KR20240086776A - Metal-organic framework (MOFs) nanoparticles coated with a fusion protein of Glutathione S-transferase and peptides targeting diseased cells and use thereof - Google Patents

Abstract

Metal-organic framework (MOFs) nanoparticles surface-coated with a fusion protein of glutathione transferase and disease cell targeting peptide, and their use for target cell-specific drug delivery and cancer prevention or treatment Related to this, according to one aspect, the metal-organic framework nanoparticle and the composition containing the same have excellent protein adsorption blocking effect, cell targeting effect, target cell specific drug delivery effect, and photoreactive ROS generation within the target cell. It can be usefully used as a target cell-specific drug carrier or disease treatment agent (e.g., cancer treatment agent) by exhibiting a cell death effect. In particular, it can induce a synergistic effect of photodynamic therapy and chemotherapy on cancer cell death.

Description

Metal-organic framework (MOFs) nanoparticles coated with a fusion protein of glutathione transferase and disease cell targeting peptide and their uses {Metal-organic framework (MOFs) nanoparticles coated with a fusion protein of Glutathione S-transferase and peptides targeting diseased cells and use thereof}

Metal-organic framework (MOFs) nanoparticles surface-coated with a fusion protein of glutathione transferase and disease cell targeting peptide, and compositions containing the same for target cell-specific drug delivery and pharmaceutical compositions for preventing or treating cancer It's about.

Metal-organic framework (MOFs) is a crystalline organic-inorganic hybrid polymer formed through coordination of metal ions or metal ion clusters with organic ligand molecules. MOFs, also called porous coordination polymers (PCPs), have a regular pore structure and high specific surface area, and can be synthesized into various structures through selection of ligand molecules and metal ions. In general, the size of the pores can be controlled by adjusting the size of the organic ligand molecule, and the surface properties of the pores can be controlled by selecting an appropriate metal ion or introducing a functional group into the organic ligand molecule. In particular, unsaturated metal sites formed inside the structure during dehydration or desolvation serve as adsorption sites for gas molecules or guest molecules. Based on these characteristics, active research is being conducted to develop fuel gas adsorption and storage materials, catalysts, sensors, synthetic media, drug delivery media, and proton conductors. However, despite much progress, its introduction out of the research field and into industry is slowing down due to its still low thermal, chemical, and mechanical stability. To solve this problem, zeolitic imidazolate frameworks (ZIFs) or covalent organic frameworks (COFs) are being developed, but these are also often obtained in the form of powders with small particle sizes, raising the issue of deactivation due to agglomeration. It is becoming. In addition, porphyrin-based MOFs have been developed recently, but this also has the problem that unnecessary proteins are adsorbed to MOFs particles in a biological environment, forming a protein corona, which interferes with the drug delivery function of MOFs particles. there is.

Therefore, research is underway to create composites by fusing MOFs with various materials as a new strategy to improve the properties of MOFs and give them new functions.

Accordingly, the present inventors developed a complex in which the surface of MOFs was coated with a fusion protein of glutathione S-transferase (GST) and a disease cell-targeting peptide, and the complex had an excellent protein adsorption blocking effect and cell targeting effect. The present invention was completed by confirming the effect, target cell-specific drug delivery effect, and target cell-specific photoreactive cell death effect.

One aspect is glutathione S-transferase (GST); and providing metal-organic framework (MOFs) nanoparticles coated on the surface with a fusion protein linked to a disease cell targeting peptide.

Another aspect is to provide a composition for target cell-specific drug delivery, comprising the metal-organic framework nanoparticles as an active ingredient.

Another aspect is to provide a composition for target cell-specific photoreactive cell death comprising the metal-organic framework nanoparticles as an active ingredient.

Another aspect is to provide a pharmaceutical composition for preventing or treating cancer, comprising the metal-organic framework nanoparticles as an active ingredient.

Another aspect is to provide a method of preventing or treating cancer comprising administering the metal-organic framework nanoparticle or the composition to a subject.

Another aspect is to provide the metal-organic framework nanoparticles for use in target cell-specific drug delivery, target cell-specific photoreactive cell killing, or cancer prevention or treatment.

Another aspect is to provide a use of the metal-organic framework nanoparticles for preparing a composition for target cell-specific drug delivery, a composition for target cell-specific photoreactive cell killing, or a composition for preventing or treating cancer. .

Other objects and advantages of the present application will become clearer from the following detailed description together with the appended claims and drawings. Contents not described in this specification can be fully recognized and inferred by a person skilled in the technical field of this application or a similar technical field, so description thereof is omitted.

Each description and embodiment disclosed in this application may also be applied to each other description and embodiment. That is, all combinations of the various elements disclosed in this application fall within the scope of this application. Additionally, the scope of the present application cannot be considered limited by the specific description described below.

One aspect is glutathione S-transferase (GST); and metal-organic framework (MOFs) nanoparticles (GST-Afb-MOFs nanoparticles) coated on the surface with a fusion protein (GST-Afb) linked to a disease cell targeting peptide (Afb).

The term "Glutathione S-transferase (GST)" refers to a detoxification enzyme that catalyzes the transfer of aliphatic, aromatic, or hetero groups from organic sulfur, nitrile, or halogen compounds to glutathione. It is also called glutathione-S-transferase.

The glutathione transferase may include the amino acid sequence of SEQ ID NO: 1. In addition, not only the amino acid sequence consisting of SEQ ID NO: 1, but also an amino acid sequence that is at least 80%, specifically at least 90%, more specifically at least 95%, even more specifically at least 98%, and most specifically at least 99% of the above sequence. An amino acid sequence showing homology includes, without limitation, any amino acid sequence that is substantially the same as that of the enzyme or shows a corresponding effect. In addition, it is clear that, as long as the amino acid sequence has such homology, amino acid sequences in which some sequences are deleted, modified, substituted, or added are also included within the scope of the present invention.

The term “homology” refers to the degree of similarity between the base sequence encoding a protein or the amino acid sequence constituting the protein. If the homology is sufficiently high, the expression product and protein of the corresponding gene may have the same or similar activity. Additionally, homology can be expressed as a percentage based on the degree of matching to a given amino acid or base sequence. In this specification, a given amino acid or nucleotide sequence and its homologous sequence having the same or similar activity are indicated as “% homology”. For example, standard software for calculating parameters such as score, identity and similarity, specifically BLAST 2.0, or hybridization performed under defined stringent conditions. It can be confirmed by comparing sequences experimentally, and appropriate hybridization conditions defined are within the scope of the relevant technology and methods well known to those skilled in the art (e.g., J. Sambrook et al., Molecular Cloning, A Laboratory Manual, 2nd Edition, Cold Spring Harbor Laboratory press, Cold Spring Harbor, New York, 1989; F.M. Ausubel et al., Current Protocols in Molecular Biology, John Wiley & Sons, Inc., New York.

According to one embodiment, the fusion protein (GST-Afb) may be one in which the N terminus of the glutathione transferase (GST) and the C terminus of the disease cell targeting peptide (Afb) are connected. More specifically, the fusion protein (GST-Afb) may be one in which the N terminus of the glutathione transferase (GST) and the C terminus of the disease cell targeting peptide (Afb) are connected by a linker. The connection may be, for example, a peptide bond (amide-type covalent bond), but is not limited thereto.

The linker may include the amino acid sequence of SEQ ID NO: 6. In addition, not only the amino acid sequence consisting of SEQ ID NO: 6, but also at least 80%, specifically at least 90%, more specifically at least 95%, more specifically at least 98%, and most specifically at least 99% of the above sequence. As an amino acid sequence showing homology, any amino acid sequence that is substantially the same as that of the linker or shows a corresponding effect is included without limitation. In addition, it is clear that, as long as the amino acid sequence has such homology, amino acid sequences in which some sequences are deleted, modified, substituted, or added are also included within the scope of the present invention.

The term “Metal-Organic framework (MOFs)” refers to a crystalline organic-inorganic hybrid polymer formed by connecting metal ions or metal nodes with organic molecules.

Specifically, the metal-organic framework may have a porous three-dimensional structure in which metal ions or metal nodes are connected between organic molecules.

The metal node may refer to a metal ion cluster containing a plurality of metal ions. Therefore, the metal-organic framework may have a structure in which organic molecules and metal ions within metal nodes are connected. The connection between the organic molecule and the metal ion or metal node (metal ion cluster) may be through a coordination bond.

The metal ion may be a pure metal or an electrolytic metal, and examples of pure metals or electrolytic metals include, but are not limited to, magnesium (Mg), calcium (Ca), manganese (Mn), iron (Fe), copper (Cu), and zinc (Zn). ), gallium (Ga), germanium (Ge), selenium (Se), strontium (Sr), zirconium (Zr), molybdenum (Mo), silver (Ag), or platinum (Pt).

Accordingly, the metal node may include one or more of the metal ions listed above. According to one embodiment, the metal node may be a zirconium (Zr) ion cluster containing a plurality of zirconium (Zr) ions. Preferably, the metal node may be Zr ₆ , but is not limited thereto.

According to one embodiment, in the GST-Afb-MOFs nanoparticle, the portion of the glutathione transferase (GST) that is not connected to the disease cell targeting peptide (Afb) is the metal-organic framework (MOFs). ), a fusion protein (GST-Afb) linked to the glutathione transferase and disease cell targeting peptide may be coated on the surface of the MOFs by being connected to the metal node exposed on the surface of the MOFs. Specifically, the connection between the portion of the GST that is not connected to the Afb and the metal node exposed on the surface of the MOFs may be due to electrostatic interaction.

