Relapsed Ewing Sarcoma Research Articles

Consensus recommendations for systemic therapies in the management of relapsed Ewing sarcoma: A report from the National Ewing Sarcoma Tumor Board.

Ewing sarcoma (ES) is a malignant tumor of bone and soft tissue that most often occurs in children, adolescents, and young adults. Debate and controversy remain in the management of relapsed/refractory ES (RR-ES). The authors leveraged the expertise assembled by the National Ewing Sarcoma Tumor Board, a multidisciplinary virtual tumor board that meets monthly to discuss challenging cases of ES. In this review, they focus on select topics that apply to the management of patients with RR-ES. The specific topics covered include the initial approach of such patients and discussion of the goals of care, the role of molecular testing, chemotherapy regimens and novel agents to consider, the role of maintenance therapy, and the use of high-dose chemotherapy with autologous stem cell rescue. The data referenced are often limited to subgroup analyses and/or compiled from multiple sources. Although not intended to replace the clinical judgement of treating physicians, these guidelines are intended to support clinicians and provide some clarity and recommendations for the management of patients with RR-ES. PLAIN LANGUAGE SUMMARY: Ewing sarcoma (ES) is a bone and soft tissue cancer that most often occurs in teenagers and young adults. This article uses the experience of the National Ewing Sarcoma Tumor Board, a multi-institution, multidisciplinary virtual tumor board that meets monthly to discuss challenging cases of ES and to address questions related to the treatment of patients with relapsed ES. Although not intended to replace the clinical judgement of treating physicians and limited by available data, these consensus recommendations will support clinicians who treat patients with this challenging malignancy, made even more difficult when it recurs.

MERTK Is a Potential Therapeutic Target in Ewing Sarcoma.

Outcomes are poor in patients with advanced or relapsed Ewing sarcoma (EWS) and current treatments have significant short- and long-term side effects. New, less toxic and more effective treatments are urgently needed. MER proto-oncogene tyrosine kinase (MERTK) promotes tumor cell survival, metastasis, and resistance to cytotoxic and targeted therapies in a variety of cancers. MERTK was ubiquitously expressed in five EWS cell lines and five patient samples. Moreover, data from CRISPR-based library screens indicated that EWS cell lines are particularly dependent on MERTK. Treatment with MRX-2843, a first-in-class, MERTK-selective tyrosine kinase inhibitor currently in clinical trials, decreased the phosphorylation of MERTK and downstream signaling in a dose-dependent manner in A673 and TC106 cells and provided potent anti-tumor activity against all five EWS cell lines, with IC50 values ranging from 178 to 297 nM. Inhibition of MERTK correlated with anti-tumor activity, suggesting MERTK inhibition as a therapeutic mechanism of MRX-2843. Combined treatment with MRX-2843 and BCL-2 inhibitors venetoclax or navitoclax provided enhanced therapeutic activity compared to single agents. These data highlight MERTK as a promising therapeutic target in EWS and provide rationale for the development of MRX-2843 for the treatment of EWS, especially in combination with BCL-2 inhibitors.

Treatment with novel topoisomerase inhibitors in Ewing sarcoma models reveals heterogeneity of tumor response.

The topoisomerase 1 (TOP1) inhibitor irinotecan is a standard-of-care agent for relapsed Ewing sarcoma (EWS), but its efficacy is limited by chemical instability, rapid clearance and reversibility, and dose-limiting toxicities, such as diarrhea. Indenoisoquinolines (IIQs) represent a new class of clinical TOP1 inhibitors designed to address these limitations. In this study, we evaluated the preclinical efficacy of three IIQs (LMP400, LMP744, and LMP776) in relevant models of EWS. We characterized the pharmacokinetics of IIQs in orthotopic xenograft models of EWS, optimized the dosing regimen through tolerability studies, and tested the efficacy of IIQs in a panel of six molecularly heterogeneous EWS patient-derived xenograft (PDX) models. For each PDX, we conducted whole genome and RNA sequencing, and methylation analysis. We show that IIQs potently inhibit the proliferation of EWS cells in vitro, inducing complete cell growth inhibition at nanomolar concentrations via induction of DNA damage and apoptotic cell death. LMP400 treatment induced ≥30% tumor regression in two of six PDX models, with more durable regression compared to irinotecan treatment in one of these models. RNA sequencing of PDX models identified a candidate predictive biomarker gene signature for LMP400 response. These data, along with pharmacogenomic data on IIQs in sarcoma cell lines, are available at a new interactive public website: https://discover.nci.nih.gov/rsconnect/EwingSarcomaMinerCDB/. Our findings suggest that IIQs may be promising new agents for a subset of EWS patients.

