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Although hematogenous spread of osteosarcoma is well known, the imaging findings of cardiovascular involvement by osteosarcoma are seldom reported and can be difficult to recognize. The enhanced resolution of modern CT and MRI scanners may lead to better detection of cardiovascular involvement. To describe the key imaging findings and clinical behavior of cardiovascular involvement by osteosarcoma. We retrospectively reviewed the imaging findings and clinical characteristics of 20 patients with cardiovascular involvement by osteosarcoma identified by two pediatric radiologists from a review of imaging studies at our institution from 2007 to 2013. At initial diagnosis, the median age of the patients was 15.1 years (range 4.8–24.6 years), and 7 (35%) patients had detectable metastases. Median time to detection of cardiovascular metastases was 1.8 years (range 0–7.3 years). Sixteen patients died of disease; 4 have survived a median of 7.4 years since initial diagnosis. The sites of cardiovascular involvement were the systemic veins draining the primary and metastatic osteosarcoma, pulmonary arteries, pulmonary veins draining the pulmonary metastases, and heart. A dilated and mineralized terminal pulmonary arteriole is an early sign of metastatic osteosarcoma in the lung. Unfamiliarity with the imaging features resulted in under-recognition and misinterpretation of intravascular tumor thrombus as bland thrombus. Knowledge of imaging findings in the era of modern imaging modalities has enhanced our ability to detect cardiovascular involvement and lung metastases early and avoid misinterpreting tumor thrombus in draining systemic veins or pulmonary arteries as bland thrombus.

The pediatric bone sarcomas osteosarcoma and Ewing sarcoma represent a tremendous challenge for the clinician. Though less common than acute lymphoblastic leukemia or brain tumors, these aggressive cancers account for a disproportionate amount of the cancer morbidity and mortality in children, and have seen few advances in survival in the past decade, despite many large, complicated, and expensive trials of various chemotherapy combinations. To improve the outcomes of children with bone sarcomas, a better understanding of the biology of these cancers is needed, together with informed use of targeted therapies that exploit the unique biology of each disease. Here we summarize the current state of knowledge regarding the contribution of receptor tyrosine kinases, intracellular signaling pathways, bone biology and physiology, the immune system, and the tumor microenvironment in promoting and maintaining the malignant phenotype. These observations are coupled with a review of the therapies that target each of these mechanisms, focusing on recent or ongoing clinical trials if such information is available. It is our hope that, by better understanding the biology of osteosarcoma and Ewing sarcoma, rational combination therapies can be designed and systematically tested, leading to improved outcomes for a group of children who desperately need them.

PURPOSE Survival of patients with osteosarcoma lung metastases has not improved in 20 years. We evaluated the efficacy of combining natural killer [NK] cells with aerosol interleukin-2 [IL-2] to achieve organ-specific NK cell migration and expansion in the metastatic organ, and to decrease toxicity associated with systemic IL-2.

Purpose The purpose of this study was to characterize the complications associated with surgical treatment of pediatric melanoma. Methods We retrospectively reviewed all pediatric patients who received surgical treatment for melanoma at our institution between 1992 and 2010. We compared complications between three groups: wide local excision only (WLE), WLE and sentinel lymph node biopsy (SLNB), and WLE and completion lymph node dissection (CLND). Results One hundred twenty-five patients were identified: 37 patients received WLE only, 47 received WLE and SLNB, and 41 patients had WLE and CLND. Complication rates differed between the three groups: 19% in WLE, 11% in WLE+SLNB, and 39% in WLE+CLND ( P =.006). The risk of complications was significantly lower among patients having WLE+SLNB versus WLE+CLND (OR 0.19, 95% CI 0.06–0.57, P =.0032). Lymphedema was a common complication with a higher incidence in the CLND group compared to the SLNB group (19.5% vs. 2.1%, P =.01). Complications were more frequent in inguinal compared to axillary dissections (52.0% vs. 17.1%, P =.006). Conclusions In the surgical treatment of pediatric melanoma, the addition of a completion lymph node dissection significantly increases complication risk. Thus, it is critical to determine which patients truly benefit from this procedure.

