Fusion protein comprising apolipoprotein and fc region-binding peptide and uses thereof

Abstract

The present invention relates to a fusion protein and a use thereof, wherein the fusion protein comprises: an apolipoprotein or a fragment thereof; and an Fc-binding peptide (Fc BP) or a fragment thereof. According to one aspect, a lipid carrier comprising the fusion protein was found to exhibit remarkably improved targeting efficiency, and thus can be used to deliver an active substance to a desired target.

Description

Fusion protein comprising apolipoprotein and FC region binding peptide and use thereof

[0001] The present invention relates to a fusion protein comprising an apolipoprotein or a fragment thereof; and an Fc region binding peptide or a fragment thereof, and to uses thereof.

[0002] As most people know, developing a single drug requires significant investment and time. Therefore, various methods are being studied to improve the safety and efficacy of existing drugs, building on existing ones. One such area is drug delivery system (DDS) research. DDS aims to efficiently deliver drugs to their target, minimizing side effects and maximizing efficacy.

[0003] Drug delivery can be broadly divided into two concepts. The first is the concept of a prodrug. A prodrug is a drug that has no effect outside the body but becomes effective upon entry into the body after being metabolized by metabolic enzymes. In other words, this involves altering the chemical structure of the drug to make it effective at its target. The second concept involves directly delivering the drug to the target molecule using a drug delivery system. This can be further divided into polymer-based drug delivery systems and drug delivery device systems utilizing fusion technologies.

[0004] Drug delivery systems can be classified by route of administration, type of delivery technology, and type of drug. Generally, they are classified by route of administration into oral, injection, pulmonary inhalation, transdermal, and implantable types. They are further classified by delivery technology into enhanced absorption, sustained-release, targeted, and intelligent DDS. Microparticles or microspheres can be used as carriers for drug delivery in the above-mentioned drug delivery systems. These drug delivery systems efficiently deliver disease-treating drugs to the treatment site, thereby reducing side effects, increasing patient compliance, and maximizing drug efficacy. It is important to design the formulation to maximize the effectiveness and efficacy of the drug.

[0005] Accordingly, the inventors of the present invention developed a delivery system capable of effectively delivering a drug to a target site, thereby completing the present invention.

[0006] One aspect provides a fusion protein comprising 1) an apolipoprotein or a fragment thereof; and 2) an Fc binding peptide (FcBP) or a fragment thereof.

[0007] Another aspect provides a polynucleotide comprising a sequence encoding the fusion protein.

[0008] Another aspect provides an expression vector comprising the polynucleotide.

[0009] Another aspect provides a targeting delivery system comprising the fusion protein, targeting and lipid-based delivery systems.

[0010] Another aspect provides a composition for drug delivery comprising the targeting carrier and a drug of interest.

[0011] Another aspect provides a pharmaceutical composition comprising the targeting delivery system and a drug of interest.

[0012] Another aspect provides an Fc binding peptide (FcBP) comprising one or more amino acid sequences selected from the group consisting of SEQ ID NOs: 12 to 14.

[0013] One aspect is to provide a fusion protein comprising 1) an apolipoprotein or a fragment thereof; and 2) an Fc binding peptide (FcBP) or a fragment thereof.

[0014] The above apolipoprotein may include at least one selected from the group consisting of apolipoprotein A (ApoA protein), apolipoprotein B (ApoB protein), apolipoprotein C (ApoC protein), apolipoprotein D (ApoD protein), apolipoprotein E (ApoE protein), apolipoprotein F (ApoF protein), apolipoprotein H (ApoH protein), apolipoprotein L (ApoL protein), apolipoprotein M (ApoM protein), and apolipoprotein(a) (Apo(a) protein).

[0015] The above apolipoprotein or fragment thereof may comprise an ApoE-derived peptide.

[0016] The above ApoA protein may include at least one selected from the group consisting of Apo-AI, Apo-A2, Apo-A4, and Apo-A5.

[0017] The above ApoB protein may include at least one selected from the group consisting of Apo-B48 and Apo B-100.

[0018] The above ApoC protein may include one or more selected from the group consisting of ApoC-I, ApoC-II, ApoC-III, and ApoC-IV.

[0019] The above ApoE protein may include at least one selected from the group consisting of ApoE1, ApoE2, ApoE3, ApoE4, ApoE5, ApoE6, ApoE7, ApoE8, ApoE9, ApoE10 and ApoE11, and may be specifically ApoE3 or ApoE4.

[0020] The apolipoprotein or a fragment thereof may include a lipid-binding region of the apolipoprotein. The lipid-binding region may refer to the 244th to 272nd amino acids from the N-terminus of the apolipoprotein protein, specifically, the 244th to 272nd amino acids from the N-terminus of the ApoE protein, and more specifically, may be a peptide represented by the 244th to 272nd amino acid sequence of SEQ ID NO: 1 or SEQ ID NO: 2.

[0021] The fragment of the above apolipoprotein may have a deleted receptor-binding region. The receptor-binding region may refer to the 136th to 150th amino acids from the N-terminus of the apolipoprotein, specifically, the 136th to 150th amino acids from the N-terminus of the ApoE protein, and more specifically, may be a peptide represented by the 136th to 150th amino acid sequence of SEQ ID NO: 1 or SEQ ID NO: 2.

[0022] The fragment of the above apolipoprotein may be one in which the receptor-binding region and the hinge region are deleted. The hinge region may refer to the 168th to 205th amino acids from the N-terminus of the apolipoprotein, specifically, the 168th to 205th amino acids from the N-terminus of the ApoE protein, and more specifically, may be a peptide represented by the 168th to 205th amino acid sequence of SEQ ID NO: 1 or SEQ ID NO: 2.

[0023] The fragment of the above apolipoprotein may be a fragment of the apolipoprotein in which a portion of the apolipoprotein is deleted, and may be a fragment in which 100 to 150 amino acids are sequentially deleted from the N-terminus of the apolipoprotein. Specifically, it may include a polypeptide expressed by an amino acid sequence in which 100 to 150, 110 to 150, 120 to 150, 130 to 150, 140 to 150, or 145 to 150 amino acids are deleted from the N-terminus.

[0024] The fragment of the above apolipoprotein is a fragment of the apolipoprotein in which a part of the apolipoprotein is deleted, and may be a fragment in which 151 to 220 amino acids are sequentially deleted from the N-terminus of the apolipoprotein, specifically, 151 to 220, 160 to 220, 170 to 220, 180 to 220, 190 to 220, 195 to 220, 200 to 220, 205 to 220, 210 to 220, 215 to 220, 151 to 215, 160 to 215, 170 to 215, 180 to 215, 190 to It may comprise a polypeptide represented by an amino acid sequence in which 215, 195 to 215, 200 to 215, 205 to 215, 151 to 210, 160 to 210, 170 to 210, 180 to 210, 190 to 210, 195 to 210, 200 to 210, 205 to 210, 151 to 205, 160 to 205, 170 to 205, 180 to 205, 190 to 205, 195 to 205, or 200 to 205 amino acids are deleted.

[0025] In one embodiment, the apolipoprotein fragment may comprise a polypeptide represented by an amino acid sequence in which 150 amino acids are sequentially deleted from the N-terminus of the ApoE protein, or a polypeptide represented by an amino acid sequence in which 205 amino acids are deleted.

[0026] In one embodiment, the apolipoprotein may include a polypeptide represented by the amino acid sequence of SEQ ID NO: 1, and specifically may be composed of a polypeptide represented by the amino acid sequence of SEQ ID NO: 1. In addition, any amino acid sequence that exhibits a homology of 80% or more, specifically 90% or more, more specifically 95% or more, even more specifically 98% or more, and most specifically 99% or more with the above sequence, and which is substantially the same as or exhibits a corresponding effect to the apolipoprotein, is included without limitation.

[0027] In one embodiment, the apolipoprotein may include a polypeptide represented by the amino acid sequence of SEQ ID NO: 2, and specifically may be composed of a polypeptide represented by the amino acid sequence of SEQ ID NO: 2. In addition, any amino acid sequence that exhibits a homology of 80% or more, specifically 90% or more, more specifically 95% or more, even more specifically 98% or more, and most specifically 99% or more with the above sequence, and which is substantially the same as or exhibits a corresponding effect to the apolipoprotein, is included without limitation.

[0028] In one embodiment, the fragment of the apolipoprotein may include a polypeptide represented by an amino acid sequence in which 100 to 220 amino acids are deleted from the N-terminus of SEQ ID NO: 2, specifically, 100 to 150, 110 to 150, 120 to 150, 130 to 150, 140 to 150, 145 to 150, 151 to 220, 160 to 220, 170 to 220, 180 to 220, 190 to 220, 195 to 220, 200 to 220, 205 to 220, 210 to 220, 215 to 220, 151 to 215, 160 to 215, 170 to 215, 180 to 215, 190 to 215, 195 to 215, 200 to 215, 205 to 215, 151 to 210, 160 to 210, 170 to 210, 180 to 210, 190 to 210, 195 to 210, 200 to 210, 205 to 210, 151 to 205, 160 to 205, 170 to 205, 180 to It may comprise a polypeptide represented by an amino acid sequence having 205, 190 to 205, 195 to 205, or 200 to 205 amino acids deleted, and more specifically, it may comprise a polypeptide represented by an amino acid sequence having 150 amino acids sequentially deleted from the N-terminus of the amino acid sequence of SEQ ID NO: 2, or a polypeptide represented by an amino acid sequence having 205 amino acids sequentially deleted from the N-terminus.

[0029] In one embodiment, the fragment of the apolipoprotein may include a polypeptide represented by the amino acid sequence of SEQ ID NO: 3, and specifically may be composed of a polypeptide represented by the amino acid sequence of SEQ ID NO: 3. In addition, any amino acid sequence that exhibits a homology of 80% or more, specifically 90% or more, more specifically 95% or more, even more specifically 98% or more, and most specifically 99% or more with the sequence, and which is substantially the same as or exhibits a corresponding effect to the fragment of the apolipoprotein, is included without limitation.