Outcomes of Patients Treated at First Transplant Center in Dubai, UAE

Introduction: Autologous stem cell transplant (ASCT) is a well-known treatment modality for decades with curative potential in the field of hematological malignancies such as lymphoma (Relapsed Hodgkin's Lymphoma) and multiple myeloma (MM), but access in the Middle East, specifically in the metropolitan city of Dubai, United Arab Emirates (UAE) has been limited. This is despite the fact that UAE is a high-income country according to the Gross national income (GNI) and has a population of approximately 9.5 million. In 2021, we developed the first and only ASCT unit in Dubai, United Arab Emirates (UAE). This was a milestone achievement to offer ASCT to residents. This service was available since approval by Dubai Health Authority (DHA) in September 2021 and capacity for 2 ASCT at a time. The unit comprises 2 positive pressure isolation rooms, with in-house apheresis, stem cell storage facilities. This is the first report of the safety and efficacy outcomes of patients (pts) who were transplanted to date at our center. Despite the modest volume, this data will serve as a resource for policymakers, healthcare professionals and is important for the ASCT community as we advance care of ASCT recipients in regions with previously limited access. Methods: This is a single-center retrospective study performed at American Hospital Dubai, UAE. We identified and included pts who received ASCT for lymphoma and MM between September 2021-June 2023. Per institutional protocols, mobilization was done with granulocyte colony-stimulating factor (GCSF). The stem cells were collected via Optia®spectra apharesis system. MM pts collected for 2 ASCT while those with relapsed HL collected for 1 ASCT. Conditioning chemotherapy administration was outpatient for melphalan (for MM) and inpatient for rHL with BEAM. Monitoring was per institutional standards. Descriptive analysis is reported. Results: There have been 17 ASCTs performed with at least one month follow-up. The most common indication was MM (9) followed by Relapsed lymphoma (8) and 1 case of multiply relapsed Ewing's Sarcoma. (Table 1) Mobilization:100% pts received granulocyte colony-stimulating factor (GCSF) mobilization with one patient required plerixafor in addition. The median days of collection was 1 (Range 1-2 days). All pts collected successfully. Neutropenic Events: Almost all patients (95%) had expected neutropenic fever by day +8.Median time to onset of Grade 4 neutropenia (absolute neutrophil count <500) was 6 days (3-8); Median time to onset of neutropenic fever was 7 days (3-10). Causative organisms were identified in 2 (11%) (Both bacterial infections). Antibiotic of choice was meropenem. ICU Admissions: Only one (6%) patient was admitted in ICU for neutropenic typhilitis on Day +3. That pt spent 20 days in ICU and total of 45 days in hospital with recovery to baseline. Resource Utilization: ASCT for MM was 100% outpatient chemotherapy and cell infusions. The patients were admitted on Day 6 post-transplant for neutropenic care, except 1 patient who presented at the emergency department with febrile neutropenia and diarrhea on day +3. The patients planned for ASCT for rHL were admitted for conditioning chemotherapy, stem cell infusion and neutropenic care until count recovery. Median duration of hospital stay was 17 days (8-45 days). The median days of GCSF post-cell infusion per patient was 8 days (6-15). Efficacy Outcomes: Of the 9 MM pts, all achieved complete response (CR). With a median follow up of 21 months, 100% continued to be on maintenance. For the rHL cohort, response rate was 100%, all CR, and continue to have ongoing responses. The patient with relapsed refractory Ewing's sarcoma has progression-free survival (PFS) of 5 months after ASCT. Overall survival (OS) was 100% for all diseases post-transplant. Conclusion: Our study describes promising outcomes of pts treated at our center, which is the first and only center offering ASCT in Dubai, UAE. This demonstrates that with available infrastructure and expertise, ASCT programs in countries similar to UAE are feasible and increase access to this therapy. Continued efforts at monitoring outcomes, education of the local population, and outreach partnerships with colleagues are essential to increase access to patients in the region.

Phase 1 study of cabozantinib in combination with topotecan-cyclophosphamide for patients with relapsed Ewing sarcoma or osteosarcoma.

Phase 1 study assessing the safety and toxicity of cabozantinib in combination with topotecan and cyclophosphamide for relapsed osteosarcoma and Ewing sarcoma. Oral cabozantinib (25mg/m2 ) was administered daily for 21 (dose level 1) or 14 (dose level -1B) days. Topotecan (0.75mg/m2 ) and cyclophosphamide (250mg/m2 ) were administered intravenously (IV) on days 1-5. A modified 3+3 design based upon first cycle dose-limiting toxicities (DLT) was used for dose escalation. Twelve patients with a median age of 15 (12.9-33.2) years were enrolled (seven with Ewing sarcoma; five with osteosarcoma); all were evaluable for toxicity. At dose level 1, three of six patients developed first cycle DLT: grade 3 epistaxis, grade 3 transaminitis, and prolonged grade 2 thrombocytopenia. Six patients were enrolled on dose level -1B (interrupted cabozantinib, given days 8-21), with one first cycle DLT (grade 3 pneumothorax) observed. Of the 10 response evaluable patients, one had partial response (Ewing sarcoma), seven had stable disease, and two had progressive disease. The recommended phase 2 doses and schedules for this combination are topotecan 0.75mg/m2 IV days 1-5, cyclophosphamide 250mg/m2 IV days 1-5, and cabozantinib 25mg/m2 days 8-21. Non-concomitant administration of cabozantinib with cytotoxic therapy in this population has acceptable toxicity, while allowing for potential disease control.

