No abstract

Background: Multimodality treatment of Ewing sarcoma (EwS), a highly malignant bone and soft tissue tumor, provides a cure in most patients, but in the metastatic stage the prognosis remains bleak; new systemically effective therapeutic options are urgently needed. The multikinase inhibitor lestaurtinib (CEP701) is an orphan drug that has been clinically studied in various carcinomas, refractory neuroblastoma, and acute myeloid leukemia, but has not yet established itself as a standard therapy. Methods: We performed an in vitro screening of 142 compounds with different mechanisms of action at two different concentrations and measured cell viability 120 hours after the first of two days of treatment. Lestaurtinib was further tested as a lead compound in various cell biology assays in a group of EwS cell lines genetically representing clinically predominant tumor characteristics. Results: Lestaurtinib was among the 21 of 142 (14.8%) compounds tested that were sensitive, i.e., reduced cell viability to below 20% at 1 µM and/or 10 µM in at least four of five EwS cell lines tested. In an expanded group of six other EwS cell lines, 72 hours of treatment with lestaurtinib resulted in significantly reduced cell growth compared to human mesenchymal stem cells. In several colony formation assays, a significant reduction in the number or size of colonies was observed, both when treatment occurred prior to colony growth and when colonies that had already formed were treated. Lestaurtinib induced apoptosis with a significant increase in PI+/FTIC+-driven cells, significantly promoted anoikis, and prevented cell migration by FACS, low adherence flow cytometry analysis, and scratch assays, respectively. Cell cycle analyses by FACS showed significant G2-M cell cycle arrest after 24 hours of treatment. Lestaurtinib also displayed a synergistic cytotoxic effect with doxorubicin, an established agent for the treatment of EwS, and exhibited a favorable dose-reduction index in all EwS cell lines tested. Notably, treatment with lestaurtinib significantly reduced doxorubicin dose at high efficacy levels (Fa 0.97) and low combination index values. Conclusion: Our results highlight the in vitro efficacy of lestaurtinib in EwS and warrant further testing in an in vivomodel as a single agent and in combination with doxorubicin. Citation Format: Samet Dayi, Christiane Schaefer, Anne Albers, Kiruthiga Balusamy, Marc Hotfilder, Maximilian Kerkhoff, Leonhard Koch, Hatice Önder, Claudia Rössig, Uta Dirksen, Stefan K. Zöllner. Preclinical testing of lestaurtinib (CEP701) as a single and combination agent for the treatment of 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 6730.

Hemangioblastomas (HBs) of the central nervous system are highly vascular neoplasms that occur sporadically or as a manifestation of von Hippel-Lindau (VHL) disease. Despite their benign nature, HBs are clinically heterogeneous and can be associated with significant morbidity due to mass effects of peritumoral cysts or tumor progression. Underlying molecular factors involved in HB tumor biology remain elusive. We investigated genome-wide DNA methylation profiles and clinical and histopathological features in a series of 47 HBs from 42 patients, including 28 individuals with VHL disease. Thirty tumors occurred in the cerebellum, 8 in the brainstem and 8 HBs were of spinal location, while 1 HB was located in the cerebrum. Histologically, 12 HBs (26%) belonged to the cellular subtype and exclusively occurred in the cerebellum, whereas 35 HBs were reticular (74%). Unsupervised clustering and dimensionality reduction of DNA methylation profiles revealed two distinct subgroups. Methylation cluster 1 comprised 30 HBs of mainly cerebellar location (29/30, 97%), whereas methylation cluster 2 contained 17 HBs predominantly located in non-cerebellar compartments (16/17, 94%). The sum of chromosomal regions being affected by copynumber alterations was significantly higher in methylation cluster 1 compared to cluster 2 (mean 262 vs. 109 Mb, p = 0.001). Of note, loss of chromosome 6 occurred in 9/30 tumors (30%) of methylation cluster 1 and was not observed in cluster 2 tumors (p = 0.01). No relevant methylation differences between sporadic and VHL-related HBs or cystic and non-cystic HBs could be detected. Deconvolution of the bulk DNA methylation profiles revealed four methylation components that were associated with the two methylation clusters suggesting cluster-specific celltype compositions. In conclusion, methylation profiling of HBs reveals 2 distinct subgroups that mainly associate with anatomical location, cytogenetic profiles and differences in cell type composition, potentially reflecting different cells of origin.