WO2022159319A2

WO2022159319A2 - A porous aluminum nanoparticulate structure

Info

Publication number: WO2022159319A2
Application number: PCT/US2022/012224
Authority: WO
Inventors: Michael Farber
Original assignee: Mountain Valley Md Inc
Priority date: 2021-01-19
Filing date: 2022-01-13
Publication date: 2022-07-28
Also published as: WO2022159319A3

Abstract

A porous aluminum nanoparticulate structure comprising an aluminum salt, a capping agent and a water-soluble polymer from 0.001-2% of an adjuvant solution is disclosed. Moreover, the capping agent is selected from group consisting of Hydroxypropylbetacyclodextrin (HPBCD), Sulfobutyletherbetacyclodextrin (SBECD) or combination with water- soluble polymers such as PAA, Polysorbate 80 and further the capping agent is associated with the aluminum salt. Particularly, the size of the porous aluminum nanoparticulate ranges from about 1 nm to about 200 nm. Additionally the porous aluminum nanoparticulate structure is stable in a liquid formulation at about 0°C to about 8°C for at least about 1 month, at least about 6 months, or at least about 1 year. The disclosed technology further relates to compositions and methods of preparing a porous aluminum nanoparticulate structure.

Description

TITLE

A Porous Aluminum Nanoparticulate structure

TECHNICAL FIELD

[1] The disclosed technology relates to the field of pharmaceutical and vaccine formulations. More specifically, the disclosed technology described herein relate to porous aluminum nanoparticulate structures, compositions comprising the porous aluminum nanoparticulates, and methods of preparing and using the porous aluminum nanoparticulates.

BACKGROUND

[2] Vaccination is a biological process in which the antigenic materials are stimulated in an individual’s immune system in order to develop immunity against a specific pathogen. Vaccination has been reported to be the most effective tool to prevent and treat illness from infectious diseases causing death. Furthermore, vaccination has been a major contributor to the global control of infectious diseases in the human population. According to the estimated data in the United States, vaccination has prevented more than 100 million cases of infectious diseases in people since the 1920s. In the recent past, advancements in the field of biotechnology and pharmaceutical have led to an increasingly rapid identification of new vaccine antigens and the generation of more effective and safer vaccines. However, the highly purified recombinant antigens are poorly immunogenic and hence require adjuvants in order to increase the level and duration of protection induced by vaccines. [3] As numerous recombinant and synthetic antigens are poorly immunogenic, adjuvant is added to vaccine formula in order to potentiate the immune responses, offer better protection against pathogens and decrease the amount of antigens needed for protective immunity. During the early stages of the development of a potential new vaccine, the need for adjuvants is critically assessed. Till date, there are about 100 different types of adjuvants for 400 different vaccines. The adjuvants possess an excellent safety profile and are further associated with minimal reactogenicity and are inexpensive too. Also, they provide a practical advantage by offering the possibility of creating single liquid vials or prefilled syringe formats. Furthermore, the immune enhancement effect of precipitation of diphtheria toxoid with insoluble aluminum salts was first reported in 1926. Since then, aluminum-containing adjuvants have been incorporated into billions of doses of vaccines and administered to millions of people.

[4] Aluminum salts have been used in vaccines for over eight decades due to their good safety profile and their ability to induce an enhanced immune response to adsorbed vaccine antigens. As one of the few classes of adjuvants approved by the US FDA, aluminum salts have an established regulatory pathway as opposed to more novel adjuvant formulations. When dispersed in an aqueous solution, aluminum salts form heterogeneous aggregate particulates of -0.5- 10 microns (pm) in size, which may make them difficult to characterize for quality control compared to formulations with monodisperse size populations such as oil-in-water emulsions. This complexity is compounded by the fact that there are multiple types of aluminum salts available with distinct properties, including aluminum phosphate, aluminum hydroxyphosphate sulfate, and aluminum oxyhydroxide.

[5] Several studies have proposed that the average particle size of an adjuvant formulation is a critical factor that can affect the biological activity of the vaccine. Recently, novel synthetic approaches have been employed using aluminum salts to de novo manufacture new synthetic formulations containing alum nanoparticles. These synthetic nanoparticles have been described to generate a stronger immune response while decreasing inflammation at the injection site, when compared to micro particles. Nevertheless, in each of these studies, a bottom-up synthetic approach was employed to manufacture the aluminum particles, and no comparison was made to clinical aluminum salt adjuvants such as Alhydrogel®, making it difficult to interpret the value of the novel formulations versus the clinically approved material.

[6] Additionally, from a regulatory perspective, clinical aluminum -based micro particles are not capable of being terminally sterilized by filtration through 0.20 micron filters, and are only sterlizable by radiation or autoclave; making their manufacture not amenable to a terminal sterilization step when combined with antigens or adjuvants. There exists a need to provide aluminum-based nanoparticles that display little to no aggregation, or reduced aggregation, and are capable of being terminally sterilized prior to being vialed.

[7] Presently, numerous nanomaterial-based adjuvants are available that have shown to be effective in order to enhance the immune responses induced by a variety of antigens. However, there are some challenges on safety and molecular mechanisms, which needs to be addressed. [8] A number of solutions were introduced in order to solve the abovementioned problems. In one of the closest relevant arts, EP2550235B1 which discloses a method for manufacturing a highly porous, stable metal oxide, wherein the metal oxide is an aluminum oxide catalyst support, comprising: mixing an aluminum alkoxide with water to form a solvent deficient precursor mixture; allowing the aluminum alkoxide and the water to react in the solvent deficient precursor mixture to form an intermediate hydroxide product; causing the intermediate hydroxide product to form intermediate nanoparticles; and calcining the intermediate nanoparticles to yield a stable aluminum oxide having a pore structure. The porous materials are made in a three-step process. In a first step, solvent deficient precursor mixtures is formed from an aluminium alkoxide and water the aluminium alkoxide and the water react in the solvent deficient precursor mixture to form an intermediate hydroxide product.

[9] Another prior art reference, EP 259844 IB 1 discloses a superficially porous material comprising a substantially nonporous core material and one or more layers of uniform thickness of a porous shell material surrounding the core material, wherein the porous shell material is a porous inorganic /organic hybrid material. The material of the disclosed technology comprises of superficially porous particles, wherein the material of the disclosed technology comprises a composite material, the composite material comprises a magnetic additive material. Moreover, the magnetic additive material has a mass magnetization at room temperature greater than 15 emu/g. The magnetic additive material is a ferromagnetic material. The substantially nonporous core material is a composite material that comprises a magnetic additive material; a composite material that comprises a high thermal conductivity additive selected from the group consisting of crystalline or amorphous silicon carbide, aluminum, gold, silver, iron, copper, titanium, niobium, diamond, cerium, carbon, zirconium, barium, cobalt, europium, gadolinium, nickel, samarium, silicon, zinc, boron, and oxide or a nitride thereof, and combinations thereof.

[10] Yet another prior art reference, US9765271B2 discloses an energetic nanoparticle compositions and materials containing silicon and other energetic elements, and methods of manufacturing the same, including reacting silicon nanoparticles and unsaturated alkene or alkyne to form covalently bonded surface coatings passivated against surface oxidation, for combination with a fuel, explosive or oxidizer, nanoparticles, compositions, manufacture and applications, particularly including silicon nanoparticle manufacture, designed multilayered energetic nanoparticles, and graphene nanoenergetics, as well as and their uses, compositions, composites including structural and energetic composites, and methods for making such materials.

[11] Yet another reference, CN103276389B relates to a kind of aluminum oxide and strengthen aluminum-based in-situ composite materials and preparation method thereof with zirconium diboride, by ZrO₂ Granule and B or B₂O₃ granule mixing and ball milling; hybrid particles after ball milling is added in matrix; the particles filled region of matrix is stirred friction processing; stirring tool rotary speed is 600~3000r/min; gait of march is 30~ 60mm/min, and volume under pressure is 0.05 ~ 0.6mm, and inclination angle is 0~3 °; processing number of times is 3~8 times, ZrO in the course of processing, B or B₂O₃. Jointly there is chemical reaction in granule and aluminum substrate, finally gives aluminium oxide and strengthen aluminum- based in-situ composite materials with zirconium diboride. In aluminum matrix composite prepared by the cited art, original position synthesis particle A1₂O₃ and ZrB₂. It is respectively provided with high hardness and heat stability so that such composite has higher high temperature wear resistant performance.