Harnessing immunomodulation during DNA damage in Ewing sarcoma.

Ewing sarcoma is a fusion-oncoprotein-driven primary bone tumor most commonly diagnosed in adolescents. Given the continued poor outcomes for patients with metastatic and relapsed Ewing sarcoma, testing innovative therapeutic approaches is essential. Ewing sarcoma has been categorized as a 'BRCAness' tumor with emerging data characterizing a spectrum of DNA damage repair defects within individual Ewing tumors, including the presence of EWSR1::FLI1 itself, recurrent somatic mutations, and rare germline-based defects. It is critical to understand the cumulative impact of various DNA damage repair defects on an individual Ewing tumor's response to therapy. Further, in addition to DNA-damage-directed therapies, subsets of Ewing tumors may be more susceptible to DNA-damage/immunotherapy combinations given the significant cross-talk between DNA damage and inflammatory pathways in the tumor microenvironment. Here we review potential approaches utilizing DNA-damaging agents as modulators of the Ewing tumor immune microenvironment, with a focus on radiation and opportunities during disease metastasis and relapse.

Abstract 1086: PEGylated talazoparib enhances therapeutic window of its combination with temozolomide in Ewing sarcoma

Abstract Introduction: The Ewing family of sarcomas is the fourth most common highly malignant childhood cancer. Current standard therapy is relatively ineffective for relapsed and metastatic Ewing sarcoma predominantly due to development of resistance to chemotherapeutics. Nanoparticle-formulated drugs are reported to increase uptake into tumor tissue, while reducing drug access to normal tissues, because of reduced permeability of normal vasculature. This potentially leads to lower dose exposure in normal tissues and reduced toxicity. The purpose of this study was to evaluate the antitumor activity of the PEGylated poly(ADP) ribose polymerase 1/2 (PARP1/2) inhibitor talazoparib combined with the DNA alkylating agent temozolomide (TMZ) in Ewing sarcoma xenograft models and other pediatric cancers. Method:. We evaluated the preclinical efficacy of PEGylated talazoparib (PEG~TLZ), alone or in combination with TMZ, in Ewing sarcoma and glioblastoma patient derived xenograft (PDX) models. Additional solid tumor models, such as rhabdomyosarcoma, Wilms tumor, malignant rhabdoid tumor, osteosarcoma, and synovial sarcoma were evaluated using a novel single mouse testing approach. PEG~TLZ was administered as a single administration intraperitoneally to mice bearing subcutaneous tumors at a single dose of 5 - 20 µmol/kg alone, or at 10 µmol/kg on day 1 combined with TMZ administered orally at 40 mg/kg starting on day 3 or 4 for 5 consecutive days. Results: Mice bearing Ewing sarcoma and MGMT-deficient glioblastoma xenografts tolerated PEG~TLZ+TMZ with minimal toxicity and achieved maintained complete response (MCR). The time-separated drug administration schedule of the single dose of PEG~TLZ followed by the 5-day TMZ treatment demonstrated objective responses in Ewing sarcoma, rhabdoid tumor, rhabdomyosarcoma, and glioblastoma PDX models. Conclusion: We evaluated the range of PEG~TLZ+TMZ activity among pediatric solid tumor panels using conventional and single-mouse testing approaches, and demonstrated that PEG~TLZ combined with delayed TMZ administration enhances the therapeutic window of the treatment compared to free TLZ. From the clinical standpoint, the single intravenous administration of PEG~TLZ could be advantageous for treating infants and young children. Citation Format: Vanessa Del Pozo, Andrew J. Robles, Shaun Fontaine, Qianqian Liu, Joel E. Michalek, Peter J. Houghton, Raushan Kurmasheva. PEGylated talazoparib enhances therapeutic window of its combination with temozolomide in Ewing sarcoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1086.