Therefore, according to one embodiment, in the GST-Afb-MOFs nanoparticle, the Afb is located at the outermost part of the nanoparticle and is exposed to the outside of the nanoparticle, and the Afb of GST linked to the Afb The unconnected portion may be connected to the surface of the MOFs.

More specifically, in the GST-Afb-MOFs nanoparticles, the G-site site (e.g., amino acid residue or amino acid functional group of the G-site site) of the GST is exposed on the surface of the MOFs. The metal nodes (e.g., zirconium (Zr) ion clusters, preferably, Zr ₆ ) or metal ions (e.g., zirconium (Zr) ions) are connected by electrostatic interaction, thereby allowing the GST-Afb to bind to the MOFs. It may be coated on the surface.

The term "G-site of glutathione transferase" refers to the glutathione binding site in glutathione transferase and the thioredoxin-like domain of glutathione transferase. -like domain) (cf. Dirr et al., Eur. J. Biochem. 220:645, 1994; Armstrong, Chem. Res. Toxicol. 10:2, 1997; ADANG et al., Biochem. J (1990) 269, 47-54).

According to one embodiment, in the GST-Afb-MOFs nanoparticles, the G-site region of the GST is connected to the metal node or metal ion exposed on the surface of the MOFs, thereby allowing the GST-Afb to form the MOFs. It can be coated more stably and more uniformly over the entire surface area. Because of this, the GST-Afb-MOFs nanoparticles exhibit superior protein adsorption blocking ability in a biological environment, targeting target cells with high efficiency and being absorbed into the target cells. As a result, the GST-Afb-MOFs nanoparticles can exhibit a significantly excellent target cell-specific drug delivery effect and a cell death effect by generating photoreactive ROS within target cells.

In the GST-Afb-MOFs nanoparticles, the organic molecules constituting the MOFs connect the metal nodes to form gaps between the metal nodes, thereby forming empty regions, that is, pores, inside the MOFs. Any organic molecule can be used without limitation.

In addition, the organic molecules constituting the MOFs may be photosensitizers that generate reactive oxygen species (ROS) in response to light.

The term “photosensitizer” may be a substance that is activated by light and can undergo a photochemical reaction. For example, the photosensitizer is an organic molecule used in photocatalysis, photodynamic therapy, etc., and may be an organic molecule that is mainly introduced into the human body in cancer treatment and then kills cells by generating singlet oxygen through light irradiation.

According to one embodiment, in the GST-Afb-MOFs nanoparticles, the organic molecules constituting the MOFs are porphyrins or polymers, chlorins or polymers, or bacteriochlorins. Or it may contain at least one selected from the group consisting of polymers, phtalocyanine compounds or polymers, naphthalocyanines compounds or polymers, and 5-aminoevuline esters compounds or polymers. More specifically, the organic molecule constituting the MOFs may be meso-tetra(4-carboxyphenyl)porphine (TCPP) or a derivative thereof, but is limited thereto. Rather, any organic molecule that can form the MOFs and function as a photosensitizer can be used without limitation.

In the GST-Afb-MOFs nanoparticles, the metal-organic framework (MOFs) may include porphyrin-based MOFs. Specifically, the MOFs include MIL-88A, MIL-88Bt, MIL-89, MIL-127, MIL-101, MIL-100, MIL-53, MOF-74, UiO-66, UiO-67, ZIF-8 , ZIFs, HKUST-1, M2 (dobpdc), NU-1000, PCN-222, and PCN-224.

In the GST-Afb-MOFs nanoparticle, the disease cell targeting peptide (Afb) is a peptide that targets disease cells, and the GST-Afb-MOFs nanoparticle moves to the targeted disease cell and is absorbed into the cell. refers to a peptide that induces

The disease cells are abnormal cells with characteristics distinct from normal cells and may refer to cells that cause disease. For example, the diseased cells may be cancer cells. The cancers include brain tumor, benign astrocytoma, malignant astrocytoma, pituitary adenoma, meningioma, brain lymphoma, oligodendroglioma, intracranial tumor, ependymoma, brainstem tumor, head and neck tumor, laryngeal cancer, oropharyngeal cancer, nasal cavity/paranasal sinus cancer, nasopharyngeal cancer, Salivary gland cancer, hypopharyngeal cancer, thyroid cancer, oral cancer, thoracic tumor, small cell lung cancer, non-small cell lung cancer, thymus cancer, mediastinal tumor, esophageal cancer, breast cancer, male breast cancer, abdominal tumor, stomach cancer, liver cancer, gallbladder cancer, biliary tract cancer, pancreatic cancer, small intestine cancer. , colon cancer, anal cancer, bladder cancer, kidney cancer, prostate cancer, cervical cancer, endometrial cancer, ovarian cancer, uterine sarcoma, and skin cancer. In addition, the cancer may be any one or more selected from the group consisting of stomach cancer, breast cancer, lung cancer, liver cancer, esophageal cancer, and prostate cancer that has resistance to anticancer drugs (e.g., multidrug resistance).

Specifically, the Afb specifically targets specific factors (e.g., markers, receptors, proteins, etc.) present in the diseased cells (e.g., expressed on the surface of the diseased cells) and associated with the disease, or are associated with the disease. It may be a peptide that specifically binds to a specific factor. For example, a specific factor associated with the disease may be a receptor expressed on the surface of cancer cells. Examples of receptors expressed on the surface of the cancer cells include, but are not limited to, CD44, CD133, CD166, CD19, CD20, CD21, CD22, CD45, BCMA, MART-1, MAGE-A3, glycoprotein 100 (gp100), NY-ESO-1, HER2 (ErbB2), IGF2B3, EGFRvIII, Kallikrein 4, KIF20A, Lengsin, Meloe, MUC-1, MUC5AC, MUC-16, B7-H3, B7-H6, CD70, CEA, CSPG4, EphA2 , EpCAM, EGFR family, FAP, FRα, glupican-3, GD2, GD3, HLA-A1+MAGE1, IL-11Rα, IL-23Rα2, Lewis-Y, mesothelin, NKG2D ligand, PSMA, ROR1, survivin , TAG72, or VEGFR2.

According to one embodiment, the Afb specifically targets HER2 (human epidermal growth factor receptor 2) and/or EGFR (epidermal growth factor receptor) on disease cells (e.g., expressed on the surface of cancer cells), or It may specifically bind to HER2 and/or EGFR, but is not limited thereto, and the Afb may be used in various ways to specifically target or specifically bind to the target, depending on the target. can be designed

According to one embodiment, the Afb may include the amino acid sequence of SEQ ID NO: 2 (HER2 target Afb) or SEQ ID NO: 3 (EGFR target Afb). In addition, not only the amino acid sequence consisting of SEQ ID NO: 2 or SEQ ID NO: 3, but also 80% or more, specifically 90% or more, more specifically 95% or more, more specifically 98% or more, most specifically is an amino acid sequence showing 99% or more homology, and includes without limitation any amino acid sequence that is substantially the same as the above-mentioned peptide or shows a corresponding efficacy. In addition, it is clear that, as long as the amino acid sequence has such homology, amino acid sequences in which some sequences are deleted, modified, substituted, or added are also included within the scope of the present invention.

According to one embodiment, in the GST-Afb-MOFs nanoparticle, the fusion protein (GST-Afb) in which the glutathione transferase (GST) and the disease cell targeting peptide (Afb) are linked is SEQ ID NO: 4 ( It may include the amino acid sequence of GST-HER2) or SEQ ID NO: 5 (GST-EGFR). In addition, not only the amino acid sequence consisting of SEQ ID NO: 4 or SEQ ID NO: 5, but also 80% or more, specifically 90% or more, more specifically 95% or more, more specifically 98% or more, most specifically is an amino acid sequence showing more than 99% homology, and includes without limitation any amino acid sequence that is substantially the same as that of the protein or shows a corresponding effect. In addition, it is clear that, as long as the amino acid sequence has such homology, amino acid sequences in which some sequences are deleted, modified, substituted, or added are also included within the scope of the present invention.

According to one embodiment, the GST-Afb-MOFs nanoparticles include a first fusion protein (first GST-Afb) linked to GST and Afb targeting HER2 on diseased cells, and Afb targeting GST and EGFR on diseased cells. A linked second fusion protein (second GST-Afb), or a combination thereof, may be coated on the surface of the MOFs.

That is, in the GST-Afb-MOFs nanoparticles, there may be two or more types of fusion proteins (GST-Afb) coated on the surface of the MOFs. For example, in the case of the two or more types of GST-Afb, each may include Afb targeting a different target. Because of this, the GST-Afb-MOFs nanoparticles may be capable of multiple targeting.

Therefore, according to one embodiment, the GST-Afb-MOFs nanoparticles surface-coated with the two or more types of GST-Afb are at least one of a plurality of factors (e.g., disease-related factors) targeted by the Afb. It targets a single type of diseased cell or multiple types of diseased cells and can be absorbed into the corresponding cell.

The GST-Afb-MOFs nanoparticles are specifically absorbed by the disease cells, that is, disease cells targeted by the Afb (e.g., cells with disease-related factors targeted by the Afb) and emit light within the cells. By generating active oxygen in response, only the diseased cells may be specifically killed.

Therefore, according to one embodiment, the GST-Afb-MOFs nanoparticles may exhibit a target cell-specific photoreactive cell death effect.

The term “reactive oxygen species (ROS)” refers to a chemically reactive molecule containing an oxygen atom. Active oxygen is a compound of oxygen generated within living organisms that is highly reactive due to unpaired electrons and has a strong oxidizing power that attacks biological tissues and damages cells.

The active oxygen may include hydrogen superoxide ion, hydrogen peroxide, hydroxy radical, singlet oxygen, etc., and according to one embodiment, the GST-Afb-MOFs nanoparticles react to light within the cell to produce singlet oxygen. It may kill cells by producing .

