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Osteogenic Sarcoma (osteosarcoma) is a bone forming cancer. It is the most frequent type of bone tumour and is most common between the ages of 15 to 25. Over 90% of tumours are located in the metaphysis (the growing ends of the bone), the most common sites are the bones around the knee which account for 80% of cases. Osteosarcomas vary greatly in radiological and pathological features and therefore needs careful diagnosis to differentiate this from other bone tumours. Most are high grade intramedullary osteosarcomas, about 5% are low grade lesions, some are secondary osteosarcomas (for example those caused by radiation therapy).

Figure 1. Radiograph showing an osteolytic and osteoblastic intra-medullary tumor characteristic of osteosarcoma. From Layfield J et al. Clin Med Pathol. 2008; 1: 55-59. Available under a Creative Commons CC-BY-3.0 license.

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See also: Genetic features of Osteosarcoma

Latest Research Publications

This list of publications is regularly updated (Source: PubMed).

Choi LE, Healey JH, Kuk D, Brennan MF
Analysis of outcomes in extraskeletal osteosarcoma: a review of fifty-three cases.
J Bone Joint Surg Am. 2014; 96(1):e2 [PubMed] Related Publications
BACKGROUND: Extraskeletal osteosarcoma is a rare soft-tissue sarcoma about which little is known. The objectives of this study were to describe the clinical features and natural history of extraskeletal osteosarcoma and to investigate factors affecting outcomes.
METHODS: A retrospective review of a prospectively maintained database of patients diagnosed with soft-tissue sarcoma was conducted. Patients with pathologically confirmed extraskeletal osteosarcoma from 1982 to 2012 were identified and were included in the analysis. Medical records were reviewed for clinical features, treatment, and outcomes.
RESULTS: Fifty-three patients were identified from the database: forty-two presented with localized disease, two presented with metastatic disease, and nine presented with recurrent (local and/or distant) disease. The median patient age at diagnosis was sixty-four years, with a median follow-up time of thirty-four months (range, one to 290 months) for survivors. Of the fifty-three patients who were identified, forty-one had lesions in the extremities, fifty-one had high-grade lesions, forty had lesions >5 cm, and forty-two had deep lesions. For patients presenting with localized disease, the median survival was 45.8 months with a three-year cumulative incidence of death due to disease of 39%. All patients with localized disease were managed with surgical resection of the primary tumor: nineteen with surgery only, ten with adjuvant radiation, five with adjuvant chemotherapy, and eight with both radiation and chemotherapy. Eighteen patients relapsed: two patients had local recurrences, ten patients had distant metastases, and six patients had local recurrences and distant metastases. In log-rank analysis, patients with superficial tumors and negative margins at resection had a higher three-year event-free survival. No significant association of disease-specific or event-free survival was found with the addition of radiation, chemotherapy, or both to surgery.
CONCLUSIONS: For patients presenting with localized extraskeletal osteosarcoma, three-year event-free survival was higher for patients with superficial tumors and negative margins at resection. Radiation and chemotherapeutic treatment were not associated with a lower incidence of death due to disease or a longer event-free survival.

Zhang Y, Ma Q, Liu T, et al.
Tumor self-seeding by circulating tumor cells in nude mouse models of human osteosarcoma and a preliminary study of its mechanisms.
J Cancer Res Clin Oncol. 2014; 140(2):329-40 [PubMed] Related Publications
PURPOSE: The purpose of this study is to determine whether and how tumor self-seeding by circulating tumor cells (CTCs) plays a role in the initiation and progression of osteosarcoma.
METHODS: Two different nude mouse models of human osteosarcoma were established for detecting tumor self-seeding by fluorescently labeled CTCs. Various tumor growth indicators were quantitated for seeded and unseeded groups. Growth mechanisms were characterized using cell proliferation assays and immunohistochemical staining. Conditioned media of primary osteosarcoma cells was characterized in a Transwell migration assay and enzyme-linked immunosorbent assay. The effect of cytokines secreted by primary tumor cells was verified by small interfering RNA and recombinant human cytokine experiments.
RESULTS: Red fluorescent protein-labeled CTCs seeded primary tumors in both models. Seeded primary tumors groups grew faster than control groups (P < 0.05), which was partially attributed to the CTCs having a higher proliferation rate and higher vascular endothelial growth factor expression after self-seeding. Conditioned media of primary osteosarcoma cells attracted CTCs, through an IL-6-dependent mechanism.
CONCLUSIONS: CTC tumor self-seeding occurs in osteosarcoma and promotes the growth of primary osteosarcoma. CTCs appear to be recruited by cytokines secreted by primary osteosarcoma cells, particularly IL-6.

Related: Bone Cancers VEGFA

Yagishita S, Horinouchi H, Yorozu T, et al.
Secondary osteosarcoma developing 10 years after chemoradiotherapy for non-small-cell lung cancer.
Jpn J Clin Oncol. 2014; 44(2):191-4 [PubMed] Related Publications
A 53-year-old female patient was admitted with pain and a progressively enlarging mass in the right upper chest. Chest computed tomography revealed a mass lesion in the region of the right upper ribs. Ten years prior to this admission, the patient had undergone right lobectomy for lung adenocarcinoma. One year after the surgery, follow-up computed tomography had revealed tumor recurrence in the mediastinal and supraclavicular lymph nodes, and the patient had been treated by chemoradiotherapy. Thereafter, regular follow-up had revealed no evidence of recurrence of the non-small-cell lung cancer. Histopathological findings revealed proliferation of spindle-shaped malignant tumor cells in a background of osteoid, consistent with the diagnosis of osteosarcoma. The location of the tumor was consistent with the radiation field. Based on the clinicopathological findings, the patient was diagnosed as having secondary osteosarcoma occurring as a result of the chemoradiotherapy administered previously for the recurrent non-small-cell lung cancer. Unfortunately, the patient died of rapid progression of the osteosarcoma within a week of admission to the hospital. The autopsy revealed contiguous invasion by the tumor of the heart, with massive thrombus formation. The peripheral pulmonary arteries were diffusely occluded by metastatic tumors. Our case serves to highlight the risk of development of secondary sarcoma as a life-threatening late complication after chemoradiotherapy for locally advanced non-small-cell lung cancer, even after complete cure of the primary tumor.

