CROSS-REFERENCE TO RELATED APPLICATION

This application claims priority from Korean Patent Application No. 10-2014-0093458, filed on Jul. 23, 2014, contents of which are incorporated by reference herein in its entirety.

BACKGROUND OF INVENTION

1. Field of Invention

The present invention relates to a molecule delivery technology and a carrier technology, which can selectively deliver a material to a desired specific cell and living tissue.

The field for which the technology of the present invention may be utilized is largely the field of a drug carrier which effectively delivers an imaging probe and a therapeutic agent to an affected part.

2. Background of Invention

Examples of the existing drug delivery technology include a liposome carrier, a polymer-based carrier, a peptide carrier, and the like. However, the liposome carrier has low vesicle stability, relatively low drug-encapsulation rate and limited size reproducibility, and the polymer-based carrier is disadvantageous in that it has low biodegradability and limited size reproducibility, its in vivo drug metabolism is unclear, as well as it shows immunogenicity. The peptide carrier is also disadvantageous in having low thermal stability, and limited modification possibility and immunogenicity.

On the other hand, unlike the existing carriers, a carrier composed of nucleic acids is advantageous in that it is completely biodegradable into non-toxic small molecules, the size and shape thereof can be easily adjusted to a suitable level, it can be self-assembled and has monodispersity and non-immunogenicity, as well as it can be easily conjugated with functional molecules. Accordingly, DNA nanocarriers are an emerging class in drug delivery technologies.

Previously, it has been shown that biocompatible DNA could self-assemble to construct various three dimensional (3D) DNA nanostructures including tetrahedron, pentahedron, cube, double pyramid, octahedron, dodecahedron, and fullerene-like structures. Among them, the DNA tetrahedron has been considered one of the most practical DNA nanostructures since it can be assembled simply from four DNA strands and prepared in high yield. A principle in which the DNA tetrahedron is assembled from 4 DNA strands is by mutual hybridization, and a schematic view explaining the principle is illustrated inFIG. 1.

A recent report in which DNA nanostructures are taken up into mammalian cells has opened an opportunity that the nanostructures may serve an important role in being applied to biomedicine. They are often introduced into mammalian cells even without transfection agents. Further, it is known that DNA nanostructures are significantly resistant to nucleases.

Due to these characteristics, recently, DNA nanostructures have been utilized in intracellular delivery of bioactive molecules such as anticancer agents, aptamers, antisenses, immunogenic molecules, and siRNA.

Throughout the present specification, a plurality of documents are referenced, and citations thereof are indicated. The disclosure of each of the cited documents is incorporated herein by reference in its entirety to describe the level of the technical field to which the present invention pertains and the content of the present invention more apparently.

SUMMARY OF INVENTION

The present inventors have studied and tried to develop a drug carrier for systemic administration, which may specifically deliver a pharmaceutically active ingredient only to a specific tissue by in vivo administration. As a result, the present inventors have found that a self-assembled 3-D nucleic acid nanostructure not only has specificity for specific cells and tissues, but also can be specifically delivered only to the corresponding tissue by in vivo administration, particularly without targeting ligands, thereby completing the present invention.

Accordingly, an object of the present invention is to provide a self-assembled 3-D nucleic acid nanoparticle having selectivity for specific tissues to cells.

Another object of the present invention is to provide a drug carrier for systemic administration having the nucleic acid nanostructure, which is specific for a specific tissue.

The other objects and advantages of the present invention will be more apparent from the following detailed description, claims and drawings of the invention.

An aspect of the present invention is to provide a method of in vivo delivering a pharmaceutically active ingredient to a specific tissue, using a self-assembled 3-D nucleic acid nanostructure as a drug carrier.

In the method of present invention, the 3-D nucleic acid nanostructure drug earner can deliver a pharmaceutically active ingredient specifically only to a target tissue by in vivo administration, and has cell or tissue specificity particularly without targeting ligands.