The term "singlet oxygen ( ¹ O ₂ )" may be oxygen in which electrons have been transferred to only one of the two π* orbitals in the oxygen molecule, and because it is in an excited state, it may be very reactive.

The GST-Afb-MOFs nanoparticles may have a drug loaded therein. Specifically, the GST-Afb-MOFs nanoparticles may have a drug loaded inside the metal-organic framework (MOFs).

In one embodiment, the drug may be an anti-cancer agent, but is not limited thereto, and may be included without limitation as long as it is an active substance that regulates biological reactions such as genes such as DNA and RNA, and proteins, and affects physiological functions. . Specifically, the drug is a compound or a variant or derivative thereof, a peptide or a variant or derivative thereof, an aptamer or a variant or derivative thereof, an antibody or a variant or derivative thereof, or a drug that treats diseases by regulating the physiological activity of cells. It may be a therapeutic agent, a gene therapy, etc.

The anticancer agent includes abarelix (Plenaxis depot ^® )]; aldesleukin (Prokine ^® )]; Aldesleukin (Proleukin ^® )]; Alemtuzumab (Campath ^® )]; alitretinoin (Panretin ^® )]; allopurinol (Zyloprim ^® )]; altretamine (Hexalen ^® )]; amifostine (Ethyol ^® )]; anastrozole (Arimidex ^® )]; arsenic trioxide (Trisenox ^® )]; asparaginase (Elspar ^® )]; azacitidine (Vidaza ^® )]; bevacuzimab (Avastin ^® )]; bexarotene capsules (Targretin ^® )]; bexarotene gel (Targretin ^® )]; bleomycin (Blenoxane ^® )]; bortezomib (Velcade ^® )]; busulfan intravenous (Busulfex ^® )]; oral busulfan [busulfan oral (Myleran ^® )]; calusterone (Methosarb ^® )]; capecitabine (Xeloda ^® )]; carboplatin (Paraplatin ^® )]; carmustine (BCNU ^® , BiCNU ^® )]; carmustine (Gliadel ^® )]; Carmustine and Polifeprosan 20 Implant (Gliadel Wafer ^® ); celecoxib (Celebrex ^® )]; cetuximab (Erbitux ^® )]; chlorambucil (Leukeran ^® )]; cisplatin (Platinol ^® )]; cladribine (Leustatin ^® ) 2-CdA ^® ]; clofarabine (Clolar ^® )]; cyclophosphamide (Cytoxan ^® , Neosar ^® )]; cyclophosphamide (Cytoxan Injection ^® )]; cyclophosphamide (Cytoxan Tablet ^® )]; cytarabine (Cytosar-U ^® )]; cytarabine liposomal (DepoCyt ^® )]; dacarbazine (DTIC-Dome ^® )]; dactinomycin, actinomycin D (Cosmegen ^® )]; Darbepoetin alfa (Aranesp ^® )]; daunorubicin liposomal (Danuoxome ^® )]; daunorubicin, daunomycin (Daunorubicin ^® )]; daunorubicin, daunomycin (Cerubidine ^® )]; Denileukin diftitox (Ontak ^® )]; dexrazoxane (Zinecard ^® )]; docetaxel ( ^Taxotere® )]; doxorubicin (Adriamycin PFS ^® )]; doxorubicin (Adriamycin ^® , Rubex ^® )]; doxorubicin (Adriamycin PFS Injection ^® )]; doxorubicin liposomal (Doxil ^® )]; dromostanolone propionate (dromostanolone ^® )]; dromostanolone propionate (masterone injection ^® )]; Elliott's B Solution (Elliott's B Solution ^® )]; epirubicin (Ellence ^® )]; Epoetin alfa (epogen ^® )]; erlotinib (Tarceva ^® )]; estramustine (Emcyt ^® )]; etoposide phosphate (Etopophos ^® )]; etoposide, VP-16 (Vepesid ^® ); exemestane (Aromasin ^® )]; Filgrastim (Neupogen ^® )]; floxuridine (intraarterial) (FUDR ^® )]; fludarabine (Fludara ^® )]; Fluorouracil [Fluorouracil, 5-FU (Adrucil ^® )]; folvestrant (Faslodex ^® )]; gefitinib (Iressa ^® )]; gemcitabine (Gemzar ^® )]; gemtuzumab ozogamicin (Mylotarg ^® )]; goserelin acetate (Zoladex Implant ^® )]; goserelin acetate (Zoladex ^® )]; histrelin acetate (Histrelin implant ^® )]; hydroxyurea (Hydrea ^® )]; Ibritumomab Tiuxetan (Zevalin ^® )]; idarubicin (Idamycin ^® )]; ifosfamide (IFEX ^® )]; imatinib mesylate (Gleevec ^® )]; interferon alfa-2a (Roferon A ^® )]; Interferon alfa-2b (Intron A ^® )]; irinotecan (Camptosar ^® )]; lenalidomide (Revlimid ^® )]; letrozole (Femara ^® )]; leucovorin (Wellcovorin ^® , Leucovorin ^® )]; Leuprolide Acetate (Eligard ^® )]; levamisole (Ergamisol ^® )]; lomustine-CCNU [lomustine-CCNU (CeeBU ^® )]; meclorethamine, nitrogen mustard (Mustargen ^® ); megestrol acetate ( ^Megace® )]; melphalan, L-PAM (Alkeran ^® ); mercaptopurine [mercaptopurine, 6-MP (purinethol ^® )]; mesna (Mesnex ^® )]; mesna [mesna (Mesnex tabs ^® )]; methotrexate (methotraxate ^® )]; methoxsalen (Uvadex ^® )]; mitomycin C (Mutamycin ^® )]; mitotane (Lysodren ^® )]; mitoxantrone (Novantrone ^® )]; nandrolone phenpropionate (Durabolin-50 ^® )]; nelarabine (Arranon ^® )]; Nofetumomab (Verluma ^® )]; Oprelvekin (Neumega ^® )]; oxaliplatin (Eloxatin ^® )]; paclitaxel (Paxene ^® )]; paclitaxel (Taxol ^® )]; paclitaxel protein-bound particles (Abraxane ^® )]; palifermin (Kepivance ^® )]; pamidronate (Aredia ^® )]; pegademas [pegademase (Adagen (Pegademase Bovine) ^® )]; pegaspargase [pegaspargase (Oncaspar ^® )]; Pegfilgrastim (Neulasta ^® )]; pemetrexed disodium (Alimta ^® )]; pentostatin (Nipent ^® )]; pipobroman (Vercyte ^® )]; plicamycin, mithramycin (Mithracin ^® ); porfimer sodium (Photofrin ^® )]; procarbazine (Matulane ^® )]; quinacrine (Atabrine ^® )]; Rasburicase ( ^Elitek® )]; Rituximab (Rituxan ^® )]; sargramostim (Leukine ^® )]; Sargramostim (Prokine ^® )]; sorafenib (Nexavar ^® )]; streptozocin (Zanosar ^® )]; sunitinib maleate (Sutent ^® )]; Talc [talc (Sclerosol ^® )]; tamoxifen (Nolvadex ^® )]; temozolomide (Temodar ^® )]; teniposide, VM-26 (Vumon ^® )]; testolactone (Teslac ^® )]; Thioguanine [thioguanine, 6-TG (thioguanine ^® )]; thiotepa (thioplex ^® )]; topotecan (Hycamtin ^® )]; toremifene (Fareston ^® )]; Tositumomab (Bexxar ^® )]; Tositumomab/I-131 Tositumomab (Bexxar ^® ); Trastuzumab (Herceptin ^® )]; Tretinoin [tretinoin, ATRA ( ^Vesanoid® )]; Uracil Mustard (Uracil Mustard Capsules ^® )]; valrubicin (Valstar ^® )]; vinblastine (Velban ^® )]; vincristine (Oncovin ^® )]; vinorelbine (Navelbine ^® )]; It may include zoledronate (Zometa ^® )], its pharmaceutically acceptable salt, and mixtures thereof.

The GST-Afb-MOFs nanoparticles are specifically absorbed into the disease cells, that is, disease cells targeted by the Afb (e.g., cells having a disease-related factor targeted by the Afb) and stored within the cells. It may be releasing drugs.

Therefore, according to one embodiment, the GST-Afb-MOFs nanoparticles may exhibit a target cell-specific drug delivery effect.

According to one embodiment, the average diameter of the GST-Afb-MOFs nanoparticles is about 100 to 500, 150 to 400, 150 to 350, 150 to 300, 200 to 400, 200 to 350, or 200 to 300 nm. You can. Most preferably, the average diameter of the GST-Afb-MOFs nanoparticles may be about 200 to 300 nm. When the average diameter of the GST-Afb-MOFs nanoparticles is within the above numerical range, the GST-Afb-MOFs nanoparticles have excellent stability, protein adsorption blocking ability, cell targeting ability, cell uptake ability, drug encapsulation rate, drug delivery ability, Alternatively, it may exhibit photoreactive ROS generation ability, thereby increasing protein adsorption blocking effect, cell targeting effect, target cell specific drug delivery effect, or target cell specific photoreactive cell death effect. In addition, if the average diameter of the GST-Afb-MOFs nanoparticles is less than the above numerical range, the drug encapsulation rate or cell targeting ability may be reduced, and if it is greater than the above numerical range, stability, protein adsorption blocking ability, and cell uptake ability may be reduced. , or drug delivery ability may be reduced. Therefore, when the average diameter of the GST-Afb-MOFs nanoparticles is outside the above numerical range, the protein adsorption blocking effect, cell targeting effect, target cell-specific drug delivery effect, or target cell effect of the GST-Afb-MOFs nanoparticles The specific photoreactive cell killing effect may be reduced.

