A Novel Orally Bioavailable Heat Shock Protein 90 Inhibitor Induces Apoptosis and Synergises with Bortezomib in Human Multiple Myeloma.

Abstract 1838

Poster Board I-864

Heat shock proteins (HSP) are ubiquitously expressed in human cells. The expression of HSP proteins is transcriptionally regulated and escalates in response to stress, furthermore, HSPs assist in the folding and establishment of new proteins as well as removing unwanted protein aggregation by transportation to the proteasome. In cancer, HSPs stabilise and protect oncoproteins such as AKT and p53, increase angiogenesis by VEGF induction and support metastasis by assisting MMP2. We evaluated the therapeutic potential of a novel orally bioavailable HSP90 inhibitor (HSP90i) against a panel of human myeloma cell lines (HMCL) and primary tumour samples from multiple myeloma (MM) patients. 8 genetically heterogenous HMCL were screened by MTS assay and found to be sensitive to HSP90i with an IC₅₀ of 50–100nM at 72 hours. This was associated with accumulation of cells in the G₀-G₁ phase and decreasing number of cells in the S and G₂/M phases indicative of cell cycle arrest. Apoptosis was confirmed by the presence of increasing numbers of propidium iodide stained cells as measured by flow cytometry and PARP cleavage on immunoblot analysis within 24 hours of HSP90i treatment. Three HMCLs, NCI H929, an IL6 independent cell line and OCI-MY1 and U266, two IL6-dependent cell lines were used for mechanistic studies. HSP90i was found to down regulate the client proteins IL-6R, STAT3, MEK, AKT, BID, MCL-1, NFkB and IkB conversely the chaperone proteins HSP70 and HSP27 were up regulated in all 3 HMCL. Importantly HSP90i treatment was associated with a rapid reduction in the phosphorylated forms of STAT3, MEK and NFkB. Furthermore, the down-regulation of ^P-STAT3 and IL6-R protein was not reversed by exposure to exogenous IL-6. Combination studies with bortezomib, a proteasome inhibitor, were undertaken to define the extent of possible synergy and schedule dependency. The combination of bortezomib with HSP90i displayed strong synergism in both IL-6 dependent and IL-6 independent HMCL with maximal killing seen with simultaneous drug exposure (CI = 0.089 — 0.338). Finally, HSP90i was also tested against primary MM tumour cells from patients with relapsed/refractory MM with killing of CD38^+veCD45^−ve cells, as evidenced by Apo2.7 staining, seen with concentrations of HSP90i from 100nM to 1000nM at 24 and 48 hours following treatment. These in vitro findings justify the further evaluation of HSP90i as a potential therapeutic agent for MM.