Metastatic Ewing Sarcoma Research Articles

S2027 Ewing's Sarcoma: A Difficult Diagnosis to Swallow

INTRODUCTION: Ewing sarcoma family of tumors (EFT) compromises of a spectrum of similar histological undifferentiated cells. EFT includes Ewings Sarcoma (ES) and Primitive Neuroectodermal Tumor (PNET). These tumors are rare and aggressive with a tendency of early metastatic spread. Peak incidence are between ages of 10-15 years of age; however, 30% can occur in adults over the age of 20. EFT affect mainly Caucasians and are extremely uncommon in other races. Most common origin occurs from the bone of the extremities and axial skeleton. These tumors are not commonly found in the gastrointestinal tract. Our case demonstrates a rare presentation of metastatic Ewing Sarcoma found during evaluation for dysphagia. CASE DESCRIPTION/METHODS: 69 year old female who is a daily smoker with history of large cell lymphoma (previously treated with Adriamycin) and Barrett's esophagus presented with dysphagia. Labs were relevant for iron deficiency anemia. EGD revealed a fungating mass in an area of Barretts esophagus. The mass was not bleeding, non obstructing and partially circumferential. Biopsies from the mass revealed Ewing sarcoma (ES) based on histological stains. Microscopically the cells were arranged in cords suggestive of a neuroectodermal tumor. Fluorescent in situ hybridization (FISH) testing was negative for EWSR fusion protein. Cells were strongly positive for CD-99 and weakly positive for FLI1 also highly suggestive of ES. Further imaging revealed metastatic disease to the liver. Liver lesions were also biopsied confirming diagnosis of extra-skeletal Ewings Sarcoma. DISCUSSION: Soft tissue Ewing Sarcoma patients are older and more likely to be female. These tumors do express high levels of cell surface glycoproteins such as CD99, MIC2, and p30. MIC2 expression is a sensitive diagnostic marker for ES however it lacks specificity. Typically ES can be diagnosed with immunohistochemical stains for CD99 in the right clinical setting. However, if the presentation is atypical, such as in our case, further confirmatory testing with Next Generation Sequencing to detect translocation or point mutations are needed. Because of the vast differentials molecular genetics plays a large role in diagnosis. Translocations in EWSR1 gene on chromosome 22q12 are found in 85% of cases and can be detected using FISH analysis. This case illustrates the importance of timely establishment of type of esophageal cancer in order to obtain prompt molecular genetic assessments in order to confirm histological diagnosis.Figure 1.: Lower esophageal fungating, non obstructing, partially circumferential mass found on EGD.Figure 2.: The cells are small to medium sized, and are arranged in cords, interspersed with smooth muscle fibers suggestive of a neuroectodermal tumor.Figure 3.: Immunohistochemical stain positive for CD99 (cell surface glycoprotein) found in Ewings Sarcoma.

LBA68 Results of the randomized, placebo (PL)-controlled phase II study evaluating the efficacy and safety of regorafenib (REG) in patients (pts) with metastatic relapsed Ewing sarcoma (ES), on behalf of the French Sarcoma Group (FSG) and UNICANCER

REGOBONE is non-comparative phase II, double-blind, PL-controlled trial designed to evaluate the activity and safety of REG, in 5 independent cohorts of sarcoma originating in bone. We report here the ES cohort results. Key-eligibility criteria were histologically confirmed diagnosis of bone ES, age ≥10 years, confirmed measurable PD not amenable to curative-intent, 1-2 previous chemotherapy (CT) regimens for metastatic disease, ECOG 0-1. Eligible ES pts were randomized (2:1) to receive either REG (160 mg/d, 21/28 d) or matched PL with optional cross-over at the time of centrally confirmed progressive disease (PD). 24 pts were planned in the REG arm based on a A'Hern's single-stage design for phase II trials (1-sided α=0.05, and 80% power) to detect a 27% improvement in the progression-free rate (PFR) at 8 weeks (P0=40%) as evaluated by central review per RECIST1.1. Secondary endpoints included PFS, OS and safety. From September 2014 to November 2019, 41 ES pts were included. Five pts were not eligible for efficacy analysis. Of 36 efficacy-evaluable pts (13 in PL arm and 23 in REG arm); 28 were men, median age was 32 (16-59) years. 13/23 pts in REG arm (56.5%; one-sided CI95% = [37.5-[) were non-progressive at 8 weeks vs. 1/13 pts in PL arm (7.7%; CI95% = [0.4-[). Median PFS was 11.4 (CI95% = 4.6-22.9) vs. 3.9 (CI95%= 3.3-7.3) weeks for REG and PL arms, respectively. 5 (21.7%) partial responses were observed on REG. Median OS was 34.9 (CI95%=17.6-58.7) and 30.4 (CI95%=10.0-NE) weeks for REG and PL arms, respectively. Ten out of 13 pts crossed-over to REG after centrally-confirmed PD on PL. The most common ≥ Gr3 REG-related AEs during the double blind period were diarrhea (13%), hand-foot skin reaction (13%), asthenia (9%), thrombocytopenia (9%), mucosal inflammation (9%) and febrile neutropenia (9%), with one toxic death due to thrombocytopenia. Despite a PFR at 8 weeks lower than expected, this randomized non comparative study shows a promising signal of benefit of REG in relapsed ES, with a median of PFS of 11.4 weeks, and a moderate toxicity.