In order to specifically deliver nucleic acid to a specific cell such as a tumor cell in the related art, there has been an attempt to deliver nucleic acid using a ligand such as antibody or folate. However, it was impossible to achieve in vivo targeting without an additional process of binding the ligand, and also there was a problem that the ligand could cause an immune reaction and the like in an organism.

Under these circumstances, the present inventors provide an in vivo drug delivery technology in which only a target tissue is specifically targeted by administration of nucleic acid nanostructures only which are completely biodegraded into non-toxic small molecules in the organism.

In an exemplary embodiment, a drug carrier of the present invention has an L-DNA nanostructure, and has in vivo cancer tissue specificity.

The L-DNA nanoparticle may have various polyhedron structures such as, for example, a tetrahedron, a pentahedron, a cube, an octahedron, a dodecahedron, a cuboctahedron, and a dodecicosahedron, a double pyramid or fullerene structure, and the like, but is not necessarily limited thereto, and may preferably have a tetrahedron structure composed of 4 strands of L-DNA 55 mer or 4 strands of L-DNA 92 mer, but is not necessarily limited thereto.

The L-DNA polyhedron structure may be formed when a plurality of single stranded nucleic acid molecules, for example, 4 to 100, 4 to 50, or 4 to 20 stranded nucleic acid molecules are self-assembled by a hybridization principle.

In another exemplary embodiment, the drug carrier of the present invention has a D-DNA nanostructure, and has specificity for liver, skin, or kidney tissue during in vivo administration.

The carrier preferably may have in vivo liver tissue specificity as a tetrahedron nanostructure composed of 4 strands of D-DNA 55 mer, or have in vivo kidney tissue specificity as a tetrahedron nanostructure composed of 4 strands of D-DNA 92 mer, but is not necessarily limited thereto.

Meanwhile, a pharmaceutically active ingredient to be delivered specifically to an organism tissue or cell by the drug carrier of the present invention may be delivered while being encapsulated in a nucleic acid nanoparticle having a polyhedron structure of the present invention, or may also be delivered while being bonded to the backbone of nucleic acid.

The pharmaceutically active ingredient may be encapsulated in a polyhedron structure, or be allowed to be bonded to the backbone of nucleic acid by an action in which linear nucleic acid strands are assembled into a 3-D polyhedron nanostructure, for example, in a state where the linear nucleic strands are mixed with the active ingredient.

The pharmaceutically active ingredient, which may be delivered into cells using the drug carrier of the present invention, is not limited to a specific kind of ingredient, and examples thereof include an anticancer agent, a contrast medium, a hormone drug, an anti-hormone drug, a vitamin preparation, a calcium agent, an inorganic preparation, a sugar preparation, an organic acid preparation, a protein amino acid preparation, an antidote, an enzyme preparation, a metabolic preparation, a diabetes combination drug, a drug for tissue repair, a chlorophyll preparation, a pigment preparation, a tumor drug, a tumor therapeutic agent, a radioactive medicine, a tissue cell diagnostic agent, a tissue cell therapeutic agent, an antibiotic preparation, an antiviral drug, a complex antibiotic preparation, a chemotherapeutic agent, a vaccine, a toxin, a toxoid, an antitoxin, leptospira serum, a blood preparation, a biological preparation, an analgesic, an immunogenic molecule, an antihistamine, an allergy drug, a non-specific immunogenic preparation, an anesthetic, an antihypnotic, a neuropsychiatric solvent, nucleic acid, an aptamer, an antisense nucleic acid, an oligonucleotide, peptide, siRNA, micro RNA, and the like, but are not necessarily limited thereto.

Another aspect of the present invention relates to a method of in vivo delivering an anticancer agent specifically to cancer cells, comprising administering a pharmaceutical composition including the aforementioned 3-D nucleic acid nanostructure drug carrier and an anticancer agent.

