WO2017217744A1 - Human antibody specifically binding to acne bacteria using phage display technique, and use thereof - Google Patents
Human antibody specifically binding to acne bacteria using phage display technique, and use thereof Download PDFInfo
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- WO2017217744A1 WO2017217744A1 PCT/KR2017/006156 KR2017006156W WO2017217744A1 WO 2017217744 A1 WO2017217744 A1 WO 2017217744A1 KR 2017006156 W KR2017006156 W KR 2017006156W WO 2017217744 A1 WO2017217744 A1 WO 2017217744A1
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Classifications
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K39/00—Medicinal preparations containing antigens or antibodies
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K39/00—Medicinal preparations containing antigens or antibodies
- A61K39/395—Antibodies; Immunoglobulins; Immune serum, e.g. antilymphocytic serum
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K8/00—Cosmetics or similar toiletry preparations
- A61K8/18—Cosmetics or similar toiletry preparations characterised by the composition
- A61K8/30—Cosmetics or similar toiletry preparations characterised by the composition containing organic compounds
- A61K8/64—Proteins; Peptides; Derivatives or degradation products thereof
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61Q—SPECIFIC USE OF COSMETICS OR SIMILAR TOILETRY PREPARATIONS
- A61Q19/00—Preparations for care of the skin
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K16/00—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies
- C07K16/12—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from bacteria
Definitions
- the present invention relates to a human antibody that specifically binds to acne bacteria using phage display technology, and more particularly, to specifically bound to Propioni-bacterium acnes (P. acnes ).
- the present invention relates to an antibody and a cosmetic composition and a pharmaceutical composition using the same.
- Acne has not yet been identified as a cause, but increased sebum secretion in the face, neck, chest, back, and shoulders, abnormal keratinization of hair follicles, acne-causing bacteria Propioni bacterium acnes (P. acnes) It is a chronic chronic inflammatory skin disease that is formed by an inflammatory response due to a complex cause, such as colony formation.
- Acne is a common skin disease that occurs in 85% of teenagers and 11% of adults.It usually occurs in early teens and decreases in adulthood. In some cases. Acne in adolescence occurs more frequently in men, but adult acne occurs more frequently in women, which is due to changes in female hormones, which stimulates the release of male hormones as women increase the amount of progesterone associated with premenstrual implantation. In many cases, this causes the symptoms of adult acne to worsen. Acne-causing bacteria P. acnes is an air-resistant anaerobic bacterium found in the skin of most healthy adults. P. acnes is an opportunistic pathogen that does not cause disease in normal people. It is associated with various inflammatory diseases, including stomatitis and bacteremia.
- P. acnes secretes various enzymes under anaerobic conditions in the pores and breaks down the fatty acid and triglyceride of sebum in hair follicles into short chain fatty acid and propionic acid. As P. acnes colonizes the pores and damages the surrounding cells and breaks down sebum, various by-products block the pores, causing acne to occur when the inflammatory response is caused by the human immune system.
- antibiotics and hormones such as lindamycin, erythromycin, etc.
- oral administration such as retinoids
- topical coatings may cause skin irritation, flushing, and dermatitis.
- penetration is not smooth.
- surgical incision has a problem that the damage due to the skin incision and scars can remain, there is a limitation that the UV treatment causes the skin damage by the ultraviolet rays and special equipment.
- phage display technology was first developed in 1990 by the Medical Research Council in the United Kingdom, and produced a human antibody library, expressed in the form of antibody fragments (Fab, ScFv) on the surface of the bacteriophage antibody clones for specific antigens. It is a technique to screen them. The possibility of screening almost any type of human recombinant monoclonal antibody that specifically reacts with an antigen from a single pot antibody library system has been proposed, which can be applied to phage display antibody technology for various diagnostics and treatments in the body. Fragments (in the form of Fab or ScFv) can be obtained.
- One object of the present invention propionyl sludge tumefaciens arc Ness-: to provide a monoclonal antibody specifically binding to (Propioni bacterium acnes P. acnes).
- Another object of the present invention is to provide a use of the monoclonal antibody.
- the present invention provides a monoclonal antibody that specifically binds to Propioni bacterium acnes (P. acnes ).
- the present invention also provides an expression vector comprising a polynucleotide encoding the monoclonal antibody.
- the present invention also provides a transformant transformed with the expression vector.