[12] The conventional methods of manufacturing highly porous, stable metal oxide and nanoparticle compositions as disclosed in the prior arts does not provide a stable aluminum-based nanoparticle that displays little to no aggregation, or reduced aggregation, and is capable of being terminally sterilized prior to being vialed.

[13] Moreover, the disclosed technology further provides an improved Aluminum salts used as co-adjuvants are advantageous because they have a good safety record, augment antibody responses, stabilize antigens, and are relatively simple for large-scale production and the aluminum salt is Alhydrogel®, an aluminum hydroxide or aluminum oxyhydroxide. Alhydrogel® has an overall positive charge and can readily adsorb negatively charged moieties. Furthermore, the disclosed technology provides an aluminum-based nanoparticle that displays little to no aggregation, or reduced aggregation, and is capable of being terminally sterilized prior to being vialed.

[14] All the problems, disadvantages and the limitations of the above mentioned relevant and conventional arts being overcome by the structure, composition and method of the disclosed technology which has various technical advancements and certainly economic benefits over the conventional arts. SUMMARY OF THE DISCLOSED TECHNOLOGY

[15] An objective of the present disclosed technology is to provide a porous aluminum nanoparticulate structure, compositions comprising the porous aluminum nanoparticulates, and methods of preparing and using the porous aluminum nanoparticulates. Moreover, the porous aluminum nanoparticulate structures are useful in the field of pharmaceuticals and/or vaccine formulations. Provided here are compositions comprising a plurality of porous aluminum nanoparticulate structures comprising an aluminum salt, a capping agent, wherein the capping agent is selected from group consisting of Hydroxypropylbetacyclodextrin (HPBCD) , Sulfobutyletherbetacyclodextrin (SBECD) or combination with water-soluble polymers such as PAA, Polysorbate 80. Furthermore, the capping agent is associated with the aluminum salt and the size of the particles in the composition is less than 1 pm. The term porous aluminum nanoparticulate is used herein to denote that the particle comprises aluminum and has a size measured in nanometers, typically from 1 nm to about 200 nm. In some embodiments of the disclosed technology, the composition is for a terminal sterilization by filter for products according to FDA regulation (such as use of a <0.20 micron filter). In some embodiments of the disclosed technology, the size of the particulates present in the composition ranges from about Inm to about 200nm. In some embodiments of the disclosed technology, the average size of the porous aluminum nanoparticulate in the composition ranges from about Inm to about 200nm. In some embodiments, the average size of the porous aluminum nanoparticulate in the composition ranges from about Inm to about 200nm. Porous aluminum nanoparticulate structures compositions described here may be produced by processing or milling aluminum hydroxide in the presence of the capping agent by standard techniques known in the art including, but not limited to, microfluidization, sonication, and high shear mixing. High shear mixing can be performed using a high shear mixer. Silverson is one company that produces high shear mixers that can be used in the present methods.

[16] The porous aluminum nanoparticulate in the compositions are stable and display little to no aggregation, or reduced aggregation, and are amenable to a terminal sterilization step prior to vialing. The porous aluminum nanoparticulate structures provided herein are useful for the delivery of an agent, such as a polypeptide or a polynucleotide or a capsid such as Inactivated Polio to an individual. By way of example only, the provided herein are useful for the delivery of antigens and/or adjuvants to a host in order to generate an immune response.

[17] In one of the embodiments, the disclosed technology mainly pertains to a porous aluminum nanoparticulates structure comprising an aluminum salt, a capping agent, wherein the capping agent is selected from group consisting of Hydroxypropylbetacyclodextrin (HPBCD) , Sulfobutyletherbetacyclodextrin (SBECD) or combination with water-soluble polymers such as PAA, Polysorbate 80, and the capping agent is associated with said aluminum salt. Moreover, the size of the porous aluminum nanoparticulate ranges from about 1 nm to about 200 nm. The porous aluminum nanoparticulate further comprises a water-soluble polymer from 0.001 - 2% of the adjuvant solution, wherein the porous aluminum nanoparticulate structure is stable in a liquid formulation at about 0°C to about 8°C for at least about 1 month, at least about 6 months, or at least about 1 year.

[18] In another embodiment, the disclosed technology discloses that the average size of the particles is the Z-average as determined by dynamic light scattering.

[19] In another important embodiment, the disclosed technology discloses that the aluminum salt is selected from the group consisting of aluminum hydroxide, aluminum hydroxide gel, A1PO₄, AIO(OH), A1(OH)(PO₄) and KAI (S0₄)₂ .In yet another important technical aspect of the disclosed technology, the porous aluminum nanoparticulate structure is in a liquid formulation which is filter-sterilized and is stable after repeated freeze-thaw cycles. Furthermore, the adjuvant solution is dried and reconstituted forming a nano-porous structure as illustrated on the electron micrographs. As a result, the structures remain stable during three freeze-thaw cycles.

[20] In certain embodiments of the disclosed technology, the porous aluminum nanoparticulate structure is in a liquid formulation which is filter- sterilized.

[21] In certain embodiments of the disclosed technology, the porous aluminum nanoparticulates structure is stable in a liquid formulation at about 0°C to about 8°C for at least about 1 month, at least about 6 months, or at least about 1 year. In certain embodiments, the porous aluminum nanoparticulates structure is stable in a liquid formulation at about 37°C for at least about 1 month. [22] In one of the embodiment of disclosed technology, the disclosed technology discloses a method of preparing a porous aluminum nanoparticulate structure comprising (a) subjecting an aluminum salt to a high energy source in the presence of a capping agent whereby a porous aluminum nanoparticulates structure is produced, with a size ranging from about 1 nm to about 200 nm, wherein aluminum salt comprises of particles ranging from 0.5 to 10 pm in size or 0.5 to 20 pm in size; and (b) mixing a capping agent with the porous aluminum nanoparticulate structure after step (a); and (c) filter- sterilizing the porous aluminum nanoparticulate structure and wherein the size of the porous aluminum nanoparticulate ranges from about 1 nm to about 200 nm.

[23] As will be appreciated by skilled artisan, the porous aluminum nanoparticulates of the disclosed technology can be produced from larger particles of micrometer size. Accordingly, the present disclosure provides a method of preparing the described porous aluminum nanoparticulates from precursor aluminum salt particles, commonly known as Alhydrogel or Adju- phos that are 0.5 pm to 20 pm in size or 0.5 pm to 10 pm in size.

[24] In certain embodiments of the disclosed technology, the high energy source is generated from a microfluidizer, an extruder, a sonicator, high shear mixer (e.g., silverson mixer), or a homogenizer. Moreover, two or more high energy sources can be used. For example, the high energy source can be generated from a microfluidizer and a high shear mixer and the mixture comprising the aluminum salt and capping agent can be passed through the microfluidizer for one or more passes (e.g., from one pass to about 30 or more passes). In certain embodiments, the high energy source is generated from a microfluidizer, and the mixture comprising the aluminum salt and capping agent is passed through the microfluidizer from one pass to about 15 passes. In certain embodiments, the aluminum salt is selected from the group consisting of aluminum hydroxide, aluminum hydroxide gel, A1PO₄, AIO(OH), A1(OH)(PO₄), and Kal(S0₄)₂ .

[25] In yet another embodiment of the disclosed technology, a porous aluminum nanoparticulate structures is obtainable or produced by a method disclosed herein, wherein the size of the porous aluminum nanoparticulate ranges from about 1 nm to about 200 nm.

[26] In another embodiment, the disclosed technology provides a composition comprising the porous aluminum nanoparticulates structures disclosed herein.

