# US 11,433,142 B2 · Compositions and methods for delivery of RNA-lipid nanoparticle complexes encoding for viral RNA polymerase region and protein antigen

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- Number: US 11,433,142 B2
- Document ID: US11433142B2
- Status: Granted
- Kind: B2
- Application number: 17/702,730
- Filed: 2022-03-23
- Published: 2022-08-11
- Granted: 2022-09-06
- Earliest claimed: 2020-03-23
- Assignees: HDT Bio Corp.
- Applicants: HDT Bio Corp.
- Inventors: Amit Khandhar; Steven Reed; Malcolm Duthie; Jesse Erasmus; Darrick Carter; Bryan J. Berube
- Web page: https://patentgrep.com/patent/US11433142B2

## CPC

- A61K 39/12: Viral antigens
- A61K 47/6907: {the form being a microemulsion, nanoemulsion or micelle}
- A61K 9/1075: {Microemulsions or submicron emulsions; Preconcentrates or solids thereof; Micelles, e.g. made of phospholipids or block copolymers (A61K9/0026 takes precedence)}
- A61K 9/127: Synthetic bilayered vehicles, e.g. liposomes or liposomes with cholesterol as the only non-phosphatidyl surfactant
- A61K 9/5115: {Inorganic compounds}
- A61K 9/5123: {Organic compounds, e.g. fats, sugars}
- A61K 31/7105: Natural ribonucleic acids, i.e. containing only riboses attached to adenine, guanine, cytosine or uracil and having 3'-5' phosphodiester links
- A61K 31/711: Natural deoxyribonucleic acids, i.e. containing only 2'-deoxyriboses attached to adenine, guanine, cytosine or thymine and having 3'-5' phosphodiester links
- A61K 31/713: Double-stranded nucleic acids or oligonucleotides
- A61K 39/215: Coronaviridae, e.g. avian infectious bronchitis virus
- A61K 39/39: characterised by the immunostimulating additives, e.g. chemical adjuvants
- A61K 47/02: Inorganic compounds
- A61K 47/44: Oils, fats or waxes according to two or more groups of A61K47/02-A61K47/42; Natural or modified natural oils, fats or waxes, e.g. castor oil, polyethoxylated castor oil, montan wax, lignite, shellac, rosin, beeswax or lanolin (synthetic glycerides, e.g. medium-chain triglycerides, A61K47/14)
- A61K 47/6911: {the form being a liposome}
- A61K 47/6923: {the form being an inorganic particle, e.g. ceramic particles, silica particles, ferrite or synsorb}
- A61K 49/183: {having a (super)(para)magnetic core coated or functionalised with an inorganic material or being composed of an inorganic material entrapping the MRI-active nucleus, e.g. silica core doped with a MRI-active nucleus}
- A61K 49/1833: {having a (super)(para)magnetic core coated or functionalised with a small organic molecule (oligomeric, polymeric, dendrimeric A61K49/1851)}
- A61P 31/14: for RNA viruses
- C07K 14/165: Coronaviridae, e.g. avian infectious bronchitis virus
- C07K 16/116: Togaviridae (F); Matonaviridae (F); Flaviviridae (F)
- C12N 15/86: Viral vectors
- C12N 15/88: using microencapsulation, e.g. using {amphiphile} liposome vesicle
- A61K 47/10: Alcohols; Phenols; Salts thereof, e.g. glycerol; Polyethylene glycols [PEG]; Poloxamers; PEG/POE alkyl ethers
- A61K 47/26: Carbohydrates, e.g. sugar alcohols, amino sugars, nucleic acids, mono-, di- or oligo-saccharides; Derivatives thereof, e.g. polysorbates, sorbitan fatty acid esters or glycyrrhizin
- A61K 47/547: {Chelates, e.g. Gd-DOTA or Zinc-amino acid chelates; Chelate-forming compounds, e.g. DOTA or ethylenediamine being covalently linked or complexed to the pharmacologically- or therapeutically-active agent}
- A61K 2039/53: {DNA (RNA) vaccination}
- A61K 2039/55505: {Inorganic adjuvants}
- A61K 2039/55516: {Proteins; Peptides}
- A61K 2039/55555: {Liposomes; Vesicles, e.g. nanoparticles; Spheres, e.g. nanospheres; Polymers}
- A61K 2039/55561: {CpG containing adjuvants; Oligonucleotide containing adjuvants}
- B82Y 5/00: Nanobiotechnology or nanomedicine, e.g. protein engineering or drug delivery
- B82Y 25/00: Nanomagnetism, e.g. magnetoimpedance, anisotropic magnetoresistance, giant magnetoresistance or tunneling magnetoresistance
- B82Y 30/00: Nanotechnology for materials or surface science, e.g. nanocomposites
- B82Y 40/00: Manufacture or treatment of nanostructures
- C12N 2770/20034: Use of virus or viral component as vaccine, e.g. live-attenuated or inactivated virus, VLP, viral protein
- C12N 2770/36143: viral genome or elements thereof as genetic vector
- Y02A 50/30: Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change
- A61K: PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES (devices or methods specially adapted for bringing pharmaceutical products into particular physical or administering forms A61J3/00; chemical aspects of, or use of materials for deodorisation of air, for disinfection or sterilisation, or for bandages, dressings, absorbent pads or surgical articles A61L; soap compositions C11D)
- A61P: SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- C07K: PEPTIDES (peptides in foodstuffs A23; obtaining protein compositions for foodstuffs, working-up proteins for foodstuffs A23J; preparations for medicinal purposes A61K; peptides containing beta-lactam rings C07D; cyclic dipeptides not having in their molecule any other peptide link than those which form their ring, e.g. piperazine-2,5-diones, C07D; ergot alkaloids of the cyclic peptide type C07D519/02; macromolecular compounds having statistically distributed amino acid units in their molecules, i.e. when the preparation does not provide for a specific; but for a random sequence of the amino acid units, homopolyamides and block copolyamides derived from amino acids C08G69/00; macromolecular products derived from proteins C08H1/00; preparation of glue or gelatine C09H; single cell proteins, enzymes C12N; genetic engineering processes for obtaining peptides C12N15/00; compositions for measuring or testing processes involving enzymes C12Q; investigation or analysis of biological material G01N33/00)
- C12N: MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA (microbiological testing media C12Q1/00)
- B82Y: SPECIFIC USES OR APPLICATIONS OF NANOSTRUCTURES; MEASUREMENT OR ANALYSIS OF NANOSTRUCTURES; MANUFACTURE OR TREATMENT OF NANOSTRUCTURES
- Y02A: TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE

