Bexarotene Normalizes Chemotherapy-Induced Myelin Decompaction And
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Chiang et al. acta neuropathol commun (2020) 8:193 https://doi.org/10.1186/s40478-020-01061-x RESEARCH Open Access Bexarotene normalizes chemotherapy-induced myelin decompaction and reverses cognitive and sensorimotor defcits in mice Angie C. A. Chiang1, Alexandre V. Seua1, Pooja Singhmar1, Luis D. Arroyo1, Rajasekaran Mahalingam1, Jian Hu2,3, Annemieke Kavelaars1 and Cobi J. Heijnen1* Abstract Frequently reported neurotoxic sequelae of cancer treatment include cognitive defcits and sensorimotor abnormali- ties that have long-lasting negative efects on the quality of life of an increasing number of cancer survivors. The underlying mechanisms are not fully understood and there is no efective treatment. We show here that cisplatin treatment of mice not only caused cognitive dysfunction but also impaired sensorimotor function. These functional defcits are associated with reduced myelin density and complexity in the cingulate and sensorimotor cortex. At the ultrastructural level, myelin abnormalities were characterized by decompaction. We used this model to examine the efect of bexarotene, an agonist of the RXR-family of nuclear receptors. Administration of only fve daily doses of bex- arotene after completion of cisplatin treatment was sufcient to normalize myelin density and fber coherency and to restore myelin compaction in cingulate and sensorimotor cortex. Functionally, bexarotene normalized performance of cisplatin-treated mice in tests for cognitive and sensorimotor function. RNAseq analysis identifed the TR/RXR pathway as one of the top canonical pathways activated by administration of bexarotene to cisplatin-treated mice. Bexarotene also activated neuregulin and netrin pathways that are implicated in myelin formation/maintenance, synaptic func- tion and axonal guidance. In conclusion, short term treatment with bexarotene is sufcient to reverse the adverse efects of cisplatin on white matter structure, cognitive function, and sensorimotor performance. These encouraging fndings warrant further studies into potential clinical translation and the underlying mechanisms of bexarotene for chemobrain. side efects of treatment, including cognitive impairment and sensory and motor abnormalities [28, 40, 41, 50, 51, Introduction 53, 54, 60]. Tere are no FDA-approved drugs to prevent During the last decade cancer treatment has become or reverse these neurotoxicities. Terefore, development more and more successful, but unfortunately a large num- of novel therapeutic strategies is urgently needed. ber of cancer survivors reports long lasting neurotoxic We have recently shown that treatment of mice with cisplatin induces a profound and long lasting impairment *Correspondence: [email protected] in performance in tasks of spatial memory and execu- 1 Division of Internal Medicine, Department of Symptom Research, tive functioning [5, 7, 8, 35, 65]. At the structural level, University of Texas M.D. Anderson Cancer Center, 6355 MD Anderson these behavioral defcits are accompanied by a decrease Blvd, Unit 1055, Houston, TX 77030, USA Full list of author information is available at the end of the article in dendritic spine density in the cingulate cortex, and a © The Author(s) 2020. 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The Creative Commons Public Domain Dedication waiver (http://creat iveco mmons .org/publi cdoma in/zero/1.0/) applies to the data made available in this article, unless otherwise stated in a credit line to the data. Chiang et al. acta neuropathol commun (2020) 8:193 Page 2 of 15 reduction in the expression of markers of synaptic integ- bexarotene reverses these white matter alterations. As a rity like PSD95 and synaptophysin in prefrontal cortex functional readout we analyzed not only cognitive func- and hippocampus [5, 7, 35, 65]. In addition, we observed tion but also sensorimotor function in cisplatin-treated an increase in the coherency of fbers after staining for mice. Potential white matter damage including changes myelin basic protein in the cingulate cortex, indicating a in myelin structure were investigated in the sensorimo- reduction in arborization and complexity of myelinated tor cortex as well in view of the abnormalities in sen- axons [7]. sorimotor function reported by patients treated with Restoration of myelin damage as a result of cerebral chemotherapy. Bexarotene is already FDA-approved as insults or neurodegenerative processes is key to restora- a treatment for cutaneous T cell lymphoma and is not tion of brain function. In the brain, myelin is produced likely to interfere with the efcacy of cancer treatment by oligodendrocytes, while astrocytes, T