[27] In certain embodiments of the disclosed technology, the composition further comprises a bioactive agent, wherein more than 75% of the bioactive agent is associated with said porous aluminum nanoparticulate structure as determined by gel electrophoresis. In certain embodiments, the bioactive agent is associated with the porous aluminum nanoparticulate structure in the composition. Furthermore, the composition of the disclosed technology further comprises an adjuvant, wherein the adjuvant is selected from the group consisting of a AS-2, monophosphoryl lipid A, 3-de-O-acylated monophosphoryl lipid A, IFA, QS21, CWS, TOM, AGPs, CpG-containing oligonucleotides, Toil-like receptor (TLR) agonists, Leif, saponins, saponin mimetics, biological and synthetic lipid A, imiquimod, gardiquimod, resiquimod, polyLC, ilagellin, GLA, SLA, STING, and combinations thereof. Furthermore, the composition further comprises a liposome, wherein the average size of the porous aluminum nanoparticulates in the composition ranges from about 1 nm to about 200 nm and wherein the composition is capable of being filtered through a 0.20 micron-sized filter. Moreover, the composition is capable of being terminally sterilized prior to vialing.

[28] In certain embodiments of the disclosed technology, the bioactive agent is a polypeptide, a polynucleotide, an antigen, an adjuvant, a diagnostic agent, a therapeutic agent, or an organism.

[29] In yet another embodiment of the disclosed technology, the composition is stable at about 0°C to about 8°C for at least about 1 month, at least about 6 months, or at least about 1 year. In certain embodiments, the composition is stable at about 37°C for at least about 1 month. In certain embodiments, the average size of the porous aluminum nanoparticulates in the composition is from about 1 nm to about 200 nm.

[30] In another embodiment, the disclosed technology discloses a method of stimulating an immune response in a subject comprising administering the composition disclosed herein to a subject, whereby stimulating an immune response in the subject involves the activation of B-cells, activation of T cells, production of antibodies, or release of cytokines.

[31] In certain embodiments of the disclosed technology, the immune response is a non-specific immune response and/or an antigen-specific immune response and/or a TH1 immune response and/or a TH2 immune response and/or both TH1 and TH2 immune response.

[32] In certain embodiments of the disclosed technology, the composition is used for the treatment of allergy, addiction, cancer, or autoimmunity. [33] In certain embodiments of the disclosed technology, the route of administration of the composition is oral, intravenous, intradermal, transdermal, nasal, subcutaneous, or anal.

[34] In another embodiment, the disclosed technology provides a method of delivering a bioactive agent to a cell in a subject comprising administering to the subject a composition comprising (a) a porous aluminum nanoparticulates structures comprising an aluminum salt and a capping agent, wherein the size of the particle ranges from about 1 nm to about 200 nm and (b) a bioactive agent, thereby delivering the bioactive agent to the cell in the subject.

[35] In yet another embodiment, the present disclosure provides a method of preparing a composition comprising, (a) subjecting an aluminum salt to a high energy source in the presence of a capping agent, whereby a porous aluminum nanoparticulate is produced, and wherein the size of the porous aluminum nanoparticulate ranges from about 1 nm to about 200 nm, and (b) mixing the porous aluminum nanoparticulate produced in step (a) with a bioactive agent.

[36] Any device or step to a method described in this disclosure can comprise, or consist of, that which it is a part of, or the parts which make up the device or step. The term “and/or” is inclusive of the items which it joins linguistically and each item by itself. Any element or described portion of the devices shown can be “substantially” as such, if used in the claims in this manner. Where used, “substantially” is defined as “within a 5% tolerance level thereof.”

BRIEF DESCRIPTION OF THE DRAWINGS [37] FIG. 1 is an electron micrograph depicting various embodiments of the disclosed technology comprising a porous aluminum nanoparticulate structure.

[38] FIG. 2 is another electron micrograph depicting various embodiments of the disclosed technology comprising a porous aluminum nanoparticulate structure.

[39] FIG. 3 is another electron micrograph depicting various embodiments of the disclosed technology comprising a porous aluminum nanoparticulate structure.

DETAILED DESCRIPTION OF THE DISCLOSED TECHNOLOGY

[40] The embodiments herein and the various features and advantageous details thereof are explained more fully with reference to the non-limiting embodiments that are illustrated in the accompanying drawings and detailed in the following description. Descriptions of well-known components and processing techniques are omitted so as to not unnecessarily obscure the embodiments herein. The description used herein are intended merely to facilitate an understanding of ways in which the embodiments herein may be practiced and to further enable those of skill in the art to practice the embodiments herein. Accordingly, the description or explanation should not be construed as limiting the scope of the embodiments herein.

[41] An embodiment of the disclosed technology is to provide a porous aluminum nanoparticulate structure, compositions comprising the porous aluminum nanoparticulates, and methods of preparing and using the porous aluminum nanoparticulates. Moreover, the porous aluminum nanoparticulate structures are useful in the field of pharmaceuticals and/or vaccine formulations. Provided herein are compositions comprising a plurality of porous aluminum nanoparticulate structures comprising an aluminum salt, a capping agent, wherein the capping agent is selected from group consisting of Hydroxypropylbetacyclodextrin (HPBCD) , Sulfobutyletherbetacyclodextrin (SBECD) or combination with water-soluble polymers such as PAA, Polysorbate 80. Furthermore, the capping agent is associated with the aluminum salt and the size of the particles in the composition is less than 1 pm. The term porous aluminum nanoparticulate is used herein to denote that the particle comprises aluminum and has a size measured in nanometers, typically from 1 nm to about 200 nm.

[42] In some embodiments of the disclosed technology, the composition is for a terminal sterilization by filter for products according to FDA regulation (such as use of a <0.20 micron filter).

[43] In some embodiments of the disclosed technology, the size of the particulates present in the composition ranges from about 1 nm to about 200 nm. In some embodiments of the disclosed technology, the average size of the porous aluminum nanoparticulate in the composition ranges from about Inm to about 200 nm.

[44] In some embodiments, the average size of the porous aluminum nanoparticulate in the composition ranges from about nm to about 200nm. Porous aluminum nanoparticulate structures compositions described here is produced by processing or milling aluminum hydroxide in the presence of the capping agent by standard techniques known in the art including, but not limited to, microfluidization, sonication, and high shear mixing. High shear mixing can be performed using a high shear mixer. Silverson is one company that produces high shear mixers that can be used in the present methods.

[45] The present disclosure described herein provides porous aluminum nanoparticulate structures, compositions comprising the porous aluminum nanoparticulates, and methods of preparing and using the porous aluminum nanoparticulates.

[46] The following terms have the following meanings unless otherwise indicated. Any undefined terms have their art recognized meanings.

[47] In the present description, the terms “about” and "consisting essentially of mean within 20% of the indicated range, value, or structure, unless otherwise indicated. In some embodiments, the terms "about" and "consisting essentially of mean within 5% of the indicated range, value, or structure, unless otherwise indicated.

[48] The use of the alternative (e.g., "or") should be understood to mean either one, both, or any combination thereof of the alternatives.

[49] As used herein, the terms "include," "have" and "comprise" are used synonymously, which terms and variants thereof are intended to be construed as non-limiting.

[50] As used herein and in the appended claims, the singular forms "a," "an," and "the" include plural reference unless the context clearly indicates otherwise.

[51] The term "bioactive agent" as used herein refers to any material to be delivered by the porous aluminum nanoparticulate structure formulations of the present disclosure and include without limitation macromolecules, peptides, proteins, peptidomimetics, nucleic acids, oligonucleotides, deoxyribonucleotides, ribonucleotides, mRNA, RNAi, Rigl, replicon RNA, adjuvants including TLR agonists (for example TLR2, TLR3, TLR4, TLR 7, TLR8, and TLR9 agonists), saponins, whole viral particles, viral fragments, cellular fragments. Also included within the term bioactive agent are, for example, aptamers, carbohydrates, conjugated carbohydrates and vims-like particles.