BACKGROUND: The UBE4B gene, which is located on chromosome 1p36, encodes a ubiquitin ligase that interacts with hepatocyte growth factor-regulated tyrosine kinase substrate (Hrs), a protein involved in epidermal growth factor receptor (EGFR) trafficking, suggesting a link between EGFR trafficking and neuroblastoma pathogenesis. The authors analyzed the roles of UBE4B in the outcomes of patients with neuroblastoma and in neuroblastoma tumor cell proliferation, EGFR trafficking, and response to EGFR inhibition. METHODS: The association between UBE4B expression and the survival of patients with neuroblastoma was examined using available microarray data sets. UBE4B and EGFR protein levels were measured in patient tumor samples, EGFR degradation rates were measured in neuroblastoma cell lines, and the effects of UBE4B on neuroblastoma tumor cell growth were analyzed. The effects of the EGFR inhibitor cetuximab were examined in neuroblastoma cells that expressed wild-type and mutant UBE4B. RESULTS: Low UBE4B gene expression is associated with poor outcomes in patients with neuroblastoma. UBE4B overexpression reduced neuroblastoma tumor cell proliferation, and UBE4B expression was inversely related to EGFR expression in tumor samples. EGFR degradation rates correlated with cellular UBE4B levels. Enhanced expression of catalytically active UBE4B resulted in reduced sensitivity to EGFR inhibition. CONCLUSIONS: The current study demonstrates associations between UBE4B expression and the outcomes of patients with neuroblastoma and between UBE4B and EGFR expression in neuroblastoma tumor samples. Moreover, levels of UBE4B influence neuroblastoma tumor cell proliferation, EGFR degradation, and response to EGFR inhibition. These results suggest UBE4B-mediated growth factor receptor trafficking may contribute to the poor prognosis of patients who have neuroblastoma tumors with 1p36 deletions and that UBE4B expression may be a marker that can predict responses of neuroblastoma tumors to treatment. Cancer 2013. © 2012 American Cancer Society.

PURPOSE OF REVIEW The survival curves for many pediatric sarcomas have remained flat for the past 2 decades or more and novel therapeutics - those small molecule medicines that selectively inhibit specific signaling molecules - have been slow to enter into pediatric practice. The preclinical basis for their use is reviewed here. RECENT FINDINGS Preclinical and phase I studies showing efficacy of antiinsulin-like growth factor receptor 1 therapies for Ewing sarcoma have led to numerous ongoing clinical trials using these agents for Ewing and other sarcomas. Early studies of ERBB signaling as a target in sarcoma therapy have been tantalizing, but progress in this area has been controversial. In-vitro analysis of Src inhibitors suggested that these agents would prevent metastasis in osteosarcoma, whereas in-vivo analysis showed no effect on metastasis, underscoring the need for thorough preclinical investigations of promising new therapies to guide future clinical trials. Antiangiogenic and immunomodulatory therapies are gaining momentum in the pediatric arena and should be tested in combination with traditional cytotoxic agents for recurrent and high-risk primary pediatric sarcomas. SUMMARY Pediatric sarcomas have diverse biology and distinct signaling pathways, making detailed preclinical evaluation of small molecule inhibitors essential to guiding the design of further clinical investigations.

Cetuximab, which blocks ligand binding to epidermal growth factor receptor (EGFR), is currently being studied as a novel treatment for non–small cell lung cancer (NSCLC). However, its mechanisms of action toward metastasis, and markers of drug sensitivity, have not been fully elucidated. This study was conducted to (a) determine the effect of Cetuximab on invasion and NSCLC-metastasis; (b) investigate urokinase-type plasminogen activator receptor (u-PAR), a major molecule promoting invasion and metastasis, as a target molecule; (c) delineate molecular mediators of Cetuximab-induced metastasis inhibition; and (d) identify biomarkers of drug sensitivity in NSCLC. Cetuximab treatment resulted in reduced growth and Matrigel invasion of H1395 and A549 NSCLC cell lines, in parallel with reduced u-PAR mRNA and protein. u-PAR down-regulation was brought about by suppressing the binding of JunD and c-Jun to u-PAR promoter motif −190/−171 in vivo, and an inhibition of MAP/ERK kinase signaling. Furthermore, Cetuximab inhibited NSCLC proliferation and metastasis to distant organs in vivo as indicated by the chicken embryo metastasis assay. Low E-cadherin and high u-PAR, but not EGFR, was associated with resistance to Cetuximab in seven NSCLC cell lines. Furthermore, siRNA knockdown of u-PAR led to a resensitization to Cetuximab. Moreover, low E-cadherin and high u-PAR was found in 63% of resected tumor tissues of NSCLC patients progressing under Cetuximab therapy. This is the first study to show u-PAR as a target and marker of sensitivity to Cetuximab, and to delineate novel mechanisms leading to metastasis suppression of NSCLC by Cetuximab. [Cancer Res 2009;69(6):2461–70]