- the monoclonal antibody may be any one selected from the group consisting of (a) to (d):
- a heavy chain variable region comprising a heavy chain CDR1 as set out in SEQ ID NO: 1, a heavy chain CDR2 as set out in SEQ ID NO: 2, and a heavy chain CDR3 as set out in SEQ ID NO: 3, a light chain CDR1 as set out in SEQ ID NO: 4, a light chain CDR2 as set out in SEQ ID NO: 5, and An antibody comprising a light chain variable region comprising a light chain CDR3 set forth in SEQ ID NO: 6;
- a heavy chain variable region comprising a heavy chain CDR1 as set out in SEQ ID NO: 7, a heavy chain CDR2 as set out in SEQ ID NO: 8, and a heavy chain CDR3 as set out in SEQ ID NO: 9, a light chain CDR1 as set out in SEQ ID NO: 10, a light chain CDR2 as set out in SEQ ID NO: 11, and An antibody comprising a light chain variable region comprising a light chain CDR3 set forth in SEQ ID NO: 12;
- a heavy chain variable region comprising a heavy chain CDR1 as set out in SEQ ID NO: 13, a heavy chain CDR2 as set out in SEQ ID NO: 14, and a heavy chain CDR3 as set out in SEQ ID NO: 15, a light chain CDR1 as set out in SEQ ID NO: 16, a light chain CDR2 as set out in SEQ ID NO: 17, and An antibody comprising a light chain variable region comprising a light chain CDR3 set forth in SEQ ID NO: 18;
- a heavy chain variable region comprising a heavy chain CDR1 as set out in SEQ ID NO: 19, a heavy chain CDR2 as set out in SEQ ID NO: 20, and a heavy chain CDR3 as set out in SEQ ID NO: 21, a light chain CDR1 as set out in SEQ ID NO: 22, a light chain CDR2 as set out in SEQ ID NO: 23, and An antibody comprising a light chain variable region comprising the light chain CDR3 set forth in SEQ ID NO: 24.
- the monoclonal antibody may be any one selected from the group consisting of (a) to (d):
- an antibody comprising a heavy chain variable region as set out in SEQ ID NO: 31 and a light chain variable region as set out in SEQ ID NO: 32.
- the monoclonal antibody may be a human antibody.
- the present invention also provides a cosmetic composition for preventing or improving acne, comprising the monoclonal antibody as an active ingredient.
- the present invention provides a pharmaceutical composition for preventing or treating acne, comprising the monoclonal antibody as an active ingredient.
- the monoclonal antibody according to the present invention can specifically recognize and bind to acne bacteria. Therefore, it is expected that by using a compound having an antimicrobial activity against acne bacteria or an antibody combined with an enzyme, acne symptoms may be alleviated or eliminated without side effects.
- the monoclonal antibody of the present invention has the effect of providing a cosmetic composition and pharmaceutical composition for improving acne without resistance and side effects even when used as a raw material in the long term.
- RCM Reinforced clostridial medium
- FIG. 2 is a diagram of a phage display technology according to an embodiment of the present invention.
- Figure 3 is a photograph of the panning titration of the tertiary P. acnes according to an embodiment of the present invention.
- Figure 4 is a graph of the results of Poly-scFv-phage ELISA according to an embodiment of the present invention.
- FIG. 5 is a graph of the results of Mono-scFv-phage ELISA according to an embodiment of the present invention.
- FIG. 6 is a diagram of a method for converting to an IgG form according to an embodiment of the present invention.
- Figure 8 is a graph of the ELISA results between the human antibody prepared according to an embodiment of the present invention and P. acnes .
- FIG. 9 is a graph showing the results of affinity analysis by ELISA between human antibodies and P. acnes prepared according to an embodiment of the present invention.
- FIG. 10 is a graph showing the results of confirming the growth inhibitory effect of P. acnes of the human antibody prepared according to an embodiment of the present invention.
- antibody includes immunoglobulin molecules that are immunologically reactive with specific antigens and is a concept that includes both polyclonal and monoclonal antibodies.
- the term also includes forms produced by genetic engineering such as chimeric antibodies (eg, humanized murine antibodies) and heterologous antibodies (eg, bispecific antibodies).
- the term further encompasses single chain antibodies, caps, derivatives of antibody constant regions and artificial antibodies based on protein scaffolds that have a binding function to FcRn.
