Ewing Sarcoma Patient-derived Xenografts Research Articles

Abstract 6249: Low Bcl-2 is a robust biomarker of sensitivity to nab-paclitaxel in Ewing sarcoma

Abstract Nanoparticle albumin-bound paclitaxel (nab-paclitaxel) shows potent preclinical anticancer activity in pediatric solid tumors such as Ewing sarcoma, rhabdomyosarcoma and neuroblastoma, but responses in clinical trials have been modest. In this work, we aimed to discover a rational biomarker-based approach to select the right candidate patients for this treatment. We assessed the efficacy of nab-paclitaxel in 27 patient-derived xenografts (PDX), including 14 Ewing sarcomas, five rhabdomyosarcomas and several other pediatric solid tumors. Response rate (partial or complete response) was remarkable in rhabdomyosarcomas (four of five) and Ewing sarcomas (four of 14). We addressed several predictive factors of response to nab-paclitaxel such as the expression of the secreted protein acidic and rich in cysteine (SPARC), chromosomal stability of cancer cells and expression of antiapoptotic members of the B-cell lymphoma-2 (Bcl-2) family of proteins such as Bcl-2, Bcl-xL, Bcl-W and Mcl-1. Protein (immunoblotting) and gene expression of SPARC correlated positively, while immunoblotting and immunohistochemistry expression of Bcl-2 correlated negatively with the efficacy of nab-paclitaxel in Ewing sarcoma PDX. The negative correlation of Bcl-2 immunoblotting signal and activity was especially robust (r = 0.8352; P = 0.0007; Pearson correlation). Consequently, we evaluated pharmacological strategies to inhibit Bcl-2 during nab-paclitaxel treatment. We observed that the Bcl-2 inhibitor venetoclax improved the activity of nab-paclitaxel in highly resistant Bcl-2-expressing Ewing sarcoma PDX. Overall, our results suggest that low Bcl-2 expression could be used to select patients with Ewing sarcoma sensitive to nab-paclitaxel, and Bcl-2 inhibitors could improve the activity of this drug in Bcl-2-expressing Ewing sarcoma. Citation Format: Guillem Pascual-Pasto, Claudia Resa-Pares, Angel M. Carcaboso. Low Bcl-2 is a robust biomarker of sensitivity to nab-paclitaxel in Ewing sarcoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 6249.

Low Bcl-2 is a robust biomarker of sensitivity to nab-paclitaxel in Ewing sarcoma

Nanoparticle albumin-bound paclitaxel (nab-paclitaxel) shows potent preclinical anticancer activity in pediatric solid tumors such as Ewing sarcoma, rhabdomyosarcoma and neuroblastoma, but responses in clinical trials have been modest. In this work, we aimed to discover a rational biomarker-based approach to select the right candidate patients for this treatment. We assessed the efficacy of nab-paclitaxel in 27 patient-derived xenografts (PDX), including 14 Ewing sarcomas, five rhabdomyosarcomas and several other pediatric solid tumors. Response rate (partial or complete response) was remarkable in rhabdomyosarcomas (four of five) and Ewing sarcomas (four of 14). We addressed several predictive factors of response to nab-paclitaxel such as the expression of the secreted protein acidic and rich in cysteine (SPARC), chromosomal stability of cancer cells and expression of antiapoptotic members of the B-cell lymphoma-2 (Bcl-2) family of proteins such as Bcl-2, Bcl-xL, Bcl-W and Mcl-1. Protein (immunoblotting) and gene expression of SPARC correlated positively, while immunoblotting and immunohistochemistry expression of Bcl-2 correlated negatively with the efficacy of nab-paclitaxel in Ewing sarcoma PDX. The negative correlation of Bcl-2 immunoblotting signal and activity was especially robust (r = 0.8352; P = 0.0007; Pearson correlation). Consequently, we evaluated pharmacological strategies to inhibit Bcl-2 during nab-paclitaxel treatment. We observed that the Bcl-2 inhibitor venetoclax improved the activity of nab-paclitaxel in highly resistant Bcl-2-expressing Ewing sarcoma PDX. Overall, our results suggest that low Bcl-2 expression could be used to select patients with Ewing sarcoma sensitive to nab-paclitaxel, and Bcl-2 inhibitors could improve the activity of this drug in Bcl-2-expressing Ewing sarcoma.

