Translation of Angiotensin-Converting Enzyme 2 upon Liver- and Lung-Targeted Delivery of Optimized Chemically Modified mRNA

Schrom, Eva; Huber, Maja; Aneja, Manish; Dohmen, Christian; Emrich, Daniela; Geiger, Johannes; Hasenpusch, Günther; Herrmann-Janson, Annika; Kretzschmann, Verena; Mykhailyk, Olga; Pasewald, Tamara; Oak, Prajakta; Hilgendorff, Anne; Wohlleber, Dirk; Hoymann, Heinz-Gerd; Schaudien, Dirk; Plank, Christian; Rudolph, Carsten; Kubisch-Dohmen, Rebekka

Fibrotic diseases are major causes of mortality and morbidity worldwide, leading to a serious economic burden and challenges for health services. 1, 2 Fibrosis is caused by repetitive noxious stimuli leading to cell stress, injury, and apoptosis, causing organ dysfunction and ultimately organ failure. 3, 4 Fibrosis can affect nearly every organ, with liver and lung fibrosis showing a rising prevalence due to lifestyle changes or unfavorable environmental conditions. [5] [6] [7] Currently, there are several treatment options under evaluation for liver and lung fibrosis; however, these have had limited therapeutic success, raising the need for new therapeutic approaches. [8] [9] [10] Investigations of the underlying mechanisms of fibrotic diseases showed that dysregulation of the organ-specific renin-angiotensin system (RAS) plays a critical role in disease onset by triggering excessive pro-inflammatory and profibrotic signaling. 1 Angiotensin-converting enzyme 2 (ACE2) is a family member of the RAS and acts as a metallo-carboxypeptidase cleaving angiotensin II (AngII) to Ang-(1-7). AngII acts on the AngII type I receptor (AT1R), leading to pro-inflammatory and pro-fibrotic signaling, while Ang-(1-7) acts on the Mas oncogenic receptor (Mas receptor), leading to anti-inflammatory and anti-fibrotic signaling. Thus, ACE2 can shift the RAS balance by reducing the amount of AngII and at the same time increasing the amount of Ang-(1-7) molecules. Therefore, ACE2 not only re-establishes the physiologic balance of the RAS, but it also clearly shifts it toward resolution of inflammation and fibrosis. [11] [12] [13] ACE2 levels are markedly increased in patients with liver fibrosis as well as in experimental models, which may be due to a counter-regulatory response to RAS upregulation. 14 ACE2 knockout studies revealed that the loss of ACE2 leads to exacerbation of liver injury, which can be attenuated by administration of recombinant ACE2 15 or by adeno-associated ACE2 gene therapy. 16 Similar therapeutic effects were shown by administration of Ang-(1-7) 17 and as a side-effect of AngII receptor blocker (ARB) therapy. 18, 19 Interestingly, unlike observations in liver fibrosis, ACE2 levels are markedly decreased in lung tissue from patients suffering from idiopathic pulmonary fibrosis (IPF). 20 In murine models, two studies showed that intraperitoneal injection of exogenous ACE2 protein in bleomycin-induced lung fibrosis led to re-establishment of local ACE2 levels and reduced levels of lung injury. 21, 22 The same protective effect of ACE2 could be shown in a bleomycin-induced mouse model by intratracheal administration of lentiviral packaged Ang-(1-7) fusion gene or ACE2 cDNA. 23 RNA transcript therapy (RTT) has gained substantial attention as a newly evolving therapeutic approach. mRNA exerts its function in the cytoplasm, leading to high and reliable transfection efficiency in proliferating as well as quiescent cells without the risk of insertional mutagenesis faced by viral vectors or plasmid DNA (pDNA). In comparison to recombinant protein therapy, RTT is not limited to secreted proteins, making it an interesting alternative for translation of intracellular or membrane-bound protein. Due to enzymatic mRNA degradation mechanisms in the cytoplasm, protein translation is controllable, as it is naturally self-limited. [24] [25] [26] Many of the initial obstacles of mRNA therapy, such as RNA instability and immunogenicity, have been solved, offer