Inhibition of H3K9 methyltransferases G9a/GLP prevents ototoxicity and ongoing hair cell death

Abstract

Sensorineural hearing loss (SNHL) is one of the most common sensory defects in humans. Hair cells are vulnerable to various ototoxic insults. Effective prevention of hair cell loss remains an unmet medical need. Apoptotic hair cell death, which involves active regulation of transcription, accounts for the majority of aminoglycoside-induced hair cells loss. As one of the important epigenetic covalent modifications, histone methylation is involved in the regulation of gene expression, development and reaction to injury. In particular, H3K9 dimethylation (H3K9me2) is critical for euchromatin gene silencing. In the present study, we examined the roles of two highly homologous histone methyltransfereases responsible for this modification, G9a/G9a-like protein (GLP), in the reaction to aminoglycoside-induced hair cell damage. We observed a rapid increase of H3K9me2 upon hair cell damage in organotypic cochlear cultures. Treatment with the G9a/GLP-specific inhibitors, BIX01294 or UNC0638, reduced the level of H3K9me2 and prevented hair cells from death. Local delivery of BIX01294 also prevented neomycin-induced in vivo auditory hair cell loss in the organ of Corti in a mouse damage model. It is unlikely that BIX01294 functions through blocking aminoglycoside absorption as it does not interfere with aminoglycoside uptaking by hair cells in the organotypic cochlear cultures. Our data revealed a novel role of histone methylation in otoprotection, which is of potential therapeutic value for SNHL management.

Inhibition of H3K4me2 Demethylation Protects Auditory Hair Cells from Neomycin-Induced Apoptosis

“…2). Neomycin exposure caused a significant reduction in the number of hair cells in the middle and basal turns of the untreated cochleae in accordance with previous findings for neomycin damage [11,20,21]. In contrast, pretreatment with 20 μM S2101 significantly prevented neomycin-induced hair cell death in middle and basal segments compared to the neomycin-only controls (P<0.001 for hair cells in middle and basal sections; Student's t test) (Supplemental Fig.…”

“…The detailed methods for the ABR test were given previously [11]. Briefly, ABR was performed in anesthetized (100 mg/kg ketamine and 25 mg/kg xylazine sodium, i.p.)…”

“…Two days after administration of the drug, neomycin was injected subcutaneously once per day for five consecutive days. The detailed protocol for neomycin administration was given previously [11]. Two weeks after exposure to neomycin, the hearing in the mice was evaluated by measuring the ABR thresholds.…”

“…Recent progress has highlighted the specific functional roles of histone methylation in a variety of cellular processes such as cell differentiation, survival, and death [8][9][10], and we have previously shown that the G9a inhibitor BIX01294 protects against neomycin-induced hair cell loss by inhibiting dimethylation of H3K9 [11].…”

Probing the epigenome

“…UNC0638, a second-generation, less-toxic chemical probe for G9a and GLP, was also found to alter patterning of dimethylated histone H3 Lys9 and chromatin structure at CpG islands in hematopoietic progenitor cells 25 . In another example, the G9a inhibitors BIX-01204 and UNC0638 protected hair cells from neomycin-induced damage in vitro, and from aminoglycoside-induced damage in vivo, thus suggesting a potential avenue for the treatment of sensorineural hearing loss 26 . Finally, prolonged exposure of lymphoma cells to a chemical inhibitor of EZH2 revealed resistance mechanisms that should be addressed by next-generation inhibitors 27 .…”