EWSR1-PBX1 gene fusion was also detected. This is to our knowledge, the first fully documented case of biphasic sarcomatoid BCC with myoepithelial carcinoma differentiation.

A ganglion-rich gastrointestinal stromal tumor: A case report.

We report a case of an extremely rare type of duodenal gastrointestinal stromal tumor (GIST) that included neuronal components. Although gastrointestinal autonomic nerve tumors (GANTs), a subtype of GISTs, exhibit ultrastructural features of the nerve plexus, neuronal cells have not been observed within GANTs or GISTs. GISTs originate from interstitial cells of Cajal (ICCs), which are markedly different from the progenitor cells of neural elements and neural-crest-derived stem cells. This may explain why GISTs typically lack neuronal elements. It remains unclear that the neuronal components of this tumor are neoplastic or hyperplastic, but proliferation and survival of ICCs have recently been reported to be closely related to neurons. Although we could not find the KIT, PDGFR, and BRAF mutation as far as we examined, it may have had a rare mutation in NF1, a fusion of EVT6-NTRK3, or an as-yet-unknown KIT mutation that affected neurogenesis. Further investigation of related genetic mutations and accumulation of data from other similar cases is needed.

Sarcomeric disorganization and nemaline bodies in muscle biopsies of patients with EXOSC3-related type 1 pontocerebellar hypoplasia.

INTRODUCTION: Mutations in the EXOSC3 gene are responsible for type 1 pontocerebellar hypoplasia, an autosomal recessive congenital disorder characterized by cerebellar atrophy, developmental delay, and anterior horn motor neuron degeneration. Muscle biopsies of these patients often show characteristics resembling classic spinal muscle atrophy, but to date, no distinct features have been identified. METHODS: Clinical data and muscle biopsy findings of 3 unrelated patients with EXOSC3 mutations are described. RESULTS: All patients presented as a severe congenital cognitive and neuromuscular phenotype with short survival, harboring the same point mutation (c.92G>C; p.Gly31Ala). Muscle biopsies consistently showed variable degrees of sarcomeric disorganization with myofibrillar remnants, Z-line thickening, and small nemaline bodies. CONCLUSIONS: In this uniform genetic cohort of patients with EXOSC3 mutations, sarcomeric disruption and rod structures were prominent features of muscle biopsies. In the context of neonatal hypotonia, ultrastructural studies might provide early clues for the diagnosis of EXOSC3-related pontocerebellar hypoplasia. Muscle Nerve 59:137-141, 2019.

A rare subtype of meningioma: Case series of anaplastic meningioma and review of the literature.

RATIONALE: Anaplastic meningioma, a rare subtype of meningioma, has malignant morphological characteristics and a World Health Organization (WHO) grade of III. PATIENT CONCERNS: In this report, we present findings from 6 cases of anaplastic meningioma. DIAGNOSES: Pathological examination of the tumors, including hematoxylin and eosin staining and immunohistochemical staining, was performed. Of the six cases of anaplastic meningioma, two were recurrent tumors from original seminoma with a WHO grade of I. Histologically, three cases had carcinoma-like morphology, one case had sarcoma-like morphology, and two had two kinds of tissue structures: carcinoma-like tumor cell nests and areas with spindle tumor cells. Necrosis was detected in most cases (5/6). Ki67 index was high and varied from 20% to 70%. INTERVENTIONS: All the patients received surgery. 3 patients received adjuvant radiotherapy. 1 patient received chemotherapy. OUTCOMES: 4 patients had no recurrence at follow-up of 19, 30, 46 and 54 months after the last surgery. 1 patient had recurrence 3 months after the last surgery. 1 patient died 12 days after the last surgery. LESSONS: This malignant subtype can be secondary to a WHO grade I meningioma after a long quiescent period. Necrosis was common in the tumor tissues, and Ki67 index was usually high. For patients with a history of meningioma, including benign cases, regular physical examination is important for early detection of tumor recurrence and malignant transformation.

Raf Kinase Inhibitor Protein Expression and Prognostic Value in Soft Tissue Sarcomas.

OBJECTIVE: Soft tissue sarcomas (STSs) are heterogeneous tumors displaying multiple and complex molecular abnormalities with no specific pattern. Despite current therapeutic advances, the patients with STS still have a poor outcome, which makes it necessary to find out new prognostic markers. The Raf kinase inhibitory protein (RKIP) has been associated with prognosis in several human neoplasms; however, its role in STS is unknown. METHODS: In the present study RKIP expression was assessed by immunohistochemistry in a series of 87 STSs, and its expression profile was associated with the patients' pathological parameters. RESULTS: We found that RKIP is expressed in the cytoplasm of the great majority of cases, and absent in only approximately 18% of cases (16/87). Importantly, we observed that loss of RKIP expression was associated with poor outcome, constituting an independent prognostic marker. CONCLUSION: This is the first study assessing RKIP expression levels in STS. We showed that loss of RKIP expression is present in a small subset of cases; however, its absence was associated with poor survival and may be a potential marker for STS prognosis.

