Carrier-base complexes and methods of use thereof

Abstract

The present disclosure relates to complexes comprising carrier-bases and single stranded nucleic acid. Specifically, the carrier base can comprise i) an organic polymer comprising one or more monomeric units; and ii) at least one nucleobase; wherein at least one covalent bond binds the one or more monomeric units to the nucleobase(s), and wherein the single stranded nucleic acid comprises at least one hydrogen bond which binds the ssNA to the carrier-base to form the complex. Also disclosed are methods of making and using these complexes.

Description

[0001] CARRIER-BASE COMPLEXES AND METHODS OF USE THEREOF

[0002] CROSS-REFERENCE TO RELATED APPLICATION

[0003] This PCT application claims priority to, and the benefit of, U.S. Provisional Patent Application No. 63 / 508,636, filed June 16th, 2024, entitled “CARRIER-BASE COMPLEXES AND METHODS OF USE THEREOF,” which is incorporated by reference herein in its entirety.

[0004] FIELD

[0005] The present disclosure relates to compositions and methods comprising carrier-bases and single stranded nucleic acid.

[0006] BACKGROUND

[0007] Development of carrier complexes for the gene protection and delivery is a major goal of gene therapeutic strategies. Significant progress has been made in the construction of non-viral vectors that comprises different functions required for gene transfer and gene therapies. Current approaches rely on complexing nucleic acid sequences, (such as, for example RNA or DNA) with organic materials, such as cationic lipids and polycations, primarily through electrostatic interactions. The complex interactions between nucleic acid sequences and polycationic polymers requires specific monomeric units, such as lysine, arginine, or other amine containing moieties, carrying positive charges to bind the negatively charged nucleic acids. This reliance of positive charge provides shortcomings in certain delivery strategies. Also, there is limited availability of carrier complexes comprising a diversity of amino acids. Moreover, the stability of nucleic acids before complexing positive charged polymers during manufacture is a major concern.

[0008] Given the structural limitations of current carrier complex compositions, there is need to address the aforementioned problems mentioned above by developing compositions that are not reliant on electrostatic interactions to form carrier complexes. The compounds, compositions, and methods disclosed herein address these and other needs.

[0009] SUMMARY

[0010] The present disclosure provides a complex comprising an organic polymer and at least one nucleobase. The present disclosure also provides a therapeutic composition comprising the complex further comprising an organic polymer and at least one nucleobase. The present disclosure also provides methods preparing, forming, delivery, or using the composition or therapeutic composition comprising said complex.

[0011] In one aspect, disclosed herein is a complex comprising a carrier-base, wherein said carrierbase comprises i) an organic polymer comprising one or more monomeric units and ii) at least one nucleobase, wherein at least one covalent bond binds the one or more monomeric units to the nucleobase(s), and a single stranded nucleic acid (ssNA), wherein at least one hydrogen bond binds the ssNA to the carrier-base to form the complex.

[0012] In some embodiments, the complex comprises an RNA-plex complex. In some embodiments, the organic polymer comprises 2 or more monomeric units. In some embodiments, the monomeric units are selected from an amino acid, a monosaccharide, a nucleotide, and a fatty acid.

[0013] In some embodiments, the organic polymer comprises a combination of at least two of an amino acid, a monosaccharide, a nucleotide, or a fatty acid. In some embodiments, the organic polymer comprising 2 or more monomeric units comprises a poly-amino acid, a polysaccharide, a polynucleotide, a lipid, or combinations thereof.

[0014] In some embodiments, the poly-amino acid comprises 2 or more amino acids, or derivatives thereof. In some embodiments, the 2 or more amino acids comprise a L-, D-, or DL-amino acid. In some embodiments, the poly-amino acid ranges between about 1 kDa to about 50 kDa. In some embodiments, the polymer comprises a silica-base nanoparticle.

[0015] In some embodiments, the complex comprises one, two, three, four, five, six, or more nucleobases. In some embodiments, the complex comprises six nucleobases selected from thymine, uracil, cytosine, adenine, guanine, and hypoxanthine. In some embodiments, the complex comprises adenine, guanine, or a combination thereof. In some embodiments, the combination of adenine and guanine includes, but is not limited to a 1:1 mixture. In some embodiments, complex comprises a ratio of about 1:1 to about 1:100 of monomeric units relative to nucleobases.

[0016] In some embodiments, the ssNA comprises single-stranded DNA, single-stranded RNA, or a combination thereof. In some embodiments, the ssNA comprises messenger RNA (mRNA), transfer RNA (tRNA), ribosomal RNA (rRNA), microRNA (miRNA), long non-coding RNA (IncRNA), RNA aptamers, circular RNA (circRNA), short-interfering RNA (siRNA), small guide RNA (sgRNA), antisense DNA oligonucleotides, or antisense RNA oligonucleotides.

[0017] In some embodiments, the at least one covalent bonds comprises an ester bond, an ether bond, or an amide bond. In one aspect, disclosed herein is a therapeutic composition comprising the complex of any preceding aspect wherein the ssNA has a therapeutic benefit. In some embodiments, the method further comprises a pharmaceutically acceptable carrier selected from an excipient, a diluent, a salt, a buffer, a stabilizer, a lipid, an emulsion, and a nanoparticle.

[0018] In some embodiments, the therapeutic composition further comprises a transfection system selected from a lipofectamine composition, a lipid nanoparticle (LNP), in vivo jetRNA plus, or polyethylenimine (PEI).

[0019] In one aspect, disclosed herein is a method of treating or preventing a disease or disorder in a subject, the method comprising administering an effective amount of the therapeutic composition of any preceding aspect, wherein the ssNA is internalized and released into a cell of the subject to treat or prevent the disease or disorder.

[0020] In one aspect, disclosed herein is a method of forming a complex comprising a single stranded nucleic acid (ssNA) and a carrier-base, the method comprising providing the carrier-base, wherein said carrier-base comprises i) an organic polymer comprising one or more monomeric units and ii) at least one nucleobase, wherein at least one covalent bond binds the one or more monomeric unit to the nucleobase(s), providing the ssNA, and incubating the carrier-base and the ssNA, under conditions which allow the carrier-base molecule and the ssNA to form the complex through a hydrogen bond.

[0021] In one aspect, disclosed herein is a method of delivering a complex comprising a singlestranded nucleic acid (ssNA) into a cell, the method comprising providing a carrier-base comprising an organic polymer comprising one or more monomeric units and at least one nucleobases, wherein at least one covalent bond binds the one or more monomeric unit to the nucleobase(s); providing the ssNA, incubating the carrier-base and the ssNA, under conditions which allow the carrier-base and the ssNA to form the complex through a hydrogen bond, and introducing the complex into the cell, wherein the complex is internalized inside the cell and the ssNA is thereafter released from the complex.

[0022] In some embodiments, the method further comprises the step of storing the complex about 25 °C or less. In some embodiments, the complex comprises an RNA-plex complex.

[0023] In some embodiments, the at least one covalent bond is added by a crosslinking reagent. In some embodiments, the crosslinking reagent comprises dicyclohexyl carbodiimide (DCC). In some embodiments, the ssNA, when in complex with the carrier-base, shows reduced degradation in cellular conditions compared to a ssNA which is not protected by a carrier-base. In some embodiments, the carrier-base and ssNA are incubated in a nuclease-free solution. In some embodiments, the nuclease-free solution comprises DNase-free water, RNase-free water, or combinations thereof.

[0024] In some embodiments, the carrier-base and ssNA are incubated above 60°C for at least one minute; and thereafter the complex is then cooled to 25 °C or lower. In some embodiments, the carrier-base and ssNA are incubated between about 60°C to about 105 °C. In some embodiments, the carrier-base and ssNA are incubated for one, two, three, four, five, or more minutes. In some embodiments, the complex protects the ssNA from degradation by high temperatures above 4°C.

[0025] In some embodiments, the complex protects the ssNA from degradation by a nuclease. In some embodiments, the nuclease comprises an endonuclease or an exonuclease. In some embodiments, the nuclease is a DNase or an RNase.

[0026] In some embodiments, the complex is stored at 25°C, 4°C, -20°C, or -80°C. In some embodiments, the complex is stored in the nuclease-free solution or the complex is dehydrated by removing the nuclease-free solution prior to storing.

[0027] In some embodiments, a gene therapy method is used to deliver the complex into the cell. In some embodiments, the complex is transfected into the cell. In some embodiments, the method of any preceding aspect stabilizes the ssNA (including, but not limited to messenger RNA (mRNA), transfer RNA (tRNA), ribosomal RNA (rRNA), microRNA (miRNA), long non-coding RNA (IncRNA), RNA aptamers, circular RNA (circRNA), short-interfering RNA (siRNA), small guide RNA (sgRNA), antisense DNA oligonucleotides, and antisense RNA oligonucleotides) for cellular mechanisms, including but not limited to translation and antigen presentation.

[0028] BRIEF DESCRIPTION OF FIGURES

[0029] The accompanying figures, which are incorporated in and constitute a part of this specification, illustrate several aspects described below.

[0030] FIG 1 shows the schematic diagram of the chemical structure of carrier-bases.

[0031] FIG. 2 shows the 'H-NMR spectrum of carrier-bases.

[0032] FIG. 3 shows the13C-NMR spectrum of carrier-bases.

[0033] FIGS. 4A, 4B, and 4C show the mass spectrometry (MS) spectrum of carrier-bases (positive bombardment).

[0034] FIGS. 5A, 5B, and 5C show the MS spectrum of carrier-bases (negative bombardment).

[0035] FIG. 6 shows the ultraviolet absorbance of carrier bases.

[0036] FIG. 7 shows the schematic figure of the RNA-plex structure. FIG. 8 shows the protection of OVA mRNA against hydrolysis at high temperature in RNase free water.

[0037] FIG. 9 shows the protection of OVA mRNA against enzymatic digestion with RNase A.

[0038] FIGS. 10A and 10B show the heat resistance of RNA-plex. Naked EGFP mRNA or EGFP RNA-plex was heated at room temperature (25 °C) or 95 °C, and then transfected to 293T cells with lipo2000. GFP expression was analyzed with fluorescent microscope and Flow cytometry.

[0039] FIG. 11 shows the structure of carrier-base macromolecule is required to achieve mRNA protection effect. Single chemical components or their physical mixture cannot avoid EGFP mRNA from degradation. Only EGFP mRNA protected by carrier-base with covalent conjugation of bases on poly-amino acid expressed strong signal.

[0040] FIG. 12 shows the expression sustainability of RNA-plex in HEK293 cells in vitro. Expression sustainability is similar between naked mRNA and RNA-plex, indicating RNA is released from RNA-plex intracellularly, and RNA expression is not affected by the inclusion of carrier-base.

[0041] FIG. 13 shows the storage stability of RNA-plex at RT. Quantification of GFP expression on HEK293T cells transfected naked EGFP mRNA or EGFR RNA-plex using fluorescence microscope (upper panel) and flow cytometry (bottom panel).

[0042] FIG. 14 shows the storage stability of RNA-plex at 4°C. Quantification of GFP expression on HEK293T cells transfected naked EGFP mRNA or EGFR RNA-plex using fluorescence microscope.

[0043] FIG. 15 shows the protection of mCherry mRNA by RNA-plex technique.

[0044] FIG. 16 shows the broad application of mRNA-plex technique in other RNA delivery systems. EGFP RNA-plex was compatible with positively charged polymer (PEI, upper panel) and liposomal mRNA delivery system for in vivo applications (in vivo-jetRNA+, bottom panel).

[0045] FIG. 17 shows the protection of Luc mRNA by RNA-plex technique. Luc RNA-plex protected and delivered with different carrier-base and in vivo-jetRNA+ratios were incubated with HEK293 cells in vitro.

[0046] FIG. 18 shows the in vivo application of RNA-plex technique. In vivo-jetRNA+formulated naked Luc mRNA or Luc RNA-plex were administrated to healthy athymic nude mice via footpad injection. Luc expression was analyzed with an IVIS-200 system.

[0047] FIG. 19 shows the stabilization of EGFP mRNA by carrier-bases comprising guanine (G), adenine (A), or A / G mixtures. EGFP mRNA was protected by carrier-A, carrier-G, and carrier- A / G for mRNA transfection test in vitro. EGFP fluorescence was detected after 24 hours. Bar = 400pm.

[0048] FIGS. 20 A, 20B, and 20C show the storage stability of mRNA at room temperature (RT). Naked EGFP mRNA and EGFP-RNA-plex in RNase free water were stored at RT for up to 2 week, followed by integrity analysis by electrophoresis.

[0049] FIG. 21 shows mRNA protection by RNA-plex. EGFP (first panel), OVA (second panel), and luciferase (last panel) mRNAs were mixed with carrier-base at different ratios (w / w), followed by 25°C or 95 °C incubation for 5 minutes. mRNA integrity was analyzed by gel electrophoresis.

[0050] FIG. 22 shows the protection of circRNA with RNA-plex technology. GFP circRNA was mixed with carrier-base at different ratios (w / w), and incubated at RT or 95 °C for 5 minutes. GFP circRNA integrity was analyzed by agarose gel electrophoresis.

[0051] FIG. 23 shows the delivery of EGFP mRNA by lipid nanoparticles (LNP). EGFP / LNP or EGFP-plex / LNP was transfected to 293T cells, followed by imaging with fluorescence microscope after 24 hours. Bar = 400pm.

[0052] FIG. 24 shows the time-dependent mRNA translation in DC2.4 cells. Naked EGFP or EGFP-RNA-plex was delivered to DC2.4 cells using lipo2000 in vitro. EGFP positive cells were detected using flow cytometry.

[0053] FIGS. 25A and 25B show the antigen presentation of OVA by a dendritic cell line DC2.4. DC2.4 cells were transfected with naked OVA mRNA or OVA-RNA-plex using Lipo2000 for 24 hours, followed by flow cytometry assays.

[0054] FIGS. 26A and 26B show the mRNA protection by silica-base. Figure 26A shows the synthesis of silica-base. Nucleobases were conjugated to carboxylic acid functionalized silica nanoparticles using N,N’-Dicyclohexylcarbodimide (DCC) linker at RT to construct silica-base, followed by removing extra free reactants with dialysis. Figure 26B shows the evaluation of mRNA integrity. GFP mRNA was mixed with silica-base at different ratios (w / w) followed by RT or 95°C incubation for 5 minutes. mRNA integrity was anlyzed by agarose gel electrophoresis.

[0055] FIGS. 27A and 27B show the immunogenicity of RNA-plex. RNA-plex, naked mRNA, and the other components of RNA-plex were condensed using jetRNA, and incubated with BMDCs at Ipg mRNA / mL or equivalent amount of formulations for 24 hours. TLR9 agonist CpG1826 and STING agonist ADU-S 100 (both Ipg / mL) were used as positive controls. TNFa and IFNP cytokines in media were measured with ELISA.

[0056] FIG. 28 A, 28B, 28C, 28D, and 28E show the evaluation of therapeutic efficacy and safety of RNA-plex-based mRNA cancer vaccine. Naked OVA mRNA or OVA-RNA-plex was formulated using jetRNA. Figure 28A shows the treatment schedule. C57BL / 6 mice were subcutaneously inoculated with OVA-engineered melanoma cell line B16 (B16-0VA) (0.1 x 106per mouse), and then received either PBA, jetRNA-naked OVA mRNA, or jetRNA-OVA-RNA- plex via foot pad vaccination at day 3 and day 10 post tumor inoculation. Figures 28B and 28C show the tumor size (Figure 28B) and body weight (Figure 28C) were monitored every 2-3 days with a caliper and balance, respectively. Figure 28D shows the percentage of IFNy+CD8+T cells in the spleen after the vaccination. Figure 28 E shows the percentage of OVA antigen specific CD8+T cells in the spleen after the vaccination.

[0057] DETAILED DESCRIPTION

[0058] The following description of the disclosure is provided as an enabling teaching of the disclosure in its best, currently known embodiment(s). To this end, those skilled in the relevant art will recognize and appreciate that many changes can be made to the various embodiments of the invention described herein, while still obtaining the beneficial results of the present disclosure. It will also be apparent that some of the desired benefits of the present disclosure can be obtained by selecting some of the features of the present disclosure without utilizing other features. Accordingly, those who work in the art will recognize that many modifications and adaptations to the present disclosure are possible and can even be desirable in certain circumstances and are a part of the present disclosure. Thus, the following description is provided as illustrative of the principles of the present disclosure and not in limitation thereof.