Phase 2 trial of palbociclib and ganitumab in patients with relapsed Ewing sarcoma.

e23507 Background: Ewing sarcoma (EwS) is an aggressive sarcoma with few treatment options for patients with relapsed disease. CDK4 is a known genomic vulnerability in EwS, but an unappealing monotherapy target given propensity for innate resistance. In laboratory studies to identify targets synergistic with CDK4 inhibition, IGF-1R scored highly. We present the results of a phase 2 study combining palbociclib (CDK4/6 inhibitor) with ganitumab (IGF-1R monoclonal antibody) for patients with relapsed or refractory EwS. Methods: This was a prospective open-label, non-randomized, single-center, phase 2 study (NCT04129151) for patients with relapsed EwS and RECIST measurable disease. Patients with relapsed EwS ≥12 years and a documented fusion consistent with EwS were eligible. Patients initially received Palbociclib 125 mg on days 1-21 and ganitumab 18 mg/kg on days 1 and 15 of a 28-day cycle. The primary endpoints were objective response (complete or partial) per RECIST and toxicity by CTCAE. Secondary endpoints included progression-free survival (PFS) and overall survival (OS). A one-stage design required ≥4 responders out of 15 to evaluate an alternative hypothesis of 40% response rate against a null of 10%. Results: Ten evaluable patients enrolled between 5/2019-8/2021. The study closed 12/2021 due to lack of ganitumab drug supply. The median age at enrollment was 25.7 years (range 12.3-40.1; Table). The median duration of therapy was 2.5 months (range = 0.9-10.8). There were no complete or partial responders. Three of 10 patients had stable disease for > 4 cycles and 2 had stable disease at completion of planned therapy or study closure. 6-month PFS was 30±14.5%. Two patients had cycle 1 hematologic DLTs triggering the pre-defined toxicity rule; the palbociclib dose was subsequently reduced to 100 mg daily for 21 days. Among the remaining 8 patients, two had cycle 1 hematologic DLTs at the 100 mg dose. 80% of patients had grade 3/4 AEs; the most commons AEs were neutropenia (n = 8), white blood cell decreased (n = 7), and thrombocytopenia (n = 5). Total IGF-1 testing demonstrated intrapatient elevations in serum levels after a single cycle of therapy. Analysis of serial ctDNA samples is ongoing. Conclusions: Four responders out of 15 were required to reject the null hypothesis of a 10% response rate. With 0/10 responders, we concluded that this combination lacked adequate therapeutic activity for further study. The combination was tolerable at a palbociclib dose of 100 mg, with primarily hematologic toxicity in patients with relapsed EwS. Clinical trial information: NCT04129151. [Table: see text]

Irinotecan plus temozolomide in relapsed Ewing sarcoma: an integrated analysis of retrospective studies

BackgroundThe prognosis of patients with relapsed Ewing sarcoma is poor. In this study, we aimed to pooled-analyze the efficacy and safety of the combination of irinotecan and temozolomide in treating patients with relapsed Ewing sarcoma.MethodsPubMed, Cochrane CENTRAL, Web of Science, and EMBASE were systematically searched on September 27, 2021. The primary outcomes were rates of objective response and disease control, and the secondary outcomes were toxicities.ResultsSix retrospective studies with 184 patients were enrolled in the analysis. The median age ranged from 14 to 21. The integrated rates were 44% (95% confidence interval [CI] 31–58) for objective response and 66% (55–77) for disease control. Grade 3–4 neutropenia, thrombocytopenia, and diarrhea occurred in 8% (3–16), 7% (3–11), and 8% (5–10) of chemotherapeutic cycles, respectively. 18% (7–32) and 6% (2–11) of patients suffered grade 3–4 neutropenia and thrombocytopenia after irinotecan plus temozolomide treatment.ConclusionIrinotecan plus temozolomide combination chemotherapy showed antitumor activity and an acceptable safety profile in patients with relapsed Ewing sarcoma. More future prospective studies are needed to confirm the retrospective results.

Whole-genome sequencing facilitates patient-specific quantitative PCR-based minimal residual disease monitoring in acute lymphoblastic leukaemia, neuroblastoma and Ewing sarcoma

BackgroundMinimal residual disease (MRD) measurement is a cornerstone of contemporary acute lymphoblastic leukaemia (ALL) treatment. The presence of immunoglobulin (Ig) and T cell receptor (TCR) gene recombinations in leukaemic clones allows widespread use of patient-specific, DNA-based MRD assays. In contrast, paediatric solid tumour MRD remains experimental and has focussed on generic assays targeting tumour-specific messenger RNA, methylated DNA or microRNA.MethodsWe examined the feasibility of using whole-genome sequencing (WGS) data to design tumour-specific polymerase chain reaction (PCR)-based MRD tests (WGS-MRD) in 18 children with high-risk relapsed cancer, including ALL, high-risk neuroblastoma (HR-NB) and Ewing sarcoma (EWS) (n = 6 each).ResultsSensitive WGS-MRD assays were generated for each patient and allowed quantitation of 1 tumour cell per 10−4 (0.01%)–10–5 (0.001%) mononuclear cells. In ALL, WGS-MRD and Ig/TCR-MRD were highly concordant. WGS-MRD assays also showed good concordance between quantitative PCR and droplet digital PCR formats. In serial clinical samples, WGS-MRD correlated with disease course. In solid tumours, WGS-MRD assays were more sensitive than RNA-MRD assays.ConclusionsWGS facilitated the development of patient-specific MRD tests in ALL, HR-NB and EWS with potential clinical utility in monitoring treatment response. WGS data could be used to design patient-specific MRD assays in a broad range of tumours.