Abstract 6211: Targeting the tumor microenvironment of metastasis to treat metastatic Ewing sarcoma

Abstract Introduction: Ewing sarcoma (EWS) is the second most common bone cancer in adolescents and young adults. Intensive multimodal treatment cures almost 75% of patients who present with localized disease. However, only 25% of patients who present with metastases become long term survivors, and those who suffer a metastatic relapse are almost never cured. Recently, a 3-cell assembly consisting of an invasive cancer cell, a Tie2-high expressing macrophage, and an endothelial cell (termed the Tumor Microenvironment of Metastasis, or TMEM), has been described as the doorway through which breast cancer cells enter the vasculature and disseminate to distant sites. We hypothesized that EWS cells use a similar mechanism for dissemination, and that targeting TMEM offers a novel therapeutic opportunity to prevent metastatic recurrence. Methods: Formalin-fixed, paraffin embedded tissue samples were obtained from 16 patients with EWS treated at our institution, and these were evaluated for the presence of TMEM structures by immunohistochemistry. We also evaluated samples obtained from our novel, clinically relevant murine model of spontaneous distant EWS metastasis. Fragments of EWS either from cell line-derived tumors or patient-derived xenografts (PDX) orthotopically or subcutaneously. After tumors grew, mice were injected with fluorescent dextran in the tail vein and tumors harvested 60 minutes later. Intravascular dextran remains inside blood vessels because the molecular weight is too large to allow diffusion between endothelial cells. As TMEM disrupts the endothelial barrier, dextran leaks out of blood vessels and can be detected and quantified using fluorescence microscopy. Tumors were evaluated for the presence of TMEM structures by immunohistochemistry (IHC), and for the presence of localized vascular leakiness, (the hallmark of functional TMEM), by fluorescence microscopy. Results: We found TMEM assemblies in all patient samples, at numbers well in excess of those seen in samples of metastatic breast cancer. Interestingly, although there was no difference in TMEM counts from patients who were survivors compared with those who died from metastatic disease, there was an increase in TMEM counts in samples from tumors that were exposed to chemotherapy compared to diagnostic biopsy samples, something also reported in breast cancer patients. We also identified TMEM assemblies and localized vascular leakiness in EWS PDX tumors implanted orthotopically. Discussion: TMEM assemblies are present in both EWS patient samples and PDX models, suggesting that this important mechanism of tumor intravasation into the vasculature may be contributing to distant metastasis. Future work will focus on identifying drugs that inhibit the function of TMEM, with the goal of translating this work into clinical trials aimed at diminishing the burden of metastasis in EWS patients. Citation Format: Rachel Offenbacher, Yu Lin, George Karagiannis, Maja Oktay, David Loeb. Targeting the tumor microenvironment of metastasis to treat metastatic 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 6211.

Abstract 6245: 3D-models of pediatric bone sarcomas for personalized therapeutic screening

