Both NXNL1 and NXNL2 are important for photoreceptor survival in the mouse

Abstract

Purpose : Retinitis pigmentosa (RP) is characterized by progressive rod then cone degeneration, leading to blindness in severe cases with no therapeutic option. The nucleoredoxin-like 1 (NXNL1) gene codes for two alternative splice isoforms: a short form, rod-derived cone viability factor (RdCVF), promoting cone aerobic glycolysis and a long form (RdCVFL) modulating oxidative damage. A phase I/II trial (NCT05748873) of gene therapy with both isoforms in RP is ongoing. NXNL2, a paralogue of NXNL1, also encodes two isoforms with cone-protective function. We investigated whether both NXNL1 and NXNL2 products are required for photoreceptor survival in mouse retina. We also studied the benefit of combined both short forms in photoreceptor protection by using the rd10 mouse model of RP.

Methods : Double knockout Nxnl1^-/-Nxnl2^-/- mice were generated. Their retinal phenotype was investigated using electroretinogram (ERG) recording and optical coherence tomography (OCT) at different ages, compared to Nxnl1^-/- or Nxnl2^-/- and wild-type (wt) mice.
The effect of combined or separate overexpression of RdCVF and RdCVF2 was tested using AAV2/8 subretinal delivery in rd10 mice at PN14 compared with rd10 injected with AAV2/8-GFP. Visual function was evaluated by ERG recording and a quantitative optomotor response (qOMR) system 1 month post injection.

Results : Nxnl1^-/-Nxnl2^-/- mice showed severer rod dysfunction at age 3, 6, and 12 months compared to Nxnl1^-/-, Nxnl2^-/-, and wt mice. The overall average reduction of scotopic ERG response is Nxnl1^-/-Nxnl2^-/- 29.0%(n≥7), Nxnl1^-/- 16.3% (n≥8), Nxnl2^-/- 10.7% (n≥5), compared to wt (n=11) mice. In addition, the outer nuclear layer (ONL) thickness was reduced on OCT at all ages, the average reduction is Nxnl1^-/-Nxnl2^-/- 15.8% (n≥9), Nxnl1^-/- 13.3% (n≥9), Nxnl2^-/- 7.1%(n≥5), compared to wt (n≥11) mice.
Rd10 mice injected with a combination of AAV-RdCVF and AAV-RdCVF2 (n=3) showed increased visual acuity on the qOMR compared to controls injected with AAV-RdCVF (n=2), AAV-RdCVF2 (n=3), and AAV-GFP (n=3). Preliminary data revealed larger photopic ERG responses in rd10 mice injected with the combination compared to the other 3 groups.

Conclusions : Both Nxnl1 and Nxnl2 are essential for photoreceptor survival in mice. The combination of RdCVF and RdCVF2 promoted vision rescue in rd10 mice, showing clinical potential for RP. Further investigation will decipher the underlying mechanism of RdCVF2 in cone rescue.

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