Abstract 6158: The relevance of endoglin and MMP14 to the metastatic potential of Ewing sarcoma cells

Abstract Background: Ewing sarcoma (ES) is a developmental bone/soft tissue neoplasia, for which mesenchymal stem cell (MSC) is the putative cell of origin. Although the outcome of ES patients bearing primary tumors has improved significantly, patients with metastatic ES still have a poor prognosis. The MSC marker endoglin/CD105 (ENG) and the ENG-shedding protease MMP14 have been, respectively, described as cell migration and invasion regulators. In our attempt to better understand the mechanism of ES progression, we are focusing on the role of ENG and MMP14 in ES cells. Methods: In vitro ENG-knockdown (ENG-KD; RM82 and SKNMC cell lines) and ENG-overexpressing (pENG; TC71 cell lines) models were generated by shRNA technology and pDisplay vectors, respectively, and confirmed by qRT-PCR, western blotting and flow cytometry. In vitro MMP14-knockout (MMP14-KO; RM82 and SKNMC) models were generated by CRISPR-Cas9 and confirmed by western blotting and Sanger sequencing. Transwell migration/invasion, adhesion, clonogenicity and proliferation assays were performed in vitro. Actin filament organization was evaluated by immunofluorescence. Extracellular vesicles (EVs) were isolated by ultracentrifugation and density gradient separation. Transcriptomic profiling was performed by Clarion S Human arrays followed by Gene Set Enrichment Analysis. HiPathia tool was used to recode the transcriptomic data into measurements of cell activity by applying mechanistic models. Reverse Phase Protein Array (RPPA) analysis was evaluated on the ENG-KD RM82 model. ENG-targeting immunoprecipitations (IP) from wild type RM82 protein extracts were analyzed by mass spectrometry (MS). Results: The impaired clonogenic potential of ENG-KD RM82 and SKNMC models suggests a role of ENG on the stemness capacity of ES, without affecting the proliferation rate. The reduced migratory and adhesive phenotype of the ENG-KD RM82 model and the transcriptomic profiling of both ENG-KD RM82 and SKNMC cells, suggest a role of ENG in ES cell migration and adhesion. The altered organization of actin filaments in ENG-KD and pENG models could support a role of ENG in actin filament remodeling. The transcriptomic profiling of ENG-KD models and the RPPA results underscore the regulation of FAK signaling by ENG. The IP-MS analysis of ENG partners identifies Integrin-Linked Kinase-Associated serine/threonine Phosphatase (ILKAP) as one of the main interactors of ENG in RM82 cells, suggesting its possible association with the focal adhesion member ILK. ENG is enriched in EVs from different ES cell lines. The transcriptomic profiling of MMP14-KO SKNMC cells highlights the involvement of MMP14 activity in the extracellular matrix organization of ES cells. Conclusion: ENG favors a pro-migratory phenotype in ES cells. These findings help to better understand the molecular landscape of tumoral cells from ES patients, and might have potential clinical implications. Citation Format: Pilar Puerto-Camacho, Ana Teresa Amaral, Juan Diaz-Martin, Alfonso Bolado-Carrancio, Carmen Jordan-Perez, Carmen Salguero-Aranda, Marina Esteban-Medina, Inmaculada Alamo-Alvarez, Joaquin Dopazo, Carmelo Bernabeu, Adam Byron, Valerie G. Brunton, Enrique de Álava. The relevance of endoglin and MMP14 to the metastatic potential of Ewing sarcoma cells [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 6158.

Abstract DDT02-03: AMG 509: A novel, humanized, half-Life extended, bispecific STEAP1 × CD3 T cell recruiting XmAb® 2+1 antibody

