WO2025054542A1

WO2025054542A1 - Methods, apparatus and systems for fabricating a carrier for competitively inhibiting pathogens in a host

Info

Publication number: WO2025054542A1
Application number: PCT/US2024/045709
Authority: WO
Inventors: Pei Zhang
Original assignee: Finncure Oy
Current assignee: Finncure Oy
Priority date: 2023-09-08
Filing date: 2024-09-06
Publication date: 2025-03-13
Anticipated expiration: 2026-03-08

Abstract

According to some embodiments, a method of fabricating nano and/or micro sized materials using a microfluidics device to be used for competitively inhibiting an agent. A method of manufacturing a carrier, including forming a core using a microfluidics device, combining the core with at least one protrusion, maintaining the core and the at least one protrusion to a temperature from about 3 ºC to about 5 ºC; and functionalizing the core by securing the at least one protrusion to the core. Also described are apparatus and systems for manufacturing a carrier.

Description

METHODS, APPARATUS AND SYSTEMS FOR FABRICATING A CARRIER FOR COMPETITIVELY INHIBITING PATHOGENS IN A HOST

CROSS-REFERENCE TO RELATED APPLICATIONS

[0001] This application claims the benefit of U.S. Provisional Patent Application Number 63/537475 filed September 8, 2023, entitled METHODS OF FABRICATING PATHOGENMIMICKING MATERIALS, the disclosures of which are incorporated herein by reference in their entirety. The contents of PCT Application PCT/FI2021/050259, filed April 9, 2021 , and published on October 14, 2021, as PCT Publ. WO 2021/205077, entitled MIMETIC NANOPARTICLES FOR PREVENTING THE SPREADING AND LOWERING THE INFECTION RATE OF NOVEL CORONAVIRUSES, is incorporated by reference herein in its entirety and made part of the present application. Any and all applications for which a foreign or domestic priority claims is identified in the Application Data Sheet as filed with the present application are hereby incorporated by reference under 37 C.F.C. § 1 .57.

BACKGROUND

Field

[0002] The present application generally relates to synthetic materials and compositions, and more specifically, apparatus, systems and methods for manufacturing nano and/or micro-sized particles.

SUMMARY

[0003] According to some embodiments, a method of manufacturing a carrier comprises forming a core using, at least in part, microfluidics, wherein the core comprises at least one component, wherein the core comprises a maximum size in at least one dimension in a nanometer or a micrometer range, including at least one additive within, on and/or to the core, and functionalizing the core with at least one protrusion, wherein the carrier is capable of binding to targeted areas of a host.

[0004] According to some embodiments, forming the core comprises using a microfluidic device.

[0005] According to some embodiments, the microfluidic device comprises a first glass capillary and at least a second glass capillary. In some embodiments, the first glass capillary comprises an inner diameter of 500 to 600 pm and an outer diameter of around 900 to 1200 pm. In some embodiments, the first glass capillary comprises a tip that is polished and its diameter enlarged to around 100 pm. In some embodiments, the second glass capillary comprises an outer cylindrical capillary with an inner diameter of 1120 pm and an outer diameter of 1500 pm. In some embodiments, the second glass capillary comprises an outer cylindrical capillary with an inner diameter of 1000 to 1200 pm and an outer diameter of 1400 to 1600 pm. In some embodiments, the inner capillary was coaxially inserted into the outer cylindrical capillary. In some embodiments, the capillaries were fixed on a glass slide and sealed as required.

[0006] According to some embodiments, a carrier is manufactured, at least in part, using one or more methods disclosed herein.

[0007] According to some embodiments, a core of the carrier comprises at least one of the following: a lipid, a cholesterol, a polymer, a monomer, an amino acid, a protein, a salt, a mineral, another organic and/or inorganic component and/or another molecule.

[0008] According to some embodiments, the core comprises at least one additive. In some embodiments, the at least one additive comprises at least one of the following: an API, a drug, a protein, an amino acid, a lipid, a cholesterol, a polymer, a monomer, an amino acid, a protein, a salt, a mineral, a molecule, RNA, DNA, another active and/or inactive substance.

[0009] According to some embodiments, the core is functionalized with a plurality of protrusions that at least partially mimic naturally occurring protrusions of a target pathogen or agent. In some arrangements, the core is functionalized such that the protrusions are attracted to the core. In some embodiments, the core is functionalized such that the protrusions are attracted to the core using at least one condition favorable for attracting the protrusions to the core. In some embodiments, the conditions favorable for attracting the protrusions to the core comprises subjecting the carrier to a temperature or temperature range around room temperature or below room temperature but over freezing point. In some embodiments, attracting the protrusions to the core is based on, at least in part, at least one of the following: covalent binding, non-covalent interactions, van der Waals forces, hydrophobic interactions, electrostatic interactions and polar interactions.

[0010] According to some embodiments, the carrier is configured to bind to targeted areas of a target pathogen or agent to at least partly inhibit entry of the target pathogen or agent into the host’s cells. In some embodiments, the targeted areas comprise at least one receptor, target molecule, amino acid or nucleotide.

[0011] According to some embodiments, the target pathogen or agent comprises at least one of a virus, a bacterium, a parasite, an antigen, a protein, a prion, a mold, a fungus, a toxin, a poison and an allergen. In some embodiments, the virus comprises at least one of the following: a coronavirus, a SARS-CoV-2 virus, an influenza virus, a rhinovirus, a norovirus, a respiratory syncytial virus (RSV), another virus that impacts the respiratory system and any other type of virus.

[0012] According to some embodiments, at least partially inhibiting entry of the target pathogen or agent into the host’s cells comprises competitive inhibition.

[0013] According to some embodiments, a method of manufacturing a carrier comprises forming a core using microfluidics made from at least one component, wherein the core comprises a maximum size in at least one dimension in a nanometer or a micrometer range, loading the core with at least one material; and

[0014] functionalizing the core with a plurality of protrusions, wherein the carrier is capable of binding to targeted areas a host, wherein the targeted areas comprise target areas of cell structure of the host’s cells, and wherein the carrier binding to target areas of the host at least partially inhibits pathogen entry into the host to prevent injection or disease that may otherwise be caused by a pathogen of interest.

[0015] According to some embodiments, the method comprises using a microfluidic device. In some embodiments, the microfluidic device comprises coaxially aligned assembling of two glass capillaries. In some embodiments, a first glass capillary of the two glass capillaries comprises an inner capillary having an inner diameter of 500 to 700 pm (e.g., 580 pm) and outer diameter of 800 to 1200 pm (e.g., 100 pm). In some embodiments, a tip of the first glass capillary comprises a smooth or substantially smooth surface.

[0016] According to some embodiments, a second glass capillary of the two glass capillaries comprises an outer cylindrical capillary with an inner diameter of 1000 to 2000 pm (e.g., 1120 pm) and an outer diameter of 1300 to 2000 pm (e.g., 1500 pm). In some embodiments, the inner capillary is configured to be coaxially inserted into the outer cylindrical capillary.

[0017] According to some embodiments, the capillaries are fixed on a glass slide or other glass surface and at least partially sealed.

[0018] According to some embodiments, the core is made of or comprises organic or inorganic components, lipid, cholesterol, polymers, monomers, amino acids, proteins, salts, minerals, other molecules or any combination thereof.

[0019] According to some embodiments, the core is configured to receive at least one material. In some embodiments, the at least one material comprises one or more of the following: an API, a drug, a protein, an amino acid, RNA, DNA, lipid, cholesterol, polymers, monomers, amino acids, proteins, salts, minerals, a molecule, active substance, any other material any combination thereof.

[0020] According to some embodiments, the core is functionalized with a plurality of protrusions that at least partially mimic naturally occurring protrusions of a target pathogen or agent.

[0021] According to some embodiments, the core is functionalized using conditions favorable for attracting the protrusions to said core. In some embodiments, the conditions favorable for attracting the protrusions to said core is performed using temperatures around room temperatures or below room temperature but over freezing point. In one embodiment, attracting the protrusions to said core is based on, at least in part, covalent binding, non- covalent interactions, van der Waals forces, hydrophobic interactions, electrostatic interactions, polar interactions and/or the like.

[0022] According to some embodiments, the protrusions are configured to at least partially bind to targeted areas of interest of the host.

[0023] According to some embodiments, binding to said targeted areas of interest at least partly inhibits pathogen entry to host’s cells.

[0024] According to some embodiments, binding to multiple targeted areas of said interest at least partly inhibits an entry of two or more different pathogens or agents to host’s cells.

[0025] According to some embodiments, target areas of interest comprise at least one of a receptor, a target molecule, an amino acid and a nucleotide.

[0026] According to some embodiments, the pathogen of interest comprises a virus, a bacterium, a parasite, an antigen, a protein, a prion, a mold, a fungus, a toxin, a poison or an allergen. In some embodiments, the virus is one or more of the following: a coronavirus, a SARS-CoV-2 virus, an influenza virus, a rhinovirus, a norovirus, a respiratory syncytial virus (RSV), another virus that impacts the respiratory system and any other type of virus.

[0027] According to some embodiments, inhibiting pathogen entry to host’s cells comprises of competitive inhibition.

[0028] According to some embodiments, methods of fabrication of man-made, synthetic particles, compositions and/or other materials, as well as the manufactured particles, materials and/or other compositions, are disclosed herein. In some embodiments, such particles or other materials comprise nano- and/or micro-sized particles that are used to reduce the likelihood of (e.g., help prevent) the infection and/or spread of infection by a pathogen (e.g., virus, bacteria, etc.) and/or other agent that may lead to disease or other condition. In some embodiments, such particles or other materials are used for targeted therapy of a subject. In some embodiments, the manufactured particles or other materials include nanoparticles and/or microparticles. Such nanoparticles and/or microparticles can be fabricated and/or otherwise manufactured to include any desired shape or configuration, such as, for example and without limitation, spheroid or substantially spheroid, cubical, cigar-shaped, elongated, triangular, particles having smooth outer or exterior surfaces, particles having sharp and/or pointy (e.g., non-smooth) outer or exterior surfaces, a sheet, a film and/or any configuration or shape, as desired or required.

[0029] According to some embodiments, the fabricated material, which may form the core structure of a carrier, is manufactured or otherwise obtained using, at least in part, one or more of the following methods or techniques: three-dimension (3D) printing, microfluidics, sol-gel methods, other bottom-up methods of fabrication, other top-down methods of fabrication and/or the like. Such manufacturing methods or processes can include but are not limited to, by way of example and without limitation, genetically engineered organism producing specific proteins or amino acids that can either self-assemble, such as, for example, ferritin protein particles or conjugate to larger entities, any other method or technique and/or combinations thereof.

[0030] According to some embodiments, a core and/or any other material or member produced using one or more of the various methods disclosed herein can be strategically coated, decorated and/or otherwise functionalized. In some embodiments, such functionalization comprises performing one or more of the following to the core: coating, applying, decorating, binding and/or doing another procedure to the fabricated (e.g., manmade) particle or material (e.g., the core). In some embodiments, such functionalization can increase an affinity of the core or other particle (e.g., once functionalized) to an area of interest or targeted area or targeted entity or targeted structure favoring the binding of the functionalized carrier to the said area. For example, in some arrangements, functionalization can increase the likelihood of the particle (e.g., the functionalized core) to bind to specific sites of host cells, to pathogens and/or other target agents and/or the like.

[0031] According to some embodiments, the core or other base structure of a carrier is obtained by 3D printing, microfluidics, sol-gel method or other bottom-up and/or top-down method of fabrication and/or the like.

[0032] According to some embodiments, nano and/or micro-sized particles or other materials are manufactured or otherwise fabricated using microfluidics (e.g., using one or more microfluidic device). In some embodiments, nano and/or micro-sized particles or other materials are manufactured or otherwise fabricated using microfluidics (e.g., using one or more microfluidic device) in combination with one or more other top-down and/or bottom-up method of synthetizing man-made materials.

[0033] According to some embodiments, the particles or other materials disclosed herein, including one or more methods of fabrication or manufacture disclosed herein, are used to produce pharmaceutical products, biologies, medical devices, over-the-counter drugs, consumer products preventing, reducing the likelihood and/or reducing the spread of pathogens or other disease or disorder-causing agents (e.g., coronaviruses (e.g., the Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2)), influenzas, viruses causing respiratory infection, diarrhea, common cold, cytokine storm, general discomfort and/or death, bacteria, other pathogens, other agents, etc.). In some embodiments, the present application relates to nano- and/or micro-sized or based carriers, particles and/or other materials. In some embodiments, carriers are manufactured, fabricated, functionalized and/or otherwise configured to at least partially mimic pathogens and/or other targeted agents, to reduce the likelihood of infection, entry and/or spread of pathogens and infectious agents (e.g., viruses (e.g., influenzas, rhinoviruses, noroviruses, respiratory syncytial virus (RSV), SARS-CoV-2, future strains and/or types of coronaviruses derived thereof, etc.), bacteria, parasites, antigens, prions, mold, fungi, toxins, poisons, allergens and/or the like).

[0034] According to some embodiments, prevention, treatment and/or other targeting of two or more pathogens (and/or stains of pathogens) and/or other agents comprises functionalizing the surface of the core with several a plurality (e.g., two or more) different protrusions and/or surface features or configurations that at least partially mimic that of the target pathogen and/or other agent. In some embodiments, at least one active pharmaceutical ingredient (“API”) and/or other component (e.g., minerals, anti-viral or other anti-agents, nucleic acids, proteins and/or the like) can be loaded inside and/or otherwise included to a carrier system capable of delivering the drug and/or other component specifically to target cells and/or tissues of a host to whom the carrier or similar device is administered. In some embodiments, this application relates to the fabrication, the resulting man-made materials (e.g., in the nano- and/or microscale) and/or the use thereof to, at least partially, saturate and/or bind to receptors, proteins and/or macromolecules at the cellular level. In some embodiments, such configurations can help prevent and/or minimize (or reduce the likelihood of) pathogens and/or other targeted agents binding to target areas of the cells (e.g., novel coronaviruses binding and entering a host’s cells and/or tissues) via, at least in part, competitive inhibition. [0035] According to some embodiments, targeted drug delivery of at least one active pharmaceutical ingredient (“API”) and/or another component is loaded inside and/or otherwise secured to a carrier system capable of delivering a drug and/or other loaded material(s) to target cells and/or tissues of a host. In some embodiments, man-made materials (e.g., carrier or other materials in the nano- and/or microscale range) described herein (and the related methods of manufacturing or fabricating the same) at least partially reduce or prevent disease progression by binding to target sites of host cells and/or tissues. Accordingly, APIs and/or other materials can be strategically delivered to host cells in order to enter such host cells for targeted delivery with minimal or reduced off-target effects or impacts. In some embodiments, increasing the therapeutic window of drugs and/or other components or materials loaded inside the carrier, especially using drugs and/or other components or materials that have a favorable solubility (e.g., relatively low solubility), have a favorable permeability (e.g., relatively low permeability) and/or are prone to aggregate without a composition or carrier.

[0036] According to some embodiments, a method comprises the steps of (a) providing a core material, e.g., a nano- and/or micro-material including nanoparticles, microparticles or any other object as disclosed herein, (b) coating or otherwise functionalizing the core material with molecules, polymers, amino acids, proteins, API, drugs or other material as disclosed herein, (c) optionally loading the object with compounds, molecules, drugs, API, DNA or RNA, and/or the like, (d) coating a second protective or functional layer on top of the object in particular for increasing its resistance that could be important in extreme environments such as the acidic environment in the stomach, and (e) providing a device, medical device, inhalation device or aerosol, sanitation product or consumer product that on- demand will release the containing synthetic material, particle or object for administration.

[0037] Certain non-limiting embodiments of the present technology are provided below and/or other portions of this application. The inclusion of such embodiment is not intended to create a non-exclusive list of embodiments, and as such, should not be seen to limit, in any way, the various inventions disclosed herein.

[0038] According to some embodiments, a method of preventing or reducing pathogen binding to target areas of cell surfaces of a host selected from mammals, comprising providing administering to the mammal a carrier comprising biocompatible particles having a maximum size in at least one dimension in the nanometer or micrometer range, forming a core, and further having a functionalized surface capable of binding to said target areas of said cell surfaces to at least temporarily block said target areas to prevent or minimize pathogen binding and thus, reducing the risk of the host contracting a disease caused by said pathogen.

[0039] According to some embodiments, a carrier (e.g., in accordance with any of the carrier embodiments disclosed herein or equivalents thereof, any carrier fabricated, ats least in part, using any of the fabrication embodiments disclosed herein, etc.) can be configured to function, at least in part, as a targeted vaccination, thereby minimizing or at least reducing the ability of the target virus or other pathogen to bind and entry to the host lowering the risk of contracting the specific infectious agent.

[0040] According to some embodiments, the core or other base structure of the carrier is being obtained by 3D printing, microfluidics, sol-gel method or other bottom-up and/or top-down method of fabrication.

[0041] According to some embodiments, the core comprises one or more of the following: organic or inorganic components, lipid droplets, amino acids, proteins, salts and minerals or other molecules or wherein the core material comprises mesoporous silica nanoparticles, in particular mesoporous silica particles with ordered mesostructures of pores that preferably are capable of being loaded with drugs.

[0042] According to some embodiments, the core material is functionalized with substance selected from the group consisting of amino acids, epitopes, peptides, proteins and/or protein fragments, chemical agents, active pharmaceutical ingredients (API), organic or inorganic polymers or molecules and combinations thereof.

[0043] According to some embodiments, the core material is functionalized with substance selected from the group consisting of peptides, proteins such as antibodies, chemical agents, active pharmaceutical ingredients (API), organic or inorganic polymers or molecules and combinations thereof.

[0044] According to some embodiments, the carrier functionalized for specifically binding to receptors, proteins and macromolecules at the cellular level in order to prevent and minimize pathogen entry to the host target tissues by competitive inhibition.

[0045] According to some embodiments, the synthetic nanoparticle and/or microparticle is used for reducing the spread of SARS-CoV-2 virus or other viruses that causes a respiratory infection, diarrhea, common cold, influenzas or generally discomfort or a combination thereof.

[0046] According to some embodiments, the synthetic nanoparticle has a 3D- configuration generally matching the characteristics of the SARS-CoV-2 virus or other viruses that causes a respiratory infection, diarrhea, common cold, influenzas or generally discomfort or a combination thereof., in particular the particle is fabricated to a size of around 100 nm and coated with similar amino acids as the glycoprotein spikes or protruding proteins at the surface of the viral particle or similar molecules that mimic the surface of the viral envelope.

[0047] According to some embodiments, the synthetic nanoparticle resembles the SARS-CoV-2 virus, influenza viruses, rhinoviruses, common cold viruses and/or noroviruses or is optimized for competitive inhibition.

[0048] According to some embodiments, the synthetic carrier exhibits a modified particle morphology, size or surface properties to achieve increased affinity for the target receptor angiotensin converting enzyme 2 (ACE-2), compared with the SARS-CoV-2 virus, and/or other viruses that causes a respiratory infection, diarrhea, common cold, in particular for increasing the binding affinity for the specific receptor e.g., silicid sialic acid, histo-blood group antigens, ICAM-1, IGF1R blocking the internalization of the viral envelope more efficiently and potentially prolonging the gained viral protection.

[0049] According to some embodiments, the synthetic nanoparticle resembling the SARS-CoV-2 virus, influenza viruses, rhinoviruses, common cold viruses and/or noroviruses is adapted for personalized medicine.

[0050] According to some embodiments, the synthetic nanoparticle resembling the SARS-CoV-2 virus, influenza viruses, rhinoviruses, common cold viruses and/or noroviruses is loaded into or onto the nanoparticle for further enhancing the anti-viral properties.

[0051] According to some embodiments, the synthetic nanoparticle resembling the SARS-CoV-2 virus, influenza viruses, rhinoviruses, common cold viruses and/or noroviruses is loaded with vehicles or proteome inhibitors for efficiently delivering the compounds in the target tissues with minimal off-target effects.

[0052] According to some embodiments, the synthetic nanoparticle is decorated with molecules that have high affinity towards the SARS-CoV-2 virus or any other pathogen of interest such as influenza viruses, rhinoviruses, common cold viruses and/or noroviruses in order to bind and immobilize the infectious agent preventing or minimizing the potential risk of host entry.

[0053] According to some embodiments, the synthetic nanoparticle resembling the SARS-CoV-2 virus or any other pathogen such as influenza viruses, rhinoviruses, common cold viruses and/or noroviruses is coated or decorated with epitopes to be used as a vaccination at target cell populations.

[0054] According to some embodiments, the carrier is loaded, stored or dispersed in a device or vessel capable of on-demand release of the carrier to the end-user. [0055] According to some embodiments, the carrier system is loaded inside a dispenser such as an inhalation device, tablet, injectable substance, cream or ointment.

[0056] According to some embodiments, the man-made materials are used for immobilizing specific pathogens by adding the synthetic material in sanitation products and disinfectants.

[0057] According to some embodiments, a method for preventing or reducing pathogen binding to target areas of cell structures of a host comprises minimizing or reducing the spread of diverse pathogens by binding to the target molecule in the host body or binding to the infectious agent itself and potently inhibit the spread of the disease.

[0058] According to some embodiments, a synthetic carrier for use in a method of preventing or reducing pathogen binding to target areas of cell surfaces of a host, said carrier comprising biocompatible particles having a maximum size which, in at least one dimension, is in the nanometer or micrometer range, forming a core, and further having a functionalized surface capable of binding to said target areas of said cell surfaces so as to at least temporarily block said target areas to prevent or minimize pathogen binding and, thus, reducing the risk of the host contracting a disease caused by said pathogen.

[0059] According to some embodiments, the carrier comprises the capacity of eliciting a protective immunological reaction against said pathogen thus at least partially hindering the ability of the pathogen to bind to and enter the host, thereby lowering the risk of contracting the specific infectious agent and the related disease.

[0060] According to some embodiments, the carrier comprises a capacity of binding and encapsulating (e.g., is configured to bind and encapsulate) the pathogen, thus immobilizing the pathogens’ ability to bind and entry to the host lowering the risk of contracting the specific infectious agent.

[0061] According to some embodiments, the core or other base member or structure of the carrier is obtained by (e.g., manufactured and/or fabricated using) 3D printing, microfluidics, sol-gel method or other bottom-up and/or top-down method of fabrication.

[0062] According to some embodiments, the core material comprises organic or inorganic components, lipid droplets, amino acids, proteins, salts and minerals or other molecules.

[0063] According to some embodiments, the core material comprises inorganic silica nanoparticles, in particular mesoporous silica particles, such particles preferably having ordered mesostructures of pores that preferably are capable of being loaded with drugs. [0064] According to some embodiments, the core material is functionalized with substance selected from the group consisting of amino acids, epitopes, peptides, proteins or fragments of proteins, chemical agents, active pharmaceutical ingredients (API), organic or inorganic polymers or molecules and combinations thereof.

[0065] According to some embodiments, the core material is functionalized with substance selected from the group consisting of peptides, proteins such as antibodies, chemical agents, active pharmaceutical ingredients (API), organic or inorganic polymers or molecules and combinations thereof.

[0066] According to some embodiments, the carrier (e.g., with its functionalization) is used for specifically binding to receptors, proteins and macromolecules at the cellular level in order to prevent and minimize pathogen entry to the host target tissues by competitive inhibition.

[0067] According to some embodiments, a method of preventing or reducing a likelihood of pathogen binding to target areas of cell structures of a host comprises loading or otherwise adding one or more of the following to a carrier: drugs, API, molecules, peptides inside or onto the carrier system.

[0068] According to some embodiments, a carrier is used for targeted drug delivery of anti-pathogenic, anti-viral or anti-microbial compounds in order to decrease the growth of the pathogen, such as infectious agent.

[0069] According to some embodiments, a method of preventing or reducing pathogen binding to target areas of cell structures of a host comprises using synthetic nanoparticles and/or microparticles used for reducing the spread of S ARS-CoV-2 virus or other viruses that causes a respiratory infection, diarrhea, common cold, influenzas or generally discomfort or a combination thereof.