## Abstract

The disclosure provides nanoemulsion compositions and methods of making and using thereof to deliver a bioactive agent such as a nucleic acid to a subject. Compositions of the disclosure include RNA-lipid nanoparticle complexes encoding for viral RNA polymerase region and protein antigen. Compositions are further described where the lipid nanoparticles are characterized as having a z-average diameter particle size measurement of 20 nm to 80 nm when measured using dynamic light scattering. The nanoemulsion compositions of the disclosure comprises a hydrophobic core, optionally including inorganic nanoparticles, in a lipid nanoparticle that allows imaging as well as delivering nucleic acids. Methods of using these particles for treatment and vaccination are also provided.

## Claims

### Claim 1

A composition, wherein the composition comprises:
RNA molecules, wherein the RNA molecules comprise a sequence encoding for:
an RNA polymerase complex region from an RNA virus; and
a protein antigen; and
lipid nanoparticles, wherein the lipid nanoparticles are characterized as having a z-average diameter particle size measurement of 20 nm to 80 nm when measured using dynamic light scattering, and wherein the lipid nanoparticles comprise:
a surface comprising cationic lipids; and
a hydrophobic core, wherein the hydrophobic core comprises liquid oil,
wherein lipids present in hydrophobic core are in liquid phase at 25 degrees Celsius, and
wherein the RNA molecules are complexed to the cationic lipids to form RNA-lipid nanoparticle complexes.

### Claim 2 (depends on 1)

The composition of claim 1, wherein the protein antigen is derived from a virus.

### Claim 3 (depends on 2)

The composition of claim 2, wherein the virus is an RNA virus.

### Claim 4 (depends on 3)

The composition of claim 3, wherein the RNA virus is a Human Immunodeficiency Virus (HIV) virus or a hepatitis virus.

### Claim 5 (depends on 3)

The composition of claim 3, wherein the RNA virus is a Coronavirus.

### Claim 6 (depends on 5)

The composition of claim 5, wherein the Coronavirus is a MERS Coronavirus or a SARS Coronavirus.

### Claim 7 (depends on 5)

The composition of claim 5, wherein the Coronavirus is SARS-CoV-2 virus.

### Claim 8 (depends on 3)

The composition of claim 3, wherein the RNA virus is a mosquito-borne virus.

### Claim 9 (depends on 8)

The composition of claim 8, wherein the mosquito-borne virus is Venezuelan equine encephalitis (VEE) virus.