[0030] In one embodiment, the fragment of the apolipoprotein may include a polypeptide represented by the amino acid sequence of SEQ ID NO: 4, and specifically may be composed of a polypeptide represented by the amino acid sequence of SEQ ID NO: 4. In addition, an amino acid sequence that exhibits a homology of 80% or more, specifically 90% or more, more specifically 95% or more, even more specifically 98% or more, and most specifically 99% or more with the above sequence, and an amino acid sequence of a protein that exhibits substantially the same or corresponding efficacy as the fragment of the apolipoprotein, is included without limitation.

[0031] The term "homology" as used herein refers to the degree of similarity between a base sequence or amino acid sequence encoding a protein. If the homology is sufficiently high, the expression product of the corresponding gene may have identical or similar activity. Furthermore, homology may be expressed as a percentage based on the degree of identity with a given amino acid sequence or base sequence. In this specification, a homologous sequence that has identical or similar activity to a given amino acid sequence or nucleotide sequence is expressed as "% homology." For example, the sequences can be compared using standard software that calculates parameters such as score, identity and similarity, specifically BLAST 2.0, or by hybridization experiments performed under defined stringent conditions, and the defined appropriate hybridization conditions are within the scope of the relevant technology and can be determined by 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).

[0032] In addition, the apolipoprotein or fragment thereof may have conservative substitutions in 1, 2, 3, 4, 5, 6, 7, 8, 9, 10 or more amino acids in the amino acid sequence selected from SEQ ID NOs: 1 to 4, but is not limited thereto.

[0033] The term "conservative substitution" as used herein refers to the replacement of one amino acid with another amino acid having similar structural and / or chemical properties. The apolipoprotein or fragment thereof may have, for example, one or more conservative substitutions while still retaining the biological activity of the native or unmutated signal peptide. Such amino acid substitutions may generally be based on similarities in the polarity, charge, solubility, hydrophobicity, hydrophilicity, and / or amphipathic nature of the residues. For example, positively charged (basic) amino acids include arginine, lysine, and histidine; negatively charged (acidic) amino acids include glutamic acid and aspartic acid; aromatic amino acids include phenylalanine, tryptophan, and tyrosine; and hydrophobic amino acids include alanine, valine, isoleucine, leucine, methionine, phenylalanine, tyrosine, and tryptophan. Additionally, amino acids can be classified into those with electrically charged side chains and those with uncharged side chains. Amino acids with charged side chains include aspartic acid, glutamic acid, lysine, arginine, and histidine, while amino acids with uncharged side chains can be further classified as nonpolar amino acids or polar amino acids. Nonpolar amino acids include glycine, alanine, valine, leucine, isoleucine, methionine, and proline; polar amino acids can include serine, threonine, cysteine, asparagine, and glutamine. Conservative substitutions with amino acids having similar properties as described above can be expected to exhibit the same or similar activities.

[0034] In one embodiment, the Fc region binding peptide or fragment thereof may comprise one or more amino acid sequences selected from the group consisting of SEQ ID NOs: 12 to 14, and specifically, the Fc region binding peptide or fragment thereof may be composed of one or more amino acid sequences selected from the group consisting of SEQ ID NOs: 12 to 14.

[0035] In one embodiment, the Fc region-binding peptide or fragment thereof may comprise a polypeptide represented by an amino acid sequence selected from the group consisting of SEQ ID NOs: 12 to 14, and specifically may be composed of a polypeptide represented by an amino acid sequence selected from the group consisting of SEQ ID NOs: 12 to 14. In addition, an amino acid sequence that exhibits a homology of 80% or more, specifically 90% or more, more specifically 95% or more, even more specifically 98% or more, and most specifically 99% or more with the above sequence, and an amino acid sequence of a protein that exhibits substantially the same or corresponding efficacy as the Fc region-binding peptide or fragment thereof, is included without limitation.

[0036] Additionally, the Fc region binding peptide or fragment thereof may have conservative substitutions in 1, 2, 3, 4, 5, 6, 7, 8, 9, 10 or more amino acids in the amino acid sequence selected from SEQ ID NOs: 12 to 14, but is not limited thereto.

[0037] In the above fusion protein, the apolipoprotein or fragment thereof and the Fc region-binding peptide or fragment thereof may be linked via a linker, or may be directly connected. The linker is not particularly limited as long as it exhibits the effect of maintaining or enhancing the function and / or activity of the protein to which it is linked. Specifically, the linker may be one or more amino acids or peptides.

[0038] In one embodiment, the linker may include a peptide represented by the amino acid sequence of SEQ ID NO: 11 (TSAEAAAKEAAAKEAAAKEAAAKALEAEAAAKEAAAKEAAAKEAAAKAGT), and specifically may be composed of a peptide represented by the amino acid sequence of SEQ ID NO: 11. In addition, an amino acid sequence that exhibits a homology of 80% or more, specifically 90% or more, more specifically 95% or more, even more specifically 98% or more, and most specifically 99% or more to the sequence, and an amino acid sequence of a protein that exhibits substantially the same or corresponding efficacy as the linker is included without limitation.

[0039] In the above fusion protein, the Fc region binding peptide or fragment thereof may be linked to the N-terminus or C-terminus of an apolipoprotein (specifically, an ApoE protein) or a fragment thereof, and may be specifically linked to the N-terminus of an ApoE protein or a fragment thereof.

[0040] In one embodiment, the fusion protein may comprise a polypeptide consisting of the following sequences (the left end refers to the N`-terminus and the right end refers to the C`-terminus).

[0041] 1) Fc region binding peptide (or fragment thereof) - apolipoprotein (or fragment thereof)

[0042] 2) Fc region binding peptide (or fragment thereof) - linker - apolipoprotein (or fragment thereof)

[0043] 3) FcBP(K1)(SEQ ID NO: 12)-ApoE3(SEQ ID NO: 1)

[0044] 4) FcBP(K1)(SEQ ID NO: 12)-ApoE4(SEQ ID NO: 2)

[0045] 5) FcBP(K1)(SEQ ID NO: 12)-ApoE(151-299)(SEQ ID NO: 3)

[0046] 6) FcBP(K1)(SEQ ID NO: 12)-ApoE(206-299)(SEQ ID NO: 4)

[0047] 7) FcBP(K1)(SEQ ID NO: 12)-Linker(SEQ ID NO: 11)-ApoE3(SEQ ID NO: 1)

[0048] 8) FcBP(K1)(SEQ ID NO: 12)-Linker(SEQ ID NO: 11)-ApoE4(SEQ ID NO: 2)

[0049] 9) FcBP(K1)(SEQ ID NO: 12)-Linker(SEQ ID NO: 11)-ApoE(151-299)(SEQ ID NO: 3)

[0050] 10) FcBP(K1)(SEQ ID NO: 12)-Linker(SEQ ID NO: 11)-ApoE(206-299)(SEQ ID NO: 4)

[0051] 11) FcBP(J4)(SEQ ID NO: 13)-ApoE3(SEQ ID NO: 1)

[0052] 12) FcBP(J4)(SEQ ID NO: 13)-ApoE4(SEQ ID NO: 2)

[0053] 13) FcBP(J4)(SEQ ID NO: 13)-ApoE(151-299)(SEQ ID NO: 3)

[0054] 14) FcBP(J4)(SEQ ID NO: 13)-ApoE(206-299)(SEQ ID NO: 4)

[0055] 15) FcBP(J4)(SEQ ID NO: 13)-Linker(SEQ ID NO: 11)-ApoE3(SEQ ID NO: 1)

[0056] 16) FcBP(J4)(SEQ ID NO: 13)-Linker(SEQ ID NO: 11)-ApoE4(SEQ ID NO: 2)

[0057] 17) FcBP(J4)(SEQ ID NO: 13)-Linker(SEQ ID NO: 11)-ApoE(151-299)(SEQ ID NO: 3)

[0058] 18) FcBP(J4)(SEQ ID NO: 13)-Linker(SEQ ID NO: 11)-ApoE(206-299)(SEQ ID NO: 4)

[0059] 19) FcBP (KJ5) (SEQ ID NO: 14)-ApoE3 (SEQ ID NO: 1)

[0060] 20) FcBP (KJ5) (SEQ ID NO: 14)-ApoE4 (SEQ ID NO: 2)

[0061] 21) FcBP(KJ5)(SEQ ID NO: 14)-ApoE(151-299)(SEQ ID NO: 3)

[0062] 22) FcBP(KJ5)(SEQ ID NO: 14)-ApoE(206-299)(SEQ ID NO: 4)

[0063] 23) FcBP (KJ5) (SEQ ID NO: 14) - Linker (SEQ ID NO: 11) - ApoE3 (SEQ ID NO: 1)

[0064] 24) FcBP (KJ5) (SEQ ID NO: 14) - Linker (SEQ ID NO: 11) - ApoE4 (SEQ ID NO: 2)

[0065] 25) FcBP (KJ5) (SEQ ID NO: 14) - Linker (SEQ ID NO: 11) - ApoE (151-299) (SEQ ID NO: 3)

[0066] 26) FcBP (KJ5) (SEQ ID NO: 14) - Linker (SEQ ID NO: 11) - ApoE (206-299) (SEQ ID NO: 4)

[0067] The fusion protein may have a protecting group attached to the N- or C-terminus of the fusion protein to obtain chemical stability, enhanced pharmacological properties (half-life, absorbability, potency, efficacy, etc.), altered specificity (e.g., broad biological activity spectrum), and reduced antigenicity. In one embodiment, the N-terminus of the fusion protein may be attached to any one protecting group selected from the group consisting of an acetyl group, a fluoreonylmethoxycarbonyl group, a formyl group, a palmitoyl group, a myristyl group, a stearyl group, a butoxycarbonyl group, an aryloxycarbonyl group, and polyethylene glycol (PEG); And / or the C-terminus of the fusion protein may be linked to any one protecting group selected from the group consisting of an amino group (-NH2), a tertiary alkyl group, and an azide (-NHNH2). In addition, the fusion protein may optionally additionally include a targeting sequence, a tag, a labeled residue, an amino acid sequence manufactured for a specific purpose to increase half-life or protein stability.