New investigational agents and chemotherapy regimens including cyclophosphamide + topotecan, temozolomide + irinotecan, and anti-IGF-1R antibodies in Ewing's sarcoma (ES) and liposomal muramyltripeptide phosphatidylethanolamine (L-MTP-PE), aerosol therapy, and bone-specific agents in osteosarcoma (OS) may improve survival and/or quality of life on 'continuation' therapy.Review of investigational approaches and control paradigms for recurrent or metastatic primary bone tumors.Analyze temozolomide + irinotecan data and review in the context of other newer approaches including antiangiogenesis, anti-IGF-1R antibodies and bisphosphonates for ES. Review some current state-of-the-art approaches for OS including L-MTP-PE, anti-IGF-1R inhibition, aerosol therapies and bone specific agents.L-MTP-PE with chemotherapy in OS has been shown to improve survival; compassionate access is available for recurrence and/or metastases. Aerosol therapy (granulocyte-macrophage colony stimulating factor, cisplatin, gemcitabine) for lung metastases is a promising approach to reduce systemic toxicity. The bone-specific agents including denosumab (anti-receptor activator of NF-kappaB ligand antibody) and bisphosphonates may have benefit against giant cell tumor, ES and OS. Anti-IGF-1R antibody SCH717454 has preclinical activity in OS but best effectiveness will most likely be in combination with chemotherapy earlier in therapy. Both temozolomide + irinotecan and cyclophosphamide + topotecan combinations are very active in ES and are likely to be tested with anti-IGF-1R antibodies against ES.

Although sarcoma surgery is very important for cancer control, it is not always possible or practical to offer in some situations, including sarcoma recurrences, metastatic disease and/or unacceptable loss of function. We review some pragmatic approaches and examples of how to balance indications, risks and alternatives to control cancer in young people with sarcomas that are no longer using ‘front-line’ therapy. Radiotherapy combined with chemotherapy and outpatient ‘continuation’ chemotherapy regimens using drugs that cause less alopecia can improve function and quality of life. Some effective strategies to help cope when cure is not possible may include tumor ablation techniques performed in interventional radiology and percutaneous nerve blocks. Family centered care and effective problem solving of difficult issues can be greatly facilitated by consultation with a multidisciplinary team experienced in the management of very difficult cases. Treatment of young people with recurrent, relapsed and/or metastatic sarcoma still remains an art very much in the realm of compassion not protocol and persistent advocacy is required for the young person for whom cure may not be possible. A reduction of suffering and assistance in writing more chapters of a rich life narrative is the goal.

Purpose Chemotherapy during radiation and/or bone-seeking radioisotope therapy (153-samarium; 1 mCi/kg) during radiation may improve osteosarcoma cancer control. Patients and Methods We analyzed our preliminary radiation experience in high-risk, metastatic, and/or recurrent patients during a consecutive period of 20 months (May 2005–December 2006). Results Thirty-nine high-risk osteosarcoma patients had radiotherapy; 119 sites were irradiated. A median four sites were irradiated per patient (range 1–14). The median radiation dose and number of fractions of radiation was 30 Gy in 10 fractions (range 10–70 Gy in 4–35 fractions). Chemotherapy, most commonly ifosfamide or methotrexate, was used in 80% (100/119) radiotherapy courses. Of 38 painful sites, 29 had improvement (76%), 4 had no change (10%), and 5 had more pain (13%). Objective and potentially durable responses were documented using PET-CT and bone scans with persistent and sustained reduction of standard uptake values (SUVs; initial SUV of indication lesion 9.5 became

Causes of lytic bone lesions in children include benign, malignant, and infectious processes. Here, we present the case of a 3-year-old boy presenting with a lytic bone lesion and surrounding soft tissue mass sent for evaluation of possible malignancy versus osteomyelitis. Biopsy revealed granulomatous osteomyelitis, and subsequent purified protein derivative resulted in 20-mm induration. Lesion cultures eventually identified pan-sensitive Mycobacterium tuberculosis. We emphasize that tuberculosis can cause primary lytic bone lesions in children in the United States, even in the absence of pulmonary symptoms or known exposure, and advise clinicians to include mycobacterial cultures when analyzing biopsies of lytic bone lesions.

To evaluate the impact of treated extra-pulmonary metastatic disease on overall (OS) and event-free survival (EFS) for pediatric osteosarcoma patients undergoing pulmonary metastatectomy.We retrospectively reviewed pediatric patients who were treated for osteosarcoma at our institution from 2001 to 2011 and received pulmonary metastatectomy (n=76). We compared OS and EFS between patients with metastases limited to the lungs (Group A, n=58) to those with treated extra-pulmonary metastases (Group B, n=18) at the time of first pulmonary metastatectomy.The estimated median OS and EFS from first pulmonary metastatectomy were 2.0years (95% CI 1.5-2.8years) and 5.5months (95% CI 3.0-8.1months), respectively. Median OS was significantly greater for Group A (2.6years, 95% CI 1.9-3.8) compared to Group B (0.9years, 95% CI 0.6-1.5) (log rank p=0.0001). Median EFS was significantly greater for Group A (7.9months, 95% CI 5.0-10.7) compared to Group B (1.6months, 95% CI 0.8-2.7) (log rank p0.0001). Independent predictors of OS included extra-pulmonary metastatic disease at the time of first thoracotomy, bilateral pulmonary metastases, and4 nodules resected at first thoracotomy (all p0.001).Osteosarcoma patients with treated extra-pulmonary metastatic disease at the time of pulmonary metastatectomy have significantly worse survival compared to those with disease limited to the lungs.

