midostaurin and Sarcoma--Ewing

Improvements in survival for Ewing sarcoma pediatric and adolescent patients have been modest over the past 20 years. Combinations of anticancer agents endure as an option to overcome resistance to single treatments caused by compensatory pathways. Moreover, combinations are thought to lessen any associated adverse side effects through reduced dosing, which is particularly important in childhood tumors. Using a parallel phenotypic combinatorial screening approach of cells derived from three pediatric tumor types, we identified Ewing sarcoma-specific interactions of a diverse set of targeted agents including approved drugs. We were able to retrieve highly synergistic drug combinations specific for Ewing sarcoma and identified signaling processes important for Ewing sarcoma cell proliferation determined by EWS-FLI1 We generated a molecular target profile of PKC412, a multikinase inhibitor with strong synergistic propensity in Ewing sarcoma, revealing its targets in critical Ewing sarcoma signaling routes. Using a multilevel experimental approach including quantitative phosphoproteomics, we analyzed the molecular rationale behind the disease-specific synergistic effect of simultaneous application of PKC412 and IGF1R inhibitors. The mechanism of the drug synergy between these inhibitors is different from the sum of the mechanisms of the single agents. The combination effectively inhibited pathway crosstalk and averted feedback loop repression, in EWS-FLI1-dependent manner. Mol Cancer Ther; 16(1); 88-101. ©2016 AACR.

Topics: Animals; Antigens, CD; Antineoplastic Agents; Cell Line, Tumor; Computational Biology; Disease Models, Animal; Drug Discovery; Drug Evaluation, Preclinical; Drug Interactions; Drug Screening Assays, Antitumor; Humans; Molecular Targeted Therapy; Oncogene Proteins, Fusion; Phosphorylation; Protein Kinase Inhibitors; Proteomics; Proto-Oncogene Protein c-fli-1; Receptor, IGF Type 1; Receptor, Insulin; Receptors, Somatomedin; RNA-Binding Protein EWS; Sarcoma, Ewing; Signal Transduction; Staurosporine; Xenograft Model Antitumor Assays

Ewing's sarcoma family of tumors (EFT) is characterized by the presence of chromosomal translocations leading to the expression of oncogenic transcription factors such as, in the majority of cases, EWS/FLI1. Because of its key role in Ewing's sarcoma development and maintenance, EWS/FLI1 represents an attractive therapeutic target. Here, we characterize PHLDA1 as a novel direct target gene whose expression is repressed by EWS/FLI1. Using this gene and additional specific well-characterized target genes such as NROB1, NKX2.2 and CAV1, all activated by EWS/FLI1, as a read-out system, we screened a small-molecule compound library enriched for FDA-approved drugs that modulated the expression of EWS/FLI1 target genes. Among a hit-list of nine well-known drugs such as camptothecin, fenretinide, etoposide and doxorubicin, we also identified the kinase inhibitor midostaurin (PKC412). Subsequent experiments demonstrated that midostaurin is able to induce apoptosis in a panel of six Ewing's sarcoma cell lines in vitro and can significantly suppress xenograft tumor growth in vivo. These results suggest that midostaurin might be a novel drug that is active against Ewing's cells, which might act by modulating the expression of EWS/FLI1 target genes.

Topics: Animals; Antineoplastic Agents; Apoptosis; Caveolin 1; Cell Line, Tumor; Cell Survival; Enzyme Inhibitors; Gene Expression Regulation, Neoplastic; Homeobox Protein Nkx-2.2; Homeodomain Proteins; Humans; Mice; Mice, Inbred NOD; Nuclear Proteins; Oncogene Proteins, Fusion; Proto-Oncogene Protein c-fli-1; Random Allocation; RNA Interference; RNA-Binding Protein EWS; RNA, Small Interfering; Sarcoma, Ewing; Small Molecule Libraries; Staurosporine; Transcription Factors; Xenograft Model Antitumor Assays; Zebrafish Proteins