[0059] Reference will now be made in detail to the embodiments of the invention, examples of which are illustrated in the drawings and the examples. This invention may, however, be embodied in many different forms and should not be construed as limited to the embodiments set forth herein.

[0060] Terminology

[0061] Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood to one of ordinary skill in the art to which this disclosure belongs. The term “comprising” and variations thereof as used herein is used synonymously with the term “including” and variations thereof and are open, non-limiting terms. Although the terms “comprising” and “including” have been used herein to describe various embodiments, the terms “consisting essentially of’ and “consisting of” can be used in place of “comprising” and “including” to provide for more specific embodiments and are also disclosed. As used in this disclosure and in the appended claims, the singular forms “a”, “an”, “the”, include plural referents unless the context clearly dictates otherwise.

[0062] The following definitions are provided for the full understanding of terms used in this specification.

[0063] The terms "about" and "approximately" are defined as being “close to” as understood by one of ordinary skill in the art. In one non-limiting embodiment the terms are defined to be within 10%. In another non-limiting embodiment, the terms are defined to be within 5%. In still another non- limiting embodiment, the terms are defined to be within 1%.

[0064] Ranges can be expressed herein as from “about” one particular value, and / or to “about” another particular value. When such a range is expressed, another embodiment includes from the one particular value and / or to the other particular value. Similarly, when values are expressed as approximations, by use of the antecedent “about,” it will be understood that the particular value forms another embodiment. It will be further understood that the endpoints of each of the ranges are significant both in relation to the other endpoint, and independently of the other endpoint. It is also understood that there are a number of values disclosed herein, and that each value is also herein disclosed as “about” that particular value in addition to the value itself. For example, if the value “10” is disclosed, then “about 10” is also disclosed. It is also understood that when a value is disclosed that “less than or equal to” the value, “greater than or equal to the value” and possible ranges between values are also disclosed, as appropriately understood by the skilled artisan. For example, if the value “10” is disclosed the “less than or equal to 10”as well as “greater than or equal to 10” is also disclosed. It is also understood that the throughout the application, data is provided in a number of different formats, and that this data represents endpoints and starting points, and ranges for any combination of the data points. For example, if a particular data point “10” and a particular data point 15 are disclosed, it is understood that greater than, greater than or equal to, less than, less than or equal to, and equal to 10 and 15 are considered disclosed as well as between 10 and 15. It is also understood that each unit between two particular units are also disclosed. For example, if 10 and 15 are disclosed, then 11, 12, 13, and 14 are also disclosed.

[0065] As used herein, the terms "may," "optionally," and "may optionally" are used interchangeably and are meant to include cases in which the condition occurs as well as cases in which the condition does not occur. Thus, for example, the statement that a formulation "may include an excipient" is meant to include cases in which the formulation includes an excipient as well as cases in which the formulation does not include an excipient. “Composition” refers to any agent that has a beneficial biological effect. Beneficial biological effects include both therapeutic effects, e.g., treatment of a disorder or other undesirable physiological condition, and prophylactic effects, e.g., prevention of a disorder or other undesirable physiological condition. The terms also encompass pharmaceutically acceptable, pharmacologically active derivatives of beneficial agents specifically mentioned herein, including, but not limited to, a vector, polynucleotide, cells, salts, esters, amides, proagents, active metabolites, isomers, fragments, analogs, and the like. When the term “composition” is used, then, or when a particular composition is specifically identified, it is to be understood that the term includes the composition per se as well as pharmaceutically acceptable, pharmacologically active vector, polynucleotide, salts, esters, amides, proagents, conjugates, active metabolites, isomers, fragments, analogs, etc.

[0066] The term “comprising”, and variations thereof as used herein is used synonymously with the term “including” and variations thereof and are open, non-limiting terms. Although the terms “comprising” and “including” have been used herein to describe various embodiments, the terms “consisting essentially of’ and “consisting of’ can be used in place of “comprising” and “including” to provide for more specific embodiments and are also disclosed.

[0067] An "increase" can refer to any change that results in a greater amount of a symptom, disease, composition, condition, or activity. An increase can be any individual, median, or average increase in a condition, symptom, activity, composition in a statistically significant amount. Thus, the increase can be a 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, or 100% increase so long as the increase is statistically significant.

[0068] A "decrease" can refer to any change that results in a smaller amount of a symptom, disease, composition, condition, or activity. A substance is also understood to decrease the genetic output of a gene when the genetic output of the gene product with the substance is less relative to the output of the gene product without the substance. Also, for example, a decrease can be a change in the symptoms of a disorder such that the symptoms are less than previously observed. A decrease can be any individual, median, or average decrease in a condition, symptom, activity, composition in a statistically significant amount. Thus, the decrease can be a 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, or 100% decrease so long as the decrease is statistically significant.

[0069] By “prevent” or other forms of the word, such as “preventing” or “prevention,” is meant to stop a particular event or characteristic, to stabilize or delay the development or progression of a particular event or characteristic, or to minimize the chances that a particular event or characteristic will occur. Prevent does not require comparison to a control as it is typically more absolute than, for example, reduce. As used herein, something could be reduced but not prevented, but something that is reduced could also be prevented. Likewise, something could be prevented but not reduced, but something that is prevented could also be reduced. It is understood that where reduce or prevent are used, unless specifically indicated otherwise, the use of the other word is also expressly disclosed.

[0070] The term “subject” refers to any individual who is the target of administration or treatment. The subject can be a vertebrate, for example, a mammal. In one aspect, the subject can be human, non-human primate, bovine, equine, porcine, canine, or feline. The subject can also be a guinea pig, rat, hamster, rabbit, mouse, or mole. Thus, the subject can be a human or veterinary patient.

[0071] The term “amino acid,” includes but is not limited to amino acids contained in the group consisting of alanine (Ala or A), cysteine (Cys or C), aspartic acid (Asp or D), glutamic acid (Glu or E), phenylalanine (Phe or F), glycine (Gly or G), histidine (His or H), isoleucine (He or I), lysine (Lys or K), leucine (Leu or L), methionine (Met or M), asparagine (Asn or N), proline (Pro or P), glutamine (Gin or Q), arginine (Arg or R), serine (Ser or S), threonine (Thr or T), valine (Vai or V), tryptophan (Trp or W), and tyrosine (Tyr or Y) residues. The term “amino acid residue” also may include amino acid residues contained in the group consisting of homocysteine, 2- Aminoadipic acid, N-Ethylasparagine, 3-Aminoadipic acid, Hydroxylysine, P-alanine, -Amino- propionic acid, allo-Hydroxylysine acid, 2-Aminobutyric acid, 3-Hydroxyproline, 4- Aminobutyric acid, 4-Hydroxyproline, piperidinic acid, 6-Aminocaproic acid, Isodesmosine, 2- Aminoheptanoic acid, allo-Isoleucine, 2-Aminoisobutyric acid, N-Methylglycine, sarcosine, 3- Aminoisobutyric acid, N-Methylisoleucine, 2-Aminopimelic acid, 6-N -Methyllysine, 2,4- Diaminobutyric acid, N-Methylvaline, Desmosine, Norvaline, 2,2'-Diaminopimelic acid, Norleucine, 2,3-Diaminopropionic acid, Ornithine, and N-Ethylglycine. Typically, the amide linkages of the peptides are formed from an amino group of the backbone of one amino acid and a carboxyl group of the backbone of another amino acid. It should also be noted that amino acids, and derivatives thereof (with exception of glycine) occurs in two isomeric forms: L-forms or D- forms. The L- and D- forms represent the same atoms of an amino acid, however the atoms can have different arrangements, which can impact the amino acid properties and functions. The two forms are similar in that they both occur naturally and comprise a central carbon atom, at least one hydrogen atom, a carboxylic group, an amine group, and a variable group. The two forms differ in that they are usually mirrored images of each other, wherein the location of the amine group varies. L-amino acids are used in protein synthesis, while D-amino acid are less common in protein synthesis. L-amino acids rotate counterclockwise or to the left in a process known as levorotation. D-amino acids rotate clockwise or to the right in a process known as dextrorotation. L-amino acids are used to synthesize proteins, while D-amino acids are found in the cell walls of bacteria.

[0072] As used herein, a ’’nucleobase” refers to a nitrogen containing compound comprising a heterocyclic aromatic features that bind to a 5 -carbon sugar to further form a nucleotide. Nucleobases can be grouped into two major forms: purines (adenine (A) and guanine (G)) and pyrimidines (cytosine (C), thymine (T), and uracil (U)).

[0073] A “nucleotide” is a compound consisting of a nucleoside, which consists of a nitrogenous base and a 5-carbon sugar, linked to a phosphate group forming the basic structural unit of nucleic acids, such as DNA or RNA. The of nucleotides are adenine (A), cytosine (C), guanine (G), thymine (T), and uracil (U), each of which are bound together by a phosphodiester bond to form a nucleic acid molecule.

[0074] A “nucleic acid” is a chemical compound that serves as the primary information-carrying molecules in cells and make up the cellular genetic material. Nucleic acids comprise nucleotides, which are the monomers made of a 5-carbon sugar (usually ribose or deoxyribose), a phosphate group, and a nitrogenous base. A nucleic acid can also be a deoxyribonucleic acid (DNA) or a ribonucleic acid (RNA). Nucleic acids can be double stranded nucleic acids (dsNA) or single stranded nucleic acids (ssNA). DNA and RNA can be single stranded nucleic acids.

[0075] A ’’fatty acid” refers to a hydrocarbon molecule comprising one or more chains of carbon and hydrogen atoms. Fatty acid molecules can exist as saturated (single bonded carbons) molecules or unsaturated (double or triple bonded carbons) molecules. Fatty acids exists as monomeric units of triglycerides, phospholipids, and cholesteryl esters, and are also the major component of lipids.

[0076] As used herein, the term “lipid” or “lipid-like” refers to a macromolecule that is soluble in nonpolar solvents. These molecules are usually hydrophobic or amphiphilic molecules; the amphiphilic nature of some lipids allows formation of structures such as vesicles, liposomes, membranes, and nanoparticles. Lipids can be categorized into fatty acids, glycerolipids, glycerophospholipids, sphingolipids, saccharolipids, polyketides, sterol lipids, and prenol lipids.

[0077] As used herein, a “carbohydrate” or a “saccharide” refers to a large family of organic compounds including, but not limited to sugars, starch, and cellulose, containing hydrogen and oxygen in similar ratios to water (2: 1) and used as structural materials in numerous biomolecules, such as DNA or RNA, and for energy storage with living tissues. Carbohydrates can be categorized into four groups including monosaccharides, disaccharides, oligosaccharides, and polysaccharides, wherein monosaccharides and disaccharides are the smallest forms of carbohydrates, commonly referred to as sugars, whereas oligosaccharides and polysaccharides are larger, complex structures used for energy storage of the structural foundation of nucleic acids and nucleotides.

[0078] As used herein, an “organic polymer” refers to a macromolecule, or a large molecule comprising hundreds or thousands of covalently bonded atoms, that is important to one or more biochemical / biophysical processes. The most common organic polymers include, but are not limited to nucleic acids (DNA or RNA), proteins, lipids, and carbohydrates. Organic polymers can also be natural organic polymers or synthetic organic polymers. Organic polymers comprise “monomeric units”, which refers to repeating molecular subunits covalently bound together to form chains of monomers. It should be understood that one subunit refers to a monomer, two subunits refer to a dimer, three subunits refer to a trimer, and so forth. Further, a polymer can comprise two or more monomers, dimers, trimer, and so forth. Thus, a polymer can comprise 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19,

[0079] 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45,

[0080] 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71,

[0081] 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97,

[0082] 98, 99, 100, 101, 102, 103, 104, 105, 106, 107, 108, 109, 110, 111, 112, 113, 114, 115, 116, 117, 118, 119, 120, 121, 122, 123, 124, 125, 126, 127, 128, 129, 130, 131, 132, 133, 134, 135, 136,

[0083] 137, 138, 139, 140, 141, 142, 143, 144, 145, 146, 147, 148, 149, 150, 151, 152, 153, 154, 155,

[0084] 156, 157, 158, 159, 160, 161, 162, 163, 164, 165, 166, 167, 168, 169, 170, 171, 172, 173, 174,

[0085] 175, 176, 177, 178, 179, 180, 181, 182, 183, 184, 185, 186, 187, 188, 189, 190, 191, 192, 193,

[0086] 194, 195, 196, 197, 198, 199, 200, 201, 202, 203, 204, 205, 206, 207, 208, 209, 210, 211, 212,

[0087] 213, 214, 215, 216, 217, 218, 219, 220, 221, 222, 223, 224, 225, 226, 227, 228, 229, 230, 231,

[0088] 232, 233, 234, 235, 236, 237, 238, 239, 240, 241, 242, 243, 244, 245, 246, 247, 248, 249, 250,

[0089] 251, 252, 253, 254, 255, 256, 257, 258, 259, 260, 261, 262, 263, 264, 265, 266, 267, 268, 269,

[0090] 270, 271, 272, 273, 274, 275, 276, 277, 278, 279, 280, 281, 282, 283, 284, 285, 286, 287, 288,

[0091] 289, 290, 291, 292, 293, 294, 295, 296, 297, 298, 299, 300, 301, 302, 303, 304, 305, 306, 307,

[0092] 308, 309, 310, 311, 312, 313, 314, 315, 316, 317, 318, 319, 320, 321, 322, 323, 324, 325, 326,

[0093] 327, 328, 329, 330, 331, 332, 333, 334, 335, 336, 337, 338, 339, 340, 341, 342, 343, 344, 345,

[0094] 346, 347, 348, 349, 350, 351, 352, 353, 354, 355, 356, 357, 358, 359, 360, 361, 362, 363, 364,

[0095] 365, 366, 367, 368, 369, 370, 371, 372, 373, 374, 375, 376, 377, 378, 379, 380, 381, 382, 383,

[0096] 384, 385, 386, 387, 388, 389, 390, 391, 392, 393, 394, 395, 396, 397, 398, 399, 400, 401, 402, 403, 404, 405, 406, 407, 408, 409, 410, 41 1 , 412, 413, 414, 415, 416, 417, 418, 419, 420, 421 ,

[0097] 422, 423, 424, 425, 426, 427, 428, 429, 430, 431, 432, 433, 434, 435, 436, 437, 438, 439, 440,

[0098] 441, 442, 443, 444, 445, 446, 447, 448, 449, 450, 451, 452, 453, 454, 455, 456, 457, 458, 459,

[0099] 460, 461, 462, 463, 464, 465, 466, 467, 468, 469, 470, 471, 472, 473, 474, 475, 476, 477, 478,

[0100] 479, 480, 481, 482, 483, 484, 485, 486, 487, 488, 489, 490, 491, 492, 493, 494, 495, 496, 497,

[0101] 498, 499, 500, 501, 502, 503, 504, 505, 506, 507, 508, 509, 510, 511, 512, 513, 514, 515, 516,

[0102] 517, 518, 519, 520, 521, 522, 523, 524, 525, 526, 527, 528, 529, 530, 531, 532, 533, 534, 535,

[0103] 536, 537, 538, 539, 540, 541, 542, 543, 544, 545, 546, 547, 548, 549, 550, 551, 552, 553, 554,

[0104] 555, 556, 557, 558, 559, 560, 561, 562, 563, 564, 565, 566, 567, 568, 569, 570, 571, 572, 573,

[0105] 574, 575, 576, 577, 578, 579, 580, 581, 582, 583, 584, 585, 586, 587, 588, 589, 590, 591, 592,

[0106] 593, 594, 595, 596, 597, 598, 599, 600, 601, 602, 603, 604, 605, 606, 607, 608, 609, 610, 611,

[0107] 612, 613, 614, 615, 616, 617, 618, 619, 620, 621, 622, 623, 624, 625, 626, 627, 628, 629, 630,

[0108] 631, 632, 633, 634, 635, 636, 637, 638, 639, 640, 641, 642, 643, 644, 645, 646, 647, 648, 649,

[0109] 650, 651, 652, 653, 654, 655, 656, 657, 658, 659, 660, 661, 662, 663, 664, 665, 666, 667, 668,

[0110] 669, 670, 671, 672, 673, 674, 675, 676, 677, 678, 679, 680, 681, 682, 683, 684, 685, 686, 687,

[0111] 688, 689, 690, 691, 692, 693, 694, 695, 696, 697, 698, 699, 700, 701, 702, 703, 704, 705, 706,