[0068] The term "stability" in this specification may mean not only in vivo stability, which protects the fusion protein from attack by in vivo protein cleavage enzymes, but also storage stability (e.g., room temperature storage stability).

[0069] In one embodiment, the fusion protein may be cross-linked with a targeting protein, and specifically, the Fc region binding peptide of the fusion protein may be cross-linked with the targeting protein.

[0070] The above targeting protein may include a protein or a fragment thereof that can bind to a protein (receptor, antigen, etc.) expressed on the surface of a target tissue or target cell.

[0071] In one embodiment, the targeting protein may be a protein that targets cancer tissue or cancer cells. Specifically, the targeting protein may comprise an antibody or a fragment thereof (antigen-binding fragment) capable of binding to a protein (antigen) expressed on the surface of cancer tissue or cancer cells. Furthermore, the targeting protein may comprise a ligand or a peptide derived from the ligand capable of binding to a receptor expressed on the surface of cancer tissue or cancer cells.

[0072] 일 구현예에 있어서, 상기 표적화 단백질은 PD-L1(Programmed Death-Ligand 1), PD-L2, MAGE-1(Melanoma-associated antigen 1), MAGE-2, MAGE-3, MAGE-12, BAGE(B melanoma antigen), GAGE(G Antigen), NY-ESO-1(New York esophageal squamous cell carcinoma 1), tyrosinase, TRP-1(Tyrosinase-related protein 1), TRP-2, gp100(glycoprotein 100), MART-1(Melanoma Antigen Recognized by T-cells 1), MCIR(melanocortin 1 receptor), Ig idotype, CDK4(Cyclin-dependent kinase 4), Caspase-B, beta-catenin, CLA(Conjugated Linoleic Acid), BCR / ABL(Breakpoint Cluster Region / Abelson Tyrosine Kinase), mutated p21 / ras, mutated p53, proteinase 3, WT1(Wilms Tumor 1), MUC-1(Mucin 1), Her2 / neu(human epidermal growth factor receptor 2), PAP(Prostatic Acid Phosphatase), PSA((prostate specific antigen), PSMA(Prostate-Specific Membrane Antigen), G250, HPV E6 / E7, EBV LMP2a(Epstein-Barr virus latent membrane protein 2), CEA(Carcinoembryonic Antigen), alpha-Fetoprotein, 5T4(Trophoblast glycoprotein), CD47(Cluster of Differentiation 47),It may comprise an antibody or antigen-binding fragment thereof capable of binding to one or more selected from the group consisting of CD70 and onco-trophoblast glycoprotein.

[0073] The term "antibody" in this specification refers to a general term for proteins that selectively act on antigens and participate in biological immunity, and the type thereof is not particularly limited. The heavy and light chains of the antibody have an antigen-binding site that recognizes an epitope that includes a variable region, and antigen specificity is expressed depending on the sequence change of the variable region. The variable region of the antigen-binding site is divided into a framework region (FR) with little variableness and a complementarity determining region (CDR) with great variableness, and both the heavy chain and the light chain have three CDR regions, divided into CDR1, 2, and 3, and four FR regions. The CDRs of each chain are typically named CDR1, CDR2, and CDR3 sequentially starting from the N-terminus, and are also identified by the chain on which the specific CDR is located.

[0074] As used herein, the term “complementarity determining region” refers to a region among the variable regions of an antibody that confers binding specificity to an antigen.

[0075] As used herein, the term “epitope” refers to a specific stereomolecular structure within an antigen molecule to which an antibody can specifically bind.

[0076] The above antibodies include monoclonal antibodies, multispecific antibodies, human antibodies, humanized antibodies, and chimeric antibodies (e.g., humanized murine antibodies). In addition, the antibodies may include diabodies, triabodies, and tetrabodies.

[0077] As used herein, the term "antibody" includes an "antigen-binding fragment" or "antibody fragment" of an antibody having antigen-binding ability. The antigen-binding fragment may be selected from the group consisting of antibody fragments including one or more complementarity-determining regions, for example, scFv, (scFv)2, scFv-Fc, Fab, Fab', and F(ab')2. Among antibody fragments, Fab has one antigen-binding site in a structure having variable regions of light and heavy chains, constant region of light chain, and first constant region (CH1) of heavy chain. Fab' differs from Fab in that it has a hinge region including one or more cysteine ​​residues at the C-terminus of the heavy chain CH1 domain. F(ab')2 antibody is generated when cysteine ​​residues in the hinge region of Fab' form a disulfide bond. Fv is the smallest antibody fragment having only heavy chain variable region and light chain variable region. In a two-chain Fv, the heavy chain variable region and the light chain variable region are non-covalently linked. In a single-chain Fv (single-chain Fv: scFv), the heavy chain variable region and the light chain variable region are covalently linked, usually via a peptide linker, or directly linked at the C-terminus, so that they can form a dimer-like structure like a two-chain Fv.

[0078] In one embodiment, the fusion protein or the Fc region binding peptide within the fusion protein may be mutually associated with the targeting protein (specifically, an antibody or an antigen-binding fragment thereof) to form a complex or conjugate, or may not be mutually associated but may be mixed with each other to form a non-covalent complex or conjugate. Specifically, the fusion protein-antibody complex / conjugate may be one in which the fusion protein and the targeting protein are chemically associated or physically associated, for example, covalently or non-covalently associated.

[0079] In one embodiment, the fusion protein may be intended to enhance delivery efficiency to a target cell or target tissue that expresses a protein (such as an antigen, receptor, or ligand) capable of binding to the targeting protein of the delivery system comprising the fusion protein.

[0080]

[0081] Another aspect is to provide a polynucleotide comprising a sequence encoding the fusion protein. The same parts as described above also apply to the polynucleotide.

[0082] The term "polynucleotide" as used herein refers to a polymer of deoxyribonucleotides or ribonucleotides, either single-stranded or double-stranded. It encompasses RNA genomic sequences, DNA (gDNA and cDNA), and RNA sequences transcribed therefrom, and unless otherwise specified, also includes analogs of natural polynucleotides.

[0083] The polynucleotide may be composed of DNA, RNA, or a modified form thereof, for example, cDNA, mRNA, or a modified form thereof. If the polynucleotide is DNA or a modified DNA, it may be composed of A (Adenine), T (Thymine), G (Guanine), C (Cytosine), and / or a modified form thereof, and if the polynucleotide is RNA or a modified RNA, it may be composed of A, U (Uridine), G, C, and / or a modified form thereof. For example, the modified DNA or RNA may be 5-methylcytidine (5mC), N6-methyladenosine (m6A), 3,2'-O-dimethyluridine (m4U), 2-thiouridine (s2U), 2' fluorouridine, pseudouridine, 2'-O-methyluridine (Um), 2' deoxyuridine (2' dU), 4-thiouridine (s4U), 5-methyluridine (m5U), 2'-O-methyladenosine (m6A), N6,2'-O-dimethyladenosine (m6Am), N6,N6,2'-O-trimethyladenosine (m62Am), 2'-O-methylcytidine (Cm), 7-methylguanosine (m7G), 2'-O-methylguanosine (Gm), It may include at least one selected from the group consisting of N2,7-dimethylguanosine (m-2,7G), N2,N2,7-trimethylguanosine (m-2,2,7G) and N1-methyl-pseudouridine.

[0084] Accordingly, when the polynucleotide is in the form of RNA, T of the polynucleotide sequence encoding the fusion protein in the present specification may be expressed by being replaced with U, and when the polynucleotide is in the form of modified DNA or modified RNA, A, T, G, C and U of the polynucleotide sequence encoding the fusion protein in the present specification may be expressed by being replaced with their modified forms, respectively.

[0085] The polynucleotide encoding the above fusion protein can have various modifications in the coding region within a range that does not change the amino acid sequence of the protein expressed from the coding region, taking into account the codon preferred in the organism that is to express the fusion protein due to the degeneracy of the codons. Therefore, the polynucleotide can be included without limitation as long as it is a polynucleotide sequence encoding the fusion protein. In addition, a probe that can be prepared from a known sequence, for example, a sequence that hybridizes under stringent conditions with a complementary sequence to all or part of the polynucleotide sequence, and encodes a protein having the activity of the fusion protein can be included without limitation.

[0086] The above "stringent conditions" refer to conditions that enable specific hybridization between polynucleotides. For example, conditions in which genes with high homology, for example, genes with a homology of 40% or more, specifically 90% or more, more specifically 95% or more, even more specifically 97% or more, and particularly specifically 99% or more, hybridize and genes with lower homology do not hybridize, or conditions in which washing is performed once, specifically twice or three times, at a salt concentration and temperature corresponding to the washing conditions of conventional Southern hybridization, such as 60°C, 1XSSC, 0.1% SDS, specifically 60°C, 0.1XSSC, 0.1% SDS, and more specifically 68°C, 0.1XSSC, 0.1% SDS.

[0087] Hybridization requires that two polynucleotides have complementary sequences, although mismatches between bases are possible depending on the stringency of hybridization. The term "complementary" is used to describe the relationship between nucleotide bases that can hybridize with each other. For example, in DNA, adenosine is complementary to thymine, and cytosine is complementary to guanine. Therefore, the present application may also encompass isolated polynucleotide fragments that are complementary in their entirety, as well as substantially similar polynucleotide sequences.

