Ewing sarcoma is characterized by multiple deregulated pathways that mediate cell survival and proliferation.
Heat shock protein 90 (HSP90) is a critical component of the multi-chaperone complexes that regulate the disposition and activity of a large number of
proteins involved in cell-signaling systems. We tested the efficacy of
PU-H71, a novel HSP90 inhibitor in
Ewing sarcoma cell lines, primary samples, benign mesenchymal stromal cells and hematopoietic stem cells. We performed cell cycle analysis, clonogenic assay, immunoblot analysis and reverse phase
protein array in Ewing cell lines and in vivo experiments in NSG and nude mice using the A673 cell line. We noted a significant therapeutic window in the activity of
PU-H71 against Ewing cell lines and benign cells.
PU-H71 treatment resulted in G2/M phase arrest. Exposure to
PU-H71 resulted in depletion of critical
proteins including AKT, pERK, RAF-1, c-MYC, c-KIT, IGF1R, hTERT and
EWS-FLI1 in Ewing cell lines. Our results indicated that
Ewing sarcoma tumor growth and the metastatic burden were significantly reduced in the mice injected with
PU-H71 compared to the control mice. We also investigated the effects of
bortezomib, a
proteasome inhibitor, alone and in combination with
PU-H71 in
Ewing sarcoma. Combination index (CI)-Fa plots and normalized isobolograms indicated synergism between
PU-H71 and
bortezomib.
Ewing sarcoma xenografts were significantly inhibited when mice were treated with the combination compared to vehicle or either
drug alone. This provides a strong rationale for clinical evaluation of
PU-H71 alone and in combination with
bortezomib in
Ewing sarcoma.