[0112] 707, 708, 709, 710, 711, 712, 713, 714, 715, 716, 717, 718, 719, 720, 721, 722, 723, 724, 725,

[0113] 726, 727, 728, 729, 730, 731, 732, 733, 734, 735, 736, 737, 738, 739, 740, 741, 742, 743, 744,

[0114] 745, 746, 747, 748, 749, 750, 751, 752, 753, 754, 755, 756, 757, 758, 759, 760, 761, 762, 763,

[0115] 764, 765, 766, 767, 768, 769, 770, 771, 772, 773, 774, 775, 776, 777, 778, 779, 780, 781, 782,

[0116] 783, 784, 785, 786, 787, 788, 789, 790, 791, 792, 793, 794, 795, 796, 797, 798, 799, 800, 801,

[0117] 802, 803, 804, 805, 806, 807, 808, 809, 810, 811, 812, 813, 814, 815, 816, 817, 818, 819, 820,

[0118] 821, 822, 823, 824, 825, 826, 827, 828, 829, 830, 831, 832, 833, 834, 835, 836, 837, 838, 839,

[0119] 840, 841, 842, 843, 844, 845, 846, 847, 848, 849, 850, 851, 852, 853, 854, 855, 856, 857, 858,

[0120] 859, 860, 861, 862, 863, 864, 865, 866, 867, 868, 869, 870, 871, 872, 873, 874, 875, 876, 877,

[0121] 878, 879, 880, 881, 882, 883, 884, 885, 886, 887, 888, 889, 890, 891, 892, 893, 894, 895, 896,

[0122] 897, 898, 899, 900, 901, 902, 903, 904, 905, 906, 907, 908, 909, 910, 911, 912, 913, 914, 915,

[0123] 916, 917, 918, 919, 920, 921, 922, 923, 924, 925, 926, 927, 928, 929, 930, 931, 932, 933, 934,

[0124] 935, 936, 937, 938, 939, 940, 941, 942, 943, 944, 945, 946, 947, 948, 949, 950, 951, 952, 953,

[0125] 954, 955, 956, 957, 958, 959, 960, 961, 962, 963, 964, 965, 966, 967, 968, 969, 970, 971, 972,

[0126] 973, 974, 975, 976, 977, 978, 979, 980, 981, 982, 983, 984, 985, 986, 987, 988, 989, 990, 991,

[0127] 992, 993, 994, 995, 996, 997, 998, 999, 1000, or more monomers, dimers, trimer, and so forth. A polymer can also comprise thousands, tens of thousands, hundreds of thousands, millions, or more monomers, dimers, trimers, and so forth.

[0128] The term “administer,” “administering”, or derivatives thereof refer to delivering a composition, substance, inhibitor, or medication to a subject or object by one or more the following routes: oral, topical, intravenous, subcutaneous, transcutaneous, transdermal, intramuscular, intrajoint, parenteral, intra-arteriole, intradermal, intraventricular, intracranial, intraperitoneal, intralesional, intranasal, rectal, vaginal, by inhalation or via an implanted reservoir. The term “parenteral” includes subcutaneous, intravenous, intramuscular, intra-articular, intra-synovial, intrastemal, intrathecal, intrahepatic, intralesional, and intracranial injections or infusion techniques.

[0129] The terms "cell," "cell line" and "cell culture" include progeny. It is also understood that all progenies may not be precisely identical in DNA content, due to deliberate or inadvertent mutations. Variant progeny that have the same function or biological property, as screened for in the originally transformed cell, are included. The "host cells" used in the present invention generally are prokaryotic or eukaryotic hosts.

[0130] The term “covalent bond” refers to a sharing of pairs of electrons between the atoms of two or more hydrophilic (water soluble) polymers or, a sharing of pairs of electrons between atoms of the hydrophilic polymer and a composition comprising the polypeptide, or vice-versa).

[0131] A “hydrogen bond” refers to a force of attraction or covalent interaction between a hydrogen atom and a negatively charged atom. Hydrogen bonds can be either intermolecular (occurring between two or more separate molecules) hydrogen bonds or intramolecular (occurring among atoms of the same molecule) hydrogen bonds. Hydrogen bonds can also occur between inorganic molecules, such as water (H2O) or organic molecules, including but not limited to nucleobases, nucleotides, nucleic acids, amino acids, peptides, polypeptides, proteins, carbohydrates, fatty acids, and lipids.

[0132] An "ester bond” refers to a covalent interaction or linkage between an atom that is double bonded to an oxygen atom bearing any alkyl or aryl group. As used herein, unless otherwise indicated, the term "alkyl" alone or in combination refers to a monovalent saturated aliphatic hydrocarbon radical having the indicated number of carbon atoms. The radical may be a linear or branched chain and, where specified, optionally substituted with one to three suitable substituents as defined above. Illustrative examples of alkyl groups include, but are not limited to, methyl, ethyl, n-propyl, n-butyl, n-pentyl, n-hexyl, isopropyl, isobutyl, isopentyl, amyl, sec-butyl, tertbutyl, tert-pentyl, n-heptyl, n-octyl, n-nonyl, n-decyl, n-dodecyl, n-tetradecyl, n-hexadecyl, n- octadecyl, n-eicosyl and the like. Preferred alkyl groups include methyl, ethyl, n-propyl and isopropyl. Preferred optional suitable substituents include halo, methoxy, ethoxy, cyano, nitro and amino. As used herein, unless otherwise indicated, the term "aryl" alone or in combination refers to a monovalent aromatic hydrocarbon radical having six to ten carbon atoms forming a carbocyclic ring and, where specified, optionally substituted with one to three suitable substituents as defined above. Illustrative examples of aryl groups include, but are not limited to, phenyl, naphthyl, tetrahydronaphthyl, indanyl and the like. Preferred aryl groups are phenyl and naphthyl, optionally mono- or disubstituted by identical or different suitable substituents selected from halo, cyano, C.sub.l-C.sub.3 alkyl, C.sub.3-C.sub.6 cycloalkyl, difluoromethyl, trifluoromethyl, C.sub.l-C.sub.3 alkoxy, difluoromethoxy and trifluoromethoxy.

[0133] An “amide bond” or commonly referred to a “peptide bond” refers to the covalent interaction or linkage between the amine group of one amino acid and the carboxyl group of another amino acid, resulting in the loss or release of a water molecule. Amide bonds are commonly found in peptides, polypeptides, and proteins, but can also occur in DNA or RNA.

[0134] An “ether bond” refers to the covalent interaction or linkage between an oxygen atom bonded to two alkyl groups, two aryl groups, or any combinations thereof.

[0135] The terms “treat,” “treating,” and grammatical variations thereof as used herein, include partially or completely delaying, alleviating, mitigating, or reducing the intensity of one or more attendant symptoms of a disorder or condition and / or alleviating, mitigating, or impeding one or more causes of a disorder or condition. Treatments according to the disclosure may be applied preventively, prophy tactically, palliatively, or remedially. Treatments are administered to a subject prior to onset (e.g., before obvious signs of the disease or disorder), during early onset (e.g., upon initial signs and symptoms of the disease or disorder), or after an established development of the disease or disorder.

[0136] A “therapeutic composition” refers to at least one substance, molecule, or compound suitable for administering to a subject, wherein the composition further includes a pharmaceutical carrier. A non-limiting example include a therapeutic composition comprises a nucleobase-poly- amino acid carrier and a sterile water-based solution.

[0137] As used herein, a “therapeutic benefit” refers to a benefit or effect obtained as a result of administering a treatment or therapeutic composition. Generally, the therapeutic benefit is a positive result that occurs as a result of a method used to treat or prevent a disease or disorder.

[0138] An "effective amount" is an amount sufficient to affect beneficial or desired results. An effective amount can be administered in one or more administrations, applications, or dosages. “Effective amount” encompasses, without limitation, an amount that can ameliorate, reverse, mitigate, prevent, or diagnose a symptom or sign of a medical condition or disorder. Unless dictated otherwise, explicitly or by context, an “effective amount” is not limited to a minimal amount sufficient to ameliorate a condition. The severity of a disease or disorder, as well as the ability of a treatment to prevent, treat, or mitigate, the disease or disorder can be measured, without implying any limitation, by a biomarker or by a clinical parameter.

[0139] "Pharmaceutically acceptable carrier" means a carrier or excipient that is useful in preparing a pharmaceutical or therapeutic composition that is generally safe and non-toxic and includes a carrier that is acceptable for veterinary and / or human pharmaceutical or therapeutic use. The terms "pharmaceutically acceptable carrier" can include, but are not limited to, phosphate buffered saline solution, water, emulsions (such as an oil / water or water / oil emulsion) and / or various types of wetting agents.

[0140] "Pharmaceutically acceptable" component can refer to a component that is not biologically or otherwise undesirable, i.e., the component may be incorporated into a pharmaceutical formulation of the invention and administered to a subject as described herein without causing significant undesirable biological effects or interacting in a deleterious manner with any of the other components of the formulation in which it is contained. When used in reference to administration to a human, the term generally implies the component has met the required standards of toxicological and manufacturing testing or that it is included on the Inactive Ingredient Guide prepared by the U.S. Food and Drug Administration.

[0141] As used herein, “pharmaceutically acceptable carrier” encompasses any excipient, diluent, filler, salt, buffer, stabilizer, solubilizer, lipid, stabilizer, or other material well known in the art for use in pharmaceutical formulations. The choice of a carrier for use in a composition will depend upon the intended route of administration for the composition. The preparation of pharmaceutically acceptable carriers and formulations containing these materials is described in, e.g., Remington's Pharmaceutical Sciences, 21st Edition, ed. University of the Sciences in Philadelphia, Lippincott, Williams & Wilkins, Philadelphia, PA, 2005. Examples of physiologically acceptable carriers include saline, glycerol, DMSO, buffers such as phosphate buffers, citrate buffer, and buffers with other organic acids; antioxidants including ascorbic acid; low molecular weight (less than about 10 residues) polypeptides; proteins, such as serum albumin, gelatin, or immunoglobulins; hydrophilic polymers such as polyvinylpyrrolidone; amino acids such as glycine, glutamine, asparagine, arginine or lysine; monosaccharides, disaccharides, and other carbohydrates including glucose, mannose, or dextrins; chelating agents such as EDTA; sugar alcohols such as mannitol or sorbitol; salt- forming counterions such as sodium; and / or nonionic surfactants such as TWEEN™ (ICI, Inc.; Bridgewater, New Jersey), polyethylene glycol (PEG), and PLURONICS™ (BASF; Florham Park, NJ). To provide for the administration of such dosages for the desired therapeutic treatment, compositions disclosed herein can advantageously comprise between about 0.1% and 99% by weight of the total of one or more of the subject compounds based on the weight of the total composition including carrier or diluent.

[0142] The term “interaction” refers to an action that occurs as two or more objects have an effect on one another either with or without physical contact. In terms of biological interactions, cell, proteins, and other macromolecules can have said effects on one another to impact biological functions, such as cell / tumor growth, cell death, and cell signaling pathways.

[0143] The terms “percent identity” and “% identity,” as applied to polynucleotide sequences, refer to the percentage of residue matches between at least two polynucleotide sequences aligned using a standardized algorithm. Such an algorithm may insert, in a standardized and reproducible way, gaps in the sequences being compared in order to optimize alignment between two sequences, and therefore achieve a more meaningful comparison of the two sequences. Percent identity for a nucleic acid sequence may be determined as understood in the art. (See, e.g., U.S. Pat. No. 7,396,664, which is incorporated herein by reference in its entirety). A suite of commonly used and freely available sequence comparison algorithms is provided by the National Center for Biotechnology Information (NCBI) Basic Local Alignment Search Tool (BLAST) (Altschul, S. F. et al. (1990) J. Mol. Biol. 215:403 410), which is available from several sources, including the NCBI, Bethesda, Md., at its website. The BLAST software suite includes various sequence analysis programs including “blastn,” that is used to align a known polynucleotide sequence with other polynucleotide sequences from a variety of databases. Also available is a tool called “BLAST 2 Sequences” that is used for direct pairwise comparison of two nucleotide sequences. “BLAST 2 Sequences” can be accessed and used interactively at the NCBI website. The “BLAST 2 Sequences” tool can be used for both blastn and blastp (discussed above).

[0144] Percent identity may be measured over the length of an entire defined polynucleotide sequence or may be measured over a shorter length, for example, over the length of a fragment taken from a larger, defined sequence, for instance, a fragment of at least 20, at least 30, at least 40, at least 50, at least 70, at least 100, or at least 200 contiguous nucleotides. Such lengths are exemplary only, and it is understood that any fragment length may be used to describe a length over which percentage identity may be measured.

[0145] A “variant,” “mutant,” or “derivative” of a particular nucleic acid sequence may be defined as a nucleic acid sequence having at least 50% sequence identity to the particular nucleic acid sequence over a certain length of one of the nucleic acid sequences using blastn with the “BLAST 2 Sequences” tool available at the National Center for Biotechnology Information's website. (See Tatiana A. Tatusova, Thomas L. Madden (1999), “Blast 2 sequences — a new tool for comparing protein and nucleotide sequences”, FEMS Microbiol Lett. 174:247-250). In some embodiments a variant polynucleotide may show, for example, at least 60%, at least 70%, at least 80%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% or greater sequence identity over a certain defined length relative to a reference polynucleotide.

[0146] A “nuclease” is an enzyme capable of cleaving the phosphodiester bonds between nucleotides of nucleic acids. Nuclease can possess properties to cause double or single stranded breaks to target nucleic acids. Nucleases are commonly used in CRISPR technology to modify a host genome to express or inhibit a target gene. Nucleases can be classified into two general categories: endonucleases and exonucleases. As used herein, an “endonuclease” refers to a nuclease enzyme which cleaves a nucleic acid or polynucleotide sequence by separating the inner nucleotides of the sequence. As used herein, an “exonuclease” refers to a nuclease enzyme which removes successive nucleotides from the end of a nucleic acid or polynucleotide sequence.

[0147] A “gene therapy” refers to a medical / clinic approach to treat or prevent a disease, condition, or disorder by correcting an underlying genetic issue. This therapeutic technique involves introducing a gene to a subject or patient to replace or alleviate consequences of the dysfunction gene(s) causing the disease, condition, or disorder.

[0148] The terms “transfect”, “transfecting”, “transfected”, “transfection”, or variations thereof refer to a process in which a cellular organism comprising DNA takes up additional DNA, e.g., an expression vector, by the cell or organism whether or not any coding sequences are in fact expressed. The terms "transfected host cell" and "transformed" refer to a cell in which DNA was introduced.

[0149] The term “incubate” refers to a process of establishing or maintaining an environment under conditions favorable to induce an event, such as an interaction, a reaction, or a development.

[0150] A “crosslinking reagent” or a “crosslinker” refers to molecules that contain two or more reactive groups capable of chemically attaching to functional groups of other proteins, nucleic acids, fatty acids, carbohydrates, or derivatives thereof. Carrier-base-ssNA Compositions

[0151] Polymeric delivery systems have been developed as techniques enabling introduction of a therapeutic substance, such as nucleic acids, into the body with minimal to no harmful side effects. Specifically, gene therapeutic methods are utilizing polymeric delivery systems to deliver or transfer genes to target cells or tissues. However, current gene therapies require cationic polymers to interact with nucleic acids thereby limiting the options of polymeric molecules to be used as carriers. As used herein, a “cationic polymer” refers to an organic polymer molecule comprising hundreds or thousands of covalently bonded atoms arranged into one or more monomeric units, wherein said atoms and / or monomeric units carry a positive (+) electrostatic charge. Therefore, it should be understood that for a cationic polymer to interact with nucleic acids, said polymer must form an electrostatic bond with the nucleic acid comprising an opposite, negative (-) charge. Because of the limiting nature of current polymeric carrier molecules in gene therapy practices, there remains a need to develop a non-limiting carrier that can form a complex with nucleic acids for delivery, treatment, and / or prevention of gene diseases and / or disorders.

[0152] The present disclosure provides a carrier-base composition comprising an organic polymer and at least one nucleobase forming a complex through hydrogen bonds with a single stranded nucleic acid (ssNA).

[0153] In one aspect, disclosed herein is a complex comprising a carrier-base, wherein said carrierbase comprises i) an organic polymer comprising one or more monomeric units and ii) at least one nucleobase, wherein at least one covalent bond binds the one or more monomeric units to the nucleobase(s), and a single stranded nucleic acid (ssNA), wherein at least one hydrogen bond binds the ssNA to the carrier-base to form the complex.