[52] The term "macromolecule" as used herein refers to large molecules exemplified by, but not limited to, peptides, proteins, oligonucleotides and polynucleotides of biologic or synthetic origin. Also included within the term macromolecule are, for example, carbohydrates.

[53] The terms "polypeptide", "peptide", and "protein" are used interchangeably herein to refer to polymers of amino acids of any length. The polymer may be linear or branched, it may comprise modified amino acids, and it may be interrupted by non-amino acids. The terms also encompass an amino acid polymer that has been modified naturally or by intervention; for example, disulfide bond formation, glycosylation, lipidation, acetylation, phosphorylation, or any other manipulation or modification, such as conjugation with a labeling component. Also included within the definition are, for example, polypeptides containing one or more analogs of an amino acid (including, for example, unnatural amino acids, etc.), as well as other modifications known in the art which includes peptidomimetic compounds which are derived from peptides and proteins by structural modification using unnatural amino acids.

[54] The term "isolated" means the molecule has been removed from its natural environment. [55] "Purified" means that the molecule has been increased in purity, such that it exists in a form that is more pure than it exists in its natural environment and/or when initially synthesized and/or amplified under laboratory conditions. Purity is a relative term and does not necessarily mean absolute purity. In some embodiments, purified can mean 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% more pure than it exists in its natural environment and/or when initially synthesized and/or amplified under laboratory conditions.

[56] A "polynucleotide" or "nucleic acid," as used interchangeably herein, refers to polymers of nucleotides of any length, include DNA and RNA. The nucleotides can be deoxyribonucleotides, ribonucleotides, modified nucleotides or bases, and/or their analogs, or any substrate that can be incorporated into a polymer by DNA or RNA polymerase, or by a synthetic reaction. A polynucleotide may comprise modified nucleotides, such as methylated nucleotides and their analogs. If present, modification to the nucleotide structure may be imparted before or after assembly of the polymer. The polynucleotides of the present disclosure include ribonucleotides (for example RNA, RNAi, tRNA, and mRNA as terms well known in the art.) and deoxyribonucleotides (DNA) know in the art and may be single or double stranded molecules.

[57] "Oligonucleotide," as used herein, generally refers to short, generally single stranded, generally synthetic polynucleotides that are generally, but not necessarily, less than about 200 nucleotides in length. The terms

“oligonucleotide” and "polynucleotide” are not mutually exclusive. The description above for polynucleotides is equally and fully applicable to oligonucleotides. Examples include Rig I agonists.

[58] A "replicon" as used herein includes any genetic element, for example, a plasmid, cosmid, bacmid, phage or virus that is capable of replication largely under its own control. A replicon may be either RNA or DNA and may be single or double stranded.

[59] An "individual" or a "subject" is any mammal. Mammals include, but are not limited to humans, primates, farm animals, sport animals, pets (such as cats, dogs, horses), and rodents.

[60] "Alkyl" is a straight or branched saturated hydrocarbon. For example, an alkyl group can have 1 to 30 carbon atoms (i.e., (Ci-C _3o)alkyl) or 1 to 20 carbon atoms (i.e., (Ci C₂o alkyl) or 1 to 10 carbon atoms (i.e., (Ci-Cio)alkyi) or 1 to 8 carbon atoms (i.e., (C 1-C )alkyl) or 1 to 6 carbon atoms (i.e., (C 1- C ) alkyl) or 1 to 4 carbon atoms (i.e., (Ci-C ₄) alkyl). This term includes, by way of example, linear and branched hydrocarbyl groups such as methyl (CH3-), ethyl (CH3CH2-), n-propyi (CH3C CH -), isopropyl ((CH3)2CH-), n-butyl (CH CH2CH2CH2-), isobutyl ((CH3)2CHCH -), sec-butyl ((CH3)(CHCH2) (CH3)), t-butyl ((CH3)3C-), n-pentyl (CH3CH2CH2CH2CH2-), neopentyl (CH3)3C(CH-)' ! ) '( ' !,-). and n-hexyl (CH3(CH2)5-).

[61] "Halo" or "halogen" refers to fluoro, chloro, bromo, and iodo.

[62] "Hydroxy" or "hydroxyl" refers to the group -OH.

[63] "Alkoxy" refers to the group — Q-aikyi, wherein alkyl is as defined herein. Alkoxy includes, by way of example, methoxy, ethoxy, n-propoxy, isopropoxy, n-butoxy, t-butoxy, sec-butoxy, n-pentoxy, and the like. [64] "Carboxyl ester" or "earboxy ester" refers to the groups -C(0)0-alkyl and -C (O)O-substituted alkyl, wherein alkyl and substituted alkyl are as defined herein.

[65] The practice of the present disclosure will employ, unless otherwise indicated, conventional techniques of molecular biology, recombinant DNA, biochemistry, and chemistry, which are within the skill of the art. Such techniques are explained fully in the literature. See, e.g., Molecular Cloning A Laboratory Manual, 2nd Ed , Sambrook et al., ed., Cold Spring Harbor Laboratory Press: (1989); DNA Cloning, Volumes I and II (D. N. Glover ed., 1985); Oligonucleotide Synthesis (M. J Gait ed., 1984); Mullis et al., U.S. Pat. No: 4,683, 195; Nucleic Acid Hybridization (B. D. Hames & S. J. Higgins eds. 1984); B. Perbai, A Practical Guide To Molecular Cloning (1984); the treatise, Methods In Enzymology (Academic Press, Inc., N.Y.), and in Ausubel et al., Current Protocols in Molecular Biology, John Wiley and Sons, Baltimore, Maryland (1989).

[66] The porous aluminum nanoparticulate structure provided herein comprise an aluminum salt (interchangeably referred to as an alum) and a capping agent, wherein the size of the particle ranges from about 1 nm to about 200 nm. Discussion of the aluminum salts and capping agents are provided below.

[67] The compositions described herein can comprise an aluminum salt, which can be referred to herein as alum. Suitable aluminum salts include aluminum hydroxide, aluminum Trihydrate, aluminum oxyhydroxide, aluminum phosphate, aluminum hydroxyphosphate, aluminum hydroxyphosphate sulfate, and potassium aluminum sulfate. Aluminum salts can also be referred to by the formulae: Ai(OH)₃, A1H₃O ₃, A1HO₃, AIO(OH),

A1(OH)(PO), and K I S i}. The skilled artisan will appreciate that aluminum hydroxyphosphate is nonstoichiometric and although it is represented herein as A1(OH)(PO), the ratio of surface hydroxyis to phosphates vary depending on the manufacturing conditions and as such is more accurately represented by the formula: Al (OH) (P0₄).

[68] In certain embodiments, the aluminum salt is Alhydrogel®, an aluminum hydroxide or aluminum oxyhydroxide. Alhydrogel® has an overall positive charge and can readily adsorb negatively charged moieties. Alhydrogel® can also be referred to as Amphojel; Aluminum hydroxide gel; hydrated alumina: Aluminum trihydroxide; or Aiugelibye.

[69] In certain embodiments, the aluminum salt is AdjuPhos®, an aluminum phosphate AdjuPhos® has an overall negative charge and can readily adsorb positively charged moieties.

[70] In some embodiments, the size of the porous aluminum nanoparticulate structure is maintained because the capping agent reduces, blocks, or retards the aggregation of the processed or milled aluminum salt, when compared to a porous aluminum nanoparticulate structure comprising an aluminum salt in the absence of a capping agent. The capping agent is either hydroxypropyl beta cyclodextrin or sulfo-butyletherbetacyclodextrin or combination with water-soluble polymers such as PAA, Polysorbate 80.