According to one embodiment, the surface charge (or zeta potential) of the GST-Afb-MOFs nanoparticles is about -10 to -1, -9 to -1, -8 to -1, -7 to -1, -6. to -1, -5 to -1, -10 to -2, -9 to -2, -8 to -2, -7 to -2, -6 to -2, -5 to -2, -10 to - It may be 3, -9 to -3, -8 to -3, -7 to -3, -6 to -3, or -5 to -3 mV. When the surface charge (or zeta potential) of the GST-Afb-MOFs nanoparticles is within the above numerical range, the GST-Afb-MOFs nanoparticles have excellent stability, protein adsorption blocking ability, cell targeting ability, cell uptake ability, and drug encapsulation rate. , drug delivery ability, or photoreactive ROS generation ability, etc. may increase the protein adsorption blocking effect, cell targeting effect, target cell specific drug delivery effect, or target cell specific photoreactive cell killing effect. In addition, if the surface charge (or zeta potential) of the GST-Afb-MOFs nanoparticles is less than the above numerical range, stability, cell uptake ability, drug delivery ability, or photoreactive ROS generation ability may be reduced, and if it exceeds the above numerical range, In this case, stability, protein adsorption blocking ability, or cell targeting ability may be reduced. Therefore, when the surface charge (or zeta potential) of the GST-Afb-MOFs nanoparticles is outside the above numerical range, the protein adsorption blocking effect, cell targeting effect, and target cell-specific drug delivery of the GST-Afb-MOFs nanoparticles The effect, or target cell-specific photoreactive cell death effect, may be reduced.

Another aspect provides a target cell-specific drug delivery composition (or drug delivery pharmaceutical composition) containing the GST-Afb-MOFs nanoparticles as an active ingredient.

Another aspect provides a composition for target cell-specific photoreactive cell death (or pharmaceutical composition for cell death) comprising the GST-Afb-MOFs nanoparticles as an active ingredient.

The target cell may be a disease cell targeted by the Afb (eg, a cell having a disease-related factor targeted by the Afb), and the disease cell may be a cancer cell.

Another aspect provides a pharmaceutical composition for preventing or treating cancer containing the GST-Afb-MOFs nanoparticles as an active ingredient.

The GST-Afb-MOFs nanoparticles are as described above.

The GST-Afb-MOFs nanoparticles may have a drug loaded therein, and the drug is as described above.

According to one embodiment, the GST-Afb-MOFs nanoparticles themselves exhibit target cell-specific photoreactive cytotoxicity and can kill targeted cancer cells, so even if no drug is loaded therein, the GST- Afb-MOFs nanoparticles may be included as an active ingredient in the target cell-specific photoreactive cell killing composition (or cell killing pharmaceutical composition) or cancer prevention or treatment pharmaceutical composition.

In addition, according to one embodiment, in the case of the GST-Afb-MOFs nanoparticles with a drug loaded therein, the synergistic effect of chemotherapy by the drug and photodynamic therapy (PDT) by photoreactive cytotoxicity It can induce a significantly better killing effect on target cells (eg, target cancer cells).

The cancers include brain tumor, benign astrocytoma, malignant astrocytoma, pituitary adenoma, meningioma, brain lymphoma, oligodendroglioma, intracranial tumor, ependymoma, brainstem tumor, head and neck tumor, laryngeal cancer, oropharyngeal cancer, nasal cavity/paranasal sinus cancer, nasopharyngeal cancer, Salivary gland cancer, hypopharyngeal cancer, thyroid cancer, oral cancer, thoracic tumor, small cell lung cancer, non-small cell lung cancer, thymus cancer, mediastinal tumor, esophageal cancer, breast cancer, male breast cancer, abdominal tumor, stomach cancer, liver cancer, gallbladder cancer, biliary tract cancer, pancreatic cancer, small intestine cancer. , colon cancer, anal cancer, bladder cancer, kidney cancer, prostate cancer, cervical cancer, endometrial cancer, ovarian cancer, uterine sarcoma, and skin cancer. In addition, the cancer may be any one or more selected from the group consisting of stomach cancer, breast cancer, lung cancer, liver cancer, esophageal cancer, and prostate cancer that has resistance to anticancer drugs (e.g., multidrug resistance).

The term “effective ingredient” means an appropriately effective amount of an ingredient that affects a beneficial or desirable clinical or biochemical outcome. Specifically, it may refer to an effective amount of GST-Afb-MOFs nanoparticles.

The effective amount may be administered once or more and may prevent the disease, treat the disease state, including but not limited to, alleviate symptoms, reduce the extent of the disease, stabilize the disease state (i.e., not worsen), delay disease progression, or It may be an appropriate amount for reduction of the rate, or improvement or temporary relief and alleviation (partial or total) of the disease state.

The term “prevention” refers to any action that inhibits or delays the onset of a disease, such as cancer, due to administration of the composition.

The term “treatment” refers to any action that improves or benefits the symptoms of a disease that has already been caused, such as cancer, by administering the composition.

The composition may further include pharmaceutically acceptable excipients, diluents, or carriers. The term “pharmaceutically acceptable excipient, diluent or carrier” may mean an excipient, diluent or carrier that does not irritate living organisms and does not inhibit the biological activity and properties of the injected compound or active ingredient. Here, “pharmaceutically acceptable” means that it does not inhibit the activity of the active ingredient and does not have toxicity beyond what is acceptable for the subject of application (prescription).

As the pharmaceutically acceptable carrier, binders, lubricants, disintegrants, excipients, solubilizers, dispersants, stabilizers, suspending agents, colorants, flavors, etc. can be used for oral administration, and buffers and preservatives for injections. , analgesics, solubilizers, isotonic agents, stabilizers, etc. can be mixed and used, and for topical administration, bases, excipients, lubricants, preservatives, etc. can be used.

The composition is formulated in the form of oral dosage forms such as powders, granules, tablets, capsules, suspensions, emulsions, syrups, and aerosols, external preparations, suppositories, unit dosage ampoules, or injections in the form of multiple dosages according to conventional methods. can be used When formulating the composition, it may be prepared by adding diluents or excipients such as commonly used fillers, extenders, binders, wetting agents, disintegrants, or surfactants.

When the composition is prepared as a parenteral formulation, it can be formulated in the form of injections, transdermal administration, nasal inhalation, and suppositories along with a suitable carrier according to methods known in the art. When formulated as an injection, suitable carriers include sterilized water, ethanol, polyols such as glycerol or propylene glycol, or mixtures thereof, preferably Ringer's solution, PBS (phosphate buffered saline) containing triethanolamine, or sterile injectable carrier. Isotonic solutions such as water or 5% dextrose can be used. When formulated for transdermal administration, it can be formulated in the form of ointments, creams, lotions, gels, external solutions, paste preparations, linear preparations, and aerol preparations. In the case of nasal inhalation, it can be formulated in the form of an aerosol spray using suitable propellants such as dichlorofluoromethane, trichlorofluoromethane, dichlorotetrafluoroethane, and carbon dioxide. When formulated as a suppository, the base is Wethepsol ( witepsol), Tween 61, polyethylene glycols, cocoa fat, laurel paper, polyoxyethylene sorbitan fatty acid esters, polyoxyethylene stearates, sorbitan fatty acid esters, etc. can be used. In addition, the composition may additionally contain or be mixed with various pharmaceutically acceptable carriers, such as physiological saline or organic solvents, and may be used in combination with carriers such as glucose, sucrose, or dextran to increase stability or absorption. Carbohydrates, antioxidants such as ascorbic acid or glutathione, chelating agents, low molecular weight proteins or other stabilizers may be additionally included or used in combination with them.

The composition may be administered through any general route as long as it can reach the target tissue. Specifically, the composition may be administered for intramuscular administration, subcutaneous administration, intraperitoneal administration, intravenous administration, oral administration, or dermal administration. It may be selected from the group consisting of compositions for administration, ocular administration, and intracerebral administration.

The composition may be administered in a pharmaceutically effective amount, where the term "pharmaceutically effective amount" means an amount sufficient to treat or prevent a disease with a reasonable benefit/risk ratio applicable to medical treatment or prevention, The effective dose level is determined by the severity of the disease, the activity of the drug, the patient's age, weight, health, gender, the patient's sensitivity to the drug, the administration time, route of administration and excretion rate of the composition of the present invention used, the treatment period, and the present invention used. It can be determined based on factors including drugs combined or used simultaneously with the composition and other factors well known in the medical field. The composition can be administered alone or in combination with ingredients known to exhibit preventive or therapeutic effects on certain known diseases (eg, cancer). It is important to consider all of the above factors and administer the amount that will achieve the maximum effect with the minimum amount without side effects.

The dosage of the composition can be determined by a person skilled in the art in consideration of the purpose of use, the degree of addiction of the disease, the patient's age, weight, gender, medical history, or the type of substance used as an active ingredient. For example, the composition of the present invention can be administered at about 0.1 ng to about 1,000 mg/kg, preferably 1 ng to about 100 mg/kg per adult, and the frequency of administration of the composition of the present invention is not particularly limited. However, it can be administered once a day, or the dose can be divided and administered several times. The above dosage or frequency of administration does not limit the scope of the present invention in any way.

Among the terms or elements mentioned in the composition, those mentioned in the description of the nanoparticles are understood to be the same as previously mentioned in the description of the nanoparticles.