Abstract AMG509, a novel STEAP1 x CD3 T cell-recruiting XmAb® 2+1 bispecific immune therapy for the treatment of prostate cancer and Ewing sarcoma. Metastatic castration resistant prostate cancer (mCRPC) and Ewing sarcoma (EWS) are both diseases with unmet medical needs. Six Transmembrane Epithelial Antigen of the Prostate 1 (STEAP1) is a cell surface antigen that is induced by androgens and highly expressed in prostate cancer. STEAP1 is also highly expressed in EWS, and it is a target of the EWS/FLI1 fusion, the primary oncogenic driver in this pediatric malignancy. AMG 509 is a novel, half-life extended, cynomolgus monkey cross-reactive, STEAP1 x CD3 T cell-recruiting XmAb® 2+1 bispecific antibody designed to treat STEAP1-expressing cancers. AMG 509 contains two identical humanized anti-STEAP1 Fab domains that bind STEAP1-expressing cells, an anti-CD3 scFv domain that binds T cells, and an Fc domain engineered to lack effector function and extend serum half-life. We found that membranous STEAP1 expression, as measured by IHC, was absent from most normal tissues (FDA normal organ tissue array, n = 72) except the prostate epithelium and a subset of lung macrophages. In contrast, we found membranous STEAP1 expression by IHC in 80% of primary prostate cancer tumors (n = 80) and moderate to high expression in 83% of metastatic prostate cancer lesions (n = 181) and 77% of bone metastases (n = 31). In addition, we found STEAP1 expression by IHC in all EWS cell lines (n=13) and xenograft models tested (n=12). AMG 509 bound to recombinant human and cynomolgus monkey CD3ϵ with a KD of 27.6 nM and 25.8 nM, respectively, and to 293T cells transfected with human or cynomolgus monkey STEAP1 with an EC50 of 3.8 nM and 3.1 nM, respectively. AMG 509 triggered potent T cell-redirected lysis of STEAP1-positive cancer cells, with a median EC50 of 18.9 pM across 17 different STEAP1-expressing prostate, EWS, gastric, and melanoma cancer cell lines. AMG 509 was 50-fold more potent in inducing the redirected lysis of C4-2B prostate cancer cells in vitro than an XmAb® molecule with a single anti-STEAP1 Fab domain. AMG 509 was also specific, as it showed no activity against C4-2B cancer cells engineered to lack STEAP1 expression. AMG 509 exhibited hallmarks of T-cell engagement in vitro, including T-cell proliferation and cytokine release. It also induced robust in vivo anti-tumor activity in prostate cancer and EWS xenograft models, with concomitant CD8+ T-cell activation and expansion in tumors. Data from a nonclinical safety study conducted in the cynomolgus monkey suggest that AMG 509 can be administered safely at doses that are predicted to be clinically active. AMG 509 is a potent and specific first-in-class T cell-recruiting antibody for the treatment of STEAP1-positive malignancies. We are currently evaluating the safety, tolerability, pharmacokinetics, and efficacy of AMG 509 in a phase 1, first-in-human study in patients with mCRPC (NCT04221542). Citation Format: Olivier Nolan-Stevaux. AMG 509: A novel, humanized, half-Life extended, bispecific STEAP1 × CD3 T cell recruiting XmAb® 2+1 antibody [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr DDT02-03.

High Response Rates and Promising Outcomes of Patients with Relapsed Ewing Sarcoma, Especially in Adolescents and Young Adults Treated on a Novel Hybrid Salvage Chemotherapy Regimen

Purpose: About 30%-35% of nonmetastatic and 60%-80% of metastatic Ewing Sarcoma (ES) will relapse post-treatment and outcomes after relapse continue to be poor over last several decades. Prognostic factors affecting survival after relapse of ES are also not robustly known. We present outcomes using a novel hybrid salvage protocol of four active chemotherapeutic agents in our cohort of patients after relapse of ES. Methods: This is a retrospective analysis of all consecutive relapsed ES patients treated with curative intent over 4 years (January 2012 to December 2015). All received 12-cycles of hybrid chemotherapy regimen with surgery/radiotherapy done after first 4 cycles. Event-free survival (EFS)/overall survival (OS) estimates were analyzed by Kaplan-Meier product-limit estimator. Cox regression analysis was performed to identify prognostic factors predicting outcome in relapsed ES. Results: Salvage regimen was given to 53/108 relapsed ES patients with the rest having opted for palliation upfront. Median age of the treated patients was 19 years (range: 4-40); male:female ratio was 2.7:1. Median time to first relapse was 18.8 months (range: 2.2-91). While 41/53 patients (77%) completed salvage therapy, 6 (11.3%) progressed and 6 (11.3%) abandoned treatment. Median follow-up of the study cohort is 31 months (range: 4-81). Of the analyzable cohort (n = 47), 30 (64%) had a second relapse or progression on salvage treatment. At last follow-up, 31 patients had died (including one due to toxicity and rest due to disease) and 16 patients were alive (14 with no active disease and 2 with disease). The 4-year EFS and OS are 28% and 37%, respectively, for the entire cohort. While adolescents and young adult patients (AYA) had a better survival (p-0.041), relapsed ES patients with shorter disease-free interval (DFI) (<24 months) had a poorer survival (p-0.004). The type of relapse (local or metastatic or combined) after primary treatment did not affect outcome after salvage therapy. Conclusions: We have used a novel hybrid chemotherapy protocol using four active agents in relapsed ES, which is well tolerated and shows promising results. Older age (≥15 years) and longer DFI (>24 months) portend better survival post-relapse. In our cohort of relapsed ES, AYAs fared better than others and type of relapse after primary treatment did not affect outcome after salvage therapy.

Abstract A48: YB-1-based oncolytic virotherapy in combination with CDK4/6-inhibitors against Ewing sarcoma

