Abstract
In single-gene disorders, like alpha-1-antitrypsin deficiency (AATD), a gene mutation causes missing or dysfunctional protein synthesis. This, in turn, can lead to serious complications for the patient affected. Furthermore, single-gene disorders are associated with severe early-onset conditions and necessitate expensive lifelong care. Until nowadays, therapeutic treatment options are still limited, cost-intensive, or lack effectiveness. For these reasons, we aim to develop a novel mRNA-based therapeutic strategy for the treatment of single-gene disorders, such as AATD, which is based on the induction of de novo synthesis of the functional proteins. Therefore, an alpha-1-antitrypsin (AAT) encoding mRNA was generated by in vitro transcription. After in vitro delivery of the mRNA to different cells, protein expression and functionality, as well as adverse effects and mRNA serum stability, were analyzed. Our results show that the AAT mRNA-transfected cells express the AAT protein in high amounts within the first 24 h. Moreover, the expressed AAT protein is highly functional, since the activity of elastase is significantly inhibited. Our data also show that mRNA concentrations up to 1 μg per 150,000 cells have no adverse effects on cell viability and immune activation. Furthermore, the encapsulated AAT encoding mRNA is stable and functional in human serum for up to 30 min. Overall, the proposed project provides an innovative, highly promising, and safe therapeutic approach and, thus, promises a novel progress in the treatment of single-gene disorders, whereby affected patients could greatly benefit.