Abstract Osteosarcoma (OS) and Ewing sarcoma (ES) are the most common bone cancers in children. They are rare cancers and thus difficult to study due to scarcity of patient material, large genomic instability and a wide histological heterogeneity (in OS) or a lack of satisfactory transgenic animal model and availability of preclinical tests (in ES). There is a dire need for new models and novel therapeutic approaches. Although patient-derived xenografts (PDXs) may recapitulate human tumor biology and predict drug response, propagating PDXs in mice limits its use as a drug-testing platform. We have established and standardized ES and OS spheroid culture and developed a semi-automated drug-screening platform in tumor spheroids. We established several robust techniques for spheroid formation, with clear pathophysiological gradients, but without central necroses at the onset of drug treatment. We performed RNA-seq comparing spheroid transcription profiles to 2D culture and observed dramatic changes in overall expression patterns. We observed upregulation of genes shown to correlate with poor prognosis in OS patients. We saw upregulation of processes associated with regulation of cell migration, negative regulation of proliferation and modulation of the extracellular matrix (ECM). In addition to ES spheroid models, we created bioprinted 3D-models of ES cell lines and of cells obtained from ES PDXs, using extrusion bioprinting techniques (where cells are encapsulated within the cross-linked polymers, thus allowing homogeneous distribution and high cell density). PDX-derived cells were kept in liquid culture and as 3D-bioprinted constructs, while their transcription profiles were compared with the initial PDX. The mevalonate pathway was the most overrepresented in all ES 3D-models, consistent with predominant upregulation of this metabolic pathway integral to tumor growth and progression. After 15 days in 3D-bioprinted culture, we observed pronounced upregulation of genes involved in ECM signaling, suggesting that the construct promoted in vivo-like tumor-ECM interactions, without further promoting main proliferation and cell survival pathways, which was observed in liquid culture. Furthermore, we showed potential for combinatorial treatment with statins and confirmed feasibility of drug testing in patient-derived 3D models. Finally, as our spheroid models showed upregulation of many processes involved in metastasis (genes associated with invasion, migration, angiogenesis and hypoxia), we focused on lung as the most common site of metastasis in ES and OS patients. We are thus establishing mixed airway organoid/tumoroid cultures, to investigate further the lung metastatic niche, with a goal to provide proof of concept for patient-specific 3D-models of lung metastatic tumors to guide personalized drug selection for patients with advanced disease. Citation Format: Branka Radic Sarikas, Mathias Ilg, Marica Markovic, Caterina Sturtzel, Eva Scheuringer, Justine Zulini, Martin Metzelder, Florian Halbritter, Martin Distel, Didier Surdez, Olivier Delattre, Aleksandr Ovsianikov, Heinrich Kovar. 3D-models of pediatric bone sarcomas for personalized therapeutic screening [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 6245.

SPARC-mediated long-term retention of nab-paclitaxel in pediatric sarcomas

Secreted protein acidic and rich in cysteine (SPARC) is a matricellular glycoprotein overexpressed by several cancers. Because SPARC shows high binding affinity to albumin, we reasoned that pediatric sarcoma xenografts expressing SPARC would show enhanced uptake and accumulation of nanoparticle albumin-bound (nab)-paclitaxel, a potent anticancer drug formulation. We first evaluated the expression of SPARC in patient-derived xenografts (PDXs) of Ewing sarcoma, rhabdomyosarcoma and osteosarcoma, finding variable SPARC gene expression that correlated well with SPARC protein measured by immunoblotting. We revealed that the activity of the fusion gene chimera EWSR1-FLI1, the genetic driver of Ewing sarcoma, leads to lower expression of the gene SPARC in these tumors, likely due to enriched acetylation marks of the histone H3 lysine 27 at regions including the SPARC promoter and potential enhancers. Then, we used SPARC-edited Ewing sarcoma cells (A673 line) to demonstrate that SPARC knocked down (KD) cells accumulated significantly less amount of nab-paclitaxel in vitro than SPARC wild type (WT) cells. In vivo, SPARC KD and SPARC WT subcutaneous xenografts in mice achieved similar maximum intratumoral concentrations of nab-paclitaxel, though drug clearance from SPARC WT tumors was significantly slower. We confirmed such SPARC-mediated long-term intratumoral accumulation of nab-paclitaxel in Ewing sarcoma PDX with high expression of SPARC, which accumulated significantly more nab-paclitaxel than SPARC-low PDX. SPARC-high PDX responded better to nab-paclitaxel than SPARC-low tumors, although these results should be taken cautiously, given that the PDXs were established from different patients that could have specific determinants predisposing response to paclitaxel. In addition, SPARC KD Ewing sarcoma xenografts responded better to soluble docetaxel and paclitaxel than to nab-paclitaxel, while SPARC WT ones showed similar response to soluble and albumin-carried drugs. Overall, our results show that pediatric sarcomas expressing SPARC accumulate nab-paclitaxel for longer periods of time, which could have clinical implications for chemotherapy efficacy.