Related: Bone Cancers Non-Small Cell Lung Cancer Lung Cancer

Gobin B, Battaglia S, Lanel R, et al.
NVP-BEZ235, a dual PI3K/mTOR inhibitor, inhibits osteosarcoma cell proliferation and tumor development in vivo with an improved survival rate.
Cancer Lett. 2014; 344(2):291-8 [PubMed] Related Publications
Despite recent improvements in chemotherapy and surgery, the problem of non-response osteosarcoma to chemotherapy remains, and is a parameter that is critical for prognosis. The present work investigated the therapeutic value of NVP-BEZ235, a dual class I PI3K/mTOR inhibitor. NVP-BEZ235 inhibited osteosarcoma cell proliferation by inducing G0/G1 cell cycle arrest with no caspase activation. In murine pre-clinical models, NVP-BEZ235 significantly slowed down tumor progression and ectopic tumor bone formation with decreased numbers of Ki67(+) cells and reduced tumor vasculature. Finally, NVP-BEZ235 considerably improved the survival rate of mice with osteosarcoma. Taken together, the results of the present work show that NVP-BEZ235 exhibits therapeutic interest in osteosarcoma and may be a promising adjuvant drug for bone sarcomas.

Related: Bone Cancers

Cheng DD, Zhao HG, Yang YS, et al.
GSK3β negatively regulates HIF1α mRNA stability via nucleolin in the MG63 osteosarcoma cell line.
Biochem Biophys Res Commun. 2014; 443(2):598-603 [PubMed] Related Publications
Hypoxia-inducible factor 1α (HIF1α) is a transcription factor involved in the growth, invasion and metastasis of malignant tumors. Glycogen synthase kinase 3 beta (GSK3β) is a protein kinase involved in a variety of signaling pathways, such as the Wnt and NF-κB pathways; this kinase can affect tumor progress through the regulation of transcription factor expression and apoptosis. Recent studies showed that GSK3β was involved in the expression of HIF1α. However, the effect of GSK3β on HIF1α expression in osteosarcoma cells remains unknown. To understand the relationship between GSK3β and HIF1α comprehensively, small RNA interference techniques, Western blot analyses, quantitative real-time PCR analyses and luciferase assays were used in our study. Experimental data revealed that inhibition of GSK3β could increase HIF1α protein levels and expression of its target genes by increasing the stability of the HIF1α mRNA, not by affecting the HIF1α protein stability, and that this process could be mediated by nucleolin.

Related: HIF1A

Johal S, Ralston S, Knight C
Mifamurtide for high-grade, resectable, nonmetastatic osteosarcoma following surgical resection: a cost-effectiveness analysis.
Value Health. 2013; 16(8):1123-32 [PubMed] Related Publications
OBJECTIVES: Mifamurtide is an immune macrophage stimulant that when added to standard chemotherapy has demonstrated survival benefit for newly diagnosed osteosarcoma. The objectives of this study were to investigate the cost-effectiveness of adding mifamurtide to standard three- or four-agent chemotherapy for high-grade, resectable, nonmetastatic osteosarcoma following surgical resection and the issues of obtaining robust cost-effectiveness estimates for ultra-orphan drugs, given the shortage of data.
METHODS: An economic evaluation was conducted from the perspective of the UK's National Health Service as part of the manufacturer's submission to the National Institute for Health and Care Excellence. The disease process was simplified to a transition through a series of health states, modeled by using a Markov approach. Data to inform the model were derived from patient-level data of Study INT-0133, published literature, and expert opinion. The final efficacy measure was life-years gained (LYG), and utilities were used to obtain quality-adjusted life-years (QALYs).
RESULTS: For a 60-year time frame and a discount rate of 3.5% for outcomes, patients receiving mifamurtide benefited from an average additional 1.57 years of life and 1.34 QALYs, compared with patients receiving chemotherapy alone, giving an incremental cost-effectiveness ratio (ICER) of £58,737 per LYG and £68,734 per QALY. Because treatment effects were both substantial in restoring health and sustained over a very long period, the National Institute for Health and Care Excellence changed its guidance to allow a discount of 1.5% for outcomes to be applied in these special circumstances. By using this discount factor, it was found that patients receiving mifamurtide had an average additional 2.58 years of life and 2.20 QALYs compared with patients receiving chemotherapy alone, resulting in an ICER of £35,765 per LYG and £41,933 per QALY.
CONCLUSION: Mifamurtide's ICER is cost-effective compared with that of other orphan and ultra-orphan drugs, for which prices and corresponding cost-effectiveness estimates are high.

Related: Bone Cancers

Fu Z, Deng B, Liao Y, et al.
The anti-tumor effect of shikonin on osteosarcoma by inducing RIP1 and RIP3 dependent necroptosis.
BMC Cancer. 2013; 13:580 [PubMed] Related Publications
BACKGROUND: Osteosarcoma is the most frequent primary malignant bone tumor, notorious for its lung metastasis. Shikonin, an effective constituent extracted from Chinese medicinal herb, was demonstrated to induce necroptosis in some cancers.
METHODS: MTT assay was performed to detect cell survival rate in vitro. Flow cytometry was used to analyze cell cycle and cell death. Western blot was performed to determine the expression levels of RIP1, RIP3, caspase-3, caspase-6 and PARP. The tibial primary and lung metastatic osteosarcoma models were used to evaluate the anti-tumor effect of shikonin in vivo.
RESULTS: The cell survival rate was decreased in a dose and time dependent manner when treated with shikonin. No major change in cell cycle was observed after shikonin treatment. The cell death induced by shikonin could be mostly rescued by specific necroptosis inhibitor necrostatin-1, but not by general caspase inhibitor Z-VAD-FMK. The number of necrotic cells caused by shikonin was decreased after being pretreated with Nec-1 detected by flow cytometry in K7 cells. After 8-hour treatment of shikonin, the expression levels of RIP1 and RIP3 were increased while caspase-3, caspase-6 and PARP were not activated in K7 and U2OS cells determined by Western blot. Size of primary tumor and lung metastasis in shikonin treated group were significantly reduced. The protein levels of RIP1 and RIP3 in primary tumor tissues were increased by shikonin. The overall survival of lung metastatic models was longer compared with control group (p < 0.001).
CONCLUSIONS: Shikonin had prompt but profound anti-tumor effect on both primary and metastatic osteosarcoma, probably by inducing RIP1 and RIP3 dependent necroptosis. Shikonin would be a potential anti-tumor agent on the treatment of primary and metastatic osteosarcoma.