Abstract Oncolytic virotherapy is associated with immunogenic cell death and antitumor inflammatory immune responses. Thereby, a local as well as systemic immunosuppressive microenvironment might be abrogated, resulting in efficient phagocytic and antigen-presenting properties. Consecutively, priming of naive T cells against a multitude of de-repressed tumor and viral antigens is claimed to elicit strong antitumor immune responses accompanied by tumor-infiltrating T cells, especially in combination with immune checkpoint blockade. These features might be beneficial for patients with relapsed and metastatic Ewing sarcoma, who have poor prognosis and lack innovative therapy concepts, and whose tumor biopsies are characterized by low or absent T-cell infiltration. Here, we explore the efficacy of the oncolytic YB-1-selective adenovirus XVir-N-31 in combination with the CDK4/6-inhibitor abemaciclib (LY2835219) in established Ewing sarcoma cell lines. We demonstrate enhanced viral replication, particle formation, and oncolysis when combined with abemaciclib in vitro. Priming of tumor cells with abemaciclib before viral infection leads to cell cycle arrest in G1 phase, with consecutive decrease in Rb, pRb, and E2F1 protein levels. Interestingly, in the Rb-mutated hence CDK4/6-inhibitor-resistant cell line SK-N-MC, the combination with abemaciclib does not result in enhanced viral replication oor oncolysis. However, the same effects as for CDK4/6-inhibitor sensitive cell lines can be induced by siRNA-mediated E2F1 knockdown, indicating the crucial role of E2F1 as a repressor for initiation of viral replication. In summary, our results further support the hypothesis of E2F1 being a repressor of adenoviral replication (Holm et al., manuscript in preparation) and suggest the use of CDK4/6-inhibitors to boost oncolytic adenoviral efficacy. Further in vivo confirmation is planned in order to prepare a clinical phase I study of XVir-N-31 in combination with a CDK4/6-inhibitor and immune checkpoint blockade for treatment-refractory pediatric sarcoma patients. Citation Format: Sebastian J. Schober, Uwe Thiel, Melanie Thiede, Maximilian Ehrenfeld, Roman Nawroth, Guenther HS Richter, Stefan E.G. Burdach, Per Sonne Holm. YB-1-based oncolytic virotherapy in combination with CDK4/6-inhibitors against Ewing sarcoma [abstract]. In: Proceedings of the AACR Special Conference on the Advances in Pediatric Cancer Research; 2019 Sep 17-20; Montreal, QC, Canada. Philadelphia (PA): AACR; Cancer Res 2020;80(14 Suppl):Abstract nr A48.

Abstract A64: Functional genomics of metastatic Ewing sarcoma

Abstract Ewing sarcoma (ES) is a poorly differentiated bone and soft tissue tumor of high metastatic potential. ES mainly affects children, adolescents, and young adults (AYAs) at a frequency of ~1.5 cases per million globally. Ewing neoplasms strikingly converge on a single recurrent initiating event, which is a chromosomal translocation that generates a fusion transcript between the EWSR1 gene and a gene of the ETS family of transcription factors, most commonly FLi1 (85%). After 20 years since the discovery of the EWS-FLi1 fusion, ES remains a clinical challenge with unacceptably low survival rates, primarily due to metastasis. The bulk of research conducted to date (&amp;gt;95%) being focused on the primary tumor has resulted in a critical knowledge gap. To address this issue and unravel the dysregulated pathways in ES tumor evolution and metastatic dissemination, we harnessed the genome-wide insertional mechanism of transposons and the transduction efficiency of lentiviruses to engineer ES cell models. Human mesenchymal stem cells (hMSC), the putative cells of origin of ES, were engineered to constitutively express EWS-Fli1 accompanied by the inducible expression of a highly active transposase. Upon activation of the former, transposon-mediated mutagenesis will activate oncogenes and inactivate tumor suppressor genes, to mediate transformation of the transduced that can now engraft when implanted in recipient mice. Xenografted tumors are resected and the mice observed for development of distant metastases. Matching primary and metastatic tumors are sequenced to uncover the genes commonly affected by transposition in the two compartments. Pathway-oriented bioinformatic analysis will further reveal candidate metastatic driver genes. Matching of the targeted pathways with drugs will be used to validate the candidates by in vitro and in vivo metastasis assays. Citation Format: Wajih Jawhar, Paul Waterhouse, Rama Khokha, Takeaki Ishii, Robert Turcotte, Nada Jabado, Livia Garzia. Functional genomics of metastatic Ewing sarcoma [abstract]. In: Proceedings of the AACR Special Conference on the Advances in Pediatric Cancer Research; 2019 Sep 17-20; Montreal, QC, Canada. Philadelphia (PA): AACR; Cancer Res 2020;80(14 Suppl):Abstract nr A64.

The Role of Tyrosine Kinases as a Critical Prognostic Parameter and Its Targeted Therapies in Ewing Sarcoma.

Ewing sarcoma (ES) is a rare, highly aggressive, bone, or soft tissue-associated tumor. Although this sarcoma often responds well to initial chemotherapy, 40% of the patients develop a lethal recurrence of the disease, with death recorded in 75–80% of patients with metastatic ES within 5 years, despite receiving high-dose chemotherapy. ES is genetically well-characterized, as indicated by the EWS-FLI1 fusion protein encoded as a result of chromosomal translocation in 80–90% of patients with ES, as well as in ES-related cancer cell lines. Recently, tyrosine kinases have been identified in the pathogenesis of ES. These tyrosine kinases, acting as oncoproteins, are associated with the clinical pathogenesis, metastasis, acquisition of self-renewal traits, and chemoresistance of ES, through the activation of various intracellular signaling pathways. This review describes the recent progress related to cellular and molecular functional roles of tyrosine kinases in the progression of ES.

Primary and Metastatic Intracranial Ewing Sarcoma at Diagnosis: Retrospective International Study and Systematic Review.

