Tanespimycin Prevents Bortezomib Toxicity and Preserves Neuronal Morphology in Primary Rat Dorsal Root Ganglion Cultures.

Tanespimycin, an inhibitor of Hsp90, is in phase 3 clinical trials in combination with bortezomib in patients with relapsed/refractory multiple myeloma (MM). The combination of tanespimycin and bortezomib produces synergistic antitumor activity and enhanced proteasome inhibition in primary MM cells (Mitsiades, Blood, 2006). In a phase 1/2 study in 72 patients with relapsed/refractory myeloma, tanespimycin + bortezomib produced durable responses in patients including bortezomib-refractory patients. Bortezomib-induced peripheral neuropathy (PN) is the key dose-limiting toxicity in patients with MM. In rat models of bortezomib-induced PN, tanespimycin is neuroprotective and can reverse bortezomib-induced PN. No Grade 3/4 PN was reported in the phase 1/2 study.

To explore the mechanism of tanespimycin-mediated neuroprotection from bortezomib-induced neurotoxicity in primary rat dorsal root ganglion (DRG) cells.

Differentiated DRG cultures prepared from rat fetuses were treated for up to 24 hours with tanespimycin alone, bortezomib alone, or the combination of tanespimycin + bortezomib. Total cell ATP, caspase 3/7 induction, calpain activity, and chymotrypsin-like activity of the proteasome were analyzed as measures of cell viability, apoptosis, neuron-specific protease activity, and proteasome function, respectively. Neuronal morphology was evaluated by light microscopy.

At concentrations ≥100 nM, both bortezomib and tanespimycin induced a rapid increase in apoptosis (up to a fivefold increase in caspase activity) associated with reduced cell viability (based on ATP levels) to 20% of control values. In addition, bortezomib at ≥100 nM visibly shortened neurite extensions from DRG cell bodies. When DRG cells were co-exposed to both tanespimycin (500 nM) and bortezomib (320 nM–2.5 μM), the inhibitory effects of bortezomib on cell viability and neurite extension were ameliorated. At concentrations as low as 100 nM, tanespimycin completely and reproducibly abrogated the induction of caspase activity at all bortezomib concentrations tested (ie, 0.15–100 μM). When administered alone, bortezomib and tanespimycin had opposite effects on proteasome and calpain activity; bortezomib inhibited all proteasome and calpain activity at concentrations >1 μM, while tanespimycin (100 nM–10 μM) induced calpain and proteasome activity by up to fivefold and threefold, respectively. When DRG cells were exposed to both bortezomib (<1 μM) and tanespimycin, tanespimycin (10 nM–1 μM) induction of the proteasome ameliorated bortezomib proteasome inhibition. Similarly, but to a lesser extent, tanespimycin also overcame the bortezomib-induced inhibition of calpain activity.

In primary rat DRG cells, tanespimycin prevented bortezomib-induced apoptosis and loss of cell viability in cultures and restored neuronal morphology. These neuroprotective effects of tanespimycin are consistent with the effects observed in rat models of bortezomib-induced PN and with the lack of severe PN observed in patients in the phase 1/2 study of tanespimycin + bortezomib in MM. The mechanism for the protective effect of tanespimycin on bortezomib-induced neuronal toxicity in MM is currently being explored.

Flint:Bristol-Myers Squibb: Employment, Equity Ownership. Moulin:Bristol-Myers Squibb: Employment. Oberdoerster:Bristol-Myers Squibb: Employment. Berman:Bristol-Myers Squibb: Employment.