Haploidentical Hematopoietic Cell Transplantation Followed By Donor Natural Killer Cell Infusion and Sirolimus Maintenance Can Successfully Treat Patients with High-Risk Ewing Sarcoma

Background: Approximately 30-40% of patients with newly diagnosed localized Ewing Sarcoma (EWS), and 60-80% with disseminated EWS, will eventually relapse. Patients with late relapse have a 5 year overall survival (OS) of 30%, while patients with early relapse (<2 years from initial diagnosis) have a more dismal 5 year OS of <10% (Stahl Ped Blood Cancer 2011). Unfortunately, conventional treatments including chemotherapy, autologous transplantation, surgery, and radiation therapy often lead to additional morbidities without substantial gains in disease remissions or survival. Allogeneic hematopoietic cell transplantation (HCT) using human leukocyte antigen (HLA)-matched related donors has shown some success for patients with high-risk EWS (Baird BBMT 2012). Additionally, EWS cells are exquisitely sensitive to natural killer (NK) cell cytotoxic activity (Cho CCR 2010), and many EWS tumors have upregulated the mammalian target of rapamycin (mTOR) pathway making them susceptible to anti-mTOR agents (Wan et al The Oncologist 2007). With growing literature supporting the safety and efficacy of HLA-haploidentical HCT with post-transplant cyclophosphamide (CY) for hematological malignancies, we developed a novel treatment regimen adding a single infusion of donor NK cells on day +7 after HLA-haploidentical HCT, followed by sirolimus as part of a GVHD prophylaxis strategy. We hypothesized that this multi-faceted immunotherapy approach, in combination with an mTOR-inhibitor post-transplant, would be well-tolerated and efficacious in patients with relapsed EWS.Methods: Patients received reduced-intensity conditioning using fludarabine (150 mg/m2), CY (29 mg/kg) and 3 Gy total body irradiation, followed by infusion of HLA-haploidentical related marrow (n=4) or peripheral blood stem cells (n=2). Post-transplant immune suppression consisted of CY (50 mg/kg, days +3 and +4), tacrolimus (beginning day +5, and transitioned to sirolimus from day +28 to 6 months), and mycophenolate mofetil (MMF) (day +9 to +84). MMF initiation was delayed until day +9 to decrease concerns of inhibiting NK activity. For NK cell therapy, non-mobilized peripheral blood mononuclear cells were collected from the same haplo donors on day +6 using apheresis. NK cells were selected on day +7 using the Miltenyi CliniMACS system (CD3 depletion followed by CD56 selection) and were administered as a single, fresh infusion that same day without prior culturing, expansion, or exogenous cytokine exposure.Results: Four pediatric and 2 adult patients with median age of 18.5 (10.5-34.0) years having relapsed EWS were enrolled on this Phase II study. All patients had stable or undetectable gross disease at HCT. Patients had 1 (n=3), 2 (n=2), or 5 (n=1) relapses prior to HCT. Median time from EWS diagnosis to first relapse was 22.5 (15-56) months. Two patients had prior autologous HCTs. Median HCT-CI score at time of transplant was 4 (2-5), demonstrating increased risk for transplant-related mortality (TRM). Patients received a median CD34 dose of 5.24 (1.3-8.3) x 106/kg. NK cell products had a median log T cell depletion of 6.25 (5.18-6.42), median NK recovery of 54% (47-71%), and median NK purity of 91% (78-96%). Excellent viability (>99%) was seen in all NK products. Donor NK infusions were well-tolerated without fevers or other adverse reactions. Sustained full donor chimerism (>95% CD3) was achieved in all 6 patients. One patient developed stage 1 (grade II) gut graft-versus-host disease (GVHD) at day +52 which was treated with budesonide. To date, no patients have developed chronic GVHD nor have died from TRM. With a median follow-up of 1.3 years (range, 180 days - 2.7 years), 1 year overall survival is estimated at 75% (see figure). Three patients have relapsed or progressed at 187, 284, and 455 days after HCT, of which 1 patient (who had 5 prior relapses), died of progressive disease at day +350. The other 2 patients have received salvage treatments. They remain alive with full-donor chimerism and low-level/absent disease at 1.7 and 2.7 years post-HCT.Summary: This dual immunotherapy approach, using HLA-haploidentical HCT with donor NK cell infusion, followed by sirolimus maintenance, was well-tolerated by all patients. While longer follow-up is needed, preliminary results of this ongoing novel approach demonstrate feasibility in EWS patients, with better than expected 1 year OS and promising disease control. [Display omitted] DisclosuresThakar:Miltenyi Biotec: Other: funding of partial costs - disposable agents. Hari:Janssen: Honoraria; Sanofi: Honoraria, Research Funding; Millennium Pharmaceuticals, Inc., a wholly owned subsidiary of Takeda Pharmaceutical Company Limited: Honoraria, Research Funding; Celgene: Consultancy, Honoraria, Research Funding; Novartis: Consultancy; Takeda Pharmaceuticals: Consultancy, Honoraria, Research Funding; Bristol Myers Squibb: Consultancy, Research Funding; Spectrum: Consultancy, Research Funding; Amgen: Consultancy, Honoraria, Research Funding.