Another aspect is the GST-Afb-MOFs nanoparticles, or a composition for target cell-specific drug delivery (or pharmaceutical composition for drug delivery) containing the same as an active ingredient, or a composition for target cell-specific photoreactive cell killing ( or a pharmaceutical composition for cell death), or a pharmaceutical composition for cancer prevention or treatment is provided to a subject.

The GST-Afb-MOFs nanoparticles and the composition containing the same as an active ingredient are as described above.

The above-mentioned entities include mammals including cattle, horses, sheep, pigs, goats, camels, antelopes, dogs, cats, rats, livestock, humans, farmed fish, and mammals other than humans that develop or are at risk of developing cancer. etc. may be included without limitation.

The term “administration” means introducing a predetermined substance into an individual by an appropriate method, and the composition of the present invention can be administered through any general route as long as it can reach the target tissue. It may be administered intraperitoneally, intravenously, intramuscularly, subcutaneously, intradermally, orally, topically, intranasally, intrapulmonaryly, or rectally, but is not limited thereto. However, when administered orally, proteins are digested, so it is desirable for oral compositions to be coated with the active agent or formulated to protect them from decomposition in the stomach. Additionally, pharmaceutical compositions can be administered by any device that can transport the active agent to target cells.

The composition may be administered as an individual therapeutic agent or in combination with other therapeutic agents, and may be administered sequentially or simultaneously with conventional therapeutic agents. And it can be administered single or multiple times. Considering all of the above factors, it is important to administer an amount that can achieve maximum effect with the minimum amount without side effects, and can be easily determined by a person skilled in the art.

Among the terms or elements mentioned in the above method, those mentioned in the description of the nanoparticles and composition are understood to be the same as previously mentioned in the description of the nanoparticles and composition.

Another aspect provides the use of the GST-Afb-MOFs nanoparticles for target cell-specific drug delivery, target cell-specific photoreactive cell killing, or cancer prevention or treatment.

Another aspect is a composition for target cell-specific drug delivery (or pharmaceutical composition for drug delivery), a composition for target cell-specific photoreactive cell death (or pharmaceutical composition for cell death) of the GST-Afb-MOFs nanoparticles. ), or for preparing a pharmaceutical composition for preventing or treating cancer.

The GST-Afb-MOFs nanoparticles are as described above.

Among the terms or elements mentioned in the above uses, those mentioned in the description of the nanoparticles, compositions, and methods are understood to be the same as previously mentioned in the description of the nanoparticles, compositions, and methods.

According to one aspect, metal-organic framework nanoparticles exhibit excellent protein adsorption blocking effect, cell targeting effect, target cell-specific drug delivery effect, and cell killing effect by generating photoreactive ROS within target cells, thereby killing target cells. It can be useful as a specific drug carrier or disease treatment agent (eg, cancer treatment), and in particular, it can induce a synergistic effect of photodynamic therapy and chemotherapy on cancer cell death.

Figure 1 is a schematic diagram schematically showing the structure, protein adsorption blocking function, cell targeting function, target cell-specific drug delivery function, and photoreactive ROS generation function in target cells of a GST-Afb-MOFs complex according to an embodiment. am.
Figure 2 shows GST-H-PCN coated on the surface of PCN with GST-HER2, a GST-Afb-MOFs complex according to an embodiment; And this is a schematic diagram schematically showing the form of GST-E-PCN with GST-EGFR coated on the surface of PCN.
Figure 3 is a TEM image performed on GST-H-PCN and GST-E-PCN, which are GST-Afb-MOFs complexes according to an example.
Figure 4 is a graph showing the results of PXRD analysis on GST-Afb-MOFs complexes, GST-H-PCN and GST-E-PCN, and a control group (PCN) according to an example.
Figure 5 is a graph showing the results of measuring the sizes of GST-Afb-MOFs complexes, GST-H-PCN and GST-E-PCN, and the control group (PCN) according to an example using a particle size analyzer (DLS).
Figure 6 shows the results of measuring the zeta potential of GST-Afb-MOFs complexes, GST-H-PCN and GST-E-PCN, and the control group (PCN, GST-HER2, and GST-EGFR) according to an example. It's a graph.
Figure 7 is a schematic diagram showing the difference in protein adsorption degree between GST-H-PCN and GST-E-PCN cultured in serum medium, which are GST-Afb-MOFs complexes according to an example, and the control group (PCN) (a); An image of the gel electrophoresis result (b), and a graph of the result of measuring the intensity of fluorescence emission by FITC-modified serum protein (c).
Figure 8 is a molecular dynamics (MD) simulation schematic diagram predicting the connection (or adsorption) structure of the MOFs of the GST-Afb-MOFs complex and GST-Afb according to one embodiment (a: connection site in GST-Afb; b: Linkage site in MOFs; c: predicted linkage structure for GST-Afb-PCN complex).
Figure 9 is a molecular dynamics (MD) simulation schematic diagram showing the connection (or adsorption) structure of PCN and GST-Afb of the GST-Afb-PCN complex, which is a GST-Afb-MOFs complex according to an embodiment.
Figure 10 shows the electrostatic interaction between the G-site of GST and the Zr ₆ node of PCN in the GST-Afb-PCN complex, a GST-Afb-MOFs complex according to one embodiment, using the radial distribution function (RDF). This is a graph showing the results of analyzing the residues in GST that induce its action.
Figure 11 shows a GST-Afb-PCN complex, a GST-Afb-MOFs complex according to an embodiment, between a specific region of GST (G-site or side region) and a specific region (frame or pore) on the surface of PCN. This is a graph showing the results of analyzing the interaction energy.
Figure 12 shows the total interaction energy between the hinge region, GST region, or Afb region of GST-Afb and the PCN surface in the GST-Afb-PCN complex, which is a GST-Afb-MOFs complex according to an embodiment. This is the result graph.
Figure 13 shows the targeting ability of GST-Afb-MOFs complex, GST-H-PCN and GST-E-PCN, on SK-BR-3 cells and MDA-MB-468 cells, respectively, according to an example. This image shows the results of fluorescence staining imaging analysis.
Figure 14 is a cell fluorescence showing the multi-cell targeting ability of GST-E/H-PCN, a GST-Afb-MOFs complex according to one embodiment, targeting both SK-BR-3 cells and MDA-MB-468 cells simultaneously. This image shows the results of dye imaging analysis.
Figure 15 is a flow cytometry showing the targeting ability of GST-H-PCN and GST-E-PCN, which are GST-Afb-MOFs complexes according to an example, against SK-BR-3 cells and MDA-MB-468 cells, respectively. This is an image showing the analysis measurement results.
Figure 16 is an image showing the results of analyzing the photoreactive ROS generation ability in cells of GST-E-PCN, a GST-Afb-MOFs complex according to an example.
Figure 17 shows the drug delivery and release ability of GST-Afb-MOFs complexes, GST-H-PCN and GST-E-PCN, according to an embodiment, and the cancer cell killing effect by cytotoxicity due to light irradiation. This is the result graph showing.
Figure 18 shows the target cell-specific drug delivery effect of the GST-Afb-MOFs complex according to one embodiment, the cell death effect by photoreactive ROS generation in the target cell, and the resulting photodynamic therapy for cancer cell death. and a schematic diagram schematically showing the synergistic effect of chemotherapy.
Figure 19 is an image showing the results of in vivo optical fluorescence imaging analysis on a tumor animal model injected with GST-H-PCN, a GST-Afb-MOFs complex according to an example.
Figure 20 is a graph showing the results of in vitro fluorescence detection analysis of organs and tumors extracted from a tumor animal model injected with GST-H-PCN, a GST-Afb-MOFs complex according to an embodiment.

Hereinafter, the present invention will be described in more detail through experimental examples and examples. However, these experimental examples and examples are for illustrative purposes only and the scope of the present invention is not limited to these experimental examples and examples.

In addition, unless there is a special definition in this specification, all scientific and technical terms used in this specification may have the same meaning as commonly understood by a person skilled in the art in the technical field to which the present invention pertains.

Example 1. Preparation and characterization of MOFs nanoparticles

As MOFs nanoparticles, PCN-224 was prepared.

Specifically, about 100 mg of meso-tetra(4-carboxyphenyl)porphine (TCPP), about 300 mg of ZrOCl ₂ ·8H ₂ O, and about 2.8 g of benzoic acid were mixed into a round shape. A mixture was obtained by dissolving in about 100 mL of dimethylformamide (DMF) in a bottom flask (250 mL). The mixture was heated to about 90°C for about 5 hours with uniform stirring, then cooled to room temperature, centrifuged, and the collected product was washed three times with fresh DMF and four times with acetone, then incubated at about 90°C. PCN-224 was obtained by drying under vacuum for about 12 hours.

TEM (Transmission Electron Microscope) imaging and nuclear magnetic resonance (H ¹ NMR) analysis were performed on the obtained PCN. As a result, the obtained PCN has the form of a spherical particle with a diameter of about 90 ± 10 nm, and is an organic ligand molecule, meso-tetra (4-carboxyphenyl) porphine: TCPP) and as a metal ion, it was confirmed to contain ZrOCl ₂ ·8H ₂ O.

In addition, as a result of nitrogen sorption analysis, it was confirmed that the obtained PCN had a large Brunauer-Emett-Teller surface area of about 2,300 ± 50 m ² /g and a basic pore size of about 1.8 ± 0.2 nm. did.

Through this example, PCN-224, a MOFs nanoparticle, was manufactured, and the PCN-224 was confirmed to have a porous three-dimensional structure formed by combining the organic ligand molecule of TCPP and the metal ion of Zr.

