Abstract
Purpose :
Vision loss due to geographic atrophy (GA) in advanced macular degeneration (AMD) patients is associated with loss of photoreceptors. Gene therapy has proven to be a promising approach for treating retinal degenerative diseases by transduction of light sensitive proteins (e.g. Multi-Characteristic Opsin, MCO) in retinal cells. Viral vector-based gene delivery method is often confronted with limitations due to packaging size limit, immunogenetic reaction, unintended non-targeted delivery, non-feasibility of re-dosing due to immunorejection, and complex manufacturing process. Here, we report Optical Coherence Tomography (OCT) guided non-viral laser approach that involves detection of GA areas, followed by spatially targeted delivery of therapeutic MCO-genes into targeted higher order neurons in GA.
Methods :
For targeted delivery of MCO, a near-infrared (NIR) laser beam was coupled with the source beam of the OCT. Mixture of MCO plasmids and gold nanorods (GNRs) was intravitreally injected in retinal degenerated (rd1), wild type mice and African green monkeys. The efficient localized field enhancement (hot spots), by surface plasmon resonance (SPR) of GNRs in the safe NIR spectrum, allows site-specific delivery of MCO-plasmids. Evaluation of safety and toxicity associated with the laser-transfection of MCO was examined by electrophysiology, OCT, intra-ocular pressure (IOP) and immunohistochemistry.
Results :
The NIR laser delivery of MCO plasmids resulted in (i) cell and layer-specific expression by use of functionalized GNRs and cell-specific promoter, (ii) spatially-targeted expression of MCO in retina by use of directed laser, allowing visually evoked activities, (iii) no noticeable cell death, (iv) no increase in IOP or ocular damage (by OCT), (v) clearing of fGNRs from retina after 1 wk, (vi) non-detectable immunotoxicity; (vii) robust delivery through internal limiting membrane (ILM) in NHP; (viii) activation of MCO-sensitized retina at ambient light; (ix) expression lasting at least 24 wks; (x) no damage to existing photoreceptors (measured by ERG); and (xi) enhanced electrophysiological signal from degenerated retina.
Conclusions :
The non-viral laser delivery approach provides efficient, targeted delivery of therapeutic MCO-molecules into spatially targeted retinal cells in GA areas without eliciting inflammatory or immunological reactions.
This abstract was presented at the 2023 ARVO Annual Meeting, held in New Orleans, LA, April 23-27, 2023.