Bortezomib alters microtubule polymerization and axonal transport in rat dorsal root ganglion neurons

Abstract

Bortezomib is part of a newer class of chemotherapeutic agents whose mechanism of action is inhibition of the proteasome-ubiquitination system. Primarily used in multiple myeloma, bortezomib causes a sensory-predominant axonal peripheral neuropathy in approximately 30% of patients. There are no established useful preventative agents for bortezomib-induced peripheral neuropathy (BIPN), and the molecular mechanisms of BIPN are unknown. We have developed an in vitro model of BIPN using rat dorsal root ganglia neuronal cultures. At clinically–relevant dosages, bortezomib produces a sensory axonopathy as evidenced by whole explant outgrowth and cell survival assays. This sensory axonopathy is associated with alterations in tubulin and results in accumulation of somatic tubulin without changes in microtubule ultrastructure. Furthermore, we observed an increased proportion of polymerized tubulin, but not total or acetylated tubulin, in bortezomib-treated DRG neurons. Similar findings are observed with lactacystin, an unrelated proteasome-inhibitor, which argues for a class effect of proteasome inhibition on dorsal root ganglion neurons. Finally, there is a change in axonal transport of mitochondria induced by bortezomib in a time-dependent fashion. In summary, we have developed an in vitro model of BIPN that recapitulates the clinical sensory axonopathy; this model demonstrates that bortezomib induces an alteration in microtubules and axonal transport. This robust model will be used in future mechanistic studies of BIPN and its prevention.

Toxic Effects of Bortezomib on Primary Sensory Neurons and Schwann Cells of Adult Mice

“…When evaluating cultured sensory neurons exposed to bortezomib, we observed a clear alteration in the arborization of the neurons, as previously described (Staff et al 2013). The neurites were thinner and their processes were shorter than in control neurons.…”

“…Values are expressed as mean and SEM. *p \ 0.05 Neurotox Res higher than the ones of our study (Staff et al 2013). However, a previous study already demonstrated that embryonic and adult DRG neurons had different sensitivity to bortezomib (Meregalli et al 2013).…”

“…However, a previous study already demonstrated that embryonic and adult DRG neurons had different sensitivity to bortezomib (Meregalli et al 2013). Therefore, to study the chemotherapy-induced neurotoxicity in vitro, it seems more adequate the use of adult primary neuronal cultures since it is closer to the in vivo model and, thus, to the clinical situation (Staff et al 2013;Meregalli et al 2013).…”

“…Conversely, other studies proposed that BIPN was mainly due to dysfunctions at the neuronal level, and changes in axons and myelin might be secondary to this initial event (Bruna et al 2010). Other studies reported neuronal alterations after bortezomib exposure, proposing that the neurotoxicity is related either to microtubule polymerization and stabilization (Poruchynsky et al 2008;Staff et al 2013;Meregalli et al 2013) or to nuclear accumulation of ubiquitylated proteins and poly(A)RNAs in nuclear granules, with consequent reduction of transcription and alteration of mRNA translation (Casafont et al 2010;Palanca et al 2013).…”

Stem Cell-Derived Immature Human Dorsal Root Ganglia Neurons to Identify Peripheral Neurotoxicants

“…In the past, rodent sensory neurons/DRG have allowed important studies of the mechanisms of peripheral neurotoxicity/degeneration [45,46]. However, the overwhelming majority of reports in this area focused on the underlying mechanisms and never compared more than three toxicants.…”