Abstract
Purpose :
PKa has been previously reported as a VEGF-independent mediator of diabetic macular edema. Here we investigate the effects of 2 novel and structurally diverse oral PKa inhibitors, KV998052 and KV998054 on VEGF-induced retinal thickening in mice.
Methods :
PKa enzyme activity: Effects of KV998052 and KV998054 were assayed using H-D-Pro-Phe-Arg-AFC. Inhibitor selectivity was assessed using a panel of closely-related serine proteases. Effects on high molecular weight kininogen (HK) cleavage was measured in whole plasma stimulated with dextran sulfate (DXS) using a capillary-based immunoassay.
Pharmacodynamics: Retinal thickening was measured in a mouse model of VEGF induced retinal edema with KV998052/KV998054/vehicle administered s.c. at doses up to 2μg/kg/day or orally 50mg/kg bid. Plasma concentrations were determined by LC-MS/MS. Retinal thickness was measured with optical coherence tomography, at baseline and time points post intravitreal injections of VEGF (100ng/eye) or PBS vehicle in both prevention and intervention studies using Bioptigen Diver 3.3.7.
Results :
The Ki of KV998054 and KV998052 for purified human PKa are 4nM and 3nM, respectively. These inhibitors displayed >500 fold selectively for PKa compared with closely related serine proteases. Both blocked DXS-stimulated HK cleavage in a dose dependent manner. Infusion of KV998054 at 2, 1 and 0.5μg/kg/day s.c. resulted in plasma exposure of 32.4, 24.6 and 8.3ng/ml, respectively (n=6) in mice. KV998054 reduced VEGF-stimulated edema at 24h by 40% at 1μg/kg/day, p=0.0037 and 54% at 2μg/kg/day, p=0.0001 (n=10 per dose). Oral pharmacokinetics of KV998054 in mice revealed plasma Cmax of 837ng/ml at 0.5h and exposure of 28.5ng/ml at 10h post-gavage. Oral administration of KV998054 initiated before VEGF injection, resulted in 59% (p=0.008) decrease in VEGF retinal thickening at 24h. KV998052 administered s.c. at 2μg/kg/day decreased VEGF-induced edema by 37% (n=12, p=0.0018). Oral KV998052 provided 24h after intravitreal injection of VEGF accelerated resolution of edema at 72h by 83% (p=0.015) compared with controls orally given vehicle. Retinal segment analysis revealed that reduction in thickening by PKa inhibition occurred in multiple layers, including an 87% decrease (p =0.004) in the inner nuclear layer.
Conclusions :
Oral PKa inhibitors prevent and reverse VEGF-induced retinal edema in mice.
This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.