- monoclonal antibody refers to a highly specific antibody directed against a single antigenic site as it is known in the art. Typically, unlike polyclonal antibodies that include different antibodies directed against different epitopes (antigenic determinants), monoclonal antibodies are directed against a single determinant on an antigen. Monoclonal antibodies have the advantage of improving the selectivity and specificity of diagnostic and analytical assays using antigen-antibody binding, and also have another advantage of not being contaminated by other immunoglobulins because they are synthesized by hybridoma culture.
- immunoglobulins typically have heavy and light chains, each heavy and light chain comprising a constant region and a variable region (the site is also known as a "domain").
- the variable regions of the light and heavy chains comprise three variable regions and four "framework regions” called “complementarity determining regions” (hereinafter referred to as "CDRs").
- CDRs mainly serve to bind epitopes of antigens.
- the CDRs of each chain are typically called CDR1, CDR2, CDR3, starting sequentially from the N-terminus and also identified by the chain in which the particular CDR is located.
- variable is used to refer to the fact that the sequence of a particular region differs greatly between antibodies and is used to indicate the binding specificity of each particular antibody for a particular antigen.
- the variability of the antibody is concentrated in the CDRs rather than uniformly distributed throughout the variable domains of the antibodies.
- the heavy and light chains of monoclonal antibodies each have three CDRs and these regions form an antigen-antibody complex by recognizing the surface antigens of the acne bacterium.
- Such CDRs have a characteristic sequence for each monoclonal antibody and some or all of these six CDRs may interact in order for one monoclonal antibody to recognize a particular epitope.
- phage display technique refers to a technique for selecting scFv exhibiting binding ability with an antigen by selecting only phage showing significant binding ability with a target antigen from a phage library.
- panning refers to a surface of a peptide that has the property of binding to a target molecule (antibody, enzyme, cell surface receptor, etc.) from a phage library that displays the peptide on the outer coat of the phage. It refers to the process of selecting only phage expressing. This procedure can be repeated 3 to 10 times to select scFvs that show significant binding capacity to the target antigen, and finally the selected scFvs can be prepared as humanized monoclonal antibodies.
- the present invention provides a monoclonal antibody that specifically binds to Propioni bacterium acnes (P. acnes ).
- the monoclonal antibody may comprise any one sequence selected from the group consisting of the following (a) to (d):
- a heavy chain variable region comprising a heavy chain CDR1 as set out in SEQ ID NO: 1, a heavy chain CDR2 as set out in SEQ ID NO: 2, and a heavy chain CDR3 as set out in SEQ ID NO: 3, a light chain CDR1 as set out in SEQ ID NO: 4, a light chain CDR2 as set out in SEQ ID NO: 5, and An antibody comprising a light chain variable region comprising a light chain CDR3 set forth in SEQ ID NO: 6;
- a heavy chain variable region comprising a heavy chain CDR1 as set out in SEQ ID NO: 7, a heavy chain CDR2 as set out in SEQ ID NO: 8, and a heavy chain CDR3 as set out in SEQ ID NO: 9, a light chain CDR1 as set out in SEQ ID NO: 10, a light chain CDR2 as set out in SEQ ID NO: 11, and An antibody comprising a light chain variable region comprising a light chain CDR3 set forth in SEQ ID NO: 12;
- a heavy chain variable region comprising a heavy chain CDR1 as set out in SEQ ID NO: 13, a heavy chain CDR2 as set out in SEQ ID NO: 14, and a heavy chain CDR3 as set out in SEQ ID NO: 15, a light chain CDR1 as set out in SEQ ID NO: 16, a light chain CDR2 as set out in SEQ ID NO: 17, and An antibody comprising a light chain variable region comprising a light chain CDR3 set forth in SEQ ID NO: 18;
- a heavy chain variable region comprising a heavy chain CDR1 as set out in SEQ ID NO: 19, a heavy chain CDR2 as set out in SEQ ID NO: 20, and a heavy chain CDR3 as set out in SEQ ID NO: 21, a light chain CDR1 as set out in SEQ ID NO: 22, a light chain CDR2 as set out in SEQ ID NO: 23, and An antibody comprising a light chain variable region comprising the light chain CDR3 set forth in SEQ ID NO: 24.
- the monoclonal antibody is preferably any one antibody selected from the group consisting of (a) to (d).
- an antibody comprising a heavy chain variable region as set out in SEQ ID NO: 31 and a light chain variable region as set out in SEQ ID NO: 32.