Related: Apoptosis Bone Cancers

Incesu Z, Hatipoğlu I, Sivas H, et al.
Effects of fibronectin and type IV collagen on osteosarcoma cell apoptosis.
Indian J Exp Biol. 2013; 51(10):789-96 [PubMed] Related Publications
The aims of this study are the investigation of the effects of fibronectin and type IV collagen extracellular matrix proteins and the role of caspase-3 and -9 on cis-platin induced U2-OS apoptosis were studied. First the cytotoxic effects of cis-platin on cell system were investigated by colorimetric method and than morphological and ELISA analysis were used for determination of cell apoptosis when induced with cis-platin. In addition, after adhering the cells to fibronection or type IV collagen proteins, the apoptotic rate and the effects of caspase-3 and -9 were also investigated by ELISA in presence of specific inhibitors. U2-OS cells showed 20% cytotoxicity after treatment with 2.4 microM of cis-platin for 48 h. Morphological and the numerical data showed that cis-platin was able to induced apoptosis on cells as a dose-dependent manner. Caspase-3 and -9 inhibitors inhibited cis-platin-induced apoptosis in U2-OS cells, respectively. The binding of cells to 10 microg/mL of fibronectin but not type IV collagen enhanced the apoptosis about 2.5 fold that effects inhibited with caspase-3 inhibitor. The caspase-3 and -9 are involved in the apoptotic signals induced by cis-platin in U2-OS. The binding to fibronectin, but not type IV collagen enhanced the apoptotic response of U2-OS and fibronectin-dependent apoptosis was activated by caspase-3. These finding might be useful for patients to fight against osteosarcoma.

Related: Apoptosis Bone Cancers Cisplatin

Jeon DG, Kong CB, Cho WH, et al.
Examination of the cutoff value of postchemotherapy increase in tumor volume as a predictor of subsequent oncologic events in stage IIB osteosarcoma.
J Surg Oncol. 2014; 109(3):275-9 [PubMed] Related Publications
BACKGROUND: Tumor enlargement after chemotherapy is a predictor of a poor histological response, poor survival, and local recurrence. However, the cutoff point of tumor enlargement for predicting subsequent oncologic events has not been determined.
METHODS: We retrospectively reviewed 567 patients who were treated at our institute for stage IIB osteosarcoma. We used receiver operating characteristic (ROC) curve analysis of tumor volume increase for the prediction of subsequent metastasis or local recurrence, and calculated diagnostic indices for different cutoff values.
RESULTS: A tumor volume increase of >15% predicted subsequent metastasis or local recurrence with a sensitivity of 64.7%, a specificity of 81.5%, a positive predictive value of 71.6%, and a negative predictive value of 76.1%. Increases in tumor volumes based on this cutoff value were able to predict subsequent oncologic events in all clinical subgroups, except in cases of rare pathologic subtypes. However, for tumors in the proximal humerus, a cutoff value of 25% had optimal predictive value.
CONCLUSIONS: This study shows that a cutoff value of 15% for tumor volume increase is useful for predicting subsequent metastasis or local recurrence. Our results suggest that tumor enlargement after chemotherapy serves as an easily assessable clinical parameter for risk-adapted therapy.

Related: Bone Cancers

Kansara M, Leong HS, Lin DM, et al.
Immune response to RB1-regulated senescence limits radiation-induced osteosarcoma formation.
J Clin Invest. 2013; 123(12):5351-60 [PubMed] Free Access to Full Article Related Publications
Ionizing radiation (IR) and germline mutations in the retinoblastoma tumor suppressor gene (RB1) are the strongest risk factors for developing osteosarcoma. Recapitulating the human predisposition, we found that Rb1+/- mice exhibited accelerated development of IR-induced osteosarcoma, with a latency of 39 weeks. Initial exposure of osteoblasts to carcinogenic doses of IR in vitro and in vivo induced RB1-dependent senescence and the expression of a panel of proteins known as senescence-associated secretory phenotype (SASP), dominated by IL-6. RB1 expression closely correlated with that of the SASP cassette in human osteosarcomas, and low expression of both RB1 and the SASP genes was associated with poor prognosis. In vivo, IL-6 was required for IR-induced senescence, which elicited NKT cell infiltration and a host inflammatory response. Mice lacking IL-6 or NKT cells had accelerated development of IR-induced osteosarcomas. These data elucidate an important link between senescence, which is a cell-autonomous tumor suppressor response, and the activation of host-dependent cancer immunosurveillance. Our findings indicate that overcoming the immune response to senescence is a rate-limiting step in the formation of IR-induced osteosarcoma.

Related: Bone Cancers Cytokines RB1

Tsagaraki I, Phenekos C, Tsilibary E, Tzinia A
Calcitonin-induced NF-κB activation up-regulates fibronectin expression in MG63 osteosarcoma cells.
Anticancer Res. 2013; 33(11):4901-6 [PubMed] Related Publications
Salmon calcitonin has been used extensively as a therapeutic tool in the regulation of bone remodeling. However, there is a growing body of evidence indicating that the calcitonin peptides are involved in regulation of cell growth, differentiation, survival and tissue development. In the present study, we investigated the effect of calcitonin in cell matrix interactions in MG63 cell line. Our results demonstrated that calcitonin increases cell growth of MG63 osteosarcoma cells in parallel with serine/threonine protein kinase B (AKT/PKB) activation. Moreover, calcitonin induced up-regulation of fibronectin expression in a nuclear factor-kappa B (NF-κB)-dependent manner, accompanied by enhanced enzymatic activity of matrix metalloproteinase-9 (MMP-9) and increased expression of tissue inhibitors of MMP-1 and -2. MMP-9 stimulation with calcitonin was accompanied by an increase in protein expression of the α5β1 integrin receptor. To our knowledge, our results demonstrate, for the first time, that calcitonin is a potent inducer of fibronectin, an extacellular matrix component that is suggested to have a pro-oncogenic and healing effect, in a NF-κB-dependent manner.

