Abstract
Purpose :
The cornea is continuously vulnerable to external insults, and severe injuries can lead to exacerbated inflammation and scar tissue formation causing corneal opacity and impaired vision. Currently, there are limited options available in the clinic for preventing corneal inflammation, scarring and promoting regeneration. Decorin is a proteoglycan that strongly binds and inactivates TGF-b1. Challenges in decorin production, primarily due to inherent protein instability, have hindered its pharmaceutical exploitation. Herein, we have pioneered the development of a range of novel decorin mimetics that promote corneal regeneration, minimize scarring, and dampen inflammation.
Methods :
We have developed a general deep-learning framework for protein design that enabled the redesign of the unstable extremities of decorin. This study developed several decorin mimetics with different characteristics using a two-stage binder design approach. These were validated based on selecting favorable interacting amino acids against TGF-β1. The decorin mimetics were synthesized and binding affinity to decorin was measured using Microscale thermophoresis. Thereafter, the therapeutic effects of the novel decorin mimetics were validated in vitro and in vivo. The decorin mimetics were able to inhibit fibroblast transdifferentiation in vitro and promote scarless corneal wound healing in vivo using an alkali burn mouse model.
Results :
Novel decorin mimetics were designed targeting TGF-β1 based on the decorin:TGF-β1 binding complex to increase the affinity of decorin to TGF-β1. Thereafter, the C- and N-terminal regions of decorin were modified without affecting the decorin:TGF-β1 binding interface, as evidenced by circular dichroism. The stable decorin mimetics produced allow for scaling-up production, and presented significantly higher affinity to TGF-β1 compared to decorin, with the Kd of decorin in the low micromolar range while the Kd of decorin mimetics reached the nanomolar range (500nM). The decorin mimetics inhibited TGF-β1 induced keratocyte transdifferentiation in vitro and promoted corneal regeneration following alkali burn, with decreased inflammation and scarring.
Conclusions :
We have successfully designed a set of hyperstable soluble decorin mimetics that significantly promote scarless corneal wound healing in vivo. Our established platform has the potential to identify and generate novel drug targets for treating a plethora of corneal diseases.
This abstract was presented at the 2024 ARVO Annual Meeting, held in Seattle, WA, May 5-9, 2024.