[0154] As used herein, “cargo” refers to any biologically active molecule capable of forming at least one hydrogen bond with a carrier-base, wherein said cargo and carrier-base form a reversible, nano-sized complex. Following formation of the complex, said complex may enter into a cell by any cellular uptake mechanism known to one of ordinary skill in the art, and said complex releases the cargo within the cell. As used herein, “biologically active” refers to a substance that can alter the chemical structure and / or activity of a cell in a desirable manner, or can selectively alter the chemical structure and / or activity of a cell type relative to another cell type in a desirable manner. In some embodiments, the biologically active substance is a single stranded nucleic acid (ssNA) including, but not limited to single-stranded DNA, single- stranded RNA, or a combination thereof. In some embodiments, the ssNA comprises messenger RNA (mRNA), transfer RNA (tRNA), ribosomal RNA (rRNA), microRNA (miRNA), long non-coding RNA (IncRNA), RNA aptamers, circular RNA (circRNA), short-interfering RNA (siRNA), small guide RNA (sgRNA), antisense DNA oligonucleotides, or antisense RNA oligonucleotides. In some embodiments, the carrier-base is biodegradable and / or biocompatible.

[0155] In some embodiments, the carrier-base comprises 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40,

[0156] 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66,

[0157] 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92,

[0158] 93, 94, 95, 96, 97, 98, 99, 100, or more covalent bonds.

[0159] In some embodiments, the complex comprises 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14,

[0160] 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40,

[0161] 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66,

[0162] 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92,

[0163] 93, 94, 95, 96, 97, 98, 99, 100, or more hydrogen bonds.

[0164] In some embodiments, the complex comprises an RNA-plex complex.

[0165] In some embodiments, the organic polymer comprises 2 or more monomeric units. In some embodiments, the organic polymer comprises 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17,

[0166] 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43,

[0167] 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69,

[0168] 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95,

[0169] 96, 97, 98, 99, 100, 101, 102, 103, 104, 105, 106, 107, 108, 109, 110, 111, 112, 113, 114, 115, 116, 117, 118, 119, 120, 121, 122, 123, 124, 125, 126, 127, 128, 129, 130, 131, 132, 133, 134,

[0170] 135, 136, 137, 138, 139, 140, 141, 142, 143, 144, 145, 146, 147, 148, 149, 150, 151, 152, 153,

[0171] 154, 155, 156, 157, 158, 159, 160, 161, 162, 163, 164, 165, 166, 167, 168, 169, 170, 171, 172,

[0172] 173, 174, 175, 176, 177, 178, 179, 180, 181, 182, 183, 184, 185, 186, 187, 188, 189, 190, 191,

[0173] 192, 193, 194, 195, 196, 197, 198, 199, 200, 201, 202, 203, 204, 205, 206, 207, 208, 209, 210,

[0174] 211, 212, 213, 214, 215, 216, 217, 218, 219, 220, 221, 222, 223, 224, 225, 226, 227, 228, 229,

[0175] 230, 231, 232, 233, 234, 235, 236, 237, 238, 239, 240, 241, 242, 243, 244, 245, 246, 247, 248,

[0176] 249, 250, 251, 252, 253, 254, 255, 256, 257, 258, 259, 260, 261, 262, 263, 264, 265, 266, 267,

[0177] 268, 269, 270, 271, 272, 273, 274, 275, 276, 277, 278, 279, 280, 281, 282, 283, 284, 285, 286,

[0178] 287, 288, 289, 290, 291, 292, 293, 294, 295, 296, 297, 298, 299, 300, 301, 302, 303, 304, 305,

[0179] 306, 307, 308, 309, 310, 311, 312, 313, 314, 315, 316, 317, 318, 319, 320, 321, 322, 323, 324,

[0180] 325, 326, 327, 328, 329, 330, 331, 332, 333, 334, 335, 336, 337, 338, 339, 340, 341, 342, 343,

[0181] 344, 345, 346, 347, 348, 349, 350, 351, 352, 353, 354, 355, 356, 357, 358, 359, 360, 361, 362, 363, 364, 365, 366, 367, 368, 369, 370, 371 , 372, 373, 374, 375, 376, 377 , 378, 379, 380, 381 , 382, 383, 384, 385, 386, 387, 388, 389, 390, 391, 392, 393, 394, 395, 396 , 397, 398, 399, 400, 401, 402, 403, 404, 405, 406, 407, 408, 409, 410, 411, 412, 413, 414, 415 , 416, 417, 418, 419, 420, 421, 422, 423, 424, 425, 426, 427, 428, 429, 430, 431, 432, 433, 434 , 435, 436, 437, 438, 439, 440, 441, 442, 443, 444, 445, 446, 447, 448, 449, 450, 451, 452, 453 , 454, 455, 456, 457, 458, 459, 460, 461, 462, 463, 464, 465, 466, 467, 468, 469, 470, 471, 472 , 473, 474, 475, 476, 477, 478, 479, 480, 481, 482, 483, 484, 485, 486, 487, 488, 489, 490, 491 , 492, 493, 494, 495, 496, 497, 498, 499, 500, 501, 502, 503, 504, 505, 506, 507, 508, 509, 510 , 511, 512, 513, 514, 515, 516, 517, 518, 519, 520, 521, 522, 523, 524, 525, 526, 527, 528, 529 , 530, 531, 532, 533, 534, 535, 536, 537, 538, 539, 540, 541, 542, 543, 544, 545, 546, 547, 548 , 549, 550, 551, 552, 553, 554, 555, 556, 557, 558, 559, 560, 561, 562, 563, 564, 565, 566, 567 , 568, 569, 570, 571, 572, 573, 574, 575, 576, 577, 578, 579, 580, 581, 582, 583, 584, 585, 586 , 587, 588, 589, 590, 591, 592, 593, 594, 595, 596, 597, 598, 599, 600, 601, 602, 603, 604, 605 , 606, 607, 608, 609, 610, 611, 612, 613, 614, 615, 616, 617, 618, 619, 620, 621, 622, 623, 624 , 625, 626, 627, 628, 629, 630, 631, 632, 633, 634, 635, 636, 637, 638, 639, 640, 641, 642, 643 , 644, 645, 646, 647, 648, 649, 650, 651, 652, 653, 654, 655, 656, 657, 658, 659, 660, 661, 662 , 663, 664, 665, 666, 667, 668, 669, 670, 671, 672, 673, 674, 675, 676, 677, 678, 679, 680, 681 , 682, 683, 684, 685, 686, 687, 688, 689, 690, 691, 692, 693, 694, 695, 696, 697, 698, 699, 700 , 701, 702, 703, 704, 705, 706, 707, 708, 709, 710, 711, 712, 713, 714, 715, 716, 717, 718, 719 , 720, 721, 722, 723, 724, 725, 726, 727, 728, 729, 730, 731, 732, 733, 734, 735, 736, 737, 738 , 739, 740, 741, 742, 743, 744, 745, 746, 747, 748, 749, 750, 751, 752, 753, 754, 755, 756, 757 , 758, 759, 760, 761, 762, 763, 764, 765, 766, 767, 768, 769, 770, 771, 772, 773, 774, 775, 776 , 777, 778, 779, 780, 781, 782, 783, 784, 785, 786, 787, 788, 789, 790, 791, 792, 793, 794, 795 , 796, 797, 798, 799, 800, 801, 802, 803, 804, 805, 806, 807, 808, 809, 810, 811, 812, 813, 814 , 815, 816, 817, 818, 819, 820, 821, 822, 823, 824, 825, 826, 827, 828, 829, 830, 831, 832, 833 , 834, 835, 836, 837, 838, 839, 840, 841, 842, 843, 844, 845, 846, 847, 848, 849, 850, 851, 852 , 853, 854, 855, 856, 857, 858, 859, 860, 861, 862, 863, 864, 865, 866, 867, 868, 869, 870, 871 , 872, 873, 874, 875, 876, 877, 878, 879, 880, 881, 882, 883, 884, 885, 886, 887, 888, 889, 890 , 891, 892, 893, 894, 895, 896, 897, 898, 899, 900, 901, 902, 903, 904, 905, 906, 907, 908, 909 , 910, 911, 912, 913, 914, 915, 916, 917, 918, 919, 920, 921, 922, 923, 924, 925, 926, 927, 928 , 929, 930, 931, 932, 933, 934, 935, 936, 937, 938, 939, 940, 941, 942, 943, 944, 945, 946, 947 , 948, 949, 950, 951, 952, 953, 954, 955, 956, 957, 958, 959, 960, 961, 962, 963, 964, 965, 966 , 967, 968, 969, 970, 971, 972, 973, 974, 975, 976, 977, 978, 979, 980, 981, 982, 983, 984, 985 , 986, 987, 988, 989, 990, 991 , 992, 993, 994, 995, 996, 997, 998, 999, 1000, or more monomeric units. In some embodiments, organic polymer comprises 2 or more dimers. In some embodiments, the polymer comprises 2 or more trimers. In some embodiments, the organic polymer comprises 2 or more tetramers. In some embodiments, the organic polymer comprises 2 or more pentamers. In some embodiments, the organic polymer comprises 2 or more hexamers. In some embodiments, the organic polymers comprises 2 or more heptamers. In some embodiments, the organic polymer comprises 2 or more octamers. In some embodiments, the organic polymer comprises 2 or more oligomers.

[0182] In some embodiments, the monomeric units are selected from an amino acid, a monosaccharide, a nucleotide, and a fatty acid.

[0183] In some embodiments, the organic polymer comprises a combination of at least two of an amino acid, a monosaccharide, a nucleotide, or a fatty acid.

[0184] In some embodiments, the monomeric units comprise at least two amino acids selected from alanine, cysteine, aspartic acid, glutamic acid, phenylalanine, glycine, histidine, isoleucine, lysine, leucine, methionine, asparagine, proline, glutamine, arginine, serine, threonine, valine, tryptophan, tyrosine, homocysteine, 2-Aminoadipic acid, N-Ethylasparagine, 3-Aminoadipic acid, Hydroxylysine, 0-alanine, -Amino-propionic acid, allo-Hydroxylysine acid, 2-Aminobutyric acid, 3-Hydroxyproline, 4-Aminobutyric acid, 4-Hydroxyproline, piperidinic acid, 6- Aminocaproic acid, Isodesmosine, 2-Aminoheptanoic acid, allo-Isoleucine, 2-Aminoisobutyric acid, N-Methylglycine, sarcosine, 3-Aminoisobutyric acid, N-Methylisoleucine, 2-Aminopimelic acid, 6-N-Methyllysine, 2,4-Diaminobutyric acid, N-Methylvaline, Desmosine, Norvaline, 2,2'- Diaminopimelic acid, Norleucine, 2,3-Diaminopropionic acid, Ornithine, -Ethylglycine, and any combinations thereof.

[0185] In some embodiments, the monomeric units comprise at least two monosaccharides including, but not limited to glucose, sucrose, ribose, deoxyribose, glyceraldehyde, pyranose, galactose, hexose, mannose, xylose, dihydroxyacetone, psicose, fructose, pentose, rhamnose, furanose, altrose, and combinations thereof.

[0186] In some embodiments, the monomeric units comprise at least two nucleotides including, but not limited to adenosine, guanosine, thymidine, uridine, cytidine, inosine, and combinations thereof.

[0187] In some embodiments, the monomeric units comprise at least two fatty acids including, but not limited to linoleic acid, myristic acid, palmitoleic acid, caproic acid, behenic acid, omega-6 fatty acid, omega-3 fatty acid, palmitic acid, lauric acid, arachidic acid, butyric acid, lignoceric acid, carboxylic acid, linolenic acid, capric acid, arachidonic acid, eicosapentaeonic acid, caprylic acid, y-linolenic acid, erucic acid, stearic acid, oleic acid, propionic acid, valeric acid, enanthic acid, pelargonic acid, undecylic acid, tridecylic acid, pentadecylic acid, margaric acid, nonadecylic acid, heneicosylic acid, tricosylic acid, pentacosylic acid, cerotic acid, carboceric acid, montanic acid, nonacosylic acid, melissic acid, hentriacontylic acid, lacceroic acid, psyllic acid, geddic acid, ceroplastic acid, hexatriacontylic acid, heptatriacontylic acid, octatriacontylic acid, nonatriacontylic acid, tetracontylic acid, and any combinations thereof.

[0188] In some embodiments, the organic polymer comprising 2 or more monomeric units comprises a poly-amino acid, a polysaccharide, a polynucleotide, a lipid, or combinations thereof.

[0189] In some embodiments, the poly-amino acid comprises 2 or more amino acids, or derivatives thereof. In some embodiments, the poly-amino acid comprises 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13,

[0190] 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 2i , 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 52 , 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 7< 80. 81. 82. 83. 84. 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98 , 99, 100, 101, 102, 103, 104, 105, 106, 107 , 108, 109, 110, 111, 112, 113, 114, 115, 116, 117, 118, 119, 120, 121, 122, 123, 124, 125, 126, 127, 128, 129, 130, 131, 132, 133, 134, 135, 136, 137, 138, 139, 140, 141, 142, 143, 144, 145, 146, 147, 148, 149, 150, 151, 152, 153, 154, 155, 156, 157, 158, 159, 160, 161, 162, 163, 164, 165, 166, 167, 168, 169, 170, 171, 172, 173, 174, 175, 176, 177, 178, 179, 180, 181, 182, 183, 184, 185, 186, 187, 188, 189, 190, 191, 192, 193, 194, 195, 196, 197, 198, 199, 200, 201, 202, 203, 204, 205, 206, 207, 208, 209, 210, 211, 212, 213, 214, 215, 216, 217, 218, 219, 220, 221, 222, 223, 224, 225, 226, 227, 228, 229, 230, 231, 232, 233, 234, 235, 236, 237, 238, 239, 240, 241, 242, 243, 244, 245, 246, 247, 248, 249, 250, 251, 252, 253, 254, 255, 256, 257, 258, 259, 260, 261, 262, 263, 264, 265, 266, 267, 268, 269, 270, 271, 272, 273, 274, 275, 276, 277, 278, 279, 280, 281, 282, 283, 284, 285, 286, 287, 288, 289, 290, 291, 292, 293, 294, 295, 296, 297, 298, 299, 300, 301, 302, 303, 304, 305, 306, 307, 308, 309, 310, 311, 312, 313, 314, 315, 316, 317, 318, 319, 320, 321, 322, 323, 324, 325, 326, 327, 328, 329, 330, 331, 332, 333, 334, 335, 336, 337, 338, 339, 340, 341, 342, 343, 344, 345, 346, 347, 348, 349, 350, 351, 352, 353, 354, 355, 356, 357, 358, 359, 360, 361, 362, 363, 364, 365, 366, 367, 368, 369, 370, 371, 372, 373, 374, 375, 376, 377, 378, 379, 380, 381, 382, 383, 384, 385, 386, 387, 388, 389, 390, 391, 392, 393, 394, 395, 396, 397, 398, 399, 400, 401, 402, 403, 404, 405, 406, 407, 408, 409, 410, 411, 412, 413, 414, 415, 416, 417, 418, 419, 420, 421, 422, 423, 424, 425, 426, 427, 428, 429, 430, 431, 432, 433, 434, 435, 436, 437, 438, 439, 440, 441. 442. 443. 444. 445. 446. 447. 448. 449. 450, 451, 452, 453, 454, 455, 456, 457, 458, 459, 460, 461 , 462, 463, 464, 465, 466, 467, 468, 469, 470, 471 , 472, 473, 474, 475, 476, 477, 478, 479, 480, 481, 482, 483, 484, 485, 486, 487, 488, 489, 490, 491, 492, 493, 494, 495, 496, 497, 498, 499, 500, or more amino acids, or derivatives thereof. In some embodiments, the 2 or more amino acids comprise a L-, D-, or DL-amino acid.