[0070] According to some embodiments, a method of preventing or reducing a likelihood of pathogen binding to target areas of cell structures of a host comprising using a synthetic nanoparticle that includes a 3D-configuration generally or substantially matching, at least in part, one or more of the SARS-CoV-2 virus or influenza viruses, rhinoviruses, common cold viruses and/or noroviruses, in particular the particle is fabricated to a size of around 100 nm and coated with similar amino acids as the glycoprotein spikes or other protruding proteins at the surface of the viral particle or similar molecules that mimic the surface of the viral envelope and thus binds to the same target receptor as the virus. [0071] According to some embodiments, the synthetic nanoparticle resembles the SARS-CoV-2 virus or influenza viruses, rhinoviruses, common cold viruses and/or noroviruses or is optimized for competitive inhibition.

[0072] According to some embodiments, the synthetic carrier exhibits a modified particle morphology, size or surface properties to achieve increased affinity for the target receptor angiotensin converting enzyme 2 (ACE-2) compared with the SARS-CoV-2 virus or influenza viruses, rhinoviruses, common cold viruses and/or noroviruses, in particular for increasing the binding affinity for the specific receptor e.g., silicid sialic acid, histo-blood group antigens, ICAM-1, IGF1R blocking the internalization of the viral envelope more efficiently and potentially prolonging the gained viral protection.

[0073] According to some embodiments, the synthetic nanoparticle resembling the SARS-CoV-2 virus, or influenza viruses, rhinoviruses, common cold viruses and/or noroviruses is adapted for personalized medicine.

[0074] According to some embodiments, the synthetic nanoparticle resembling the SARS-CoV-2 virus or influenza viruses, rhinoviruses, common cold viruses and/or noroviruses is loaded into or onto the nanoparticle for further enhancing the anti-viral properties.

[0075] According to some embodiments, the synthetic nanoparticle resembling the SARS-CoV-2 virus, or influenza viruses, rhinoviruses, common cold viruses and/or noroviruses is loaded with vehicles or proteome inhibitors for efficiently delivering the compounds in the target tissues with minimal off-target effects.

[0076] According to some embodiments, the synthetic nanoparticle is decorated with molecules that mimics naturally occurring protrusion of SARS-CoV-2 virus or any other pathogen of interest for example influenza viruses, rhinoviruses, common cold viruses and/or noroviruses in order to bind to target receptor of said host and thus preventing or minimizing the potential risk of the infectious agent entry to host.

[0077] According to some embodiments, the synthetic nanoparticle is decorated with molecules that have high affinity towards the SARS-CoV-2 virus or any other pathogen of interest for example influenza viruses, rhino viruses, common cold viruses and/or noroviruses in order to bind and immobilize the infectious agent preventing or minimizing the potential risk of host entry.

[0078] According to some embodiments, the synthetic nanoparticle resembling the SARS-CoV-2 virus or any other pathogen for example or influenza viruses, rhinoviruses, common cold viruses and/or noroviruses is coated or decorated with epitopes to be used as a vaccination at target cell populations making the administration potentially easier for the end user e.g., inhalation compared to intra muscular injection used in traditional vaccinations.

[0079] According to some embodiments, the carrier is loaded, stored or dispersed in a device or vessel capable of on-demand release of the carrier to the end-user.

[0080] According to some embodiments, the carrier system is loaded inside a dispenser such as an inhalation device, tablet, injectable substance, cream or ointment.

[0081] According to some embodiments, man-made materials (e.g., carriers) are used for immobilizing specific pathogens by adding the synthetic material in sanitation products and disinfectants.

[0082] According to some embodiments, a method of preventing or reducing a likelihood of pathogen binding to target areas of cell structures of a host comprises minimizing or otherwise reducing the spread of diverse pathogens by binding to the target molecule in the host body or binding to the infectious agent itself and potently inhibit the spread of the disease.

[0083] According to some embodiments, a method of producing a synthetic carrier comprises the steps of providing a core material (e.g., a nano- and/or micro-material including nanoparticles, microparticles or any other object as disclosed herein), coating or functionalizing the core material with molecules, polymers, amino acids, proteins, API, drugs or other material as disclosed herein, loading the object with compounds, molecules, drugs, API, DNA or RNA etc., and coating a second protective and/or functional layer on top of the object in particular for increasing its resistance that could be important in extreme environments such as the acidic environment in the stomach. In some embodiments, the method further comprises providing a small device, medical device, inhalation device or aerosol, sanitation product or consumer product that on-demand will release the containing synthetic material, particle or object for administration.

[0084] According to some embodiments, a method of preventing or reducing pathogen binding, in particular of preventing or reducing binding of SARS-CoV-2 or influenza viruses, rhinoviruses, common cold viruses and/or noroviruses and viral strains thereof, to target areas of cell surfaces of a host selected from mammals, comprises administering to the mammal a carrier comprising biocompatible particles having a maximum size in at least one dimension in the nanometer or micrometer range, forming a core, and further having a functionalized surface capable of binding to said target areas of said cell surfaces to at least temporarily block said target areas to prevent or minimize pathogen binding and thus, reducing the risk of the host contracting a disease caused by said pathogen. [0085] According to some embodiments, the carrier has the capacity of binding (e.g., is configured to bind, at least partially) and encapsulating (e.g., is configured to encapsulate, at least partially) the pathogen, thus immobilizing the ability of pathogens to bind and entry to the host lowering the risk of contracting the specific infectious agent.

[0086] According to some embodiments, the carrier is configured to at least partly hinder (e.g., prevent, slow, etc.) pathogens from binding and entering host cells, wherein the carrier is capable of binding to several said target areas of cell surfaces to at least temporarily and/or partially block viral entry, thereby giving the carrier dual targeting strategies and at least partially (e.g., partially, significantly, etc.) hindering the ability of one or more target pathogens to bind to and/or enter the host. In some embodiments, such carriers lower the risk of contracting the specific infectious agent.

[0087] According to some embodiments, the carrier is configured to (e.g., has the capacity to), at least partially, bind and/or encapsulate pathogens, thereby at least partially immobilizing the pathogens ability to bind and enter the host and capable of binding to said target areas of said cell surfaces to at least temporarily block viral entry, thus having dual targeting strategies thus significantly hinder the pathogens ability to bind and entry to the host lowering the risk of contracting the specific infectious agent.

[0088] According to some embodiments, the core structure of the carrier is obtained by 3D printing, microfluidics, sol-gel method or other bottom-up and/or top-down method of fabrication.

[0089] According to some embodiments, the core material comprises organic or inorganic components, lipid droplets, amino acids, proteins, salts and minerals or other molecules or wherein the core material comprises mesoporous silica nanoparticles, in particular mesoporous silica particles with ordered mesostructures of pores that preferably are capable of being loaded with drugs.

[0090] According to some embodiments, the core material is functionalized with substance selected from the group consisting of amino acid, epitopes, peptides, proteins and/or fragments of proteins, chemical agents, active pharmaceutical ingredients (API), organic or inorganic polymers or molecules and combinations thereof.

[0091] According to some embodiments, the core material is functionalized with substance selected from the group consisting of peptides, proteins such as antibodies, chemical agents, active pharmaceutical ingredients (API), organic or inorganic polymers or molecules and combinations thereof. [0092] According to some embodiments, the carrier is functionalized for specifically binding to receptors, proteins and macromolecules at the cellular level in order to prevent and minimize pathogen entry to the host target tissues by competitive inhibition.

[0093] According to some embodiments, the synthetic nanoparticle and/or microparticle is used for reducing the spread of SARS-CoV-2 virus or other viruses that causes a respiratory infection, diarrhea, common cold, influenzas or generally discomfort or a combination thereof.

[0094] According to some embodiments, the synthetic nanoparticle comprises a 3D-configuration generally matching the characteristics of the SARS-CoV-2 virus or other viruses that causes a respiratory infection, diarrhea, common cold, influenzas or generally discomfort or a combination thereof, in particular the particle is fabricated to a size of around 100 nm and coated with similar amino acids as the glycoprotein spikes at the surface of the viral particle or similar molecules that mimic the surface of the viral envelope.

[0095] According to some embodiments, the synthetic nanoparticle resembles, at least partially, one or more aspects and/or properties of the SARS-CoV-2 virus or other viruses that causes a respiratory infection, diarrhea, common cold, influenzas or generally discomfort or a combination thereof, or is optimized for competitive inhibition.

[0096] According to some embodiments, the synthetic carrier exhibits a modified particle morphology, size or surface properties to achieve increased affinity for the target receptor angiotensin converting enzyme 2 (ACE2) compared with the SARS-CoV-2 virus or other viruses that causes a respiratory infection, diarrhea, common cold, influenzas or generally discomfort or a combination thereof, in particular for increasing the binding affinity for the specific receptor e.g., silicid sialic acid, histo-blood group antigens, ICAM-1, IGF1R blocking the internalization of the viral envelope more efficiently and potentially prolonging the gained viral protection.

[0097] According to some embodiments, the carrier (e.g., the synthetic nanoparticle) resembles, at least in part, the SARS-CoV-2 virus or other viruses that causes a respiratory infection, diarrhea, common cold, influenzas or generally discomfort or a combination thereof. In some embodiments, the carrier is adapted for use as personalized medicine and/or targeted therapy.

[0098] According to some embodiments, the synthetic nanoparticle resembles, at least in part, the SARS-CoV-2 virus or other viruses that causes a respiratory infection, diarrhea, common cold, influenzas or generally discomfort or a combination thereof. In some embodiments, one or more components are added to the carrier (e.g., the nanoparticle) in order to further enhance anti-viral properties and/or any other desired properties or functions.

[0099] According to some embodiments, the synthetic nanoparticle that resembles, at least in part, the SARS-CoV-2 virus or other viruses that causes a respiratory infection, diarrhea, common cold, influenzas or generally discomfort or a combination thereof is loaded with vehicles or proteome inhibitors for efficiently delivering the compounds in the target tissues with minimal off-target effects.

[0100] According to some embodiments, the carrier (e.g., the synthetic nanoparticle) is decorated with molecules that have high affinity towards the SARS-CoV-2 virus or any other pathogen of interest for example influenzas, rhinoviruses and viruses causing respiratory infection in order to bind and immobilize the infectious agent preventing or minimizing the potential risk of host entry.

[0101] According to some embodiments, the synthetic nanoparticle that resembles, at least in part, the SARS-CoV-2 virus or any other pathogen for example influenzas, rhinoviruses and viruses causing respiratory infection, is coated or decorated with epitopes to be used as a vaccination at target cell populations.

[0102] According to some embodiments, the carrier is loaded, stored or dispersed in a device or vessel capable of on-demand release of the carrier to the end-user.

[0103] According to some embodiments, the carrier system is loaded inside a dispenser such as an inhalation device, tablet, injectable substance, cream or ointment.

[0104] According to some embodiments, the carriers or other man-made materials are used for immobilizing specific pathogens by adding the synthetic material in sanitation products and disinfectants.

[0105] According to some embodiments, a method for preventing or reducing a likelihood of pathogen binding to target areas of cell structures of a host comprises minimizing or reducing the spread of diverse pathogens by binding to the target molecule in the host body or binding to the infectious agent itself and potently inhibit the spread of the disease.

[0106] In some embodiments, a synthetic carrier for use in a method of preventing or reducing binding of a pathogen to target areas of cell structures of a host, said carrier comprising biocompatible particles having a maximum size in at least one dimension in the nanometer or micrometer range, forming a core, and further having a functionalized surface, which preferably mimics that of the pathogen capable of binding to said target areas of said cell surfaces to at least temporarily block said target areas to prevent or minimize pathogen binding and thus, reducing the risk of the host contracting a disease caused by said pathogen. [0107] In some embodiments, the pathogen is a coronavirus, in particular SARS- CoV-2 or viral strains derived thereof. In some embodiments, the cell structures are selected from ACE2 and TMPRSS2 receptors and combinations thereof.

[0108] In some embodiments, the synthetic carrier includes the capacity of binding to the pathogen’s co-receptors (e.g., high-density lipoprotein (HDL) scavenger receptor B type 1 (SR-B1)), thus at least partially hindering the ability of the pathogens to bind and enter the host. Advantageously, this can lower the risk of contracting the specific infectious agent and the disease resulting therefrom.

[0109] In some embodiments, the synthetic carrier includes the capacity of binding and encapsulating the pathogens co-receptors e.g., high-density lipoprotein (HDL) scavenger receptor B type 1 (SR-B1), thus immobilizing the pathogens’ ability to bind and entry to the host lowering the risk of contracting the specific infectious agent.

[0110] In some embodiments, the synthetic carrier comprises a core or other base structure that is manufactured using, at least in part, 3D printing, microfluidics, sol-gel method or other bottom-up and/or top-down method of fabrication, and wherein the core material comprises organic or inorganic components, lipid droplets, amino acids, proteins, salts and minerals or other molecules.

[0111] In some embodiments, the synthetic carrier for use in a method of preventing or reducing pathogen, in particular coronaviruses binding to target areas of cell structures of a host according to any one of the preceding embodiments, wherein the core material comprises inorganic silica nanoparticles, in particular mesoporous silica particles, such particles preferably having ordered mesostructures of pores that preferably are capable of being loaded with drugs.

[0112] In some embodiments, the synthetic carrier comprises a core that is functionalized with one or more substances selected from the group consisting of amino acids, epitopes, peptides, proteins and/or fragments of proteins, chemical agents, active pharmaceutical ingredients (API), organic or inorganic polymers or molecules and combinations thereof, and wherein said carrier with its functionalization is preferably used for specifically binding to receptors, proteins and macromolecules at the cellular level in order to prevent and minimize novel coronaviruses such as SARS-CoV-2 entry to the host target cells by competitive inhibition.

[0113] In some embodiments, the synthetic carrier comprises a core that is functionalized with substance selected from the group consisting of peptides, proteins such as antibodies, chemical agents, active pharmaceutical ingredients (API), organic or inorganic polymers or molecules and combinations thereof, and wherein said carrier with its functionalization is preferably used for specifically binding to receptors, proteins and macromolecules at the cellular level in order to prevent and minimize novel coronaviruses such as SARS-CoV-2 entry to the host target cells by competitive inhibition.

[0114] In some embodiments, a method of preventing or reducing a likelihood of coronaviruses binding to target areas of cell structures of a host comprises loading or otherwise providing drugs, API, molecules, peptides inside or onto the carrier system, wherein the carrier preferably comprises a functionalized and drug loaded carrier, said carrier being used for targeted drug delivery of anti-viral in order to decrease the replication of the virus inside the host cell.

[0115] In some embodiments, the synthetic carrier (e.g., nanoparticle and/or microparticle) is used for reducing the spread of SARS-CoV-2 or other coronaviruses strains and/or types derived from the SARS-CoV-2 that causes a respiratory infection, diarrhea, common cold, cytokine storm, death or generally discomfort or a combination thereof.

[0116] In some embodiments, the synthetic carrier includes a 3D-configuration generally matching the characteristics of the SARS-CoV-2 virus or future variants thereof, in particular the particle is fabricated to a size of around 100-120 nm and coated with similar amino acids as the glycoprotein spikes at the surface of the viral particle or similar molecules that mimic the surface of the viral envelope e.g. spike protein and thus binds to the same target receptor as the virus.

[0117] In some embodiments, the synthetic carrier (e.g., nanoparticle, microparticle, etc.) resembles, at least in part, the SARS-CoV-2 virus or is optimized for competitive inhibition, wherein preferably the synthetic carrier exhibits a modified particle morphology, size or surface properties to achieve increased affinity for the target receptor ACE2 and/or TMPRSS2, compared with the SARS-CoV-2 virus, in particular for increasing the binding affinity for the specific receptor blocking the internalization of the viral envelope more efficiently and potentially prolonging the gained viral protection.

[0118] In some embodiments, the synthetic carrier resembling the SARS-CoV-2 virus is adapted for personalized medicine. In some embodiments, the synthetic carrier resembling the SARS-CoV-2 virus is adapted for personalized medicine in the case of ACE2 receptor polymorphism or different animal host organisms for achieving receptor interaction.

[0119] In some embodiments, the synthetic carrier resembling the SARS-CoV-2 virus is loaded into or onto the nanoparticle for further enhancing the anti-viral properties, or wherein said synthetic nanoparticle resembling the SARS-CoV-2 virus is loaded with vehicles or proteome inhibitors for efficiently delivering the compounds in the target cell and tissues with minimal off-target effects.

[0120] In some embodiments, the synthetic carrier is loaded, stored or dispersed in a device or vessel capable of on-demand release of the carrier to the end-user, wherein said carrier system is preferably loaded inside a dispenser such as an inhalation device, tablet, injectable substance, cream or ointment.

[0121] In some embodiments, the synthetic carrier is a self-assembling recombinant protein-based nanoparticle construct, such as a SpyTag™/SpyCatcher™ system.

[0122] In some embodiments, a method of producing a synthetic carrier comprises the steps of: providing a core material, e.g., a nano- and/or micro-material including nanoparticles, microparticles or any other object as disclosed herein, coating or otherwise functionalizing the core material with molecules, polymers, amino acids, proteins, API, drugs or other material as disclosed herein, loading the object with compounds, molecules, drugs, API, DNA or RNA etc., coating a second protective and/or functional layer on top of the object in particular for increasing its resistance that could be important in extreme environments such as the acidic environment in the stomach, and providing a small device, medical device, inhalation device or aerosol, sanitation product or consumer product that on-demand will release the containing synthetic material, particle or object for administration.

[0123] According to some embodiments, forming a core using a microfluidics device, wherein the core comprises a mean size ranging from 100 nm to 500 nm; combining the core with at least one protrusion, wherein the protrusion is adapted to bind to a target area of a host; maintaining the core and the at least one protrusion to a temperature from about 3 °C to about 5 °C; and functionalizing the core by securing the at least one protrusion to the core.

[0124] According to some embodiments, further comprising adding an additive to the core, wherein the additive comprises an API, a drug, a protein, an amino acid, a lipid, a cholesterol molecule, a polymer, a monomer, an amino acid, a protein, a salt, a mineral, RNA, DNA, or any combination thereof; wherein functionalizing comprises incubating the mixture for about 2 hours to about 5 hours; and wherein the at least one protrusion is adapted to mimic an agent, the agent is a ligand, an antigen, an agonist, a pathogen or part thereof, a virus or part thereof, a bacterium or part thereof, a fungus or part thereof, a protein, a prion, a nucleic acid, an enzyme, a toxin, an allergen, a xenobiotic agent, or any combination thereof.

[0125] According to some embodiments, functionalizing comprises incubating the mixture for about 2 hours to about 5 hours. [0126] According to some embodiments, further comprising adding an additive to the core.

[0127] According to some embodiments, the additive comprises an API, a drug, a protein, an amino acid, a lipid, a cholesterol molecule, a polymer, a monomer, an amino acid, a protein, a salt, a mineral, RNA, DNA, or any combination thereof.

[0128] According to some embodiments, forming comprises vesiculating a raw material, the raw material comprising a lipid, a cholesterol molecule, a polymer, a monomer, an amino acid, a protein, a salt, a mineral, or any combination thereof.

[0129] According to some embodiments, the at least one protrusion is adapted to at least partially mimic an agent.

[0130] According to some embodiments, the agent is a ligand, an antigen, an agonist, a pathogen or part thereof, a virus or part thereof, a bacterium or part thereof, a fungus or part thereof, a protein, a prion, a nucleic acid, an enzyme, a toxin, an allergen, a xenobiotic agent, or any combination thereof.

[0131] According to some embodiments, the virus or part thereof comprises a coronavirus, an influenza virus, a rhinovirus, a norovirus, a respiratory syncytial virus (RSV), or any virus that impacts the respiratory system.

[0132] According to some embodiments, further comprising administering the carrier to the host.

[0133] According to some embodiments, administering comprises competitively inhibiting the agent from binding to a host cell.

[0134] According to some embodiments, competitively inhibiting comprises preventing entry of the agent into the host cell.

[0135] According to some embodiments, functionalizing comprises securing the core with a plurality of protrusions.

[0136] According to some embodiments, functionalizing comprises ligating the at least one protrusion to an outer surface of the core.

[0137] According to some embodiments, securing comprises ligating the at least one protrusion to an outer surface of the core.

[0138] According to some embodiments, ligating comprises covalent binding, non- covalent interactions, van der Waals forces, hydrophobic interactions, electrostatic interactions, or polar interactions.

[0139] According to some embodiments, the temperature is greater than 0 °C to less than about 10 °C. [0140] According to some embodiments, the target area of a host is a host cell.

[0141] According to some embodiments, the target comprises at least one receptor, target molecule, amino acid or nucleotide.

[0142] According to some embodiments, further comprising loading the carrier with at least one material.

[0143] According to some embodiments, the material comprises a drug, API, or other pharmaceutical composition.

[0144] According to some embodiments, the pharmaceutical composition comprises Celastrol, zinc, anti-viral compounds, Interferon-Gamma modulators, antibodies, proteins, or any combination thereof.

[0145] According to some embodiments, administering comprises using an inhalation device, an oral tablet, or an injection device.

[0146] According to some embodiments, the core comprises synthetic polymers.

[0147] According to some embodiments, the synthetic polymers comprise polyolefins, polyesters, polylactides, polycaprolactones, polyamides, polyimides, polynitriles, or any combination thereof.

[0148] According to some embodiments, the carrier is biocompatible.

[0149] According to some embodiments, forming comprises 3D printing, microfluidics, sol-gel methods, genetically engineering organisms to produce vesiculating biopolymers, or any combination thereof.

[0150] According to some embodiments, forming comprises casting the core from polydimethylsiloxane (PDMS) using soft lithography.

[0151] According to some embodiments, further comprising purifying the carrier, wherein purifying comprises dialyzing the carrier in a solution to remove an excess of the at least one protrusion that is uncoupled with the core.

[0152] According to some embodiments, a microfluidic device for forming a plurality of particles or cores, comprising a first inlet capillary having a first longitudinal axis and configured to receive a first component of particles or cores; a second inlet capillary having a second longitudinal axis, and configured to receive a second component of particles or cores; an outlet capillary configured to receive the first and second components, wherein the first longitudinal axis is angled relative to the second longitudinal axis to promote mixing of the first and second components.

[0153] According to some embodiments, the particles or cores are shaped by a distal tip of the outlet capillary, wherein a size distribution of the particles or cores is from 100 nm to 500 nm, and wherein the microfluidic device is configured to operate at from 0 °C to 10 °C.

[0154] According to some embodiments, further comprising a connector or luer in fluid communication with the first inlet capillary, the second inlet capillary, and the outlet capillary, wherein the connector or luer is configured to promote mixing of the first and second components.

[0155] According to some embodiments, the microfluidic device is configured for use on a tabletop.

[0156] According to some embodiments, the microfluidic device is constructed, at least in part, of glass.

[0157] According to some embodiments, the first inlet capillary is positioned substantially perpendicular to the second inlet capillary.

[0158] According to some embodiments, the first and second inlet capillaries comprise a flow velocity of from 0.5 mL/min to 3 mL/min.

[0159] According to some embodiments, the first and second inlets are configured to deliver a fluid to the outlet capillary at a pressure higher than atmospheric pressure.

[0160] According to some embodiments, the pressure is from 50 psi to 300 psi.

[0161] According to some embodiments, the first and second components are delivered to the first and second inlets, respectively, by a mechanical device.

[0162] According to some embodiments, the mechanical device is a syringe or an electromechanically powered device,

[0163] According to some embodiments, the first and second inlet are configured to receive a fluid from a first reservoir and a second reservoir, respectively.

[0164] According to some embodiments, the first and second reservoirs comprise the first and second components.

[0165] According to some embodiments, the first inlet and the second inlet comprise an inner diameter of 500 to 700 pm.

[0166] According to some embodiments, the first inlet and the second inlet comprise an outer diameter of 800 to 1200 pm.

[0167] According to some embodiments, the distal tip is polished and smooth.

[0168] According to some embodiments, further comprising external equipment, wherein the microfluidic device is configured to engage the external equipment. [0169] According to some embodiments, the external equipment comprises fluid pumps, compressed gas lines, computers, microscopes, electrical supply outlets, or any combination thereof.

[0170] According to some embodiments, a system for functionalizing a carrier, comprising the microfluidic device and a third reservoir, wherein the third reservoir comprises a functionalization medium.

[0171] According to some embodiments, the collection chamber comprises a volume of an aqueous solution.

[0172] According to some embodiments, the third reservoir is configured to engage the collection chamber.

[0173] According to some embodiments, the functionalization medium comprises a solution.

[0174] According to some embodiments, the functionalization medium is a solid.

[0175] According to some embodiments, the functionalization medium comprises at least one protrusion.

[0176] According to some embodiments, further comprising a pump.