Long-Term Follow-up of High-Dose Chemotherapy with Autologous Stem Cell Transplantation in Children and Young Adults with Metastatic or Relapsed Ewing Sarcoma: A Single-Institution Experience

Forty-seven patients with metastatic disease at diagnosis or recurrent Ewing sarcoma (EWS) received high-dose chemotherapy (HDC) followed by tandem (n=20, from February 13, 1997, to October 24, 2002) or single (n=27, from October 1, 2004, to September 5, 2018) autologous hematopoietic stem cell transplantation (ASCT). To our knowledge, this is the largest single-institution study with sustained long-term follow-up exceeding 10 years. All patients who underwent single ASCT received a novel conditioning regimen with busulfan, melphalan, and topotecan. The overall survival (OS) and disease-free survival (DFS) were 46% and 37% at 10 years and 42% and 37% at 15 years, respectively. Disease status at transplant and the time to disease relapse prior to ASCT were identified as important prognostic factors in OS, DFS, and risk of relapse. At 10 years, patients who underwent transplantation in first complete response (1CR) had an excellent outcome (OS 78%), patients in 1CR/second complete response (2CR)/first partial response (1PR) had an OS of 66%, and patients at third or more complete response, second or more partial response, or advanced disease had an OS of 26%. Ten-year OS for patients without a history of relapse, with late relapse (≥2 years from diagnosis), or with early relapse (<2 years from diagnosis) was 75%, 50%, and 18%, respectively. Selected patients in 1CR, 2CR, 1PR, and with late relapse had excellent, sustained 10- and 15-year OS and DFS.

Chemoimmunotherapy for the salvage treatment of Ewing sarcoma: A case report

Although, immune check-point inhibitors changed the course of many cancers, the outcomes in sarcomas were rather disappointing with less than 10% response rates. Ewing sarcoma is a poorly differentiated and aggressive tumor mostly seen in the children and adolescents. It's a distinct type of sarcoma with prominent chemosensitivity in the early stages. However, the relapsing disease has a poor prognosis with limited treatment options. Herein, we represent a case of relapsed Ewing sarcoma treated with multiple lines of chemotherapy.Management & outcome: The patient had a very good response to salvage treatment with a combination of paclitaxel and nivolumab which lasted for twelve months after the cessation of treatment. We think that chemotherapy plus immunotherapy can be an option for Ewing sarcoma patients treated with multiple lines of chemotherapy.

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.

Irinotecan and temozolomide chemotherapy in paediatric and adult populations with relapsed Ewing Sarcoma.

Irinotecan and temozolomide (IT) is a widely used regimen for relapsed Ewing sarcoma (ES), although studies are largely limited to paediatric populations. We retrospectively reviewed paediatric (< 18years) and adult patients (≥ 18years) treated with salvage IT at two institutions. Haematologic toxicities were graded according to common terminology criteria of adverse events. Survival was estimated by the Kaplan-Meier method and compared by the Log Rank test. Fifty-three patients were treated with IT from Jan, 2010 to Dec, 2018 (n = 16 paediatric; n = 37 adult). IT was given as second-line (n = 34; 64%) or ≥ third-line (n = 19; 36%). There was no difference in ≥ grade 3/4 haematologic toxicity between paediatrics and adults (31% vs. 35% respectively; p = 0.76). The frequency of diarrhoea of any grade was similar (38% in each group). Of 43 patients assessable for response, 12 (28%) had objective response (1 CR, 11 PR), 12 (28%) stable disease and 19 (44%) disease progression. Objective response rate did not differ between the two groups (36% in paediatrics vs. 25% in adults; p = 0.47). Median PFS was superior in paediatrics vs. adults (7.4 vs. 2.2months, p = 0.039). Irinotecan and temozolomide (IT) chemotherapy has activity for relapsed ES, with favourable toxicity and equally observed objective responses in the paediatric and adult populations. The observed superior PFS for the paediatric cohort requires further confirmation in future studies.