[0191] In some embodiments, the poly-amino acid ranges between about IkDa to about 200kDa. In some embodiments, the poly amino acid ranges between about IkDa to about 150kDa. In some embodiments, the poly-amino acid ranges between about IkDa to about lOOkDa. In some embodiments, the poly-amino acid ranges between about 1 kDa to about 50 kDa. In some embodiments, the poly-amino acid is 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19,

[0192] 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45,

[0193] 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71,

[0194] 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97,

[0195] 98, 99, 100, 101, 102, 103, 104, 105, 106, 107, 108, 109, 110, 111, 112, 113, 114, 115, 116, 117, 118, 119, 120, 121, 122, 123, 124, 125, 126, 127, 128, 129, 130, 131, 132, 133, 134, 135, 136,

[0196] 137, 138, 139, 140, 141, 142, 143, 144, 145, 146, 147, 148, 149, 150, 151, 152, 153, 154, 155,

[0197] 156, 157, 158, 159, 160, 161, 162, 163, 164, 165, 166, 167, 168, 169, 170, 171, 172, 173, 174,

[0198] 175, 176, 177, 178, 179, 180, 181, 182, 183, 184, 185, 186, 187, 188, 189, 190, 191, 192, 193,

[0199] 194, 195, 196, 197, 198, 199, 200kDa, or more.

[0200] In some embodiments, the polysaccharide comprises 2 or more monosaccharides. In some embodiments, the polysaccharide comprises 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18,

[0201] 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44,

[0202] 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70,

[0203] 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96,

[0204] 97, 98, 99, 100, 101, 102, 103, 104, 105, 106, 107, 108, 109, 110, 111, 112, 113, 114, 115, 116, 117, 118, 119, 120, 121, 122, 123, 124, 125, 126, 127, 128, 129, 130, 131, 132, 133, 134, 135,

[0205] 136, 137, 138, 139, 140, 141, 142, 143, 144, 145, 146, 147, 148, 149, 150, 151, 152, 153, 154,

[0206] 155, 156, 157, 158, 159, 160, 161, 162, 163, 164, 165, 166, 167, 168, 169, 170, 171, 172, 173,

[0207] 174, 175, 176, 177, 178, 179, 180, 181, 182, 183, 184, 185, 186, 187, 188, 189, 190, 191, 192,

[0208] 193, 194, 195, 196, 197, 198, 199, 200, 201, 202, 203, 204, 205, 206, 207, 208, 209, 210, 211,

[0209] 212, 213, 214, 215, 216, 217, 218, 219, 220, 221, 222, 223, 224, 225, 226, 227, 228, 229, 230,

[0210] 231, 232, 233, 234, 235, 236, 237, 238, 239, 240, 241, 242, 243, 244, 245, 246, 247, 248, 249,

[0211] 250, 251, 252, 253, 254, 255, 256, 257, 258, 259, 260, 261, 262, 263, 264, 265, 266, 267, 268,

[0212] 269, 270, 271, 272, 273, 274, 275, 276, 277, 278, 279, 280, 281, 282, 283, 284, 285, 286, 287, 288, 289, 290, 291 , 292, 293, 294, 295, 296, 297, 298, 299, 300, 301 , 302, 303, 304, 305, 306,

[0213] 307, 308, 309, 310, 311, 312, 313, 314, 315, 316, 317, 318, 319, 320, 321, 322, 323, 324, 325,

[0214] 326, 327, 328, 329, 330, 331, 332, 333, 334, 335, 336, 337, 338, 339, 340, 341, 342, 343, 344,

[0215] 345, 346, 347, 348, 349, 350, 351, 352, 353, 354, 355, 356, 357, 358, 359, 360, 361, 362, 363,

[0216] 364, 365, 366, 367, 368, 369, 370, 371, 372, 373, 374, 375, 376, 377, 378, 379, 380, 381, 382,

[0217] 383, 384, 385, 386, 387, 388, 389, 390, 391, 392, 393, 394, 395, 396, 397, 398, 399, 400, 401,

[0218] 402, 403, 404, 405, 406, 407, 408, 409, 410, 411, 412, 413, 414, 415, 416, 417, 418, 419, 420,

[0219] 421, 422, 423, 424, 425, 426, 427, 428, 429, 430, 431, 432, 433, 434, 435, 436, 437, 438, 439,

[0220] 440, 441, 442, 443, 444, 445, 446, 447, 448, 449, 450, 451, 452, 453, 454, 455, 456, 457, 458,

[0221] 459, 460, 461, 462, 463, 464, 465, 466, 467, 468, 469, 470, 471, 472, 473, 474, 475, 476, 477,

[0222] 478, 479, 480, 481, 482, 483, 484, 485, 486, 487, 488, 489, 490, 491, 492, 493, 494, 495, 496,

[0223] 497, 498, 499, 500, or more monosaccharides, or derivatives thereof.

[0224] In some embodiments, the polynucleotide comprises 2 or more nucleotides. In some embodiments, the polynucleotide comprises 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18,

[0225] 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44,

[0226] 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70,

[0227] 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96,

[0228] 97, 98, 99, 100, 101, 102, 103, 104, 105, 106, 107, 108, 109, 110, 111, 112, 113, 114, 115, 116, 117, 118, 119, 120, 121, 122, 123, 124, 125, 126, 127, 128, 129, 130, 131, 132, 133, 134, 135,

[0229] 136, 137, 138, 139, 140, 141, 142, 143, 144, 145, 146, 147, 148, 149, 150, 151, 152, 153, 154,

[0230] 155, 156, 157, 158, 159, 160, 161, 162, 163, 164, 165, 166, 167, 168, 169, 170, 171, 172, 173,

[0231] 174, 175, 176, 177, 178, 179, 180, 181, 182, 183, 184, 185, 186, 187, 188, 189, 190, 191, 192,

[0232] 193, 194, 195, 196, 197, 198, 199, 200, 201, 202, 203, 204, 205, 206, 207, 208, 209, 210, 211,

[0233] 212, 213, 214, 215, 216, 217, 218, 219, 220, 221, 222, 223, 224, 225, 226, 227, 228, 229, 230,

[0234] 231, 232, 233, 234, 235, 236, 237, 238, 239, 240, 241, 242, 243, 244, 245, 246, 247, 248, 249,

[0235] 250, 251, 252, 253, 254, 255, 256, 257, 258, 259, 260, 261, 262, 263, 264, 265, 266, 267, 268,

[0236] 269, 270, 271, 272, 273, 274, 275, 276, 277, 278, 279, 280, 281, 282, 283, 284, 285, 286, 287,

[0237] 288, 289, 290, 291, 292, 293, 294, 295, 296, 297, 298, 299, 300, 301, 302, 303, 304, 305, 306,

[0238] 307, 308, 309, 310, 311, 312, 313, 314, 315, 316, 317, 318, 319, 320, 321, 322, 323, 324, 325,

[0239] 326, 327, 328, 329, 330, 331, 332, 333, 334, 335, 336, 337, 338, 339, 340, 341, 342, 343, 344,

[0240] 345, 346, 347, 348, 349, 350, 351, 352, 353, 354, 355, 356, 357, 358, 359, 360, 361, 362, 363,

[0241] 364, 365, 366, 367, 368, 369, 370, 371, 372, 373, 374, 375, 376, 377, 378, 379, 380, 381, 382,

[0242] 383, 384, 385, 386, 387, 388, 389, 390, 391, 392, 393, 394, 395, 396, 397, 398, 399, 400, 401, 402, 403, 404, 405, 406, 407, 408, 409, 410, 41 1 , 412, 413, 414, 415, 416, 417, 418, 419, 420,

[0243] 421, 422, 423, 424, 425, 426, 427, 428, 429, 430, 431, 432, 433, 434, 435, 436, 437, 438, 439,

[0244] 440, 441, 442, 443, 444, 445, 446, 447, 448, 449, 450, 451, 452, 453, 454, 455, 456, 457, 458,

[0245] 459, 460, 461, 462, 463, 464, 465, 466, 467, 468, 469, 470, 471, 472, 473, 474, 475, 476, 477,

[0246] 478, 479, 480, 481, 482, 483, 484, 485, 486, 487, 488, 489, 490, 491, 492, 493, 494, 495, 496,

[0247] 497, 498, 499, 500, or more nucleotides, or derivative thereof.

[0248] In some embodiments, the lipid comprises 2 or more fatty acids. In some embodiments, the lipid comprises 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24,

[0249] 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50,

[0250] 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76,

[0251] 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, 101,

[0252] 102, 103, 104, 105, 106, 107, 108, 109, 110, 111, 112, 113, 114, 115, 116, 117, 118, 119, 120,

[0253] 121, 122, 123, 124, 125, 126, 127, 128, 129, 130, 131, 132, 133, 134, 135, 136, 137, 138, 139,

[0254] 140, 141, 142, 143, 144, 145, 146, 147, 148, 149, 150, 151, 152, 153, 154, 155, 156, 157, 158,

[0255] 159, 160, 161, 162, 163, 164, 165, 166, 167, 168, 169, 170, 171, 172, 173, 174, 175, 176, 177,

[0256] 178, 179, 180, 181, 182, 183, 184, 185, 186, 187, 188, 189, 190, 191, 192, 193, 194, 195, 196,

[0257] 197, 198, 199, 200, 201, 202, 203, 204, 205, 206, 207, 208, 209, 210, 211, 212, 213, 214, 215,

[0258] 216, 217, 218, 219, 220, 221, 222, 223, 224, 225, 226, 227, 228, 229, 230, 231, 232, 233, 234,

[0259] 235, 236, 237, 238, 239, 240, 241, 242, 243, 244, 245, 246, 247, 248, 249, 250, 251, 252, 253,

[0260] 254, 255, 256, 257, 258, 259, 260, 261, 262, 263, 264, 265, 266, 267, 268, 269, 270, 271, 272,

[0261] 273, 274, 275, 276, 277, 278, 279, 280, 281, 282, 283, 284, 285, 286, 287, 288, 289, 290, 291,

[0262] 292, 293, 294, 295, 296, 297, 298, 299, 300, 301, 302, 303, 304, 305, 306, 307, 308, 309, 310,

[0263] 311, 312, 313, 314, 315, 316, 317, 318, 319, 320, 321, 322, 323, 324, 325, 326, 327, 328, 329,

[0264] 330, 331, 332, 333, 334, 335, 336, 337, 338, 339, 340, 341, 342, 343, 344, 345, 346, 347, 348,

[0265] 349, 350, 351, 352, 353, 354, 355, 356, 357, 358, 359, 360, 361, 362, 363, 364, 365, 366, 367,

[0266] 368, 369, 370, 371, 372, 373, 374, 375, 376, 377, 378, 379, 380, 381, 382, 383, 384, 385, 386,

[0267] 387, 388, 389, 390, 391, 392, 393, 394, 395, 396, 397, 398, 399, 400, 401, 402, 403, 404, 405,

[0268] 406, 407, 408, 409, 410, 411, 412, 413, 414, 415, 416, 417, 418, 419, 420, 421, 422, 423, 424,

[0269] 425, 426, 427, 428, 429, 430, 431, 432, 433, 434, 435, 436, 437, 438, 439, 440, 441, 442, 443,

[0270] 444, 445, 446, 447, 448, 449, 450, 451, 452, 453, 454, 455, 456, 457, 458, 459, 460, 461, 462,

[0271] 463, 464, 465, 466, 467, 468, 469, 470, 471, 472, 473, 474, 475, 476, 477, 478, 479, 480, 481,

[0272] 482, 483, 484, 485, 486, 487, 488, 489, 490, 491, 492, 493, 494, 495, 496, 497, 498, 499, 500, or more fatty acids, or derivatives thereof. In some embodiments, the polymer comprises a silica- base nanoparticle.

[0273] In some embodiments, the complex comprises one, two, three, four, five, six, or more nucleobases. In some embodiments, the complex comprises six nucleobases selected from thymine, uracil, cytosine, adenine, guanine, and hypoxanthine. In some embodiments, the complex comprises adenine, guanine, or a combination thereof. In some embodiments, a combination of any nucleobase of any preceding aspect includes, but is not limited to a 1: 1 mixture, a 1:2 mixture, and a 2: 1 mixture. In some embodiments, the combination of adenine and guanine includes, but is not limited to a 1:1 mixture, a 1:2 mixture, and a 2:1 mixture.

[0274] In some embodiments, the complex comprises a ratio of about 1: 1 to about 1:1000 of monomeric units relative to nucleobases. In some embodiments, the complex comprises a ratio of about 1:1 to about 1:500 of monomeric units relative to nucleobases. In some embodiments, the complex comprises a ratio of about 1: 1 to about 1:250 of monomeric units relative to nucleobases. In some embodiments, the complex comprises a ratio of about 1 :1 to about 1:100 of monomeric units relative to nucleobases. In some embodiments, the complex comprises a ratio of about 1:1 to about 1:50 of monomeric units relative to nucleobases. In some embodiments, the complex comprises a ratio of about 1:1 to about 1:25 of monomeric units relative to nucleobases. In some embodiments, the complex comprises a ratio of about 1:1 to about 1:10. In some embodiments, the complex comprises a ratio of about 1:1 to about 1:5.

[0275] In some embodiments, the ssNA comprises single-stranded DNA, single-stranded RNA, or a combination thereof. In some embodiments, the ssNA comprises messenger RNA (mRNA), transfer RNA (tRNA), ribosomal RNA (rRNA), microRNA (miRNA), long non-coding RNA (IncRNA), RNA aptamers, circular RNA (circRNA), short-interfering RNA (siRNA), small guide RNA (sgRNA), antisense DNA oligonucleotides, or antisense RNA oligonucleotides.

[0276] In some embodiments, the at least one covalent bonds comprises an ester bond, an ether bond, or an amide bond.

[0277] In one aspect, disclosed herein is a therapeutic composition comprising the complex of any preceding aspect wherein the ssNA has a therapeutic benefit. As used herein, a “therapeutic benefit” refers to a benefit or effect obtained as a result of treatment. Further, the therapeutic benefit comprises a positive result that occurs as a result of a method of treating and / or preventing a disease or disorder. In some embodiments, the therapeutic benefit may be referred to as a therapeutic effect. In some embodiments, the therapeutic composition further comprises a pharmaceutically acceptable carrier selected from an excipient, a diluent, a salt, a buffer, a stabilizer, a lipid, an emulsion, and a nanoparticle.

[0278] In some embodiments, the therapeutic composition further comprises a transfection system selected from a lipofectamine composition (including, but not limited to Lipofectamine 2000), a lipid nanoparticle (LNP), in vivo jetRNA plus, or polyethylenimine (PEI).

[0279] Methods of treating and / or preventing diseases or disorders

[0280] In one aspect, disclosed herein is a method of treating or preventing a disease or disorder in a subject, the method comprising administering an effective amount of the therapeutic composition of any preceding aspect, wherein the ssNA is internalized and released into a cell of the subject to treat or prevent the disease or disorder.

[0281] The therapeutic composition may be administered in such amounts, time, and route deemed necessary in order to achieve the desired result. The exact amount of the therapeutic composition will vary from subject to subject, depending on the species, age, and general condition of the subject, the severity of the disease or disorder, the particular composition, its mode of administration, its mode of activity, and the like. The therapeutic composition is preferably formulated in dosage unit form for ease of administration and uniformity of dosage. It will be understood, however, that the total daily usage of the therapeutic composition will be decided by the attending physician within the scope of sound medical judgment. The specific therapeutically effective dose level for any particular subject will depend upon a variety of factors including the disease or disorder being treated and the severity of the disease or disorder; the activity of the composition employed; the specific composition employed; the age, body weight, general health, sex and diet of the patient; the time of administration, route of administration, and rate of excretion of the specific composition employed; the duration of the treatment; drugs used in combination or coincidental with the specific composition employed; and like factors well known in the medical arts.

[0282] The therapeutic composition may be administered by any route. In some embodiments, the therapeutic composition is administered via a variety of routes, including oral, intravenous, intramuscular, intra-arterial, intramedullary, intrathecal, subcutaneous, intraventricular, transdermal, interdermal, rectal, intravaginal, intraperitoneal, topical (as by powders, ointments, creams, and / or drops), mucosal, nasal, buccal, enteral, sublingual; by intratracheal instillation, bronchial instillation, and / or inhalation; and / or as an oral spray, nasal spray, and / or aerosol. In general, the most appropriate route of administration will depend upon a variety of factors including the nature of the therapeutic composition (e.g., its stability in the environment of the tissue or organ), the condition of the subject (e.g., whether the subject is able to tolerate the selected route of administration), etc.

[0283] The exact amount of therapeutic composition required to achieve a therapeutically or prophylactically effective amount will vary from subject to subject, depending on species, age, and general condition of a subject, severity of the side effects, identity of the particular compound(s), mode of administration, and the like. The amount to be administered to, for example, a child or an adolescent can be determined by a medical practitioner or person skilled in the art and can be lower or the same as that administered to an adult.

