Multiple myeloma is a clonal plasma cell malignancy accompanied by characteristic bone lesions, cytopenias, renal insufficiency and immune deficiency. The last decade has witnessed significant advances in anti-myeloma therapy with median survival extending from 2-3 years to over 7 years for patients younger than 50 years .
Heat shock protein 90 (HSP90) inhibitors are an emerging class of targeted agents in cancer therapy. HSP90 inhibition would make intuitive sense as anti-tumor therapy in cell types that depend on sustained protein homeostasis for their survival. A number of HSP90 inhibitors have demonstrated anti-myeloma activity in pre-clinical studies and at least three compounds have been evaluated in Phase I trials for relapsed/refractory myeloma [2–5]. PU-H71 is a novel purine scaffold HSP90 inhibitor that has shown pre-clinical activity in triple negative breast cancer , Bcl6 dependent lymphoma , hepatocellular carcinoma  and myeloproliferative disorders .
HSP90 family of proteins are ubiquitous molecular chaperones that are involved in folding, activation, maturation and assembly of many proteins (referred to as HSP90 client proteins or HSP90 clientele) that include essential mediators of signal transduction and cell cycle progression . The mammalian HSP90 family members include the cytosolic HSP90, the HSP90 paralogue gp96 (also known as grp94, endoplasmin, HSP90B1) in the endoplasmic reticulum (ER) and the mitochondrial protein TRAP1. There have been recent significant progresses in the understanding of both the structure and function of gp96. It serves as an obligate master chaperone for multiple Toll-like receptors [11, 12] and integrins [13, 14], neither of which could function properly in the absence of gp96. More recently, gp96 has been observed to play a critical role in lymphopoeisis in that deletion of gp96 leads to a transitional block from pro-B to pre-B cells and the inability of thymocytes to develop beyond the CD4(-)CD8(-) stage . gp96 also maintains the fidelity of the endoplasmic reticulum protein synthesis by mediating the unfolded protein response (UPR) . It shares ~50% homology at the amino acid level with its cytosolic HSP90 paralogue, with a similar domain organization consisting of an N-terminal ATP-binding domain, a charged middle domain and a C-terminal homodimerization domain .
The unfolded protein response (UPR) is a highly conserved eukaryotic protein homeostasis mechanism that is especially important for secretory cell types (e.g., hepatocytes, plasma cells, etc.) . In response to cellular stress, UPR leads to increased ER chaperones such as grp78, gp96 and calreticulin to deal with the increased load of unfolded and nascent proteins in the ER. In response to sustained cellular stress, the UPR activates the apoptotic pathway. It has been previously demonstrated that, a geldanamycin derived HSP90 inhibitor, can activate the unfolded protein response in myeloma cells .
Herein, we evaluated the in vitro anti-myeloma activity of PU-H71, a novel purine scaffold HSP90 inhibitor. We also determined if the anti-tumor activity of HSP90 inhibitors is achieved via targeting both cytosolic HSP90 and the endoplasmic reticulum HSP90 paralogue gp96.