[71] In some embodiments the capping agent is added during the processing aluminum salt by high energy input such as high shear mixing, sonication or microfluidization to achieve the desired porous aluminum nanoparticulate size. In some embodiments the capping agent is added after processing aluminum salt by high energy input such as sonication or microfluidization to achieve the desired porous aluminum nanoparticulate size. In some embodiments when the capping agent is added after processing aluminum salt to achieve the desired porous aluminum nanoparticulates size by high energy input such as sonication or microfluidization the capping agent is added immediately after processing or about 0.5 minutes, 0.5- 1.0 minute, 1.0-1 .5 minutes, 1.5-2.0 minutes, 2.0-2.5 minutes, 2.5-3.0 minutes, 3.0-3.5 minutes, 3.5-4.0 minutes, 4.0-4.5 minutes, 4.5-5.0 minutes, 5.05-5.5 minutes, 5.5-6.0 minutes, 6.0-6.5 minutes, 6.5-7.0 minutes, 7.0-7.5 minutes, 7.5-8.0 minutes, 8.0-8.5 minutes, 8.5-9.0 minutes, about 10 minutes, about 12 minutes, about 14 minutes, about 16 minutes, about 8 minutes, about 20 minutes, about 22 minutes, about 24 minutes, about 26 minutes, about 28 minutes, about 30 minutes after processing aluminum salt to achieve the desired porous aluminum nanoparticulates size.

[72] In some embodiments, the capping agent is one that changes the surface properties of the aluminum salt. In some embodiments, the capping agent is one that stabilizes the size of the aluminum salt. In some embodiments, the capping agent is one that stabilizes or protects a bioactive agent. Examples of bioactive agents include, but are not limited to an antigen, adjuvant, TLR agonist, peptide mimetic, peptide, polypeptide, protein, nucleotide, polynucleotide, RNA, DNA, whole viral genome, and whole virus. The bioactive agent can be delivered by the porous aluminum nanoparticulate structure formulations of the present disclosure. In some embodiments, the capping agent protects or shields the bioactive agent from oxidation. In some embodiments, the capping agent protects or shields the bioactive agent from heat stress, which can include factors of heat temperature and time. In some embodiments the capping agent protects or shields the bioactive agent from cold stress, which can include factors of cold temperature and time. In some embodiments, the capping agent protects or shields the bioactive agent from degradation. In some embodiments, the capping agent is one that shields or protects the bioactive agent to be delivered by the porous aluminum nanoparticulate structure formulations of the present disclosure from degradation or inactivation when exposed to serum or blood components. In some embodiments the capping agent protects or shields the bioactive agent such that the agent may be formulated with the porous aluminum nanoparticulates as a stable single via formulation.

[73] In some embodiments, the presence of the capping agent reduces, blocks, or retards the aggregation or re-aggregation of the aluminum salt by at least 50%, at least 60%, at least 70%, at least 75%, at least 80%, at least 90%, or even blocks the aggregation or re-aggregation of the aluminum salt by nearly 100% as compared to a nanoalum particle formed in the absence of the capping agent.

[74] In the porous aluminum nanoparticulate structure provided herein, the capping agent is associated with the aluminum salt. In some embodiments, the sizing agent is adsorbs to the aluminum salt. In some embodiments the capping agent is adsorbed to the porous aluminum nanoparticulate structure.

[75] Provided herein is a porous aluminum nanoparticulate structure comprising an aluminum salt and a capping agent, wherein the size of the porous aluminum nanoparticulate ranges from about 1 nm to 200 nm. The present disclosure provides methods for preparing such porous aluminum nanoparticulate structures.

[76] The method for preparing a porous aluminum nanoparticulate structures comprises subjecting an aluminum salt to a high energy source or high energy sheer force in the presence of a capping agent, whereby the size of the aluminum salt is reduced and a porous aluminum nanoparticulate structure is produced, and wherein the size of the porous aluminum nanoparticulate ranges from about 1 nm to 200 nm.

[77] In certain embodiments, the alum is processed or milled in the presence of the capping agent or that the capping agent is added to the milled alum at least seconds, minutes, after processing. In some embodiments, the capped alum is dried most with nitrogen flow with or without antigen.

[78] In some embodiments the high energy source provides at least 5000 PSI , at least 10,000 PSI, at least 15,000 PSI at least 20,000 PSI, at least 25,000 PSI, at least 30,000 PSI, at least 35,000 PSI, at least 40,000 PSI, at least 45,000 PSI, or at least 50,000 PSI. In some embodiments the high energy source provides about 5000 to 50000; 5000 to 10000; 5000 to

[79] 15000; 5000 to 20000; 5000 to 25000; 5000 to 30000; 5000 to 35000, 5000 to 40000; 5000 to 45000; or 5000 to 50000 PSI. In some embodiments the high energy source provides about 45000 to 50000, 40000 to 50000; 35000 to 50000, 30000 to 50000; 25000 to 50000, 20000 to 50000; 15000 to 50000; 10000 to 50000; or 5000 to 50000 PSI.

[80] In some embodiments the high energy source provides about 25000 to 35000; 25000 to 30000; or 30000 to 35000 PSI In some embodiments the high energy source provides about 30000 PSI. In some embodiments, the high energy source is a high shear source. In some embodiments the high energy source is a microfluidizer. Microfluidization is used to describe a process in which compositions are exposed to high shear force. In some embodiments the high energy source is an extruder. In some embodiments the high energy source is a sonicator. In some embodiments the high energy source is a homogenizer.

[81] In some embodiments, the aluminum salt and the capping agent are subjected to least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, 25, 30, 50, or 100 passes of the high shear force. In some embodiments the aluminum salt and the sizing agent are subjected to 1-5, 6-10, 1 1-15, 16- 20, 21-30, 3 1-40, 41-50, 51-60, 61-70, 71-80, 81-90, or 91-100 passes of the high shear force. In some embodiments, the aluminum salt and the sizing agent are subjected to 3, 6, or 0 passes of the high shear force.

[82] In some embodiments, the method for preparing the porous aluminum nanoparticulate structures of the present disclosure is performed at 0°C, at 4°C, at 25°C, at 30°C, at 50 , C, or at 55°C. In some embodiments, the method for preparing the porous aluminum nanoparticulate of the present disclosure is performed at 0-4, 5-10, 10- 15, 16-20, 21-25, 26-30, 31-35, 36-40, 41-45, 46-50, 51-55, or 56-60°C. In some embodiments, the method for preparing the porous aluminum nanoparticulate of the present disclosure is performed between 4°C and approximately 55 deg Celsius with or without drying under nitrogen flow.

[83] In some embodiments, the starting concentration of the aluminum salt is 10 mg/ml. In some embodiments, the starting concentration of the aluminum salt is 4mg/ml. In some embodiments, the starting concentration of the aluminum salt is 2mg/ml. In some embodiments, the starting concentration of the aluminum salt is 0.5 to 10 mg/ml, 1 to 10 mg/ml, 0.5 to 5mg/ml; 1 to 5mg/ml; 0 5 to 4mg/ml; 0.5 to 3mg/ml; or 0.5 to 3mg/ml. In some embodiments, the starting size of the aluminum salt is 1 pm. In some embodiments, the starting size of the aluminum salt is 0.5 to 20 pm.

[84] In some embodiments, a porous aluminum nanoparticulate described herein is produced by milling or processing according to methods described herein in the presence of a capping agent, and has an average particle size of 1-200 nm. In certain embodiments, a synthetic porous aluminum nanoparticulate may include synthetic alum as has been described in the art which is de novo synthesized to produce the appropriate alum particle size to which a capping agent of the present disclosure has been added to create a stable aqueous porous aluminum nanoparticulate structure formulation. Porous aluminum nanoparticulate structure of the formulation may be mixed with pharmaceutically acceptable excipients known in the art to produce porous aluminum nanoparticulate compositions or formulations.