Related: Bone Cancers MMP2 MMP9: matrix metallopeptidase 9 AKT1 Signal Transduction

Lim JB, Sharma H, MacDuff E, Reece AT
Primary osteosarcoma of the spine: a review of 10 cases.
Acta Orthop Belg. 2013; 79(4):457-62 [PubMed] Related Publications
The authors describe 10 cases of osteosarcoma of the spine treated between January 1951 and December 2010, and obtained from the Tumour Registry of their hospital. The mean age at presentation was 38.8 years (range: 16-73 years); the mean duration of symptoms was 5.1 months (range: 3 weeks-1 year). Pain was the commonest complaint (9 patients), followed by neurological compromise (6 patients). The thoracic spine and male gender were predominant. Seven patients underwent marginal resection, 3 underwent intralesional resection. All, except one, had adjuvant chemotherapy and radiotherapy, pre- and/or postoperatively. This rare sarcoma has a dismal prognosis : the median survival period was only 23 years. The 1-year, 3-year and 5-year survival rates were 80%, 40% and 20%. Astonishingly, marginal resection (7 cases) did not lead to a longer survival than intralesional resection (3 cases): respectively 30 months and 42 months. Quite logically, local recurrence in 6 patients was linked to a survival of only 36 months, while the other 4 patients survived 52 months. Age below 40 was a positive factor, but not significantly. All patients had a reasonable quality of life with outcomes consistent with the available literature. Recent literature stresses that there is a trend toward improved survival with en bloc resection.

Tu B, Peng ZX, Fan QM, et al.
Osteosarcoma cells promote the production of pro-tumor cytokines in mesenchymal stem cells by inhibiting their osteogenic differentiation through the TGF-β/Smad2/3 pathway.
Exp Cell Res. 2014; 320(1):164-73 [PubMed] Related Publications
Mesenchymal stem cells (MSCs) are among the most important components of the osteosarcoma microenvironment and are reported to promote tumor progression. However, the means by which osteosarcoma cells modulate MSC behavior remains unclear. The aim of this study was to determine the effects of osteosarcoma cells on both the production of pro-tumor cytokines by mesenchymal stem cells (MSCs) and the osteogenic differentiation of MSCs. High level of transforming growth factor-β (TGF-β) was detected in three osteosarcoma cell lines. Conditioned media (CM) from the osteosarcoma cell lines Saos-2 and U2-OS were used to stimulate the cultured MSCs. We found that osteosarcoma cells promoted the production of IL-6 and VEGF in MSCs by inhibiting their osteogenic differentiation. Furthermore, TGF-β in tumor CM was proved to be an important factor. The TGF-β neutralizing antibody antagonized the effects induced by osteosarcoma CM. The inhibition of Smad2/3 by siRNA significantly decreased the production of IL-6 and VEGF in MSCs and induced their osteogenic differentiation. We also found that Smad2/3 enhanced the expression of β-catenin in MSCs by decreasing the level of Dickkopf-1 (DKK1). Although the inhibition of β-catenin did not affect the production of IL-6 or VEGF, or the gene expression of the early osteogenic markers Runx2 and ALP, it did enhance the gene expression of osteocalcin. Taken together, our data indicate that osteosarcoma cells secrete TGF-β to maintain the stemness of MSCs and promote the production of pro-tumor cytokines by these cells.

Related: Cytokines

Habel N, Hamidouche Z, Girault I, et al.
Zinc chelation: a metallothionein 2A's mechanism of action involved in osteosarcoma cell death and chemotherapy resistance.
Cell Death Dis. 2013; 4:e874 [PubMed] Free Access to Full Article Related Publications
Osteosarcoma is the most common primary tumor of bone occurring in children and adolescents. The histological response to chemotherapy represents a key clinical factor related to survival. We previously showed that statins exhibit antitumor effects in vitro, inducing apoptotic cell death, reducing cell migration and invasion capacities and strengthening cytotoxic effects in combination with standard drugs. Comparative transcriptomic analysis between control and statin-treated cells revealed strong expression of several genes, including metallothionein (MT) 2A. MT2A overexpression by lentiviral transduction reduced bioavailable zinc levels, an effect associated with reduced osteosarcoma cell viability and enhanced cell differentiation. In contrast, MT2A silencing did not modify cell viability but strongly inhibited expression of osteoblastic markers and differentiation process. MT2A overexpression induced chemoresistance to cytotoxic drugs through direct chelation of platinum-containing drugs and indirect action on p53 zinc-dependent activity. In contrast, abrogation of MT2A enhanced cytotoxic action of chemotherapeutic drugs on osteosarcoma cells. Finally, clinical samples derived from chemonaive biopsies revealed that tumor cells expressing low MT2A levels correspond to good prognostic (good responder patients with longer survival rate), whereas high MT2A levels were associated with adverse prognosis (poor responder patients). Taken together, these data show that MT2A contributes to chemotherapy resistance in osteosarcoma, an effect partially mediated by zinc chelation. The data also suggest that MT2A may be a potential new prognostic marker for osteosarcoma sensitivity to chemotherapy.

Related: TP53

Norimatsu Y, Ohsaki H, Masuno H, et al.
Efficacy of CytoLyt® hemolytic action on ThinPrep® LBC using cultured osteosarcoma cell line LM8.
Acta Cytol. 2014; 58(1):76-82 [PubMed] Related Publications
OBJECTIVE: The removal of blood components is necessary to improve the quality of the liquid-based cytology (LBC) preparations. In ThinPrep® (TP) samples a cell suspension in a methanol-based fixative undergoes a vacuum filtration method, whereas in SurePath™ (SP) samples a cell suspension in an ethanol-based fixative is processed through a density gradient centrifugation system prior to gravity deposition of the specimen onto a glass slide. We compared the cyto-architectural features for the cytologic diagnosis of endometrial adenocarcinoma using parallel TP and SP preparations in a previous publication.
STUDY DESIGN: We performed our study on LM8 cells (a cultured osteosarcoma cell line). LM8 cells at a concentration of 1.25 × 10(3) cell/cm(2) were seeded on a 35-mm plate in culture medium, which contained 10% fetal bovine serum (FBS), 100 units/ml penicillin, and 100 μ/ml streptomycin in Dulbecco's modified Eagle's medium (DMEM), and aliquots of the cell suspension obtained in this way were compared after the addition of a hemolytic agent, i.e. Cytolyt® (CyL). LBC preparations were then obtained on cell suspensions treated with CyL after different time intervals of hemolysis.
RESULTS: Treatment with CyL did not alter the cellularity of the preparation, but reduction of the nuclear area and a tendency towards nuclear chromatin condensation with a subsequent higher brightness were found. Because CyL is a 25% methanol-buffered solution, its alcoholic concentration is low; it was our impression that, while its fixative effect was weak, its hemolytic effect was high. Water influx or efflux through the cell membrane is controlled by osmotic pressure changes induced by the buffer solution in the CyL solution. While CyL was not shown to alter the cell shape, nuclear shrinkage was thought to be probably due to the increasing cell dehydration caused by longer exposure intervals to methanol.
CONCLUSION: This study has allowed us to make significant observations on the hemolytic properties of CyL, and on its combined effects with PreservCyt on the cytomorphology of cells suspensions.