[0177] According to some embodiments, further comprising a heat exchanger.

[0178] According to some embodiments, the heat exchanger is configured to maintain a temperature of the collection chamber of about 3 °C to about 5 °C.

[0179] According to some embodiments, the microfluidic device is configured to fabricate a core.

BRIEF DESCRIPTION OF THE DRAWINGS

[0180] FIG. 1 schematically illustrates one embodiment of a blocking mechanism created by a virus-like particles (VLP) against a viral (e.g., SARS-CoV-2) infection.

[0181] FIGS. 2 A to 2F schematically illustrate various aspects related to the precise control of a lipid nanoprecipitation process enabled by microfluidic devices and methods.

[0182] FIGS. 3 A to 31 illustrate various aspects related to the functionalization of SLN with Spike SI RBD programmed by streptavidin-biotin interaction.

[0183] FIGS. 4 A to 4D provide data to illustrate the excellent biocompatibility of SLN and VLPs manufactured using one or more of the embodiments disclosed herein.

[0184] FIGS. 5 A to 5 J illustrate various aspects related to the enhanced cell uptake of VLPs (e.g., such as VLP embodiments disclosed herein and/or fabricated, at least in part, by methods disclosed herein) mediated by the interaction with ACE2 receptor. [0185] FIGS. 6A to 6E illustrate various aspects related to the ability of VLPs to help prevent or reduce the likelihood of a SARS-CoV-2 infection through interaction with ACE2 in a dose-dependent manner.

[0186] FIG. 7 illustrates example intensity size distribution curves of VLPO and VLPD according to one embodiment.

[0187] FIGS. 8 A and 8B illustrate examples of long-term stability of fabricated particles, such as those disclosed herein or fabricated using, at least in part, one or more methods disclosed herein.

[0188] FIGS. 9 A to 9D illustrate various aspects related to the enhanced uptake kinetics of functionalized particles produced using at least one or more of the methods disclosed herein.

[0189] FIG. 10 illustrates example quantitative confocal microscopy images of particle endocytosis.

[0190] FIG. 11 illustrates example quantitative confocal microscopy images of particle endocytosis.

[0191] FIG. 12 is one example of a reconstructed confocal microscopy 3D-image of cells internalizing particles according to one embodiment.

[0192] FIG. 13 is a table of data related to RBD proteins in VLPO and VLPD.

[0193] FIG. 14 is one example of the microfluidics device for the fabrication of nanoparticles or microparticles.

[0194] FIG. 15 illustrates example quantitative laser scanning confocal microscopy images of particle endocytosis of Cy5.5-labelled SLN and VLPD by Caco-2 cells.

[0195] FIG. 16 illustrates an example schematic of the microfluidics device for the fabrication of nanoparticles or microparticles.

DETAILED DESCRIPTION

[0196] These and other features, aspects and advantages of the present application are described with reference to drawings of certain embodiments, which are intended to illustrate, but not to limit, any of the inventions disclosed herein. It is to be understood that these drawings are for the purpose of illustrating the various concepts disclosed herein and may not be to scale.

[0197] FIG. 1 schematically illustrates one embodiment of a blocking mechanism created by a virus-like particles (VLP) 10 against a viral (e.g., SARS-CoV-2) infection. As shown, in some embodiments, viral (e.g., the SARS-CoV-2) pathogenesis can be initialized by the binding of one or more portions or components (e.g., Spike SI receptor-binding domain (RBD)) of the virus to one or more receptors 20 (e.g., ACE2 receptors) of the host cell. The host-directed VLPs can bind to the ACE2 receptor, thereby at least partially blocking viral entry into host cells and helping prevent or reduce the likelihood of viral infection.

[0198] FIGS. 2 A to 2F schematically illustrate various aspects related to the precise control of a lipid nanoprecipitation process enabled by microfluidic devices, systems and methods. For example, FIGS. 2A and 2B schematically and generally illustrate the fabrication of solid lipid nanoparticles (SLN) under a bulk condition and a microfluidic condition, respectively. FIG. 2C illustrates one embodiment of the intensity size distribution curves of SLN prepared by bulk and microfluidic method. Data for average particle size, Polydispersity Index (PDI) and zeta potential according to one embodiment are illustrated graphically in FIG. 2D, FIG. 2E and FIG. 2F, respectively. For the embodiments related to FIGS. 2D to 2F, SLN was prepared by bulk and microfluidic methods in different batches. Statistical analysis was done using student’s t-test (n=10; microfluidic group vs. the corresponding bulk group; ***P < 0.001, n.s., not significant). The box plots indicate the minimum value, first quartile, median, third quartile and maximum value.

[0199] FIGS. 3 A to 31 illustrate various aspects related to the functionalization of SLN with Spike S 1 RBD programmed by streptavidin-biotin interaction. For example, FIG. 3 A schematically illustrates the functionalization of SLN with Spike SI RBD through streptavidinbiotin interaction. The influence of the functionalization with spike SI RBD on the particle size, PDI, and zeta potential are illustrated graphically in FIG. 3B, FIG. 3C and FIG. 3D, respectively. Statistical analysis was done using one-way ANOVA with post-hoc Bonferroni’s test (n=3; *P < 0.05, ** P < 0.01, n.s., not significant). Example transmission electron microscope images of SLN, VLPO and VLPD are illustrated in FIG. 3E. Example conjugation efficiency and mass fraction data of Spike S 1 RBD in VLPO and VLPD are illustrated in FIG. 3F and FIG. 3G, respectively. Statistical analysis was done using student’s t-test; n=3; VLPD group vs. the corresponding VLPO; n.s., not significant). An example histogram showing particle size distribution against nanoparticles concentration of VLPO and VLPD obtained by nanoparticles tracking analysis is illustrated in FIG. 3H. Example data for the number of Spike SI RBD molecules carried on the surface of VLPO and VLPD are illustrated in FIG. 31. Statistical analysis was done using student’s t-test (n=3; VLPD group vs the corresponding VLPO; **P < 0.01).

[0200] FIGS. 4A to 4D provide data to illustrate the excellent biocompatibility of SLN and VLPs manufactured using one or more of the embodiments disclosed herein. Example cytotoxicity data for (A) SLN, VLPO, and VLPD on A549, (B) A549-ACE2/T, (C) Calu-3 and (D) Caco-2 cells are provided on FIG. 4A, FIG. 4B, FIG. 4C and FIG. 4D, respectively.

[0201] FIGS. 5 A to 5 J illustrate various aspects related to the enhanced cell uptake of VLPs (e.g., such as VLP embodiments disclosed herein and/or fabricated, at least in part, by methods disclosed herein) mediated by the interaction with ACE2 receptor. Data related to cell uptake of FITC-labelled SLN, VLPO, and VLPD by (A) A549, (B) A549-ACE2/T, (C) Calu- 3 and (D) Caco-2 cells after different incubation times (0.5 hours, 1 hour, 3 hours, 6 hours and 24 hours) quantified by flow cytometry are provided in FIG. 5A, FIG. 5B, FIG. 5C and FIG. 5D, respectively. Statistical analysis was done using one-way ANOVA with post-hoc Bonferroni’s test (n=3; VLPO and VLPD groups were compared with the corresponding SLN group; *P < 0.05, **P < 0.01 and ***P < 0.001). Examples of laser scanning confocal microscope images revealing the cell uptake of FITC-labelled SLN, VLPO, and VLPD (20 pg/mL) by (i) A549 and (ii) A549-ACE2/T cells after 3 hours of treatment are illustrated, according to some embodiments, in FIGS. 5E and FIG. 5F, respectively. An example orthogonal view of laser scanning confocal microscope image revealing the subcellular localization of FITC-labelled VLPO (20 g/mL) in Calu-3 cells after 3 hours of treatment is illustrated in FIG. 5G. An example three-dimensional (3D) surface and dot reconstruction revealing the internalization of FITC-labelled VLPO (20 pg/mL) in Calu-3 cells after 3 hours of treatment is illustrated in FIG. 5H. In some embodiments, the wheat surface identifies cytomembrane stained with CellMask Deep Red, while the steel blue represents FITC-labelled VLPO. An example distance distribution of VLPO from the Calu-3 cell membrane in illustrated in FIG. 51. An example of apparent permeability coefficient of SLN, VLPO and VLPD (100 and 200 pg/mL) after 18 hours of incubation at 37 °C is illustrated in FIG. 5J. Statistical analysis was done using one-way ANOVA with post-hoc Bonferroni’s test (n=4; ***P < 0.001 , n.s., not significant).

[0202] FIGS. 6 A to 6E illustrate various aspects related to the ability of VLPs to help prevent or reduce the likelihood of a SARS-CoV-2 infection through interaction with ACE2 in a dose-dependent manner. One example of a pseudoviral infection assay is schematically illustrated in FIG. 6A. The effect of nanoparticles (SLN and VLPs) on blocking SARS-CoV-2 pseudoviral infection at a concentration of 20 pg/mL, 100 pg/mL and Spike SI RBD is shown in FIG. 6B, FIG. 6C and FIG. 6D, respectively. Statistical analysis was done using one-way ANOVA with post-hoc Bonferroni’s test (n=3; *P < 0.05, **P < 0.01 and ***P < 0.001, n.s., not significant). The effect of Spike SI RBD (Avi-His-Tag, 1 pg/pL) on the cell uptake of FITC-labelled VLPO and VLPD by A549-ACE2/T cells after 1 hour of incubation is illustrated in FIG. 6E. Statistical analysis was done using one-way ANOVA with post-hoc Bonferroni’s test (n=3; the 0.5, 1, 3 and 6 h groups were compared with the corresponding 0 h group, **P < 0.01).

[0203] FIG. 7 illustrates example intensity size distribution curves of VLPO and VLPD according to one embodiment.

[0204] FIGS. 8 A and 8B illustrate examples of long-term stability of fabricated particles, such as those disclosed herein or fabricated using, at least in part, one or more methods disclosed herein. Example normalized size and PDI data of SLN, VLPO and VLPD stored at 4°C, are illustrated graphically in FIG. 8A and FIG. 8B, respectively.

[0205] FIGS. 9 A to 9D illustrate various aspects related to the enhanced uptake kinetics of functionalized particles produced using at least one or more of the methods disclosed herein. FIG. 9 A, FIG. 9B, FIG. 9C and FIG. 9D provide example positive event data of A549, A549-ACE2/T, Calu-3 and Caco-2 cells, respectively, treated with FITC-labelled SLN, VLPO and VLPD for different time periods (0.5 hours, 1 hour, 3 hours, 6 hours and 24 hours) quantified by flow cytometry. Statistical analysis was done using one-way ANOVA with post-hoc Bonferroni’s test (n=3; VLPO and VLPD groups were compared with the corresponding SLN group; *P < 0.05, **P < 0.01 and ***P < 0.001).

[0206] FIG. 10 illustrates example quantitative confocal microscopy images of particle endocytosis. The depicted embodiments were validated with laser scanning confocal microscope images revealing the cell uptake of FITC-labelled SLN, VLPO and VLPD (20 pg/mL) by Calu-3 cells after 3 hours of incubation.

[0207] FIG. 11 illustrates example quantitative confocal microscopy images of particle endocytosis. The depicted embodiments were validated with laser scanning confocal microscope images revealing the cell uptake of FITC-labelled SLN, VLPO and VLPD (20 pg/mL) by Caco-2 cells after 3 hours incubation.

[0208] FIG. 12 is one example of a reconstructed confocal microscopy 3D-image of cells internalizing particles according to one embodiment. The reconstructed image of FIG. 12 uses confocal Z-stack images of a Calu-3 cell treated with FITC-labelled VLPO (20 pg/mL) for 3 hours.

[0209] FIG. 13 is a table of data related to RBD proteins in VLPO and VLPD at Day 0 and Day 110 validating the long storage possibilities of the functionalized particles fabricated, at least in part, by one or more of the methods or other embodiments disclosed herein. [0210] FIG. 14 is one example of the microfluidics device for the fabrication of nanoparticles or microparticles. The device includes two inlets and one outlet, as shown.

[0211] FIG. 15 illustrates example quantitative laser scanning confocal microscopy images of particle endocytosis of Cy5.5-labelled SLN and VLPD (20 pg/mL) by Caco-2 cells after 3 hours incubation.

[0212] FIG. 16 illustrates an example schematic of the microfluidics device for the fabrication of nanoparticles or microparticles. The microfluidics device includes a base 1610, on which a first inlet capillary 1620 is disposed parallel to the base 1610, and a second inlet capillary 1630 that is disposed perpendicular to the base 1610, and an outlet capillary 1640 disposed parallel to the base 1620. The outflow capillary 1660 slides over an outlet capillary 1640. The orifice 1640 is shaped such as to increasingly narrow circumferentially towards the distal end of the outlet capillary, thereby shaping the particles or cores flowing therethrough. The first inlet capillary 1620, the second inlet capillary 1630 and the outlet capillary 1640 are configured to engage a connector or luer 1650 in which one or more components are mixed. For example, a first component can flow from the first inlet capillary 1620 into the connector or luer 1650 and mix with the second component flowing into the connector or luer 1650 from the second inlet capillary 1630. The outlet capillary 1640 comprises a distal tip that is substantially smooth and uniform such as to uniformly shape the particle or core. The outflow capillary 1660 is configured to collect the particles or cores, and comprises a distal end 1670 from which the particles or cores can flow into a reservoir.

[0213] In some embodiments, the the first and second inlet capillaries comprise a flow velocity of 0.5, 1.0, 1.5, 2.0, 2.5, 3.0, 3.5, 4.0, 4.5, 5.0, 5.5, 6.0, 6.5, 7.0, 7.5, 8.0, 8.5, 9.0, 9.5, 10.0 mL/min, or any range of values therebetween. In some embodiments, the first and second inlets are configured to deliver a fluid to the outlet capillary at a pressure higher than atmospheric pressure. In some embodiments, the first and second inlets are configured to deliver a fluid to the outlet capillary at 10, 20, 30, 40, 50, 60, 70, 80, 90, 100, 1 10, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, 250, 260, 270, 280, 290, 300 psi, or any range of values therebetween. In some embodiments, the connector or luer is configured to promote mixing by promoting a non-laminar flow of fluids therein. In some embodiments, the outlet capillary is configured to promote a laminar flow therethrough.

[0214] In the present context, the term “around” means, when used in connection with numerical values, that a variation of ±25 %, in particular ±20 %, for example ±10 %, or ±5 %, of the exact value is included by a literal reading of that value. [0215] In the present context, the term “about” means, when used in connection with numerical values, that a variation of ±25 %, in particular ±20 %, for example ±10 %, or ±5 %, of the exact value is included by a literal reading of that value.

[0216] The term “polymer” is used herein in a broad sense and refers to materials, compounds, amino acids and proteins characterized by repeating moieties or units.

[0217] The term “functionalization” is used herein in a broad sense and refers to conjugating, coating, covalently and/or otherwise adding (e.g., allosterically adding) materials, compounds, drugs, amino acids and/or proteins to the synthetized particle (e.g., core) or object.

[0218] The term “biocompatible” refers herein to the ability of a material to perform with an appropriate host response in a specific application.

[0219] The term “host” refers herein to, but is not necessarily limited to, an individual mammal, in particular a human or an animal.

[0220] The terms “mimic,” “mimics,” “mimicking” and other conventional variations thereof refer herein to, but not limited to, a molecule, compound, agent or other object that is adapted to exhibit, at least partially, structural, functional, and/or reactive properties of another molecule, compound, agent or other object. By way of a non-limiting example, a molecule may be adapted to mimic the spike protein of the SARS-CoV-19 virus such that the molecule binds to ACE2 receptors of a human cell.

[0221] According to some embodiments, fabrication of nano and micro sized materials has received significant attention in the field of targeted drug delivery systems, vaccinations, formulations, adjuvants, composition materials and/or the like. As discussed herein, nano and micro sized particles and/or other materials can be delivered to a subject to prevent or reduce the likelihood of infection and/or spread of infection using competitive inhibition and/or immobilization. For example, the onset and/or spread of viral infections can be prevented, slowed and/or impacted in the favor of a subject when nanoparticles and/or microparticles are delivered to a subject such that such particles bind to sites (e.g., ACE2) of host cells that otherwise would provide an entry point into host cells by viruses. In some embodiments, such nano and/or micro particles are functionalized to assist with the binding of the particles to certain host cell sites, viruses, other pathogens or agents and/or the like.

[0222] Traditional fabrication methods restrict or limit the use and application of nano and micro materials, as their fabrication is not sufficiently accurate to produce consistently sized and/or shaped materials. In some circumstances, this is at least partly due to the lack of controllability of traditional fabrication methods available. Even though many of the used building-blocks of these materials are considered non-toxic, biocompatible and/or are “generally recognized as safe” (GRAS) by the FDA, nano and micro sized materials can have difficulties getting approval due to their size and shape variability. In some instances, this variability can give rise to toxicity, other unwanted effects or impacts and/or the like. As a result, the use of such materials has been relatively restricted or limited.

[0223] Generally, nano and micro sized particles or materials are produced using one or two techniques: (1) using a top-down manufacturing or fabrication approach or (2) using a bottom-up manufacturing or fabrication approach. In some arrangements, top-down approaches use building materials that have larger dimensions than the final, desired product, meaning that the material undergoes physical and/or other force or stress to reduce its size. This physical reduction in size can lead to surface imperfections, unevenness, fractures in the material and/or other negative results or conditions. Furthermore, the top-down method of fabrication is usually relatively cost inefficient as some of the building materials are wasted during production.

[0224] On the other hand, bottom-up methods of manufacture or fabrication typically begin by introducing smaller building blocks (e.g., materials in a medium/solution that is configured to react and transform, over time, to the desired final product). In certain cases, bottom-up methods provide a more cost- and material-efficient way of producing nano and/or micro sized materials. In addition, the consistency (e.g., in size, with respect to repeatability, etc.) of products produced using bottom-up methods is improved, meaning that in some circumstances, the final product has fewer defects and/or other quality control issues.

[0225] In some arrangements, traditional bottom-up methods of manufacture pose certain challenges, especially in scaling up the production, as variations in pH, compositions, temperature, mixing ratios, mixing speed and/or other factors may negatively influence the structure, size and/or surface features of the final product. There exist several different traditional bottom-up methods for the manufacture of particles, including but not limited to, the following: co-precipitation, template synthesis, sol-gel method (e.g., where the starting materials, for instance, the building blocks, often are copolymers, colloids, lipids, liquid crystals, inorganic materials and/or the like), etc.

[0226] Traditional methods of manufacturing also face significant challenges in functionalizing nano or micro sized materials, primarily due to the complex, multi-step processes required for achieving desired functionality. Methods often involve multiple crosslinking steps and other intricate chemical techniques, which must be carried out at elevated temperatures, often significantly above room temperature. The high-temperature conditions not only increase energy consumption but also raise concerns regarding the thermal stability of sensitive materials used in the functionalization process. As a result, the particles produced may suffer from reduced functional integrity and geometric uniformity, limiting their effectiveness for various applications. Moreover, the multiple crosslinking steps and other intricate chemical techniques often result in chemical residues that, e.g., reduce host cell viability when treated with said nano or micro sized materials. These limitations necessitate the development of more stable, scalable and cost-effective methods for producing functionalized nano and micro sized materials.

[0227] In certain circumstance, by improving the reproducibility and controllability of nano and micro sized materials it is possible to create new therapeutics, probiotics, over-the- counter products, therapeutics, theragnostic solutions and/or other products or strategies.

[0228] There are virtually endless functionalization possibilities by covalently attaching, adhering, saturating or binding (e.g., allosterically binding) molecules, polymers, proteins, amino acids, compounds and/or drugs onto the nano or micro sized materialsfor achieving active targeting. One of the major advantages of functionalizing a smaller molecule to a larger entity, e.g., proteins, epitopes, amino acid sequences or hydrophobic molecules to a nanomaterial, is to increase the combined object’s stability and/or solubility and/or to possibly minimize unwanted immunologic effects.

[0229] Described herein are carriers (e.g., fabricated nano or micro sized materials or other carriers), and methods of manufacturing and/or using the same. The carriers may be used for inhibiting or improving the ability to inhibit, at least partially, entry of certain pathogens or other unwanted organisms into the cells of a host organism, in particular, coronaviruses (e.g., SARS-CoV-2), influenzas, rhinoviruses, noroviruses, respiratory syncytial viruses and other viruses causing respiratory infection to the host organism. Accordingly, such nano or micro sized materials or other carriers can be advantageously used to limit, competitively inhibit, or reduce the replication and spread of a pathogen or other unwanted organism.

[0230] Embodiments disclosed herein have capabilities of carrying, delivering or providing anti-pathogenic pharmaceuticals or other materials, such as anti-viral compositions, in the carrier (e.g., nanomaterial) to reduce the replication and growth of the pathogen or other unwanted organism.

[0231] Embodiments disclosed herein have capabilities of carrying, delivering or providing drugs, pharmaceuticals, APIs, or other molecules, such as viscosity modulators, antihistamines and/or immunosuppressors, in the carrier (e.g., nanomaterial) to provide targeted treatment of a specific disease caused by the pathogen or other unwanted organism or agent.

[0232] Embodiments disclosed herein pertain to the fabrication of man-made (e.g., fabricated) carriers or materials (e.g., in the nano- and/or microscale) that are configured to at least partially saturate and bind to receptors, proteins and/or macromolecules of a host cell to reduce the likelihood (e.g., prevent) and reduce (e.g., minimize) pathogen (e.g., coronavirus) binding to host cell receptors and/or targeting tissues of the host. In some embodiments, the synthesized carrier can he stored and loaded onto a medical device capable of releasing the synthesized carrier system to specific tissue. For example, the medical device can administer the carrier on-demand and/or with specific dosages). Such medical devices or other devices or systems include, without limitation, inhalation devices, oral tablets, injection devices, lotions, creams and/or any other device, system or component, as desired or required.

[0233] In one embodiment, a synthetic carrier including a core. In some embodiments, the core can be functionalized. For example, a core can be functionalized such as to produce a carrier. In some embodiments, functionalization can include ligating, binding or otherwise coupling with the core one or more protrusion. In some embodiments, the protrusions are configured to bind one or more host cell surface molecule. For example, the protrusions can be specific to a host cell receptor to which a virus is adapted to specifically bind. By way of further example, the protrusions can be configured to specifically bind the ACE2 receptors displayed by human host cells, thereby selectively inhibiting binding and subsequent infection of said human host cells with SARS-CoV-2.

[0234] In some embodiments, a synthetic carrier, particle or object is configured to at least partially hinder or impede the spread of a pathogen or other unwanted organism by competitive inhibition and to deliver an API, drug or molecule to targeted host cells and/or host tissues with increased or improved efficacy. For example, the pathogen or other unwanted organism can be CO VID- 19. In some embodiments, the use of such carriers, particles or objects is configured to have few or minimal side effects in the host while simultaneously creating a hostile environment for pathogen or other unwanted organism. According to some arrangements, the carrier or particle is functionalized with, in one non-limiting example, hexapeptide resembling that of the RBD from SARS-CoV-2. This can, according to some embodiments, allow high binding affinity to the ACE2 receptor at the lining of the respiratory system, thus inhibiting host infection by SARS-CoV-2. In some embodiments, competitive inhibition of a pathogen or other unwanted organism, e.g., a virus, eliminates risk of adaptive mutation of the pathogen or other unwanted organism by preventing reproduction of, e.g., the virus. This may be especially significant, for instance, in light of the impactful SARS-CoV-2 mutations that have appeared starting in 2021 and beyond, which have and will have a significant impact on the state of world health. In some embodiment, alternatively or simultaneously, ACE2 receptor binding moieties or antibodies designed to bind and immobilize the virus at specific sites can be used.

[0235] Therefore, in some embodiments, as noted above and discussed in greater detail herein, carriers (e.g., particles, obstacles, etc.) are configured to prevent or reduce the likelihood of infection by pathogens using one or more principles or mechanisms. For example, in some arrangements, the carriers are sized, shaped and otherwise configured to prevent or reduce the likelihood of pathogen infection by competitive inhibition (e.g., blocking receptors).