Ring chromosomes, breakpoint clusters, and neocentromeres in sarcomas.

Gene amplification is relatively common in tumors. In certain subtypes of sarcoma, it often occurs in the form of ring and/or giant rod-shaped marker (RGM) chromosomes whose mitotic stability is frequently rescued by ectopic novel centromeres (neocentromeres). Little is known about the origin and structure of these RGM chromosomes, including how they arise, their internal organization, and which sequences underlie the neocentromeres. To address these questions, 42 sarcomas with RGM chromosomes were investigated to detect regions prone to double strand breaks and possible functional or structural constraints driving the amplification process. We found nine breakpoint cluster regions potentially involved in the genesis of RGM chromosomes, which turned out to be significantly enriched in poly-pyrimidine traits. Some of the clusters were located close to genes already known to be relevant for sarcomas, thus indicating a potential functional constraint, while others mapped to transcriptionally inactive chromatin domains enriched in heterochromatic sites. Of note, five neocentromeres were identified after analyzing 13 of the cases by fluorescent in situ hybridization. ChIP-on-chip analysis with antibodies against the centromeric protein CENP-A showed that they were a patchwork of small genomic segments derived from different chromosomes, likely joint to form a contiguous sequence during the amplification process.

Is mammary not otherwise specified-type sarcoma with CD10 expression a distinct entity? A rare case report with immunohistochemical and ultrastructural study.

Mammary sarcoma is extremely rare and the diagnosis is established only after metaplastic carcinomas and malignant phyllodes tumours are excluded. A rare case of not otherwise specified-type sarcoma with CD10 expression in the left breast in a 45-year-old female was presented. It was a high-grade tumour composed of spindle cells histologically. The immunohistochemical results showed that CD10, vimentin and EGFR were positive diffusely and SMA presented focally, whereas epithelial markers and other myoepithelial or myogenic markers were all negative. The electron microscope investigation demonstrated fibroblast-like features. The exact entity of the tumour remains to be studied because it resembles undifferentiated sarcoma or sarcomatoid metaplastic carcinoma to some degree, as well as high-grade malignant phyllodes tumour in particular. VIRTUAL SLIDES: The virtual slide(s) for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/9019879588725702.

Methylglyoxal induces mitochondria-dependent apoptosis in sarcoma.

In the preceding paper (A. Ghosh et al. (2011) Biochemistry (Moscow), 76, 1051-1060), using several comparable tissue materials, it has been convincingly demonstrated that methylglyoxal, a normal metabolite, inhibits mitochondrial complex I of specifically malignant cells. This suggests a distinct alteration of complex I, a highly important enzyme for energy (ATP) production, in malignancy. The present paper shows that as a consequence of this inhibition mitochondrial membrane potential is drastically reduced in sarcoma tissue but not in normal skeletal muscle. This was estimated spectrofluorimetrically using the dye rhodamine 123. As a consequence, cytochrome c was released from the sarcoma mitochondria as evidenced by Western blot analysis. Moreover, on treatment with methylglyoxal membrane potential collapse of sarcoma 180 cells was also indicated by fluorescence-activated cell sorter analysis. Atomic force microscopic study demonstrated gross structural alteration specifically of tumor mitochondria on methylglyoxal treatment. All these studies suggest that methylglyoxal might initiate an apoptotic event in malignant cells.

Immunoglobulin G locus events in soft tissue sarcoma cell lines.

Recently immunoglobulins (Igs) have been found to be expressed by cells other than B lymphocytes, including various human carcinoma cells. Sarcomas are derived from mesenchyme, and the knowledge about the occurrence of Ig production in sarcoma cells is very limited. Here we investigated the phenomenon of immunoglobulin G (IgG) expression and its molecular basis in 3 sarcoma cell lines. The mRNA transcripts of IgG heavy chain and kappa light chain were detected by RT-PCR. In addition, the expression of IgG proteins was confirmed by Western blot and immunofluorescence. Immuno-electron microscopy localized IgG to the cell membrane and rough endoplasmic reticulum. The essential enzymes required for gene rearrangement and class switch recombination, and IgG germ-line transcripts were also identified in these sarcoma cells. Chromatin immunoprecipitation results demonstrated histone H3 acetylation of both the recombination activating gene and Ig heavy chain regulatory elements. Collectively, these results confirmed IgG expression in sarcoma cells, the mechanism of which is very similar to that regulating IgG expression in B lymphocytes.

Detection of SRS19-6MuLV in mouse dendritic cell sarcoma and its tumorigenesis.