Example 2. Preparation and characterization of enzyme-MOFs complex

Enzyme-MOFs interaction using existing enzymes such as trypsin (Tr), hyaluronidase (HA), transferrin (Tf), and beta-galactosidase (β-gal) The action was tested. Specifically, each of the above enzyme proteins was mixed with MOFs nanoparticles, i.e., PCN obtained in Example 1, in PBS at about 4°C for about 20 minutes, then vigorously stirred and centrifuged to purify the collected particles. By doing so, an enzyme-MOFs complex (enzyme-PCN complex) in which the enzyme protein was coated on the surface of the PCN was obtained.

As a result of measuring the obtained enzyme-PCN complex using a particle size analyzer (Dynamic Light Scattering: DLS), the initial hydrodynamic diameter of the PCN (about 180 ± 20 nm) was reduced to about 390 ± 30 nm after mixing with the enzyme protein. It was confirmed that there was an increase. Through this, it was found that the obtained enzyme-PCN complex was one in which the enzyme protein was coated on the surface of the PCN.

In addition, the surface charge, i.e., zeta potential, of the obtained enzyme-PCN complex was measured using a ZETASIZER NANO ^TM (Malvern) instrument. As a result, the initial surface charge (or zeta potential) of the PCN before mixing with the enzyme protein was measured. ) was about 15 to 35 mV (specifically, about 23 ± 4 mV), but in the case of Tr-PCN (trypsin-PCN complex), the surface charge changed to about 6.06 ± 2 mV, and HA-PCN (hyaluronidase -PCN complex), the surface charge changed to about -13.4±2 mV, for Tf-PCN (transferrin-PCN complex), the surface charge changed to about -15.2±3 mV, and β-gal-PCN (beta In the case of -galactosidase-PCN complex), the surface charge changed to about -14.2±2 mV, confirming that the surface charge of the obtained enzyme-PCN complex had a similar value to the surface charge of each enzyme. Through this, it was found that the obtained enzyme-PCN complex was one in which the enzyme protein was coated on the surface of the PCN.

In addition, as a result of performing powder X-ray diffraction (PXRD) pattern analysis on the obtained enzyme-PCN complex, it was confirmed that the characteristic crystallinity of PCN was well maintained after enzyme coating.

In addition, the enzyme activity of the obtained enzyme-PCN complex was measured using an enzyme assay kit (Beta-Gal assay kit, Thermo Fisher). Typically, experiments were performed on β-gal-PCN. As a result, it was confirmed that the β-gal activity shown by β-gal-PCN was reduced by about 40% compared to the activity of β-gal not attached to PCN. This is understood that in the case of the enzyme-PCN complex, the enzyme activity decreases as the enzyme protein attaches to the surface of the MOFs particles, that is, PCN.

Example 3. Preparation and characterization of GST-Afb-MOFs complex

As Example 2 demonstrated that proteins were firmly coated on the surface of MOFs particles, i.e., PCN, without chemical attachment, an effective disease treatment nanosystem was constructed using cell-targeting peptide (Afb).

Specifically, two types of Afb proteins targeting HER2 (H) or EGFR (E) receptors that are overexpressed in various cancer types were used, and each of the Afb proteins was activated by glutathione S-transferase (GST). was fused with to obtain GST-Afb fusion proteins (GST-HER2 and GST-EGFR). More specifically, the gene encoding HER2 or EGFR containing the linker sequence (SEQ ID NO: 6: GGGLVPRGSGGGCGGGGTGGGSGGG) was inserted into the IPTG-derived pETDuet plasmid vector, which is a plasmid vector containing a hexahistidine tag at the N-terminus and a GST-encoding gene. , the vector was introduced into E. coli strain, BL21 (DE) to obtain a transformed strain. The transformed strain was induced to overexpress GST-HER2 or GST-EGFR by culturing it in LB medium at about 30°C for about 16 hours. After incubation, the culture was centrifuged at approximately 5000 g for approximately 10 min at approximately 4°C, and the collected pellet was resuspended in phosphate buffer (approximately 50 mM sodium phosphate and approximately 100 mM sodium chloride, pH 6.5). The suspension was incubated with lysozyme for approximately 20 minutes at room temperature, sonicated for approximately 10 minutes with approximately 30 seconds between each incubation, and then centrifuged at approximately 12000 g for approximately 1 hour at approximately 4°C. The supernatant obtained after centrifugation was purified by immobilized metal affinity chromatography (1 mL HisTrap FF column, GE HealthCare) using FPLC (Fast protein liquid chromatography) to produce GST-Afb fusion proteins GST-HER2 and GST-EGFR. Obtained. The obtained GST-HER2 and GST-EGFR were dialyzed in PBS (pH 7.4) overnight.

Then, about 0.5 mg of MOFs nanoparticles (PCN obtained in Example 1 above) and about 0.5 mg of GST-HER2 or GST-EGFR were incubated in about 2 mL of neutral PBS at about 4°C for about 20 minutes. After mixing, unreacted free proteins were removed by centrifugation (3220 xg, Eppendorf Centrifuge 5415R), and GST-Afb-MOFs complex (GST-Afb-PCN complex) was separated and collected. Specifically, GST-Afb-PCN complex with GST-HER2 coated on the surface of PCN (GST-H-PCN) and GST-Afb-PCN complex with GST-EGFR coated on the surface of PCN (GST-E-PCN). was obtained (Figure 2). The obtained GST-Afb-PCN complex was washed once with water and redispersed in PBS.

As a result of TEM imaging of the GST-Afb-PCN complex, it was confirmed that the GST-Afb-PCN complex had the shape of a spherical particle, as shown in Figure 3.

In addition, as a result of performing PXRD analysis on the GST-Afb-PCN complex, as shown in Figure 4, the crystalline framework of the GST-Afb-PCN complex was not damaged even though GST-Afb was coated on the surface of the GST-Afb-PCN complex. It was confirmed that it was well maintained.

In addition, as a result of measuring the GST-Afb-PCN complex using a particle size analyzer (Dynamic Light Scattering: DLS), as shown in Figure 5, the initial hydrodynamic diameter of the PCN before mixing with the GST-Afb (about 180 ± 20 nm) was confirmed to have further increased after mixing with GST-Afb (GST-H-PCN: approximately 260±20 nm; GST-E-PCN: approximately 250±20 nm). Through this, it was found that the GST-Afb-PCN complex consisted of GST-Afb coated on the surface of the PCN.

In addition, as a result of measuring the surface charge, i.e., zeta potential, of the GST-Afb-PCN complex using a ZETASIZER NANO ^TM (Malvern) instrument, as shown in Figure 6, the PCN before mixing with the GST-Afb The initial surface charge (or zeta potential) of was about 15 to 35 mV (specifically, about 20 to 30 mV), whereas for GST-H-PCN, the surface charge was about -6 to -1 mV (specifically, about - 3.85 ± 1 mV), and in the case of GST-E-PCN, the surface charge changed to about -8 to -1 mV (specifically, about -4.3 ± 2.5 mV), so that the surface charge of the GST-Afb-PCN complex It was confirmed that the charge had a similar value to the surface charge of GST-Afb (GST-HER2: about -5.34 ± 2 mV; GST-EGFR: about -3.8 ± 1 mV). Through this, it was found that the GST-Afb-PCN complex consisted of GST-Afb coated on the surface of the PCN.

Example 4. Confirmation of protein adsorption blocking effect of GST-Afb-MOFs complex

In the case of existing drug delivery systems using MOFs nanoparticles, there is a problem in that unnecessary proteins are adsorbed to MOFs nanoparticles in a biological environment, forming a protein corona, which disrupts the drug delivery function of MOFs nanoparticles. Therefore, in the case of the GST-Afb-MOFs complex obtained in Example 3, it was confirmed whether the above problem could be solved, that is, whether it had the ability to block protein adsorption in a biological environment.

Specifically, the PCN before being coated with the GST-Afb, the GST-Afb-PCN complex (GST-H-PCN) surface coated with the GST-HER2, and the GST-Afb- surface coated with the GST-EGFR. Each PCN complex (GST-E-PCN) was cultured in approximately 50% serum medium for approximately 1 hour at approximately 37°C, purified by centrifugation, and analyzed by gel electrophoresis.

As a result, as shown in Figure 7, a strong serum protein signal was detected in the case of PCN before coating with GST-Afb, and the GST-Afb-PCN complex (GST-H-PCN and GST-E-PCN) In this case, it was confirmed that a weak serum protein signal decreased by about 9 times compared to the PCN was detected (Figure 7b).

In addition, to further clarify the protein adsorption blocking ability of the GST-Afb-PCN complex, each of the PCN, GST-H-PCN, and GST-E-PCN was incubated with FITC modified serum albumin for about 8 hours. After this, the fluorescence emission from the generated particles was measured.

As a result, as shown in Figure 7, a strong fluorescence signal was detected in the case of PCN before coating with GST-Afb, and in the case of the GST-Afb-PCN complex (GST-H-PCN and GST-E-PCN) , it was confirmed that a significantly reduced fluorescence signal was detected compared to the PCN (Figure 7c).

Through this example, in the case of PCN before being coated with GST-Afb, many serum proteins were adsorbed to the surface of the PCN, whereas the GST-Afb-PCN complex (GST-H-PCN and GST-E-PCN) In the case of , it was confirmed that the adsorption of serum proteins was significantly reduced. Through this, it was confirmed that the GST-Afb-MOFs complex exhibits a significantly excellent protein adsorption blocking ability in a biological environment and can block the formation of a protein corona. As a result, it was confirmed that the GST-Afb-MOFs complex can be absorbed into target cells with high efficiency and exhibit excellent drug delivery effects, and as a result, the problems of existing technology can be resolved.