[85] As used herein, the terms "milling," "capping," or "processing" refers to a process of treating a solution of alum in order to achieve porous aluminum nanoparticulate. The process includes processing an alum composition (including formulation) via high energy source or input to reduce the aggregation of the alum particles as measured by a reduced average particle size below 0.5-10 pm. Suitable examples of energy input to achieve porous aluminum nanoparticulate compositions include, but are not limited to, high shear mixing (such as ultrasonication or high shear mixing with a Silverson high shear mixer), extrusion , homogenization and microfluidization. In some embodiments, high shear mixing is performed at 1000, 2000, 5000, or 10,000 rpms for 1 minute, 2 minutes, 5 minutes, or 10 minutes. In some embodiments, the raicrofluidizer is a Microfluidics M 1 IOP (Newton, MA), equipped with a diamond F 2Y interaction chamber followed by a ceramic H30Z auxiliary processing module. In some embodiments, the alum compositions are microfluidized at pressures of 3,000 PSI 5,000PSI, 10,000 PSI, 15,000 PSI, or 30,000 PSI. In some embodiments, the solution of alum is processed by a microfluidizer with a recirculation water temperature of 60°C, 40°C, 20°C or 4°C to achieve porous aluminum nanoparticulate structures. In some embodiments, the solution of alum is milled or processed at least about 1, 3, 6, 10, 15, 20, or 30 passages to reproducibly achieve nanoparticulates of the present disclosure having an average particle size of 1-200 nm in size. In some embodiments, the solution of alum is microfluidized for up to 10 passages at 30,000 PSI with a recirculating 4°C water to prevent temperature increase during processing. In some embodiment, the solution of alum is processed in the presence of the capping agent. In some embodiments, the ratio of the capping agent to alum is 30: 1, 20: 1, 15: 1, 10: 1, 7.5: 1, 4: 1, 3: 1, 2: 1, 1.5: 1, 0.5: 1, or 0.25: 1. In some embodiments, the ratio of the capping agent to alum is 7 5: 1, 4: 1, 3: 1, 2: 1 or 1: 1.

[86] It is understood that certain variables can be controlled in a method of preparing a porous aluminum nanoparticulate structures of the embodiments. Certain variables include, but are not limited to, the capping agent, the type of high energy source, the pressure exerted by the high energy source, the number of passes of the mixture through the high energy source, the temperature at which the process takes place, the concentration of capping agent, the point in the method wherein the capping agent is added to the aluminum, and the ratio of the aluminum salt to the capping agent by weight.

[87] As provided herein, the size of the porous aluminum nanoparticulate structure comprising an aluminum salt and a capping agent ranges from about 1 nm to about 200 nm.

[88] In some embodiments the size of the porous aluminum nanoparticulate ranges from about 50nm to 75nm. In some embodiments the size of the porous aluminum nanoparticulate ranges from about 50nm to 100 nm. In some embodiments the size of the porous aluminum nanoparticulate ranges from about 50nm to 150nm. In some embodiments the size of the porous aluminum nanoparticulate ranges from about 50nm to 200nm. In some embodiments the size of the porous aluminum nanoparticulate ranges from about 20nm to lOOnm. In some embodiments the size of the porous aluminum nanoparticulate ranges from about 20nm to 50nm. In some embodiments the size of the porous aluminum nanoparticulate ranges from about lOnm to 200nm. In some embodiments the size of the porous aluminum nanoparticulate ranges from about lOnm to 100 nm. In some embodiments the size of the porous aluminum nanoparticulate ranges from about 10 nm to 50nm. In some embodiments the size of the nanoalum particle is about 1 nm, is about 5nm, is about lOnm, is about 15nm, is about 20nm, is about 25nm, is about 30nm, is about 35nm, is about 40nm, is about 45nm, is about 50nm, is about 55nm, is about 60nm, is about 65nm, is about 70nm, is about 75nm, is about 80nm, is about 85nm, is about 90nm, is about 95nm, is about lOOnm, is about 105nm, is about l lOnm, is about 115nm, is about 120nm, is about 125nm, is about 130nm, is about 135nm, is about 140nm, is about 145nm, is about 150nm, is about 155nm, is about 160nm, is about 165nm, is about 170nm, is about 175nm, is about 180nm, is about 185nm, is about 190nm, is about 195nm, is about 200nm. In some embodiments, the porous aluminum nanoparticulate is capable of being filtered through a 0.20 micron filter.

[89] In some embodiments provided herein, the 1-200 nm size of the porous aluminum nanoparticulate comprising an aluminum salt and a sizing agent is stable, in that the porous aluminum nanoparticulate's size of less than 200 nm is maintained, and in that the aluminum salt exhibits reduced aggregation, or no aggregation, when compared to an aluminum salt in the absence of a capping agent.

[90] In some embodiments, "stable" refers to a porous aluminum nanoparticulate structure formulation or composition comprised of porous aluminum nanoparticulate which does not "aggregate" displays little to no aggregation, or reduced aggregation and or demonstrates little to no overall increase in average particle size or polydispersity of the formulation over time compared to the initial particle size.

[91] The stability of the porous aluminum nanoparticulate structure can be measured by techniques familiar to those of skill in the art. In some embodiments, the stability is observed visually. Visual inspection can include inspection for particulates, flocculence, or aggregates. In some embodiments, the stability is determined by the size of the porous aluminum nanoparticulate. For example, the size can be assessed by known techniques in the art, including but not limited to, x-ray and laser diffraction, dynamic light scattering (DLS), CryoEM, or Malvern Zetasize. In some embodiments, the size of the porous aluminum nanoparticulate refers to the Z-average diameter. In some embodiments, the stability is determined by assessing the percentage aggregation of the aluminum salts in the porous aluminum nanoparticulate structures. In some embodiments, the stability is assessed by the ability of the aluminum nanoparticulate structures to pass through a filter of a particular size, for example through a 0.20, 0.35, and 0.45 micron filter. In some embodiments, stability is determined by measurement of the polydispersity index (Pdl), for example with the use of the dynamic light scattering (DLS) technique.

[92] In some embodiments, the Z-average diameter of the nanoparticle increases less than 50%, less than 40%, less than 30%, less than 25%, less than 20%, less than 15%, less than 12%, less than 0%, less than 7%, less than 5%, less than 3%, less than 1% over time period assayed. In some embodiments, the polydispersity index (Pdl) of the nanoparticlulate increases less than 50%, less than 40%, less than 30%, less than 25%, less than 20%, less than 5%, less than 12%, less than 10%, less than 7%, less than 5%, less than 3%, less than 1% over time period assayed.

[93] In some embodiments, the porous aluminum nanoparticulate structure is stable at 0-8°C. In some embodiments, the porous aluminum nanoparticulate structure is stable at 0°C, 1°C, 2°C, 3°C, 4°C, 5°C, 6°C, TC, or 8°C for at least 1 minute, for at least 5 minutes, for at least 0 minutes, for at least 5 minutes, for at least 20 minutes, for at least 25 minutes, for at least 30 minutes, for at least 35 minutes, for at least 40 minutes, for at least 45 minutes, for at least 50 minutes, for at least 55 minutes, for at least 1 hour, for at least 2 hours, for at least 6 hours, for at least 12 hours, for at least 8 hours, for at least 24 hours, for at least 48 hours, for at least 72 hours, for at least 1 week, for at least 2 weeks, for at least 3 weeks, for at least 1 month, for at least 2 months, for at least 3 months, for at least 4 months, for at least 5 months, for at least 6 months, for at least 7 months, for at least 8 months, for at least 9 months, for at least 10 months, for at least 11 months, for at least 1 year, for at least 2 years, or for at least 5 years.

[94] In some embodiments, the porous aluminum nanoparticulate structure is stable at 20-30°C. In some embodiments, the porous aluminum nanoparticulate structure is stable at 25°C for at least 1 minute, for at least

5 minutes, for at least 10 minutes, for at least 5 minutes, for at least 20 minutes, for at least 25 minutes, for at least 30 minutes, for at least 35 minutes, for at least 40 minutes, for at least 45 minutes, for at least 50 minutes, for at least 55 minutes, for at least 1 hour, for at least 2 hours, for at least 6 hours, for at least 2 hours, for at least 8 hours, for at least 24 hours, for at least 48 hours, for at least 72 hours, for at least 1 week, for at least 2 weeks, for at least 3 weeks, for at least 1 month, for at least 2 months, for at least 3 months, for at least 4 months, for at least 5 months, for at least

6 months, for at least 7 months, for at least 8 months, for at least 9 months, for at least 10 months, for at least 11 months, for at least 1 year, for at least 2 years, or for at least 5 years.