SRS19-6MuLV is a member of the MuLV family originally isolated from the Tianjin-Shanghai-Zunyi complex of murine leukemia. A notable characteristic of this virus is that it induces tumors of multiple hematopoietic lineages, including myeloid, erythroid, T-lymphoid and B-lymphoid. In a previous study, a sequence with high homology to SRS19-6MuLV in a murine dendritic cell sarcoma (DCS) was identified through cDNA expression screening with mAb 983D4. To investigate the relationship between SRS19-6MuLV and DCS, the existence of a specific SRS19-6MuLV DNA fragment in DCS cells, 15 murine tumor cells, 2 murine tumor tissues, 12 normal murine cells/tissues, 11 human tumor cell lines and SRSV/3T3 (NIH/3T3 cells infected with SRS cell supernatant) was detected by PCR. The specific fragment of SRS19-6MuLV was detected in DCS, mouse fore-gastric cancer cells, LⅡ tumor tissue from which DCS is derived and SRSV/3T3. In addition, the integration sites of SRS19-6MuLV in the positive cells were examined by inverse PCR. Thus, 7 integration sites for SRS19-6MuLV were detected in DCS and 3 in SRSV/3T3. Analysis of sequences by BLAST revealed that some of the integration sites were associated with common fragile sites and some Ras-regulating miRNAs. Our results indicate that SRS19-6MuLV not only induced four types of leukemia, but also induced DCS. This virus does not infect human cells. Multiple integration of SRS19-6MuLV into chromosomes around fragile sites accounts for its carcinogenic effects.

Histone deacetylase inhibitor vorinostat suppresses the growth of uterine sarcomas in vitro and in vivo.

BACKGROUND: Uterine sarcomas are very rare malignancies with no approved chemotherapy protocols. Histone deacetylase (HDAC) inhibitors belong to the most promising groups of compounds for molecular targeting therapy. Here, we described the antitumor effects of suberoylanilide hydroxamic acid (SAHA; vorinostat) on MES-SA uterine sarcoma cells in vitro and in vivo. We investigated effects of vorinostat on growth and colony forming ability by using uterine sarcoma MES-SA cells. We analyzed the influence of vorinostat on expression of different HDACs, p21(WAF1) and activation of apoptosis. Finally, we examined the antitumor effects of vorinostat on uterine sarcoma in vivo. RESULTS: Vorinostat efficiently suppressed MES-SA cell growth at a low dosage (3 microM) already after 24 hours treatment. Decrease of cell survival was even more pronounced after prolonged treatment and reached 9% and 2% after 48 and 72 hours of treatment, respectively. Colony forming capability of MES-SA cells treated with 3 microM vorinostat for 24 and 48 hours was significantly diminished and blocked after 72 hours. HDACs class I (HDAC2 and 3) as well as class II (HDAC7) were preferentially affected by this treatment. Vorinostat significantly increased p21(WAF1) expression and apoptosis. Nude mice injected with 5 x 106 MES-SA cells were treated for 21 days with vorinostat (50 mg/kg/day) and, in comparison to placebo group, a tumor growth reduction of more than 50% was observed. Results obtained by light- and electron-microscopy suggested pronounced activation of apoptosis in tumors isolated from vorinostat-treated mice. CONCLUSIONS: Our data strongly indicate the high therapeutic potential of vorinostat in uterine sarcomas.

Transformation potential of bone marrow stromal cells into undifferentiated high-grade pleomorphic sarcoma.

PURPOSE: Bone marrow adherent cells contain conventional bone marrow stromal cells and mesenchymal stem cells and these cells constitute the hematopoietic microenvironment. Mesenchymal stem cells have the capacity to give rise to multiple mesenchymal lineage cells and even ectodermal lineage cells. In the present study, we investigated what types of tumor cells are inducible from BM adherent cells by chemical carcinogens. METHODS: Bone marrow cells from neonatal C3H/HeN mice were collected within 24 h after birth and then cultured. Four days later, bone marrow adherent cells were obtained and the cells were treated with 3-methylcholanthrene. RESULTS: By this treatment, some transformed clones consisting of large spindle cells were obtained. The transformed cells were highly positive for CD44 and were positive for Sca-1, CD49d and CD106, whereas the cells were negative for hematolymphoid markers. The cell clones had the ability to support hematopoiesis in vitro. These results indicate that the transformed cell lines have the characteristics of BM stromal cells/mesenchymal stem cells. Moreover, during culture of the transformed cells, spontaneous bone nodule formation was observed. When the transformed cells were inoculated into immunodeficient mice subcutaneously, the neoplasms grew in the subcutaneous tissue of the mice. Microscopically and ultrastructurally, the neoplasms showed the typical morphology of undifferentiated high-grade pleomorphic sarcoma (UHGPS). Bone-related genes have been found to be expressed in both transformed cells and UHGPSs. CONCLUSION: The present study suggests that UHGPSs are derived from BM stromal cells, probably mesenchymal stem cells.