[0236] In another aspect or embodiment, a carrier is configured to deliver “cargo” (e.g., component, material, other content, etc.) to one or more targeted cell populations. For example, in some arrangements, the carrier comprises a core that is configured to be loaded or otherwise provided with one or more of the following: a drug, API and/or other molecule or material, etc. Such substances that are strategically delivered to a targeted area of the subject’s anatomy via a carrier can be targeted with higher efficacy to specific cells and tissues using, for example, functionalization. For example, functionalization can include ligating, bonding, or otherwise coupling protrusions to the core, where the protrusions are capable of binding to host cell structures such as receptors, thereby facilitating carrier uptake by the cells and thereby enabling targeted therapeutics. Thus, potentially, the therapeutic effect of the drug can be improved, increased or otherwise enhanced, e.g., by accumulating the local dosage in specific cells, reducing side-effects of the drug, e.g., by reducing off-target effect in unwanted cells and/or the like.

[0237] In some embodiments, as noted above and discussed in greater detail herein, carriers can include protrusions that are configured to improve a therapeutic effect of the loaded material(s) that can help alleviate or improve the symptoms, progression, discomfort and/or other undesirable impacts associate with a specific disease or disorder. For example, the carriers can be functionalized with protrusions that are functionally identical or sufficiently similar to binding molecules of pathogens, other unwanted organisms, viruses or other agents. In some embodiments, the protrusions are ligated, bound or otherwise coupled with a core or base portion of the carrier. In some embodiments, the disease causing agent can be, for example and without limitation, one or more of the following: an infectious agent, chemical agent, a pathogen, a virus, a bacterium, a parasite, an antigen, a protein, a prion, a mold, a fungus, a toxin, a poison or an allergen, electromagnetic radiation, carcinogens, molecules, hormones, inflammation, enzymes, replication machinery, DNA repair machinery, another agent or material and/or the like.

[0238] In some embodiments, the carrier is loaded with or otherwise includes an API, drug, mineral, molecule and/or other material, that is intended to be delivered to target cells and tissues with increased efficacy and with minimal or reduced side effects. In some embodiments, this is accomplished by creating an environment where the disease progression halters or at least partly slows or stops. As noted herein, in some arrangements, the size of the carrier (e.g., the particle or object) is similar or substantially similar to the size of the virus or other pathogen being targeted. In some embodiments, the cross-sectional dimensions of the carrier can be about 50% to about 200%, about 50% to about 100%, about 50% to about 150%, about 50% to about 200%, about 100% to about 150%, about 150% to about 200%, or any range of values therebetween, of the cross-sectional dimension of a pathogen, unwanted organism, virus, or other agent for which the carrier is configured to inhibit and/or impede. For example, a diameter or other cross-sectional dimension of the carrier can be 50% to 200% of the diameter or other cross-sectional dimension of influenza or other targeted virus or pathogen.

[0239] In some embodiments, the carrier is loaded with or otherwise includes a cargo. For example, the cargo can include an API, a drug, a molecule and/or other material to be delivered to target cells and tissues. In some embodiments, loading the carrier with the cargo can increase efficacy and reduced side effects of the cargo, while creating a hostile environment for the pathogen, other unwanted organism, virus or other agents. However, in other arrangements, the carrier does not contain a cargo. In other words, for any of the embodiments disclosed herein, a carrier does not include components and/or other materials that are intended to be delivered a site within the host. Even in such embodiments, carriers or particles can be configured to reduce the likelihood of infection. For example, the carrier alone can be adapted to prevent or reduce the likelihood of a virus from binding to and infecting targeted cells of the host, which can be accomplished by, at least in part, blocking receptors (or other binding sites or portions) of host cells. In some embodiments, the carrier can be functionalized with, for example and without limitation, protein fragments resembling HA and NA binding moieties. In such embodiments, high binding affinity to the host sialic acid receptors at the lining of the respiratory system is facilitated, thereby blocking a route (e.g., the primary route) of infection. Additionally, in some embodiments, the protrusions can be configured to elect an immune response against said pathogen.

[0240] In some embodiments, a carrier can be adapted to to hinder the spread of the major causative agent of the common cold by competitive inhibition. For example, the carrier can be functionalized such that it is adapted to bind host cell receptors to which a Rhinovirus binds. In some embodiments, the appropriate API, drug or molecule is delivered to target cells and tissues with increased efficacy and with minimal or reduced side effects while creating a hostile environment for the virus. The mimetic particle can be functionalized with, for example, but not limited to, VP1 and VP2 capsid protein allowing high binding affinity to the ICAM-1 and other related receptors at the lining of the respiratory system. Thus, the primary route of infection can be, at least partially, blocked. Additionally, in some embodiments, the protrusions are at least partially capable of electing or adapted to elect an immune response against said pathogen.

[0241] In other embodiments, a carrier mimics or substantially mimics (or is configured to mimic or substantially mimic), at least approximately or substantially, respiratory syncytial virus (RSV) in order to hinder, at least partially, the spread of influenza or the “flu,” a respiratory disease, by competitive inhibition. In some embodiments, with the use of such carriers, the appropriate (e.g., desired, required, etc.) API, drug or molecule is delivered to target cells and tissues with increased efficacy and with minimal or reduced side effects while creating a hostile environment for the virus. In some embodiments, the carrier (e.g., mimetic particle or object) is functionalized with, for example and without limitation, receptor attachment protruding glycoprotein (G) allowing high binding affinity to the IGF1R receptor at the lining of the respiratory system. Accordingly, such a carrier can be configured to block, at least partially, the primary route of infection and/or slow or stop the progression of infection. Additionally, in some embodiments, the protrusions are configured to selectively elect an immune response against a target pathogen or group of pathogens and/or other disease-causing agents or conditions.

[0242] In another aspect, a carrier (e.g., a synthetic particle or object) mimics or is configured or adapted to mimic), at least partially, Noroviruses. In such embodiments, the carrier can hinder, at least partially, the spread of “stomach flu” a gastroenteritis disease by competitive inhibition. In some embodiments, with the use of such carriers, the appropriate (e.g., desired, required, etc.) API, drug or molecule is delivered to the target cells and tissues with increased efficacy and with minimal or reduced side effects while creating a hostile environment for the virus. The carrier (e.g., the envisioned mimetic particle) functionalized with, for example and without limitation, VP1 containing the P2 subdomain allowing high binding affinity to the including sialic acid and histo-blood group antigens at the lining of the respiratory system. Thus, the carriers can block, at least partially, the primary route of infection. Additionally, in some embodiments, the protrusions are at least partially capable of electing or configured to elect an immune response against said pathogen.

[0243] In some embodiments, carriers can be further functionalized with protrusions on the surface of a core or particle (e.g., micro and/or nano-particle) of a carrier for targeting, blocking, immobilization and/or other purpose(s). Functionalization can include the addition of multiple (e.g., 2, 3, 4 different, more than 4 different, etc.) proteins resembling, at least partially, diverse different pathogens. Thus, by way of an example, the lipid particle (e.g., carrier, nanomaterial, micromaterial, etc.) can be loaded with (or supplemented with) one or more active pharmaceutical ingredients (API), such as, e.g., Celastrol, Zinc, anti-viral compounds, Interferon-Gamma modulators, antibodies, proteins etc., and then used in inhalation devices, oral tablets or injectables, or other devices or tools of administering the carriers, to name just a few. Such nanomaterials and/or micromaterials can be used to hinder, at least partially, the entry of novel coronaviruses, and/or hinder multiple pathogens within host cells, thereby reducing or minimizing the spreading of the disease.

[0244] Embodiments disclosed herein allow for decreasing the risk of a pathogen or pathogens, such as coronaviruses, influenzas, rhinoviruses, other viruses causing respiratory infection (e.g., SARS-CoV-2), entering its host for a temporary or prolonged duration. Accordingly, such embodiments can advantageously give a targeted treatment for the specific disease caused by the infectious agent.

[0245] According to some embodiments, a synthetic carrier disclosed herein and/or manufactured, at least in part, using methods and/or other arrangements disclosed herein, comprises biocompatible particles having a maximum size in at least one dimension in the nanometer or micrometer range. In some embodiments, said maximum size in at least one dimension is 10 nanometers to 500 nanometers, 10 nanometers to 50 nanometers, 10 nanometers to 100 nanometers, 50 nanometers to 100 nanometers, 1 nanometer to 100 nanometers, 100 nanometers to 200 nanometers, 200 nanometers to 300 nanometers, 300 nanometers to 400 nanometers, 400 nanometers to 500 nanometers, 200 nanometers to 400 nanometers, or any range of values therebetween. In one preferred embodiment, the maximum size in at least one dimension is 10 nanometer to 500 nanometers. In some embodiments, such carriers form or include a core and include a functionalized surface capable of binding to target areas of cell surfaces of a host. Advantageously, such binding can at least temporarily block the target areas to prevent or minimize pathogens (e.g., influenzas, rhinoviruses, coronaviruses including but not limited to SARS-CoV-2, other viruses causing respiratory infection, thereby reducing the risk of the host contracting the disease caused by the pathogen (e.g., the CO VID- 19 disease, diarrhea, respiratory infections, common cold, etc.).

[0246] As schematically illustrated in FIG. 1, the carrier includes a core and a plurality of surface features related to the core. As disclosed herein, the surface features can include protrusions that resemble or mimic, at least partially, spike proteins or other protrusions or features of a target virus or other pathogen. With continued reference to FIG. 1 , the carrier can be “loaded” or otherwise provided with one or more materials or other substances (e.g., APIs, other pharmaceuticals or agents, etc.). As disclosed herein, such materials can be delivered by the carrier to or near the site of a targeted virus or other pathogen for improved treatment (e.g., therapeutic treatment, infection prevention or mitigation, etc.).

[0247] In some embodiments, the synthetic carrier comprises a “nano” material which can be of nano- or micrometer or larger size. In some arrangements, the synthetic carrier has a size in at least one dimension which is in the nanometer scale. In some arrangements, the synthetic carrier has a size in at least one dimension which is in the micrometer scale. In other embodiments, the carrier has a size in at least one dimension that is outside the nanometer or micrometer range, as desired or required. For instance, the carrier can have a size in at least one dimension which is smaller than one nanometer (e.g., in the picometer range or smaller) or greater than one millimeter, depending on the targeted pathogen or other factors. The nanomaterial or other synthetic carrier can be formed as a particle, spheroid, cubical, cigarshaped, elongated, triangle, sharp and pointy, a sheet and film and/or any configuration or shape.

[0248] In some embodiments, the synthetic carrier has a maximum size in at least one dimension which is smaller than 2500 pm (e.g., less than 2500 pm, less than 2000 pm, less than 1500 pm, less than 1000 pm, less than 500 pm, less than 100 pm, less than 50 pm, less than values between the foregoing, etc.). In one embodiment, the material has a maximum size in at least one dimension which is smaller than 10 pm (e.g., less than 10 pm, less than 5 pm, less than 1 pm, less than values between the foregoing, etc.). In one embodiment, the material in particular nanomaterial has a maximum size in at least one dimension which is smaller than 1000 nm, in particular smaller than around 500 nm or around 100 nm or smaller than around lOnm or smaller than around 0.2nm.

[0249] In one embodiment, the synthetic carrier is biocompatible. For example, according to some arrangements, such a material is configured to cause no reaction or only a minor unwanted reaction in the end-user (e.g., toxicity, off-target effects, etc.). [0250] Generally, in some embodiments, the carriers disclosed herein are synthetic, which is used interchangeably with “synthesized” to denote that they are man-made or nonnatural.

[0251] Embodiments of the carriers comprise organic or inorganic materials, protein based, ferritin protein particles, lipid droplets, micelles, solid lipids, or any combination of these.

[0252] The synthetic material can be selected from inorganic and organic, monomeric and polymeric materials capable of forming biocompatible nano- or micro-sized particles as explained herein.

[0253] Examples of materials for the carriers comprise one or more of the following: synthetic polymers (e.g., thermoplastic or thermosetting materials, such as polyolefins, polyesters, including biodegradable polyesters (e.g., polylactides, polycaprolactones, etc.), polyamides, polyimides, polynitriles, etc.). Further non-limiting examples of possible materials include, for example and without limitation, silica, polysiloxanes, silicone materials which optionally may contain organic and metal residues, and/or the like. In some embodiments, silica particles are preferred, but not in all embodiments.

[0254] In one embodiment, the material, which forms the core structure of the carrier, is manufactured or otherwise obtained using one or more of the following: 3D printing, microfluidics, sol-gel methods (e.g., bottom-up methods or top-down methods of fabrication), genetically engineered organism producing specific proteins or amino acids that can either selfassembly such as ferritin protein particles or conjugate to larger entities any other method or technique, and/or combinations thereof.

[0255] In one embodiment, the core material comprises one or more materials, such as, for example, mesoporous silica nanoparticles with ordered mesostructures of pores. Such pores can be loaded with different drugs. The most common methods for drug loading to particles is either by physical adsorption using a highly saturated drug solution (e.g., a hydrophobic solvent such as cyclohexane with a hydrophobic drug) or an aqueous solution for water-soluble drugs. In some embodiments, loading further includes covalently conjugating the molecule to the particle surface using, for example, thiol chemistry and/or attracting the cargo molecule by having a different charge than the particle (e.g., particles having a positive charge which will allow loading of negatively charged drug, RNA/DNA molecules).

[0256] The carriers (e.g., particles or objects) disclosed herein can be synthetized in various sizes and shapes. In one embodiment, the material forming the core of the carrier contains pores with diameters between 1 and 75 nm, such as, for example, 2 to 50 nm, 2.5 to 30 nm, 2 to 5.5 nm, other values or ranges within the foregoing. In some embodiments, determining the hydrodynamic size using dynamic light scattering (DLS) makes it possible to confirm redispersibility of particle. In some arrangements, the morphology and particle diameter can be measured by either scanning electron microscope (SEM) or transmission electron microscopy (TEM). In order to determine surface area, pore size and pore volume, N2-sorption measurements can be used. The size and volume of the of the mesopores can be detected using small angle X-ray (S AXRD), according to some embodiments. The drug loading is, in some embodiments, measured by Thermogravi metric analysis (TGA) and alternatively or additionally measured by UV/vis spectroscopy measurements at a wavelength of 425 nm. Any alternative method or technology of forming the carriers and/or determining measurements can be used, either in addition to or in lieu of those disclosed herein, as desired or required.

[0257] In one embodiment, the core material comprises mesoporous silica nanoparticles (MSNs).

[0258] In one embodiment, the material compromises a nanoparticle core with coated targeting ligands with a possibility of (or configured to allow for) loading the particle with API, drugs, molecules, proteins and amino acids, RNA or DNA and compounds of interest.

[0259] In one embodiment, the material compromises a nanoparticle core and/or microparticle core with coated and/or functionalized targeting ligands with a possibility of (or configured to allow for) loading the particle into or onto with API, drugs, molecules, proteins and amino acids, RNA or DNA and compounds of interest.

[0260] Thus, in one embodiment, a nano and/or micro sized particle for example solid lipid particle (e.g., palmitate-based or stearylamine and the matrix lipid Compritol) having a net positive charge can be decorated/coated with negative molecules, such as RNA or DNA encoding for example interferon gamma for targeted delivery.

[0261] Thus, in one embodiment, the nano and/ or micro material compromises a core particle or object functionalized with targeting moieties, drugs, amino acids, protein or any combination thereof, such as hybrid materials containing but not limited to 1 ,2-Dioleoyl- 3 -trimethylammonium propane (DOTAP), Cholesterol (Choi), Dioleoylphosphatidylethanolamine (DOPE) and/or l,2-Distearoyl-sn-glycero-3- phosphorylethanolamine (DSPE), polyethylene glycol (PEG) (e.g. DOTAP:Chol:DOPE:PEG or DOTAP:Chol:DSPE:PEG) loaded with, for example and without limitation, RNA and/or DNA. In some embodiments, the object is preferably loaded with an active substance, drug or API.

[0262] In some embodiments of the present application and the technologies disclosed herein, two ways of synthetizing nanomaterials or other carriers are in particular employed. These include the top-down and the bottom-up approach or hybrid approach where some of the particle components are done with one approach and another component with another approach. In other embodiments, however, carriers can be synthesized or otherwise manufactured using other methods or approaches, as desired or required.

[0263] In the top-down approach, for example, the building materials have larger dimensions than the final product, which means that the materials undergo physical stresses, such as, e.g., grinding, milling etc., in order to be reduced in size. This process can lead to surface imperfections that could give rise to some variations in the final product that affect particle distribution in the host and binding kinetics.

[0264] In some embodiments, the bottom-up method starts by using smaller building blocks in solution transforming gradually to the final product, which can provide a more cost-efficient way of producing nanomaterials and/or micromaterials. Common bottom- up methods include, for example, co-precipitation, template synthesis and sol-gel method where the building blocks are often copolymers, colloids and liquid crystals and selfassembling components such as ferritin protein particles.

[0265] According to some embodiments, microfluidics can be used as a bottom-up method for fabricating nano or micro sized materials. Certain benefits can be obtained by using microfluidics as the design of the device. The device (e.g., microfluidic chip) can take several manifestations, materials and combinations and the inlets (e.g., channels inside the chip) can be made to different diameters in order to produce materials at a certain size and morphology. Other advantages of using microfluidics include, by way of example and without limitation, uniformly producing nano or micro sized materials (e.g., with narrow size distribution of particles). Another advantage of using microfluidics is the scalability potential as micro pumps can be used for creating constant input of building materials inside the chip allowing for relatively large-scale fabrication of materials.

[0266] In other embodiments, microfluidics is used as a combination or as part of a larger fabrication processes (e.g., one that include different steps). For example, the core material can be produced using, at least in part, a microfluidic device by bottom-up methods. Some elements can be introduced to (e.g., into, within, along an interior, along an exterior, etc.) the core materials that have been fabricated using one or more other methods and/or technologies (e.g., top-down methods). Alternatively, producing first components to be used as at least partly in the core using a top-down method to be then incorporated in the microfluidics fabrication producing hybrid materials that have elements produced in different ways.

[0267] In some embodiments, a microfluidic device (e.g., a microfluidic chip) is configured to produce, fabricate, synthetize and/or otherwise make nano or micro sized materials. The microfluidic device can be fabricated, arranged and/or otherwise configured by coaxially aligning and assembling of two capillaries (e.g., two glass capillaries). In some embodiments, an inner capillary with inner diameter (ID) of, for example and without limitation, 500 to 600 pm (e.g., 580, 500 to 520, 520 to 540, 540 to 560, 560 to 580, 580 to 600 pm, values between the foregoing ranges and/or values, etc.) and outer diameter (OD) of, for example and without limitation, 800 to 1200 pm (e.g., 1000, 800 to 900, 900 to 1000, 950 to 1050, 1000 to 1100, 1100 to 1200 pm, values between the foregoing ranges and/or values, etc.).

[0268] In some embodiments, the inner capillary is tapered (e.g., using a micropipette puller or similar device or technology). The tip (e.g., fine tip) can be polished or otherwise make smooth. In some arrangements, its diameter is further enlarged to, for example, around 100 pm (e.g., 100, 50 to 150, 90 to 110, 100 to 105, 100 to 1 10, 95 to 105, 95 to 100, 90 to 100, 80 to 120, 60 to 140, 70 to 130 pm values between the foregoing ranges and/or values, etc.). An inner capillary can be coaxially inserted into the outer cylindrical capillary with an ID of, for example and without limitation, 1000 to 1200 pm (e.g., 1120 pm) and an OD of, for example and without limitation, 1400 to 1600 pm (e.g., 1500 pm). In some embodiments, the diameter or cross-sectional dimension of the inner capillary is 10% to 200% (e.g., 10-200, 50-200, 50-100, 50-150, 50-200, 100-150, 150-200%, values and ranges between the foregoing, etc.) of the diameter or cross-sectional dimension of the foregoing embodiments describing the inner diameter and /or outer diameter.

[0269] According to some embodiments, the capillaries can be fixed or otherwise positioned on a glass slide and sealed, at least partially, employing, for example and without exclusion or limitation, a transparent epoxy resin to produce solid lipid nanoparticles (SLN). In some arrangements, the building material for the SLN can include, among other things, 1,2- dioleoyl-3-trimethylammoniumpropane (DOTAP), Cholesterol (Choi), dioleoylphosphatidylethanolamine (DOPE) and/or distearoyl phosphoethanolaminepolyethylene glycol-biotin (DSPE-PEG-Biotin).

[0270] In some embodiments, the microfluidic device can be cast from polydimethylsiloxane (PDMS) using soft lithography. In other embodiments, microfluidic device can be produced from thermoplastics (including, but not limited to, polystyrene, polycarbonate, and cyclic olefin copolymer) and silicon. In some embodiments, chip fabrication includes one or more of various technologies, such as hot embossing, laser cutting, wax printing for paper-based devices, 3D printing, or similar technologies.

[0271] In some embodiments, the microfluidic device can be configured to operate with external equipment. For example, the microfluidic device can be configured to operate with fluid pumps, compressed gas lines, computers, microscopes, electrical supply outlets, or similar external equipment.

[0272] In some embodiments, various chip geometries can be employed to fabricate nano- or micro-sized materials with different structures. For example, geometries such as coflow, flow focusing, T-junction, and Y-junction, and their different combinations, such as coflow combing flow focusing and co-flow combing co-flow, can be used to fabricate nano or micro-sized materials.

[0273] In some embodiments, therapeutics, including biomacromolecules, hydrophobic small molecules, hydrophilic small molecules, and similar compounds, can be loaded into the nano or micro sized materials during the microfluidic fabrication process through coprecipitation, sequential precipitation, emulsion method, and like processes.

[0274] According to some embodiments, the conjugation, functionalization, binding and/or coating of the carrier or other nano and/or micro sized materials (e.g., core) can be performed using click chemistry. For instance, streptavidin coated and/or bound protrusion(s) (e.g., SARS-CoV-2 Spike SI) can react with biotin containing materials, such as SLN in order to produce solid lipid nanoparticles. Such nanoparticles can include the original and/or variant (e.g., delta) form of SARS-CoV-2 Spike SI receptor binding domain (RBD), Avi-His-Tag, biotin-labeled). The conjugation procedure can further include adding protein protrusion (e.g., in a dropwise or other fashion) to an aqueous solution containing streptavidin. The mixture can be stirred for several hours (e.g., 1 to 2 hours, 2 to 3 hours, 3 to 5 hours, 1 to 10 hours, 1 to 10 hours, 1 to 12 hours, 1 to 24 hours, hour values between the foregoing, etc.) at room temperature (e.g., 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 15 to 25, 20 to 22, 18 to 22, 20 to 24, 19 to 21, 20 to 25 °C, temperatures between the foregoing values and ranges, etc.). In some embodiments, the mixture is stirred for 2 hours or less at a temperature of approximately 4 °C (e.g., 2 to 6, 3 to 5, 4, 0 to 5, 0 to 10, 0.1 to 5, 0.1 to 10, 0.1 to 20 °C, values between the foregoing values or ranges, etc.). In some embodiments, the mixture is maintained at a temperature above the freezing point of water (e.g., 0 °C), e.g., under normal atmospheric pressure.

[0275] According to some embodiments, to remove unconjugated protrusions from the carrier, a dialysis procedure can be performed (e.g., for a duration of several hours, preferably around and/or more than 4 hours, more preferably around and/or more than 12 hours, even more preferably around and/or 24 hours, even more preferably around and/or 48 hours), as desired or required.

[0276] Another embodiment where the SLN can or other or nano or micro material contain a tracer into or onto, a fluorescent molecule. For example and without limitation, fluorescein isothiocyanate (FITC) labelled SLN (FITC-SLN) can be prepared by changing part of the of DOTAP with Distearoyl phosphoethanolamine-polyethylene glycol-fluorescein isothiocyanate (DSPE-PEG-FITC).

[0277] Another embodiment where the SLN or other nano or micro material can contain a tracer inside the composition (part of the composition) of the core. For example, and without limitation, Cyanine 5.5 (Cy 5.5) labelled SLN (Cy 5.5-SLN) can be prepared by changing part of the of DOTAP with Distearoyl phosphoethanolamine-polyethylene glycol- Cyanine 5.5 (DSPE-PEG- Cy 5.5).