The purpose of this study was to identify health disparities in children with non-CNS solid tumor malignancies and examine their impact on disease presentation and outcome.We examined the records of all children (age≤18years) diagnosed with a non-CNS solid tumor malignancy and enrolled in the Texas Cancer Registry between 1995 and 2009 (n=4603). The primary outcome measures were disease stage and overall survival (OS). Covariates included gender, age, race/ethnicity, year of diagnosis, socioeconomic status (SES), and driving distance to the nearest pediatric cancer treatment facility. Statistical analyses included life table methods, logistic, and Cox regression. Statistical significance was defined as p0.05.Children with advanced-stage disease were more likely to be male,10years old, and Hispanic or non-Hispanic Blacks (all p0.05). Distance to treatment and SES did not impact stage of disease at presentation. However, Hispanic and non-Hispanic Blacks and patients in the lowest SES quartile had the worst 1- and 5-year survival (all p0.05). The adjusted OS differed by age, race, and stage, but not SES or distance to the nearest treatment facility.Race/ethnicity plays an important role in survival for children with non-CNS solid tumor malignancies. Future work should better define these differences to establish mechanisms to decrease their impact.

The two most common primary bone malignancies, osteosarcoma and Ewing sarcoma, are both aggressive, highly metastatic cancers that most often strike teens, though both can be found in younger children and adults. Despite distinct origins and pathogenesis, both diseases share several mechanisms of progression and metastasis, including neovascularization, invasion, anoikis resistance, chemoresistance and evasion of the immune response. Some of these processes are well-studies in more common carcinoma models, and the observation from adult diseases may be readily applied to pediatric bone sarcomas. Neovascularization, which includes angiogenesis and vasculogenesis, is a clear example of a process that is likely to be similar between carcinomas and sarcomas, since the responding cells are the same in each case. Chemoresistance mechanisms also may be similar between other cancers and the bone sarcomas. Since osteosarcoma and Ewing sarcoma are mesenchymal in origin, the process of epithelial-to-mesenchymal transformation is largely absent in bone sarcomas, necessitating different approaches to study progression and metastasis in these diseases. One process that is less well-studied in bone sarcomas is dormancy, which allows micrometastatic disease to remain viable but not growing in distant sites – typically the lungs – for months or years before renewing growth to become overt metastatic disease. By understanding the basic biology of these processes, novel therapeutic strategies may be developed that could improve survival in children with osteosarcoma or Ewing sarcoma.

Bone sarcomas cause disproportionate morbidity and mortality and desperately need new therapies as there has been little improvement in outcomes in 20 years. Identification of critical signaling pathways, including type 1 insulin-like growth factor receptor (IGF-1R) for Ewing sarcoma and possibly osteosarcoma, and the ERBB and the Wnt signaling pathways for osteosarcoma, have emerged as receptors mediating vital signals for bone sarcoma. Akt, mammalian target of rapamycin (mTOR), phosphoinositide 3-kinases, mitogen-activated protein kinase kinase, extracellular signal-regulated kinases, and Ras pathway play key roles in at least some tumors, and inhibition of mTOR in particular will likely lead to improved survival, although clinical trials are still underway. The Notch pathway and ezrin are essential for osteosarcoma metastasis, and Fas down-regulation is necessary for survival of metastases in lungs. As little is known about chondrosarcoma signaling, more preclinical work is needed. By defining vital signaling pathways in bone sarcomas, small molecule inhibitors can be applied rationally, leading to longer survival and reducing morbidity and late effects from intensive chemotherapy.

Background: Conventional therapy for osteosarcoma has reached a plateau of 60 – 70%, a 5-year survival rate that has changed little in two decades, highlighting the need for new approaches. Objective: To review the alternative means of delivering effective therapy for osteosarcoma that reach beyond the central venous catheter. Methods: Drawing on the author's own experiences providing care to high-risk osteosarcoma patients and reviewing the last two decades of literature describing sarcoma therapy, available information is summarized about potential osteosarcoma treatments that deliver therapy by a less conventional route. Results/conclusions: Intra-arterial chemotherapy has a limited impact on survival, but may help to achieve a better limb salvage. Intrapleural chemotherapy is important for managing malignant effusions. The development of inhalation therapies, treatments that target new bone formation such as bisphosphonates, chemically targeted radiation and antibody-based therapies all have potential...