Abstract 2079: Collection of patient-derived xenografts (PDX) to study the biology and therapy of bone sarcomas

Abstract Osteosarcoma (OS) and Ewing's sarcoma (EWS) are the two most common pediatric solid tumors, after brain tumors. Multimodal treatments have significantly improved prognosis in localized disease but outcome is still poor in metastatic patients, for whom therapeutic options are often inadequate. Preclinical drug testing to identify promising treatment options that match the molecular make-up of these tumors is hampered by the lack of appropriate and molecularly well-characterized patient-derived models. To address this need, a panel of patient-derived- xenografts (PDXs) was established by subcutaneous implantation of fresh, surgically resected OS and EWS tumors in NSG mice. Tumors were re-transplanted to next mice generations and fragments were collected for histopathological and molecular characterization. A model was considered established after observing stable histological and molecular features for at least three passages. To evaluate the similarity of the model with primary tumor, we performed a global gene expression profiling and tissue microarrays (TMA), to assess tumor specific biomarkers on tissues from OS/EWS tumors and their PDXs (1st and 3rd passage). Moreover, we verified the feasibility of these models for preclinical drug testing. We implanted 61 OS and 29 EWS samples: 14/38 (37%) primary OS and 9/23 (39%) OS lung metastases successfully engrafted; while among EWS, 5/26 (19%) primary samples and 1/3 (33%) metastases were established. Comparison between patient samples and PDXs, highlighted that histology and genetic characteristics of PDXs were stable and maintained over passages. In particular, correlative analysis between OS and EWS samples and their PDXs was extremely high (Pearson's r range r=0.94-0.96), while patient-derived primary cultures displayed reduced correlation with human samples (r=0.90-0.93), indicating that in vitro adaptation superimpose molecular alterations that create genetic diversion from original tumors. No significant differentially expressed gene profile was observed from the comparison between EWS samples and PDXs (fold change > 2, adjusted p <0.05 at paired t-test). In OS, the comparison between OS patient-derived tumors and PDX indicated differences in 397 genes, mostly belonging to immune system functional category. This is in line with the idea that human immune cells are gradually replaced by murine counterparts upon engraftment in the mouse. As proof-of concept, two EWS PDX and one OS PDX have been treated with conventional and innovated drugs to test their value in terms of drug-sensitivity prediction. Overall, our study indicated that PDX models maintained the histological and genetic markers of the tumor samples and represent reliable models to test sensitivity to novel drug associations. Grants AIRC (IG 18451 to KS; IG15324 to P-LL), Italian Ministry of Health (ER2017 2364984-ERANET. to KS; RF-2013-02357552 to P-LL) Citation Format: Maria Cristina Manara, Giordano Nicoletti, Manuela Ferracin, Camilla Cristalli, Alberto Righi, Mauro Magnani, Michela Pasello, Piero Picci, Patrizia Nanni, Pier-Luigi Lollini, Katia Scotlandi. Collection of patient-derived xenografts (PDX) to study the biology and therapy of bone sarcomas [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2079.

Abstract 1625: Ureidomustine, a novel DNA-crosslinking agent shows activity in sarcoma preclinical models and lacks toxicity in normal tissues

Abstract Ureidomustine (BO-1055) is a water-soluble N- mustard derivative that has antitumor activity against a number of human tumors including prostate, colon cancers and gliomas. In this study, we investigated its pre-clinical activity in sarcoma models. We tested the activity of BO-1055 on early passages from Ewing sarcoma patient samples and patient-derived xenografts, preclinical models of rhabdomyosarcoma, desmoplastic small round cell tumor (DSRCT) and osteosarcoma. We used cell proliferation, viability, clonogenicity and cell death assays, and immunoblotting for differential expression of proteins in cells that are sensitive and resistant to BO-1055. We studied the toxicity of BO-1055 on cardiac myocytes, hematopoietic stem and progenitor cells (HSPCs), epithelial, mesenchymal and endothelial cells, and performed animal toxicity experiments in mice. Efficacy of BO-1055 was tested in NSG mice bearing A673 xenografts (Ewing sarcoma) treated at doses of 10mg/kg, 20mg/kg and 30mg/kg, Q2D×5, via i.v. injection after the tumors grew to 250-500 mm3. We also investigated the efficacy of this agent in NSG mice bearing A204 xenografts and Ewing sarcoma patient-derived xenografts that were resistant to cyclophosphamide. The antitumor effects of BO-1055 in combination with standard chemotherapeutic agents were also evaluated. Ureidomustine exhibited significant cytotoxicity against the in vitro growth of Ewing sarcoma and rhabdomyosarcoma with sub μM IC50 values and was more cytotoxic than conventional DNA damaging agents. BO-1055 had less cytotoxicity against a panel of normal HSPCs. BO-1055 had moderate activity (IC50 = 2-4 μM) on DSRCT cells and had only weak activity (IC50>10μM) against osteosarcoma cell lines. Notably, this agent has no cross-resistance to Taxol and Vinblastine. Growth arrest in G2/M phase was noted at 24h and maximal apoptosis and cell death at 48h and 72h after exposure of A673 cells. Apoptosis was due to the induction of caspase 3 and 7 activity in a dose- and time- dependent manner. Cardiotoxicity was evaluated by binding of BO-1055 to hERG, which was significantly less than that of Astemizole indicating that BO-1055 has less propensity for QTc prolongation. BO-1055 significantly suppressed Ewing sarcoma patient-derived xenografts that were resistant to cyclophosphamide. Remarkably, this agent induced complete suppression of tumors in nude mice bearing A204 xenografts, without significant weight loss. In two-drug combination studies, BO-1055 exhibited synergism with etoposide, SN-38 (active metabolite of irinotecan), doxorubicin and PU-H71 (HSP-90 inhibitor). Because of its broad therapeutic window, lack of cross-resistance, better safety profile and synergistic cytotoxicity with standard drugs, BO-1055 has high potential for clinical application for treatment of Ewing sarcoma and rhabdomyosarcoma patients. Citation Format: Srikanth R. Ambati, Shieh JaeHung, Benet Pera, Elissa W.P. Wong, Eloisi Caldas Lopes, Elizabeth Peguero, Tsann-Long Su, Malcolm A.S. Moore. Ureidomustine, a novel DNA-crosslinking agent shows activity in sarcoma preclinical models and lacks toxicity in normal tissues. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 1625. doi:10.1158/1538-7445.AM2015-1625