[0088] The sequence encoding the above fusion protein may include a sequence encoding an apolipoprotein or a fragment thereof, and a sequence encoding an Fc region binding peptide or a fragment thereof.

[0089] In one embodiment, the sequence encoding the Fc region-binding peptide or a fragment thereof may include a polynucleotide represented by a base sequence selected from the group consisting of SEQ ID NOs: 15 to 17, and specifically may be composed of a polynucleotide represented by a base sequence selected from the group consisting of SEQ ID NOs: 15 to 17. In addition, any amino acid sequence that exhibits a homology of 80% or more, specifically 90% or more, more specifically 95% or more, even more specifically 98% or more, and most specifically 99% or more with the sequence, and that encodes a protein that exhibits substantially the same or corresponding efficacy as the Fc region-binding peptide or a fragment thereof, is included without limitation.

[0090] In one embodiment, the sequence encoding the apolipoprotein may include a polynucleotide represented by the base sequence of SEQ ID NO: 5 or SEQ ID NO: 6, and specifically may be composed of a polynucleotide represented by the base sequence of SEQ ID NO: 5 or SEQ ID NO: 6. In addition, a base sequence showing a homology of 80% or more, specifically 90% or more, more specifically 95% or more, even more specifically 98% or more, and most specifically 99% or more with the sequence, and a coding sequence of a protein showing an effect substantially identical to or corresponding to the apolipoprotein is included without limitation.

[0091] In one embodiment, the sequence encoding the fragment of the apolipoprotein may include a polynucleotide represented by the base sequence of SEQ ID NO: 7 or SEQ ID NO: 8, and specifically may be composed of a polynucleotide represented by the base sequence of SEQ ID NO: 7 or SEQ ID NO: 8. In addition, a base sequence showing a homology of 80% or more, specifically 90% or more, more specifically 95% or more, even more specifically 98% or more, and most specifically 99% or more with the sequence, and a coding sequence of a protein showing an effect substantially identical to or corresponding to the fragment of the apolipoprotein is included without limitation.

[0092]

[0093] Another aspect is to provide an expression vector comprising the polynucleotide. The same parts described above also apply to the expression vector.

[0094] The term "expression vector" as used herein refers to a recombinant vector capable of expressing a target protein when introduced into a suitable host cell or living organism, and a genetic construct containing essential regulatory elements operably linked to enable expression of the gene insert. The term "operably linked" means that a nucleic acid expression regulatory sequence and a nucleic acid sequence encoding the target protein are functionally linked to perform a general function. The operably linked vector can be produced using genetic recombination techniques well known in the art, and site-specific DNA cleavage and ligation can be easily performed using enzymes generally known in the art.

[0095] Suitable expression vectors of the present invention may include signal sequences for membrane targeting or secretion, in addition to expression control elements such as a promoter, initiation codon, termination codon, polyadenylation signal, and enhancer. The initiation and termination codons are generally considered to be part of the nucleotide sequence encoding the protein of interest, and must be functional in a subject when the genetic construct is administered, and must be in frame with the coding sequence. Common promoters can be constitutive or inducible and include, but are not limited to, the lac, tac, T3 and T7 promoters in prokaryotes, and the simian virus 40 (SV40), mouse mammary tumor virus (MMTV) promoters, human immunodeficiency virus (HIV), e.g., the long terminal repeat (LTR) promoter of HIV, Moloney virus, cytomegalovirus (CMV), Epstein-Barr virus (EBV), Rous sarcoma virus (RSV) promoters, as well as the β-actin promoter, human hemoglobin, human muscle creatine and human metallothionein promoters in eukaryotes.

[0096] Additionally, the expression vector may include a selectable marker for selecting host cells containing the vector. The selectable marker is used to select cells transformed with the vector, and markers that confer selectable phenotypes such as drug resistance, nutrient requirements, cytotoxic agent resistance, or expression of surface proteins may be used. In an environment treated with a selective agent, only cells expressing the selectable marker survive, allowing the transformed cells to be selected. Furthermore, if the vector is a replicable expression vector, it may include a replication origin, which is a specific nucleic acid sequence where replication begins.

[0097] Recombinant expression vectors for inserting foreign genes can take many forms, including plasmids, viruses, and cosmids. The type of recombinant vector is not particularly limited, as long as it can express the desired gene and produce the desired protein in various host cells, both prokaryotic and eukaryotic. However, vectors that possess a highly active promoter and strong expression capacity, while also being capable of mass-producing foreign proteins in a form similar to that of the native form, are particularly useful.

[0098] To express the protein of the present invention, various combinations of hosts and vectors can be utilized. Expression vectors suitable for eukaryotic hosts include, but are not limited to, expression control sequences derived from SV40, bovine papillomavirus, adenovirus, adeno-associated virus, cytomegalovirus, and retrovirus. Expression vectors that can be used in bacterial hosts include, but are not limited to, bacterial plasmids obtained from Escherichia coli, including pET21a, pET, pRSET, pBluescript, pGEX2T, pUC vectors, col E1, pCR1, pBR322, pMB9 or derivatives thereof, plasmids having a wider host range such as RP4, phage DNA such as phage lambda derivatives such as λgt10, λgt11 or NM989, and other DNA phages such as M13 and filamentous single-stranded DNA phages. In yeast cells, a second-order plasmid or derivatives thereof can be used, and in insect cells, pVL941 can be used, etc.

[0099] The above cells, for example, eukaryotic cells, can be cells of yeast, fungi, protozoa, plants, higher plants and insects, or amphibians, or mammalian cells such as CHO, HeLa, HEK293, and COS-1, and can be, for example, cultured cells (in vitro), graft cells and primary cell cultures (in vitro and ex vivo), and in vivo cells, and also mammalian cells including humans, which are commonly used in the art. Furthermore, the organisms can be yeast, fungi, protozoa, plants, higher plants and insects, amphibians, or mammals.

[0100]

[0101] Another aspect provides a targeting composition or use comprising the fusion protein and the targeting protein. The same portions described above also apply to the composition or use.

[0102] In one embodiment, the targeting composition may comprise a complex or conjugate comprising the fusion protein and the targeting protein.

[0103] The targeting composition comprising the above fusion protein and targeting protein may be for targeting or improving targeting efficiency of a lipid-based carrier, and specifically may be for improving targeting or improving targeting efficiency of the carrier to a specific cell or tissue.

[0104] In one embodiment, the apolipoprotein (specifically, ApoE) or fragment thereof of the fusion protein may be linked to the lipid portion of the lipid-based transporter.

[0105]

[0106] Another aspect is to provide a targeting delivery system comprising the fusion protein, the targeting protein, and the lipid-based delivery system. The same principles as described above apply to the delivery system.

[0107] The lipid-based carrier may include at least one selected from the group consisting of a lipid nanoparticle (LNP), a liposome, a nano-structured lipid carrier (NLC), a lipid microemulsion (LME), a lipid nanoemulsion (LNE), a solid lipid nanoparticle (SLN), a self-emulsifying delivery system (SEDDS), a lipid sphere (Liposphere), a solid lipid microparticle (SLM), a lipid drug conjugate (LDC), an exosome, a niosome, a pharmacosome, a phytosome, an ethosome, and a herbosome.

[0108] The lipid-based carrier may comprise at least one selected from the group consisting of phospholipids, sterols, PEG (polyethylene glycol)-lipids, and ionizable lipids. In addition, the ionizable lipid may be a cationic lipid, and the sterol may be cholesterol.

[0109] In the above targeting carrier, the fusion protein may be linked or combined with a lipid-based carrier. Specifically, a region of the fusion protein comprising an apolipoprotein (specifically, an ApoE protein) or a fragment thereof may be linked or combined with a lipid constituting the lipid-based carrier.

[0110] The targeting protein or fragment thereof included in the above targeting carrier can bind to a cell or tissue to be targeted, and through the above mechanism, the targeting carrier of the present invention can be effectively delivered or moved to the cell or tissue to be targeted.

[0111] The above targeting delivery system may target cancer, and may specifically enhance delivery efficiency to cancer tissues or cancer cells.

[0112] The above targeting delivery vehicle may additionally comprise a drug of interest to be delivered. Accordingly, the targeting delivery vehicle may be a drug delivery vehicle.

[0113] The term "drug carrier" or "drug delivery system (DDS)" in this specification refers to a series of technologies or materials that control the delivery and release of pharmacologically active substances into cells, tissues, organs, and systems to achieve optimal efficacy by using various physicochemical technologies, and refers to a technology that optimizes drug treatment by designing a formulation to efficiently deliver the required amount of drugs while minimizing side effects of existing drugs and maximizing efficacy and effectiveness.

[0114]

[0115] Another aspect provides a drug delivery composition comprising the targeting carrier. The same principles as described above apply to the composition.

[0116] The above composition may additionally contain a desired drug.

[0117] The above composition can be delivered or transported to a target cell or tissue to which the desired drug is to be targeted using a targeting delivery vehicle.

[0118] The above drug is not particularly limited as long as it can exhibit pharmacological activity and disease treatment effect in vivo, in tissues and / or in vivo as an active ingredient, and may be a drug or a physiologically active substance in particular. For example, it may include one or more selected from the group consisting of proteins including enzymes, ligands, antibodies, etc., compounds, nucleic acids including siRNA, microRNA, mRNA, plasmids, genes, etc., aptamers, genetic material, fats, carbohydrates, dyes, photosensitizers, anticancer agents, antibiotics, chemical compounds, and combinations thereof, but may not be limited thereto.

[0119]

[0120] Another aspect provides a pharmaceutical composition comprising the targeting delivery system and the desired drug. The same principles described above apply to the composition.

[0121] The above-mentioned targeting delivery vehicle may be for delivering a desired drug. Accordingly, the pharmaceutical composition may be a composition for drug delivery.