Example 5. Prediction of GST-Afb binding (or adsorption) site on the surface of MOFs nanoparticles of GST-Afb-MOFs complex

In the GST-Afb-MOFs complex obtained in Example 3, the connection (or adsorption) structure of MOFs nanoparticles and GST-Afb was theoretically predicted using molecular dynamics (MD) simulation.

As a result, as shown in Figure 8, various regions of GST-Afb (G-site, Side, Terminal, and Hinge of GST-Afb) , and Afb regions) (a in Figure 8) and two regions (Frame and Pore) (b) on the surface of PCN, which is a MOFs nanoparticle, were confirmed to be connected, and GST-Afb In the -PCN complex, five likely linking structures were predicted as follows (Figure 8c):

1. Linkage of the G-site of GST and the surface pore of PCN

2. Association of the G-site of GST with the surface frame of PCN

3. Connection of the lateral regions of GST with the surface frame of PCN

4. Connection of the hinge region of GST-Afb with the surface frame of PCN

5. Connection of the Afb region with the superficial frame of the PCN.

In addition, as a result of analysis using the Radial Distribution Function (RDF), as shown in Figure 9, in the case of the GST-Afb-PCN complex, Zr ₆ mainly exposed to the G-site of GST and the surface of PCN. It was confirmed that the strongest electrostatic interaction occurred between nodes, and that PCN and GST-Afb had a connected structure. In particular, the carboxylate group of glutamic acid (GLU) at the G-site of GST was confirmed to be located close to the surface Zr ₆ node of PCN through electrostatic interaction. In addition, the strongest electrostatic interaction between the G-site of GST and the Zr ₆ node exposed on the surface of PCN is due to the polar amino acid residues (SER, THR, ASN, GLN, GLY, and PRO) of GST and the negative charge. It was predicted to be due to amino acid residues (GLU and ASP) (Figure 10). In addition, the interaction energy between the G-site of GST and the surface pore of PCN was found to be larger than that between the G-site of GST and the surface frame of PCN or the lateral region of GST and the surface pore of PCN ( 11), the total interaction energy between the hinge region or Afb region of GST-Afb and the PCN surface was confirmed to be significantly lower than the total interaction energy between GST and the PCN surface (FIG. 12).

Through this example, it was confirmed that in the case of the GST-Afb-MOFs complex, the GST-Afb fusion protein is linked to the surface of the MOFs nanoparticle through GST, and Afb has a structure exposed to the outside. In particular, in the case of the GST-Afb-MOFs complex, it was confirmed that the G-site of GST interacted with the surface pore area of the MOFs nanoparticle, resulting in a structure in which GST-Afb and MOFs were connected. More specifically, in the case of the GST-Afb-MOFs complex, the electrostatic interaction occurs between the G-site of GST and the metal node exposed on the surface of the MOFs nanoparticle (e.g., Zr ₆ node in the case of PCN). , it was confirmed that GST-Afb and MOFs had a connected structure.

Through these results, it was found that the GST-Afb-MOFs complex was well organized in a specific direction with the GST-Afb fusion protein on the surface of the MOFs nanoparticle. Therefore, it was found that the GST-Afb-MOFs complex can be sufficiently protected from the adsorption of unnecessary proteins such as serum proteins in a biological environment, and thus, excellent targeting and drug delivery capabilities can be well maintained.

Example 6. Confirmation of cell targeting effect of GST-Afb-MOFs complex

The cell targeting efficiency of the GST-Afb-MOFs complex obtained in Example 3 was analyzed.

Specifically, the GST-Afb-PCN complex obtained in Example 3, GST-H-PCN (a fusion protein of Afb and GST targeting the HER2 receptor (GST-HER2) is coated on the surface of PCN) and GST -E-PCN (a fusion protein of Afb and GST targeting the EGFR receptor (GST-EGFR) is coated on the surface of the PCN) was prepared by loading DiI dye (loading capacity = ~ 20 wt%). Additionally, human breast cancer cells, SK-BR-3 or MDA-MB-468 (overexpressing HER2 and EGFR receptors, respectively) cells were seeded at approximately 4 × 10 per well in an 8-well chambered cover-glass (Lab Tek II, Thermo Scientific). They were seeded at a density of ⁴ cells and cultured for about 24 hours in 10% FBS conditions. Afterwards, each of GST-H-PCN and GST-E-PCN loaded with the dye was cultured with the cultured SK-BR-3 or MDA-MB-468 cells in 50% FBS for about 3 hours. Afterwards, confocal fluorescence imaging of the cultured cells was performed using a multiphoton LSM780 confocal microscope to detect the intracellular uptake of GST-H-PCN and GST-E-PCN.

As a result, as shown in Figure 13, strong DiI fluorescence was detected in SK-BR-3 cells cultured with GST-H-PCN, while MDA-MB-468 cells cultured with GST-H-PCN It was confirmed that no fluorescence was detected (top of Figure 13). In addition, strong DiI fluorescence was detected in MDA-MB-468 cells cultured with GST-E-PCN, whereas no fluorescence was detected in SK-BR-3 cells cultured with GST-E-PCN. (bottom of Figure 13). Through this, it was confirmed that the GST-Afb-PCN complex can only target cells targeted by Afb, thereby enabling target cell-specific intracellular uptake.

In addition, DiI dye was loaded onto GST-E/H-PCN, a GST-Afb-PCN complex prepared by coating both GST-HER2 and GST-EGFR simultaneously on the surface of PCN, and the dye-loaded GST-E After culturing /H-PCN with SK-BR-3 or MDA-MB-468 cells, respectively, confocal fluorescence imaging was performed on the cultured cells to detect intracellular uptake of GST-E/H-PCN.

As a result, as shown in Figure 14, it was confirmed that DiI fluorescence was detected in both SK-BR-3 and MDA-MB-468 cells. Through this, it was confirmed that the GST-Afb-PCN complex is capable of targeting multiple cells.

In addition, for flow cytometry measurements, SK-BR-3 or MDA-MB-468 cells were seeded in a 6-well plate at a density of about 2x10 ⁵ cells per well and cultured for about 24 hours, and then the cultured cells were loaded with DiI dye. GST-H-PCN and GST-E-PCN were each treated and further cultured for about 3 hours. After incubation, cells were trypsinized, collected, and analyzed using a BD FACSVerse flow cytometer (BD Biosciences, USA).

As a result, as shown in Figure 15, it was confirmed that stronger red DiI fluorescence was detected in cells treated with DiI-loaded GST-H-PCN or GST-E-PCN compared to the control group (cells treated with DiI-loaded PCN). did. In particular, in the case of the control group, red DiI fluorescence was detected at a significantly low level, which was confirmed to be the same level as measured in pure cells without DiI and particle treatment. This means that, in the case of PCN not coated with GST-Afb, almost no cellular uptake occurred, unlike GST-H-PCN and GST-E-PCN coated with GST-Afb.

Through this example, it was confirmed that the GST-Afb-MOFs complex exhibits excellent stability and cell specificity for cellular uptake. Specifically, the GST-Afb-MOFs complex blocks the adsorption of unnecessary proteins in the biological environment by surface-coating GST-Afb, which not only makes it easier to absorb into cells compared to existing MOFs nanoparticles, but also allows Afb to target only the cells. It was confirmed that target cell-specific intracellular absorption was possible.

Example 7. Confirmation of photoreactive ROS generation ability of GST-Afb-MOFs complex

When the cells into which the GST-Afb-MOFs complex obtained in Example 3 is absorbed are irradiated with light, reactive oxygen species (reactive oxygen species: ROS) was confirmed to be generated.

Specifically, MDA-MB-468 cells were inoculated into Lab Tek II slide chambers at a density of about ² A PCN complex, GST-E-PCN (a fusion protein of Afb and GST targeting the EGFR receptor (GST-EGFR) coated on the surface of PCN) was treated and cultured for about 2 hours. Afterwards, the cell culture medium was replaced and light irradiation (light density = 40 mW·cm ^-2 ) was performed for about 10 minutes using a solar simulator, and then additional culture was performed for about 2 hours. Afterwards, the cultured cells were washed with PBS, stained with DHE (dihydroethidium), a ROS indicator dye, and imaged using a Carl Zeiss LSM 780 NLO multiphoton microscope. As a control, PCN before coating with GST-Afb was used.

As a result, as shown in Figure 16, MDA-MB-468 cells cultured with PCN before being coated with GST-Afb and then irradiated with light, and MDA-MB cultured with GST-E-PCN but not irradiated with light. In the case of -468 cells, there was no staining by DHE at all, whereas in the case of MDA-MB-468 cells cultured with GST-E-PCN and then irradiated with light, it was confirmed that they were stained by DHE and emit fluorescence. Through this, PCN before being coated with GST-Afb hardly experienced intracellular uptake, whereas GST-E-PCN was absorbed into the target cell, MDA-MB-468 cells, and then worked within the cells by light irradiation. It was found that ROS such as singlet oxygen were generated.

Through this example, it was confirmed that the GST-Afb-MOFs complex exhibits the ability to generate photoreactive ROS within the target cells after being absorbed into the target cells with excellent efficiency. As a result, the GST-Afb-MOFs complex was used for photodynamic therapy (photodynamic therapy). It was confirmed that it can be used for therapy (PDT).

Example 8. Confirmation of target cell-specific drug delivery and cell death effects of GST-Afb-MOFs complex

The synergistic effect of PDT and chemotherapy on cancer cell death shown by the drug-encapsulated GST-Afb-MOFs complex was confirmed.