The PARP inhibitor olaparib enhances the sensitivity of Ewing sarcoma to trabectedin.

Recent preclinical evidence has suggested that Ewing Sarcoma (ES) bearing EWSR1-ETS fusions could be particularly sensitive to PARP inhibitors (PARPinh) in combination with DNA damage repair (DDR) agents. Trabectedin is an antitumoral agent that modulates EWSR1-FLI1 transcriptional functions, causing DNA damage. Interestingly, PARP1 is also a transcriptional regulator of EWSR1-FLI1, and PARPinh disrupts the DDR machinery. Thus, given the impact and apparent specificity of both agents with regard to the DNA damage/DDR system and EWSR1-FLI1 activity in ES, we decided to explore the activity of combining PARPinh and Trabectedin in in vitro and in vivo experiments. The combination of Olaparib and Trabectedin was found to be highly synergistic, inhibiting cell proliferation, inducing apoptosis, and the accumulation of G2/M. The drug combination also enhanced γH2AX intranuclear accumulation as a result of DNA damage induction, DNA fragmentation and global DDR deregulation, while EWSR1-FLI1 target expression remained unaffected. The effect of the drug combination was corroborated in a mouse xenograft model of ES and, more importantly, in two ES patient-derived xenograft (PDX) models in which the tumors showed complete regression. In conclusion, the combination of the two agents leads to a biologically significant deregulation of the DDR machinery that elicits relevant antitumor activity in preclinical models and might represent a promising therapeutic tool that should be further explored for translation to the clinical setting.