cells and mac- [16, 17]. Terefore, bexarotene could represent a promis- rophages/microglia can all modulate myelin formation ing new and safe therapeutic strategy to reverse the nega- [15, 29, 45]. Retinoid X receptor (RXR) is a member of tive consequences of cancer treatment for brain health the NR2B nuclear receptor family. As a common bind- without negative interference with cancer treatment. ing partner of many other nuclear receptors, it mainly functions as a ligand-dependent transcription factor and Materials and methods regulates many physiological processes. Activation of the Mice RXR family of receptors can promote (re)myelination Male and female C57BL/6 J mice (Jackson Laboratory) either via their anti-infammatory efect, their efects on were housed at 22 ± 2 °C on a 12/12 h reverse dark–light monocyte/macrophage phagocytosis of myelin to remove cycle with water and food ad libitum. All experiments myelin debris, and for their capacity to directly stimulate were conducted at Te University of Texas MD Anderson oligodendrocyte precursor proliferation/diferentiation Cancer Center and approved by the Institutional Animal [9, 12, 39]. RXR activation, either via genetic manipula- Care and Use Committee of Te University of Texas MD tion or pharmacologic interventions, increased oligoden- Anderson Cancer Center in Houston, TX. Mice were drocyte diferentiation and remyelination in models of randomly assigned to treatment groups and investigators toxin-induced demyelination in rats [25]. Moreover, tran- were blinded to treatment. scripts encoding RXRγ were upregulated during remy- elination and expressed by cells of the oligodendrocyte Chemotherapy and bexarotene treatment lineage [25]. At 9 weeks of age, mice received cisplatin (Fresenius Kabi Bexarotene is a synthetic retinoid modulator of RXRs USA) or phosphate-bufered saline (PBS) administered that binds the RXR receptor subtypes RXRα, RXRβ, and intraperitoneally (i.p.) in 2 rounds consisting of 5 daily RXRγ with high afnity. Te drug has been explored as doses of 2.3 mg/kg, followed by 5 days of rest without an add on for cancer therapies. Although some studies injections. Te cumulative dose of 23 mg/kg is equivalent indicated stabilization of advanced small cell lung carci- to 70 mg/m2 in humans [37], which is within the range noma in patients receiving add on bexarotene, there was of one cycle of cisplatin treatment in humans [55]. We no detectable increase in survival. Importantly, there is showed previously that this treatment regimen has anti- no evidence that bexarotene negatively interferes with tumor efects in the mouse and induces cognitive defcits anti-cancer efects of chemotherapy [16, 36, 43]. RXR are [7, 8, 35]. expressed in all cell types in the brain, including neurons, Bexarotene in 10% DMSO in sunfower seed oil was oligodendrocytes, astrocytes and microglia (http://dropv delivered at a fnal dose of 100 mg/kg/day for 5 consecu- iz.org/). In vitro, RXR stimulation by the agonist bex- tive days by oral gavage starting 24 h after the last dose of arotene restored the age-related defciencies in myelin cisplatin. debris phagocytosis by macrophages, a key process in myelin maintenance [38]. A more recent study showed Tissue processing and Black Gold II staining that bexarotene also promotes myelin formation in a Mice were sacrifced after behavioral testing using brief genetic model of myelin loss [66]. Moreover, RXR ago- CO2 exposure, followed by intracardial perfusion with nists like bexarotene can reduce the cognitive defcits and ice-cold PBS. Brains were removed and post-fxed in brain damage that develop in rodent models of cerebral 4% PFA for 48 h, cryoprotected in sucrose, and cut at ischemia, subarachnoid hemorrhage, and traumatic brain 25 μm in the coronal plane on a sliding microtome. For injury [9, 58, 63, 67]. each animal, 4 sections were used for Black Gold II (Mil- Te aim of this study was to better characterize the lipore, #AG105) staining according to manufacturer’s white matter damage that develops in mice treated with instructions. Briefy, sections were mounted onto slides cisplatin and to determine whether the RXR agonist and dried overnight at room temperature. Te next day, Chiang et al. acta neuropathol commun (2020) 8:193 Page 3 of 15 slides were rehydrated in ddH2O before immersion in of ice cold MtBE (Methyl tert-butyl ethe)/Methanol/ Black Gold II solution at 60 °C for 15 min. After wash- Water. Samples were centrifuged at 17,000 g for 5 min ing in ddH2O, slides were incubated in pre-warmed 1% at 4 °C, and the organic top layer was transferred to a sodium thiosulfate solution at 60 °C for 3 min. Slides clean tube, followed by evaporation to dryness under were then rinsed with ddH2O and dehydrated through a nitrogen.