[0122] The above-mentioned target drug may be a drug for treating, preventing or improving cancer, and specifically, may be an anticancer agent or an anticancer adjuvant agent.

[0123] The above pharmaceutical composition may be for treating or preventing cancer. The cancer may include at least one selected from the group consisting of liver cancer, lung cancer, pancreatic cancer, non-small cell lung cancer, colon cancer, colon cancer, bone cancer, skin cancer, head or neck cancer, cutaneous or intraocular melanoma, uterine cancer, ovarian cancer, rectal cancer, stomach cancer, anal cancer, breast cancer, fallopian tube carcinoma, endometrial carcinoma, cervical carcinoma, vaginal carcinoma, vulvar carcinoma, Hodgkin's disease, esophageal cancer, small intestine cancer, endocrine cancer, thyroid cancer, parathyroid cancer, adrenal cancer, soft tissue sarcoma, urethral cancer, penile cancer, prostate cancer, chronic or acute leukemia, lymphocytic lymphoma, bladder cancer, kidney or ureter cancer, renal cell carcinoma, renal pelvic carcinoma, central nervous system (CNS) tumor, primary central nervous system lymphoma, spinal cord tumor, brainstem glioma, and pituitary adenoma.

[0124] The pharmaceutical composition may include a pharmaceutically acceptable carrier. The term "pharmaceutically acceptable carrier" may refer to a carrier or diluent that does not stimulate a living organism and does not inhibit the biological activity or properties of the injected compound. Here, "pharmaceutically acceptable" means that the carrier does not inhibit the activity of the active ingredient and does not exhibit toxicity exceeding the adaptability of the subject of application (prescription). Any carrier commonly used in the art and pharmaceutically acceptable for the pharmaceutical composition may be used. Non-limiting examples of the above carriers include lactose, dextrose, maltodextrin, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, glycerol, ethanol, starch, acacia gum, alginate, gelatin, calcium phosphate, calcium silicate, cellulose, methyl cellulose, microcrystalline cellulose, polyvinyl pyrrolidone, water, saline solution, sterile water, Ringer's solution, buffered saline, albumin injection solution, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate, or mineral oil. These may be used alone or in combination of two or more. The pharmaceutical composition may be prepared as an oral formulation or a parenteral formulation according to the route of administration by a conventional method known in the art, including a pharmaceutically acceptable carrier in addition to the active ingredient. The above pharmaceutical composition can be formulated and used in the form of oral formulations such as powders, granules, tablets, capsules, suspensions, emulsions, syrups, aerosols, external preparations, suppositories, or sterile injection solutions, each according to a conventional method.

[0125] When formulating the above pharmaceutical composition, it may be prepared using a diluent or excipient such as a generally used filler, bulking agent, binder, wetting agent, disintegrant, or surfactant, but may not be limited thereto.

[0126] When the above pharmaceutical composition is manufactured into an oral dosage form, it can be manufactured into a dosage form such as powder, granules, tablets, pills, dragees, capsules, liquids, gels, syrups, suspensions, wafers, etc., using a suitable carrier according to a method known in the art. At this time, examples of suitable pharmaceutically acceptable carriers include sugars such as lactose, glucose, sucrose, dextrose, sorbitol, mannitol, and xylitol; starches such as corn starch, potato starch, and wheat starch; celluloses such as cellulose, methylcellulose, ethylcellulose, sodium carboxymethylcellulose, and hydroxypropylmethylcellulose; polyvinyl pyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, magnesium stearate, mineral oil, malt, gelatin, talc, polyols, and vegetable oils. In case of formulation, the formulation may include diluents and / or excipients such as fillers, bulking agents, binders, wetting agents, disintegrants, and surfactants, as needed.

[0127] When the above pharmaceutical composition is prepared as a parenteral dosage form, it can be formulated in the form of injections, transdermal administration, nasal inhalation, and suppositories using a suitable carrier according to a method known in the art. When formulated as an injection, suitable carriers include sterile water, ethanol, polyols such as glycerol or propylene glycol, or mixtures thereof, and preferably, Ringer's solution, phosphate buffered saline (PBS) containing triethanolamine, sterile water for injection, and isotonic solutions such as 5% dextrose can be used. When formulated as a transdermal dosage form, it can be formulated in the form of ointments, creams, lotions, gels, external solutions, pastes, liniments, aerosols, etc. In the case of nasal inhalation, it can be formulated in the form of an aerosol spray using a suitable propellant such as dichlorofluoromethane, trichlorofluoromethane, dichlorotetrafluoroethane, or carbon dioxide, and in the case of formulating it as a suppository, the base can be witepsol, tween 61, polyethylene glycol, cacao butter, laurin butter, polyoxyethylene sorbitan fatty acid ester, polyoxyethylene stearate, sorbitan fatty acid ester, etc.

[0128] The pharmaceutical composition may be administered in a pharmaceutically effective amount. The term "pharmaceutically effective amount" refers to an amount sufficient to treat or prevent a disease at a reasonable benefit / risk ratio applicable to medical treatment or prevention, and the effective dosage level may be determined based on the severity of the disease, the activity of the drug, the patient's age, weight, health, sex, the patient's sensitivity to the drug, the time of administration of the composition of the present invention used, the route of administration and the excretion rate, the treatment period, the drug used in combination with or concurrently with the composition of the present invention used, and other factors well known in the medical field. The pharmaceutical composition of the present invention may be administered alone or in combination with a component known to exhibit a therapeutic effect on a known disease. It is important to take all of the above factors into consideration and administer an amount that can achieve the maximum effect with the minimum amount without side effects.

[0129] The dosage of the pharmaceutical composition may be determined by a person skilled in the art in consideration of the intended use, the degree of toxicity of the disease, the patient's age, weight, sex, medical history, or the type of substance used as the active ingredient. For example, the pharmaceutical composition of the present invention may 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 disclosure is not particularly limited thereto, but may be administered once a day or administered in divided doses several times. The dosage or frequency of administration does not limit the scope of the present disclosure in any way.

[0130]

[0131] Another aspect provides a method for delivering a desired drug to a subject, comprising administering to the subject a targeting delivery system, drug delivery system, or drug delivery composition. The same aspects described above also apply to the composition.

[0132] The term "subject" in this specification may include, without limitation, mammals including dogs, cats, rats, livestock, humans, birds, reptiles, farmed fish, etc.

[0133] The above method may be for delivering a desired drug to cells, tissues, and / or organs expressing a ligand, receptor, and / or antigen capable of binding to the targeting protein of the present invention within a subject. Specifically, the method may be for delivering a desired drug to cancer cells and / or cancer tissues.

[0134]

[0135] Another aspect provides an Fc binding peptide (FcBP) comprising one or more amino acid sequences selected from the group consisting of SEQ ID NOs: 12 to 14. The same portions as described above also apply to the protein.

[0136] In one embodiment, the Fc region binding peptide may comprise one or more amino acid sequences selected from the group consisting of SEQ ID NOs: 12 to 14, and specifically, the Fc region binding peptide or fragment thereof may be composed of one or more amino acid sequences selected from the group consisting of SEQ ID NOs: 12 to 14.

[0137] In one embodiment, the Fc region-binding peptide may comprise a polypeptide represented by an amino acid sequence selected from the group consisting of SEQ ID NOs: 12 to 14, and specifically may be composed of a polypeptide represented by an amino acid sequence selected from the group consisting of SEQ ID NOs: 12 to 14. In addition, an amino acid sequence that exhibits a homology of at least 80%, specifically at least 90%, more specifically at least 95%, even more specifically at least 98%, and most specifically at least 99% with the above sequence, and is substantially the same as or equivalent to the amino acid sequence of a protein exhibiting an effect as the Fc region-binding peptide or a fragment thereof, is included without limitation.

[0138] Additionally, the Fc region binding peptide may be one in which conservative substitutions have occurred in 1, 2, 3, 4, 5, 6, 7, 8, 9, 10 or more amino acids in an amino acid sequence selected from SEQ ID NOs: 12 to 14, but is not limited thereto.

[0139] The peptide may have a protecting group bonded to the N- or C-terminus of the peptide to obtain chemical stability, enhanced pharmacological properties (half-life, absorbability, potency, efficacy, etc.), altered specificity (e.g., broad biological activity spectrum), and reduced antigenicity. In one embodiment, the N-terminus of the peptide may be bonded to any one protecting group selected from the group consisting of an acetyl group, a fluoreonylmethoxycarbonyl group, a formyl group, a palmitoyl group, a myristyl group, a stearyl group, a butoxycarbonyl group, an aryloxycarbonyl group, and polyethylene glycol (PEG); And / or the C-terminus of the peptide may be bonded to any one protecting group selected from the group consisting of an amino group (-NH2), a tertiary alkyl group, and an azide (-NHNH2). In addition, the peptide may optionally additionally include a targeting sequence, a tag, a labeled residue, an amino acid sequence manufactured for a specific purpose to increase half-life or protein stability.

[0140]

[0141] Another aspect provides a polynucleotide or expression vector comprising a sequence encoding the Fc region binding peptide (FcBP). The same portions as described above also apply to the polynucleotide or expression vector.

[0142] In one embodiment, the sequence encoding the Fc region-binding peptide may include a polynucleotide represented by a base sequence selected from the group consisting of SEQ ID NOs: 15 to 17, and specifically may be composed of a polynucleotide represented by a base sequence selected from the group consisting of SEQ ID NOs: 15 to 17. In addition, any amino acid sequence that exhibits a homology of 80% or more, specifically 90% or more, more specifically 95% or more, even more specifically 98% or more, and most specifically 99% or more with the sequence, and that encodes a protein that exhibits substantially the same or corresponding efficacy as the Fc region-binding peptide, is included without limitation.

[0143]

[0144] Another aspect is to provide a fusion protein comprising an Fc binding peptide (FcBP) or a fragment thereof. The same portions described above also apply to the protein.