Intracranial Ewing sarcoma (EwS) is rare and publications on primary or metastatic intracranial EwS are minimal. The aim of this study was to describe incidence, clinical behavior, treatment, and factors associated with outcome in patients with primary intracranial EwS or patients with a primary extracranial EwS and cerebral metastases at diagnosis. We reviewed all patients with primary or with metastatic intracranial EwS at diagnosis registered in the International Clinical Trial Euro-E.W.I.N.G.99 (EE99). In total, 17 of 1435 patients (1.2%) presented with primary intracranial EwS; 3 of them had metastatic disease. Four patients (0.3%) with primary extracranial EwS presented with intracranial metastatic lesions. The 3-year event-free survival (EFS) was 64% and overall survival (OS) was 70% in patients with a primary intracranial EwS. Local control in patients with primary intracranial EwS consisted of surgery (6%), radiotherapy (RT) (18%), or both modalities (76%). Univariate analysis showed that patients < 15 years of age had significantly better outcome (EFS: 72%; OS: 76%) compared to those aged above 15 years (EFS: 13%; OS: 25%). In conclusion, primary intracranial EwS and extracranial EwS with cerebral metastases at diagnosis is rare, yet survival is comparable with local and metastatic EwS elsewhere in the body. Age and stage of disease are important prognostic factors. Besides chemotherapeutic treatment, local control with surgical resection combined with RT is recommended whenever feasible.

Maintenance therapy with oral cyclophosphamide plus celecoxib in patients with metastatic Ewing sarcoma: Results of the Italian Sarcoma Group/AIEOP EW-2 study.

10517 Background: The prognosis of patients with metastatic Ewing sarcoma remains poor. The primary aim of the ISG/AIEOP EW2 Study (EUDRACT# 2009-011197-15) was to evaluate the feasibility and efficacy of maintenance therapy with oral cyclophosphamide plus celecoxib. Methods: From June 1st 2009 to Nov 22nd 2019, 112 patients with metastatic Ewing sarcoma at onset entered the ISG/AIEOP EW2 study, consisting of induction chemotherapy, radiotherapy and/or surgery at the site of the primary tumor, a consolidation phase with high-dose busulphan/melphalan + autologous stem cell rescue, whole-lung irradiation (12-15Gy), and a maintenance phase of 180 days with cyclophosphamide 50 mg daily (35 mg/mq daily if age &lt; 14 years) plus celecoxib 400 mg twice daily (250 mg/mq twice daily if age &lt; 14 years). Exclusion criteria from the maintenance phase were disease progression, cardiac or gastro-intestinal comorbidity. For CTCAE v4.0 grade 3-4 toxicities a temporary interruption was planned. Results: Seventy-one patients were eligible and entered the maintenance phase. Median age was 16 years (range 13-41); sites of metastases were lung or single bone (n = 56) and multicentric metastatic spread (n = 15). Sixty-one patients terminated the maintenance phase, 4 patients are still on treatment, 1 patient interrupted the treatment due to auto-immune thrombocytopenia at 4 months, 5 patients were withdrawn throughout maintenance due to disease progression/relapse. The duration of maintenance therapy was 89% of the scheduled days, with a median suspension length of 12 days (range 1-44 days). Causes of temporary suspension were hematological toxicity (19 episodes), infections (12 episodes), gastrointestinal disorders (9 episodes), fluid retention/distal oedema (3 episodes), renal disorders (3 episodes). Median follow-up was 42 months. The 3-year EFS of patients who entered the maintenance phase was 0.79 ± 0.09 for lung or single bone, and 0.19 ± 0.11 for those with multicentric metastatic spread. Conclusions: This schedule of maintenance phase is feasible, despite previous intensive treatment. A longer follow-up is needed to monitor side effects and to evaluate clinical outcome of patients with lung or single bone metastases, while the outcome remains dismal for multicentric metastatic Ewing sarcoma. Clinical trial information: NCT02727387.

The outcome of vincristine, dactinomycin D, ifosfamide and doxorubicin (VAIA) as first-line therapy for adult-patients with metastatic ewing sarcoma; a single-center experience.

e23501 Background: Ewing sarcoma family of tumors (ESFT) is a rare malignancy among adults. Most studies from western countries have reported improvement in outcomes following multi-agent chemotherapy. We report our experience in the management of this disease among Arab ethnicity. The aim of this study is to assess the outcome of VAIA combination as a first-line treatment in Adult-patients with metastatic Ewing sarcoma. Methods: Patients who were newly diagnosed as metastatic Ewing sarcoma between 03/1997 and 11/2016 at King Faisal Specialist Hospital and Research Center, who received VAIA as first-line therapy were eligible. The patient's characteristics were summarized using Medians with interquartile ranges (IQR) and frequencies for continuous and categorical variables, respectively. Variables including age, sex, primary tumor size, site (skeletal vs extraskeletal) and extent of metastasis in correlation with progression and overall survival were analyzed using the Kaplan–Meier method and Cox proportional hazards regression. Results: Thirty-nine patients were identified. Male (26, 66.7%), Female (13, 33.3%). Skeletal (27, 69.2%), Extraskeletal (12, 30.8%). The median longest diameter of the primary tumor 9.75 (IQR 8-15). The most common metastatic sites were Lungs (22, 56.4%) &amp; Bone (10, 25.6%), however, the least common sites were Bone Marrow (3, 7.7%) and liver (2, 5.1%). The median number of VAIA cycles was 10 cycles (IQR 5-14). Objective response rate (ORR) was noticed in 20 patients (51.2%) (Complete Remission (7, 17.9%), Partial Remission (13, 33.3%). One patient had stable disease and 12 (30.8%) patients had progressive disease. The assessment was not feasible in 3 (7.7%) patients. With a median follow up duration of 18 months (1-44).20 patients died (62.5%). The median PFS and OS was 10,18 months respectively with 3,5 years overall survival rate of;35.7%,26.9% respectively. Univariate analysis correlation among different variables were insignificant. Conclusions: VAIA chemotherapy combination showed poor outcomes among our patients in comparison to literature further improvement is needed in this aggressive malignancy in our region.

