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Microtubule-Targeting Agents Eribulin and Paclitaxel Differentially Affect Neuronal Cell Bodies in Chemotherapy-Induced Peripheral Neuropathy

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Abstract

Chemotherapy-induced peripheral neuropathy (CIPN) is a common side effect of anticancer treatment with microtubule-targeted agents (MTAs). The frequency of severe CIPN, which can be dose limiting and even life threatening, varies widely among different MTAs. For example, paclitaxel induces a higher frequency of severe CIPN than does eribulin. Different MTAs also possess distinct mechanisms of microtubule-targeted action. Recently, we demonstrated that paclitaxel and eribulin differentially affect sciatic nerve axons, with paclitaxel inducing more pronounced neurodegenerative effects and eribulin inducing greater microtubule stabilizing biochemical effects. Here, we complement and extend these axonal studies by assessing the effects of paclitaxel and eribulin in the cell bodies of sciatic nerve axons, housed in the dorsal root ganglia (DRG). Importantly, the microtubule network in cell bodies is known to be significantly more dynamic than in axons. Paclitaxel induced activating transcription factor 3 expression, a marker of neuronal stress/injury. Paclitaxel also increased expression levels of acetylated tubulin and end binding protein 1, markers of microtubule stability and growth, respectively. These effects are hypothesized to be detrimental to the dynamic microtubule network within the cell bodies. In contrast, eribulin had no significant effect on any of these parameters in the cell bodies. Taken together, DRG cell bodies and their axons, two distinct neuronal cell compartments, contain functionally distinct microtubule networks that exhibit unique biochemical responses to different MTA treatments. We hypothesize that these distinct mechanistic actions may underlie the variability seen in the initiation, progression, persistence, and recovery from CIPN.

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Acknowledgements

We are grateful to Nichole LaPointe, Brett Cook, Benjamin Reese, Patrick Keeley, Jennifer Smith, and Mary Raven for valuable discussions and to Benjamin Reese for sharing equipment with us. We also acknowledge the use of instruments in the NRI-MCDB Microscopy Facility at UCSB (supported in part by the Office of The Director, National Institutes of Health of the NIH under Award # S10OD010610). This work was supported by grants from EISAI (to SCF, BSS, MAJ, and LW) and the NIH/NCI (R01CA161056) to BSS.

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Authors and Affiliations

  1. Neuroscience Research Institute, University of California, Santa Barbara, CA, 93106, USA

    Sarah J. Benbow, Bridget Kulesh, April Savage, Mary Ann Jordan, Leslie Wilson & Stuart C. Feinstein

  2. Department of Molecular, Cellular and Developmental Biology, University of California, Santa Barbara, CA, USA

    Sarah J. Benbow, Bridget Kulesh, April Savage, Mary Ann Jordan, Leslie Wilson & Stuart C. Feinstein

  3. Johns Hopkins Drug Discovery Program, Johns Hopkins School of Medicine, Baltimore, MD, USA

    Krystyna M. Wozniak & Barbara S. Slusher

  4. Departments of Neurology, Psychiatry, Neuroscience, Medicine and Oncology, Johns Hopkins School of Medicine, Baltimore, MD, USA

    Barbara S. Slusher

  5. Eisai Inc., Andover, MA, USA

    Bruce A. Littlefield

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  1. Sarah J. Benbow
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Correspondence to Stuart C. Feinstein.

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Electronic Supplementary Material

Supplemental figure 1

Image set example. Each image set contains one section from each treatment condition incubated with the same antibody dilution and imaged using the same acquisition settings. VehPac = Vehicle for paclitaxel, Pac = paclitaxel, Veherb = Vehicle for eribulin, and Erb = eribulin. (GIF 150 kb).

High Resolution Image (TIFF 4642 kb).

Supplemental figure 2

Separation of neuronal cell bodies from non-neuronal cells within sectioned DRG tissue. a Representative stained DRG tissue section. Merged channel image containing Cy2 (e.g., α-tubulin, green) and βIII-tubulin (red) signals. b Cy2 channel (green, used for several proteins of interest throughout this study). c βIII-tubulin signal used as a marker for neuronal cells (cy3 channel, red). d Manually defined region of interest to separate DRG tissue from axonal tissue within a section. e Merged (cy2 (green, and βIII-tubulin (red))) image showing the identification of βIII-tubulin positive regions (grey) separating neuronal cell bodies from non-neuronal cells within the manually defined region shown in (d). f Single βIII-tubulin channel image showing the identification of βIII-tubulin positive regions (grey) separating neuronal cell bodies from non-neuronal cells as in (e) within the manually defined region shown in (d). Mean fluorescent signal for a given section were calculated using the measurements of cy2 or cy3 signal within each βIII-tubulin positive volumetric pixel as denoted by grey ROIs in (e) and (f). Scale bar 50 μm. (GIF 109 kb).

High Resolution Image (TIFF 4414 kb).

Supplemental figure 3

Normalization procedure enabling comparison between image sets. a Each image set contained a section from each of the treatment groups stained with the same antibody dilution and imaged using identical acquisition parameters. The mean fluorescence for each section, termed the “treatment mean”, (a.1-a.4) by averaging cy2 (green) fluorescent signal for βIII-tubulin positive volumetric regions (grey in images a.5-a.8). b Fluorescent signal readings in βIII-tubulin positive volumetric regions (green circles) are averaged resulting in the “treatment mean”. In order to compare across image sets of sections means were normalized within the respective image set prior to cross-set comparison, resulting in normalized treatment means. Means are normalized by dividing the treatment mean by the sum of means within the image set. The normalized treatment means were used to compose the graphs in the main text. (GIF 165 kb).

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Benbow, S.J., Wozniak, K.M., Kulesh, B. et al. Microtubule-Targeting Agents Eribulin and Paclitaxel Differentially Affect Neuronal Cell Bodies in Chemotherapy-Induced Peripheral Neuropathy. Neurotox Res 32, 151–162 (2017). https://doi.org/10.1007/s12640-017-9729-6

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