[0145] The above fusion protein may additionally comprise an apolipoprotein or a fragment thereof.

[0146]

[0147] Another aspect provides a polynucleotide or expression vector comprising a sequence encoding a fusion protein comprising an Fc binding peptide (FcBP) or a fragment thereof. The same portions as described above also apply to the polynucleotide or expression vector.

[0148]

[0149] Another aspect provides a targeting delivery system comprising a fusion protein comprising the Fc binding peptide (FcBP) or a fragment thereof and a lipid-based delivery system. The same aspects described above also apply to the delivery system.

[0150]

[0151] Another aspect provides a drug delivery composition comprising a fusion protein comprising the Fc binding peptide (FcBP) or a fragment thereof, a lipid-based carrier, and a drug of interest. The same parts as described above also apply to the composition.

[0152] According to the fusion protein according to the aspect, it was confirmed that the targeting efficacy was significantly improved in the case of a lipid transporter including the fusion protein, and thus, the active ingredient can be effectively delivered to the desired target using this.

[0153] Figure 1 is a schematic diagram showing a lipid-based delivery system using the fusion protein of the present invention.

[0154] Figure 2 is a diagram showing the description of each region of the ApoE protein.

[0155] Figure 3 is a drawing showing the results of confirming the targeting delivery efficacy of lipid nanoparticles produced using a fusion protein including the ApoE protein of the present invention.

[0156] Figure 4 is a diagram showing the structure and domains of the ApoE4 protein.

[0157] Figures 5 to 7 are drawings showing the results of confirming the targeting delivery efficacy of lipid nanoparticles produced using a fusion protein including the ApoE protein fragment of the present invention.

[0158] Figures 8 and 9 are drawings showing the results of confirming the targeting delivery efficacy of lipid nanoparticles produced using a fusion protein including an ApoE protein fragment and FcBP of the present invention.

[0159] The present invention will be described in more detail through the following examples. However, these examples are provided for illustrative purposes only and the scope of the present invention is not limited to these examples.

[0160]

[0161] Example 1: Evaluation of the lipid-based transporter targeting efficacy of a fusion protein containing the ApoE protein.

[0162] The present invention relates to a fusion protein comprising an ApoE protein and an Fc binding peptide (FcBP), as an example of an apolipoprotein, and a targeting delivery system comprising the same. To confirm the targeting delivery efficiency of a lipid-based delivery system comprising the fusion protein of the present invention, the following experiments were performed.

[0163] Specifically, in order to determine whether the targeting efficiency of a lipid-based delivery system can be improved by using a fusion protein containing an ApoE protein, a fusion protein containing an ApoE protein, ApoE3 or ApoE4, and a targeting protein was first prepared. As an example, the targeting protein of the fusion protein was PD1 protein (SEQ ID NO: 9). In addition, the fusion protein was prepared so that it was linked in the order of ApoE-PD1 (targeting protein) or PD1 (targeting protein)-ApoE from the N-terminus. Meanwhile, the structure and domain of the ApoE protein are described in FIG. 2, and the amino acid sequence and base sequence of the ApoE3 and ApoE4 proteins are described in Tables 1 and 2, respectively (when the base sequence in Table 2 is a DNA sequence, 'U' is replaced with 'T').

[0164] 단백질아미노산 서열서열번호

[0165] 단백질염기 서열 (DNA 서열 또는 mRNA 서열)서열번호ApoE3AAA GUG GAG CAA GCU GUG GAA ACG GAG CCA GAG CCG ACUGAG CAUG CA UUU CAA AGG UGG GAA CUC GCC CUU GGU AGA UUU UGG GAU UAU UUG CGG UGG GUC CAA ACA CUC AGC GAG CAG GUA CAA GAG GAA CUC CUC UCC UCC CAG GUU ACA CAG GAG CUG AGG GAGG GCU UUG GAUG AUCAG AAA AGU GAA UUG GAA GAA CAG CUU ACG CCU GUU GCU GAA GAG ACC AGA GCA AGA CUG AAG GAG CUU CAA GCU GCC CAG GCA CGA CUU GCC GCC GAC AUG GAA GAC GUU UGU GGG CAG GAG UGAU ACG GAU CA GGC CAG UCU ACA GAG GAG CUC CGG GUU CGG CUU GCG UCA CAC CUC AGA AAA UUG AGG AAG CGG CUC CUG CGC GAU GCU GAC GAU CUU CAG AAA CGA CUG GCA GUA UAU CAG GCC GGA GCA GCAG GAGGA GAA GAG GAU GAA CGC UUG GGU CCC CUU GUU GAA CAA GGG CGC GUU AGA GCC GCA ACC GUU GGG AGU UUG GCU GGU CAG CCU CUC CAG GAG CGA GCG CAA GCA UGG GGU GAG AGG UUG CGA ACC GCCG AUGG GAGG GAC GAA CUG GAU GAG GUU AAG GAA CAA GUC GCU GAG GUA CGC GCGAAA CUU GAG GAA CAA GCA CAG CAG AUU AGA CUU CAG GCA GAG GCA UUU CAG GCG CGG CUU AAG AGU UGG UUC GAG CCG CUC GUG GAA GAU AUG CAG CGC CAA UGG GCA GGA CUC GUC GAA AAA GUU CAG GCA GCU GUC GGG ACG AGU GCA GCC CCU GUC CCA UCA GAC AAU CAC5ApoE4AAG GUG GAG CAA GCG GUG GAG ACA GAG CCG GAG CCC GAG CUG CGC CAG CAG ACC GAG UGG CAG AGC GGC CAG CGC UGG GAA CUG GCA CUG GGU CGC UUU UGG GAU UAC CUG CGC UGG GUG CAG ACA CUG UCU GAG CAG GUG CAG GAG GAG CUG CUC AGC UCC CAG GUC ACC CAG GAA CUG AGG GCG CUG AUG GAC GAG ACC AUG AAG GAG UUG AAG GAC UAC AAA UCG GAA CUG GAG GAA CAA CUG ACC CCG GUG GCG GAG GAG ACG CGG GCA CGG CUG UCC AAG GAG CUG CAG GCG GCG CAG GCC CGG CUG GGC GCG GAC AUG GAG GAC GUG UGC GGC CGC CUG GUG CAG UAC CGC GGC GAG GUG CAG GCC AUG CUC GGC CAG AGC ACC GAG GAG CUG CGG CGG GUG CGC GUG CGC CUC GCC UCC CAC CUG CGC AAG CUG CGU AAG CGG CUC CUC CGC GAU GCC GAU GAC CUG CAG AAG CGC CUG GCA GUG UAC CAG GCC GGG GCC CGC GAG GGC GGC GGC GGC GGC CUC AGC GCC AUC CGC GAG CGC CUG GGG CCC CUG GUG GAA CAG GGC CGC GGG CCC CUG GUGGCC GCC ACU GUG GGC UCC CUG GCC GGC CAG CCG CUA CAG GAG CGG GCC CAG GCC UGG GGC GAG CGG CUG CGC GCG CGG AUG GAG GAG AUG GGC AGC CGG ACC CGC GAC CGC CUG GAC GAG GAG GAG GAG GCG GAG GUG CGC GCC AAG CUG GAG GAG CAG GCC CAG CAG AUA CGC CUG CAG GCC GAG GCC UUC CAG GCC CGC CUC AAG AGC UGG UUC GAG CCC CUG GUG GAA GAC AUG CAG CGC CAG UGG GCC GGG CUG GUG GAG AAG GUG CAG GCU GCC GUG GGC ACC AGC GCC GCC CCU GUG CCC AGC GAC AAU CAC6

[0166]

[0167] The fusion protein produced above was mixed with lipid nanoparticles to produce lipid nanoparticles (targeting delivery vehicles) linked to the fusion protein. Meanwhile, in order to evaluate the targeting efficacy of the lipid nanoparticles, lipid nanoparticles loaded with mRNA of Firefly Luciferase were used as an example of an active ingredient. To produce the lipid nanoparticles loaded with luciferase, ionizable lipids, non-cationic lipids, sterols, and pegylated lipids (PEG) were mixed. Next, the mRNA to be encapsulated in the lipid nanoparticles was included in a citric acid buffer solution between pH 3.0 and pH 3.5 at a concentration of N / P ratio between 4 and 20. The lipid mixture solution and the mRNA solution were mixed at a volume ratio of 1:3 and mixed at a speed of 12.0 ml / min. For the production of lipid nanoparticles, a NanoAssemblr Ignite (NIN0001) device manufactured by Precision Nanosystems was used, and the lipid and mRNA solution mixed through the device was subjected to an ethanol removal process using an Amicon 100kDa membrane (Millipore, UFC810024) using ultrafiltration.

[0168] To confirm whether the lipid nanoparticles manufactured above effectively encapsulated Firefly luciferase mRNA, the mRNA content was measured using the Ribogreen assay kit (Invitrogen, R11490). Specifically, the solution provided in the Ribogreen kit is a fluorescent substance that stains nucleic acids, and the amount of RNA in the solution can be measured to determine the encapsulation efficiency within the lipid nanoparticles. To stain the mRNA within the lipid nanoparticles, the lipid nanoparticles were degraded with a 2% Triton X-100 solution to expose the mRNA. After mixing the degraded lipid nanoparticle solution and the Ribogreen solution, the results were measured using a microplate reader at Excitation 485 nm and Emission 530 nm, confirming that the encapsulation was performed with excellent efficiency (Table 3).