### Claim 10 (depends on 8)

The composition of claim 8, wherein the mosquito-borne virus is a flavivirus.

### Claim 11 (depends on 10)

The composition of claim 10, wherein the flavivirus is Zika Virus.

### Claim 12 (depends on 1)

The composition of claim 1, wherein the protein antigen is derived from a cancer.

### Claim 13 (depends on 1)

The composition of claim 1, wherein the protein antigen is derived from a bacterium.

### Claim 14 (depends on 1)

The composition of claim 1, wherein the lipid nanoparticles are characterized as having a z-average diameter particle size measurement of 40 nm to 60 nm when measured using dynamic light scattering.

### Claim 15 (depends on 1)

The composition of claim 1, wherein the RNA-lipid nanoparticles complexes are characterized as having a z-average diameter particle size measurement of about 90 nm when measured using dynamic light scattering.

### Claim 16 (depends on 1)

The composition of claim 1, wherein the surface further comprises a hydrophobic surfactant and a hydrophilic surfactant.

### Claim 17 (depends on 16)

The composition of claim 16, wherein the hydrophobic surfactant is selected from the group consisting of sorbitan monostearate, sorbitan monooleate, and sorbitan trioleate.

### Claim 18 (depends on 16)

The composition of claim 16, wherein the hydrophilic surfactant comprises polyethylene oxide.

### Claim 19 (depends on 18)

The composition of claim 18, wherein the polyethylene oxide comprises polyoxyethylene sorbitan ester.

### Claim 20 (depends on 16)

The composition of claim 16, wherein the hydrophilic surfactant comprises polysorbate.

### Claim 21 (depends on 20)

The composition of claim 20, wherein the polysorbate comprises polysorbate 80.

### Claim 22 (depends on 1)

The composition of claim 1, wherein the cationic lipids comprise 1,2-dioleoyloxy-3-(trimethylammonium)propane (DOTAP); 3b-[N-(N′,N′-dimethylaminoethane)-carbamoyl]cholesterol (DC Cholesterol); dimethyldioctadecylammonium (DDA); 1,2-dimyristoyl-3-trimethylammoniumpropane (DMTAP); dipalmitoyl(C16:0)trimethyl ammonium propane (DPTAP); distearoyltrimethylammonium propane (DOTAP); N-[1-(2,3-dioleyloxy)propyl]-N,N,N-trimethylammonium chloride (DOTMA); N,N-dioleoyl-N,N-dimethylammonium chloride (DODAC); 1,2-dioleoyl-sn-glycero-3-ethylphosphocholine (DOEPC); 1,2-dioleoyl-3-dimethylammonium-propane (DODAP); 1,2-dilinoleyloxy-3-dimethylaminopropane (DLinDMA); or 1, r-((2-(4-(2-((2-(bis(2-hydroxydodecyl)amino)ethyl) (2-hydroxydodecyl)amino)ethyl)piperazin-1-yl)ethyl)azanediyl)bis(dodecan-2-ol) (C 12-200).

### Claim 23 (depends on 1)

The composition of claim 1, wherein the cationic lipids are DOTAP.

### Claim 24 (depends on 1)

The composition of claim 1, wherein the hydrophobic core further comprises a solid inorganic nanoparticle.

### Claim 25 (depends on 24)

The composition of claim 24, wherein the solid inorganic nanoparticle is selected from the group consisting of metal salts, metal oxides, metal hydroxides, and metal phosphates.

### Claim 26 (depends on 25)

The composition of claim 25, wherein the solid inorganic nanoparticle comprises iron oxide.

### Claim 27 (depends on 26)

The composition of claim 26, wherein the iron oxide is magnetite (Fe3O4), maghemite (y-Fe2O3), wüstite (FeO), hematite (a-Fe2O3), or any combination thereof.

### Claim 28 (depends on 1)

The composition of claim 1, wherein the liquid oil comprises vegetable oil, animal oil, or synthetically prepared oil.

### Claim 29 (depends on 1)

The composition of claim 1, wherein the liquid oil comprises squalene, sunflower oil, soybean oil, olive oil, grapeseed oil, capric/caprylic triglyceride, vitamin E, lauroyl polyoxylglyceride, or monoacylglycerol.

### Claim 30 (depends on 1)

The composition of claim 1, wherein the hydrophobic core further comprises iron oxide, wherein the surface further comprises sorbitan monostearate and polysorbate 80, wherein the liquid oil is squalene, and wherein the cationic lipid is DOTAP.