[0284] In some embodiments, the composition is administered 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38,

[0285] 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64,

[0286] 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90,

[0287] 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, or more times. In some embodiments, the composition is administered daily. In some embodiments, the composition is administered every day, every 2 days, every 3 days, every 4 days, every 5 days, every 6 days, every 7 days, or more. In some embodiments, the composition is administered every week, every 2 weeks, every 3 weeks, every 4 weeks, or more. In some embodiments, the composition is administered every month, every 2 months, every 3 months, every 4 months, every 5 months, every 6 months, every 7 months, every 8 months, every 9 months, every 10 months, every 11 months, every 12 months, or more. In some embodiments, the composition is administered every year, every 2 years, every 3 years, every 4 years, every 5 years, or more.

[0288] In some embodiments, the composition is used to treat or prevent cancer. Exemplary cancers include, but not limited to acoustic neuroma, adenocarcinoma, adrenal gland cancer, anal cancer, angiosarcoma (e.g., lymphangiosarcoma, lymphangioendotheliosarcoma, hemangiosarcoma), appendix cancer, benign monoclonal gammopathy, biliary cancer (e.g., cholangiocarcinoma), bladder cancer, breast cancer (e.g., adenocarcinoma of the breast, papillary carcinoma of the breast, mammary cancer, medullary carcinoma of the breast), brain cancer (e.g., meningioma; glioma, e.g., astrocytoma, oligodendroglioma; medulloblastoma), bronchus cancer, carcinoid tumor, cervical cancer (e.g., cervical adenocarcinoma), choriocarcinoma, chordoma, craniopharyngioma, colorectal cancer (e.g., colon cancer, rectal cancer, colorectal adenocarcinoma), epithelial carcinoma, ependymoma, endotheliosarcoma (e.g., Kaposi's sarcoma, multiple idiopathic hemorrhagic sarcoma), endometrial cancer (e.g., uterine cancer, uterine sarcoma), esophageal cancer (e.g., adenocarcinoma of the esophagus, Barrett's adenocarinoma), Ewing's sarcoma, eye cancer (e.g., intraocular melanoma, retinoblastoma), familiar hypereosinophilia, gall bladder cancer, gastric cancer (e.g., stomach adenocarcinoma), gastrointestinal stromal tumor (GIST), head and neck cancer (e.g., head and neck squamous cell carcinoma, oral cancer (e.g., oral squamous cell carcinoma (OSCC), throat cancer (e.g., laryngeal cancer, pharyngeal cancer, nasopharyngeal cancer, oropharyngeal cancer)), hematopoietic cancers (e.g., leukemia such as acute lymphocytic leukemia (ALL) (e.g., B-cell ALL, T-cell ALL), acute myelocytic leukemia (AML) (e.g., B-cell AML, T-cell AML), chronic myelocytic leukemia (CML) (e.g., B-cell CML, T-cell CML), and chronic lymphocytic leukemia (CLL) (e.g., B-cell CLL, T-cell CLL); lymphoma such as Hodgkin lymphoma (HL) (e.g., B-cell HL, T-cell HL) and non- Hodgkin lymphoma (NHL) (e.g., B-cell NHL such as diffuse large cell lymphoma (DLCL) (e.g., diffuse large B-cell lymphoma (DLBCL)), follicular lymphoma, chronic lymphocytic leukemia / small lymphocytic lymphoma (CLL / SLL), mantle cell lymphoma (MCL), marginal zone B-cell lymphomas (e.g., mucosa-associated lymphoid tissue (MALT) lymphomas, nodal marginal zone B-cell lymphoma, splenic marginal zone B-cell lymphoma), primary mediastinal B-cell lymphoma, Burkitt lymphoma, lymphoplasmacytic lymphoma (i.e., “Waldenstrom’s macroglobulinemia”), hairy cell leukemia (HCL), immunoblastic large cell lymphoma, precursor B-lymphoblastic lymphoma and primary central nervous system (CNS) lymphoma; and T-cell NHL such as precursor T-lymphoblastic lymphoma / leukemia, peripheral T-cell lymphoma (PTCL) (e.g., cutaneous T-cell lymphoma (CTCL) (e.g., mycosis fungiodes, Sezary syndrome), angioimmunoblastic T-cell lymphoma, extranodal natural killer T-cell lymphoma, enteropathy type T-cell lymphoma, subcutaneous panniculitis -like T-cell lymphoma, anaplastic large cell lymphoma); a mixture of one or more leukemia / lymphoma as described above; and multiple myeloma (MM)), heavy chain disease (e.g., alpha chain disease, gamma chain disease, mu chain disease), hemangioblastoma, inflammatory myofibroblastic tumors, immunocytic amyloidosis, kidney cancer (e.g., nephroblastoma a.k.a. Wilms' tumor, renal cell carcinoma), liver cancer (e.g., hepatocellular cancer (HCC), malignant hepatoma), lung cancer (e.g., bronchogenic carcinoma, small cell lung cancer (SCLC), non-small cell lung cancer (NSCLC), adenocarcinoma of the lung), leiomyosarcoma (LMS), mastocytosis (e.g., systemic mastocytosis), myelodysplastic syndrome (MDS), mesothelioma, myeloproliferative disorder (MPD) (e.g., polycythemia Vera (PV), essential thrombocytosis (ET), agnogenic myeloid metaplasia (AMM) a.k.a. myelofibrosis (MF), chronic idiopathic myelofibrosis, chronic myelocytic leukemia (CML), chronic neutrophilic leukemia (CNL), hypereosinophilic syndrome (HES)), neuroblastoma, neurofibroma (e.g., neurofibromatosis (NF) type 1 or type 2, schwannomatosis), neuroendocrine cancer (e.g., gastroenteropancreatic neuroendoctrine tumor (GEP-NET), carcinoid tumor), osteosarcoma, ovarian cancer (e.g., cystadenocarcinoma, ovarian embryonal carcinoma, ovarian adenocarcinoma), papillary adenocarcinoma, pancreatic cancer (e.g., pancreatic adenocarcinoma, intraductal papillary mucinous neoplasm (IPMN), Islet cell tumors), penile cancer (e.g., Paget's disease of the penis and scrotum), pinealoma, primitive neuroectodermal tumor (PNT), prostate cancer (e.g., prostate adenocarcinoma), rectal cancer, rhabdomyosarcoma, salivary gland cancer, skin cancer (e.g., squamous cell carcinoma (SCC), keratoacanthoma (KA), melanoma, basal cell carcinoma (BCC)), small bowel cancer (e.g., appendix cancer), soft tissue sarcoma (e.g., malignant fibrous histiocytoma (MFH), liposarcoma, malignant peripheral nerve sheath tumor (MPNST), chondrosarcoma, fibrosarcoma, myxosarcoma), sebaceous gland carcinoma, sweat gland carcinoma, synovioma, testicular cancer (e.g., seminoma, testicular embryonal carcinoma), thyroid cancer (e.g., papillary carcinoma of the thyroid, papillary thyroid carcinoma (PTC), medullary thyroid cancer), urethral cancer, vaginal cancer and vulvar cancer (e.g., Paget's disease of the vulva).

[0289] In some embodiments, the composition is used to treat or prevent neurodegenerative diseases. Exemplary neurodegenerative disease include, but are not limited to Alzheimer’ s disease, ataxia, Huntington’s disease, Parkinson’s disease, amyotrophic lateral sclerosis (ALS), Friedreich ataxia, Lewy body disease, spinal muscular atrophy, Alpers’ disease, Batten disease, Cerebro- oculo-facio-skeletal syndrome, Leigh syndrome, Prion diseases, monomelic amyotrophy, multiple system atrophy, striatonigral degeneration, motor neuron disease, multiple sclerosis (MS), Creutzfeldt-J akob disease, Parkinsonism, spinocerebellar ataxia, dementia, and other related diseases.

[0290] In some embodiments, the composition is used to treat or prevent an infectious disease. Exemplary infectious diseases include, but are not limited to common cold, influenza ( including, but not limited to human, bovine, avian, porcine, and simian strains of influenza), measles, acquired immune deficiency syndrome / human immunodeficiency virus (AIDS / HIV), anthrax, botulism, cholera, Campylobacter infections, chickenpox, chlamydia infections, cryptosporidosis, dengue fever, diphtheria, hemorrhagic fevers, Escherichia coli (E. coll) infections, ehrlichiosis, gonorrhea, hand-foot-mouth disease, hepatitis A, hepatitis B, hepatitis C, legionellosis, leprosy, leptospirosis, listeriosis, malaria, meningitis, meningococcal disease, mumps, pertussis, polio, pneumococcal disease, paralytic shellfish poisoning, rabies, rocky mountain spotted fever, rubella, salmonella, shigellosis, small pox, syphilis, tetanus, trichinosis (trichinellosis), tuberculosis (TB), typhoid fever, typhus, west nile virus, yellow fever, yersiniosis, and zika.

[0291] In some embodiments, the composition is used to treat or prevent a cardiovascular disease. Exemplary cardiovascular diseases include, but are not coronary artery disease, high / low blood pressure, cardiac arrest / heart failure, congestive heart failure, congenital heart defects / diseases (including, but not limited to atrial septal defects, atrioventricular septal defects, coarctation of the aorta, double-outlet right ventricle, d-transposition of the great arteries, Ebstein anomaly, hypoplastic left heart syndrome, and interrupted aortic arch), arrhythmia, peripheral artery disease, stroke, cerebrovascular disease, renal artery stenosis, aortic aneurysm, cardiomyopathies, hypertensive heart disease, pulmonary heart disease, cardiac dysrhythmias, endocarditis, inflammatory cardiomegaly, myocarditis, eosinophilic myocarditis, valvular heart diseases, rheumatic heart diseases, and other related cardiovascular diseases.

[0292] In some embodiments, the composition is used to treat or prevent a metabolic disease. Exemplary metabolic diseases include, but are not limited to diabetes mellitus Type I, diabetes mellitus Type II, familial hypercholesterolemia, Gaucher disease, Hunter syndrome, Krabbe syndrome, metachromatic leukodystrophy, Niemann-Pick syndrome, phenylketonuria (PKU), Tay-Sachs disease, Wilson’s disease, hemachromatosis, mitochondrial disorders or diseases (including, but not limited to Alpers Disease; Barth syndrome; beta. -oxidation defects :camitine- acyl-carnitine deficiency; carnitine deficiency; coenzyme Q10 deficiency; Complex I deficiency; Complex II deficiency; Complex III deficiency; Complex IV deficiency: Complex V deficiency; cytochrome c oxidase (COX) deficiency, LHON Leber Hereditary Optic Neuropathy; MM Mitochondrial Myopathy: L1MM Lethal Infantile Mitochondrial Myopathy; MMC Maternal Myopathy and Cardiomyopathy; NARP Neurogenic muscle weakness, Ataxia, and Retinitis Pigmentosa; Leigh Disease: FICP — Fatal Infantile Cardiomyopathy Plus, a MELAS-associated cardiomyopathy: MELAS Mitochondrial Encephalomyopathy with Lactic Acidosis and Strokelike episodes; LDYT Leber's hereditary optic neuropathy and Dystonia; MERRF Myoclonic Epilepsy and Ragged Red Muscle Fibers; MHCM Maternally inherited Hypertrophic CardioMyopathy; CPEO Chronic Progressive External Opthalmoplegia; KSS Kearns Sayre Syndrome; DM Diabetes Mellitus; DMDF Diabetes Mellitus+DeaFness; CIPO Chronic Intestinal Pseudoobstruction with myopathy and Opthalmoplegia; DEAF Maternally inherited DEAFness or aminoglycoside-induced DEAFness; PEM Progressive encephalopathy; SNHL SensoriNeural Hearing Loss; Encephalomyopathy; Mitochondrial cytopathy: Dilated Cardiomyopathy: GER Gastrointestinal Reflux: DEMCHO Dementia and Chorea; AMDF Ataxia, Myoclonus; Exercise Intolerance: ESOC Epilepsy, Strokes, Optic atrophy, & Cognitive decline; FBSN Familial Bilateral Striatal Necrosis: FSGS Focal Segmental Glomerulosclerosis: LIMM Lethal Infantile Mitochondrial Myopathy; MDM Myopathy and Diabetes Mellitus: MEPR Myoclonic Epilepsy and Psychomotor Regression; MERME MERRF / MELAS overlap disease; MHCM Maternally Inherited Hypertrophic CardioMyopathy; MICM Maternally Inherited Cardiomyopathy; MILS Maternally Inherited Leigh Syndrome; Mitochondrial Encephalocardiomyopathy; Multisystem Mitochondrial Disorder (myopathy, encephalopathy, blindness, hearing loss, peripheral neuropathy); NAION Nonarteritic Anterior Ischemic Optic Neuropathy; NIDDM Non-Insulin Dependent Diabetes Mellitus; PEM Progressive Encephalopathy; PME Progressive Myoclonus Epilepsy; RTT Rett Syndrome: SIDS Sudden Infant Death Syndrome: MIDD Maternally Inherited Diabetes and Deafness; and MODY Maturity- Onset Diabetes of the Young, and MNGIE), alcoholic cardiomyopathy, systemic carnitine deficiency, malonyl carboxylase deficiency, malonic aciduria, camitine-acylcarnitine translocase deficiency, carnitine palmitoyltransferase II deficiency, deficiencies to mitochondrial beta-oxidation (including, but not limited to mediumchain acyl-coenzyme A (coA) dehydrogenase (MCAD) deficiency, short-chain acyl-coA dehydrogenase (SCAD) deficiency, very-long-chain acyl-coA dehydrogenase (VLCAD) deficiency, and long-chain 3-hydroxyacyl-coA dehydrogenase (LCHAD) deficiency), deficiencies to the mitochondrial electron respiratory chain (including, but not limited to Kearns-Sayre syndrome, MELAS syndrome, MERRF syndrome, Barth syndrome, Leigh’s syndrome, Pearson syndrome, respiratory chain complex I deficiency, and Complex III deficiency), Glycogen storage disease type II (Pompe disease), Glycogen storage disease type III, Niemann-Pick disease, Gaucher disease, Lcell disease, mucopolysaccharidosis type 1 (Hurler syndrome), mucopolysaccharidosis type II (Hunter syndrome), mucopolysaccharidosis type III (Harris- Sanfilippo syndrome), mucopolysaccharidosis type IV (Morquio syndrome), mucopolysaccharidosis type VI (Maroteaux-Lamy syndrome), GM1 gangliosidosis, galactosialidosis, carbohydrate deficient glycoprotein syndromes, Sandhoff’s disease, and other metabolic diseases.

[0293] In some embodiments, the composition is used to treat or prevent a respiratory disease. Exemplary respiratory diseases include, but are not limited to asthma, chronic obstructive pulmonary disease (COPD), pulmonary fibrosis, pneumonia, bronchitis (chronic or acute bronchitis), emphysema, cystic fibrosis / bronchiectasis, pleural effusion, acute chest syndrome, acute respiratory distress syndrome, asbestosis, aspergilosis, severe acute respiratory syndrome (including, but not limited to SARS-CoV-1 and SARS-CoV-2), respiratory syncytial virus (RSV), middle eastern respiratory syndrome (MERS), mesothelioma, pneumothorax, pulmonary arterial hypertension, pulmonary hypertension, pulmonary embolism, sarcoidosis, sleep apnea, and other respiratory diseases.

[0294] In some embodiments, the composition is used to treat or prevent a congenital disease. Exemplary congenital diseases include, but are not limited to albinism, amniotic band syndrome, anencephaly, Angelman syndrome, Barth syndrome, chromosomal abnormalities (including, but not limited to abnormalities to chromosome 9, 10, 16, 18, 20, 21, 22, X chromosome, and Y chromosome), cleft lip / palate, club foot, congenital adrenal hyperplasia, congenital hyperinsulinism, congenital sucrase-isomaltase deficiency (CSID), cystic fibrosis, De Lange syndrome, fetal alcohol syndrome, first arch syndrome, gestational diabetes, Haemophilia, heterochromia, Jacobsen syndrome, Katz syndrome, Klinefelter syndrome, Kabuki syndrome, Kyphosis, Larsen syndrome, Laurence-Moon syndrome, macrocephaly, Marfan syndrome, microcephaly, Nager’s syndrome, neonatal jaundice, neurofibromatosis, Noonan syndrome, Pallister-Killian syndrome, Pierre Robin syndrome, Poland syndrome, Prader-Willi syndrome, Rett syndrome, sickle cell disease, Smith-Lemli-Optiz syndrome, spina bifida, congenital syphilis, teratoma, Treacher Collins syndrome, Turner syndrome, Umbilical hernia, Usher syndrome, Waardenburg syndrome, Werner syndrome, Wolf-Hirschhom syndrome, Wolff-Parkinson- White syndrome, and other congenital diseases or disorders.