[0169] No.Sample IDEncapsulation Efficiency (%)Total mRNAConc. (ug / ml)Encapsulated mRNAConc. (ug / ml)1Firefly Luciferase mRNA-LNP79.395.976.0

[0170]

[0171] Next, to confirm the targeting efficacy of the lipid nanoparticle containing the fusion protein, the targeting carrier manufactured above was treated to 293T cells (control) or 293T-mPDL1 cells (293T cells expressing mouse PDL1 on the surface), and the cells were harvested after 24 hours. The cell lysate was subjected to a luciferase assay to evaluate the delivery efficiency of the targeting carrier. In addition, to evaluate the delivery efficiency of the fusion protein, lipid nanoparticles containing PD1, ApoE3, or ApoE4, respectively, were manufactured and used as controls. The luciferase assay was performed using the Promega, Dual-Luciferase® Reporter Assay System, #E1910 kit after adjusting the concentration of the protein present in the cell lysate to be the same across samples.

[0172] As a result, in the control 293T cells, regardless of the type of protein linked to the lipid nanoparticles, similar results were observed as when lipid nanoparticles were treated alone. In contrast, in 293T-mPDL1 cells expressing PDL1 on the surface that can bind to the targeting protein PD1, the delivery efficiency of LNPs produced using fusion proteins including ApoE3 and PD1 or ApoE4 and PD1 was significantly increased compared to LNPs produced using only LNPs or LNPs produced by linking only PD1, ApoE3, or ApoE4 (Fig. 3). In addition, when PD1-ApoE3 and PD1-ApoE4, in which the targeting protein (PD1) is linked to the N-terminus of ApoE, were used, the delivery efficiency was significantly improved compared to when ApoE3-PD1 and ApoE4-PD1 (in which the targeting protein is linked to the C-terminus of ApoE) were used.

[0173] Based on the above results, it can be seen that a fusion protein including ApoE and a targeting protein can improve the targeting efficacy of lipid nanoparticles, and in particular, a fusion protein in which a targeting protein is linked to the N-terminus of ApoE can significantly improve the delivery efficiency of lipid nanoparticles.

[0174]

[0175] Example 2: Evaluation of the transporter targeting efficacy of a fusion protein comprising an ApoE protein variant

[0176] To improve the targeting efficacy of a fusion protein containing the ApoE protein, the following experiments were performed.

[0177] Specifically, to improve targeting efficacy, two types of mutants in which a portion of the N-terminus of the ApoE protein was deleted were produced (Fig. 4), and a fusion protein comprising the ApoE protein mutant and the targeting protein was produced. The first mutant was composed of amino acids 151 to 299 of the ApoE protein excluding amino acids 1 to 150, and the second mutant was composed of amino acids 206 to 299 of the ApoE protein excluding amino acids 1 to 205. The amino acid sequences and base sequences of the first and second mutants are shown in Tables 4 and 5, respectively (when the base sequence in Table 5 is a DNA sequence, 'U' is replaced with 'T').

[0178] Protein amino acid sequence SEQ ID NO: 1 variant [ApoE (151-299) DADDLQKRLAVYQAGAREGAERGLSAIRERLGPLVEQGRVRAATVGSLAGQPLQERAQAWGERLRARMEEMGSRTRDRLDEVKEQVAEVRAKLEEQAQQIRLQAEAFQARLKSWFEPLVEDMQRQWAGLVEKVQAAVGTSAAPVPSDNH3 2 variant [ApoE (206-299) RAQAWGERLRARMEEMGSRTRDRLDEVKEQVAEVRAKLEEQAQQIRLQAEAFQARLKSWFEPLVEDMQRQWAGLVEKVQAAVGTSAAPVPSDNH4

[0179] 단백질염기 서열 (DNA 서열 또는 mRNA 서열)서열번호제1 변이체[ApoE(151-299)GAAU GGAGAU CCUGU CAU GAU GAC GAC CAG GCC GGA GCA AGA GAG GGU GCC GAA CGC GGG UUG AGC GCC AUC AUC CGG GAA CGC UUG GGU CCC CUU GUU GAA CAA GGG CGC GUU AGA GCC GCC GCA ACC GUU GGG AGU UUG GCU GGU CAG CAG GCA GAG CUC GAG GAG CA CGA GCC CGG AUG GAG GAA AUG GGA UCU AGG ACC CGG GAC CGC CUG GAU GAG GUU AAG GAA CAA GCU GAG GUA CGC GCG AAA CUU GAG GAA CAA GCA CAG CAG CAG AUU AGA CUU CAG GACG UUC A UG CAG GCUG GCA UGU GAG CCG CUC GUG GAA GAU AUG CAG CGC CAA UGG GGA GGA CUC GUC GAA AAA GUU CAG GCU GUC GGG ACG AGU GCA GCC CCU GUC CCA UCA GAC AAU GAC7제2 변이체[AGG29 GGCG(9AGU)CA AGG UUG CGA GCC CGG AUG GAG GAA AUG GGA UCU AGG ACC CGG GAC CGC CUG GAU GAG GUU AAG CAA GUC GCU GAG GUA CCG GCG AAA CUU GAG GAA CAA GCA CAG CAG AUU AGA GAGG CUU GCU UCA GCA UGG UUC GAG CCG CUC GUG GAA GAU AUG CAG CGC CAA UGG GGA GGA CUC GUC GAA AAA GUU CAG GCA GCU GUC GGGACG AGU GCA GCC CCU GUC CCA UCA GAC AAU CAC8

[0180]

[0181] The fusion protein containing the above ApoE protein variant was mixed with a lipid nanoparticle loaded with luciferase in the same manner as in Example 1 to produce a lipid nanoparticle (targeting carrier) to which the fusion protein was linked. After treating 293T cells (control) or 293T-mPDL1 cells, a luciferase assay was performed with cell lysate 24 hours later to evaluate the delivery efficiency of the targeting carrier. In addition, to evaluate the delivery efficiency of the fusion protein, lipid nanoparticles to which PD1, ApoE3, ApoE4, ApoE(151-299), or ApoE(206-299) proteins were linked were produced and used as a control.

[0182] As a result, in 293T-mPDL1 cells expressing PDL1 on their surface that can bind to the targeting protein PD1, it was confirmed that the delivery efficiency of LNPs produced using a fusion protein comprising ApoE3 and PD1 or ApoE4 and PD1 was improved compared to LNPs produced using only LNPs or LNPs containing only PD1, ApoE3, ApoE4, ApoE(151-299), or ApoE(206-299). In addition, the delivery efficiency was further improved in the case of LNPs containing ApoE(151-299) and PD1 in which the N-terminus of the ApoE protein was partially truncated, or containing a fusion protein comprising ApoE(206-299) and PD1, and in particular, it was confirmed that the delivery efficiency of LNPs containing a fusion protein comprising ApoE(206-299) and PD1 was the highest (Fig. 5).

[0183] Next, to confirm whether the LNP linked to the fusion protein including the ApoE protein variant and the targeting protein exhibits excellent targeting efficacy in cells expressing human PDL1 (hPDL1) on the surface, the targeting carrier manufactured above was treated on 293T cells (control) or 293T-hPDL1 cells (293T expressing hPDL1 on the surface), and then 24 hours later, a luciferase assay was performed with cell lysates to evaluate the delivery efficiency of the targeting carrier.

[0184] As a result, it was confirmed that the delivery efficiency of LNPs containing ApoE (151-299) and PD1 or a fusion protein containing ApoE (206-299) and PD1 was significantly increased (Fig. 6).

[0185] Based on the above results, it can be seen that targeting efficiency is significantly improved when a mutant in which the N-terminal portion of the ApoE protein is deleted is used.

[0186]

[0187] Example 3: Evaluation of the transporter targeting efficacy of a fusion protein comprising an ApoE protein variant and SIRPα.

[0188] In Example 1 above, the targeting efficacy of a fusion protein comprising the ApoE protein (or a variant thereof) was confirmed, and PD1 was used as an example of the targeting protein. Accordingly, to determine whether the targeting efficacy is enhanced by the ApoE protein even when various targeting proteins are used, SIRPα (SEQ ID NO: 10) was used as another example of the targeting protein.

[0189] Meanwhile, SIRPα (Signal Regulatory Protein Alpha) and CD47 are proteins that play crucial roles in the immune system. Tumor cells are known to overexpress CD47 and can bind to SIRPα on macrophages. Therefore, we aimed to confirm the targeting efficacy of a fusion protein containing SIRPα against CD47-expressing cells.

[0190] Specifically, a fusion protein containing an ApoE protein variant and SIRPα as a targeting protein was mixed with a luciferase-loaded lipid nanoparticle in the same manner as in Example 1 to produce a lipid nanoparticle (targeting carrier) to which the fusion protein was linked. After treating A549 cells (control) or A549-CD47 cells (A549 expressing hCD47 on the surface), a luciferase assay was performed with cell lysates 24 hours later to evaluate the delivery efficiency of the targeting carrier. In addition, to evaluate the delivery efficiency of the fusion protein, lipid nanoparticles to which ApoE (206-299) protein or SIRPα was linked were produced and used as a control.

[0191] As a result, it was confirmed that the delivery efficiency of LNPs conjugated with a fusion protein including ApoE (206-299) and SIRPα was significantly increased compared to other comparative groups (Fig. 7).

[0192] Based on the above results, it can be seen that targeting efficiency is significantly improved when a mutant in which a portion of the N-terminus of the ApoE protein is deleted is used, regardless of the type of target protein.

[0193]

[0194] Example 4: Evaluation of the transporter targeting efficacy of a fusion protein comprising an ApoE protein variant and FcBP.

[0195] To evaluate the targeting efficacy of a fusion protein comprising ApoE protein and FcBP, the following experiments were performed.

[0196] Specifically, a fusion protein comprising a mutant comprising the C-terminal region (206-299 aa) of the ApoE protein and an Fc binding peptide (FcBP) or a fragment thereof was constructed. Next, the constructed fusion protein and an antibody as a targeting protein were mixed and incubated overnight at 4°C to prepare a fusion protein-antibody complex. As an example of the targeting protein, an anti-PD-L1 antibody (Invivogen, Catalog Number: hpdl1-mab1) was used. The amino acid sequence and base sequence of the Fc binding peptide used in the construction of the fusion protein are shown in Tables 6 and 7, respectively (when the base sequence in Table 7 is a DNA sequence, 'U' is replaced with 'T').