Activity of front-line window therapy with temozolomide plus irinotecan in patients with primary multifocal Ewing sarcoma: ISG/AIEOP EW-2 protocol.

10516 Background: The prognosis of patients with primary multifocal metastatic Ewing sarcoma (PMES) remains dismal. So far, combination with temozolomide and irinotecan (TEMIRI) was tested in patients with refractory or relapsed disease. This study evaluates the activity and the tolerability of TEMIRI as front-line treatment in PMES. Methods: In the study-period 2012-2018, a front-line window therapy with 2 courses TEMIRI (temozolomide 100 mg/sqm/day + irinotecan 50 mg/sqm/day for 5 days every three weeks) was introduced as amendment to the ISG/AIEOP EW-2 protocol ( EUDRACT#2009-011197-15, Vers. 1.02 ) for patients with PMES. Main objective was to test the activity of TEMIRI evaluated by RECIST 1.1 criteria, with centralized revision of the radiological response. Secondary objectives included assessment of the toxicity profile and clinical benefit of the combination. A two-step study design by Simon was planned. Results: Thirty-four patients were enrolled. Median age at diagnosis was 19 years (range 3-55); males/females ratio was 2.4. Primary axial tumour was present in 24 (70%). After TEMIRI, RECIST response was as follows: partial response -20 (59%), stable disease -11 (32%), progression disease -3 (9%). After TEMIRI, amelioration in ECOG/Lansky score was achieved in 25/34 (73,5%), and reduction or disappearance of pain was observed in 31/34 patients (91%). TEMIRI toxicity was manageable: incidence of grade 3-4 nonhaematological and haematological toxicity was 3% and 3%, respectively (67/68 evaluable courses). At the time of the present analysis, 11 patients are alive; 7 of them are in complete remission and completed their treatment program (5-drug standard chemotherapy). With a median follow-up of 31 months (range 23-75), the 3-year survival estimate is 36,5%±0.09. Conclusions: Upfront TEMIRI x 2 courses showed an encouraging activity, with response rate 59% and deserves further evaluation combined with conventional treatments also in non-metastatic patients. In PMES new treatment strategies are urgently needed. Clinical trial information: NCT02727387.

Efficacy of add-on treosulfan and melphalan high-dose therapy in patients with high-risk metastatic Ewing sarcoma: Report from the International Ewing 2008R3 trial.

11501 Background: Ewing 2008R3 (EudraCT2008-003658-13) was conducted in 12 countries. It evaluated the effect of treosulfan and melphalan high dose chemotherapy followed by re-infusion of autologous hematopoietic stem cells (HDTreoMel) on event-free (EFS, primary endpoint) and overall survival (OS) in high-risk Ewing Sarcoma (EwS). Methods: Phase III, open label, prospective, multi-center, randomized controlled clinical trial. Eligible patients (pts) had disseminated EwS with metastases to bone and/or other sites, excluding pts with only pleuropulmonary metastases. Pts received 6 cycles of VIDE induction and 8 cycles of VAC consolidation therapy. Patients were randomized to receive additional HDTreoMel chemotherapy or no further treatment (control), They were further stratified by number of bone metastases (1, 2-5, &gt; 5). One-sided adaptive inverse-normal 4-stage design, changed after the 1st interim analysis via Müller-Schäfer method. Initial sample size 185 pts, type I error rate 2.5%, power 80%. Results: 109 pts were randomized between 2009 and 2018: 55 were randomized to HDTreoMel. With a median follow-up of 3.3 years, the primary endpoint EFS was not significantly different between HDTreoMel and control in the adaptive design (HR 0.85, 95% CI 0.55-1.32, intention-to-treat). 3-year (3y) EFS was 20.9 % (95% CI 11.5-37.9%) in HDTreoMel and 19.2 % (95% CI 10.8-34.4%) in control pts. Results were similar in the per protocol collective. Subgroup analyses showed that independent of treatment, male patients had a worse outcome than female patients: 3y EFS 13.3% (95% CI 5.7-31.1%) vs 25.2% (95% CI 15.5-40.8%); p = 0.07. Patients aged &lt; 14 had a better outcome when treated in the HDTreoMel group: 3y EFS 39.3% (95% CI 20.4-75.8%) vs 9% (95% CI 2.4-34%); p = 0.016; HR 0.40 (0.19-0.87). These effects were similar in the per protocol collective. Severe toxicities of hematology, gut, general condition and infection were more pronounced in the HDTreoMel group (p &lt; 0.05). Conclusions: In patients with very high risk EwS, additional HDTreoMel was of no benefit for the entire cohort of patients. HDTreoMel may be of benefit for children age &lt; 14. This observation is supported by comparable results from a non-randomized trial EE99 R3 (Ladenstein et al. JCO, 2010). Clinical trial information: NCT00987636 .