[0295] In some embodiments, the composition is used to treat or prevent a gastrointestinal disease. Exemplary gastrointestinal diseases include, but are not limited to heartburn, irritable bowel syndrome, lactose intolerance, gallstones, cholecystitis, cholangitis, anal fissure, hemorrhoids, proctitis, colon polyps, infective colitis, ulcerative colitis, ischemic colitis, Crohn’s disease, radiation colitis, celiac disease, diarrhea (chronic or acute), constipation (chronic or acute), diverticulosis, diverticulitis, acid reflux (gastroesophageal reflux (GER) or gastroesophageal reflux disease (GERD)), Hirschsprung disease, abdominal adhesions, achalasia, acute hepatic porphyria (AHP), anal fistulas, bowel incontinence, centrally mediated abdominal pain syndrome (CAPS), clostridioides difficile infection, cyclic vomiting syndrome (CVS), dyspepsia, eosinophilic gastroenteritis, globus, inflammatory bowel disease, malabsorption, scleroderma, volvulus, and other gastrointestinal diseases.

[0296] Methods of preparing and / or delivering carrier-base-ssNA compositions

[0297] The present disclosure also provides methods preparing, forming, or delivering the composition or therapeutic composition comprising said complex. In one aspect, disclosed herein is a method of forming the complex of any preceding aspect, the method comprising providing a carrier-base, wherein said carrier-base comprises i) an organic polymer comprising one or more monomeric units and ii) at least one nucleobase, wherein at least one covalent bond binds the one or more monomeric unit to the nucleobase(s), providing a ssNA, and incubating the carrier-base and the ssNA, under conditions which allow the carrier-base molecule and the ssNA to form the complex through a hydrogen bond.

[0298] In one aspect, disclosed herein is a method of delivering a complex of any preceding aspect comprising a single-stranded nucleic acid (ssNA) into a cell, the method comprising providing a carrier-base comprising an organic polymer comprising one or more monomeric units and at least one nucleobases, wherein at least one covalent bond binds the one or more monomeric unit to the nucleobase(s); providing the ssNA, incubating the carrier-base and the ssNA, under conditions which allow the carrier-base and the ssNA to form the complex through a hydrogen bond, and introducing the complex into the cell, wherein the complex is internalized inside the cell and the ssNA is thereafter released from the complex.

[0299] In some embodiments, the complex comprises an RNA-plex complex.

[0300] In some embodiments, the method further comprises storing the complex at about 25 °C or less. In some embodiments, the method further comprises storing the complex at about -100, -99, -98, - 97, -96, -95, -94, -93, -92, -91, -90, -89, -88, -87, -86, -85, -84, -83, -82, -81, -80, -79, -78, -77, -

[0301] 76, -75, -74, -73, -72, -71, -70, -69, -68, -67, -66, -65, -64, -63, -62, -61, -60, -59, -58, -57, -56, -

[0302] 55, -54, -53, -52, -51, -50, -49, -48, -47, -46, -45, -44, -43, -42, -41, -40, -39, -38, -37, -36, -35, -

[0303] 34, -33, -32, -31, -30, -29, -28, -27, -26, -25, -24, -23, -22, -21, -20, -19, -18, -17, -16, -15, -14, -

[0304] 13, -12, -11, -10, -9, -8, -7, -6, -5, -4, -3, -2, -1, 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, or 25°C. In some embodiments, the complex is stored at room temperature.

[0305] In some embodiments, the at least one covalent bond is added by a crosslinking reagent. In some embodiments, the crosslinking reagent comprises dicyclohexyl carbodiimide (DCC). In some embodiments, the crosslinking reagent comprises any one of the following including, but not limited to EDC, (l-Ethyl-3-[3-dimethylaminopropyl]carbodiimide hydrochloride), A-Cyclohexyl- A'-(2-morpholinoethyl)carbodiimide metho-p-toluenesulfonate, A,A-Diethylcarbodi imide, Bis(trimethylsilyl)carbodiimide, D1C. A ’ '-Diisopropylcarbodiimide, Carbodiimide (CH2N2), Bis(2,6-diisopropylphenyl)carbodiimide, 1,3-Diphenylcarbodiimide, bis-(4-tert.-buty1-pheny1)- carbod imide, Bis-(4-chloromethyl-phenyl)-carbodiimide, 1.3-Di(p-tolyl)carbodiimide, 1 ,3- Bis(2,2-dimethyl-l ,3-dioxolan-4-ylmethyl)carbodiimide, 1 ,3-Bis(p-bromophenyl)carbodiirft£de, Bis(3-chloro-2-methylphenyl)carbodiimide, and Bis(3,4-dipropylphenyl)carbodiimide.

[0306] In some embodiments, the ssNA, when in complex with the carrier-base, shows reduced degradation in cellular conditions compared to a ssNA which is not protected by a carrier-base. In some embodiments, ssNA degradation is reduced by 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100% compared to a ssNA which is not protected by a carrier-base.

[0307] In some embodiments, the carrier-base and ssNA are incubated in a nuclease-free solution. In some embodiments, the nuclease-free solution comprises DNase-free water, RNase-free water, or combinations thereof.

[0308] In some embodiments, the carrier-base and ssNA are incubated above 60°C for at least one minute; and thereafter the complex is then cooled to 25 °C or lower.

[0309] In some embodiments, the carrier-base and ssNA are incubated between about 60°C to about 105 °C. In some embodiments, the carrier-base and the ssNA are incubated at about 60°, 61°, 62°, 63°, 64°, 65°, 66°, 67°, 68°, 69°, 70°, 71°, 72°, 73°, 74°, 75°, 76°, 77°, 78°, 79°, 80°, 81°, 82°, 83°, 84°, 85°, 86°, 87°, 88°, 89°, 90°, 91°, 92°, 93°, 94°, 95°, 96°, 97°, 98°, 99°, 100°, 101°, 102°, 103°, 104°, or 105°.

[0310] In some embodiments, the carrier-base and ssNA are incubated for one, two, three, four, five, or more minutes. In some embodiments, the carrier-base and ssNA are incubated for about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, or 30 minutes.

[0311] In some embodiments, the complex is cooled to -100, -99, -98, -97, -96, -95, -94, -93, -92, -91, -90, -89, -88, -87, -86, -85, -84, -83, -82, -81, -80, -79, -78, -77, -76, -75, -74, -73, -72, -71, - 70, -69, -68, -67, -66, -65, -64, -63, -62, -61, -60, -59, -58, -57, -56, -55, -54, -53, -52, -51, -50, -

[0312] 49, .48, -47, -46, -45, -44, -43, -42, -41, -40, -39, -38, -37, -36, -35, -34, -33, -32, -31, -30, -29, -

[0313] 28, -27, -26, -25, -24, -23, -22, -21, -20, -19, -18, -17, -16, -15, -14, -13, -12, -11, -10, -9, -8, -7, -

[0314] 6, -5, -4, -3, -2, -1, 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23,

[0315] 24, or 25°C. In some embodiments, the complex is cooled to room temperature.

[0316] In some embodiments, the complex protects the ssNA from degradation by high temperatures above 4°C. In some embodiments, the complex protects the ssNA from degradation by high temperatures above 4°, 5°, 6°, 7°, 8°, 9°, 10°, 11°, 12°, 13°, 14°, 15°, 16°, 17°, 18°, 19°,

[0317] 20°, 21°, 22°, 23°, 24°, 25°, 26°, 27°, 28°, 29°, 30°, 31°, 32°, 33°, 34°, 35°, 36°, 37°, 38°, 39°,

[0318] 40°, 41°, 42°, 43°, 44°, 45°, 46°, 47°, 48°, 49°, 50°, 51°, 52°, 53°, 54°, 55°, 56°, 57°, 58°, 59°,

[0319] 60°, 61°, 62°, 63°, 64°, 65°, 66°, 67°, 68°, 69°, 70°, 71°, 72°, 73°, 74°, 75°, 76°, 77°, 78°, 79°, 80°, 81 °, 82°, 83°, 84°, 85°, 86°, 87°, 88°, 89°, 90°, 91 °, 92°, 93°, 94°, 95°, 96°, 97°, 98°, 99°, 100°C, or more.

[0320] In some embodiments, the complex protects the ssNA from degradation by a nuclease. In some embodiments, the nuclease comprises an endonuclease or an exonuclease. In some embodiments, the nuclease is a DNase or an RNase.

[0321] In some embodiments, the complex is stored in the nuclease-free solution. In some embodiments, the complex is dehydrated by removing the nuclease-free solution prior to storing. In some embodiments, the complex is lyophilized prior to storing. In some embodiments, the complex is hydrated or rehydrated prior to delivery into the cell.

[0322] In some embodiments, a gene therapy method is used to deliver the complex into the cell. In some embodiments, the gene therapy includes, but is not limited to RNA therapy, gene addition, gene editing (including CRISPR gene editing methods), gene silencing (including RNA interference (RNAi) methods), and cell therapy. In some embodiments, the complex is transfected into the cell. In some embodiments, the complex is transformed into the cell. In some embodiments, the method of any preceding aspect stabilizes the ssNA (including, but not limited to messenger RNA (mRNA), transfer RNA (tRNA), ribosomal RNA (rRNA), microRNA (miRNA), long non-coding RNA (IncRNA), RNA aptamers, circular RNA (circRNA), shortinterfering RNA (siRNA), small guide RNA (sgRNA), antisense DNA oligonucleotides, and antisense RNA oligonucleotides) for cellular mechanisms, including but not limited to translation and antigen presentation.

[0323] A number of embodiments of the disclosure have been described. Nevertheless, it will be understood that various modifications may be made without departing from the spirit and scope of the invention. Accordingly, other embodiments are within the scope of the following claims.

[0324] By way of non-limiting illustration, examples of certain embodiments of the present disclosure are given below.

[0325] EXAMPLES

[0326] The following examples are set forth below to illustrate the compositions, devices, methods, and results according to the disclosed subject matter. These examples are not intended to be inclusive of all aspects of the subject matter disclosed herein, but rather to illustrate representative methods and results. These examples are not intended to exclude equivalents and variations of the present invention which are apparent to one skilled in the art. Example 1: RNA-plex technology for single-stranded nucleic acid protection and delivery

[0327] A protection, stabilization, and delivery strategy, namely “RNA-plex”, was conceived to prevent ssNAs from hydrolysis and enzymatic degradation in harsh environment with a group of polymeric macromolecules “Carrier-Base”. The ssNAs include ribonucleic acid (RNA) and deoxyribonucleic acid (DNA) like messenger RNA (mRNA), transfer RNA (tRNA), ribosomal RNA (rRNA), microRNA (miRNA), long non-coding RNAs (IncRNA), single guide RNA (sgRNA) and antisense DNA or RNA oligos. This ssNA protection strategy has been validated by current experimental data.

[0328] Synthesis of carrier-base

[0329] Carrier-bases are composed by poly-amino acid (carrier) and nucleobases (base). A group of novel carrier-base macromolecules were prepared by covalently conjugating bases to polyamino acids. A representative product poly-glutamic acid / aspartic acid-Guanine / Adenine (pDE- GA) of the carrier-bases is presented in FIG. 1.

[0330] The poly-amino acid can be made from the following amino acids: aspartic acid, glutamic acid, tryptophan, histidine, serine, threonine, cysteine, tyrosine, asparagine, glutamine, lysine, arginine. The poly-amino acid can be a single amino acid or a combination of any other ratio (The percentage from 0% to 100%) of the listed amino acids. The molecular weight of the poly-amino acid ranges from 100 to 50,000.

[0331] The base part includes the following six bases (Adenine, Guanine, Uracil, Cytosine, Thymine, Hydroxy purine). The carrier-bases may contain any single base, or two bases, or three, or more.

[0332] To synthesize carrier-base, 100 mg pDE (D:E=6:4, mol / mol) was dissolved in 2 mL dimethyl sulfoxide (DMSO) , and bases such as adenine and guanine, and crosslinking reagent dicyclohexyl carbodiimide (DCC) were also dissolve in DMSO at the concentration of 10 mg / mL and 60 mg / mL, respectively. Mix 100 mg pDE, 10 mg bases and 30 mg DCC in a 20 mL glass bottle, protect from light and stir for 2 days at room temperature. Next, 1 M NaHCO3 was added to the solution to adjust the pH to pH>8.0 (approximately 1 mL). Solution was then stirred for 15 min, and dialyzed against deionized water with 8,000 Da MWCO dialysis tubing for 12 h. The final product was then filtered with a 0.45 pm filter, and dried with a lyophilizer.

[0333] The chemical structure and base content of pDE-GA were confirmed with ' H-nuclear magnetic resonance (NMR) (FIG. 2),13C-NMR (FIG. 3), mass spectrum (MS) (FIG. 4 and FIG. 5) and ultraviolet-visible spectroscopy (UV) (FIG. 6). Construction of RNA-plex mRNA (1 mg / mL) was purchased from Trilink company. Carrier-based was dissolved in RNase free water at the concentration of 1 mg / mL. To form RNA-plex, mRNA and carrier-base was mixed and diluted into RNase free water at different ratios to the final concentration of 25 pg / mL mRNA. The optimal ratio of mRNA and carrier-base may depend on the length of mRNA and needs to be optimized to achieve best protection effect. Next, the mixture was heated to 95°C for 5 min, and then cooled down to room temperature using a PCR thermocycler. mRNA protection from hydrolysis

[0334] With the exceptional performance of mRNA vaccine against CO VID- 19, the development of mRNA-based vaccine and drugs goes into a new era of pharmacy. The performance of RNA- plex technology was tested on mRNA as a representative of ssNAs. The molecular entangling of carrier-base with mRNA forms RNA-plex via hydrogen bonds (FIG. 7). RNA-plex technology shields mRNA structure against hydrolysis and enzymatic digestion. First, OVA mRNA was mixed with carrier-bases at different ratios in RNase free water, and then incubated at 95 °C for 5 minutes to form RNA-plex. Naked mRNA without carrier-base was used as control.RNA-plex was found tolerating hydrolysis at high temperature (95 °C), instead, naked mRNA was completely damaged in 5 minutes (FIG. 8). The mRNA inside the RNA-plex also escaped enzymatic degradation after the incubation with RNase A (FIG. 9).

[0335] Transfection and expression of mRNAs protected by RNA-plex technology

[0336] More important, RNA-plex technology is compatible to transfection reagents. The translation function of the mRNA protected in by RNA-plex technology (RNA-plex) was not affected after including carrier-base. There were attempts to break the RNA-plex by either heat or RNase, and then tested its translation ability on HEK293T cells comparing to naked mRNA. As a pilot test of translation ability of RNA-plex, a traditional reagent Lipo2000 was applied. Lipo2000 is widely used and accepted as a potent DNA / RNA delivery reagent. EGFP reporter mRNA was used in these experiments and EGFP expression was evaluated with fluorescence microscope and Flow cytometry. At room temperature (RT, 25°C), naked mRNA and RNA-plex showed the same performance on EGFP protein translation. However, when heating mRNA or RNA-plex at 95°C for as short as 5 min, the translation ability of naked mRNA was completely destroyed. Excitingly, the translation of RNA-plex protected EGFP mRNA was little influenced (FIG. 10). Next, it was tested whether the structure of carrier-base macromolecule with polymer backbone and covalently conjugated bases is critical for the protection effect, the expression of EGFP mRNA interacted with polymer only, bases only, physical mixture of polymer and bases or carrier-base macromolecule was compared. Result showed that only carrier-base can protect mRNAs from degradation (FIG. 11).

[0337] Finally, the time-dependent EGFP expression of naked mRNA and RNA-plex was evaluated in HEK293 cells. The expression patterns were very similar between naked mRNA and RNA-plex in cells, which indicated the releasing of mRNA from RNA-plex, followed by EGFP protein expression (FIG. 12).