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 A14: Exploiting DNA damage repair defects to enhance PD-L1 expression in Ewing sarcoma

Abstract Objective: The purpose of this work is to determine whether PD-L1 expression can be upregulated by PARP inhibitor treatment in Ewing tumor cells harboring DNA damage repair defects. Further, we aimed to determine whether enhanced PD-L1 expression renders Ewing cells more susceptible to T-cell mediated apoptosis following PD-1 inhibition. Experimental Procedures: Patient-derived tumor organoids and monolayers (named PSaRC-318) were established from a viably cryofrozen metastatic lung lesion from a patient with Ewing sarcoma and a paternally inherited germline BARD1 mutation. A673, CHLA9, and CHLA10 Ewing tumor cell lines were also employed. Sensitivity of PSaRC-318 to the PARP inhibitors olaparib and talazoparib was established by performing IncuCyte apoptosis assays. PD-L1 expression was examined using RT-PCR, Western blot, and flow cytometry. BARD1, BRCA, and RAD51D siRNA were also utilized to test the role of specific DNA damage repair proteins on this effect. T-cell/tumor cell cocultures and tumor-primed T-cells were established. T cell-induced tumor cell apoptosis was monitored in real-time using IncuCyte caspase and annexin assays. Results: PSaRC-318 organoids demonstrate exquisite sensitivity to PARP inhibitors, with an IC50 for talazoparib of 3.54nM. Treatment of PARP inhibitor-resistant Ewing cell lines, such as CHLA10, with BARD1 siRNA rendered cells sensitive to PARP inhibition. Ewing tumor cells were found to upregulate tumor cell surface expression of the checkpoint protein PD-L1 in response to treatment with PARP inhibitors. The impact of BARD1, BRCA1, and RAD51D on PD-L1 expression in Ewing tumor cells was also determined. Next, Ewing tumor cells were pretreated with PARP inhibitors and then cocultured with tumor-primed T-cells. T cell-mediated tumor cell apoptosis was monitored in real time using caspase IncuCyte assays. PD-1 blocking antibody added to T-cell/tumor cell cocultures resulted in an increase in early tumor cell death compared to IgG controls. Conclusion: Single-agent PARP inhibitors have not demonstrated significant clinical benefit in relapsed Ewing sarcoma to date, suggesting that combinatorial therapy with PARP inhibitors may be a more effective approach. Here, we have shown that Ewing tumor cells with germline mutations in DNA-damage repair proteins, such as BARD1, can upregulate PD-L1 expression following treatment with PARP inhibitors. Thus, PARP inhibitor/anti-PD-1 therapy may be a logical combination for the treatment of relapsed Ewing sarcoma. Ongoing preclinical studies are continuing to examine this combination. Citation Format: Lisa M. Maurer, Rose M. Venier, Claire Julian, Kelly M. Bailey. Exploiting DNA damage repair defects to enhance PD-L1 expression in 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 A14.

Phase II trial using haploidentical hematopoietic cell transplantation (HCT) followed by donor natural killer (NK) cell infusion and sirolimus maintenance for patients with high-risk solid tumors.

e23551 Background: Patients with relapsed solid tumors have dismal predicted OS &lt; 10-20% at 5 years with conventional salvage therapies. We hypothesized that a multi-faceted immunotherapy approach to optimize graft-versus-tumor (GVT) effects, in combination with a mammalian target of rapamycin (mTOR) inhibitor maintenance strategy, would enhance early disease-control rates (DCR), leading to improved progression-free survival (PFS) and OS for this high-risk population. Methods: Treatment consisted of HLA-haploidentical marrow (n = 12) or peripheral blood stem cell (n = 3) transplantation to optimize GVT effects, preceded by reduced-intensity conditioning (fludarabine, cyclophosphamide, and 3 Gy total body irradiation) to promote engraftment of these mismatched stem cells. Haplo-NK cells were given to boost this GVT effect. They were purified from non-mobilized donor mononuclear cells by CD3 depletion followed by CD56 selection using the Miltenyi CliniMACS system and were infused fresh on day +7 after HCT. Postgrafting immunosuppression included sirolimus maintenance, which continued until 6 months post-HCT. Results: Fifteen patients with relapsed Ewing sarcoma (EWS) (n = 9), rhabdomyosarcoma (n = 4), osteosarcoma (n = 1), and medulloblastoma (n = 1) having stable or undetectable gross disease were enrolled on this Phase II trial. Median age at HCT was 19 (4.5-37) years old and median performance status was 80%. Four patients underwent prior autologous transplants. Patients received a median NK dose of 6.5 (3.7-11.4) x 106/kg. NK cell products had a median log T cell depletion of 5.94 (5.18-6.75), median NK recovery of 62% (48-71%), and median NK purity of 92% (74%-97%). All donor NK infusions were well-tolerated without cytokine release syndrome. All patients engrafted, and all had sustained full donor chimerism ( &gt; 95% CD3). Two patients developed grade II acute graft-versus-host disease (GVHD), and 2 patients developed chronic GVHD. No patients died from transplant-related causes. With a median follow-up of 1.3 years (range, 70 days – 5 years), 6-month DCR is 72%. 1- and 2-year OS for the entire cohort is estimated at 64% and 40%, respectively, while PFS is 29% and 22%, respectively. For patients with EWS,1- and 2- year -OS is estimated at 75% and 45%, and PFS is 38% and 25%, respectively. Conclusions: This dual immunotherapy approach followed by mTOR inhibition maintenance was well-tolerated, with better than expected OS for this high-risk set of diseases. Clinical trial information: NCT02100891 .