[0278] The carrier or particle system comprising of a core and functionalization can be characterized, in some arrangements, using Scanning electron microscopy and/or electron microscopy to confirm the size, monodispersity, morphology and non-agglomerated state of the particles. In some embodiments, to find (e.g., accurately, approximately) the amount of drug loading in the particle if the drug is fluorescent, particles can be dispersed in ethanol for complete drug elution. The concentration of drug can be determined by UV/vis spectroscopy measurements at a wavelength of 425 nm, for example with Celastrol. In some embodiments, from such measurements, the drug loading amount can be calculated or approximated. The mesoscopic ordering of the particles can be detected by powder-XRD using a Kratky compact small-angle system or similar X-ray diffraction (XRD) methods. In some embodiments, the hydrodynamic size of the particles can be determined by dynamic light scattering, and the mesoporosity by nitrogen sorption measurements. Thermogravimetric analysis can be used in order to estimate the amount of Polyethyleneimines (PEI), sugar motifs, folic acid (FA) or methotrexate (MTX) or other organic content functionalized to the particle. In some embodiments, thermogravimetric analysis can be used to estimate the amount of organic contact or other molecule and/or drug content functionalized to the particle. [0279] In some embodiments, inhibiting the spread of the virus SARS-CoV-2, influenza, rhinovirus, other viruses causing respiratory infection and/or any other virus includes using a carrier (e.g., a mesoporous silica nanoparticle, lipid nanoparticle, protein-based nanoparticle or any combination thereof with similar size as the virus). In some embodiments, such nanoparticles, microparticles or other carriers are configured to be strategically provided or otherwise administered to a host in one or more ways (e.g., via inhalation, oral ingestion, intravenous injection, topical application, etc.), as desired or required. In some arrangements, the carriers (e.g., nanoparticles) include a size of 1 to 200 nm (e.g., 1 to 200, 10 to 120, 50 to 100, 90 to 110, 100 nm, values between the foregoing ranges, etc.). In some embodiments, the carriers include a size of 0.01 to 1000 nm (e.g., 0.01 to 1000, 10 to 1000, 50 to 1000, 100 to 1000, 1 to 500, 500 to 1000, 200 to 800, 400 to 600 nm, values between the foregoing ranges, etc.). In some embodiments, the carriers include a size of 0.2 to 100 nm (e.g., 0.2 to 100, 1 to 10, 2 to 20, 5 to 50, 10 to lOOnm, values between the foregoing ranges, etc.). Further, the nanoparticles can be fabricated using the bottom-up sol-gel method or top-down method.

[0280] In some embodiments, mitigating the progression of a specific disease caused by the disease-causing agent using a carrier (e.g., a mesoporous silica nanoparticle, lipid nanoparticle, protein-based nanoparticle or any combination thereof with similar size and protrusions as the mimicking pathogen). In some embodiments, such nanoparticles or other carriers are configured to be provided or otherwise administered to a host in one or more ways (e.g., via inhalation, oral ingestion, intravenous injection, topical application, etc.), as desired or required. In some arrangements, the carriers (e.g., nanoparticles) include a size of 1 to 200 nm (e.g., 1 to 200, 10 to 120, 50 to 100, 90 to 110, 100 nm, values between the foregoing ranges, etc.). In some embodiments, the carriers include a size of 0.01 to 1000 nm (e.g., 0.01 to 1000, 10 to 1000, 50 to 1000, 100 to 1000, 1 to 500, 500 to 1000, 200 to 800, 400 to 600 nm, values between the foregoing ranges, etc.). In some embodiments, the carriers include a size of 0.2 to 100 nm (e.g., 0.2 to 100, 1 to 10, 2 to 20, 5 to 50, 10 to lOOnm, values between the foregoing ranges, etc.). Further, the nanoparticles, microparticles can be fabricated using the bottom-up sol-gel method or top-down method.

[0281] In some embodiments, by using known viral genetic information, such as known viral (e.g., coronaviral, influenza viral, rhinoviral and/or other viral, etc.) genetic information, it is possible to produce similar peptides present in targeted viruses. For example, peptides or other structures can be similar or substantially similar to those found in viral glycoprotein spikes and/or protein protrusions, thus, in some arrangements, mimicking (e.g., at least substantially or approximately) at least some of the viral surface properties that assist with the binding of the carrier to certain receptors (e.g., ACE2 N-terminal helix or sialic acid, histo-blood group antigens, ICAM-1, IGF1R, other target receptors ACE2, etc.). In some arrangements, the carrier can include amino acid sequences found in the viral receptor binding domain (RBD) or the viral receptor binding motif (RBM) in the S protein, HA or NA or VP other decorated proteins that could be used or functionalizing the particle with similar (e.g., substantially similar) or identical peptides. Alternatively or additionally, the carrier’s ability to at least partially inhibit entry of viruses can be enhanced by including organic polymers as part of the protrusion (e.g., of cationic polyamidoamine dendrimer (PAMAM)) or by predicting an amino acid sequence or polymer for producing a surface coating which is similar in surface charge as the viral surface or by attaching targeting motifs which are known to bind to the target receptor allowing selective internalization in target cells.

[0282] In one embodiment, the carriers disclosed in the present application, or variations thereof comprise mesoporous silica particles. In some embodiments, such carriers preferably include a spherical or substantially spherical form or shape. In some arrangements, the particles or other carriers are provided with a plurality of protruding (e.g., relative to a spherical or substantially spherical core) peptide structures in the form of protein spikes or protein fragments/protrusions on their surfaces. In some embodiments, each of the particles include 5 to 500 protruding peptide structures (e.g., 5 to 500, 0 to 100, 100 to 200, 200 to 300, 300 to 400, 400 to 500, 100 to 500, 200 to 500, 300 to 500, 0 to 200, 0 to 300, 0 to 400, 0 to 500, values between the foregoing ranges and values, etc.). In some embodiments, each of the particles include 1 to 1000 protruding peptide structures (e.g., 1 to 1000, 0 to 100, 100 to 200, 200 to 300, 300 to 400, 400 to 500, 100 to 500, 200 to 500, 300 to 500, 100 to 600, 200 to 600,

300 to 600, 400 to 600, 500 to 600, 100 to 700, 200 to 700, 300 to 700, 400 to 700, 500 to 700,

600 to 700, 100 to 800, 200 to 800, 300 to 800, 400 to 800, 500 to 800, 600 to 800, 700 to 800,

100 to 900, 200 to 900, 300 to 900, 400 to 900, 500 to 900, 600 to 900, 700 to 900, 800 to 900,

100 to 1000, 200 to 1000, 300 to 1000, 400 to 1000, 500 to 1000, 600 to 1000, 700 to 1000, 800 to 1000, 900 to 1000, 0 to 200, 0 to 300, 0 to 400, 0 to 500, 0 to 600, 0 to 700, 0 to 800, 0 to 900, 0 to 1000, values between the foregoing ranges and values, etc.).

[0283] In one embodiment, the surface features or other members that protrude from a core of the carrier (e.g., spikes) have a length of about 1 to 200 nm (e.g., 1 to 200, 1 to 100, 2 to 80, 5 to 50, 20 to 100, 50 to 100, 100 to 200 nm, values between the foregoing, etc.). In some embodiments, the surface features or other members that protrude from a core of the carrier (e.g., spikes) have a length of 0.2 to 100 nm (e.g., 0.2 to 100, 1 to 10, 2 to 20, 5 to 50, 10 to lOOnm, values between the foregoing ranges, etc.). In some embodiments, the length includes the actual length of a spike or other protrusion is the total liner length of such a spike or protrusion. However, in other embodiments, the length includes the distance from the spherical or other core of the carrier to the outermost radial distance of the protrusion.

[0284] In some embodiments, allowing the carrier (e.g., synthetic particle) to compete with viral particles, such as coronaviruses (e.g., the SARS-CoV-2 virus, variants thereof, etc.), influenzas, rhinoviruses, Respiratory Syncytial Viruses (RSVs), noroviruses, other viruses, etc.) for the same receptor and/or other binding site or portion of a host cell (e.g., ACE2, sialic acid, histo-blood group antigens, ICAM-1 , IGF1R receptor, etc.) can function as a hindrance and/or other obstacle (e.g., allosteric regulation or hinder, other competitive or non-competitive inhibition, etc.) for the viral particle to bind to the receptor or other site or portion. This can advantageously minimize or reduce the likelihood of endocytosis of the virus or other pathogen, thereby lowering the risk of infecting the host cell.

[0285] In some embodiments, a host receptor (e.g., ACE2) is responsible for mediating the SARS-CoV-2 infection responsible for coronavirus disease 19 (e.g., CO VID- 19). In some configurations, by binding carriers (e.g., the novel synthetic nanoparticles, other particles, objects, etc.) to that reception site (e.g., receptor), to the specific host receptors motifs and/or other any other site or portions of the host cell, infection (e.g., caused by the SARS- CoV-2 viruses, other viruses, etc.) can be advantageously prevented, controlled and/or otherwise mitigated.

[0286] In some embodiments, a host receptor (e.g., ACE2) responsible for mediating the infection resulting in a specific disease is generally depicted (schematically). In some embodiments, by binding a carrier (e.g., a novel synthetic particle, object, etc.) to that specific area or to the specific host receptors motifs, the infection caused by the specific virus, viruses and/or other pathogen can be prevented and controlled (e.g., the likelihood of infection can be reduced or otherwise mitigated, etc.). The competitive inhibition can be utilized against different viruses and/or other pathogens (such as, for example and without limitation, influenzas, rhinoviruses, RSVs, noroviruses, other respiratory and gastrointestinal viruses, other viruses or pathogens, etc.).

[0287] Based on, for example, the foregoing, in an embodiment, carriers (e.g., synthetic nanoparticles, other particles, etc.) are selected such that they resemble, at least partially, coronaviruses (e.g., SARS-CoV-2), influenzas, rhinoviruses, noroviruses, other common cold viruses and/or any other viruses or pathogens, as desired or required. In some embodiments, preferably, synthetic nanoparticles are enhanced or otherwise optimized, at least partially, for competitive inhibition. For example, the particle morphology, size, surface properties and/or any other properties or features of such particles can be modified to achieve higher (or otherwise improve) affinity for the target receptor angiotensin converting enzyme 2 (ACE2) and/or TMPRSS2, sialic acid, histo-blood group antigens, ICAM-1, IGF1R or other target receptors. Thus, the binding affinity for the specific receptor can be advantageously increased, thereby blocking the internalization of the viral envelope more efficiently and potentially prolonging the gained viral protection.

[0288] A carrier system as described herein, wherein the carrier (e.g., synthetic nanoparticle, other particle or object, etc.) resembling a targeted virus (e.g., the SARS-CoV-2, other corona or spiked viruses, influenza, rhinoviruses, noroviruses, other common cold viruses, etc.) can be enhanced or optimized for personalized medicine as variations and mutations in individuals might give rise to slightly different target receptors. Thus, the surface properties and functionalization of the carrier can be changed to match or substantially match the individual properties (e.g., mutations or variations) in target receptors and/or other binding sites or locations of a host cell for tailored therapies.

[0289] One embodiment of a targeted and/or personalized medicine is schematically illustrated in Figures 5E, 5F, 10 and 11. As depicted, by way of an example, by designing a carrier (e.g., synthetic nanoparticle, other particle or object, etc.) that has features that resemble the selected or targeted virus or other pathogen (e.g., SARS-CoV-2, other corona or spiked viruses, influenzas, rhinoviruses, noroviruses, other common cold viruses, etc.). For example, the synthetic particles or other carriers can include spike protein fragments, protein protrusions, other protrusions, other surface features and/or any other feature or property. Accordingly, it is possible for targeted drug delivery at (e.g., at, to, near, etc.) host cells that are susceptible for the virus and/or other pathogen. In some embodiments, as discussed herein, the carrier can include (e.g., can be “loaded” or otherwise provided with) one or more drugs and/or other compounds (for example, fluorescent molecules), substances and/or materials (for example, anti-viral compounds, zinc, immune modulating drugs (e.g., Celastrol, other interferon-gamma or stimulating molecules, penicillium, Dalbavancin or other anti-bacterial compounds, drugs intended to combat virus-related pneumonia, voriconazole, isavuconazole, drugs intended to combat viral-associated pulmonary aspergillosis, anti-fungal compounds, etc.) and/or the like, as desired or required by a particular application or use.

[0290] In some embodiments, the synthetic particle or other carrier comprises (e.g., is provided with) a coating and/or functionalization that has higher affinity towards the receptor favoring the binding of the synthetic particle or other carrier than the viral one (e.g., the virus, other pathogenic or infectious agent or member, etc.).

- H- [0291] In some embodiments, the synthetic particle or other carrier comprises an amino acid sequence that is similar to that of the said viral protrusion having affinity for the same target receptor as the pathogen thus having competition for the same receptor.

[0292] In some embodiments, the synthetic particle or other carrier is further optimized or enhanced, at least partially, for improved binding (e.g., at least partially, at least temporarily, etc.) to said host receptor in order to achieve improved blocking effect by competitive inhibition to the said pathogen.

[0293] In some embodiments, the synthetic particle or other carrier having coating and/or functionalization of epitopes similar to that of the pathogen of interest in order to give a vaccination at target cell population.

[0294] According to some embodiments, the synthetic particle or other carrier is decorated and/or loaded with immunogenic epitopes of the pathogen or pathogens of interest for the targeted cell populations to present the said epitopes to B and/or T cells in order to elicit an immunological response towards said pathogen or pathogens. In some embodiments, this advantageously results in the infected cells to be hindered and/or eliminated by the host’s immune system.

[0295] In some embodiments, the synthetic particle (e.g., carrier) is decorated and/or loaded with polypeptide protrusions containing epitopes from several different pathogens of interest to be used in the composition of a particle-based vaccine composition. The carrier protrusions, which can include both B cell stimulating and/or T cell stimulating epitopes, can be configured to comprise a specific sequence obtained from the amino-acid sequence of the protein of interest (e.g., derived from a coronavirus, SARS-CoV-2 virus, an influenza virus, a rhinovirus, a norovirus, a respiratory syncytial virus (RSV), another virus that impacts the respiratory system, any other type of virus, etc.).

[0296] In some embodiments, the synthetic particle or other carrier is decorated and/or loaded with selected and isolated nucleic acid molecules (e.g., RNA or DNA) having a nucleic acid sequence that encodes a specific polypeptide sequence. Such a sequence can include both B cell stimulating and/or T cell stimulating epitopes that include specific sequences. In some embodiments, the sequences are obtained from the amino-acid sequence (e.g., complete sequence) of the protein of interest (e.g., derived from a coronavirus, SARS- CoV-2 virus, an influenza virus, a rhinovirus, a norovirus, a respiratory syncytial virus (RSV), another virus that impacts the respiratory system, any other type of virus, etc.). [0297] In one embodiment, the synthetic particle or other carrier is configured to prevent or at least partially hinder the ability of the SARS-CoV-2 pathogen to infect and/or replicate in the host (e.g., human, animal, etc.).

[0298] In some arrangements, the synthetic particle or carrier is synthesized using different materials and functionalization in order to match or substantially match the optimal or beneficial properties to be administered to target cell populations (e.g., hydrophobic or hydrophilic properties, depending on the intended use).

[0299] In some embodiments, silica (e.g., stable organic silica) is used as the core material that could exhibit a blocking effect that, optionally after modification of the particle, could be prolonged for hours, days or longer as it takes time for silica nanoparticles to degrade in aqueous conditions similar to the environment of the human body.

[0300] In some embodiments, solid lipid particles (e.g., fabricated by a bottom-up method using microfluidics) are used as the core material for the carrier to be further coated, functionalized and/or loaded into or onto with API, epitopes, proteins, RNA/DNA, anti-virals and immune stimulating compounds such as Celastrol, interferon gamma.

[0301] According to some embodiments, self-assembling protein particles produced by genetically engineered bacterial or mammalian cells producing proteins or protein fragments, such as ferritin heavy or light chain, are used as the core material for the carrier. Such particles can be further functionalized and/or loaded into or onto with other molecules, epitopes, API, epitopes, proteins, RNA/DNA, anti-virals and immune stimulating compounds such as Celastrol, interferon gamma.

[0302] According to some embodiments, the administration route of a carrier depends on, at least partially, the tissue that the virus has invaded. For example, if the virus or other targeted pathogen resides in the upper or lower respiratory tract, it may be preferred to use an inhalation device for administering the carriers (e.g., synthetic particles) with a desired dosage. In some arrangements, such an inhalation device can allow a desired (e.g., optimal, effective, etc.) dosage of a carrier to be provided to a targeted anatomical location on demand.

[0303] According to some embodiments, an inhalation device compromises a container (e.g., a small plastic container) with dried carriers (e.g., synthetic particles, objects, etc.) like that of a dry powder inhaler or as a meter dose inhaler where the carriers (e.g., particles) are sprayed from the inhaler as an aerosol, as an vaporizer creating a fine mist of particles and solution, as an nasal spray dispersed in an aqueous solution and/or in any other form or configuration or hybrid form, as desired or required. [0304] According to some embodiments, for improving or enhancing (e.g., maximizing) the coverage of the upper respiratory tract, an inhalation mask is used. As a result, the entry of carriers (e.g., particles) into the nasal cavity and lower respiratory tract (where epithelial cells expressing ACE2, sialic acid, histo-blood group antigens, ICAM-1, IGF1R or other receptors that may also reside) can be enhanced or otherwise improved, thereby lowering (e.g. , minimizing) the risk of being infected by the virus or other pathogen, at least temporarily.

[0305] According to some embodiments, where the viral infection is (or would be) in the gastrointestinal tract, a tablet, an orally ingestible liquid and/or any other ingestible material is the preferred route of administration of the carrier to the host or subject. The synthetic particles or other carriers of such orally administered compositions can advantageously temporarily protect, at least partially, the end-user from infection by the virus (e.g., orally, via fecal-oral transmission, etc.).

[0306] In some embodiments, the carrier (e.g., nanomaterial, other particle or object, etc.) is fabricated and configured to have a high or a favorable affinity for the pathogen. In one embodiment, the carrier is configured to at least partially encapsulate and/or immobilize the threat of infection. In some arrangements, coating or otherwise functionalizing the particle with molecules to provide an increased (e.g., higher compared to a non-functionalized version) binding affinity towards the pathogen. Accordingly, such carriers could be used in and/or used as disinfecting products (e.g., cleaning solution, hand sanitizer products, disinfecting wipes, etc.).

[0307] FIG. 1 schematically illustrates one embodiment of utilization of carriers 10 (e.g., nanoparticles, other particles or objects, etc.) coated and/or otherwise provided with peptides (e.g., protrusions) resembling the binding motif of the target receptor 20, such as, e.g., ACE-2, sialic acid, histo-blood group antigens, ICAM-1, IGF1R or other receptors that the specific or targeted virus or other pathogen uses and/or are otherwise functionalized for a specific purpose. For example, such targeted viruses or other pathogens include, without limitation or restrictions, coronaviruses (e.g., SARS-CoV-2), influenzas, rhinoviruses, noroviruses and other common cold viruses and/or the like. In some embodiments, as discussed throughout this application, the carriers are configured to at least partially encapsulate and/or immobilize the virus and/or other pathogen or agent, thus minimizing or at least reducing the risk of the virus and/or other pathogen infecting the host (e.g., host cells 4).

[0308] With continued reference to FIG. 1, utilization of carriers (e.g., nanoparticles, other particles or objects, etc.) coated and/or otherwise provided with peptides that can include immune stimulating epitopes resembling the naturally occurring protrusions of the pathogen or pathogens capable of binding to targeted receptor and/or receptors, blocking viral entry and functioning as a vaccine for targeted cell populations. For example, such targeted viruses or other pathogens can include, without limitation or restrictions, coronaviruses (e.g., SARS-CoV-2), influenzas, rhinoviruses, noroviruses and other common cold viruses and/or the like. In some embodiments, as discussed throughout this application, the carriers are configured to at least partially hinder viral or pathogen spread and elicit a protective immune response against said virus and/or other pathogen, thus minimizing or otherwise reducing the risk of viral or other pathogenic infection within the host.

[0309] In some embodiments, the carrier (e.g., nanomaterial, other particle or object, etc.) are fabricated to have a favorable, increased or enhanced (e.g., higher relative to embodiments that do not have similar features) affinity for the receptor and co-receptor. Thus, the carrier 10 can be configured to bind to multiple receptors (e.g., receptors that various pathogens use for cell entry). Further, as noted herein, the synthetic particle or other carrier can be provided with a coating or similar layering or component that has higher or otherwise favorable or improved affinity towards the receptor favoring the binding of the carrier (e.g., synthetic particle). This can allow or otherwise facilitate the carrier to be used in a multiple targeting approach (e.g., further reducing (e.g., minimizing) the risk of contracting said disease or diseases (e.g., viral or pathogenic infection and the diseases originating therefrom)).

[0310] In some embodiments, the fabricated nanoparticles or other carriers are coated (or otherwise provided) with peptides resembling the binding motif of a viral protrusion protein, such as, for example, the spike protein from the SARS-CoV-2 or other coronaviruses, combined with other protrusions for example Hemagglutinin (HA) and Neuraminidase (NA) proteins from influenza A virus, etc. combined with peptides resembling the binding motif of host receptors (e.g., ACE2, sialic acid, histo-blood group antigens, ICAM-1, IGF1R or other receptors) of the subject. As a result, the carriers can advantageously be provided with multiple targeting strategies, thereby minimizing or reducing the risk of multiple viruses or other pathogens infecting the host.

[0311] According to some embodiments, the carrier (e.g., nanomaterial, particle or object, etc.) is fabricated or otherwise configured to have high or favorable affinity for the targeted pathogen(s) and/or other agent(s) (e.g., virus(es)) circulating co-receptors e.g., high- density lipoprotein (HDL) scavenger receptor B type 1 (SR-B1), etc.). Thus, a threat which that may be used as an antidote can be neutralized and/or otherwise handled, thereby preventing or reducing the likelihood of further spreading of the virus and/or another pathogen or agent in the host to which the carrier is administered. [0312] With continued reference to FIG. 1, utilization of nanoparticles or other carriers coated (and/or otherwise provided) with peptides resembling the binding motif of the spike protein from a coronavirus (e.g., the SARS-CoV-2 virus) can help at least partially bind to target receptor(s) and/or co-receptor(s). Accordingly, the presence of such carriers (in accordance with any of the embodiments disclosed herein or equivalents thereof) can decrease the mobility of the virus (and/or other pathogen and/or agent). As a result, the risk of the targeted virus or other pathogen or agent infecting the host, spreading within the host (e.g., to other organs or tissues) and/or causing any other undesirable and potentially dangerous or harmful impact to the host can be reduced or eliminated.

[0313] According to some embodiments, a carrier (e.g., synthetized carrier in the nano- or microscale or any other object that has the capacity of saturating and binding to target receptors, proteins and/or macromolecules) can help prevent the entry of a virus, other pathogen and/or other agent into host cells. Receptors and/or other binding sites to which the carrier maybe configure to bind include, for example and without limitation to, ACE2, sialic acid, histo-blood group antigens, ICAM-1, IGF1R or other receptors at the surface of cells that prevent or minimize pathogen, such as influenzas, rhinoviruses, RS Vs, noroviruses, coronaviruses (e.g., SARS-CoV-2), other viruses causing respiratory infection, binding and entry to the host lowering the risk of contracting the specific disease, such as COVID- 19 disease, diarrhea, common cold, cytokine storm, death or generally discomfort or a combination thereof.

[0314] According to some embodiments, a carrier (e.g., a synthetized carrier in the nano- or microscale or any other object that has the capacity of binding and encapsulating the pathogen and/or other agent of interest) is configured to at least partially inhibit the ability of target pathogens and/or other agents to bind (e.g., at least partially, at least temporarily, etc.) to entire the host, thereby lowering the risk of contracting the specific infectious agent).

[0315] According to some embodiments, a carrier (e.g., a carrier described herein or equivalents thereof, a carrier fabricated, at least in part, using one or more of the methods disclosed herein, etc.) comprises a core or other base structure or component that is obtained (e.g., manufactured, fabricated, etc.), at least in part, by 3D printing, microfluidics, supercritical solution method, sol-gel method, other bottom-up and/or top-down method of fabrication selfassembling components and/or any other method or technology.

[0316] According to some embodiments, a carrier (e.g., as provided above and/or herein) comprises a core or base component or material comprising one or more of the following materials: an organic component, an inorganic component, lipid droplets, micelles, cholesterol, amino acids, proteins, salts and minerals, other molecules and/or any other material.

