Nanoformulações da angiotensina-(1-7) e do remdesivir com lipossomas: eficácia terapêutica por via intranasal em camundongos K18-hACE2 infectados por SARS-CoV-2

Abstract

ABSTRACT The COVID-19 pandemic highlighted the urgent need for effective therapeutic strategies to reduce the impact of the disease and protect individuals from severe acute respiratory syndrome. Several approaches have been explored, including the search for new molecules with therapeutic potential, the repurposing of existing anti-inflammatory and antiviral drugs and the development of novel formulations for pulmonary deliveryThe endogenous peptide angiotensin-(1-7) [Ang-(1-7)] emerged as a promising anti-inflammatory agent, while remdesivir (RDV) has been widely used due to its antiviral properties, despite the limitations of its pharmaceutical formulation Veklury®, such as moderate efficacy, high toxicity, and the requirement for parenteral administration. In this context, the studies that led to this thesis were primarily aimed at developing liposomal formulations for intranasal administration of Ang-(1-7) [LAng (1-7)] and RDV [LRDV], and evaluating their efficacy in SARS-CoV-2 infection models, including K18-hACE2 transgenic mice. The LAng (1-7) and LRDV formulations were prepared by the ethanol injection method, yielding vesicles with mean diameters of 100 nm and 120 nm, respectively, and polydispersity indices below 0.1. The encapsulation efficiencies of Ang-(1-7) and RDV were 13% and 65%, respectively, with release half-lives at 37 °C of 9.3 h and 5.3 h. In vitro cytotoxicity and antiviral assays of LRDV demonstrated a 3.5-fold higher selectivity index compared to the free drug. Pharmacokinetic studies of intranasally administered LRDV confirmed efficient drug delivery to the lungs. K18-hACE2 mice infected with SARS-CoV-2 were treated every 12 hours for 5 days with intranasal LAng (1-7), intranasal LRDV, or intraperitoneal Veklury®, with control groups receiving empty liposomes, free Ang-(1-7), or PBS. Treatment with LAng (1-7) increased survival compared with control groups, while also significantly reducing viral load and levels of IL-6 and TNF in the lungs. In the case of intranasal LRDV, all mice remained alive up to 15 days post-infection (dpi), whereas control groups showed 100% mortality by 6 dpi, and the group treated with intraperitoneal Veklury® exhibited 62.5% mortality by 8 dpi. Moreover, LRDV significantly reduced viral loads in both lungs and brain, preventing SARS-CoV-2-induced inflammatory responses in the lungs, which was not observed with Veklury®. In conclusion, this thesis provides proof-of-concept for innovative COVID-19 therapeutic strategies based on intranasal liposomal nanoformulations of Ang-(1-7) and RDV. LAng (1-7) demonstrated both anti-inflammatory and antiviral potential, while LRDV outperformed conventional treatment, reducing viral burden in lungs and brain and preventing pulmonary inflammation, making it a promising alternative also for post-COVID-19 conditions. More broadly, these findings underscore the therapeutic potential of intranasal liposomal nanoformulations in the treatment of COVID-19.