Osteosarcoma (OS) is the most common primary bone cancer in children, and despite more than 20 years of clinical trials, the 5-year survival for OS patients remains essentially unchanged (∼70% for non-metastatic patients and 30% for metastatic patients), with the vast majority deaths arising from metastasis to lungs or bones. We wanted to identify signaling pathways that mediate tumor survival and metastasis in OS, and we focused on Her-4 (ERBB4), which promoted survival on neuroblastoma cells in our previous work. Purpose: To evaluate the role of Her-4 in OS cell survival, metastasis and “stemness”. Methods: Her-4 expression was measured by flow cytometry, western blot and Q-PCR. Validated OS cell lines were manipulated with shRNA to knock-down Her-4 (compared to scrambled control). In vitro responses were measured by varying culture conditions (normal, high-density or anchorage-independent growth), serum starvation, hypoxia or chemotherapy (methotrexate, cisplatin, doxorubicin and 4-OH-ifosfamide) and measuring proliferation, cell cycle, and apoptosis (by both flow cytometry and PARP cleavage). A sarcosphere assay assessed expression of “stemness” markers, or tumor initiating cells (TICs), and Her-4 expression. The luciferase-labeled CCH-OS-O xenograft model was used to assess the impact of Her-4 knock-down on metastatic potential, with weekly luciferase measures (IVIS 100, Xenogen) assessing tumor burden. Immunohistochemistry (IHC) assessed Her-4 expression in archival material and murine xenografts. Results: Most OS cells express Her-4 which upregulated with increased culture density and with serum starvation. RT-PCR showed that the proportion of cleavable isoforms increased with increasing cell density (p < .05 with One-Way ANOVA analysis), while non-cleavable isoforms decreased (p < .05). Hypoxia increased the cleavage of Her-4 to the 80KD intracellular fragment without increasing Her-4 expression. IHC analysis of OS tissue microarrays showed that Her-4 expression is higher in metastatic lesions relative to primary tumors (p>0.01). Knock-down of Her-4 in OS cell lines reduced proliferation and increased apoptosis (by sub-diploid DNA and PARP cleavage) for cells grown in conventional culture, high density or spheres. Her-4 knock-down increased sensitivity to methotrexate (60% increase in apoptosis) but not other chemotherapies. To assess the role of Her-4 in metastasis, NOD/SCID/IL2Rg-/- mice were injected in the tibia with CCH-OD-O cells expressing luciferase and either Her-4 shRNA or scrambled control. With this model, a “primary” tumor arises in the tibia, with spontaneous metastasis to the lungs. Non-invasive imaging showed reduced growth of primary tumors in knock-down mice and complete elimination of tumor signal from lungs. Xenograft lungs showed a significant reduction in metastasis in knock-down mice. IHC for Her-4 showed that, while tibial tumors of knock-down mice still expressed no Her-4, the tumors present in these mice had re-expressed Her-4 despite the shRNA. Stress resistance, chemotherapy resistance and metastatic potential are TIC-associated behaviors, so we examined the association of Her-4 with other described TIC markers for OS. Sarcosphere culture induced expression of both Her-4 and stemness markers CD117 and Stro1 over a period of 2-7 days. Her-4 expression preceded upregulation of other markers by 1-2 days. Conclusions: Expression of Her-4 confers a survival benefit to OS cells under stressed conditions, including anoikis, serum starvation and methotrexate exposure. Her-4 is necessary for OS metastasis. Her-4 expression increases in sarcophere cultures and may be a mediator of stemness. Therapies targeting Her-4 in OS should be evaluated in further preclinical models. Citation Format: Rocio K. Rivera-Valentin, Yingqi Hua, Yi Zhang, Nupur Lala, Yanwen Yang, Dennis P.M. Hughes. Her-4 mediates anoikis resistance, chemoresistance, and metastatic potential in osteosarcoma. [abstract]. In: Proceedings of the AACR Special Conference on Pediatric Cancer at the Crossroads: Translating Discovery into Improved Outcomes; Nov 3-6, 2013; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2013;74(20 Suppl):Abstract nr A71.