[0317] According to some embodiments, lipid-based micelles comprise cholesterol decorated with SARS-CoV-2 spike protein fragments and/or other protrusions that bind both to host receptor sites or other portions of the host cell (e.g., ACE2, TMPRSS2, etc.) and to cholesterol and its high-density lipoprotein (HDL) scavenger receptor B type 1 (SR-B1) that would facilitate ACE2-dependent entry of the nanoparticle and/or microparticle loaded with selected API for combating COVID- 19 disease or other disease resulting from infection by a virus or other pathogen. In some embodiments, the cholesterol recognition amino acid consensus (CRAC) motifs near the inverted cholesterol recognition motif (CARC) have been proven to bind with SARS-CoV-2 S I subunit and this HDL complex enhances viral entry to host cells facilitating replication. Therefore, by creating a carrier (e.g., nanoparticle, other particle or object, etc.) that would compete with this spike protein-HDL interaction would potentially lower the ability of SARS-CoV-2 (or the targeted pathogen for ACE2-mediated (or other receptor- mediated)) internalization, at least partially blocking viral entry to host cells and at least partially hindering replication. In one arrangements, this co-receptor incarceration could be blocked by decorating the nanoparticle with spike protein fragments from CARC- CRAC region of SARS-CoV-2 preferably but not limited to together with other amino acids for example the RBD spike fragment hexapeptide that binds to the ACE2 receptor creating a nanoparticle capable of blocking viral-host interaction on multiple positions loaded with selected API for targeted therapeutics (e.g., Celastrol, Zinc, ITX 5601, etc.).

[0318] One embodiment comprises the use of simultaneous inhibiting of multiple receptors by multiple targeting approaches, where the carrier (e.g., mimetic particle) includes protrusions (e.g., located on or along an outer surface of the carrier, extending from the outer surface, etc.) that are similar or substantially similar to those of the target virus or viruses. Such protrusions can include, for example, spike proteins, HA and NA or VP that would bind to the specific host receptor for inhibiting viral entry by competitive inhibition, etc. In some embodiments, the carrier (e.g., virus-like particle) also includes surface protrusions that mimic epitopes selected from said viruses for eliciting an immune response against said viruses.

[0319] One embodiment comprises the use of simultaneous targeting of multiple receptors by multiple targeting approaches, where the carrier (e.g., mimetic particle) includes protrusions (e.g., located on or along an outer surface of the carrier, extending from the outer surface, etc.) that are similar or substantially similar to those of the target virus or viruses. Such protrusions can include, for example, spike proteins, HA and NA or VP that would bind to the specific host receptor for inhibiting viral entry by competitive inhibition, etc. In some embodiments, the carrier (e.g., virus-like particle) also includes drugs, API or other molecules for targeted delivery to the host.

[0320] According to some embodiments, carriers are used to simultaneous inhibit (at least partially) and immobilize (at least partially) by dual targeting approaches, where the carrier (e.g., mimetic particle) includes protrusions and/or other surface features that are similar, at least in part, to those of the virus or other pathogen or agent. For example, the surface can resemble spike protein, HA and NA or VP that would bind to the specific host receptor for inhibiting viral entry by competitive inhibition. In some embodiments, the carrier (e.g., viruslike particle) also includes surface protrusions that mimic the host component (e.g., ACE2, silicid sialic acid, histo-blood group antigens, ICAM-1, IGF1R receptors, and/or antibodies such as the monoclonal antibody bebtelovimab, etc.).

[0321] According to some embodiments, self-assembling recombinant proteinbased nanoparticle constructs, such as, for example, SpyTag™/SpyCatcher™ system and ferritin-based constructs, are used. Where the constructs are expressed in E. Coli; the proteins are purified and then assembled like a two-component “superglue” into virus-like particles (VLPs) conjugated with the selected antigens, viral epitopes or fragments. The carrier could be assembled using the SpyTag™/SpyCatcher™ system or ferrtin (heavy or light chain) based particle core and then conjugated, coated and/or functionalized with the selected SARS-CoV- 2 spike protein or selected hexapeptide 438YKYRYL443 derived thereof or peptides from the CARC-CRAC region or other proteins of interest. It appears possible to develop mimetic nanoparticles or other carriers for preventing the spreading and lowering the infection rate of novel coronaviruses with higher affinity then the RBD monomer.

[0322] For any of the embodiments disclosed herein, the synthetic carrier or nanoparticle may comprise or be decorated with a polypeptide or protein having an amino acid sequence of the ACE2 binding sequence and/or the SARS-CoV-2 spike protein RBD or a fragment thereof. In an embodiment, the amino acid sequence of the ACE2 binding sequence and/or the SARS-CoV-2 spike protein RBD or a fragment thereof is optimized, for example such that it has a higher binding affinity for the ACE2 receptor and enhanced blocking properties that of the spike protein of the coronavirus interaction compared to the corresponding, unmodified spike protein. By optimizing the amino acid sequence of the ACE2 binding sequence and/or the SARS-CoV-2 spike protein RBD or a fragment thereof it is possible achieve even higher binding affinity for example with combining hexapeptides with the optimized spike protein. [0323] According to some embodiments, in a carrier as described herein or an equivalent thereof, the core or core material may be made of, for example, self-assembling virus-like protein nanoparticles that can be saturated with different drugs. Such particles can be synthesized in various sizes and/or shapes, as desired or required.

[0324] According to some embodiments, in a carrier as described herein or an equivalent thereof, the core material comprises, for example, mesoporous silica nanoparticles with ordered mesostructures of pores that can be loaded with different drugs and that these particles can be synthetized in various sizes and shapes.

[0325] According to some embodiments, in a carrier as described herein or an equivalent thereof, the core material can be functionalized with one or several of the following: amino acids, epitopes, peptides or proteins and/or protein fragments, chemical agents, active pharmaceutical ingredients (API), organic or inorganic polymers, molecules and/or the like.

[0326] According to some embodiments, in a carrier as described herein or an equivalent thereof, the carrier with its functionalization provides a method of specifically binding, at least partially, to receptors, proteins, macromolecules at the cellular level and/or other sites in order to at least partially prevent and minimize (e.g., reduce) pathogen entry to the host target tissues by competitive inhibition.

[0327] According to some embodiments, the use of a carrier as described herein or an equivalent thereof provides a method of specifically bind to receptors, proteins and macromolecules at the cellular level in order to prevent and minimize SARS-CoV-2, influenzas, rhinoviruses, respiratory syncytial virus, norovirus and other viruses causing respiratory infection entry to the host target receptors by competitive inhibition.

[0328] According to some embodiments, a method of loading drugs, API, molecules, peptides inside or onto the carrier system is provide herein.

[0329] According to some embodiments, in a carrier system as described herein or an equivalent thereof, the functionalized and drug loaded carrier system can be used for targeted drug delivery of anti-pathogenic, anti-viral or anti-microbial compounds in order to decrease the growth of the infectious agent.

[0330] According to some embodiments, in a carrier as described herein or an equivalent thereof, the functionalized and drug loaded carrier system can be used for targeted drug delivery of, anti-viral compounds in order to decrease the replication rate of the coronavirus.

[0331] According to some embodiments, a synthetic nanoparticle resembling the SARS-CoV-2 virus is loaded into or onto the nanoparticle for further enhancing the anti-viral properties of the carrier. For example, Zinc can be added to the carrier to reduce viral replication in its host cells. In some embodiments, viscosity modulators, antihistamines, Celastrol, immunosuppressors and/or any other products can be provided to the carrier for delivery to the anatomy to combat the onset and/or symptoms of COVID-19 disease (e.g., for reducing or minimizing the cytokine storm that potentially is dangerous to some patients).

[0332] According to some embodiments, synthetic nanoparticle resembling one or more target viruses, pathogens and/or other agents (e.g., SARS-CoV-2, influenzas, rhinoviruses and other viruses causing respiratory infection, etc.) are loaded or provided with proteome inhibitors or new molecular entities developed in the future for efficiently deliver the compounds in the target tissues with minimal off-target effects.

[0333] According to some embodiments, the synthetic nanoparticle of a carrier is decorated with (and/or provided with) one or more molecules that have a relatively high or enhanced affinity towards the SARS-CoV-2 virus or influenzas, rhinoviruses and viruses causing respiratory infection e.g. proteins resembling that of the ACE-2, sialic acid, histo-blood group antigens, ICAM-1, IGF1R receptor or any other pathogen of interest in order to bind and/or immobilize the infectious agents (e.g., thereby preventing or minimizing the potential risk of host entry).

[0334] According to some embodiments, a synthetic nanoparticle resembling the SARS-CoV-2 virus and/or any other pathogen or agent (e.g., influenzas, rhinoviruses and viruses causing respiratory infection, bacteria, fungus, etc.) is decorated with epitopes to be used as a vaccination (e.g., to have an effect similar to that provide by a vaccine) at target cell populations.

[0335] According to some embodiments, the carrier system is loaded, stored and/or dispersed in a device or vessel capable of on-demand release of the carrier to the end-user. In some embodiments, the carrier system is loaded inside a dispenser such as an inhalation device, tablet, injectable substance, cream, ointment and/or any other prescription and/or over the counter (e.g., consumer) product.

[0336] According to some embodiments, man-made materials (e.g., carriers) are used to at least partially immobilize specific pathogens and/or other agents by adding the synthetic material in sanitation products and disinfectants.

[0337] According to some embodiments, a carrier system (as described herein and/or manufactured using the method provided herein) for minimizing or reducing the infection and spread of infection caused by diverse pathogen populations is configured to bind, at least in part, target molecule in the host body and/or bind to the infectious agent itself to at least partially inhibit the onset and/or spread of a disease or disorder. Furthermore, as a combination treatment, the carrier can help hinder the replication of the infectious agent together with giving the immune system in the host a gained advantage to fight the disease similar to vaccines or immunoregulating drugs.

[0338] In some embodiments, methods for preparing a synthetic nanomaterial comprising a core, particle, sheet, film or spheroid, tringle, star shaped and/or shape or configuration is disclosed. Such carriers can include a coating and/or other functionalization (e.g., of organic polymers, amino acids proteins, molecules) that at least partially mimics the surface of the target pathogen(s) and/or other agent(s) (e.g., SARS-CoV-2, other coronavirus, existing and/or future variants, influenzas, rhinoviruses and other viruses causing respiratory infection, bacteria, mold, other fungi, other pathogens, other agents, etc.). In some embodiments, the carriers comprise protrusions and/or other proteins at least partially mimicking pathogens such as the SARS-CoV-2 spike protein, and variation thereof, may be produced using a vector for producing the specific protein construct to be conjugated to the virus-like nanoparticle or synthetic carrier in host cells such as mammalians or human cells.

[0339] Producing a man-made material that has the capability of mimicking the pathogen of interest that has the capability of competing with the pathogen of interest for the same host target molecule, receptor, amino acid or nucleotide. Alternatively, in some embodiments, producing a material that has the capability of binding and immobilizing the pathogen of interest minimizing the possible infection in its host. However, in other embodiments, such immobilization is not required or necessary.

[0340] Producing a man-made material that has the capability of mimicking the coronavirus of interest e.g., SARS-CoV-2 or influenzas, rhinoviruses and other viruses causing respiratory infection that has the capability of competing with the virus for the same host target molecule, receptor, amino acid or nucleotide e.g., ACE2 and/or TMPRSS2, sialic acid, histo- blood group antigens, ICAM-1, IGF1R receptors.

[0341] In some embodiments, a microfluidic device, can include a base, a first inlet capillary that is disposed parallel to the base; a second inlet capillary that is disposed perpendicular to the base, a glass capillary disposed parallel to the base, a luer configured to engage the first inlet, the second inlet, and the glass capillary; and an outlet including a distal tip. In some embodiments, the first inlet and the second inlet can include an inner diameter of 100, 150, 200, 250, 300, 350, 400, 450, 500, 550, 600, 650, 700 pm, or any range of values therebetween. For example, the first inlet and the second inlet can include an inner diameter of about 500 to about 700 pm. In some embodiments, the first inlet and the second inlet can include an outer diameter of 200, 250, 300, 350, 400, 450, 500, 550, 600, 650, 700, 750, 800, 850, 900, 950, 1000, 1050, 1100, 1150, 1200, 1250, 1300, 1350, 1400, 1450, 1500 pm. For example, first inlet and the second inlet can include an outer diameter of about 800 to about 1200 pm. In some embodiments, the distal tip can be polished, smooth, or substantially frictionless. In some embodiments, the microfluidic device can further include external equipment, the external equipment can engage the microfluidic device. In some embodiments, the external equipment can include fluid pumps, compressed gas lines, computers, microscopes, electrical supply outlets, or any combination thereof.

[0342] In some embodiments, a system for functionalizing a carrier includes a first reservoir; a microfluidics device including a base; a first inlet capillary that is disposed parallel to the base; a second inlet capillary that is disposed perpendicular to the base; a glass capillary disposed parallel to the base; a luer configured to engage the first inlet, the second inlet, and the glass capillary; and an outlet including a distal tip; a collection chamber; and a third reservoir, wherein the third reservoir includes a functionalization medium. In some embodiments, the first reservoir includes a volume of a first medium; a second reservoir. In some embodiments, the second reservoir includes a volume of a second medium. In some embodiments, the first reservoir is configured to engage the first inlet capillary, and wherein the second reservoir is configured to engage the second inlet capillary. In some embodiments, the collection chamber includes a volume of an aqueous solution. In some embodiments, the aqueous solution is water. In some embodiments, the third reservoir is configured to engage the collection chamber. In some embodiments, the functionalization medium includes a solution. In some embodiments, the functionalization medium is a solid. In some embodiments, the functionalization medium includes at least one protrusion. In some embodiments, the system further includes a pump. In some embodiments, the system further includes a heat exchanger. In some embodiments, the heat exchanger is configured to maintain a temperature of the collection chamber of about 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10 °C. For example, the heat exchanger can maintain a temperature of the collection chamber from about 3 °C to about 5 °C. In some embodiments, the microfluidic device is configured to fabricate a core.

[0343] The inventive concepts disclosed herein can be implemented in various ways. The inventions and their embodiments are not limited to the examples described above but may vary within the scope of the claims.

[0344] Although several embodiments and examples are disclosed herein, the present application extends beyond the specifically disclosed embodiments to other alternative embodiments and/or uses of the inventions and modifications and equivalents thereof. It is also contemplated that various combinations or subcombinations of the specific features and aspects of the embodiments may be made and still fall within the scope of the inventions. Accordingly, it should be understood that various features and aspects of the disclosed embodiments can be combined with or substituted for one another in order to form varying modes of the disclosed inventions. Thus, it is intended that the scope of the present inventions herein disclosed should not be limited by the particular disclosed embodiments described above but should be determined only by a fair reading of the claims that follow.

[0345] While the inventions are susceptible to various modifications, and alternative forms, specific examples thereof have been shown in the drawings and are herein described in detail. It should be understood, however, that the inventions are not to be limited to the particular forms or methods disclosed, but, to the contrary, the inventions are to cover all modifications, equivalents, and alternatives falling within the spirit and scope of the various embodiments described and the appended claims. Any methods disclosed herein need not be performed in the order recited. The methods summarized above and set forth in further detail below describe certain actions taken by a practitioner; however, it should be understood that they can also include the instruction of those actions by another party. The methods summarized above and set forth in further detail below describe certain actions taken by a user (e.g., a professional in some instances); however, it should be understood that they can also include the instruction of those actions by another party. Thus, actions such as “delivering” include “instructing delivering.” The ranges disclosed herein also encompass any and all overlap, sub-ranges, and combinations thereof. Language such as “up to,” “at least,” “greater than,” “less than,” “between,” and the like includes the number recited. Numbers proceeded by a term such as “about” or “approximately” include the recited numbers. For example, “about 10 mm” includes “10 mm.” Terms or phrases preceded by a term such as “substantially” include the recited term or phrase. For example, “substantially parallel” includes “parallel.”

EXPERIMENTAL SECTION

[0346] The experimental section that follows provides additional information regarding certain embodiments related to the subject matter of the present application. As such, it is not intended to and should not be used to restrict or otherwise limit the scope of the application and the inventions disclosed therein in any manner.

[0347] Fabrication of the microfluidic device: The micro fluidic device was fabricated through coaxially aligned assembling of two borosilicate glass capillaries (World Precision Instruments, USA) (Whitesides, G.M., The origins and the future of microfluidics. Nature, 2006; 442(7101):368-373. doi: 10.1038/nature05058.). The inner capillary with an inner diameter (i.d.) of 580 pm and outer diameter (o.d.) of 1000 pm was tapered using a micropipette puller (P-97, Sutter Instrument, USA). The fine tip was polished, and the diameter of the orifice was further enlarged to approximately 100 pm. This inner capillary was coaxially inserted into the outer cylindrical capillary with i.d. of 1120 pm and o.d. of 1500 pm. The capillaries were fixed on a glass slide and sealed as required employing a transparent epoxy resin.

[0348] Preparation of SLN through microfluidic precipitation method'. The SLN composed of DOTAP (Avanti Polar Lipids, Inc., USA), Choi (>99%; Merck Limited, USA), DOPE (Avanti Polar Lipids, Inc., USA), and DSPE-PEG2000-Biotin (Avanti Polar Lipids, Inc., USA) was prepared through nanoprecipitation method employing a microfluidic device. The inner phase, ethanol solution of DOTAP (4.84 mg/mL), Choi (0.54 mg/mL), DOPE (4 mg/mL), and DSPE-PEG-Biotin (0.62 mg/mL), was pumped into the inner capillary. The outer phase, an aqueous solution of PVA (1%, w/v), flowed through the space between the inner and outer capillary in the same direction. The lipid molecules precipitated into nanoparticles as the diffusion of water into the ethanol phase. The flow rate of the inner and outer phases was ImL/min, controlled by PHD 2000 pumps (Harvard Apparatus, USA). To thoroughly remove the ethanol and PVA, the fabricated SLN were washed with Milli-Q water employing Nanosep® Centrifugal Device with Omega™ Membrane 100K (Pall Corporation, USA) 6 times. The FITC-labelled SLN was prepared with modified method, replacing the inner phase with an ethanol solution of DOTAP (4.22 mg/mL), Choi (0.54 mg/mL), DOPE (4 mg/mL), DSPE-PEG-Biotin (0.62 mg/mL) and distearoyl phosphoethanolamine-polyethylene glycolfluorescein isothiocyanate (DSPE-PEG-FITC, 0.62 mg/mL, Nanosoft Polymers, USA). The flow velocity of the inner and outer phase was 1 and 0.82 mL/min, respectively. To thoroughly remove the ethanol and PVA, the fabricated Cy5.5-labelled SLN were washed with Milli-Q water employing Nanosep® Centrifugal Device with Omega™ Membrane 100K (Pall Corporation, USA) 6 times.. Similarly, the Cy5.5-labelled SLN was prepared by replacing the inner phase with an ethanol solution of DOTAP (4.79 mg/mL), Choi (0.54 mg/mL), DOPE (4 mg/mL), DSPE-PEG-Biotin (0.62 mg/mL) and distearoyl phosphoethanolamine-polyethylene glycol-cyanine5.5 (DSPE-PEG-Cy5.5, 0.05 mg/mL, BroadPharm, USA). The flow velocity of the inner and outer phase was 1 and 0.9 mL/min, respectively. To thoroughly remove the ethanol and PVA, the fabricated Cy5.5-labelled SLN were washed with Milli-Q water employing Nanosep® Centrifugal Device with Omega™ Membrane 100K (Pall Corporation, USA) 6 times. [0349] Preparation of SLN through bulk precipitation method: To fabricate SLN by bulk precipitation method, ethanol solution of DOTAP (4.84 mg/mL), Choi (0.54 mg/mL), DOPE (4 mg/mL), and DSPE-PEG-Biotin (0.62 mg/mL, 0.5 mL) was added dropwise into an aqueous solution of PVA (1%, w/v, 0.5 mL) under stirring at 300 rpm. The precipitated nanoparticles were purified by rinsing with Milli-Q water using Nanosep® Centrifugal Device with Omega™ Membrane 100K (Pall Corporation, USA) 6 times.

[0350] Streptavidin-biotin-programmed functionalization of SLN with Spike SI RBD: The SLN was functionalized with Spike S I RBD derived from the original and delta variant SARS-CoV-2 through biotin-streptavidin interaction to form VLPO and VLPD, respectively. Specifically, SARS-CoV-2 Spike SI RBD (original variant, Avi-His-Tag, biotin- labeled, Tebubio, France) solution (90 pg/mL, 0.5 mL) was added to streptavidin solution (120 pg/mL, 0.5 mL) dropwise. After stirring at 4 °C for 2 h, SLN suspension (1 mg/mL, 0.5 mL) was added into the mixture, which was further stirred at 4 °C for 3 h. To remove the unconjugated Spike SI RBD, the mixture was dialyzed against Milli-Q water employing Spectra/Por® dialysis membrane with molecular weight cut-off of 1000 kDa (Biotech CE tubing, Spectrum Labs, USA) for 48 h. For the functionalization of SARS-CoV-2 Spike SI RBD (B.1.617.2, Delta variant, Avi-His-Tag, biotin-labeled, Tebubio, France), the stirring time was decreased to 0.5 and 2 h, respectively. The FITC-labelled VLPO and VLPD were prepared with the same method.

[0351] Characterization of SLN and VLPs: The size and size distribution of SLN and VLPs were analyzed through dynamic light scattering using Zetasizer Nano ZS (Malvern Instruments Ltd., UK). The analysis was performed with a disposable polystyrene cuvette (Sarstedt AG & Co., Germany) at an angle of 173° at 25 °C. Their zeta-potential was measured employing Zetasizer Nano ZS equipped with a disposable folder capillary cell (DTS1070, Malvern Instruments Ltd., UK). Samples were dispersed in Milli-Q water (pH 7.4) during measurement. Morphological characteristics of the fabricated SLN and VLPs stained with ammonium molybdate (1%) were observed by JEOL JEM- 1400 transmission electron microscopy (JEOL Ltd., Japan). The conjugated amount of Spike SI RBD in VLPs was measured with Micro BCA™ Protein Assay Kit (Thermo Fisher Scientific, USA) following the manufacturer’s protocol. Conjugate efficiency, the weight percentage of conjugated Spike SI RBD among the added Spike SI RBD, was calculated as [(weight of conjugated Spike SI RBD/weight of added Spike SI RBD) x 100%]. Mass fraction, the amount of Spike SI RBD conjugated to per unit weight of VLPs, was calculated as [(weight of conjugated Spike SI RBD/weight of VLPs) x 100%]. For the stability study, SLN and VLPs were stored at 4 °C for 100 and 200 days. Afterward, their size and size distribution were measured through dynamic light scattering using Zetasizer Nano ZS (Malvern Instruments Ltd., UK).

[0352] The particle concentration of VLPs was measured through nanoparticle tracking analysis with Nanosight LM14C (Malvern Panalytical Ltd., UK). The VLPs were captured and analyzed with the built-in NanoSight Software NTA version 3.4.4 (Malvern Panalytical Ltd., UK). The camera level was fixed at 15 to ensure all VLPs were visible without signal saturation. The detection threshold was set as 5 to include most of the observed VLPs while excluding indistinct ones. VLP suspensions were diluted with Milli-Q water to concentrations of 0.5-1 pg/mL and injected into the sample chamber by a sterile syringe (HENKE- JECT®, Henke Sass Wolf GmbH, Germany) until reaching the nozzle tip. For each measurement, five consecutive videos were recorded for 30 s with a 405 nm laser (blue) and scientific CMOS camera. The number of conjugated Spike S 1 RBD per VLP was calculated following Equation (1):

[0353] where, N is the number of conjugated Spike S 1 RBD per VLP, m is the mass of conjugated Spike SI RBD, NA is Avogadro constant=6.02xl0²³, C is particle concentration of VLPs, and M_moi is the molar mass of Spike S 1 RBD.