[95] In some embodiments, the porous aluminum nanoparticulate structure is stable at 35-40°C. In some embodiments, the porous aluminum nanoparticulate structure is stable at 35°C, 36°C, 37°C, 38°C, 39°C, or 40°C for at least 1 minute, for at least 5 minutes, for at least 10 minutes, for at least 15 minutes, for at least 20 minutes, for at least 25 minutes, for at least 30 minutes, for at least 35 minutes, for at least 40 minutes, for at least 45 minutes, for at least 50 minutes, for at least 55 minutes, for at least 1 hour, for at least 2 hours, for at least 6 hours, for at least 12 hours, for at least 18 hours, for at least 24 hours, for at least 48 hours, for at least 72 hours, for at least 1 week, for at least 2 weeks, for at least 3 weeks, for at least 1 month, for at least 2 months, for at least 3 months, for at least 4 months, for at least 5 months, for at least 6 months, for at least 7 months, for at least 8 months, for at least 9 months, for at least 10 months, for at least 11 months, for at least 1 year, for at least 2 years, or for at least 5 years.

[96] In some embodiments, the porous aluminum nanoparticulate structure is stable at 57-75°C. In some embodiments, the porous aluminum nanoparticulate structure is stable at 57°C, 58°C, 59°C, 60°C, 6 1°C, or 62°C for at least 1 minute, for at least 5 minutes, for at least 10 minutes, for at least 15 minutes, for at least 20 minutes, for at least 25 minutes, for at least 30 minutes, for at least 35 minutes, for at least 40 minutes, for at least 45 minutes, for at least 50 minutes, for at least 55 minutes, for at least 1 hour, for at least 2 hours, for at least 6 hours, for at least 12 hours, for at least 18 hours, for at least 24 hours, for at least 48 hours, for at least 72 hours, for at least 1 week, for at least 2 weeks, for at least 3 weeks, for at least 1 month.

[97] In some embodiments, the porous aluminum nanoparticulate structure is stable at 55°C when wrapped in a phosphatidyl choline liposome. In some embodiments, the porous aluminum nanoparticulate structure is stable at individual transition melting point when wrapped in other class of phsopholipid liposome.

[98] In one exemplary embodiment, the porous aluminum nanoparticulate structure is stable at 4°C for at least 2 years. In one exemplary embodiment, the porous aluminum nanoparticulate structure is stable at 4°C for at least 4 years. In one exemplary embodiment, the porous aluminum nanoparticulate structure is stable at 4°C for at least 5 years. In one exemplary embodiment, the porous aluminum nanoparticulate structure is stable at 25°C for at least one month. In one exemplary embodiment, the porous aluminum nanoparticulate structure is stable at 37°C for at least two weeks. In one exemplary embodiment, the porous aluminum nanoparticulate structure is stable at 60°C for at least two weeks.

[99] In some embodiments, the porous aluminum nanoparticulate structure is stable after 1-3 freeze thaws. In some embodiments, the porous aluminum nanoparticulate structure is stable after 1, after 2, or after 3 freeze thaws.

[100] Provided herein are compositions comprising a porous aluminum nanoparticulate structures, wherein the porous aluminum nanoparticulate structure comprise an aluminum salt and a capping agent, and wherein the size of the porous aluminum nanoparticulates are about 1 nm- 200 nm in size. In some embodiments the average size of the porous aluminum nanoparticulate composition ranges from about 50 nm to 75 nm. In some embodiments the average size of the porous aluminum nanoparticulates composition ranges from about 50 nm to 100 nm. In some embodiments the average size of the porous aluminum nanoparticulates composition ranges from about 50 nm to 150 nm. In some embodiments the average size of the porous aluminum nanoparticulates composition ranges from about 50nm to 200nm. In some embodiments the average size of the porous aluminum nanoparticulates composition ranges from about 20 nm to 100 nm. In some embodiments the average size of the porous aluminum nanoparticulates composition ranges from about 20 nm to 50 nm. In some embodiments the average size of the porous aluminum nanoparticulates composition ranges from about 10 nm to 200 nm. In some embodiments the average size of the porous aluminum nanoparticulates composition ranges from about 10 nm to 100 nm. In some embodiments the average size of the porous aluminum nanoparticulates composition ranges from about 10 nm to 50 nm. In some embodiments the average size of the porous aluminum nanoparticulates composition is about Inm, is about 5 nm, is about 10 nm, is about 15 nm, is about 20 nm, is about 25 nm, is about 30 nm, is about 35nm, is about 40nm, is about 45nm, is about 50nm, is about 55nm, is about 60nm, is about 65nm, is about 70nm, is about 75nm, is about 80nm, is about 85nm, is about 90nm, is about 95nm, is about lOOnm, is about 105nm, is about l lOnm, is about 115nm, is about 120nm, is about 125nm, is about 130nm, is about 135nm, is about 140nm, is about 145nm, is about 150nm, is about 155nm, is about 160nm, is about 165nm, is about 170nm, is about 175nm, is about 180nm, is about 185nm, is about 190nm, is about 195nm, is about 200nm. In some embodiments, the average size of the porous aluminum nanoparticulate composition is no greater than about Inm, no greater than about 5nm, no greater than about lOnm, no greater than about 15nm, no greater than about 20nm, no greater than about 25nm, no greater than about

30nm, no greater than about 35nm, no greater than about 40nm, no greater than about 45nm, no greater than about 50nm, no greater than about 55nm, no greater than about 60nm, no greater than about 65nm, no greater than about 70nm, no greater than about 75nm, no greater than about 80nm, no greater than about 85nm, no greater than about 90nm, no greater than about 95nm, no greater than about lOOnm, no greater than about 105nm, no greater than about HOnm, no greater than about 115nm, no greater than about 120nm, no greater than about 125nm, no greater than about 130nm, no greater than about 135nm, no greater than about 140nm, no greater than about 145nm, no greater than about 150nm, no greater than about 155nm, no greater than about 160nm, no greater than about 165nm, no greater than about 170nm, no greater than about 175nm, no greater than about 180nm, no greater than about 185nm, no greater than about 190nm, no greater than about 195nm, no greater than about 199nm, no greater than about 200nm measured by DLS.

[101] In some embodiments, the composition is capable of being filtered through a 0.20 micron size filter. In some embodiments, the compositions are maintained as aqueous formulations. In some embodiments, the compositions are maintained as dried formulations. In some embodiments, the compositions are maintained as spray-dried or dried under nitrogen flow or under ambient conditions formulations. In some embodiments, the composition comprises a porous aluminum nanoparticulate structure and an emulsion, micelle, or liposome. In some embodiments the emulsion of the composition is water in oil emulsion. In some embodiments the emulsion of the composition is a pickering emulsion. In some embodiments the emulsion of the composition is an oil-in-water emulsion. In some embodiments the oil of the emulsion is biodegradable oil. In further embodiments the oils is a squalene. In other embodiment the oil is synthetic biodegradable oil. Liposomes and liposome derived nanovesicles known in the art and may be used with the porous aluminum nanoparticulate of the present disclosure. In some embodiments, the composition comprises a liposome containing the porous aluminum nanoparticulate structures. In some embodiments the composition comprises a porous aluminum nanoparticulate and a liposome wherein the liposome is a cationic liposome. In some embodiments the composition comprises a porous aluminum nanoparticulate and a liposome wherein the liposome is an anionic liposome. In some embodiments the composition comprises a porous aluminum nanoparticulate and a liposome wherein the liposome is a neutral liposome. In some embodiments the composition comprises a porous aluminum nanoparticulate and a liposome wherein the liposome is an archaeosome. In some embodiments, the composition comprises a porous aluminum nanoparticulate and a liposome wherein the liposome is virosome.