- the heavy chain variable region of the monoclonal antibody comprises at least one amino acid sequence selected from the group consisting of SEQ ID NOs: 25, 27, 29 and 31 or has at least 80% identity with such sequence in the CDR region, preferably Preferably at least 90%, most preferably at least 95%; Or / and their light chain variable regions comprise at least one amino acid sequence selected from the group consisting of SEQ ID NOs: 26, 28, 30 and 32 or have at least 80% identity, preferably 90% identity with such sequence in the CDR region Or more preferably 95% or more.
- Monoclonal antibodies of the invention can be used to detect Propioni -bacterium acnes (P. acnes ) by reacting with biological samples and detecting antigen-antibody complex formation.
- the "antigen-antibody complex” means a combination of the P. acnes protein antigen in the sample and the monoclonal antibody according to the present invention which recognizes the same.
- the formation of such antigen-antibody complex is performed by colormetric method and electrochemical method. detection by any method selected from the group consisting of electrochemical method, fluorimetric method, luminometry, particle counting method, visual assessment and scintillation counting method can do. However, it is not necessarily limited to these and various applications and applications are possible.
- various labels can be used to detect the antigen-antibody complex.
- Specific examples may be selected from the group consisting of enzymes, fluorescent materials, ligands, luminescent materials, microparticles, and radioisotopes, but are not limited thereto.
- the antigen-antibody complex can be detected using enzyme immunosorbent adsorption (ELISA).
- Enzyme immunosorbent methods include direct ELISA using labeled antibodies that recognize antigens attached to a solid support, and indirect ELISAs using labeled secondary antibodies that recognize capture antibodies in a complex of antibodies that recognize antigens attached to a solid support.
- Direct sandwich ELISA using another labeled antibody that recognizes the antigen in the complex of the antibody and the antibody attached to the solid support followed by reaction with another antibody that recognizes the antigen in the complex of the antibody and the antigen attached to the solid support.
- Various ELISA methods include indirect sandwich ELISAs using labeled secondary antibodies that recognize the antibody.
- the monoclonal antibody may have a detection label, and if it does not have a detection label, the monoclonal antibody may be captured and can be identified by treating another antibody having the detection label.
- the present invention also provides an expression vector comprising a polynucleotide encoding a monoclonal antibody according to the present invention.
- the present invention also provides a transformant transformed with the expression vector.
- the polynucleotide encoding the antibody of the present invention does not change the amino acid sequence of the antibody expressed from the coding region due to degeneracy of the codon or in consideration of the codon preferred in the organism to express the antibody.
- Various modifications may be made to the coding region within the region, and various modifications or modifications may be made within a range that does not affect the expression of the gene even in portions other than the coding region, and such modified genes are also included in the scope of the present invention. Will be well understood. That is, as long as the polynucleotide of the present invention encodes a protein having equivalent activity, one or more nucleic acid bases may be mutated by substitution, deletion, insertion, or a combination thereof, which are also included in the scope of the present invention.
- the sequence of such polynucleotides may be single or double chained and may be DNA molecules or RNA (mRNA) molecules.
- expression control sequences such as promoters, terminators, enhancers, and the like, sequences for membrane targeting or secretion, etc., depending on the type of host cell in which the antibody is to be produced. It can be appropriately selected and various combinations depending on the purpose.
- Expression vectors of the present invention include, but are not limited to, plasmid vectors, cosmid vectors, bacteriophage vectors, and viral vectors.
- Suitable expression vectors include signal sequences or leader sequences for membrane targeting or secretion in addition to expression control elements such as promoters, operators, initiation codons, termination codons, polyadenylation signals and enhancers and can be prepared in various ways depending on the purpose.
- the promoter of the expression vector may be constitutive or inducible.
- the signal sequence includes a PhoA signal sequence and an OmpA signal sequence when the host is Escherichia sp., And an ⁇ -amylase signal sequence and a subtilisin signal when the host is Bacillus sp.
- the MF ⁇ signal sequence, the SUC2 signal sequence, and the like may be used when the host is a yeast, and the insulin signal sequence, the ⁇ -interferon signal sequence, the antibody molecular signal sequence, or the like may be used when the host is an animal cell.
- the expression vector may also include a selection marker for selecting a host cell containing the vector, and in the case of a replicable expression vector, includes the origin of replication.