According to an exemplary embodiment of the present invention, the anticancer agent may be encapsulated in the nucleic acid drug carrier of the present invention or be bonded to the backbone of nucleic acid, and examples of the anticancer which may be used include doxorubicin, DNA aptamer, RNA aptamer, acivicin, aclarubicin, acodazole, acronycine, adozelesin, alanosine, aldesleukin, allopurinol sodium, altretamine, aminoglutethimide, amonafide, ampl igen, amsacrine, androgens, anguidine, aphidicolin glycinate, asaley, asparaginase, 5-azacitidine, azathioprine,Bacillus calmette-guerin(BCG), Baker's Antifol, beta-2-deoxythioguanosine, bisantrene HCl, bleomycin sulfate, busulfan, buthionine sulfoximine, BWA 773U82, BW 502U83/HCl, BW 7U85 mesylate, ceracemide, carbetimer, carboplatin, carmustine, chlorambucil, chloroquinoxaline-sulfonamide, chlorozotocin, chromomycin A3, cisplatin, cladribine, corticosteroids,Corynebacterium parvum, CPT-11, crisnatol, cyclocytidine, cyclophosphamide, cytarabine, cytembena, dabismaleate, dacarbazine, dactinomycin, daunorubicin HCl, deazauridine, dexrazoxane, dianhydrogalactitol, diaziquone, dibromodulcitol, didemnin B, diethyldithiocarbamate, diglycoaldehyde, dihydro-5-azacytidine, echinomycin, dedatrexate, edelfosine, eplolnitin, Elliott's solution, elsamitrucin, epirubicin, esorubicin, estramustine phosphate, estrogens, etanidazole, ethiofos, etoposide, fadrazole, fazarabine, fenretinide, filgrastim, finasteride, flavone acetic acid, floxuridine, fludarabine phosphate, 5-fluorouracil, Fluosol™, flutamide, gallium nitrate, gemcitabine, goserelin acetate, hepsulfam, hexamethylene bisacetamide, homoharringtonine, hydrazine sulfate, 4-hydroxyandrostenedione, hydrozyurea, idarubicin HCl, ifosfamide, interferon alpha, interferon beta, interferon gamma, interleukin-I alpha and beta, interleukin-3, interleukin-4, interleukin-6, 4-ipomeanol, iproplatin, isotretinoin, leucovorin calcium, leuprolide acetate, levamisole, liposomal daunorubicin, liposome encapsulated doxorubicin, lomustine, lonidamine, maytansine, mechlorethamine hydrochloride, melphalan, menogaril, merbarone, 6-mercaptopurine, mesna, methanol extract ofBacillus calmette-guerin, methotrexate, N-methylformamide, mifepristone, mitoguazone, mitomycin-C, mitotane, mitoxantrone hydrochloride, monocyte/macrophage colony-stimulating factor, nabilone, nafoxidine, neocarzinostatin, octreotide acetate, ormaplatin, oxaliplatin, paclitaxel, pala, pentostatin, piperazinedione, pipobroman, pirarubicin, piritrexim, piroxantrone hydrochloride, PIXY-321, plicamycin, porfimer sodium, prednimustine, procarbazine, progestins, pyrazofurin, razoxane, sargramostim, semustine, spirogermanium, spiromustine, streptonigrin, streptozocin, sulofenur, suramin sodium, tamoxifen, taxotere, tegafur, teniposide, terephthalamidine, teroxirone, thioguanine, thiotepa, thymidine injection, tiazofurin, topotecan, toremifene, tretinoin, trifluoperazine hydrochloride, trifluridine, trimetrexate, tumor necrosis factor (TNF), uracil mustard, vinblastine sulfate, vincristine sulfate, vindesine, vinorelbine, vinzolidine, Yoshi 864, zorubicin, mixtures thereof, and the like, but are not necessarily limited thereto.

The pharmaceutical composition may further include a pharmaceutically acceptable carrier, and the pharmaceutically acceptable earner is typically used in formulations and includes lactose, dextrose, sucrose, sorbitol, mannitol, starch, gum acacia, calcium phosphate, alginate, gelatin, calcium silicate, microcrystalline cellulose, polyvinylpyrrolidone, cellulose, water, syrup, methyl cellulose, methyl hydroxybenzoate, propyl hydroxybenzoate, talc, magnesium stearate, mineral oil, and the like, but is not limited thereto. Suitable pharmaceutically acceptable carriers and formulations are described in detail inRemington's Pharmaceutical Sciences(19th ed., 1995).