Osteosarcoma is a highly invasive bone malignancy in which metastasis accounts for the vast majority of death and morbidity in patients. Understanding the mechanisms controlling metastasis is essential for improving patient survival in this disease. In order to improve the clinical outcomes for patients with poor prognosis, it is urgent to find new therapeutic targets to block metastasis in this disease. Recent studies have shown that Metadherin (MTDH) plays an essential role in mediating tumorigenesis and metastasis in a variety of human cancers. Our study assessed the role of MTDH in osteosarcoma metastasis and elucidated the mechanisms underlying its metastasis-promoting activity. To evaluate the expression of MTDH in primary and metastatic lesions of osteosarcoma, two tissue microarrays containing patient-derived primary and metastatic tumor specimens were examined by immunohistochemical staining with anti-MTDH antibody. We also examined MTDH in a cDNA array expression database made from pretreatment diagnostic biopsies of high-grade osteosarcoma patients to further assess the correlation between MTDH expression and clinical outcome. We used western blot, qPCR, and flow cytometry to measure the expression of MTDH in a panel of osteosarcoma cell lines. In parallel experiments we used MTDH-specific shRNA to reduce endogenous MTDH expression, and blocked cell surface MTDH by anti-MTDH antibodies. The impact of MTDH inhibition was assessed in vitro using transwell migration assays and matrigel invasion assays. In addition, we developed an orthotopic xenograft mouse model to study the relationship between MTDH expression and osteosarcoma pulmonary metastasis. To investigate the role of MTDH in cell-extracellular matrix (ECM) interaction and to identify the extracellular binding partner for cell surface MTDH, a series of adhesion assays were performed, followed by bidirectional co-immunoprecipitation. We have demonstrated that MTDH is upregulated in human osteosarcoma cell lines and patient-derived specimens compared with normal human osteoblasts. Overexpression of MTDH is more profound in metastatic lesions compared to primary tumors and is correlated with poor clinical outcomes in osteosarcoma patients. MTDH knockdown and blockade of cell surface MTDH significantly reduced migration and invasion in osteosarcoma cells. In the in vivo experiments, downregulation of MTDH in osteosarcoma cells delayed primary tumor growth and prohibited pulmonary metastasis. Both in vitro and in vivo studies confirmed the critical role of MTDH in the invasive and metastatic capacity of osteosarcoma cells. More importantly, we have identified the significance of cell surface localization of MTDH in mediating osteosarcoma motility and invasiveness. We showed that MTDH exists as a type II membrane protein in osteosarcoma cells and its expression on cell surface is facilitating cell invasion by means of modulating cell adhesion to the ECM through interaction with laminin. In total, these observations establish MTDH as a promising target for therapeutic interventions in metastatic osteosarcoma. The novel connection between MTDH and extracellular laminin also establishes a new paradigm for the function of MTDH in mediating tumor cell metastasis. Citation Format: Limin Zhu, Pingyu Zhang, Yanwen Yang, Adrianna S. Buford, Wei-Lien Wang, Dafydd G. Thomas, Dennis P.M. Hughes. Metadherin functions as a laminin receptor that is essential for metastasis and is associated with poor survival in osteosarcoma. [abstract]. In: Proceedings of the AACR Special Conference on Pediatric Cancer at the Crossroads: Translating Discovery into Improved Outcomes; Nov 3-6, 2013; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2013;74(20 Suppl):Abstract nr A53.

Osteosarcoma is the most common primary tumor of bones. In the past three decades treatment paradigms and survival rates have not improved. While osteosarcoma clinical phenotype has been well characterized, the underlying tumor molecular biology remains unclear. We have previously demonstrated that IL-11Rα is overexpressed in primary and metastatic osteosarcoma. Recently investigators identified that IL-11 and IL-11Rα play a role in the development of metastasis in breast and gastrointestinal cancer. However the role of IL-11/IL-11Rα signaling in osteosarcoma is unknown. Using in vitro and in vivo osteosarcoma models we investigated the role of IL-11/IL-11Rα in osteosarcoma metastasis. The purpose of our study is to determine the effect of the knockdown of IL-11Rα in osteosarcoma. We stably knocked down IL-11Rα expression in a highly metastatic osteosarcoma cell line, using a lentiviral short hair-pin (shRNA) based approach. Knockdown of IL-11Rα led to inhibition of STAT-3(Tyr705) protein phosphorylation, cell growth, migration, invasion and colony formation in vitro. To determine the role of IL-11Rα in osteosarcoma tumorigeniesis and metastasis, we used an orthotopic metastatic model. In this model, mice injected with KRIB-IL-11Rα shRNA cells developed significantly less metastasis than control mice injected with KRIB control shRNA (P Citation Format: Valerae O. Lewis, Eswaran Devarajan, Dennis PM Hughes. Targeting IL-11Rα inhibits osteosarcoma pulmonary metastasis in an orthotopic xenograft mouse model. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 1261. doi:10.1158/1538-7445.AM2014-1261

