Evaluation of a new ROCK inhibitor in a rat model of diabetic retinopathy

Abstract

Purpose : Rho-associated kinase (ROCK) activation contributes to micro-vascular closure, retinal hypoxia and disrupted retinal pigment epithelium (RPE) barrier in Diabetic Retinopathy (DR) rat model. ROCK inhibition seems to be a promising therapeutic target as previous results with Fasudil, a clinically approved ROCK inhibitor, showed a better retinal perfusion and reduced edema. However, its short life-time in the vitreous is not compatible with a long-term treatment. In this study, we evaluate the potential of a new treatment, BIRKI, a slow-release ROCK Inhibitor from Boehringer Ingelheim.

Methods : 10 to 16 month-old male Goto-Kakizaki (GK) type 2 diabetic rats were injected intravitreously with 3µl of either BIRKI or Vehicle. The diabetic status was defined by measurement of the plasma concentration of glycosylated hemoglobin (HbA1c). Eyes were enucleated and dissected 8 or 28 days after the injection. Immunohistochemistry was performed on flat mounts and cryosections to evaluate retinal hypoxia and vasodilatation), RPE morphology and barrier disruption 28 days after the treatment. Western Blots were performed to assess ROCK activity and expression 8 days after the treatment (significance of results was evaluated using non parametric tests).

Results : ROCK activity, quantified through the specific phosphorylation of one of its substrates, MYPT1, was significantly reduced in RPE/choroid complex of BIRKI injected rats compared to Vehicle rats (p=0,028). Moreover, a pimonidazole staining showed a significant reduction of retinal hypoxia in BIRKI rats compared to Vehicle (p=0.004). This result is consistent with the dilation of retinal vessel observed in BIRKI rats (p=0.017). The RPE morphology seems to be partially restored as well as its function as barrier, through quantification of cell size, cell shape and leakage. No signs of toxicity of BIRKI have been observed so far.

Conclusions : Our results are consistent with ROCK inhibition as an interesting new therapeutic concept for diabetic retinopathy. A single injection of BIRKI showed similar effects at 28 days than 3 consecutive injections of Fasudil at 48h. Further studies are needed to confirm that BIRKI would be a good candidate in the treatment of diabetic retinopathy, especially in regard of improvement of retinal vascular infusion and protection of the outer retinal barrier.

This is a 2021 ARVO Annual Meeting abstract.