IGF2 is highly expressed in pediatric undifferentiated sarcomas and reveals two distinct cytoplasmic trafficking patterns.

Pediatric undifferentiated soft tissue sarcomas (USTSs) are tumors composed of primitive mesenchymal cells. As such, they form an attractive model for studying the early events in sarcoma development. In an effort to better understand the critical molecular aberrations leading to sarcoma development, gene expression array analysis and post-array validation techniques were applied to several USTSs; the results were consistent with upregulation of the excitatory components of the insulin-like growth factor (IGF) pathway. Particularly high expression of the insulin-like growth factor 2 (IGF2) ligand was seen and confirmed by real-time reverse transcriptase-polymerase chain reaction. Immunohistochemistry performed using antibodies against IGF2 revealed overexpression of the IGF2 protein in 19 of 21 (90%) USTSs and revealed 2 distinct staining patterns, 1 in which there was diffuse cytoplasmic staining (16/19) and 1 in which there was punctate perinuclear positivity (3/19). Using ultrastructural immunogold localization of IGF2, it was determined that IGF2 was primarily localized to Golgi-derived vesicles and multivesicular bodies in tumor cells showing the punctate pattern, and to both the cytoplasm and plasma membrane of tumor cells showing the diffuse pattern. The results suggest that upregulation of the IGF pathway in pediatric USTSs is a critical early event in the development of sarcomas. Furthermore, findings from the immunocytochemical and immunogold analyses confirm the presence of 2 different cytoplasmic trafficking patterns and storage motifs of IGF2 within this type of tumor. Given that in one subcellular pattern the IGF2 protein does not appear to reach the membrane, these findings may have functional significance.

Epithelioid sarcoma: a case report with ultrastructural confirmation of myofibroblastic differentiation based on fibronexus junctions.

Epithelioid sarcoma is an uncommon but well-described malignancy, which is found predominantly in the soft tissues of the young and middle-aged, and which pursues an indolent to aggressive course. It shows a degree of both mesenchymal and epithelial differentiation. Myofibroblastic differentiation has been recorded in epithelioid sarcoma for some time, the evidence being based mainly on the presence of smooth-muscle-type myofilaments and, more recently, on alpha-smooth-muscle actin and muscle-specific actin immunostaining. Myofibroblastic differentiation based on the stricter criterion of the fibronectin fibril/fibronexus junction has not so far been demonstrated except for a single atypical case with spindle-cell morphology and a cytokeratin-negative immunophenotype. The authors describe a conventional epithelioid sarcoma showing myofibroblastic differentiation based on the presence of fibronectin fibrils and fibronexus junctions. The patient was a 35-year-old Chinese male with a tumor that initially developed on his left forefinger: it recurred, then metastasised first to the left arm and eventually to the scalp. Histologically, the tumors had the typical features of epithelioid sarcoma: by immunohistochemistry, cytokeratin, epithelial membrane antigen, and vimentin were positive. By electron microscopy, rough endoplasmic reticulum, intermediate filaments, peripheral myofilaments, and fibronexus junctions were observed. This is the first epithelioid sarcoma of conventional histological type to show myofibroblastic differentiation on the basis of the more stringent criterion of the fibronexus. The findings are discussed in relation to the unusually varied differentiation reported for this tumor.

Sarcomatoid acquired cystic disease-associated renal cell carcinoma.

In this article, we report a rare case of hitherto undescribed acquired cystic disease (ACD)-associated renal cell carcinoma (RCC) with sarcomatoid change. A 78-year-old woman had been receiving hemodialysis for fourteen years at the time when a renal tumor was encountered on the follow-up examination of the kidney. Microscopically, oncocytic cuboidal cells proliferated with tubular, cribriform or papillary growth patterns, and atypical columnar cells with abundant cytoplasm proliferated with papillary configuration. Oxalate crystal deposition was observed in the stroma and the tumor focally resembled translocation type (TFE3) RCC. Sarcomatous neoplastic cells were also seen. The cytoplasm of oncocytic and sarcomatous neoplastic cells was diffusely positive for anti-mitochondrial antibody and the ultrastructural examination detected many mitochondria in the cytoplasm of oncocytic carcinoma cells and sarcomatous neoplastic cells. The loss of chromosomes 1p, 2q11-22, 9 and 14 was observed using comparative genomic hybridization analysis. We thus report here a case of hitherto undescribed ACD-associated RCC intermingled with oncocytic cells, translocation type RCC-like area and sarcomatoid change. This is the sixth case of sarcomatoid RCC arising in end-stage kidney disease.