Osteosarcoma is the most common type of bone malignancy in children. It is a highly invasive mesenchymal tumor in which metastasis accounts for the vast majority of mortality and morbidity in patients. Understanding the mechanisms controlling metastasis in osteosarcoma and developing novel therapeutic options targeting metastasis are essential for improving patient survival in this disease. Recent studies have shown that Metadherin (MTDH) plays a key role in mediating metastasis in a variety of human cancers. This study examined the role of MTDH in osteosarcoma metastasis and elucidated the mechanisms underlying its oncogenic activity. We used western blot, qPCR, and flow cytometry to measure the expression of MTDH in a panel of osteosarcoma cell lines. Patient-derived osteosarcoma samples were also examined for MTDH expression using immunohistochemistry. In parallel experiments we used MTDH-specific shRNA to down-regulate endogenous MTDH expression, and blocked cell surface MTDH by anti-MTDH antibodies. The impact of MTDH inhibition was assessed in vitro using migration assays and matrigel invasion assays. We also developed an osteosarcoma orthotopic xenograft model of NOD/SCID/IL2Rγ-deficient mice through intratibial injection of osteosarcoma cells with or without MTDH knockdown to study the relationship between MTDH expression and pulmonary metastatic potential of osteosarcoma cells. Compared to normal human osteoblasts, all osteosarcoma cell lines tested displayed elevated MTDH expression as shown by western blot, qPCR and flow cytometry analyses. MTDH knockdown reduced migration and decreased invasion in matrigel. Inhibition of cell surface MTDH with an anti-MTDH antibody targeting the extracellular domain of MTDH also profoundly impeded osteosarcoma cell motility and invasiveness in vitro. In an orthotopic xenograft mouse model using CCHD cells, inhibition of MTDH delayed primary tumor growth and prohibited pulmonary metastasis. All mice in the control group developed lung metastases while fewer than 20% of the mice in the MTDH-knockdown group showed sign of metastasis at six weeks after inoculation. MTDH knockdown also reduced mean lung weight and mean number of pulmonary metastasis. Immunohistochemical analysis of tumor samples showed that lung metastases had elevated MTDH expression level compared to primary tumors. Altogether these studies suggested that MTDH plays a critical role in promoting pulmonary metastasis of osteosarcoma and may serve as an attractive target for therapeutic intervention. Citation Format: Limin Zhu, Adrianna S. Buford, Yanwen Yang, Pingyu Zhang, Wei-Lien Wang, Dennis P. Hughes. Metadherin mediates osteosarcoma invasion and metastasis. [abstract]. In: Proceedings of the AACR Special Conference on Tumor Invasion and Metastasis; Jan 20-23, 2013; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2013;73(3 Suppl):Abstract nr B67.

The human epidermal growth factor receptor 2 (Her2) is overexpressed in multiple types of tumors, including breast tumors and osteosarcoma. Tumor-targeted antibody therapy is one of the safest therapeutics available, but it is often ineffective in treating primary and acquired antibody-resistant tumor cells via indirect tumor cell death and inducing direct tumor cell death. This study reveals that Herceptin conjugates, if generated via the crosslinker EDC (1-ethyl-3-(3-dimethylaminopropyl) carbodimide hydrochloride), is capable of causing Her2 positive tumor cell death. Using an HPLC system, two peaks with estimated molecular weights of antibody dimers and trimers were isolated. The Herceptin dimer and trimer separated by HPLC, especially the trimer, induces significant levels of necrotic tumor cell death. Unlike SMCC conjugated Herceptin, this study indicates that the EDC-conjugated Herceptin overcomes the acquired resistance. The Herceptin trimer is more effective than the dimer in enhancing Herceptin-resistant tumor cell death. Surprisingly different from the known cell death mechanism often resulting from antibody treatment, the Herceptin trimer causes effective and direct tumor cell death via a novel cell death mechanism: programmed cell necrosis. Inhibition of RIP, a critical regulator of programmed necrotic cell death, reversed the Herceptin trimer-induced cell death. In summary, the Herceptin conjugate especially trimer linked by EDC could overcome Her2 positive breast cancer or osteosarcoma cells as well as the resistant cell lines via program cell necrosis cell death. This conjugate has great potential for overcoming the tumor cell resistance to the Herceptin treatment. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 4559. doi:10.1158/1538-7445.AM2011-4559

10060 Background: Sarcoma metastasizing to the heart is a difficult and complicated clinical problem, yet local control is feasible and effective for select patients with cardiac metastasis. Diligence is required to detect these lesions while still amenable to treatment, and echocardiogram remains the best tool for detecting cardiac disease. Methods: We reviewed all echocardiograms (1330) performed on 307 pediatric sarcoma patients treated at UT M. D. Anderson Cancer Center between 1997 and 2008. Measures of cardiac function and pathology (including size of ventricles and atria, function of ventricles and valves, presence of metastasis, and presence of pericardial effusion) were assigned numerical values. Fisher's exact test and the Wilcoxon rank-sum test compared clinical characteristics of patients with and without cardiac metastases. Results: The prevalence of cardiac metastases was 1.6% (5/307) with 95% confidence interval 0.5% to 3.8%. The presence of cardiac metastasis positively correlated with pericardial effusion (p = 0.001) and tricuspid valve insufficiency (p = 0.014). The probability of a patient with pericardial effusion having a cardiac metastasis was 28.6% (95% CI: 3.7%-71%). Of the 5 patients with documented sarcoma metastasis to the heart, 3 had widespread refractory disease, were given no cardiac-specific therapy, and rapidly died from disease. One patient who had widespread disease controlled with chemotherapy and radiation had open resection of 2 cardiac metastases which resulted in site-specific disease control for 6 months before succumbing to progressive extra-cardiac disease. Another patient had isolated cardiac metastasis, treated with open resection of 2 metastases followed by adjuvant chemotherapy, and has been without recurrent cardiac metastases for 5 years and 9 months. Conclusions: Durable local control for sarcoma metastasizing to the heart is possible and effective for select patients. Pericardial effusion merits evaluation for potential associated cardiac metastasis. Echocardiogram monitoring of sarcoma patients remains important, and should include screening for possible cardiac metastasis. No significant financial relationships to disclose.

