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CDK 7/9 inhibition amplifies mithramycin's suppression of Ewing sarcoma cell proliferation
Author(s) -
Flores Guillermo,
Everett Joel,
Osgood Christy,
Madaj Zachary,
Grohar Patrick J
Publication year - 2017
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.31.1_supplement.178.10
Subject(s) - cyclin dependent kinase , sarcoma , cancer research , fli1 , ewing's sarcoma , cell growth , chemistry , cyclin dependent kinase 6 , synovial sarcoma , biology , transcription factor , cell , cell cycle , gene , medicine , biochemistry , pathology
Background Ewing sarcoma is driven by the fusion of the EWSR1 and FLI1 genes generated by a t(11:22) chromosomal translocation. Ewing sarcoma cells depend on the EWS‐FLI1 fusion product, a constitutively active oncogenic transcription factor, and become nonviable when EWS‐FLI1 driven transcription is inhibited. Mithramycin, a natural antineoplastic compound, has been identified as a potent EWS‐FLI1 inhibitor and is highly toxic to Ewing sarcoma cells. However, clinical trials of mithramycin in patients with Ewing sarcoma have been limited due to toxicity. To overcome this challenge, we sought to identify genes that sensitize Ewing sarcoma cells to mithramycin when inhibited. We identified RNA pol II inhibition as a potent sensitizer to mithramycin and phenocopied this with the dual cyclin dependent kinase (CDK) 7/9 inhibitors PHA‐767491 and SNS‐032. We show a marked suppression of cellular proliferation in response to mithramycin in combination with either CDK 7/9 inhibitor at concentrations significantly lower than mithramycin alone. Methods We utilized matrix drug screening with small molecule CDK inhibitors in combination with mithramycin. We measured cell viability using MTS assays and time lapse microscopy in Ewing sarcoma cells. Results were correlated with EWS‐FLI1 activity using western blot analyses and qPCR. Synergy between small molecule inhibitors and mithramycin was measured using Bliss independence. Results PHA‐767491 and SNS‐032 displayed strong synergy when combined with mithramycin. These combinations decreased the concentration of mithramycin necessary to block Ewing sarcoma cell proliferation, measured by multiple complementary assays. These results reflected effects on EWS‐FLI1 activity. Conclusions We have identified a synergistic relationship between the small molecules PHA‐767491 and SNS‐032 with mithramycin in Ewing sarcoma cells. Future work will include siRNA validation of the CDK 7/9 inhibition and mithramycin combination, analysis of genome wide expression changes, and characterization of the efficacy of a PHA‐767491 or SNS‐032 and mithramycin drug combination in vivo with the eventual goal of introducing it into the clinic.