Comparison of two chemotherapy regimens in Ewing sarcoma (ES): Overall and subgroup results of the Euro Ewing 2012 randomized trial (EE2012).

11500 Background: In 2010, different chemotherapy regimens were standard in Europe and the USA for newly diagnosed ES. In the absence of novel agents to investigate, comparison of these two strategies was considered worthwhile. Methods: Newly diagnosed localised or metastatic ES patients aged 5-50 were eligible. Patients were randomized to receive either the European regimen (Arm A) of VIDE (vincristine [V], ifosfamide [I], doxorubicin [D] and etoposide [E]) induction and VAI or VAC (V, actinomycin D and I or cyclophosphamide [C]) consolidation or the USA regimen (Arm B) of compressed VDC/IE induction and IE/VC consolidation. The primary outcome measure was event-free survival (EFS); secondary outcomes included overall survival (OS) and toxicity. The design was Bayesian with interpretation based on posterior probabilities (with non-informative priors) – i.e. probability that true hazard ratio (HR) &lt; 1.0 given the data [Pr(HR&lt;1.0|data)], with 95% credible intervals (CrI) reported. HRs were obtained from Cox models adjusted for baseline stratification parameters. Heterogeneity tests (HT) were used to investigate whether the treatment effect differed according to baseline parameters. Analysis was intention-to-treat. Results: Between December 2013 and May 2019, 640 patients were randomised (320 to each arm) from 10 European countries. Baseline stratification factors were: sex (58% male; 42% female); age (41% &lt;14 years; 59% 14+ years); disease type (74% localised, 17% lung/pleural metastasis, 9% other metastasis); tumour volume (56% &lt;200 ml, 44% &gt;200 ml); country (37% UK, 31% France, 32% other). Median follow-up was 1.7 years. The HRs (95% CrI) were 0.70 (0.51, 0.95) for EFS and 0.64 (0.42, 0.96) for OS in favour of Arm B, with posterior probabilities of 98% for both that Arm B was better. Subgroup analyses showed no evidence that this benefit differed depending the baseline features, with no HT being close to significance (table). There were no major differences in acute toxicity: 68% of patients in Arm A experienced serious adverse events and 67% in Arm B. Conclusions: VDC/IE chemotherapy is superior to VIDE for both EFS and OS, with no excess toxicity. This benefit is consistent across all baseline stratification parameters. Clinical trial information: ISRCTN92192408 . [Table: see text]

Protein phosphatase 1 regulatory subunit 1A regulates cell cycle progression in Ewing sarcoma.

Ewing sarcoma (ES) is a malignant pediatric bone and soft tissue tumor. Patients with metastatic ES have a dismal outcome which has not been improved in decades. The major challenge in the treatment of metastatic ES is the lack of specific targets and rational combinatorial therapy. We recently found that protein phosphatase 1 regulatory subunit 1A (PPP1R1A) is specifically highly expressed in ES and promotes tumor growth and metastasis in ES. In the current investigation, we show that PPP1R1A regulates ES cell cycle progression in G1/S phase by down-regulating cell cycle inhibitors p21Cip1 and p27Kip1, which leads to retinoblastoma (Rb) protein hyperphosphorylation. In addition, we show that PPP1R1A promotes normal transcription of histone genes during cell cycle progression. Importantly, we demonstrate a synergistic/additive effect of the combinatorial therapy of PPP1R1A and insulin-like growth factor 1 receptor (IGF-1R) inhibition on decreasing ES cell proliferation and migration in vitro and limiting xenograft tumor growth and metastasis in vivo. Taken together, our findings suggest a role of PPP1R1A as an ES specific cell cycle modulator and that simultaneous targeting of PPP1R1A and IGF-1R pathways is a promising specific and effective strategy to treat both primary and metastatic ES.

Ectopic ACTH Secretion in a Child With Metastatic Ewing's Sarcoma: A Case Report

Ectopic ACTH syndrome is rare in pediatric age. Sarcomas that cause Ectopic ACTH Syndrome (EAS) are even more uncommon. Currently, only three cases of EAS caused by Ewing' sarcoma have been reported. We detail a 10-year-old boy with Cushing's syndrome symptoms caused by ectopic ACTH production by a metastatic Ewing's sarcoma of the right ischio-pubic and ileo-pubic branches. The rapid appearance of cushingoid symptoms, with significant weight gain, acne, hirsutism, and hypercortisolism were implications of ectopic ACTH production as paraneoplastic Cushing's Syndrome. The very high levels of ACTH and non-suppression at the high dose dexamethasone test confirmed the clinical suspicion. We underline the possibility EAS was caused by an ACTH-secreting tumor, including soft tissue sarcomas.