[102] The porous aluminum nanoparticulate structures can be combined with any other bioactive agents that can induce an immune response, allergic response or anti-tumor response as are known in the art.

[103] In some embodiments the agent is useful for therapeutic purposes. Thus in some embodiments, the compositions described comprise the porous aluminum nanoparticulate structures provided herein, and further comprise an agent for the treatment of a disease, condition, or disorder. In some embodiments the agent is useful for the treatment or prevention of allergy, cancer, infectious disease, autoimmunity, or addiction. [104] Figure 1 shows an electron micrograph depicting various embodiments of the disclosed technology comprising a porous aluminum nanoparticulate structure. Figure 1 is obtained by a series 1 in which Alhydrogel, HPBCD and a very small percentage of PAA are added, filtered, dried out and then reconstituted.

[105] Figure 2 also shows an electron micrograph depicting various embodiments of the disclosed technology comprising a porous aluminum nanoparticulate structure.

[106] Moreover, figure 3 shows an electron micrograph depicting various embodiments of the disclosed technology comprising a porous aluminum nanoparticulate structure. Figure 3 is obtained by a series 3 in which Alhydrogel, HPBCD, very small percentage of water-soluble polymer, PS 80 are filtered, dried out and then rehydrated.

[107] It is inferred from the abovementioned figures that the Aluminum particles confirm porous with high surface area to mass. The Aluminum particles appear to look unusual and are displayed as sheets of aluminum nanoparticles embedded within these sheets. As it can be clearly seen that it looks like it is more than 1 structure. This is something out of the ordinary morphology of aluminum particles which have crystal agglomeration of rodlike structure to them. Hence, it is a novel form of adjuvant.

[108] While the disclosed technology has been taught with specific reference to the above embodiments, a person having ordinary skill in the art will recognize that changes can be made in form and detail without departing from the spirit and the scope of the disclosed technology. The described embodiments are to be considered in all respects only as illustrative and not restrictive. All changes that come within the meaning and range of equivalence of the claims are to be embraced within their scope. Combinations of any of the methods, systems, and devices described herein-above are also contemplated and within the scope of the disclosed technology.

Claims

CLAIMS What is claimed is:

1. A porous aluminum nanoparticulate structure comprising:

(a) an aluminum salt;

(b) a capping agent, wherein said capping agent is selected from group consisting of Hydroxypropylbetacyclodextrin (HPBCD), Sulfobutyletherbetacyclodextrin (SBECD) or combination with water-soluble polymers such as PAA, Polysorbate 80; and said capping agent is associated with said aluminum salt; wherein, the size of said porous aluminum nanoparticulate ranges from about 1 nm to about 200 nm; and

(c) a water-soluble polymer from 0.001 - 2% of the adjuvant solution; wherein, said porous aluminum nanoparticulate structure is stable in a liquid formulation at about 0°C to about 8°C for at least about 1 month, at least about 6 months, or at least about 1 year.

2. The porous aluminum nanoparticulate structure of claim 1, wherein the average size of said porous aluminum nanoparticulate structure is the Z- average as determined by dynamic light scattering.

3. The porous aluminum nanoparticulate structure of claim 1, wherein said aluminum salt is selected from the group consisting of aluminum

39 hydroxide, aluminum hydroxide gel, A1PO₄, AIO(OH), A1(OH)(PO₄), and KA1(SO₄)₂.

4. The porous aluminum nanoparticulate structure of claim 1, wherein said porous aluminum nanoparticulate structure is in a liquid formulation which is filter-sterilized.

5. The porous aluminum nanoparticulate structure of claim 1, wherein said porous aluminum nanoparticulate structure is stable after repeated freeze-thaw cycles.

6. The porous aluminum nanoparticulate structure of claim 1, wherein said porous aluminum nanoparticulate structure is stable in a liquid formulation at about 37°C for at least about 1 month.

7. A method of preparing a porous aluminum nanoparticulate structure comprising:

(a) subjecting an aluminum salt to a high energy source in the presence of a capping agent, whereby said porous aluminum nanoparticulate structure is produced with a size ranging from about 1 nm to about 200 nm, wherein said aluminum salt comprises of particles ranging from 0.5 to 10 pm in size or 0.5 to 20 pm in size; and

(b) mixing a capping agent with said porous aluminum nanoparticulate structure after step (a); and

(c) filter-sterilizing said porous aluminum nanoparticulate structure.

40

8. The method of claim 7, wherein said high energy source is generated from a microfluidizer, an extruder, a sonicator, a high shear mixer, or a homogenizer and said high energy source is generated from two or more of said microfluidizer, said extruder, said sonicator, said high shear mixer, or said homogenizer.

9. The method of claim 7, wherein said high energy source is generated from a microfluidizer and a high shear mixture, and the mixture comprising the aluminum salt and capping agent is passed through the microfluidizer from one pass to about 15 passes and one pass to about 30 passes.

10. The method of claim 7, wherein said aluminum salt is selected from the group consisting of aluminum hydroxide, aluminum hydroxide gel, A1PO₄, AIO(OH), Al(OH)_x(PO₄)_y, and KA1(SO₄).

11. A composition comprising a porous aluminum nanoparticulate structure produced by a method according to claim 7, wherein the composition comprises: a bioactive agent, wherein more than 75% of said bioactive agent is associated with said porous aluminum nanoparticulate structure as determined by gel electrophoresis; an adjuvant, wherein said adjuvant is selected from the group consisting of AS-2, monophosphoryl lipid A, 3-de-O-acylated monophosphoryl lipid A, IFA, QS21, CWS, TOM, AGPs, CpG-containing oligonucleotides, Toil-like receptor (TLR) agonists, Leif, saponins, saponin

41 mimetics, biological and synthetic lipid A, imiquimod, gardiquimod, resiquimod, polyLC, ilagellin, GLA, SLA, STING, and combinations thereof; and a liposome; wherein, the average size of said porous aluminum nanoparticulates in the composition ranges from about 1 nm to about 200 nm and wherein said composition is capable of being filtered through a 0.20 micron-sized filter; wherein, said composition is further capable of being terminally sterilized prior to vialing.

12. The composition of claim 11, wherein said bioactive agent is a polypeptide, a polynucleotide, an antigen, an adjuvant, a diagnostic agent, a therapeutic agent, or an organism.

13. The composition of claim 11, wherein said composition is stable at about 0°C to about 8°C for at least about 1 month, at least about 6 months, or at least about 1 year, or is stable at about 37°C for at least about 1 month.

14. The composition of claim 11, wherein the average size of said porous aluminum nanoparticulates in the composition ranges from about 1 nm to about 200 nm.

15. A method of stimulating an immune response in a subject comprising administering said composition of claim 11, to a subject, whereby stimulating said immune response in the subject response involves the activation of B-cells, activation of T cells, production of antibodies, or release of cytokines.

16. The method of claim 15, wherein said immune response is a nonspecific immune response and/or an antigen-specific immune response and/or a TH1 immune response and/or a TH2 immune response and/or both TH1 and TH2 immune response.

17. The method of claim 15, wherein said composition is used for the treatment of allergy, addiction, cancer, or autoimmunity.

18. The method of claim 15, wherein the route of administration of said composition is oral, intravenous, intradermal, transdermal, nasal, subcutaneous, or anal.

19. A method of delivering a bioactive agent to a cell in a subject comprising administering to the subject a composition comprising (a) a porous aluminum nanoparticulate structure comprising an aluminum salt and a sizing agent, wherein the size of the particle ranges from about 1 nm to about 200 nm and (b) a bioactive agent, thereby delivering the bioactive agent to the cell in the subject.

20. A method of preparing a composition comprising the steps: (a) subjecting an aluminum salt to a high energy source in the presence of a capping agent, whereby a porous aluminum nanoparticulate is produced, and wherein the size of the porous aluminum nanoparticulate ranges from about 1 nm to about 200 nm; and (b) mixing the porous aluminum nanoparticulate produced in step (a) with a bioactive agent.

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