A Promising Approach for treating SARS-CoV-2 infection Direct Lung Delivery of Targeted Antiviral Drug-loaded Nano Liposomal Suspension Product for the Treatment of SARS-CoV-2 Infection
Mahavir Bhupal Chougule1, Rudragouda Channapanavar2, and Sterling Symone Neill1,
1 Ingenious BioPharma-engineered Drugs and Biologics Delivery Laboratory (iBD2 lab), Department of Pharmaceutical Sciences, Mercer University, Atlanta, Georgia, 30341, USA.
2 Department of Veterinary Pathobiology, College of Veterinary Medicine, Oklahoma State University, Stillwater, OK, 74078, USA
COVID-19 and Liposomes
The epidemic event of coronavirus disease (COVID-19) has plagued the globe, prompting new treatments to be researched and implemented to combat COVID-19. Nanoliposomes are classified into four categories based on the number of bilayers and the liposome size. These four classification categories include small unilamellar vesicles (SUV), large unilamellar vesicles (LUV), multilamellar vesicles (MLV), and multivesicular vesicles (MVV). Lipid vesicles or liposomes are spherical and range in size with a diameter between 50 and 500 nm. The size of liposomes is not the only variable. Liposomes also vary in the number of lipid bilayers, which depends on the composition of the aqueous medium. The number of lipid bilayers affects the nanoliposome's ability to encapsulate drug molecules.
Nanoliposomes are bilayer lipid vesicles that encapsulate hydrophobic and/or hydrophilic bioactive agents, small molecule inhibitors, peptides, proteins, iRNA, mRNA, antigen, antibody, and Casper9, making them versatile nanocarriers in targeted drug delivery systems. The size and % drug load affect the aerodynamic properties of liposomal products such as mass median aerodynamic diameter (MMAD), % fine particle fraction (FPF), and Geometric Standard Deviation (GSD). It also affects the stability, fate of liposomes, and release of bioactive agents at the tissues and site of action. Many nanoliposome factors affecting the product suggest rationalized strategies, including the loading capacity, encapsulation efficiency, stability during aerosolization, and lack of target specificity. Design of Experiments (DoE) and Quality by Design (QbD) strategies are required for developing inhalable pharma or biopharma products, meeting the target profile for successful delivery at the site of action while overcoming the limitations. Also, the inhalable product delivers bioactive agents directly to the site of action while minimizing distribution in the plasma and other organs, thereby overcoming the side effects that occur in the conventional route of administration. Nanoliposomes have entered many clinical applications as nanocarriers or nano-sized materials that enable the transportation of drug molecules to target tissues. Few are FDA-approved products administered via the inhalation route.
Inhaled Nanomedicine
The inhalable bioactive drug-loaded nanoliposomes are delivered through a nebulizer, dry powder inhaler, and pressurized meter dose inhaler. A nebulizer is widely used for the delivery of nanosuspension via nebulization. It is a machine that alters liquid or suspension products to an aerosol cloud. The four types of nebulizers are used: jet, mesh, ultrasonic, and soft mist inhalers. Jet nebulizers use compressed gas to convert the nanosuspension to an aerosol cloud. In contrast, mesh nebulizers pass drugs through a mesh to create a fine mist, and ultrasonic nebulizers use high-frequency vibrations to convert drug molecules into an aerosol cloud. Nebulization is advantageous as it allows for specific pulmonary drug delivery, which can overcome biological-related obstacles, enhance aerosol lung deposition fraction, and increase bioactive agent bioavailability and delivery to the site of action. Inhalable targeted strategically designed nanoliposomes with the target profile ensure improved bioactive stability, delivery to the lungs, use of lower dose, improved patient compliance, and a broad range of applicability for both treatment and prevention. Since nanoliposomes are adaptable, they can be tailored to carry various bioactive agents, target specific tissues, and provide extended release. Inhalable nanoliposomal products have the potential to deliver a range of therapeutic agents that will aid in the treatment of various lung diseases, such as chronic obstructive pulmonary disease, cystic fibrosis, bacterial infections, viral infections including coronavirus disease 19 (COVID-19), asthma etc. COVID-19 is a disease caused by the infection virus SARS-CoV-2 in nasal and/or lung tissues. COVID-19 enters host human cells by attaching its spike proteins to the Angiotensin-converting enzyme 2 (ACE2) receptor at the host cell surface. The virus particle first binds with the ACE2 receptor and is cleaved by Transmembrane Serine Protease 2 (TMPRSS2), which enables the virus to enter human cells.
Antiviral agents loaded nanoliposomal products
Antiviral agents loaded nanoliposomal products have huge potential in the treatment and/or prophylaxis of COVID-19). SARS-CoV-2 is highly transmissible; therefore, treatment methods are required to treat and prevent the spread of the disease. Since the mortality rate is over seven million people, finding effective treatments for patients suffering from COVID-19 is essential without the interference of therapeutic agents. One of the promising strategies is to treat the diseased patient and prevent viral infection from spreading. The antiviral Camostat mesylate inhibits TMPRSS2 by inhibiting serine proteases from cleaving the spike protein, which will hinder SARS-CoV-2 from infecting other cells and proliferating within host cells. To ensure the inhibition of SARS-CoV-2 viral entry into host cells, Camostat mesylate (Camo) was encapsulated in nanoliposomes (Fig. 1), had an extended-release profile, and was PEGylated or underwent a process that covalently attached polyethylene glycol (PEG) chains, which imparted the salt stability and minimized macrophage uptakes.
Our lab developed Camo-loaded PEGylated nanoliposomes (Camo-pegNLs) using an industrially scalable ethanol injection method. Nanoliposome size, polydispersity index (PDI), and zeta potential are used to characterize or analyze the nanoliposome's physical, chemical, and biological properties. Camo-pegNLs suspension was 167.50 ± 0.90nm, PDI 0.07 ± 0.01, and zeta potential 0.48 ± 0.04 mV. The entrapment efficiency was found to be 93.9 ± 2.1%w/v. The Camo-pegNL-2 showed an extended-release profile for up to 24 hours, 4.295 ±0.1 µm MMAD, 1.915 ± 0.064 GSD, and 42.01% ± 6.90 FPF. The extended release of the antiviral Camo from Camo-pegNLs inhibits spike protein cleavage, inhibiting viral entry and replication, demonstrating the application for COVID-19 treatment.
The release of Camo from the nebulized product and interaction with TMPRSS2, which allowed the patient to exert an antiviral mechanism and effect, was depicted in Figure 2A-D. In the antiviral assay using the SARS-CoV-2 infection on Vero cells model, a significant antiviral effect was found in the Camo-pegNLs treated compared to no treatment group (p<0.01). This suggests that the developed nebulized Camo-pegNLs suspension product could be an effective treatment for COVID-19. The scientific findings of our investigation provide promising evidence for the potential of nanoliposomes in treating viral lung diseases.