[0197] Protein amino acid sequence sequence number FcBP (K1) DCAWHLGELVWCT12 FcBP (J4) GPGISAFSPGRGWYDPETGTWYDAAWHLGELVWATYYDPETGTWEPDWQRMLGQ13 FcBP (KJ5) GVVRRWSGDCAWHLGELVWCTRSSIAYGGG14

[0198] Protein base sequence (DNA sequence or mRNA sequence) SEQ ID NO: FcBP (K1) GAU UGU GCC UGG CAC CUG GGA GAG CUG GUG UGG UGU ACC15FcBP (J4) GGG CCA GGC AUU UCU GCC UUU AGC CCA GGA AGG GGC UGG UAC GAU CCU GAG ACC GGG ACA UGG UAU GAU GCC GCU UGG CAC CUG GGA GAG CUG GUG UGG GCC ACC UAC UAC GAU CCU GAA ACA GGC ACC UGG GAG CCU GAU UGG CAG AGA AUG CUG GGU CAG16FcBP(KJ5)GGG GUG GUG AGG AGA UGG AGC GGC GAU UGU GCC UGG CAC CUG GGA GAG CUG GUG UGG UGC. ACA AGA AGU AGU AUC GCU UAU GGA GGA GGG17

[0199]

[0200] The fusion protein-antibody complex was mixed with luciferase-loaded lipid nanoparticles in the same manner as in Example 1 and incubated at room temperature for 4 hours to produce lipid nanoparticles (targeting carriers) to which the fusion protein was linked. These were treated with 293T hPD-L1 cell lines, and 24 hours later, a luciferase assay was performed with cell lysates to evaluate the delivery efficiency of the targeting carrier. In addition, to evaluate the delivery efficiency of the fusion protein, lipid nanoparticles not containing the fusion protein were produced and used as a control, and the signal value was normalized as a relative value (fold change) compared to the mock group. Meanwhile, the specific compositions of the control and experimental groups used in the above experiment are as follows:

[0201] - Group 1: Mock (only DMEM) + RG-fLuc_LNP

[0202] - Group 2: anti-hPD-L1 Ab (control) + RG-fLuc_LNP

[0203] - Group 3: FcBP(K1)-ApoE (30 μg) + anti-hPD-L1 Ab (1 ug) + RG-fLuc_LNP

[0204] - Group 4: FcBP(J4)-ApoE (30 μg) + anti-hPD-L1 Ab (1 ug) + RG-fLuc_LNP

[0205] - Group 5: FcBP(KJ5)-ApoE (30 μg) + anti-hPD-L1 Ab (1 ug) + RG-fLuc_LNP

[0206] As a result of the above experiment, it was confirmed that the delivery efficiency of the LNP in which the complex of the fusion protein containing the ApoE protein and FcBP and the antibody was combined was significantly increased compared to the control group (Fig. 8).

[0207] Next, as another example of the fusion protein and targeting protein produced above, a fusion protein-antibody complex was produced using an anti-CD70 antibody (Invivogen, Catalog Number: hcd70-mab1). The lipid nanoparticles containing this complex were treated with A549 cells expressing CD70, and the delivery efficiency was evaluated. Meanwhile, the specific compositions of the control and experimental groups used in the experiment are as follows:

[0208] - Group 1: Mock (only DMEM) + RG-fLuc_LNP

[0209] - Group 2: anti-CD70 Ab (control) + RG-fLuc_LNP

[0210] - Group 3: FcBP(K1)-ApoE (30 μg) + anti-CD70 Ab + RG-fLuc_LNP

[0211] - Group 4: FcBP(J4)-ApoE (30 μg) + anti-CD70 Ab + RG-fLuc_LNP

[0212] - Group 5: FcBP(KJ5)-ApoE (30 μg) + anti-CD70 Ab + RG-fLuc_LNP

[0213] As a result of the above experiment, it was confirmed that the delivery efficiency of the LNP in which the complex of the fusion protein containing the ApoE protein and FcBP and the antibody was combined was significantly increased compared to the control group (Fig. 9).

[0214]

[0215] Based on the above results, it can be seen that, regardless of the type of antibody that is the targeting protein, a fusion protein containing ApoE protein and FcBP can significantly improve the targeting efficacy of lipid nanoparticles.

[0216]

[0217] The foregoing description of the present invention is provided for illustrative purposes only. Those skilled in the art will readily appreciate that the present invention can be readily modified into other specific forms without altering the technical spirit or essential characteristics of the present invention. Therefore, the embodiments described above should be understood as illustrative in all respects and not restrictive.

Claims

1. 1) Apolipoprotein or a fragment thereof; and 2) Fc binding peptide (FcBP) or fragment thereof A fusion protein comprising:

2. A fusion protein according to claim 1, wherein the Fc region binding peptide or fragment thereof comprises one or more amino acid sequences selected from the group consisting of SEQ ID NOs: 12 to 14.

3. A fusion protein according to claim 1, wherein the apolipoprotein comprises at least one selected from the group consisting of apolipoprotein A (ApoA protein), apolipoprotein B (ApoB protein), apolipoprotein C (ApoC protein), apolipoprotein D (ApoD protein), apolipoprotein E (ApoE protein), apolipoprotein F (ApoF protein), apolipoprotein H (ApoH protein), apolipoprotein L (ApoL protein), apolipoprotein M (ApoM protein), and apolipoprotein(a) (Apo(a) protein).

4. A fusion protein according to claim 3, wherein the ApoE protein comprises at least one selected from the group consisting of ApoE1, ApoE2, ApoE3, ApoE4, ApoE5, ApoE6, ApoE7, ApoE8, ApoE9, ApoE10, and ApoE11.

5. A fusion protein according to claim 1, wherein the apolipoprotein fragment has a receptor-binding region deleted.

6. A fusion protein according to claim 1, wherein the apolipoprotein fragment has a receptor-binding region and a hinge region deleted.

7. A fusion protein according to claim 1, wherein the apolipoprotein fragment has 100 to 150 amino acids deleted from the N-terminus of the apolipoprotein.

8. A fusion protein according to claim 1, wherein the apolipoprotein fragment has 151 to 220 amino acids deleted from the N-terminus of the apolipoprotein.

9. A fusion protein according to claim 1, wherein the apolipoprotein comprises a peptide represented by the amino acid sequence of SEQ ID NO: 1 or SEQ ID NO:

2.

10. A fusion protein according to claim 1, wherein the apolipoprotein fragment comprises a peptide represented by an amino acid sequence in which 100 to 220 amino acids are deleted from the N-terminus of SEQ ID NO:

2.

11. A fusion protein according to claim 1, wherein the apolipoprotein fragment comprises a peptide represented by the amino acid sequence of SEQ ID NO: 3 or SEQ ID NO:

4.

12. A fusion protein according to claim 1, wherein the fusion protein is mutually linked to a targeting protein.

13. In claim 12, the targeting protein comprises an antibody or an antigen-binding fragment thereof capable of binding to at least one selected from the group consisting of PD-L1, PD-L2, MAGE-1, MAGE-2, MAGE-3, MAGE-12, BAGE, GAGE, NY-ESO-1, tyrosinase, TRP-1, TRP-2, gp100, MART-1, MCIR, Ig idotype, CDK4, Caspase-B, beta-catenin, CLA, BCR / ABL, mutated p21 / ras, mutated p53, proteinase 3, WT1, MUC-1, Her2 / neu, PAP, PSA, PSMA, G250, HPV E6 / E7, EBV LMP2a, CEA, alpha-Fetoprotein, 5T4, CD70, CD47, and onco-trophoblast glycoprotein. fusion protein.

14. A polynucleotide comprising a sequence encoding a fusion protein of any one of claims 1 to 13.

15. An expression vector comprising the polynucleotide of claim 14.

16. A targeting delivery system comprising a fusion protein, a targeting protein and a lipid-based delivery system according to any one of claims 1 to 13.

17. A targeting delivery system according to claim 16, wherein the targeting delivery system targets a protein to which the targeting protein can bind or a cell expressing the protein.

18. A targeting delivery system according to claim 16, wherein the targeting delivery system targets cancer.

19. A composition for drug delivery comprising the targeting delivery system of claim 16 and a desired drug.

20. A composition according to claim 19, wherein the composition is for delivering a drug to a target cell or tissue.

21. A composition according to claim 19, wherein the composition is for delivering a drug to cancer cells or cancer lesions.

22. A composition according to claim 21, wherein the cancer is liver cancer, lung cancer, pancreatic cancer, non-small cell lung cancer, colon cancer, colon cancer, bone cancer, skin cancer, head or neck cancer, cutaneous or intraocular melanoma, uterine cancer, ovarian cancer, rectal cancer, stomach cancer, anal cancer, breast cancer, fallopian tube carcinoma, endometrial carcinoma, cervical carcinoma, vaginal carcinoma, vulvar carcinoma, Hodgkin's disease, esophageal cancer, small intestine cancer, endocrine cancer, thyroid cancer, parathyroid cancer, adrenal cancer, soft tissue sarcoma, urethral cancer, penile cancer, prostate cancer, chronic or acute leukemia, lymphocytic lymphoma, bladder cancer, kidney or ureter cancer, renal cell carcinoma, renal pelvic carcinoma, central nervous system (CNS) tumor, primary central nervous system lymphoma, spinal cord tumor, brainstem glioma, or pituitary adenoma.

23. An Fc binding peptide (FcBP) comprising one or more amino acid sequences selected from the group consisting of SEQ ID NOs: 12 to 14.

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