In the method of present invention, the pharmaceutical composition 1s systemically administered, and may be administered parenterally or orally. In case of parenteral administration, it may be administered through intravenous injection, intramuscular injection, and the like. An adequate administration amount of the pharmaceutical composition of the present invention may vary depending on factors, such as formulation method, administration method, age, weight, sex or disease condition of the patient, diet, administration time, administration route, elimination rate and response sensitivity.

Still another aspect of the present invention provides a method of in vivo delivering a contrast medium to a specific tissue for imaging, comprising administering a pharmaceutical composition including the aforementioned 3-D nucleic acid nanostructure drug carrier and a contrast medium.

In an exemplary embodiment, an inorganic or organic dye, a fluorescent substance, an isotope, a magnetic substance, a paramagnetic or superparamagnetic nanoparticle may be used as the contrast medium.

Herein, the inorganic or organic dyes are used for enhancing image contrast in fluorescent or optical image, radiation imaging such as computed tomography (CT) photography, and non-radiation image such as sonogram or MRI including the existing dyes, and may include, for example, diatrizoate, metrizoate, ioxaglate, iopamidol, iohexyl, ioxilan, iopromide, iodixanol, barium-based dye such as barium sulfate, Gastrografin®, and the like, but are not necessarily limited thereto.

The fluorescent probe may include, for example, indocyanine, NIR (near infrared) dye, fluorescein, phycoerythrin, rhodamine, lissamine, Cy3, Cy5 (Pharmacia), chromophore, chemical luminophore, mass labeling, electron dense particle, enzymes (alkaline phosphatase or horseradish peroxidase), but is not necessarily limited thereto.

In the contrast medium composition of the present invention, the composition may be a magnetic substance, and a paramagnetic or superparamagnetic nanoparticle, and in this case, may be usefully utilized particularly in imaging using MRI.

The paramagnetic nanoparticle is a material which may induce spin-lattice relaxation, and it is possible to use, for example, Magnevist (Schering, Germany), Gd chelating compounds such as Gd-DTPA (Gd-diethylene triamine pentaacetic acid), and a material such as Gd2O3(C. Riviere et al. J. Am. Chem. Soc. 2007, 129, 5076.) and MnO (T. Hyeon et al. Angew. Chem. Int. Ed. 2007, 46, 5397.).

Further, the superparamagnetic nanoparticle is a material which is magnetized by an externally given magnetic field, affects the spin-spin process of a hydrogen nuclear spin in surrounding water molecules by generating an induced magnetic field, and shows a dark or negative contrast effect typically compared with water by amplifying the magnetic resonance imaging signal, and for example, Feridex, Resovist, Combidex, MEIO (magnetism engineered iron oxide) and the like including oxidized iron components.

As described above, depending on what contrast medium is encapsulated in the nucleic acid nanoparticle drug carrier having a polyhedron structure of the present invention or is bonded to the backbone of nucleic acid, the contrast medium composition of the present invention may be used appropriately for fluorescence imaging, optical imaging, radiation imaging, computed tomography (CT) photography or MRI.

In a preferred exemplary embodiment of the present invention, the contrast medium composition may be for imaging of liver, kidney, or cancer tissues.

The 3-D nucleic nanoparticle drug carrier of the present invention an in vivo drug delivery technology which targets only a specific tissue even without including a target ligand during the administration in an organism.

The drug carrier of the present invention as described above may be completely biodegraded into non-toxic small molecules, and the size and shape thereof may be easily adjusted to a suitable level, and the carrier is advantageous in being capable of being self-assembled while having monodispersity and non-immunogenicity, and being easily conjugated with functional molecules, and simultaneously has specific cell or biotissue selectivity. Accordingly, the drug carrier of the present invention may be usefully utilized as a drug carrier.

