Specific delivering of RNAi using Spike's aptamer‐functionalized lipid nanoparticles for targeting SARS‐CoV‐2: A strong anti‐Covid drug in a clinical case study

Abstract

Coronavirus (SARS‐CoV‐2) as a global pandemic has attracted the attention of many scientific centers to find the right treatment. We expressed and purified the recombinant receptor‐binding domain (RBD) of the SARS‐CoV‐2 spike (S) protein, and specific RBD aptamers were designed using SELEX method. RNAi targeting nucleocapsid phosphoprotein was synthesized and human lung cells were inoculated with aptamer‐functionalized lipid nanoparticles (LNPs) containing RNAi. The results demonstrated that RBD aptamer having K D values of 0.290 n m possessed good affinity. Based on molecular docking and efficacy prediction analysis, siRNA molecule was showed the best action. LNPs were appropriately functionalized by aptamer and contained RNAi molecules. Antiviral assay using q‐PCR and ELISA demonstrated that LNP functionalized with 35 µ m Apt and containing 30 n m RNAi/ml of cell culture had the best antiviral activity compared to other concentrations. Applied aptamer in the nanocarrier has two important functions. First, it can deliver the drug (RNAi) to the surface of epithelial cells. Second, by binding to the SARS‐CoV‐2 spike protein, it inhibits the virus entrance into cells. Our data reveal an interaction between the aptamer and the virus, and RNAi targeted the virus RNA. CT scan and the clinical laboratory tests in a clinical case study, a 36‐year old man who presented with severe SARS‐CoV‐2, demonstrated that inhalation of 10 mg Apt‐LNPs‐RNAi nebulized/day for six days resulted in an improvement in consolidation and ground‐glass opacity in lungs on the sixth day of treatment. Our findings suggest the treatment of SARS‐CoV‐2 infection through inhalation of Aptamer‐LNPs‐RNAi.

Selection and Validation of siRNAs Preventing Uptake and Replication of SARS-CoV-2

“…Structure proteins are needed for virion assembly, and it has been shown that in particular the spike protein is under high selection pressure, resulting in high numbers of mutations ( Weisblum et al, 2020 ; Callaway, 2021 ; He et al, 2021 ; Chen et al, 2022 ). However, an approach based on siRNA targeting the N-protein gene of SARS-CoV-2 (complexed with lipid nanoparticles that coupled to ACE2 binding aptamers) has been described by Saify Nabiabad et al (2022) . Since aptamers were used in this study having an inhibitory effect on viral infection solely by blocking the binding of SARS-CoV-2 spike protein to ACE2, the proportion of efficacy of the siRNA in vivo is unknown.…”

Therapeutic siRNA: State-of-the-Art and Future Perspectives

“…Likewise, RNAi is a promising therapeutic approach to fight infections caused by pathogenic viruses featuring single-stranded RNA genomes and subgenomic RNA transcripts (e.g., hepatitis C virus, respiratory syncytial virus, influenza, coronaviruses) [13][14][15][16]. The worldwide rampant severe acute respiratory syndrome coronavirus 2 pandemic has especially catalyzed the progress in promising antiviral siRNA therapy development [17][18][19][20][21].…”

Aptamers for SARS‐CoV‐2: Isolation, Characterization, and Diagnostic and Therapeutic Developments

“…in China in 2020 where the receptor‐binding domain (RBD) of the S protein from the wildtype SARS‐CoV‐2 was used as the target [42] . RBD was also used as the target for aptamer selection in four other studies [44,51–53] . The S1 subdomain and full trimeric S proteins have also been used as targets for SELEX, as reported by three and two groups, respectively [43,45,47,48,50] .…”

“…We note that a ∼40‐nucleotide random region has been widely used in previous SELEX studies, [15] and thus it is not surprising that most of the S protein aptamer selections adopted this size. The study by Saify Nabiabad used a random domain of only 22 nucleotides [52] . Although this size of random domain is less common, small‐sized aptamers do exist.…”