Abstract
Purpose :
Primary open-angle glaucoma (POAG) is associated with changes in extracellular matrix (ECM) in the trabecular meshwork (TM) and other structural changes in the TM that cause increased resistance to aqueous humor outflow and result in increased intraocular pressure (IOP). Currently, pharmacological intervention for glaucoma therapy is limited to intraocular pressure (IOP) lowering. Durysta (bimatoprost (BIM) implant) was launched in 2020 and became the first intracameral, biodegradable sustained-release implant indicated for POAG. Clinical data demonstrate that a large proportion of Durysta treated patients experience up to 2-years of IOP benefit; the extended duration of IOP lowering is likely related to tissue remodeling in the TM. This striking response led us to further investigate the mechanism of implant levels of BIM on glaucomatous TM.
Methods :
We treated glaucomatous human primary TM cells (n=3) with different concentrations of BIM: 10µM, 100µM, and 1000µM for 24h and performed mRNA-seq. We determined differentially expressed genes for all conditions compared to control and analyzed them using Ingenuity Pathway Analysis (IPA) software. Western blot (WB) and immunofluorescence (IF) complemented the selected candidate genes for the follow-up experiments.
Results :
Autophagy activation (p=6.1e-8) was the most significant pathway altered in the 1000µM condition compared to control or 10µM level of BIM. 46 out of 216 genes involved in the autophagy pathway were mostly upregulated (5% FDR), including MAP1LC3B, AMBRA1, and SQSTM1. These results led us to hypothesize that ECM degradation via autophagy may alter flow dimensions in TM for aqueous humor outflow. In order to understand the super response phenomenon of Durysta, a follow-up experiment was performed where glaucomatous TM cells (n=3) were treated with 1000µM BIM for 24h followed by fresh media chase for 48h. Under these conditions, MAP1LC3B, AMBRA1, and SQSTM1 gene expression decreased back to their normal levels, however, the protein level of MAP1LC3B and SQSTM1 remained significantly elevated as assessed by WB. Finally, IF of glaucomatous TM cells (n=3) treated with 1000µM BIM for 24h followed by 48h fresh media chase showed a decrease of FN1 in their ECM.
Conclusions :
These results suggest that autophagy in the TM may be a main player in the extended duration of IOP mitigation by implant levels of BIM.
This abstract was presented at the 2023 ARVO Annual Meeting, held in New Orleans, LA, April 23-27, 2023.