Tavanti E, Sero V, Vella S, et al.
Preclinical validation of Aurora kinases-targeting drugs in osteosarcoma.
Br J Cancer. 2013; 109(10):2607-18 [PubMed] Article available free on PMC after 12/11/2014 Related Publications
BACKGROUND: Aurora kinases are key regulators of cell cycle and represent new promising therapeutic targets in several human tumours.
METHODS: Biological relevance of Aurora kinase-A and -B was assessed on osteosarcoma clinical samples and by silencing these genes with specific siRNA in three human osteosarcoma cell lines. In vitro efficacy of two Aurora kinases-targeting drugs (VX-680 and ZM447439) was evaluated on a panel of four drug-sensitive and six drug-resistant human osteosarcoma cell lines.
RESULTS: Human osteosarcoma cell lines proved to be highly sensitive to both drugs. A decreased drug sensitivity was observed in doxorubicin-resistant cell lines, most probably related to ABCB1/MDR1 overexpression. Both drugs variably induced hyperploidy and apoptosis in the majority of cell lines. VX-680 also reduced in vitro cell motility and soft-agar cloning efficiency. Drug association experiments showed that VX-680 positively interacts with all conventional drugs used in osteosarcoma chemotherapy, overcoming the cross-resistance observed in the single-drug treatments.
CONCLUSION: Aurora kinase-A and -B represent new candidate therapeutic targets for osteosarcoma. In vitro analysis of the Aurora kinases inhibitors VX-680 and ZM447439 indicated in VX-680 a new promising drug of potential clinical usefulness in association with conventional osteosarcoma chemotherapeutic agents.

Related: Bone Cancers

Giang AH, Raymond T, Brookes P, et al.
Mitochondrial dysfunction and permeability transition in osteosarcoma cells showing the Warburg effect.
J Biol Chem. 2013; 288(46):33303-11 [PubMed] Article available free on PMC after 15/11/2014 Related Publications
Metabolic reprogramming in cancer is manifested by persistent aerobic glycolysis and suppression of mitochondrial function and is known as the Warburg effect. The Warburg effect contributes to cancer progression and is considered to be a promising therapeutic target. Understanding the mechanisms used by cancer cells to suppress their mitochondria may lead to development of new approaches to reverse metabolic reprogramming. We have evaluated mitochondrial function and morphology in poorly respiring LM7 and 143B osteosarcoma (OS) cell lines showing the Warburg effect in comparison with actively respiring Saos2 and HOS OS cells and noncancerous osteoblastic hFOB cells. In LM7 and 143B cells, we detected markers of the mitochondrial permeability transition (MPT), such as mitochondrial swelling, depolarization, and membrane permeabilization. In addition, we detected mitochondrial swelling in human OS xenografts in mice and archival human OS specimens using electron microscopy. The MPT inhibitor sanglifehrin A reversed MPT markers and increased respiration in LM7 and 143B cells. Our data suggest that the MPT may play a role in suppression of mitochondrial function, contributing to the Warburg effect in cancer.

Related: Mitochondrial Mutations in Cancer

Kang K, Lee JH, Kim HG
Contralateral referred pain in a patient with intramedullary spinal cord metastasis from extraskeletal small cell osteosarcoma.
J Spinal Cord Med. 2013; 36(6):695-9 [PubMed] Article available free on PMC after 01/11/2014 Related Publications
CONTEXT: Referred pain has been observed in some patients after cordotomy, wherein noxious stimulus applied to a region rendered analgesic by cordotomy produces pain at a spot different from the one where the noxious stimulus is applied. We report a patient who had intramedullary spinal cord metastasis of extraskeletal small cell osteosarcoma, a rare form of metastatic disease, and experienced contralateral referred pain.
FINDINGS: Initially, the patient had a mass in the left posterior neck region and later developed a large extradural mass at the C3-C7 level. The masses were excised, and the histological findings led to a diagnosis of small cell osteosarcoma. He underwent chemotherapy and radiation therapy. He experienced numbness in his left leg; subsequently, the numbness slowly spread up the thigh to the left side of the abdomen. When pinched in the numb area on the left side of the body, he felt as though he had been pinched in both that area and the corresponding area on the right side. A magnetic resonance imaging scan showed an enhancing lesion in the right side of the cord at the C6-C7 level.
CONCLUSION/CLINICAL RELEVANCE: An intramedullary spinal cord metastasis can arise from primary extraskeletal small cell osteosarcoma and cause contralateral referred pain, especially in a mirror-image location. Contralateral referred pain may be caused by a subsidiary pathway comprising ascending chains of short neurons that link the dorsal horn neurons longitudinally and latitudinally.

Bianchi E, Artico M, Di Cristofano C, et al.
Growth factors, their receptor expression and markers for proliferation of endothelial and neoplastic cells in human osteosarcoma.
Int J Immunopathol Pharmacol. 2013 Jul-Sep; 26(3):621-32 [PubMed] Related Publications
Osteosarcoma is the most common primary malignant tumour of the bone. Although new therapies continue to be reported, osteosarcoma-related morbidity and mortality remain high. Modern medicine has greatly increased knowledge of the physiopathology of this neoplasm. Novel targets for drug development may be identified through an understanding of the normal molecular processes that are deeply modified in pathological conditions. The aim of the present study is to investigate, by immunohistochemistry, the localisation of different growth factors and of the proliferative marker Ki-67 in order to determine whether these factors are involved in the transformation of osteogenic cells and in the development of human osteosarcoma. We observed a general positivity for NGF - TrKA - NT3 - TrKC - VEGF in the cytoplasm of neoplastic cells and a strong expression for NT4 in the nuclear compartment. TGF-beta was strongly expressed in the extracellular matrix and vascular endothelium. BDNF and TrKB showed a strong immunolabeling in the extracellular matrix. Ki-67/MIB-1 was moderately expressed in the nucleus of neoplastic cells. We believe that these growth factors may be considered potential therapeutic targets in the treatment of osteosarcoma, although proof of this hypothesis requires further investigation.