[0354] Cell culture'. The in vitro cytotoxicity and uptake study were conducted with human lung carcinoma A549 cells, A549-ACE2/T cells, human non-small-cell lung cancer Calu-3 cells, and human colorectal adenocarcinoma Caco-2 cells. All cells were cultured in Dulbecco’s modified Eagle medium (DMEM) with 4.5 g/L glucose (Life Technologies Gibco, USA). For A549 and Caco-2 cells, the DMEM medium was supplemented with heat inactivated fetal bovine serum (10%, Life Technologies Gibco, USA), 1-glutamine (1%, GE Healthcare Lifesciences, USA), non-essential amino acids (1%, GE Healthcare Lifesciences, USA), penicillin (100 lU/mL, GE Healthcare Lifesciences, USA), and streptomycin (100 pg/mL, GE Healthcare Lifesciences, USA). Three more antibiotics, normocin (100 pg/mL, InvivoGen, USA), puromycin (0.5 pg/mL, InvivoGen, USA), and hygromycin B gold (300 pg/mL, Invivogen, USA), were further added into the DMEM medium for A549-ACE2/T cells. Calu- 3 cells were cultured in the DMEM medium supplemented with heat inactivated fetal bovine serum (20%), 1-glutamine (1%), penicillin (100 lU/mL), and streptomycin (100 pg/mL). The HEK-293T-ACE2 cells for in vitro pseudoviral infection assay were cultured in Growth Medium IN (BPS Bioscience, USA) and plated in Thaw Medium 1 (BPS Bioscience, USA). All cells were maintained at 37 °C under a 5% CO2 atmosphere and 95% relative humidity. The cell culture medium was changed every other day.

[0355] In vitro cytotoxicity study: To perform the in vitro cytotoxicity study, A549, A549-ACE2/T, Calu-3, and Caco-2 cells were seeded on a 96-well plate (Corning Incorporated, USA) at a density of 2xl0⁴ cells per well, followed by overnight incubation for attachment. Afterward, the culture medium was changed into the fresh medium with SLN and VLPs at different concentrations (10, 20, 50, 100, 200, and 500 pg/mL). Cell culture medium without or with cells served as the blank and positive control, respectively. After incubation for 24, 48, and 72 h, the cell viability was measured using CellTiter-Glo luminescent assay (Promega Corporation, USA) with Varioskan LUX multimode microplate reader (Thermo Fisher Scientific, USA).

[0356] In vitro cell uptake study: The in vitro cell uptake of SLN and VLPs was evaluated through confocal imaging and flow cytometry analysis. For flow cytometry analysis, A549, A549-ACE2/T, Calu-3, and Caco-2 cells were seeded in 12- well plates (Corning Incorporated, USA) at a density of 3x10^s cells per well. The cells were incubated overnight for attachment, followed by treatment with FITC-labelled SLN, VLPO, and VLPD (20 pg/mL) for 0.5, 1, 3, 6, and 24 h. Afterward, the cells were washed with PBS buffer twice and detached by trypsin (0.25%, v/v, GE Healthcare Lifesciences, USA). The detached cells were washed with PBS twice and analyzed on a BD LSR-II Cell Analyzer flow cytometer (Becton Dickinson, USA). The fluorescence on the cell surface was quenched by incubating with trypan blue (0.005%, Thermo Fisher Scientific Inc, USA) for 15 min. The cells were washed with PBS and analyzed again by BD LSR-II Cell Analyzer flow cytometer.

[0357] For confocal imaging, A549, A549-ACE2/T, Calu-3, and Caco-2 cells were seeded in Lab-Tek chambered borosilicate coverglass (8- well, Thermo Fisher Scientific, USA) at a density of 2.5xl0⁴ cells per well. After incubating overnight for attachment, the cells were cocultured with FITC-labelled SLN, VLPO, and VLPD (20 pg/mL) for 3 h. Cells treated with culture medium without any particles served as control. After washing with PBS buffer twice, the cytomembrane was stained with CellMask Deep Red™ (5 pg/mL in PBS, 200 pL, Invitrogen, USA), while the cell nucleus was stained with DAPI (2.48pg/mL, 200 pL, Thermo Fisher Scientific Inc, USA). Finally, the cells were imaged by LEICA SP8 STED confocal microscope (Leica Microsystems, Germany). The 3D image visualization and reconstruction were performed through Imaris™ (Oxford Instruments, UK). Similarly, the uptake of Cy5.5 labelled nanoparticles was evaluated through confocal imaging. Caco-2 cells were seeded in Lab-Tek chambered borosilicate coverglasses (8-well, Thermo Fisher Scientific, USA) at a density of 2.5xl0⁴ cells per well. After incubating overnight for attachment, the cells were cocultured with Cy5.5-labelled SLN and VLPD (20 pg/mL) for 3 hours. Cells treated with culture medium without any particles served as control. After washing with PBS buffer twice, the cytomembrane and cell nucleus was stained with CellMask Deep Red™ and DAPI, respectively, employing the same method as described above. Finally, the cells were imaged by LEICA SP8 STED confocal microscope (Leica Microsystems, Germany).

[0358] Permeability assay: The permeability of SLN and VLPs was evaluated using PAMPA Kit (BioAssay Systems, USA), with a 96-well filter plate as the permeation donor compartment and a 96-well receiver plate as the acceptor. Lecithin dodecane solution (4%, 5 pL) was added to the membrane of each well. Immediately after the application of the artificial membrane, the donor plates were added with test nanoparticle suspensions (100 and 200 pg/mL, 200 pL), while the acceptor plates were filled with phosphate buffered saline (300 pL). All analyzes were performed in quadruplicate. The nanoparticle-filled donor plates were carefully installed into the acceptor plate wells, ensuring the contact of the underside of the membranes with the acceptor cell solution without air bubbles. After incubating under constant shaking (150 rpm) for 18 h at 37 °C, samples were withdrawn from the acceptor plates. Nanoparticle suspensions diluted to 40 and 80 pg/mL served as equilibrium standards, and phosphate buffered saline was employed as blank control. The absorbance spectrum (200-500 nm) for acceptor solutions, equilibrium standards, and blank controls were read through a UV- 1600PC UV-Vis Spectrophotometer (VWR International Oy, Finland) in 10 nm intervals to determine the peak absorbance of test nanoparticles. The apparent permeability coefficient of the nanoparticles was calculated following Equation (2):

[0359] where, P_app is the apparent permeability coefficient, ODA, ODE. and ODB are the peak absorbance of acceptor solutions, equilibrium standards, and blank controls, respectively, and C is a constant, 7.72xl0‘⁶.

[0360] In vitro pseudoviral infection assay: Pseudoviral infection assay was conducted to evaluate the effect of VLPs on blocking the viral entry into host cells and preventing SARS-CoV-2 infection (Editorial. Let’s talk about lipid nanoparticles. Nature Reviews Materials, 2021 ; 6(2):99-99. doi: https ://doi . or g/ 10.1038/s41578 -021 -00281 -4. ) . Briefly, HEK-293T-ACE2 cells were seeded into a 96-well plate at a density of 5xl0³ cells per well and incubated overnight for attachment. The medium was replaced with fresh medium with Spike SI RBD (1 pg/pL, Avi-His-tag, biotin-labeled, BPS Bioscience, USA), SLN (20 and 100 pg/mL) and VLPs (20 and 100 pg/mL). After incubating for 0.5, 1, 2, or 6 h, Spike (SARS-CoV-2) pseudotyped lentivirus (luciferase reporter, 5 pL, BPS Bioscience, USA) was added to each well. Bald lentiviral pseudovirion (luciferase reporter, BPS Bioscience, USA) and Spike SI neutralizing antibody (SARS-CoV-2, Clone:414-1, BPS Bioscience, USA) were employed as negative controls. The neutralizing antibody (10 pL, final concentration of 100 nM) was preincubated with the pseudotyped lentivirus for 30 min before adding to the cells. Cells treated with only pseudotyped lentivirus served as the positive control. At 24 h postinfection, the pseudotyped lentivirus was removed and the cells were overlaid with fresh medium. Following additional 24 h incubation, the pseudoviral infection was measured employing One-Step luciferase assay (BPS Bioscience, USA) with POLARstar Omega microplate reader (BMG Labtech, Germany). The experiments were conducted in triplicate, and pseudoviral infection was normalized with the positive control group.

[0361] Cell uptake inhibition assay: The effect of Spike SI RBD (Avi-His-Tag) on the cell uptake of FITC-labelled VLPs was evaluated on A549-ACE2/T cells through flow cytometry analysis. Briefly, A549-ACE2/T cells were seeded in 12-well plates (Corning Incorporated, USA) at a density of 3xl0⁵ cells per well. After overnight incubation, the cells were pretreated with Spike SI RBD (Avi-His-Tag, 1 pg/pL) for 0.5, 1, 3, and 6 h, followed by the coincubation with FITC-labelled VLPs (20 pg/mL) for 1 h. The cells were detached by trypsin (0.25%, v/v, GE Healthcare Lifesciences, USA) and washed with PBS twice. The cell uptake was analyzed through a BD LSR-II Cell Analyzer flow cytometer. After analysis, the fluorescence on the cell surface was quenched by trypan blue (0.005%, Thermo Fisher Scientific Inc, USA). The quenched cells were washed with PBS and analyzed again by BD LSR-II Cell Analyzer flow cytometer.

[0362] Statistical analysis: Results are expressed as mean ± standard deviation (s.d.) for at least three independent experiments. Statistical analyses were performed (Origin 2021b, Origin Lab Corporation, USA) as indicated in the figure legends. The levels of significance were set at probabilities of *P < 0.05, **P< 0.01, and ***P < 0.001.

RESULTS

[0363] Precise control of the lipid nanoprecipitation process enabled by microfluidic

[0364] SARS-CoV-2 is an enveloped and spherical particle with a diameter of -120 nm (Robinson, P.C., et al., COVID-19 therapeutics: Challenges and directions for the future. Proceedings of the National Academy of Sciences, 2022; 119(15):e2119893119. doi: 10.1073/pnas.2119893119 To mimic the size and structure of SARS-CoV-2, we fabricated a lipid core composed of DOTAP, Choi, DOPE, and DSPE-PEG2000-Biotin with average size of around 120 nm. Microfluidic, an advanced technology that can manipulate small (10^-9 to 10“¹⁸ liters) amounts of fluids at the submillimeter scale, was employed to precisely control the physicochemical properties of the formulation. Microfluidic technologies have been identified with various advantageous features, such as precise fluid control and rapid sample processing, which made them attractive candidates to replace the conventional bulk approaches (mixing the two phases in a vessel) for the fabrication and engineering of nanomaterials (Hoffmann, M., et al., SARS-CoV-2 Cell Entry Depends on ACE2 and TMPRSS2 and Is Blocked by a Clinically Proven Protease Inhibitor. Cell, 2020; 181(2):271-280.e8. doi:

10.1016/j.cell.2020.02.052)

[0365] In this study, the SLN was fabricated by the nanoprecipitation method under conventional bulk and microfluidic conditions (Gordon, D.E., et al., A SARS-CoV-2 protein interaction map reveals targets for drug repurposing. Nature, 2020; 583(7816):459-468. doi: 10.1038/s41586-020-2286-9). For the bulk nanoprecipitation process, the lipid ethanol solution was added dropwise into an aqueous solution of polyvinyl alcohol (PVA, 1%, w/v) under continuous stirring at 300 rpm. For the microfluidic nanoprecipitation process, the lipid solution and PVA aqueous solution were pumped into the inner capillary and the space between the inner and outer capillary of a co-flow glass capillary microfluidic device, respectively. The lipid molecules are self-assembled into SLN as the diffusion of water into the ethanol phase. Although the hydrodynamic size of SLN prepared by the bulk method was close to that obtained with the microfluidic process, the deficient control of the mixing process and unstable mass transfer for bulk condition resulted in higher polydispersity and partial agglomeration with the average size of 4955 nm. By contrast, SLN fabricated by the microfluidic method revealed a monomodal and narrower size distribution, which could be attributed to precise fluid control and rapid microscale mixing in the microfluidic device.

[0366] To illustrate the precise control capability of the microfluidic method towards the nanoprecipitation process, 10 batches of SLN prepared by bulk and microfluidic process were characterized. The average hydrodynamic size of SLN fabricated through bulk method varied from 112.9 nm to 155.1 nm, while that of microfluidic varied in the range of 124.1-133.8 nm . The poly dispersity index (PDI) of SLN fabricated by microfluidic (0.139 ± 0.024) was significantly (P < 0.001) lower than that of the bulk method (0.202 ± 0.020), indicating a narrower size distribution caused by the better mixing performance in the microscale device . Furthermore, the SLN prepared by the microfluidic process displayed higher batch-to-batch reproducibility in terms of zeta potential (44.5-51.3 mV) compared with the bulk method (43.2-54.5 mV. Therefore, the developed microfluidic platform could improve the controllability and reproducibility for the SLN engineering process. To be noted, the productivity of SLN fabricated under microfluidic condition was ~ 0.6 g/h. Considering the flow rate of the ethanol phase (1 mL/min) and lipid concentration (10 mg/mL), the SLN was essentially 100% effective after the purification process.

[0367] Overall, the developed bottom-up fabrication method using our microfluidic device was sufficient to control the nanoprecipitation process of SLN and was able to fabricate SLN with unimodal and narrow size distribution, low batch-to-batch variation, and high productivity. As reproducible manufacturing and scale-up are the major challenges for the translation of nanomedicines to a clinical product, microfluidic technology exhibits significant potential to become an alternative for the conventional bulk method (Guy, R.K., et al., Rapid repurposing of drugs for COVID-19. Science, 2020; 368(6493):829-830. doi: 10.1126/science.abb9332).

[0368] Functionalization of SLN with Spike SI RBD programmed by streptavidin -biotin interaction

[0369] For DSPE-PEG2000-Biotin, the biotin moieties tend to locate on the particle surface when self-assembling because of the hydrophobicity of DSPE and hydrophilicity of PEG (Kaufmann, S.H., et al., Host-directed therapies for bacterial and viral infections. Nature Reviews Drug Discovery, 2018; 17(l):35-56. doi: 10.1038/nrd.2017.162). To mimic the structure of SARS-CoV-2, the surface of SLN immobilized with biotin moieties was functionalized with Spike SI RBD (Avi-His-Tag, biotin-labeled) protrusions derived from SARS-CoV-2 original and delta variant, respectively, through a streptavidin linker. Streptavidin is a tetrameric protein possessing high affinity with biotin (Ka = 10¹³'¹⁵ M ¹) (Beigel, J.H., et al., Advances in respiratory virus therapeutics - A meeting report from the 6th isirv Antiviral Group conference. Antiviral Research, 2019; 167:45-67. doi:

10.1016/j.antiviral.2019.04.006). It has four biotin-binding sites symmetrically located at the exterior region, which potentially lead to multiple captures of biotinylated particles on one tetravalent streptavidin, and as a result, cause particle aggregation (Beigel, J.H., et al., Advances in respiratory virus therapeutics - A meeting report from the 6th isirv Antiviral Group conference. Antiviral Research, 2019; 167:45-67. doi: 10.1016/j. antiviral.2019.04.006). To avoid particles aggregation, Spike S I RBD (Avi-His-Tag, biotin-labeled) was initially mixed with streptavidin in a mole ratio of 1.5 : 1 to form a streptavidin-Spike S 1 RBD complex. This complex was then bound to the biotin moieties on the SLN surface, and the hindrance resistance from bound Spike SI RBD is supposed to avoid multiple captures of particles onto one streptavidin and the resultant aggregation.

[0370] The average size of the SEN was 129.7 nm with a PDI of 0.159 ± 0.031, as measured by dynamic light scattering. As expected, there was no sign of particle aggregation after functionalization, indicated by the uniform peak of hydrodynamic radius at 123.4 ± 2.9 nm for VLPO and 154.3 ± 14.2 nm for VLPD. The size of VLPs was homogenous with comparable PDI (0.167 + 0.008 for SLNO and 0. 162 + 0.034 for SLND) to that of SLN. The SLN was positively charged with a zeta-potential of +51 .7 mV, which decreased (P<0.01) to about +34.6 mV for VLPO and +34.2 mV for VLPD, respectively, after functionalization. The decrease of zeta-potential demonstrated the successful conjugation of streptavidin-Spike SI RBD complex to the SLN. The SLN and VLPs were regarded as stable colloidal suspension systems because of their high zeta potential (>30 mV) and the resultant strong repulsion between nanoparticles (Trimarco, J.D., et al., Cellular glycan modification by B3GAT1 broadly restricts influenza virus infection. Nature Communications, 2022; 13(1):6456. doi: 10.1038/s41467-022-34111-0). As a result, the nanoparticles showed excellent colloidal stability at 4 °C for at least 200 days. The VLPs were observed with a buried exterior surface employing transmission electron microscopy with negative staining, which indicated the successful conjugation of Spike SI RBD to the surface of VLPs. The slight discrepancy in the particle size was observed through dynamic light scattering and transmission electron microscopy, because dynamic light scattering measures the hydrodynamic diameter of the nanoparticle, including the solvation layer whereas transmission electron microscopy presents an estimation of the projected area diameter in a dry state.

[0371] The efficient functionalization of VLPs with Spike SI RBD was measured with Micro BCA™. The conjugate efficiency of Spike SI RBD, defined as the weight percentage of conjugated molecules among the added molecules, was 43.3% for VLPO and 56.4% for VLPD, respectively. This corresponded to the mass fraction, the amount of Spike SI RBD conjugated per unit weight of VLPs, of 3.22% for VLPO and 4.20% for VLPD. The size distribution measured by nanoparticles tracking analysis showed a mean particle radius at 109.4 nm for SLNO and 145.9 nm for SLND with 90% of the particles being <145.4 nm for SLNO and <199.1 nm for SLND, confirming the narrow size distribution of the nanoparticles. This smaller particle size in comparison with dynamic light scattering results is attributed to the lower weighting function and intensity scattered by larger particles for nanoparticle tracking analysis (Wei, J., et al., Pharmacological disruption of mSWI/SNF complex activity restricts SARS-CoV-2 infection. Nature Genetics, 2023; 55(3):471-483. doi: 10.1038/s41588- 023-01307-z) . The total nanoparticles concentration was represented by the area under the curve, which equaled 3xl0⁹ particles/mL for SLNO and 2.03xl0⁹ particles/mL for SLND. Accordingly, the number of Spike SI RBD molecules carried on each VLP was calculated by considering the mass fraction of Spike SI RBD in VLPs, which corresponded to an average of 230 Spike S 1 RBD for VLPO. By contrast, a VLPD carried -444 Spike S 1 RBD on the particle surface, which is significantly (P<0.01) higher than that of VLPO. Benefiting from the remarkable affinity between streptavidin and biotin, the number of Spike SI RBD molecules bound to each VLP was obviously higher than that reported previously, which might potentially improve the blocking efficiency of the VLPs (Kumar, N., et al., Host-Directed Antiviral Therapy. Clinical Microbiology Reviews, 2020; 33(3):e00168-19. doi: 10.1128/CMR.00168- 19).

[0372] Overall, streptavidin-biotin interaction was employed to functionalize the SLN with Spike SI RBD. The fabricated VLPs revealed decreased zeta potential and buried exterior surface, which demonstrated the successful conjugation of Spike SI RBD onto the SLN surface. To be noted, compared with VLPO, VLPD carried significantly larger amount of Spike SI RBD on its surface, which may result in higher blocking efficiency against SARS- CoV-2 infection.

[0373] Enhanced cell uptake of VLPs mediated by the interaction with ACE2 receptor

[0374] The Spike protein of SARS-CoV-2 was reported to interact with human cerebrovascular cells, including endothelial cells, pericytes, and smooth muscle cells, mediated by the ACE2 (Mohsen, M.O. and M.F. Bachmann, Virus-like particle vaccinology, from bench to bedside. Cellular & Molecular Immunology, 2022; 19(9):993-1011. doi: 10.1038/s41423- 022-00897-8) . We hypothesized that the prepared VLPs functionalized with Spike SI RBD bind to the ACE2 on the cell surface, which may enhance their cell uptake into the host cells. To prove that the fabricated VLPs bind to ACE2, we evaluated the cell uptake of VLPs with human lung carcinoma A549 cell line expressing ACE2 and TMPRSS2 (A549-ACE2/T), human non-small-cell lung cancer Calu-3 cells and human colorectal adenocarcinoma Caco-2 cells, which are expressed with ACE2 and highly permissive for SARS-CoV-2 infection (Nooraei, S., et al., Virus-like particles: preparation, immunogenicity and their roles as nanovaccines and drug nanocarriers. Journal of Nanobiotechnology, 2021; 19(1):59. doi: 10.1186/s 12951-021-00806-7). By contrast, A549 cells, which express a negligible level of ACE2, and thus, are poorly permissive for SARS-CoV-2 infection, served as control (Tariq, H., et al., Virus-Like Particles: Revolutionary Platforms for Developing Vaccines Against Emerging Infectious Diseases. Frontiers in Microbiology, 2021; 12:790121. doi: 10.3389/fmicb.2021.790121).

[0375] Before evaluating the cell uptake of SLN and VLPs, a cell viability assay was performed on A549, A549-ACE2/T, Calu-3, and Caco-2 cells to test the nanoparticles biocompatibility. The cells were incubated with SLN, VLPO, and VLPD for 24, 48, and 72 h, and their viability was evaluated by CellTiter-Glo® luminescence assay. The VLPO was nontoxic against A549 and A549-ACE2/T cells at all given concentrations even after 72 h incubation, while SLN and VLPD exhibited slight cell inhibitory effect (viability >60%) at the highest tested concentration (500 pg/mL). For Calu-3 cells, the viability treated with SLN and VLPs of concentration <200 pg/mL were over 85% for all time points, and slight cytotoxic behavior was observed as the concentration increased to 500 pg/mL. Interestingly, SLN and VLPs could stimulate the proliferation of Caco-2 cells, especially after 48 and 72 h coincubation. The viability study suggests that the SLN and VLPs were nontoxic up to 200 pg/mL, and thus, can be potentially used for blocking SARS-CoV-2 infection.

[0376] To prove the interactions between VLPs and ACE2, the uptake of SLN and VLPs by A549, A549-ACE2/T, Calu-3, and Caco-2 cells was evaluated quantitatively through flow cytometry. Fluorescein isothiocyanate (FITC)-labelled DSPE-PEG was employed for the synthesis of fluorescent SLN. The FITC-labelled SLN and VLPs were taken up rapidly with >60% of the cells exhibiting nanoparticles fluorescence after 0.5 h, which may attribute to their positive surface charge. The mean fluorescence intensity (MFI) climbed with the incubation time increasing from 0.5 to 24 h, indicating the continual cell uptake of SLN and VLPs. For A549 cells with negligible levels of ACE2, there was no significant difference between the uptake levels for SLN and VLPs groups. For ACE2 expressing cells (A549-ACE2/T, Calu-3, and Caco-2 cells), the functionalization of VLPs with Spike SI RBD protrusions significantly enhanced their cellular uptake compared to the bare SLN after incubating for <6 h. This enhanced effect disappeared for Calu-3 and Caco-2 cells as the incubation time increased to 24 h, possibly due to the exhaustion of VLPs.

[0377] The enhanced cell uptake of VLPs after functionalization with Spike SI RBD protrusions was further confirmed by visualizing their cellular distribution through a laser scanning confocal microscope. To indicate the distribution of FITC-labelled nanoparticles (green), the cell nucleus was stained with 4’,6-diamidino-2-phenylindole (DAPI, blue), while the cytomembrane was stained with CellMask Deep Red. Consistent with the results of flow cytometry, the fluorescence intensity of the FITC-labelled VLPs was stronger than that of the FITC-labelled SLN group for A549-ACE2/T, Calu-3, and Caco-2 cells, while negligible difference was observed for A549 cells. Furthermore, --stack images were acquired to confirm the internalization of FITC-labelled nanoparticles. Representative images demonstrated that part of the VLPO was internalized into Calu-3 cells after incubation for 3 h, while the others adsorbed onto the cytomembrane because of their interaction with cells. 3D models of cytomemabrane (wheat) and nanoparticles (steel blue) were generated to visualize the subcellular localization of FITC-labelled nanoparticles. Isolated nuclear fragments were removed for clear visualization of the surface and dot renderings. Approximately 53% of VLPO distributed within the Calu-3 cells with distance to the cymembrane >0.5 pm, indicating successful cellular internalization of VLPO. Similarly, the fluorescence intensity of the Cy5.5- labelled VLPD was stronger than that of the Cy5.5 -labelled SLN group for Caco-2 cells after incubation for 3 hours. The cell uptake of the FITC- and Cy5.5-labelled SLN and VLPs results indicated that the fabricated nanoparticles are capable of loading cargos into or onto them.