10036 Background: Extrapulmonary osteosarcoma (OS) metastases portend poor outcome and short survival. Local control is essential for durable remission. Intracardiac, mediastinal and CNS metastases...

20006 Background: Epithelial salivary gland neoplasms (ESGN) are uncommon tumors that mainly occur in adults, only 5 percent of ESGNs occur in children. Data regarding differences in clinical presentation and biologic behavior of these tumors between adult and pediatric patients remain unclear. There is less data concerning the role of radiation therapy in the treatment of ESGNs in pediatric patients. Methods: Retrospective review of medical records of patients presenting under age 18 years from January 1988 to August 2006. Results: 45 patients were evaluable. 26 patients were female. Median age at diagnosis was 14.7 years. Mucoepidermoid carcinoma (MEC) was seen in 22 cases (49%), acinic cell carcinoma 10 cases (22%), adenocystic carcinoma (ACC) 8 cases, and pleomorphic adenoma (PA) 5 cases. The parotid area was the most common involved site (25/45; 56%) with MEC and acinic cell noted in 80% of parotid cases. 76% of MEC cases (16/21) were grade 2 or 3 tumors. Initial complete resection was achieved in 20 ESGN cases, while the remaining cases required re-excision or adjuvant therapy. Thirteen patients received postoperative radiation. Median age was 15.8 years. Radiation therapy was used in 6 ACC cases, 4 MEC and 3 adenoid cystic. Primary site and first echelon nodes dose range was 54–64 Gy, and supraclavicular dose was 50 Gy in those in whom regional nodes were treated. Positive margin status and local invasion (perineural, lymphatic, or nodal) were the most common indications. Mean follow-up for 35 patients was 60.3 months (2–175 months). Nine patients were lost to follow- up, and one patient received therapy at another institution. Two patients died from progressive disease, and both had received radiation therapy. Conclusions: MEC is the most common ESGN in pediatrics, and the parotid is the most common involved site. Complete surgical resection remains the treatment of choice. Radiation therapy may be used as adjuvant therapy, but more study is required. No significant financial relationships to disclose.

19502 Background: It has been suggested that sedentary behavior during cancer treatment or effects of the treatment itself may decrease cardiorespiratory fitness in cancer survivors. However, few studies have tested whether cancer survivors actually have impaired cardiorespiratory fitness compared to similarly aged individuals who have not had cancer. We investigated whether endometrial cancer survivors had poorer fitness than women of similar age and activity level with no cancer history. Methods: The sample included endometrial cancer survivors in a pilot exercise study; all had been diagnosed with Stage I-IIIa disease and were at least 6 months post- treatment. Controls were recruited from a local multi-speciality clinic. Both groups had no contraindications to exercise and were sedentary. Cardiorespiratory fitness was measured by predicted maximal myocardial oxygen carrying capacity (MV02 max) on a submaximal ramped cycle ergometer test. Quality of life was measured using the SF-36 (physical and mental component scores). Results: The study accrued 20 survivors (Mean age=58.6, SD=7.9; Mean BMI=30.5, SD=7.7) and 13 controls (Mean age=55.8, SD=7.6; Mean BMI=30.3, SD=7.0). In t-tests comparing survivors and controls, there were significant differences in predicted MV02 max (p=0.044) and physical aspects of quality of life (p=0.004), but not mental aspects of quality of life. After adjusting for age and BMI using ANCOVA, the p value for predicted MV02 max increased to 0.068, but differences in physical quality of life remained significant (see Table 1 ). Conclusions: Endometrial cancer survivors have diminished quality of life in the physical domain compared to women who have not had cancer; similar results were found for cardiorespiratory fitness, although the effect was attenuated after adjusting for BMI and age. The results suggest endometrial cancer survivors could benefit from exercise interventions to improve fitness and physical functioning. Table 1 : Fitness and quality of life of endometrial cancer survivors and controls. [Table: see text] No significant financial relationships to disclose.