RUNX1-Trap regulates pathologic ocular angiogenesis: proof-of-concept of an mRNA-encoded candidate therapeutic

Abstract

Purpose : The discovery of the transcription factor RUNX1 as a mediator of ocular aberrant angiogenesis has facilitated the development of novel experimental therapeutics. We developed an mRNA-encoded dominant negative inhibitor of RUNX1 function, RUNX1-Trap, based on a fusion protein of CBFb. It binds to RUNX1 and inhibits its transcriptional activity. We report results from a proof-of-concept study evaluating the in vitro and in vivo anti-angiogenic potential of RUNX1-Trap

Methods : RUNX1 overexpression was induced in HRECs by treat TNFα. Proliferation and migration were assessed and RUNX1 and RUNX1-Trap expression were measured by immunofluorescence, in induced cells. RUNX1-Trap (25 ng/μL) was administrated intravitreally in a laser-induced choroidal neovascularization (LCNV) mouse model. Leakage, lesion severity and size were measured 7 days post-injection. The eyes were collected and cryo-sectioned 24- and 48-hours post-injection. They were stained for RUNX1-Trap and images were segmented to quantify the biodistribution RUNX1-Trap. Markers for retinal pigmental epithelium, astrocytes, microglia, and blood vessels were used to co-localize RUNX1-Trap expression

Results : RUNX1-Trap was expressed in HRECs and inhibited RUNX1 expression and function. Proliferation and migration of endothelial cells were also significantly reduced. RUNX1-Trap significant reduced leakage (75%, P < 0.05) and lesion size (80%, P < 0.05) in the LCNV mice (Fig. 1A and B). Biodistribution analysis showed a significant increase of RUNX1-Trap expression at 48 hours within the inner plexiform layer, inner nuclear layer, outer plexiform layer, and outer nuclear layer. RUNX1-Trap co-localized with retinal blood vessels (Fig. 1C), astrocytes, and microglia (Fig. 1D)

Conclusions : We showed that RUNX1-Trap inhibited RUNX1 expression and function and in vitro proliferation and migration. RUNX1-Trap effectively prevented neovascularization in an LCNV mouse model. RUNX1-Trap was expressed in specific layers of the retina, and co-localized with blood vessels, astrocytes, and microglia. Expression of the mRNA-encoded RUNX1-Trap was observed in key structures and cells implicated in the development of aberrant ocular angiogenesis

This abstract was presented at the 2024 ARVO Annual Meeting, held in Seattle, WA, May 5-9, 2024.

 

Fig. 1. A, Images of leakage (top) and lesion size (bottom) in the LCNV mouse model treated. B, Leakage quantification, RUNX1-Trap (red) co-localized with C, blood vessels (green) and D, microglia (green)