[0378] To indicate nanoparticles permeability across the pulmonary epithelial epithelium, the diffuse ability of SLN and VLPs from a donor compartment, through a lecithin- infused artificial membrane, into an acceptor compartment is evaluated employing a parallel artificial membrane permeability assay (PAMPA) kit. As previously reported, the coefficients <1.5xl0^-6 cm/s demonstrated poor permeability, while higher values revealed good permeability (Hou, X., et al., Lipid nanoparticles for mRNA delivery. Nature Reviews Materials, 2021 ; 6(12): 1078-1094. doi: 10.1038/s41578-021-00358-0). The apparent permeability coefficient of SLN was 5.8xl0^-6 cm/s for 100 ug/mL and 2.6xl0^-6 cm/s for 200 ug/mL, demonstrating its good permeability. The coefficient values decreased significantly (P<0.001) after functionalization with Spike SI RBD protrusions derived from the SARS- CoV-2 original (1.3xl0^-6 cm/s for 100 ug/mL and 1.2xlO’⁶ cm/s for 200 ug/mL) and delta variant (1.2xl0^-6 cm/s for 100 ug/mL and l.lxlO'⁶ cm/s for 200 ug/mL). The significant decrease in permeability ability can be attributed to the functionalization with Spike SI RBD, which coved the hydrophobic lipid surface with hydrophilic proteins.

[0379] In summary, the fabricated SLN and VLPs reveraled excellent cell compatiability. The conjugated Spike SI RBD protrusions could enhance the cell uptake of VLPs, confirming the interaction between VLPs and ACE2 on the cytomembrane. The exterior surface of VLPs buried by Spike SI RBD ensured their significantly decreased permeability, which may lead to good compatibility, decrease side effects, and capacitate clinic application of VLPs. [0380] VLPs blocked SARS-CoV-2 infection through interaction with ACE2 in a dose-dependent manner

[0381] We hypothesized that the VLPs functionalized with Spike SI RBD protrusions could attach to and occupy the ACE2, and as a result, block the viral entry into host cells and prevent SARS-CoV-2 infection. A pseudoviral infection assay, that was developed for evaluating neutralizing antibodies against SARS-CoV-2, was modified to study the effect of VLPs on blocking SARS-CoV-2 infection (Editorial. Let’s talk about lipid nanoparticles. Nature Reviews Materials, 2021 ; 6(2):99-99. doi: https ://doi .org/ 10.1038/s41578-021 -00281 - 4). Briefly, after pretreating with nanoparticles (SLN and VLPs, 20 g/mL and 100 g/mL) and Spike SI RBD (1 pg/pL, Avi-His-tag, biotin-labeled), human embryonic kidney 293 cells expressing ACE2 (HEK293-ACE2) were infected with Spike (SARS-CoV-2) pseudotyped lentivirus containing a luciferase reporter system. The pseudoviral infection was measured employing a One-Step luciferase assay. The fewer pseudotyped lentivirus entered the cells, the lower intensity of emitted light. This pseudoviral infection assay is quantitative and sensitive, and can be carried out in biosafety level 2 facilities (Puranik, A., et al., Comparative effectiveness of mRNA-1273 and BNT162b2 against symptomatic SARS-CoV-2 infection. Med, 2022; 3(l):28-41.e8. doi: 10.1016/j.medj.202L 12.002).

[0382] In the pseudoviral infection assay, SLN and VLPs treatment suppressed SARS-CoV-2 pseudo virions infection in a dose-dependent manner, while the free Spike SI protein did not indicate any inhibitory capacity. As shown by the cell uptake study, SLN attached to and are internalized into cells rapidly because of their positive surface charge. The attachment of SLN could repel the pseudovirions from cells, and therefore, SLN revealed light inhibitory capacity (normalized infection as low as -60%) on the pseudoviral infection. When the nanoparticle concentration was 20 ug/mL, the inhibitory capacity of VLPO and VLPD was significantly (P < 0.05) higher than that of SLN with 0.5 h nanoparticles pretreatment; as the pretreatment time increased, negligible difference was observed. For nanoparticle concentration of 100 pg/mL, the VLPD maintained strong inhibitory capacity (normalized infection varied in the range of 24-30%) for 6 h and significantly (P < 0.001) lowered the pseudoviral infection compared with both SLN and VLPO. The more efficient blockade provided by VLPD could be ascribed to the higher binding affinity of delta Spike SI RBD with ACE2 as well as the larger amount of Spike SI RBD conjugated to each VLPD surface (Liu, Y.C., R.L. Kuo, and S.R. Shih, COVID-19: The first documented coronavirus pandemic in history. Biomedical Journal, 2020; 43(4):328-333. doi: 10. 1016/j.bj.2020.04.007). [0383] To further study the interaction between VLPs and ACE2 receptor, we investigated the effect of Spike SI RBD (Avi-His-Tag, 1 pg/pL) on the VLPs cell uptake into A549-ACE2/T . The cells were pretreated with Spike SI RBD (Avi-His-Tag, 1 pg/pL) for different times (0, 0.5, 1 , 3, 6 h), followed by incubation with FITC-labeled VLPs for 1 h. Spike SI RBD greatly (P < 0.005) reduced the cell uptake of VLPD after 0.5 h pretreatment, indicating ACE2, as the receptor of Spike S 1 RBD, was involved in the internalization process of VLPD; inhibition effect was barely observed as the pretreatment time increased, which may attribute to fast exhaustion of Spike SI RBD. Surprisingly, Spike S 1 RBD revealed negligible influence on the cell uptake of SLNO. This result demonstrated that SLNO might efficiently replace the Spike S 1 RBD bond to the ACE2 receptor, and as a result, could potentially serve as a blocker not only before but also during the SARS-CoV-2 infection progress.

[0384] By way of example, the bottom-up fabricated VLPs (e.g., carrier) comprising a particle having a maximum size in at least one dimension in the nanometer or micrometer range, forming a core, and further having a functionalized surface capable of binding to said target areas of said cell surfaces to at least temporarily block said target areas to prevent or minimize pathogen binding and thus, reducing the risk of the host contracting a disease caused by said pathogen, blocked, e.g., SARS-CoV-2 pseudoviral infection through the interaction with ACE2 in a dose-dependent manner. The VLPD (e.g., carrier) could efficiently decreased or hinder the normalized pseudoviral infection to 24-30% for 6 h. Bare inhibition effect of free Spike SI RBD was observed with SLNO cell uptake, indicating that there is an affinity balance towards the receptor and therefore different variants of the virus and particle functionalization will affect the uptake outcome. The fabricated VLPs functionalized with Spike SI RBD protrusions mimicking the highly transmissible Delta variant may be able to efficiently protect against SARSCoV-2 infection and prevent the global COVID- 19 pandemic from reoccurring.

[0385] By way of further example, two types of host-directed VLPs against S ARS- CoV-2 infection were fabricated and blocked virus entry into host cells through the ACE2 receptor. In some embodiments, the blocking efficacy of the host-directed VLPs is at least 10% blocking efficacy, preferably at least 20% blocking efficacy, preferably at least 30% blocking efficacy, more preferably over 40% blocking efficacy, preferably over 50% blocking efficacy, preferably at least 60% blocking efficacy, more preferably over 70% blocking efficacy, even more preferably about 80% blocking efficacy, or preferable close to 90% blocking efficacy. The core of the VLPs (e.g. carrier) was manufactured using a microfluidic platform from, e.g., lipid components. This platform was amenable to sufficient control upon the precipitation process, and as a result, the fabricated SLN (e.g., core) revealed narrow size distribution and high reproducibility. SLN (e.g., core) was functionalized, e.g., with abundant Spike SI RBD protrusions derived from original and delta variant SARS-CoV-2 because of the extraordinarily high affinity of biotin-streptavidin interaction. Specifically, the resultant VLPD carried significantly larger amount of Spike SI RBD protrusions on its surface than VLPO. The interaction between VLPs and ACE2 receptor was proved by their enhanced cell uptake into ACE2 expressing cells. Because of the interaction, VLPs blocked SARS-CoV-2 pseudoviral infection in a dose-dependent manner. VLPD (e.g., carrier) showed inhibitory capacity towards the viral entry (e.g., blocking), and efficiently maintained the normalized pseudoviral infection to 24-30% for up to 6 hours at a dosage of 100 pg/mL. These results highlight the VLPs potential as candidates against SARS-CoV-2 infection, shedding light on future prophylactic strategies against global COVID-19 pandemics and general prophylactic strategies to hinder pathogen entry to host cells. Furthermore, the host-directed VLPs may also provide protection against other coronaviruses employing ACE2 receptor for entry, such as HCoV-NL63 and SARS-CoV, and potentially offer implications for pathogenic outbreak control both locally and globally.¹³⁷¹ Furthermore, the host-directed VLPs (e.g. carriers) can provide protection against other pathogens employing host receptors for pathogen entry potentially offer implications for pathogenic outbreak control both locally and globally. Furthermore, the host-directed VLPs (e.g. pathogen mimicking carriers) can provide targeting strategies for drugs, API, molecules and compounds to be internalised at specific cells and/or tissues employing host receptors for high uptake of compounds at target site reducing the off-target effects common in these compounds (Sackmann, E.K., A.L. Fulton, and D.J. Beebe, The present and future role of microfluidics in biomedical research. Nature, 2014; 507(7491): 181-189. doi:

10.1038/naturel3118; Whitesides, G.M., The origins and the future of microfluidics. Nature, 2006; 442(7101):368-373. doi: 10.1038/nature05058; Zhang, P., et al., Controlled Interfacial Polymer Self-Assembly Coordinates Ultrahigh Drug Loading and Zero-Order Release in Particles Prepared under Continuous Flow. Advanced Materials, 2023; 35(22):2211254. doi: 10.1002/adma.202211254; Agrahari, V. and V. Agrahari, Facilitating the translation of nanomedicines to a clinical product: challenges and opportunities. Drug Discovery Today, 2018; 23(5):974-991. doi: 10.1016/j.drudis.2018.01 .047).

[0386] It will be understood that the above values and ranges are for purposes of example. In some embodiments, normalized pseudoviral infection can be at least 10%, preferably at least 20%. In some embodiments, the blocking efficacy can be at least 30% blocking efficacy, more preferably over 40% blocking efficacy, preferably over 50% blocking efficacy, preferably at least 60% blocking efficacy, more preferably over 70% blocking efficacy, more preferably about 80% blocking efficacy, more preferably approximately 90% blocking efficacy. In some embodiments, the normalized pseudoviral infection can be maintained at least 1 hour, at least 2 hours, at least 3 hours, at least 4 hours, at least 5 hours, at least 6 hours, at least 7 hours, at least 8 hours, at least 9 hours, at least 10 hours, at least 11 hours, at least 12 hours, over about 12 hours to about 24 hours, for prolonged over 24 hours, or any range of values therebetween. In some embodiments, the dosage can be 1 pg/mL, 10 pg/mL, 20 pg/mL, 30 pg/mL, 40 pg/mL, 50 pg/mL, 60 pg/mL, 70 pg/mL, 80 pg/mL, 90 pg/mL, 100 pg/mL, 110 pg/mL, 120 pg/mL, 130 pg/mL, 140 pg/mL, 150 pg/mL, 160 pg/mL, 170 pg/mL, 180 pg/mL, 190 pg/mL, 200 pg/mL, or any range of values therebetween. In some embodiments, the dosage can be administered in cell culture or in vitro.

[0243] Features, materials, characteristics, or groups described in conjunction with a particular aspect, embodiment, or example are to be understood to be applicable to any other aspect, embodiment or example described in this section or elsewhere in this specification unless incompatible therewith. All of the features disclosed in this specification (including any accompanying claims, abstract and drawings), and/or all of the steps of any method or process so disclosed, may be combined in any combination, except combinations where at least some of such features and/or steps are mutually exclusive. The protection is not restricted to the details of any foregoing embodiments. The protection extends to any novel one, or any novel combination, of the features disclosed in this specification (including any accompanying claims, abstract and drawings), or to any novel one, or any novel combination, of the steps of any method or process so disclosed.

[0387] The word “exemplary” is used herein to mean “serving as an example, instance, or illustration.” Any aspect or embodiment described herein as “exemplary” is not necessarily to be construed as preferred or advantageous over other aspects or embodiments. Various aspects of the novel systems, apparatuses, and methods are described more fully hereinafter with reference to the accompanying drawings. This disclosure may, however, be embodied in many different forms and should not be construed as limited to any specific structure or function presented throughout this disclosure. Rather, these aspects are provided so that this disclosure will be thorough and complete, and will fully convey the scope of the disclosure to those skilled in the art. Based on the teachings herein one skilled in the art should appreciate that the scope of the disclosure is intended to cover any aspect of the novel systems, apparatuses, and methods disclosed herein, whether implemented independently of, or combined with, any other aspect described. For example, an apparatus may be implemented or a method may be practiced using any number of the aspects set forth herein. In addition, the scope of the disclosure is intended to cover such an apparatus or method which is practiced using other structure, functionality, or structure and functionality in addition to or other than the various aspects of the disclosures set forth herein. It should be understood that any aspect disclosed herein may be embodied by one or more elements of a claim.

[0388] Furthermore, certain features that are described in this disclosure in the context of separate implementations can also be implemented in combination in a single implementation. Conversely, various features that are described in the context of a single implementation can also be implemented in multiple implementations separately or in any suitable subcombination. Although features may be described above as acting in certain combinations, one or more features from a claimed combination can, in some cases, be excised from the combination, and the combination may be claimed as a subcombination or variation of a subcombination.

[0389] The features and attributes of the specific embodiments disclosed above may be combined in different ways to form additional embodiments, all of which fall within the scope of the present disclosure. Also, the separation of various system components in the implementations described above should not be understood as requiring such separation in all implementations, and it should be understood that the described components and systems can generally be integrated together in a single product or packaged into multiple products.

[0390] Moreover, while operations may be depicted in the drawings or described in the specification in a particular order, such operations need not be performed in the particular order shown or in sequential order, or that all operations be performed, to achieve desirable results. Other operations that are not depicted or described can be incorporated in the example methods and processes. For example, one or more additional operations can be performed before, after, simultaneously, or between any of the described operations. Further, the operations may be rearranged or reordered in other implementations. Those skilled in the art will appreciate that in some embodiments, the actual steps taken in the processes illustrated and/or disclosed may differ from those shown in the figures. Depending on the embodiment, certain of the steps described above may be removed, others may be added. Furthermore, the features and attributes of the specific embodiments disclosed above may be combined in different ways to form additional embodiments, all of which fall within the scope of the present disclosure.

[0391] For purposes of this disclosure, certain aspects, advantages, and novel features are described herein. Not necessarily all such advantages may be achieved in accordance with any particular embodiment. Thus, for example, those skilled in the art will recognize that the disclosure may be embodied or carried out in a manner that achieves one advantage or a group of advantages as taught herein without necessarily achieving other advantages as may be taught or suggested herein.

[0392] Conditional language, such as “can,” “could,” “might,” or “may,” unless specifically stated otherwise, or otherwise understood within the context as used, is generally intended to convey that certain embodiments include, while other embodiments do not include, certain features, elements, and/or steps. Thus, such conditional language is not generally intended to imply that features, elements, and/or steps are in any way required for one or more embodiments or that one or more embodiments necessarily include logic for deciding, with or without user input or prompting, whether these features, elements, and/or steps are included or are to be performed in any particular embodiment.

[0393] Conjunctive language such as the phrase “at least one of X, Y, and Z,” unless specifically stated otherwise, is otherwise understood with the context as used in general to convey that an item, term, etc. may be either X, Y, or Z. Thus, such conjunctive language is not generally intended to imply that certain embodiments require the presence of at least one of X, at least one of Y, and at least one of Z. Thus, as used herein, a phrase referring to “at least one of X, Y, and Z” is intended to cover: X, Y, Z, X and Y, X and Z, Y and Z, and X, Y and Z.

[0394] The headings provided herein, if any, are for convenience only and do not necessarily affect the scope or meaning of the devices and methods disclosed herein.

[0395] Language of degree used herein, such as the terms “approximately,” “about,” “generally,” and “substantially” as used herein represent a value, amount, or characteristic close to the stated value, amount, or characteristic that still performs a desired function or achieves a desired result. For example, the terms “approximately”, “about”, “generally,” and “substantially” may refer to an amount that is within less than 10% of, within less than 5% of, within less than 1% of, within less than 0.1% of, and within less than 0.01% of the stated amount.

[0396] The scope of the present disclosure is not intended to be limited by the specific disclosures of embodiments in this section or elsewhere in this specification and may be defined by claims as presented in this section or elsewhere in this specification or as presented in the future. The language of the claims is to be interpreted broadly based on the language employed in the claims and not limited to the examples described in the present specification or during the prosecution of the application, which examples are to be construed as non-exclusive.

Claims

WHAT IS CLAIMED IS:

1. A method of manufacturing a carrier, comprising: forming a core using a microfluidics device, wherein the core comprises a mean size ranging from 100 nm to 500 nm; combining the core with at least one protrusion, wherein the protrusion is adapted to bind to a target area of a host; maintaining the core and the at least one protrusion to a temperature from about 3 °C to about 5 °C; and functionalizing the core by securing the at least one protrusion to the core.

2. The method of claim 1 , further comprising adding an additive to the core, wherein the additive comprises an API, a drug, a protein, an amino acid, a lipid, a cholesterol molecule, a polymer, a monomer, an amino acid, a protein, a salt, a mineral, RNA, DNA, or any combination thereof; wherein functionalizing comprises incubating the mixture for about 2 hours to about 5 hours; and wherein the at least one protrusion is adapted to mimic an agent, the agent is a ligand, an antigen, an agonist, a pathogen or part thereof, a virus or part thereof, a bacterium or part thereof, a fungus or part thereof, a protein, a prion, a nucleic acid, an enzyme, a toxin, an allergen, a xenobiotic agent, or any combination thereof.

3. The method of claim 1, wherein functionalizing comprises incubating the mixture for about 2 hour to about 5 hours.

4. The method of claim 1 or 3, further comprising adding an additive to the core.

5. The method of claim 4, wherein the additive comprises an API, a drug, a protein, an amino acid, a lipid, a cholesterol molecule, a polymer, a monomer, an amino acid, a protein, a salt, a mineral, RNA, DNA, or any combination thereof.

6. The method of claims 1-5, wherein forming comprises vesiculating a raw material, the raw material comprising a lipid, a cholesterol molecule, a polymer, a monomer, an amino acid, a protein, a salt, a mineral, or any combination thereof.

7. The method of claims 1-6, wherein the at least one protrusion is adapted to at least partially mimic an agent.

8. The method of claim 7, wherein the agent is a ligand, an antigen, an agonist, a pathogen or part thereof, a virus or part thereof, a bacterium or part thereof, a fungus or part thereof, a protein, a prion, a nucleic acid, an enzyme, a toxin, an allergen, a xenobiotic agent, or any combination thereof.

9. The method of claim 7, wherein the virus or part thereof comprises a coronavirus, an influenza virus, a rhinovirus, a norovirus, a respiratory syncytial virus (RSV), or any virus that impacts the respiratory system.

10. The method of claims 1-7, further comprising administering the carrier to the host.

11. The method of claims 6-8, wherein administering comprises competitively inhibiting the agent from binding to a host cell.

12. The method of claim 11, wherein competitively inhibiting comprises preventing entry of the agent into the host cell.

13. The method of claims 1-12, wherein functionalizing comprises securing the core with a plurality of protrusions.

14. The method of claims 1-13, wherein functionalizing comprises ligating the at least one protrusion to an outer surface of the core.

15. The method of claims 1-14, wherein securing comprises ligating the at least one protrusion to an outer surface of the core.

16. The method of claims 1-15, wherein ligating comprises covalent binding, non- covalent interactions, van der Waals forces, hydrophobic interactions, electrostatic interactions or polar interactions.

17. The method of claims 1-16, wherein the temperature is greater than 0 °C to less than about 10 °C.

18. The method of claims 1-17, wherein the target area of a host is a host cell.

19. The method of claims 1-18, wherein the target comprises at least one receptor, target molecule, amino acid or nucleotide.

20. The method of claims 1-19, further comprising loading the carrier with at least one material.

21. The method of claim 20, wherein the material comprises a drug, API or other pharmaceutical composition.

22. The method of claim 21, wherein the pharmaceutical composition comprises Celastrol, zinc, anti-viral compounds, Interferon- Gamma modulators, antibodies, proteins, or any combination thereof.

23. The method of claim 10, wherein administering comprises using an inhalation device, an oral tablet, or an injection device.

24. The method of claims 1-23, wherein the core comprises synthetic polymers.

25. The method of claim 24, wherein the synthetic polymers comprise polyolefins, polyesters, polylactides, polycaprolactones, polyamides, polyimides, polynitriles, or any combination thereof.

26. The method of claims 1-25, wherein the carrier is biocompatible.

27. The method of claims 1-26, wherein forming comprises 3D printing, microfluidics, sol-gel methods, genetically engineering organism to produce vesiculating biopolymers, or any combination thereof.

28. The method of claims 1-27, wherein forming comprises casting the core from polydimethylsiloxane (PDMS) using soft lithography.

29. The method of claims 1-28, further comprising purifying the carrier, wherein purifying comprises dialyzing the carrier in a solution to remove an excess of the at least one protrusion that are uncoupled with the core.

30. A microfluidic device for forming a plurality of particles or cores, comprising: a first inlet capillary having a first longitudinal axis and configured to receive a first component of particles or cores; a second inlet capillary having a second longitudinal axis, and configured to receive a second component of particles or cores; an outlet capillary configured to receive the first and second components, wherein the first longitudinal axis is angled relative to the second longitudinal axis to promote mixing of the first and second components.

31. The microfluidics device of claim 30, wherein the particles or cores are shaped by a distal tip of the outlet capillary, and wherein a size distribution of the particles or cores is from 100 nm to 500 nm, wherein the microfluidic device is configured to operate at from 0 °C to 10 °C.

32. The microfluidic device of claim 31, further comprising a connector or luer in fluid communication with the first inlet capillary, the second inlet capillary, and the outlet capillary, wherein the connector or luer is configured to promote mixing of the first and second components.

33. The microfluidic device of claims 30-32, wherein the microfluidic device is configured for use on a tabletop.

34. The microfluidic device of claims 30-33, wherein the first inlet capillary is positioned substantially perpendicular to the second inlet capillary.

35. The microfluidic device of claims 30-34, wherein the first and second inlet capillaries comprise a flow velocity of from 0.5 mL/min to 3 mL/min.

36. The microfluidic device of claims 30-35, wherein the first and second inlets are configured to deliver a fluid to the outlet capillary at a pressure higher than atmospheric pressure.

37. The microfluidic device of claims 36, wherein the pressure is from 50 psi to 300 psi.

38. The microfluidic device of claims 30-37, wherein the first and second components are delivered to the first and second inlets, respectively, by a mechanical device.

39. The microfluidic device of claim 38, wherein the mechanical device is a syringe or an electromechanically powered device.

40. The microfluidic device of claims 30-39, wherein the first and second inlet are configured to receive a fluid from a first reservoir and a second reservoir, respectively.

41. The microfluidic device of claim 40, wherein the first and second reservoirs comprise the first and second components.

42. The microfluidic device of claim 30-41, wherein the first inlet and the second inlet comprise an inner diameter of 500 to 700 pm.

43. The microfluidic device of claim 30-42, wherein the first inlet and the second inlet comprise an outer diameter of 800 to 1200 pm.

44. The microfluidic device of claim 30-43, wherein the distal tip is polished and smooth.

45. The microfluidic device of claims 30-44, further comprising external equipment, wherein the microfluidic device is configured to engage the external equipment.

46. The microfluidic device of claims 30-45, wherein the external equipment comprises fluid pumps, compressed gas lines, computers, microscopes, electrical supply outlets, or any combination thereof.

47. A system for functionalizing a carrier, comprising the microfluidic device of claims 30-46, and a third reservoir, wherein the third reservoir comprises a functionalization medium.

48. The system of claim 47, wherein the collection chamber comprises a volume of an aqueous solution.

49. The system of claims 35-48, wherein the third reservoir is configured to engage the collection chamber.

50. The system of claims 35-49, wherein the functionalization medium comprises a solution.

51. The system of claim 35-50, wherein the functionalization medium is a solid.

52. The system of claim 35-51, wherein the functionalization medium comprises at least one protrusion.

53. The system of claim 35-52, further comprising a pump.

54. The system of claim 35-53, further comprising a heat exchanger.

55. The system of claim 54, wherein the heat exchanger is configured to maintain a temperature of the collection chamber of about 3 °C to about 5 °C.

56. The system of claim 35-55, wherein the microfluidic device is configured to fabricate a core.