Peptide Therapy for Neuroprotection in Glaucoma

Abstract

Purpose : This study assesses the neuroprotective potential of P1-CPP, a mini-chaperone peptain-1 (P1) with cell-penetrating peptide, in in vitro, in vivo, and ex vivo glaucoma models.

Methods : Primary retinal ganglion cells (RGCs) were subjected to neurotrophic factor (NF) deprivation for 48 h or 100 nM ET-3 treatment, for 24 h receiving either 12.5 µg/ml P1-CPP or vehicle. Survival was analyzed using Live/Dead assay. Human retinal explants (n=5 postmortem donors) were treated with 12.5 µg/ml P1-CPP or vehicle for 7 days, stained with RBPMS an RGC markers, and RGC survival was analyzed.
For the in vivo study, Brown Norway (BN) rats with elevated IOP received intravitreally P1-CPP (2 µg/eye) or vehicle weekly for six weeks. RGC function was evaluated using PERG. RGC and axon counts were assessed. In another set of experiments, RGCs from hypertensive rats treated with P1-CPP or vehicle for 7 days were isolated for transcriptomic analysis. RGCs subjected to NF deprivation for 48 h were used for qPCR targets confirmation.

Results : NF deprivation led to an 83% RGC loss (p<0.0001), which was reduced to 64% with P1-CPP (p<0.0001) treatment. P1-CPP also decreased ET-3-induced RGC death by 19% (p<0.01).
In ex vivo human retinal explants, P1-CPP decreased RGC loss by 35% (p<0.0004).
IOP elevation resulted in 37% and 30% (p<0.001) of RGC loss in comparison with naïve animals in mid-peripheral and peripheral retinas compared to naïve rats. P1-CPP significantly inhibited RGC loss by 18% (p<0.03) and 36% (p<0.001) in mid-peripheral and peripheral retinas, respectively. PERG amplitude decline (41%, 4.55 µV, p<0.001) in IOP-elevated rats was mitigated by P1-CPP treatment (7.55 µV, p<0.03).
Transcriptomic analysis showed over six thousand differentially expressed genes. Upregulated pathways included CREB signaling and synaptogenesis. A 1.6-fold increase (p<0.01) in Creb1 mRNA and elevated phosphorylated Creb (125%, p=0.01) were observed in P1-CPP-treated cells compared to vehicle-treated RGCs. qPCR confirmed the RNA-seq findings of the increased expression of Creb1 in P1-CPP-treated RGCs compared to that of the vehicle-treated group.

Conclusions : P1-CPP exhibits neuroprotective effects in glaucoma models, potentially through CREB signaling enhancement, indicating its therapeutic promise in treatment.

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