Antileukemic Activity of Shepherdin, a Novel Targeted Inhibitor of the Survivin-Hsp90 Complex.

Cancer therapy now aims at disabling critical signaling pathways essential for tumor cell maintenance without affecting normal cellular functions. The molecular interaction between heat shock protein 90 (Hsp90) and survivin functions as a crossroad to preserve cell viability and cell proliferation in malignant cells, and offers tangible therapeutic prospects. We recently used structure-based rational design to identify Shepherdin, a cell permeable peptidomimetic from the survivin sequence Lys79-Leu87, which inhibits the complex between survivin and Hsp90 in tumor cells (K_d~80 nM). Here, we tested the activity of Shepherdin against a panel of acute leukemia cell lines (AML). We identified a shorter cell-permeable Shepherdin variant Lys79-Gly83 that docked in the Hsp90 ATP pocket by molecular dynamics simulations, bound recombinant Hsp90 in vitro, and competitively inhibited the survivin-Hsp90 complex. When administered in cell culture, Shepherdin rapidly (<30 min) accumulated intracellularly in all AML cells tested, as well as in quiescent or phytohemagglutinin (PHA)-stimulated normal mononuclear cells. A brief, 30-min exposure of AML cells to Shepherdin was sufficient to induce complete cell killing by apoptotic and non-apoptotic mechanisms, with associated mitochondrial dysfunction and effector caspase activity. Conversely, Shepherdin did not affect colony formation of isolated CD34⁺ hematopoietic progenitors or viability of resting or proliferating normal mononuclear cells. When compared with known Hsp90 antagonists currently in the clinic, i.e. 17-AAG, Shepherdin was superior for rapidity and efficacy of antileukemic activity. Shepherdin induced collapse of Hsp90 chaperone functions in solid tumor cell types with destabilization of multiple Hsp90 client proteins, but did not affect the levels of pro-survival Hsp70 in AML cells, which is typically upregulated following Hsp90 inhibition by 17-AAG. In conclusion, Shepherdin provides a novel approach for rational therapy of acute leukemia.