Vincristine and bortezomib use distinct upstream mechanisms to activate a common SARM1-dependent axon degeneration program
Stefanie Geisler, … , Jeffrey Milbrandt, Aaron DiAntonio
Stefanie Geisler, … , Jeffrey Milbrandt, Aaron DiAntonio
Published September 5, 2019
Citation Information: JCI Insight. 2019;4(17):e129920. https://doi.org/10.1172/jci.insight.129920.
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Research Article Cell biology Neuroscience

Vincristine and bortezomib use distinct upstream mechanisms to activate a common SARM1-dependent axon degeneration program

Abstract

Chemotherapy-induced peripheral neuropathy is one of the most prevalent dose-limiting toxicities of anticancer therapy. Development of effective therapies to prevent chemotherapy-induced neuropathies could be enabled by a mechanistic understanding of axonal breakdown following exposure to neuropathy-causing agents. Here, we reveal the molecular mechanisms underlying axon degeneration induced by 2 widely used chemotherapeutic agents with distinct mechanisms of action: vincristine and bortezomib. We showed previously that genetic deletion of SARM1 blocks vincristine-induced neuropathy and demonstrate here that it also prevents axon destruction following administration of bortezomib in vitro and in vivo. Using cultured neurons, we found that vincristine and bortezomib converge on a core axon degeneration program consisting of nicotinamide mononucleotide NMNAT2, SARM1, and loss of NAD+ but engage different upstream mechanisms that closely resemble Wallerian degeneration after vincristine and apoptosis after bortezomib. We could inhibit the final common axon destruction pathway by preserving axonal NAD+ levels or expressing a candidate gene therapeutic that inhibits SARM1 in vitro. We suggest that these approaches may lead to therapies for vincristine- and bortezomib-induced neuropathies and possibly other forms of peripheral neuropathy.

Authors

Stefanie Geisler, Ryan A. Doan, Galen C. Cheng, Aysel Cetinkaya-Fisgin, Shay X. Huang, Ahmet Höke, Jeffrey Milbrandt, Aaron DiAntonio

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Figure 2

SARM1 is required for both vincristine- and BTZ-induced axon degeneration.

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SARM1 is required for both vincristine- and BTZ-induced axon degeneratio...
(A) WT or SARM1-KO DRG neurons were treated with 40 nM vincristine or vehicle and axon degeneration determined over time. Degeneration index ranges from 0 (completely intact) to 1 (completely fragmented). A 2-way ANOVA showed significant main effects of group F (3, 16) = 76.43, P < 0.0001; time F (3, 48) = 198.8, P < 0.0001; and interaction F (9, 48) = 110.2, P < 0.0001. Tukey’s multiple-comparisons test, ***P < 0.001 WT vincristine versus SARM1-KO vincristine 24 hours, ****P < 0.0001 (n = 5). (B) WT or SARM1-KO DRG neurons were treated with 100 nM BTZ or vehicle and axon degeneration determined at indicated times. A 2-way ANOVA showed significant main effects of group F (3, 16) = 175.2, P < 0.0001; time F (3, 48) = 290.8, P < 0.0001; and interaction F (9, 48) = 180.2, P < 0.0001. Post hoc Tukey’s multiple-comparisons test, ****P < 0.0001 WT BTZ versus SARM1-KO BTZ (n = 5). (C) WT DRG neurons expressing a control vector (vector) or SARM1 dominant-negative (SARM1-DN) transgene were treated with 100 nM BTZ and axon degeneration was determined. A 2-way ANOVA showed main effects of group F (1, 4) = 1301, P < 0.001; time F (1, 4) = 1235, P < 0.0001; and interaction F (1, 4) = 832.2, P < 0.0001; Šídák’s multiple-comparisons test, ****P < 0.0001 (n = 3). (D) Representative bright-field micrographs of WT axons, WT axons expressing the SARM1-DN transgene, and SARM1-KO axons taken 72 hours after BTZ was added. The mitochondrial potential was monitored with red tetramethylrhodamine (TMRM) fluorescence in the same axons as shown above. Original magnification 200×. (E) WT or SARM1-KO mice were treated for 4 weeks with intravenous BTZ (n = 7 per group) or vehicle (n = 5 per group), and the intraepidermal nerve fiber (IENF) density of the footpads was determined. There was significantly less IENF in BTZ-treated WT mice compared with vehicle-treated WT and BTZ-treated SARM1-KO mice. WT vehicle: 28.7 ± 3.2, and n = 5; WT BTZ: 13.5 ± 3.8, and n = 7; SARM1-KO vehicle: 28.0 ± 3.4, and n = 5; SARM1-KO BTZ: 25.8 ± 2.7, and n = 7; One-way ANOVA, F (3, 20) = 5.827 and P = 0.0050; post hoc Tukey’s, *P < 0.05. (F) Representative images of IENF densities stained with the pan-neuronal marker PGP 9.5 (black, arrow) of WT (top row) or SARM1-KO (bottom row) mice treated with vehicle (left) or BTZ (right). Original magnification 400×.