Irinotecan and temozolomide upfront and in relapsed Ewing sarcoma: A translational study on MGMT (O6-methylguanine–DNA methyltransferase) and ABCG2 (MGMTLiberati).

e23564 Background: Activity of temozolomide (TEM) and irinotecan (IRI) in recurrent Ewing sarcoma (EWS) was demonstrated. Few data are available on TEMIRI use upfront. Biological predictive factors are lacking. Methods: This multi-institutional retrospective study (NCT03542097) included 59 patients with EWS. 8 patients with very high risk (HR) EWS (multivisceral ± bone marrow) received TEMIRI (TEM 100 mg/m2/day oral, and IRI 40 mg/m2/day intravenous, days 1-5, every 21 days) upfront, 51 patients after relapse (28% in 1st line, 72% ≥ 2nd line). Overall response rate (ORR: CR+PR), SD, and PD, progression-free (PFS) and overall survival (OS) were assessed. The relationship between pre-treatment expression of MGMT and ABCG2 (when FFPE tissue available) with ORR, PFS and OS was evaluated. Results: Median age was 27 years (range 4-62 years): 47 patients (80%) were adults (≥18 years), 35 (61%) had ECOG 0 and 42 (71%) presented with multivisceral disease (+ bone marrow in 5). MGMT was positive in 16/30 (53%), ABCG2 in 4/33 (12%). ORR for upfront TEMIRI (n = 8) was 50% (CR + PR = 1 + 3), with SD 50%, while in recurrent EWS (n = 49, 2 patients no measurable by RECIST) ORR was 31% (CR + PR = 4 + 11), SD 38%, and PD 31%. A better ORR was observed in adult (p = 0.008) &amp; ECOG 0 (p = 0.001) patients; MGMT &amp; ABCG2 expression did not influence ORR. 6-mos PFS was 87% after TEMIRI upfront, 43% at recurrence (p = 0.06), and 65% vs 28% (p = 0.02) for ECOG 0 vs ECOG 1-2, respectively. 6-mos PFS was 62% in MGMT+ vs 33% in MGMT- (p = 0.4) and 75% in ABCG2+ vs 50% in ABCG2- (p = 0.7). Median time to progression (TTP) with upfront TEMIRI was 9 months (range 5-28 months), with 1 patient with ongoing CR at 56 months; median TTP at relapse was 3 months (1-29 months). MGMT and ABCG2 expression did not influence 1-yr OS, whereas ECOG (0 vs 1-2) did (88% vs 31% p &lt; 0.001). Grade 3-4 diarrhea, neutropenia, and thrombocytopenia incidence was &lt; 5%, 1 patient with recurrent kidney EWS died of acute liver failure. Conclusions: TEMIRI showed promising activity in a very unfavorable cohort of EWS patients, with manageable toxicity. Performance status correlated with 6-month PFS &amp; OS whereas MGMT &amp; ABCG2 did not. Clinical trial information: NCT03542097 .

EWS-FLI1 low Ewing sarcoma cells demonstrate decreased susceptibility to T-cell-mediated tumor cell apoptosis.

Metastatic and relapsed Ewing sarcoma typically afflicts the adolescent population and is largely fatal. These bone tumors are most commonly driven by the fusion oncoprotein EWS-FLI1. Ewing tumors demonstrate significant intra-tumoral heterogeneity, and individual tumor cells can express highly variable and dynamic levels of EWS-FLI1. Recent studies revealed that the EWS-FLI1 oncoprotein level (high versus low expression) greatly influences the behavior of Ewing tumor cells. As compared to cells with high EWS-FLI1, Ewing cells in the EWS-FLI1 low state demonstrate an increased propensity for metastasis. In light of these observations, we sought to determine how tumor cell EWS-FLI1 level influences the anti-tumor cell immune response. Since ICAM-1, which can promote tumor cell/T-cell interaction and T-cell activation, is highly expressed on EWS-FLI1 low cells, we hypothesized that EWS-FLI1 low cells would be more susceptible to T-cell mediated tumor cell apoptosis as compared to cells with high EWS-FLI1. Unexpectedly, we found that EWS-FLI1 low cells are more resistant to T-cell mediated apoptosis than EWS-FLI1 high cells. We investigated the potential mechanisms by which EWS-FLI1 level might influence the T-cell anti-tumor response, and discovered that low EWS-FLI1 expression results in upregulation of PD-L1 and PD-L2, both important ligands for the PD-1 immune checkpoint receptor on T-cells. We demonstrated that blocking PD-1 results in a greater increase of T-cell mediated killing of EWS-FLI1 low tumor cells as compared to cells with higher EWS-FLI1 expression. Our studies suggest that Ewing cells in the EWS-FLI1 low expression state may serve as a niche of tumor immune-evasion.