Vitreoretinopathy: An unmet clinical need
Since its recognition in 1983 by the Retina Society Terminology Committee, Proliferative Vitreoretinopathy (PVR), has been one of the unsolved challenges in Ophthalmology. PVR is a complication after retinal detachment that is characterised by proliferation of cells in the vitreous or peri-foveal retina, resulting in membrane formation and eventually tractional re-detachment. It is devastating and accounts for 75% of surgical failures of retinal detachment repair. Despite decades of research, there are still no effective therapies for PVR. Surgical removal of membranes to relieve traction for prevention of re-detachment remains the standard treatment, albeit with guarded prognosis. This highlights a crucial need for effective therapeutics against PVR.
Unexpected observations of our biodegradable polymer
The primary prevention of PVR stems back to the success of a retinal detachment repair surgery. During retinal detachment repair, pars plana vitrectomy (whereby vitreous is removed from the eye) is performed to relieve vitreous traction and provide access to the retinal for manipulation and repair. As the vitreous humour is thought to be unable to regenerate de novo, the cavity is filled with an endotamponade agent to keep the retina attached after completion of repair. While expansile gases (such as sulphur hexafluoride) and silicone oil are usually used in clinics, multiple disadvantages such as the need for post-operation posturing, inability to proceed with air travel due to expansile nature of gas and need for additional surgery for oil removal are associated with these agents.
To overcome these inherent drawbacks with conventional substitutes, we previously adopted a rational stepwise approach in the development of a tri-component multi-block thermogelling polymer, poly(CEP) which fulfilled the clinical requirements of a vitreous substitute for retinal detachment repair and facilitated the reformation of a vitreous-like body in situ as it biodegraded. Interestingly, we observed that retinal detachment rabbit models injected with poly(CEP) did not develop retinal scarring when compared to those with saline injections. This suggested the possibility of the injected polymer being a bioactive substance in the eye. We hypothesised that the polymer not only worked in re-apposing the detached retina, but also actively suppressed PVR development post-surgery. Hence, we embarked to determine the effects of the polymer on PVR pathogenesis at a phenotypical and molecular level.
Proof of concept in rabbit PVR models
We established a disease model with severe retinal scarring phenotype using the large-eyed, New Zealand white rabbit which closely resembles PVR in the human eye. This was achieved by injecting rabbit blood and retinal pigment epithelial (RPE) cells into the vitreous cavity, to form contractile fibro-cellular membranes - a key hallmark of PVR. These eyes were then treated with either an injection of air, sulphur hexafluoride or our poly(CEP) polymer as an endotamponade agent. Eyes treated with air or sulphur hexafluoride injections developed contractile membranes and eventual retinal detachment by 10 days and 2 months, respectively. Consistent with our previous observations, poly(CEP) injections were capable of preventing both retinal detachment and secondary fibrocellular growth. This further enforced idea that poly(CEP) had a bio-functional component capable of disrupting the pathogenesis of PVR.
Poly(CEP) has a multi-pronged anti-PVR mechanism
The pathogenesis of PVR is a complicated process involving RPE cells and various aberrations of cellular processes such as epithelial-mesenchymal transition (EMT), hyper-proliferation and increased migratory behaviour. To further interrogate the mechanism of poly(CEP), we modelled the pathogenic processes in vitro. By subjecting human embryonic stem cell-RPE cells to cytokines, such as TNF-α and TGF-β, involved in PVR pathogenesis, formation of contractile membranes could be recapitulated in a dish. We found out that poly(CEP) was capable of preventing membrane formation in vitro through the inhibition of EMT, hyper-proliferation and cellular migration. To elucidate the mechanisms underlying this phenomenon, we looked at how poly(CEP) can affect RPE cellular signalling pathways.
Activation of signalling cascades
To interrogate the subcellular effects of poly(CEP), we performed genome-wide transcriptomic screens of RPE cells treated with poly(CEP) at multiple time-points. Surprisingly, genes related to EMT, and proliferation were inhibited by 8 hours, indicating the cellular response was fairly quick.
Interestingly we noticed a significant upregulation of NRF2 signalling pathway at 8 hours. The NRF2 transcription factor is an emerging target against various diseases such as neurodegeneration and cancer, but its role in RPE cell migration, proliferation and epithelial mesenchymal transition has not been described previously.
We further validated that poly(CEP) prevented PVR via the NRF2-mediated mechanism by using a FDA-approved pharmacologic activator of NRF2 using the in vitro PVR contractile membrane model. Indeed, activation of NRF2 alone was sufficient to prevent contractile membrane formation. More importantly, we confirmed by RNA sequencing that NRF2 target genes were also upregulated in rabbit eyes, which were filled with poly(CEP), suggesting that similar NRF2 dependent mechanisms were happening in vivo.
Beyond the eye
Our study showcased the bio-functionality of a synthetic polymer with intrinsic anti-scarring properties. Poly(CEP) inhibited multiple hallmarks of PVR, providing a multi-pronged approach towards preventing intraocular scarring. This presents a disruptive approach of using biomaterials alone to overcome the challenge of post-operative intraocular scarring. Beyond ophthalmology applications, poly(CEP), the polymer may also have applications in other systemic diseases such as cancer. Furthermore, the study identifies NRF2 signalling pathway as a potential therapeutic target to halt PVR pathogenesis.
Most intriguingly, the ability of poly(CEP) to interact and regulate RPE cell behaviour deviates from the traditional view of synthetic polymers being biologically inert. This opens new possibilities of studying polymers for functions that involve polymer-cell interactions. Future characterisation of this polymer-cell interaction can enable rational design of polymers to elicit therapeutic functions without the loading of any active pharmacological agents.