- the present invention comprises the steps of 1) inoculating and transforming the transformant in the medium;
- the expression vector according to the present invention may be transformed into an appropriate host cell, for example, E. coli or yeast cell, and then cultured in the transformed host cell, thereby producing a large amount of the antibody according to the present invention.
- Suitable culture methods, media conditions, and the like according to the type of host cell can be easily selected by those skilled in the art from known techniques known to those skilled in the art.
- the host cell may be a prokaryote such as E. coli or Bacillus subtilis . It may also be eukaryotic cells derived from yeast, insect cells, plant cells, animal cells such as Saccharomyces cerevisiae . More preferably, the animal cell may be a cell line derived from human.
- the expression vector introduction method into the host cell may be any method known to those skilled in the art.
- the invention also relates to P. It provides a cosmetic composition for preventing or improving acne comprising a monoclonal antibody that specifically binds to acnes as an active ingredient.
- the present invention provides a use for the use of the monoclonal antibody in the cosmetic composition for preventing or improving acne.
- the cosmetic composition of the present invention includes components commonly used in cosmetic compositions in addition to the human antibody, and includes, for example, conventional auxiliaries such as sulfates, stabilizers, solubilizers, vitamins, pigments and flavors, and carriers.
- auxiliaries such as sulfates, stabilizers, solubilizers, vitamins, pigments and flavors, and carriers.
- the monoclonal antibody of the present invention to the cosmetic composition usually contained may be added in an amount of 0.1 to 50% by weight, preferably 1 to 10% by weight.
- Cosmetic compositions of the present invention may be prepared in any formulation conventionally prepared in the art and include, for example, solutions, suspensions, emulsions, pastes, gels, creams, lotions, powders, soaps, surfactant-containing cleansing , Oils, powder foundations, emulsion foundations, wax foundations and sprays, and the like, but are not limited thereto. More specifically, it may be prepared in the form of a flexible lotion (skin), nutrition lotion (milk lotion), nutrition cream, massage cream, essence, eye cream, cleansing cream, cleansing foam, cleansing water, pack, spray or powder .
- the formulation of the present invention is a paste, cream or gel, animal oil, vegetable oil, wax, paraffin, starch, tragacanta, cellulose derivative, polyethylene glycol, silicone, bentonite, silica, talc or zinc oxide are used as carrier components.
- the formulation is a powder or spray, lactose, talc, silica, aluminum hydroxide, calcium silicate or polyamide powder may be used as the carrier component, especially in the case of a spray, additionally chlorofluorohydrocarbons.
- Propellant such as propane / butane or dimethyl ether.
- a solvent, solubilizer or emulsifier is used as the carrier component, such as water, ethanol, isopropanol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene glycol, 1 Fatty acid esters of, 3-butylglycol oil, glycerol aliphatic ester, polyethylene glycol or sorbitan.
- liquid carrier diluents such as water, ethanol or propylene glycol
- suspending agents such as ethoxylated isostearyl alcohol, polyoxyethylene sorbitol esters and polyoxyethylene sorbitan esters, and microcrystals
- Soluble cellulose, aluminum metahydroxy, bentonite, agar or tragacanta and the like can be used.
- the carrier component is aliphatic alcohol sulfate, aliphatic alcohol ether sulfate, sulfosuccinic acid monoester, isethionate, imidazolinium derivative, methyltaurate, sarcosinate, fatty acid amide.
- Ether sulfates, alkylamidobetaines, aliphatic alcohols, fatty acid glycerides, fatty acid diethanolamides, vegetable oils, lanolin derivatives or ethoxylated glycerol fatty acid esters and the like can be used.
- the invention also relates to P. It provides a pharmaceutical composition for preventing or treating acne, comprising a monoclonal antibody that specifically binds to acnes as an active ingredient.
- the present invention also provides the use of a pharmaceutical composition for the prevention or treatment of acne comprising the monoclonal antibody.
- the monoclonal antibodies of the present invention can be administered parenterally during clinical administration and can be used in the form of general pharmaceutical formulations.
- Parenteral methods of administration include, but are not limited to, intravenous, intramuscular, intraarterial, intramedullary, intradural, intracardiac, transdermal, subcutaneous, intraperitoneal, intranasal, intestinal, topical, sublingual or rectal administration. Can be.
- compositions of the present invention may be formulated according to methods known in the art in the form of injections, transdermal and nasal inhalants with suitable parenteral carriers.
