Safety and Biodistribution of Nanoligomers Targeting the SARS-CoV-2 Genome for the Treatment of COVID-19

Abstract

As the world braces to enter its fourth year of the coronavirus disease 2019 (COVID-19) pandemic, the need for accessible and effective antiviral therapeutics continues to be felt globally. The recent surge of Omicron variant cases has demonstrated that vaccination and prevention alone cannot quell the spread of highly transmissible variants. A safe and nontoxic therapeutic with an adaptable design to respond to the emergence of new variants is critical for transitioning to the treatment of COVID-19 as an endemic disease. Here, we present a novel compound, called SBCoV202, that specifically and tightly binds the translation initiation site of RNA-dependent RNA polymerase within the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) genome, inhibiting viral replication. SBCoV202 is a Nanoligomer, a molecule that includes peptide nucleic acid sequences capable of binding viral RNA with single-base-pair specificity to accurately target the viral genome. The compound has been shown to be safe and nontoxic in mice, with favorable biodistribution, and has shown efficacy against SARS-CoV-2 in vitro. Safety and biodistribution were assessed using three separate administration methods, namely, intranasal, intravenous, and intraperitoneal. Safety studies showed the Nanoligomer caused no outward distress, immunogenicity, or organ tissue damage, measured through observation of behavior and body weight, serum levels of cytokines, and histopathology of fixed tissue, respectively. SBCoV202 was evenly biodistributed throughout the body, with most tissues measuring Nanoligomer concentrations well above the compound K D of 3.37 nM. In addition to favorable availability to organs such as the lungs, lymph nodes, liver, and spleen, the compound circulated through the blood and was rapidly cleared through the renal and urinary systems. The favorable biodistribution and lack of immunogenicity and toxicity set Nanoligomers apart from other antisense therapies, while the adaptability of the nucleic acid sequence of Nanoligomers provides a defense against future emergence of drug resistance, making these molecules an attractive potential treatment for COVID-19.

Large- and Small-Animal Studies of Safety, Pharmacokinetics, and Biodistribution of Inflammasome-Targeting Nanoligomer in the Brain and Other Target Organs

“…There were no significant histologic findings/differences in the liver, kidney, or colons of mice treated with NI112. Kidney nephrons displayed no evidence of inflammation within the glomeruli, supporting the renal clearance shown previously for Nanoligomer molecules. , (C) Immune panel studies of dogs treated with 75 mg/kg NI112_dog which corresponds to 500 mg/kg dosing in mice, and is well above the MTD. Dogs were monitored for 2 weeks post administration.…”

“…We did not observe any evidence of accumulation, organ damage, toxicity, or any immune cell infiltration, as shown using H&E staining. Also, in long-term studies, there was no evidence of accumulation or resulting inflammation in the medulla or nephrons (within glomeruli, Figure B), supporting renal clearance for Nanoligomer molecules. , These results again demonstrate the high safety profile of the drug for repeated dosing over longer periods.…”

“…We first started with a maximum tolerable dose (MTD) assessment of the NF-κB and NLRP3 inhibitor combination NI112_mouse [ nfkb1 (Sequence: AGTGGTACCGTCTGCTA) and nlrp3 (Sequence: CTTCTACTGCTCACAGG)]. Prior studies have shown assessment of NI112 in relevant disease models (MS, AD, Prion/Creutzfeldt-Jacob disease, neuroinflammation, and autoimmune diseases), ,,,,, and Nanoligomer selectivity and high-binding affinity ( K D 3.17 nM, , other PNA/RNA and PNA/DNA binding studies showed K D 5–8 nM) along with rapid biodistribution and clearance. We observed a strong safety-toxicity profile in both animal models for longer-term repeat dosing (IP, 150 mg/kg, 3× a week for 12 weeks, Figures A,B, S1, and S2) as well as single-dose acute toxicity (5× MTD for IV), followed by 14 day toxicity assessment and clinical monitoring (Figures S3, C–F, S4, and S5).…”

“…), use of more traditional pharmaceutical modalities such as small-molecule-based targeting is slow and low-throughput with mixed safety profile. On the other hand, high-throughput approaches such as design-based antisense oligonucleotides can potentially improve translation rates, but suffer from delivery challenges, especially for neurological diseases, , and accumulation in first-pass organs. , The emergence of new RNA-targeting antisense modalities such as Nanoligomers can have the combined advantage of high delivery and biodistribution profile of small molecules, extremely high target-binding affinity ( K D ∼ 3–5 nM) like antibodies, with the specificity and ease of design using nucleic acid-targeting. − Nanoligomers are peptide nucleic-acid-based RNA therapeutic modality that have shown targeted up- and downregulation of proteins, with extremely high specificity, minimal off-targets, and a high safety profile. − However, more detailed safety-toxicity and biodistribution studies need to be conducted across small- and large-animal models for these new modalities to gain greater acceptance toward human clinical translation and pharma adoption.…”

“…While rodents are acceptable small-animal models for autoimmune and neurological disease modeling and induction, dogs are the preferred large-animal model for clinical drug translation to further assess the safety profile, central nervous system (CNS), PK/pharmacodynamic (PD), and biodistribution parameters . We conducted extensive small- and large-animal studies for our targeted inflammasome inhibitor, an NF-κB and NLRP3 inhibitor combination (herein described as SB_NI_112 or NI112 in short) as the active pharmaceutical ingredient (API), − to assess safety-toxicity, develop a PK and biodistribution model to obtain relevant pharmacological parameters, and obtain insights for interspecies scaling for further translation. We observed extremely high biodistribution parameters across different routes of administration (biodistribution parameter F > 0.98), excellent drug delivery across the blood–brain barrier (BBB) (brain tissue concentration ≥25–30% of blood serum bound concentration), rapid absorption across different brain sections, lack of accumulation in first-pass organs, as well as renal clearance of unbound/excess drug (or missense molecules). , These pharmacological results prove the potential for further translation and the need for the adoption of new therapeutic modalities for the benefit of millions of patients across multiple autoimmune and CNS diseases.…”