Whole Lung Irradiation for Completely Responding Pulmonary Metastases in Pediatric Ewing Sarcoma

Aim: Scarce data assessing the real value of whole lung irradiation (WLI) in Ewing's sarcoma (ES) with lung-only metastasis, with published conflicting results. We studied the impact of WLI in a homogenous pediatric population. Materials & methods: Retrospective study evaluating the survival outcomes of WLI in these patients. Results: Out of 163 metastatic ES; 41 patients were eligible for WLI. 30 patients (73.1%) received WLI (+ve) while 11 patients (26.8%) did not receive WLI (-ve). Five-year event-free survival was statistically significant in WLI (+ve). Five-year pulmonary relapse-free survival showed trend for improvement with WLI (+ve), while 5-year overall survival was not statistically significant between the two arms. Conclusion: WLI added significantly to the long term clinical outcome of metastatic ES patients, with no irreversible toxicity.

SHH Signaling Pathway Drives Pediatric Bone Sarcoma Progression.

Primary bone tumors can be divided into two classes, benign and malignant. Among the latter group, osteosarcoma and Ewing sarcoma are the most prevalent malignant primary bone tumors in children and adolescents. Despite intensive efforts to improve treatments, almost 40% of patients succumb to the disease. Specifically, the clinical outcome for metastatic osteosarcoma or Ewing sarcoma remains poor; less than 30% of patients who present metastases will survive 5 years after initial diagnosis. One common and specific point of these bone tumors is their ability to deregulate bone homeostasis and remodeling and divert them to their benefit. Over the past years, considerable interest in the Sonic Hedgehog (SHH) pathway has taken place within the cancer research community. The activation of this SHH cascade can be done through different ways and, schematically, two pathways can be described, the canonical and the non-canonical. This review discusses the current knowledge about the involvement of the SHH signaling pathway in skeletal development, pediatric bone sarcoma progression and the related therapeutic options that may be possible for these tumors.

Role of Metastatic Site Irradiation in Pediatric Patients With Metastatic Ewing Sarcoma.

The use of radiation therapy to treat metastases in patients with metastatic Ewing sarcoma (MES) has been controversial and variable. The authors report outcomes and patterns of failure after metastatic site irradiation (MSI). A total of 27 pediatric patients with MES were treated with chemotherapy and received radiation therapy to their primary site. Ten patients additionally received MSI, which consisted of whole-lung irradiation (WLI) in patients with lung metastases. Metastatic sites were followed from diagnosis to the first relapse. Median follow-up was 29 months. Seventy-eight percent of patients relapsed. Two-year progression-free survival (PFS) and overall survival with and without MSI were 30 versus 29% (log rank P=0.38) and 60 versus 70% (log rank P=0.11), respectively. The median time to relapse among patients who relapsed was 19.5 versus 12.3 months for those receiving MSI versus those who did not (P=0.04).Seven of 20 (35%) patients with lung metastases received WLI±other MSI. Two-year PFS with and without MSI was 43% versus 23% (log rank P=0.02). Among patients with a complete response to computed tomography, 5 of 14 (36%) patients received MSI. Two-year PFS with and without MSI was 60% versus 33% (log rank P=0.04).In the cohort of patients who relapsed, among all metastatic sites at diagnosis, the disease recurred at 15% of irradiated sites and 31% of unirradiated sites. On logistic regression, no factors were statistically associated with increased risk of recurrence at initial sites of metastases. Relapses frequently occur at sites of prior unirradiated disease in patients with MES. WLI may improve 2-year PFS, regardless of chemotherapy response. Further investigation of the role of MSI is warranted.

Research on Analysis of Final Diagnosis and Prognostic Factors, and Development of New Therapeutic Drugs for Malignant Tumors (Especially Malignant Pediatric Tumors)

Outcomes of treatment for malignant pediatric tumors including leukemia are improving by conventional multimodal treatment with strong chemotherapy, surgical resection, radiotherapy, and bone marrow transplantation. However, patients with advanced neuroblastoma, metastatic Ewing's sarcoma family of tumor (ESFT), and metastatic osteosarcoma continue to have an extremely poor prognosis. Therefore novel therapeutic strategies are urgently needed to improve their survival. Apoptotic cell death is a key mechanism for normal cellular homeostasis. Intact apoptotic mechanisms are pivotal for embryonic development, tissue remodeling, immune regulation, and tumor regression. Genetic aberrations disrupting programmed cell death often underpin tumorigenesis and drug resistance. Moreover, it has been suggested that apoptosis or cell differentiation proceeds to spontaneous regression in early stage neuroblastoma. Therefore apoptosis or cell differentiation is a critical event in this cancer. We extracted many compounds from natural plants (Angelica keiskei, Alpinia officiarum, Lycaria puchury-major, Brassica rapa) or synthesized cyclophane pyridine, indirubin derivatives, vitamin K3 derivatives, burchellin derivatives, and GANT61, and examined their effects on apoptosis, cell differentiation, and cell cycle in neuroblastoma and ESFT cell lines compared with normal cells. Some compounds were very effective against these tumor cells. These results suggest that they may be applicable as an efficacious and safe drug for the treatment of malignant pediatric tumors.