Development of an EFEMP1-directed Antisense Oligonucleotide for the treatment of Autosomal Dominant Drusen

Abstract

Purpose : Autosomal Dominant Drusen (ADD) is a dominantly inherited maculopathy characterized by the generation of massive deposits underneath the retinal pigment epithelium (RPE) and is associated with an irreversible and progressive loss of central visual acuity due to geographic atrophy and choroidal neovascularization. There are no approved treatments for ADD. ADD is caused by a single non-conservative missense mutation of the epidermal growth factor-containing fibrillin-like extracellular matrix protein 1 (EFEMP1) gene. Herein we describe the development and characterization of an Antisense Oligonucleotide (ASO) directed at EFEMP1 knockdown to treat the underlying cause of disease.

Methods : We have generated a novel CRISPR-edited ARPE-19 EFEMP1^R345W lines and iPSC-RPE lentiviral EFEMP1 overexpression models to test and screen antisense oligonucleotides compared to siRNA ‘pools’ as positive controls. We used qPCR, Western blotting, and immunohistochemistry to assess EFEMP1 knockdown and complement activity. Using C57B6J mice we carried out intravitreal injections of EFEMP1 targeted ASOs across a dose range and assessed knockdown efficiency by qPCR and western blotting

Results : We report that ARPE19 EFEMP1^R345W cells display over twice the expression of EFEMP1 that observed in the parental ARPE19 line after 4 weeks of differentiation. We show a significant (P<0.05) dose dependent knockdown of EFEMP1 using ASO in ARPE-19 and ARPE-19 EFEMP1^R345W cells. We report significant overexpression of EFEMP1 and C3 in IPSC-RPE transduced with CMV-EFEMP1 wild-type and mutant EFEMP1^R345W. EFEMP1-directed ASOs can significantly reduce EFEMP1 expression in iPSC-RPE overexpression models. Intravitreal injection of EFEMP1 targeting ASOs in mice leads to significant reduction (over 80%) in EFEMP1 expression in a dose dependent manner.

Conclusions : These data show preclinical efficiency of an EFEMP1 targeting ASO that shows effective knockdown of EFEMP1 in iPSC-RPE EFEMP1 R345W overexpression models and in vivo via intravitreal injection. These data indicate that ASOs targeting EFEMP1 could have potential as a treatment for ADD, a disease associated with excessive EFEMP1 accumulation.

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