Related: Bone Cancers MKI67 Signal Transduction

Jaing TH, Yang CP, Hung IJ, et al.
Phyllodes tumor in survivors of childhood osteosarcoma: a single institution's experience.
J Pediatr Hematol Oncol. 2014; 36(1):e36-8 [PubMed] Related Publications
We evaluate the incidence of second neoplasms in 86 patients with osteosarcoma (OS) of the extremities treated with different protocols of adjuvant chemotherapy. Three patients developed phyllodes tumors as the second neoplasm. One of these patients simultaneously developed a third cancer with therapy-related acute myeloid leukemia. The sites of primary OS were the tibia (2) and humerus (1). None had received prior radiotherapy before excision of phyllodes tumor. All the patients were female with a median age of 21.7 years at the time of presentation. As yet, that precise causation is unclear, but it can increase our understanding of carcinogenic processes, in general.

Related: Bone Cancers Acute Myeloid Leukemia (AML) Childhood Acute Myeloid Leukaemia AML - Molecular Biology

Lauvrak SU, Munthe E, Kresse SH, et al.
Functional characterisation of osteosarcoma cell lines and identification of mRNAs and miRNAs associated with aggressive cancer phenotypes.
Br J Cancer. 2013; 109(8):2228-36 [PubMed] Article available free on PMC after 15/10/2014 Related Publications
BACKGROUND: Osteosarcoma is the most common primary malignant bone tumour, predominantly affecting children and adolescents. Cancer cell line models are required to understand the underlying mechanisms of tumour progression and for preclinical investigations.
METHODS: To identify cell lines that are well suited for studies of critical cancer-related phenotypes, such as tumour initiation, growth and metastasis, we have evaluated 22 osteosarcoma cell lines for in vivo tumorigenicity, in vitro colony-forming ability, invasive/migratory potential and proliferation capacity. Importantly, we have also identified mRNA and microRNA (miRNA) gene expression patterns associated with these phenotypes by expression profiling.
RESULTS: The cell lines exhibited a wide range of cancer-related phenotypes, from rather indolent to very aggressive. Several mRNAs were differentially expressed in highly aggressive osteosarcoma cell lines compared with non-aggressive cell lines, including RUNX2, several S100 genes, collagen genes and genes encoding proteins involved in growth factor binding, cell adhesion and extracellular matrix remodelling. Most notably, four genes-COL1A2, KYNU, ACTG2 and NPPB-were differentially expressed in high and non-aggressive cell lines for all the cancer-related phenotypes investigated, suggesting that they might have important roles in the process of osteosarcoma tumorigenesis. At the miRNA level, miR-199b-5p and mir-100-3p were downregulated in the highly aggressive cell lines, whereas miR-155-5p, miR-135b-5p and miR-146a-5p were upregulated. miR-135b-5p and miR-146a-5p were further predicted to be linked to the metastatic capacity of the disease.
INTERPRETATION: The detailed characterisation of cell line phenotypes will support the selection of models to use for specific preclinical investigations. The differentially expressed mRNAs and miRNAs identified in this study may represent good candidates for future therapeutic targets. To our knowledge, this is the first time that expression profiles are associated with functional characteristics of osteosarcoma cell lines.

Related: Bone Cancers

Posthumadeboer J, Piersma SR, Pham TV, et al.
Surface proteomic analysis of osteosarcoma identifies EPHA2 as receptor for targeted drug delivery.
Br J Cancer. 2013; 109(8):2142-54 [PubMed] Article available free on PMC after 15/10/2014 Related Publications
BACKGROUND: Osteosarcoma (OS) is the most common bone tumour in children and adolescents. Despite aggressive therapy regimens, treatment outcomes are unsatisfactory. Targeted delivery of drugs can provide higher effective doses at the site of the tumour, ultimately improving the efficacy of existing therapy. Identification of suitable receptors for drug targeting is an essential step in the design of targeted therapy for OS.
METHODS: We conducted a comparative analysis of the surface proteome of human OS cells and osteoblasts using cell surface biotinylation combined with nano-liquid chromatography - tandem mass spectrometry-based proteomics to identify surface proteins specifically upregulated on OS cells. This approach generated an extensive data set from which we selected a candidate to study for its suitability as receptor for targeted treatment delivery to OS. First, surface expression of the ephrin type-A receptor 2 (EPHA2) receptor was confirmed using FACS analysis. Ephrin type-A receptor 2 expression in human tumour tissue was tested using immunohistochemistry. Receptor targeting and internalisation studies were conducted to assess intracellular uptake of targeted modalities via EPHA2. Finally, tissue micro arrays containing cores of human OS tissue were stained using immunohistochemistry and EPHA2 staining was correlated to clinical outcome measures.
RESULTS: Using mass spectrometry, a total of 2841 proteins were identified of which 156 were surface proteins significantly upregulated on OS cells compared with human primary osteoblasts. Ephrin type-A receptor 2 was highly upregulated and the most abundant surface protein on OS cells. In addition, EPHA2 was expressed in a vast majority of human OS samples. Ephrin type-A receptor 2 effectively mediates internalisation of targeted adenoviral vectors into OS cells. Patients with EPHA2-positive tumours showed a trend toward inferior overall survival.
CONCLUSION: The results presented here suggest that the EPHA2 receptor can be considered an attractive candidate receptor for targeted delivery of therapeutics to OS.