Specifically, before coating the surface with GST-Afb, the antitumor drug camptothecin (CPT) was encapsulated inside the PCN obtained in Example 1 (loading capacity = ~ 88 wt%). Specifically, about 5 mg of CPT and about 5 mg of PCN obtained in Example 1 were mixed in about 1 mL of DMSO solution at room temperature for about 48 hours. After mixing, the mixture was centrifuged at about 13000 rpm for about 5 minutes to separate the PCN particles encapsulated with the drug (CPT). Afterwards, GST-Afb fusion protein was coated on the surface of the PCN encapsulated with the drug in the same manner as described in Example 3, to obtain a GST-Afb-PCN complex with the drug encapsulated inside. Afterwards, SK-BR-3 or MDA-MB-468 cells were seeded in a 96-well plate at a density of about ⁵ The cultured cells were treated with PCN complex or GST-Afb-PCN complex without drug encapsulated at different concentrations (about 0.5 μg/ml, about 1.0 μg/ml, about 1.5 μg/ml, about 2.0 μg). /ml), and cultured for about 24 hours. Afterwards, the medium was replaced, light irradiation (light density = 40 mW·cm ^-2 ) was performed for about 10 minutes using a solar simulator, and additional culture was performed for about 24 hours. Next, the cultured cells were treated with 3(4,5-dimethyl-thyzoyl-2-yl)2,5 diphenyltetrazolium bromide (MTT), and cell viability was measured using an ELISA plate reader (λ = 570 nm, n = 3). The experimental groups used in this experiment were as follows:

1) Measurement of survival rate of SK-BR-3 cells

i) GST-H-PCN: Only GST-H-PCN, a GST-Afb-PCN complex coated on the surface of the PCN with a fusion protein of Afb and GST (GST-HER2) targeting the HER2 receptor, was treated (drug encapsulation and light treatment) no survey)

ii) CPT@GST-H-PCN: Process only GST-H-PCN with CPT drug encapsulated inside (no light irradiation)

iii) Light@GST-H-PCN: Light irradiation performed after GST-H-PCN treatment (no drug encapsulation)

iv) Light/CPT@GST-H-PCN: Light irradiation after treatment of GST-H-PCN with CPT drug encapsulated inside

2) Measurement of survival rate of MDA-MB-468 cells

i) GST-E-PCN: Process only GST-E-PCN, which is a GST-Afb-PCN complex coated on the surface of PCN with a fusion protein of Afb and GST (GST-EGFR) targeting the EGFR receptor (drug encapsulation and light treatment) no survey)

ii) CPT@GST-E-PCN: Process only GST-E-PCN with CPT drug encapsulated inside (no light irradiation)

iii) Light@GST-E-PCN: Light irradiation performed after GST-E-PCN treatment (no drug encapsulation)

iv) Light/CPT@GST-E-PCN: Light irradiation after treatment of GST-E-PCN with CPT drug encapsulated inside

As a result, as shown in Figure 17, it was confirmed that when GST-H-PCN or GST-E-PCN with no drug encapsulated inside was treated with cells without light irradiation, the cell viability was maintained at about 90% or more. Through this, it was confirmed that GST-H-PCN and GST-E-PCN themselves are safe particles that do not cause cytotoxicity. In addition, when GST-H-PCN or GST-E-PCN with the CPT drug encapsulated inside is treated with cells without light irradiation, and when GST-H-PCN or GST-E-PCN without the drug encapsulated inside the cells is treated with In all cases of light irradiation after treatment, the cancer cell death rate increased as the treatment concentration increased, and when the treatment concentration was about 2.0 ㎍/ml, it was confirmed that more than 25% of the cancer cells were killed. Through this, GST-H-PCN and GST-E-PCN exhibit target cancer cell-specific intracellular drug delivery and release effects, and even in the absence of drug encapsulation, GST-H-PCN exhibits photoreactive cytotoxicity within target cancer cells. -PCN and GST-E-PCN themselves were confirmed to have a cancer cell killing effect. In addition, when cells were treated with GST-H-PCN or GST-E-PCN with CPT drug encapsulated inside and then irradiated with light, the cancer cell death rate increased as the treatment concentration increased, and the treatment concentration was about 1.0 μg. In the case of /ml or more, it was confirmed that at least about 60% of cancer cells were killed, showing a significantly excellent cancer cell killing effect. Through this, the drug-encapsulated GST-H-PCN and GST-E-PCN simultaneously exhibit excellent drug delivery and release effects specific to target cancer cells and cytotoxic effects by light irradiation, creating a synergistic effect on cancer cell death. It was found that it could be induced.

Through this example, as shown in Figure 18, the GST-Afb-MOFs complex is absorbed into target cells with high efficiency by GST-Afb coated on the surface, and delivers and releases the drug encapsulated inside into the target cells. It was confirmed that the photosensitive porphyrin group present in MOFs can cause photoreactive cytotoxicity within target cells and kill them. Therefore, it was found that the GST-Afb-MOFs complex functions as a target cell-specific drug carrier and at the same time can itself function as a target cell-specific photoreactive disease treatment (eg, cancer treatment). In addition, it was found that the drug-encapsulated GST-Afb-MOFs complex can induce a synergistic effect of PDT and chemotherapy on cancer cell death.

Example 9. Confirmation of in vivo tumor cell targeting effect of GST-Afb-MOFs complex through animal model experiments

The in vivo tumor cell targeting effect of the GST-Afb-MOFs complex obtained in Example 3 was confirmed through animal model experiments.

Specifically, the GST-Afb-PCN complex obtained in Example 3, GST-H-PCN (a fusion protein of Afb and GST targeting the HER2 receptor (GST-HER2) coated on the surface of the PCN), was exposed to far-infrared rays. GST-H-PCN loaded with DiD dye was prepared by loading fluorescent DiD dye (DiD loading capacity = ~ 15 wt%), and about 20 μg of GST-H-PCN loaded with DiD dye was added to about 100 μL. Particle samples were prepared by dispersing in neutral PBS. Additionally, a tumor xenograft model was created by injecting SK-BR-3 cells into the right flank of balb/c nude female mice (Orient bio, Korea), and tumors developed to a size of ~150 mm ³ in the mice. was confirmed. The tumor-bearing mice (n = 3) were intravenously injected with particle samples containing GST-H-PCN loaded with the DiD dye and incubated at various time points (0, 2, 4, 8, 24 hours after injection). Fluorescence imaging was performed using an in vivo optical imaging system (Bruker Xtreme model) (λex = 630 nm, λem = 700 nm). Additionally, the biodistribution of each sample was assessed by ex vivo imaging of organs and tumors extracted from mice approximately 24 hours after intravenous injection. As control samples, PCN loaded with DiD dye and DiD dye itself were used, and all animal studies were performed according to protocols approved by the UNIST Animal Care and Use Committee.

As a result, as shown in Figures 19 and 20, GST-H-PCN is a MOFs nanoparticle coated on the surface of PCN with a fusion protein of Afb and GST (GST-HER2) targeting the HER2 receptor overexpressed in tumor cells. , MOFs nanoparticles designed to target cells expressing the HER2 receptor, that is, tumor cells. In fact, GST-H-PCN was confirmed to target tumor tissue with significantly high efficiency in vivo in tumor mice; In the case of PCN whose surface was not coated with GST-HER2, it was confirmed that it hardly targeted tumor tissue in vivo in tumor mice.

Through this example, the GST-Afb-MOFs complex surface-coated with GST-Afb, which blocks protein adsorption and targets tumor cells, specifically targets tumor tissue with high efficiency in vivo by GST-Afb. Therefore, it was found that it can be applied as a drug carrier or therapeutic agent for cancer treatment.

Sequence list electronic file attached

Claims (14)

Metal-organic framework (MOFs) nanoparticles coated on the surface with a fusion protein linked to glutathione S-transferase (GST) and disease cell targeting peptide. The metal-organic framework nanoparticle according to claim 1, wherein the metal-organic framework has a porous three-dimensional structure in which metal nodes are connected between organic molecules. The method of claim 2, wherein, in the fusion protein, a portion of the glutathione transferase that is not connected to the disease cell targeting peptide is connected to the metal node exposed on the surface of the metal-organic framework. Metal-organic framework nanoparticles. The metal-organic framework nanoparticle according to claim 3, wherein the G-site site of the glutathione transferase is connected to a metal node exposed on the surface of the metal-organic framework. . The metal-organic framework nanoparticle of claim 1, wherein the diseased cell is a cancer cell. The metal-organic framework nanoparticle according to claim 1, wherein the disease cell targeting peptide targets HER2 (human epidermal growth factor receptor 2) or EGFR (epidermal growth factor receptor) on disease cells. The metal-organic framework nanoparticle of claim 1, wherein the disease cell targeting peptide comprises the amino acid sequence of SEQ ID NO: 2 or SEQ ID NO: 3. The metal-organic framework nanoparticle according to claim 1, wherein the fusion protein comprises the amino acid sequence of SEQ ID NO: 4 or SEQ ID NO: 5. The metal-organic framework nanoparticle according to claim 2, wherein the organic molecule is a photosensitizer that generates reactive oxygen species (ROS) in response to light. The method of claim 9, wherein the metal-organic framework nanoparticles are specifically absorbed into the diseased cells and produce active oxygen in response to light within the cells, thereby specifically killing only the diseased cells. Organic framework nanoparticles. The metal-organic framework nanoparticle according to claim 1, wherein a drug is supported inside the metal-organic framework. The metal-organic framework nanoparticle according to claim 11, wherein the metal-organic framework nanoparticle is specifically absorbed into the diseased cell and releases the drug contained within the cell. A composition for target cell-specific drug delivery, comprising the metal-organic framework nanoparticle of any one of claims 1 to 12 as an active ingredient. A pharmaceutical composition for preventing or treating cancer, comprising the metal-organic framework nanoparticle of any one of claims 1 to 12 as an active ingredient.