[0338] Long-term storage stability of RNA-plex

[0339] More importantly, RNA-plex ensures superior mRNA stability during storage. Although mRNA drugs like mRNA vaccine showed extraordinary advantages in pharmacy, its natural of low stability, especially in aqueous environment, is one of the major problems to be solved. An eGFP RNA-plex solution was prepared and stored at either RT or 4°C for different duration, then formulated them with lipo2000 and tested EGFP expression on HEK293 cells. Naked EGFP mRNA was manipulated in parallel as control. At RT, naked mRNA started to degrade at the second day. However, RNA-plex kept intact within 1 week, and slightly loss 20% efficacy at the second week (FIG. 13). Long-term stability can be achieved by simply storing RNA-plex at 4°C. EGPF translation was not affected after up to 2 months (FIG. 14).

[0340] Universal protection effects on different mRNAs and compatibility of multiple transfection systems

[0341] Protection effects of RNA-plex technology have been tested on OVA (FIG. 8 and FIG. 9) and EGFP mRNAs (FIGS. 10-14) using lipo2000 reagent. Additional applications were extended on other mRNAs like mCherry mRNAs, and different transfection systems such as polyethylenimine (PEI) and in vivo-jetRNA+. As expected, mCherry mRNA can be protected in a similar pattern as EGFP mRNA (FIG. 15). In addition to lipo2000, we demonstrated that RNA- plex was also compatible to other widely used DNA / RNA delivery systems with different structures and applications (FIG. 16).

[0342] In vivo application of RNA-plex

[0343] Finally, mRNA translation was tested in animal. Naked firefly luciferase (Luc) mRNA or Luc mRNA-plex were formulated using in vivo-jetRNA+. Result again proved the broad-spectrum application of RNA-plex technique (FIG. 17). After administration to mice via footpad injection, which is common route for vaccine treatment, a clear trend of decreased Luc expression was detected if storing naked mRNA for 1 day at RT. In the contrary, no difference was observed if protecting mRNA with RNA-plex technique (FIG. 18). Example 2: Optimization of nucleobases for mRNA stabilization activity of carrier-base.

[0344] To test which nucleobase or nucleobase combination is optimal for mRNA stabilization, guanine (G), adenine (A) or their 1: 1 mixture (A / G) were conjugated to poly-amino acid carrier, and then mixed with enhanced green fluorescent protein (EGFP) mRNA to form RNA-plexes. Next, different RNA-plexes were heated at 95°C for 5 min to accelerate hydrolysis, followed by encapsulation using Lipofectamine 2000 (Lipo2000) and transfection to HEK293T (293T) cells. Carrier- A / G performed the best on EGFP mRNA protection and translation, which was used in the following studies (FIG. 19).

[0345] Example 3: Stabilization of mRNA during RT-storage by RNA-plex technology

[0346] RNA-plex technology aims to protect mRNA from degradation during storage in ambient environments. To prove this concept, naked EGFP mRNA and EGFP-RNA-plex were stored in RNase free water at RT for up to two weeks. mRNA integrity was then analyzed using agarose gel electrophoresis. mRNAs kept intact for up to 2 weeks when protected by RNA-plex technology, while naked mRNA was completely degraded after as short as 3 days (FIG. 20A, 20B, and 20C).

[0347] Example 4: Universal stabilization of different RNAs by RNA-plex technology

[0348] Universal protection of different linear mRNA sequences. mRNA protection by carrierbase was tested on EGFP, ovalbumin (OVA) and luciferase mRNAs. To accelerate hydrolysis, naked mRNA and RNA-plexes were heated at 95°C for 5 min, and samples stored at room temperature (RT, 25°C) were set as control. Naked mRNA (1:0) was completely degraded after heating, while RNA-plexes of all the three mRNAs at >1:3 (w / w) ratios were still intact after heating. Moreover, RNA-plex did not cause delayed mRNA migration, indicating the loose and dissociable interaction between mRNA and carrier-base (FIG. 21).

[0349] Example 5: Protection of circular mRNA (circRNA)

[0350] To expand the applicability of RNA-plex technology to other unstable RNAs beyond linear mRNA, we investigated its compatibility with circRNA, which is an existed stabilization technology with much higher stability within cells after administration. We performed the same evaluation on GFP circRNA protection. As expected, RNA-plex effectively protected GFP circRNA as linear mRNAs (FIG. 22), demonstrating a broad- spectrum application. Example 6: Compatibility of RNA-plex technology to lipid nanoparticles (LNP)

[0351] Herein, the compatibility of RNA-plex to multiple transfection systems including Lipo2000, in vzvo-jetRNA plus and polyethylenimine (PEI) is shown. LNP is the most popular mRNA delivery system currently due to its low-toxicity. To test whether RNA-plex is compatible to LNPs, naked EGFP-mRNA or EGFP-RNA-plex was loaded in LNPs composed of ionizable lipid SM-102, l,2-distearoyl-sn-glycero-3-phosphocholine (DSPC), cholesterol and 1,2- dimyristoyl-rac-glycero-3-methoxypolyethylene glycol-2000 (DMG-PEG2000) at the ratio of 50: 10:38.5: 1.5 (mol / mol) using NanoAssemblr, followed by transfection to HEK293T (293T) cells. Fresh-prepared EGFP-RNA-plex-LNP (EGFP-plex / LNP) showed the same encapsulation efficiency (>90%) as naked EGFP mRNA-LNP (EGFP / LNP), and higher translation efficiency after 24 h (FIG. 23).

[0352] Example 7: Increased intracellular mRNA translation using RNA-plex technology in vitro.

[0353] A. EGFP translation in DC2.4 cells.

[0354] To test intracellular translation and stabilization of mRNAs in RNA-plex, and its application potential on mRNA vaccine application, naked EGFP mRNA and EGFP-mRNA-plex were encapsulated using Lipo2000, and then transfected to a dendritic cell line DC2.4 at 1 ug / mL in vitro. Transfection efficiency was measured with flow cytometry after up to 8 days. EGFP positive DC2.4 cells were doubled when RNA-plex technology was applied. Moreover, cells with EGFP expression in RNA-plex group were still detectable after 8 days, while those transfected with naked mRNA completely lose EGFP expression (FIG. 24).

[0355] B. OVA mRNA protection, delivery, and antigen presentation in DC2.4.

[0356] Next, antigen presentation by DC2.4 was tested using the model antigen OVA. Naked mRNA or RNA-plex was packed with Lipo2000 at different ratios. The presentation of OVA peptide (SIINFEKL) by H-2Kb were detected by #25-D1.16 antibody using flow cytometry. Comparing antigen efficiencies of naked mRNA and RNA-plex at their optimal ratios (naked: mRNA: Lipo2000=4, RNA-plex: mRNA: Lipo2000=10), OVA antigen presenting cells were doubled when applying RNA-plex technology (FIG. 25A and 25B).

[0357] Example 8: mRNA protection by silica-base.

[0358] The extraordinary mRNA protection ability of polymer-based carrier-base in RNA-plex was demonstrated herein. Next, it was explored whether other vectors could be applied for mRNA protection in addition to polymers. Nucleobases were conjugated to silica nanoparticle to form silica-base (FIG. 26A), and evaluated GFP mRNA integrity and silica-base protection efficacy using agarose gel electrophoresis. Silica-base and GFP mRNA were mixed at different ratios, and heated at 95 °C for 5 min to accelerate hydrolysis and compared to samples prepared at RT / 25°C. Naked GFP mRNA (1:0) was completely degraded after heating, while mRNA: silica-base at >1:3 (w / w) ratios were well-protected (FIG. 26B).

[0359] Example 9: In vivo application of RNA-plex technology on therapeutic anti-cancer mRNA vaccines.

[0360] The high immunogenicity of mRNA formulations causes severe side effects after administration, and shortens mRNA expression due to the stimulation of Type-I interferon and RNase L. To evaluate the immunogenicity and potential toxicity of RNA-plex, RNA-plex and the individual components were condensed using a commercially available delivery reagent Invivo jetRNA (jetRNA), and incubated with bone marrow derived dendritic cells (BMDCs). Inflammatory cytokine TNFa and type I interferon IFN| secretion were reduced comparing jetRNA-RNA-plex to the conventional jetRNA-naked mRNA complex (FIG. 27), showing reduced immunogenicity and enhanced safety profile.

[0361] Example 10: Therapeutic efficacy in B16 melanoma murine model.

[0362] In vivo therapeutic efficacy of RNA-plex was assessed using a B16-ov albumin (OVA) melanoma murine model in vivo (FIG. 28 A). JetRNA-RNA-plex vaccination significantly inhibited tumor growth compared to controls, showing the improved therapeutic potential of RNA- plex technology (FIG. 28B). Importantly, no body weight loss was found in all groups throughout the experiment, suggesting good vaccine tolerability (FIG. 28C). In addition, the JetRNA-RNA- plex significantly increased the IFNy+and OVA antigen specific CD8 T cells in the spleen after vaccination.

[0363] It will be apparent to those skilled in the art that various modifications and variations can be made in the present disclosure without departing from the scope or spirit of the invention. Other embodiments of the disclosure will be apparent to those skilled in the art from consideration of the specification and practice of the methods disclosed herein. It is intended that the specification and examples be considered as exemplary only, with a true scope and spirit of the invention being indicated by the following claims. TABLES

[0364] Table 1.

[0365] Table 2.

[0366] Table

[0367]

Claims

CLAIMSWhat is claimed is:

1. A complex comprising: a. a carrier-base, wherein said carrier-base comprises: i) an organic polymer comprising one or more monomeric units; and ii) at least one nucleobase; wherein at least one covalent bond binds the one or more monomeric units to the nucleobase(s), and b. a single stranded nucleic acid (ssNA), wherein at least one hydrogen bond binds the ssNA to the carrier-base to form the complex.

2. The complex of claim 1, wherein the complex comprises an RNA-plex complex.

3. The complex of claim 1 or 2, wherein the organic polymer comprises 2 or more monomeric units.

4. The complex of any one of claims 1-3, wherein the monomeric units are selected from an amino acid, a monosaccharide, a nucleotide, and a fatty acid.

5. The complex of any one of claims 1-4, wherein the organic polymer comprises a combination of at least two of an amino acid, a monosaccharide, a nucleotide, or a fatty acid.

6. The complex of any one of claims 1-5, wherein the organic polymer comprising 2 or more monomeric units comprises a poly-amino acid, a polysaccharide, a polynucleotide, a lipid, or combinations thereof.

7. The complex of claim 6, wherein the poly-amino acid comprises 2 or more amino acids, or derivatives thereof.

8. The complex of claim 7, wherein the 2 or more amino acids comprise a L-, D-, or DL- amino acid.

9. The complex of any one of claims 6-8, wherein the poly-amino acid ranges between about 1 kDa to about 50 kDa.

10. The complex of any one of claims 1-9, wherein the polymer comprises a silica-base nanoparticle.

11. The complex of any one of claims 1-10, wherein the complex comprises one, two, three, four, five, six, or more nucleobases.

12. The complex of any one of claims 1-11, wherein the complex comprises six nucleobases selected from thymine, uracil, cytosine, adenine, guanine, xanthine, and hypoxanthine.

13. The complex of claim 12, wherein the complex comprises adenine, guanine, or a combination thereof.

14. The complex of claim 13, wherein the combination of adenine and guanine comprises a 1:1 mixture.

15. The complex of any one of claims 1-14, wherein the complex comprises a ratio of about 1: 1 to about 1:100 of monomeric units relative to nucleobases.

16. The complex of any one of claims 1-15, wherein the ssNA comprises single-stranded DNA, single-stranded RNA, or a combination thereof.

17. The complex of any one of claims 1-16, wherein the ssNA comprises messenger RNA (mRNA), transfer RNA (tRNA), ribosomal RNA (rRNA), microRNA (miRNA), long non-coding RNA (IncRNA), RNA aptamers, circular RNA (circRNA), short-interfering RNA (siRNA), small guide RNA (sgRNA), antisense DNA oligonucleotides, or antisense RNA oligonucleotides.

18. The complex of any one of claims 1-17, wherein the at least one covalent bonds comprises an ester bond, an ether bond, or an amide bond.

19. A therapeutic composition comprising the complex of any one of claims 1 -18, wherein the ssNA has a therapeutic benefit.

20. The therapeutic composition of claim 19, further comprising a pharmaceutically acceptable carrier selected from an excipient, a diluent, a salt, a buffer, a stabilizer, a lipid, an emulsion, and a nanoparticle.

21. The therapeutic composition of claim 19 or 20, further comprising a transfection system selected from a lipofectamine composition, a lipid nanoparticle (LNP), in vivo jetRNA plus, or polyethylenimine (PEI).

22. A method of treating or preventing a disease or disorder in a subject, the method comprising administering an effective amount of the therapeutic composition of any one of claims 19-21, wherein the ssNA is internalized and released into a cell of the subject to treat or prevent the disease or disorder.

23. A method of forming a complex comprising a single stranded nucleic acid (ssNA) and a carrier-base, the method comprising: a) providing the carrier-base, wherein said carrier-base comprises: i) an organic polymer comprising one or more monomeric units; and ii) at least one nucleobase; wherein at least one covalent bond binds the one or more monomeric unit to the nucleobase(s); b) providing the ssNA; and c) incubating the carrier-base and the ssNA, under conditions which allow the carrierbase molecule and the ssNA to form the complex through a hydrogen bond.

24. The method of claim 23, further comprising the step of: d) storing the complex at about 25°C or less.

25. The method of claim 23 or 24, wherein the complex comprises an RNA-plex complex.

26. The method of any one of claims 23-25, wherein the at least one covalent bond is added by a crosslinking reagent.

27. The method of claim 26, wherein the crosslinking reagent comprises dicyclohexyl carbodiimide (DCC).

28. The method of any one of claims 23-27, wherein the ssNA, when in complex with the carrier-base, shows reduced degradation in cellular conditions compared to a ssNA which is not protected by a carrier-base.

29. The method of any one of claims 23-28, wherein the carrier-base and ssNA are incubated in a nuclease-free solution.

30. The method of claims 29, wherein the nuclease-free solution comprises DNase-free water, RNase-free water, or combinations thereof.

31. The method of any one of claims 23-30, wherein the carrier-base and ssNA are incubated above 60°C for at least one minute; and thereafter the complex is then cooled to 25°C or lower.

32. The method of any one of claims 23-31, wherein the carrier-base and ssNA are incubated between about 60°C to about 105 °C.

33. The method of any one of claims 23-32, wherein the carrier-base and ssNA are incubated for one, two, three, four, five, or more minutes.

34. The method of any one of claims 23-33, wherein the complex protects the ssNA from degradation by high temperatures above 4°C.

35. The method of any one of claims 23-34, wherein the complex protects the ssNA from degradation by a nuclease.

36. The method of claim 35, wherein the nuclease comprises an endonuclease or an exonuclease.

37. The method of claim 35 or 36, wherein the nuclease is a DNase or an RNase.

38. The method of any one of claims 23-37, wherein the complex is stored at 25°C, 4°C, - 20°C, or -80°C.

39. The method of any one of claims 23-38, wherein the complex is stored in the nuclease-free solution or the complex is dehydrated by removing the nuclease-free solution prior to storing.

40. A method of delivering a complex comprising a single stranded nucleic acid (ssNA) into a cell, the method comprising: a. providing a carrier-base comprising: i. an organic polymer comprising one or more monomeric units; and ii. at least one nucleobases; wherein at least one covalent bond binds the one or more monomeric unit to the nucleobase(s); b. providing the ssNA; c. incubating the carrier-base and the ssNA, under conditions which allow the carrierbase and the ssNA to form the complex through a hydrogen bond; and d. introducing the complex into the cell, wherein the complex is internalized inside the cell and the ssNA is thereafter released from the complex.

41. The method of claim 40, wherein the complex comprises an RNA-plex complex.

42. The method of claim 40 or 41, wherein the at least one covalent bond is added by a crosslinking reagent.

43. The complex of claim 42, wherein the crosslinking reagent comprises dicyclohexyl carbodiimide (DCC).

44. The method of any one of claims 40-43, wherein the complex protects the ssNA from degradation by high temperatures above 25 °C.

45. The method of any one of claims 40-44, wherein the complex protects the ssNA from degradation by a nuclease.

46. The method of claim 45, wherein the nuclease comprises an endonuclease or an exonuclease.

47. The method of claim 45 or 46, wherein the nuclease is a DNase or an RNase.

48. The method of any one of claims 45-47, wherein a gene therapy method is used to deliver the complex into the cell.

49. The method of any one of claims 40-48, wherein the complex is transfected into the cell.

50. The method of any one of claims 40-49, wherein the method stabilizes the ssNA for one or more cellular mechanisms.

51. The method of claim 51, wherein the one or more cellular mechanisms comprises translation or antigen presentation.

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