Related: Bone Cancers

Tian Q, Jia J, Ling S, et al.
A causal role for circulating miR-34b in osteosarcoma.
Eur J Surg Oncol. 2014; 40(1):67-72 [PubMed] Related Publications
PURPOSE: To investigate the associations between plasma miR-34b/c expression levels and osteosarcoma (OS).
SUBJECTS AND METHODS: A case-control study was conducted in 133 patients with OS and 133 controls. MiR-34b/c levels were detected by quantitative reverse transcriptase polymerase chain reaction (qRT-PCR) assays. Genotyping of SNP rs4938723 was done using the TaqMan assay. The causal association was examined by mendelian randomization analysis.
RESULTS: Plasma miR-34b level was significantly lower in OS patients than in controls (P = 0.001). Expression levels of miR-34b in OS tissues decreased (P = 3.22 × 10(-4)) and was significantly related with its expression in plasma (r = 0.21, P = 0.004). Compared with wild-type TT genotype, the variant genotypes of rs4938723 TC/CC were significantly associated with increased OS risk (TC vs. TT: OR, 1.97 [95% CI: 1.40-2.55], P = 0.021; CC vs. TT: OR, 2.76 [95% CI: 2.00-3.53], P = 0.009; TC + CC vs. TT: OR, 2.16 [95% CI: 1.61-2.70], P = 0.006), consistent with its decreased effect on plasma miR-34b (TC vs. TT: -0.32 (-0.43, -0.21), P < 0.001; CC vs. TT: -0.70 (-0.84, -0.56), P < 0.001; TC + CC vs. TT: -0.42 (-0.53, -0.32), P < 0.001). Adjustment for miR-34b completely abolished the association between SNP rs4938723 and OS risk (P > 0.05). In addition, plasma expression levels of miR-34b were significantly decreased in the metastatic patients compared with that in the non-metastatic ones (P = 0.004).
CONCLUSION: Plasma miR-34b was causally associated with OS risk and related with its metastatic status, suggesting that plasma miR-34b might be a novel biomarker and a potential treatment target for OS.

Related: Bone Cancers

Wang Z, Cai H, Lin L, et al.
Upregulated expression of microRNA-214 is linked to tumor progression and adverse prognosis in pediatric osteosarcoma.
Pediatr Blood Cancer. 2014; 61(2):206-10 [PubMed] Related Publications
BACKGROUND: MicroRNA-214 (miR-214) expression has been demonstrated to be dysregulated in human malignancies and to play various roles in tumor progression. While previous study of miRNA expression profiling found that it was one of the most upregulated miRNAs in osteosarcoma signature, the potential role of miR-214 in osteosarcomas has been unclear. Therefore, the aim of this study was to investigate association of miR-214 expression with clinicopathologic features and prognosis in pediatric patients with osteosarcoma.
PROCEDURE: Quantitative real-time reverse transcriptase-polymerase chain reaction analysis was performed to detect expression levels of miR-214 in cancerous and noncancerous bone tissues from 92 children treated for primary osteosarcomas. Then, the clinical significance of miR-214 dysregulation in pediatric osteosarcomas was also determined.
RESULTS: Compared with noncancerous bone tissues, the expression levels of miR-214 were significantly upregulated in osteosarcoma tissues (P < 0.001). High miR-214 expression occurred more frequently in osteosarcoma tissues with large tumor size (P = 0.01), positive metastasis (P = 0.001) and poor response to pre-operative chemotherapy (P = 0.006). Moreover, high miR-214 expression was significantly associated with both shorter overall (P < 0.001) and progression-free survival (PFS; P = 0.001). Multivariate analysis by the Cox proportional hazard model further confirmed that high miR-214 expression was an independent prognostic factor of unfavorable survival in pediatric osteosarcoma (for overall survival: P = 0.008; for PFS: P = 0.01).
CONCLUSION: Our data offer evidence that upregulated expression of miR-214 may be linked to tumor progression and adverse prognosis in pediatric osteosarcoma. Further investigation in prospective studies would appear warranted.

Related: Bone Cancers

Wen X, Liu H, Yu K, Liu Y
Matrix metalloproteinase 2 expression and survival of patients with osteosarcoma: a meta-analysis.
Tumour Biol. 2014; 35(1):845-8 [PubMed] Related Publications
A number of studies investigated the impact of matrix metalloproteinase 2 (MMP2) expression on the survival of patients with osteosarcoma, but no consistent results were reported. To derive a more precise estimate of the prognostic role of MMP2 expression in patients with osteosarcoma, we systematically reviewed the published studies and carried out a meta-analysis. Cohort studies assessing the prognostic role of MMP2 expression in patients with osteosarcoma were included. Pooled risk ratio (RR) with 95% confidence intervals (95%CI) was used to assess the prognostic role of MMP2 expression. Five cohort studies were eligible in the meta-analysis. Overall, high MMP2 expression was associated with increased risk of mortality in patients with osteosarcoma during the follow-up (fixed effects RR = 2.14, 95%CI 1.66-2.75, P < 0.001). Sensitivity analysis suggested that the pooled RR was stable and omitting a single study did not change the significance of the pooled RR. There was some possibility of publication bias risk in the meta-analysis. In conclusion, the meta-analysis suggests that osteosarcoma patients with high MMP2 expression have poorer prognosis compared with those with low MMP2 expression.

Related: Bone Cancers MMP2

Botchu R, Douis H, Davies AM, et al.
Post-traumatic heterotopic ossification of distal tibiofibular syndesmosis mimicking a surface osteosarcoma.
Clin Radiol. 2013; 68(12):e676-9 [PubMed] Related Publications
AIM: To present the imaging features of post-traumatic heterotopic ossification (HO) of the distal tibiofibular syndesmosis initially suspected to be a surface osteosarcoma.
MATERIALS AND METHODS: A retrospective review was conducted of the presenting complaint and imaging features of patients with a final diagnosis of HO referred over an 8 year period to a specialist orthopaedic oncology centre.
RESULTS: Five patients with HO were identified. All were adult males with an age range of 19-41 years. There was a history of prior ankle trauma in all cases but the significance was not recognized at the time of referral to the specialist centre. There was radiographic evidence of HO arising from the inner aspects of the distal tibia and fibula approximately 3 cm proximal to the ankle joint. The HO was "kissing" in two cases and partially fused (synostosis) in two. The HO in the fifth case was arising on the inner fibular cortex alone. Magnetic resonance imaging (MRI), available in four cases, showed predominantly low signal intensity due to the dense bone formation.
CONCLUSION: The history of prior ankle trauma with ossification arising from the inner aspects of both the distal tibia and fibula is typical of post-traumatic HO and distinguish this benign condition from the rare surface osteosarcoma at this site.

