Efficacy and safety of suberoylanilide hydroxamic acid (Vorinostat) in the treatment of canine corneal fibrosis.

Abstract

OBJECTIVE: Study aims were to evaluate the safety and efficacy of the Food and Drug Administration-approved drug Vorinostat [suberoylanilide hydroxamic acid (SAHA)] in the treatment of canine corneal fibrosis using an in vitro model. METHODS: Healthy donor canine corneas were collected and used to generate primary canine corneal fibroblasts (CCFs) by growing cultures in minimal essential medium supplemented with 10% fetal bovine serum. Canine corneal myofibroblasts, used as a model for corneal fibrosis, were produced by growing CCF cultures in serum-free medium containing transforming growth factor &#x3b2;1 (1 ng/mL). Trypan blue exclusion assays were used to determine the optimal SAHA dose for this in vitro model. Four hour after culturing with TGF&#x3b2;1, CCF cultures were treated with 0.06% SAHA for 5 min (group 1) and for 24 h (group 2), representing single and multiple dose treatment regimes, respectively. Cultures were then further incubated in the presence of TGF&#x3b2;1 (1 ng/&#x3bc;L) under serum-free conditions until they reached 70% confluence. Trypan blue exclusion, immunocytochemistry, and TUNEL assays were used to evaluate the cytotoxicity of SAHA. Real-time PCR, western blot analysis, and immunocytochemistry were used to determine the efficacy of SAHA to inhibit canine corneal myofibroblast formation. RESULTS: Topical SAHA application in both treatment groups successfully decreased &#x3b1;-smooth muscle actin expression when compared to the TGF&#x3b2;1 only treatment group (P < 0.05). Tested SAHA did not affect CCF phenotype or cellular viability and did not cause significant cell death. CONCLUSIONS: Suberoylanilide hydroxamic acid safely and effectively inhibits TGF&#x3b2;1-induced CCFs transformation to myofibroblast in vitro.