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Limbal Approach-Subretinal Injection of Viral Vectors for Gene Therapy in Mice Retinal Pigment Epithelium
Published on: August 7, 2015
Less is more: Self-amplifying mRNA becomes self-killing upon dose escalation in immune-competent retinal cells
Helena Vanluchene1, Oriane Gillon2, Karen Peynshaert1
1Ghent Research Group on Nanomedicines, Laboratory of General Biochemistry and Physical Pharmacy, Faculty of Pharmaceutical Sciences, Ghent University, Ottergemsesteenweg 460, 9000 Ghent, Belgium.
Abstract:
In the last few years, mRNA therapeutics experienced a new wave of interest as therapy for retinal diseases. Nevertheless, despite the widespread use of mRNA vaccines in the COVID-19 pandemic, mRNA delivery to the eye is still in its infancy. Recently, our research group has demonstrated that after subretinal and intravitreal delivery of modified mRNA, the number of transfected retinal cells and protein expression per cell remains limited. In this study, we aimed to tackle this limitation by using self-amplifying mRNA (saRNA), which in theory will increase the duration and level of protein expression when only a few mRNA molecules reach their target cells. A one-on-one comparison between modified mRNA and saRNA in two immune-competent human retinal cell types, including Müller cells and retinal pigment epithelial cells, and in immune-deficient BHK-21 cells revealed that saRNA delivery induced an innate immune response blocking its own translation above a certain dose threshold. Removal of double-stranded (ds)RNA byproducts by cellulose-based purification and addition of the innate immune inhibitor B18R remarkably improved translation from saRNA through a reduction in innate immune response. Taken together, when saRNA is applied for retinal disease, the dose should be controlled and measures should be taken to limit immunogenicity.
Insights
Self-amplifying mRNA (saRNA) shows promise for retinal diseases but triggers an immune response. Controlling saRNA dose and limiting immunogenicity are crucial for effective therapeutic delivery to the eye.
Area of Science:
- Ophthalmology
- Molecular Biology
- Immunology
Background:
- Messenger RNA (mRNA) therapeutics are gaining interest for treating retinal diseases.
- Current mRNA delivery to the eye faces limitations in cell transfection and protein expression.
- Self-amplifying mRNA (saRNA) offers potential for enhanced protein expression in target cells.
Purpose of the Study:
- To evaluate saRNA as a therapeutic tool for retinal diseases.
- To compare the efficacy of saRNA versus modified mRNA in retinal cells.
- To investigate and overcome the immunogenicity challenges associated with saRNA delivery.
Main Methods:
- Compared modified mRNA and saRNA in human Müller cells, retinal pigment epithelial cells, and BHK-21 cells.
- Investigated the innate immune response triggered by saRNA delivery.
- Assessed the impact of double-stranded RNA (dsRNA) byproduct removal and innate immune inhibition (B18R) on saRNA translation.
Main Results:
- saRNA delivery induced an innate immune response that inhibited its own translation above a specific dose.
- Cellulose-based purification to remove dsRNA byproducts significantly improved saRNA translation.
- Addition of the innate immune inhibitor B18R further enhanced saRNA translation by reducing the immune response.
Conclusions:
- saRNA holds potential for retinal disease therapy by increasing protein expression duration and levels.
- The immunogenicity of saRNA must be managed for successful ocular application.
- Controlling saRNA dosage and implementing strategies to mitigate innate immune responses are essential for therapeutic efficacy.