Related: Bone Cancers

Tome Y, Kimura H, Maehara H, et al.
High lung-metastatic variant of human osteosarcoma cells, selected by passage of lung metastasis in nude mice, is associated with increased expression of α(v)β(3) integrin.
Anticancer Res. 2013; 33(9):3623-7 [PubMed] Related Publications
Altered expression of αvβ3 integrin is associated with tumor progression and metastasis in several types of cancer, including metastatic osteosarcoma. In this study, we demonstrate that in vivo passaging of lung metastasis in nude mice can generate an aggressive variant of human osteosarcoma cells. Experimental metastases were established by injecting 143B human osteosarcoma cells, expressing green fluorescent protein (GFP) in the nucleus and red fluorescent protein (RFP) in the cytoplasm, in the tail vein of nude mice. Lung metastases were harvested under fluorescence microscopy from nude mice to establish cell lines which were then injected via the tail vein of additional nude mice. This procedure was repeated for four passages in order to isolate highly metastatic variant sublines. When the parental and metastatic variants were transplanted orthotopically into the tibia of nude mice, the 143B-LM4 variant had the highest metastatic rate, approximately 18-fold higher than the parent (p<0.01). αvβ3 integrin expression was increased approximately 5.6-fold in 143B-LM4 compared to parental cells (p<0.05). Thus, serial passage of lung metastases created a highly metastatic variant of human osteosarcoma cells which had increased expression of αvβ3 integrin, suggesting that αvβ3 integrin plays an essential role in osteosarcoma metastasis. With this highly metastatic variant overexpressing αvβ3 integrin, it will now be possible to further investigate the mechanism by which αvβ3 integrin facilitates metastasis.

Related: Bone Cancers

Fu HL, Shao L, Wang Q, et al.
A systematic review of p53 as a biomarker of survival in patients with osteosarcoma.
Tumour Biol. 2013; 34(6):3817-21 [PubMed] Related Publications
Osteosarcoma is the most common malignant bone tumor, and the prognosis of patients with osteosarcoma is still unsatisfactory with low survival rates. There are many studies assessing the prognostic role of upregulated p53 in patients presenting osteosarcoma, and there is no consistent finding. To summarize the existing evidence about whether the presence of upregulated p53 was a biomarker of survival in patients with osteosarcoma, we performed a systematic review and meta-analysis of relevant publications. We assessed the effect of upregulated p53 on the 3-year overall survival and the 3-year disease-free survival by calculating the pooled odds ratio (OR) with corresponding 95% confidence interval (95%CI). Fifteen studies with a total of 609 patients with osteosarcoma were finally included into the systematic review and meta-analysis. Compared with osteosarcoma patients with low or undetectable p53, patients with upregulated p53 were obviously associated with decreased 3-year overall survival (OR = 0.29, 95 %CI 0.19-0.43, P < 0.001). In addition, patients with upregulated p53 were obviously associated with decreased 3-year disease-free survival (OR = 0.06, 95 %CI 0.02-0.23, P < 0.001). The results from the systematic review and meta-analysis highlight that p53 is an effective biomarker of survival in patients with osteosarcoma. In addition, more studies with a large sample size are needed to identify the effect of p53 expression in osteosarcoma patients.

Related: Bone Cancers TP53

Zhao Z, Wu MS, Zou C, et al.
Downregulation of MCT1 inhibits tumor growth, metastasis and enhances chemotherapeutic efficacy in osteosarcoma through regulation of the NF-κB pathway.
Cancer Lett. 2014; 342(1):150-8 [PubMed] Related Publications
Monocarboxylate transporter isoform 1 (MCT1) is an important member of the proton-linked MCT family and has been reported in an array of human cancer cell lines and primary human tumors. MCT1 expression is associated with developing a new therapeutic approach for cancer. In this study, we initially showed that MCT1 is expressed in a variety of human osteosarcoma cell lines. Moreover, we evaluated the therapeutic response of targeting MCT1 using shRNA or MCT1 inhibitor. Inhibiting MCT1 delayed tumor growth in vitro and in vivo, including in an orthotopic model of osteosarcoma. Targeting MCT1 greatly enhanced the sensitivity of human osteosarcoma cells to the chemotherapeutic drugs adriamycin (ADM). In addition, we observed that MCT1 knockdown significantly suppressed the metastatic activity of osteosarcoma, including wound healing, invasion and migration. Further mechanistic studies revealed that the antitumor effects of targeting MCT1 might be related to the NF-κB pathway. Immunochemistry assay showed that MCT1 was an independent positive prognostic marker in osteosarcoma patients. In conclusion, our data, for the first time, demonstrate that MCT1 inhibition has antitumor potential which is associated with the NF-κB pathway, and high MCT1 expression predicates poor overall survival in patients with osteosarcoma.

Related: Bone Cancers Signal Transduction

Poos K, Smida J, Nathrath M, et al.
How microRNA and transcription factor co-regulatory networks affect osteosarcoma cell proliferation.
PLoS Comput Biol. 2013; 9(8):e1003210 [PubMed] Article available free on PMC after 15/10/2014 Related Publications
Osteosarcomas (OS) are complex bone tumors with various genomic alterations. These alterations affect the expression and function of several genes due to drastic changes in the underlying gene regulatory network. However, we know little about critical gene regulators and their functional consequences on the pathogenesis of OS. Therefore, we aimed to determine microRNA and transcription factor (TF) co-regulatory networks in OS cell proliferation. Cell proliferation is an essential part in the pathogenesis of OS and deeper understanding of its regulation might help to identify potential therapeutic targets. Based on expression data of OS cell lines divided according to their proliferative activity, we obtained 12 proliferation-related microRNAs and corresponding target genes. Therewith, microRNA and TF co-regulatory networks were generated and analyzed regarding their structure and functional influence. We identified key co-regulators comprising the microRNAs miR-9-5p, miR-138, and miR-214 and the TFs SP1 and MYC in the derived networks. These regulators are implicated in NFKB- and RB1-signaling and focal adhesion processes based on their common or interacting target genes (e.g., CDK6, CTNNB1, E2F4, HES1, ITGA6, NFKB1, NOTCH1, and SIN3A). Thus, we proposed a model of OS cell proliferation which is primarily co-regulated through the interactions of the mentioned microRNA and TF combinations. This study illustrates the benefit of systems biological approaches in the analysis of complex diseases. We integrated experimental data with publicly available information to unravel the coordinated (post)-transcriptional control of microRNAs and TFs to identify potential therapeutic targets in OS. The resulting microRNA and TF co-regulatory networks are publicly available for further exploration to generate or evaluate own hypotheses of the pathogenesis of OS (http://www.complex-systems.uni-muenster.de/co_networks.html).

Related: Bone Cancers

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