Abstract 2733: Antitumor activity of the PARP inhibitor E7449 in Ewing's sarcoma

Abstract Ewing's sarcoma affects mostly adolescents and young adults with tumors that predominate in bone, characterized by the presence of fusion proteins created by chromosomal translocations (EWS-FLI1, EWS-ERG etc.). Recent studies demonstrated an interaction between EWS fusion proteins and PARP1; increased DNA damage and elevated PARP levels upon fusion protein expression as well as sensitivity to PARP inhibition have been reported. E7449, a potent, orally available PARP inhibitor was evaluated in various xenograft models of Ewing's sarcoma as a single agent and in combination with TMZ or irinotecan, chemotherapies often used in relapsed patients. In an RD-ES (ATCC® HTB166™) Ewing's sarcoma (EWS-FLI1) s.c. xenograft model, treatment with single agent E7449 or TMZ resulted in no antitumor activity. However, when combined exquisite synergy that resulted in tumor regression and tumor-free mice was observed, even at the low E7449 combination dose of 1 mg/kg. E7449 dose responsive re-growth of tumors was observed and 8/8 mice remained tumor-free in the highest dose group (30 mg/kg) at study termination. Enhanced toxicity as measured by body weight loss was observed in combination treatment groups but mice generally recovered well. Significant antitumor activity was observed for irinotecan alone in the RD-ES model; combination with E7449 enhanced that activity. Combining TMZ and irinotecan was also efficacious in the RD-ES model. Antitumor activity was further enhanced by the addition of E7449 which also potentiated combination toxicity. Studies are ongoing to optimize dosing of E7449 and chemotherapy and to identify a treatment schedule that maximizes activity and tolerability of the various combinations. Additionally, the activity of E7449 alone and in combination was evaluated in 2 Ewing's sarcoma patient-derived xenograft (PDx) models. In one model (CTG-0143) neither TMZ nor E7449 as single agents had significant antitumor activity. However, the combination of E7449 + TMZ resulted in striking synergy and tumor regression. Although tumors were large when treatment started, significant antitumor activity was also observed with single agent irinotecan and combination treatment. The second model (CTG-0142) was sensitive to E7449 alone at high dose and to both chemotherapies (TMZ and irinotecan) as single agents. Addition of E7449 to either TMZ or irinotecan resulted in enhanced anti-tumor activity as measured by significantly delayed tumor re-growth following regression. Biomarker analysis is ongoing, including gene expression profiling and assessment of the DNA damage response in tumors from RD-ES and PDx models following drug treatments. In conclusion, the combination of E7449 with TMZ and irinotecan resulted in highly potent anti-cancer activity against cell line and PDx models of Ewing's sarcoma. These data support the assessment of E7449 as a combination therapy for Ewing's sarcoma in clinical trials. Citation Format: Sharon McGonigle, Zhihong Chen, Jingzang Tao Miu, Kuan-Chun Huang, Donna Kolber-Simonds, Nanding Zhao, Natalie C. Twine, Qiongfang Cao, Galina Kuznetsov, Shanqin Xu, Kenichi Nomoto. Antitumor activity of the PARP inhibitor E7449 in Ewing's sarcoma. [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 2733. doi:10.1158/1538-7445.AM2014-2733

Defense Technical Information Center (DTIC): Its role in the USAF scientific and technical information program

Secreted protein acidic and rich in cysteine (SPARC) is a matricellular glycoprotein overexpressed by several cancers. Because SPARC shows high binding affinity to albumin, we reasoned that pediatric sarcoma xenografts expressing SPARC would show enhanced uptake and accumulation of nanoparticle albumin-bound (nab)-paclitaxel, a potent anticancer drug formulation. We first evaluated the expression of SPARC in patient-derived xenografts (PDXs) of Ewing sarcoma, rhabdomyosarcoma and osteosarcoma, finding variable SPARC gene expression that correlated well with SPARC protein measured by immunoblotting. We revealed that the activity of the fusion gene chimera EWSR1-FLI1, the genetic driver of Ewing sarcoma, leads to lower expression of the gene SPARC in these tumors, likely due to enriched acetylation marks of the histone H3 lysine 27 at regions including the SPARC promoter and potential enhancers. Then, we used SPARC-edited Ewing sarcoma cells (A673 line) to demonstrate that SPARC knocked down (KD) cells accumulated significantly less amount of nab-paclitaxel in vitro than SPARC wild type (WT) cells. In vivo, SPARC KD and SPARC WT subcutaneous xenografts in mice achieved similar maximum intratumoral concentrations of nab-paclitaxel, though drug clearance from SPARC WT tumors was significantly slower. We confirmed such SPARC-mediated long-term intratumoral accumulation of nab-paclitaxel in Ewing sarcoma PDX with high expression of SPARC, which accumulated significantly more nab-paclitaxel than SPARC-low PDX. SPARC-high PDX responded better to nab-paclitaxel than SPARC-low tumors, although these results should be taken cautiously, given that the PDXs were established from different patients that could have specific determinants predisposing response to paclitaxel. In addition, SPARC KD Ewing sarcoma xenografts responded better to soluble docetaxel and paclitaxel than to nab-paclitaxel, while SPARC WT ones showed similar response to soluble and albumin-carried drugs. Overall, our results show that pediatric sarcomas expressing SPARC accumulate nab-paclitaxel for longer periods of time, which could have clinical implications for chemotherapy efficacy.