- suitable parenteral carriers include, but are not limited to, solvents or dispersion media comprising water, ethanol, polyols (e.g., glycerol, propylene glycol and liquid polyethylene glycols, etc.), mixtures thereof and / or vegetable oils Can be.
- suitable carriers include Hanks' solution, Ringer's solution, phosphate buffered saline (PBS) containing triethanol amine or sterile water for injection, 10% ethanol, 40% propylene glycol and 5% dextrose Etc. can be used.
- PBS phosphate buffered saline
- various antibacterial and antifungal agents such as parabens, chlorobutanol, phenol, sorbic acid, thimerosal, and the like may be further included.
- the injection may in most cases further comprise an isotonic agent such as sugar or sodium chloride.
- transdermal administrations In the case of transdermal administrations, ointments, creams, lotions, gels, external preparations, pasta preparations, linen preparations, air rolls and the like are included.
- 'transdermal administration' means that the pharmaceutical composition is locally administered to the skin so that an effective amount of the active ingredient contained in the pharmaceutical composition is delivered into the skin.
- the compounds used according to the invention may be pressurized packs or by means of suitable propellants, for example dichlorofluoromethane, trichlorofluoromethane, dichlorotetrafluoroethane, carbon dioxide or other suitable gas. It can be delivered conveniently from the nebulizer in the form of an aerosol spray. In the case of a pressurized aerosol, the dosage unit can be determined by providing a valve to deliver a metered amount.
- gelatin capsules and cartridges for use in inhalers or blowers can be formulated to contain a mixture of the compound and a suitable powder base such as lactose or starch. Formulations for parenteral administration are described in Remington's Pharmaceutical Science, 15th Edition, 1975. Mack Publishing Company, Easton, Pennsylvania 18042, Chapter 87: Blag, Seymour, a prescription generally known in all pharmaceutical chemistries.
- the pharmaceutical composition of the present invention may provide the desired effect of preventing or treating acne when it contains an effective amount of a monoclonal antibody according to the present invention.
- the 'effective amount' refers to an amount that exhibits a higher response than a negative control, and preferably refers to an amount sufficient to exhibit an effect of killing P. acnes or reducing acne.
- the pharmaceutical composition of the present invention may contain 0.01 to 99.99% monoclonal antibody according to the present invention, the remaining amount may be occupied by a pharmaceutically acceptable carrier.
- the effective amount of the monoclonal antibody according to the present invention included in the pharmaceutical composition of the present invention will vary depending on the form in which the composition is commercialized and the like.
- the total effective amount of the pharmaceutical composition of the present invention may be administered to a patient in a single dose and may be administered by a fractionated treatment protocol which is administered in multiple doses for a long time.
- the pharmaceutical composition of the present invention may vary the content of the active ingredient depending on the extent of the disease.
- the present invention is administered in an amount of preferably 0.01 to 50 mg, more preferably 0.1 to 30 mg per kg of body weight per day, based on the monoclonal antibody according to the present invention, and when administered orally.
- Based on the monoclonal antibody according to the present invention may be administered in several divided doses so as to be administered in an amount of preferably 0.01 to 100 mg, more preferably 0.01 to 10 mg per kg body weight per day.
- the dose of the monoclonal antibody according to the present invention may be adjusted to the patient in consideration of various factors such as the age, weight, health condition, sex, severity of the disease, diet and excretion rate, as well as the route of administration and the frequency of treatment of the pharmaceutical composition.
- the effective dosage is determined, one of ordinary skill in the art will be able to determine the appropriate effective dosage for the specific use of the monoclonal antibody according to the invention for the prevention or treatment of acne.
- the pharmaceutical composition according to the present invention is not particularly limited to its formulation, route of administration and method of administration as long as the effect of the present invention is shown.
- composition of the present invention may be used alone or in combination with methods using surgery, radiation therapy, hormone therapy, chemotherapy or biological response modifiers.
- the pharmaceutical composition of the present invention may also be provided in the formulation of an external preparation containing the monoclonal antibody according to the present invention as an active ingredient.
- the pharmaceutical composition of the present invention is used as an external preparation for skin, it is additionally used for fatty substances, organic solvents, solubilizers, thickening and gelling agents, emollients, antioxidants, suspending agents, stabilizers, foaming agents, fragrances, interfaces.
- Skin such as active agents, water, ionic emulsifiers, nonionic emulsifiers, fillers, metal ion sequestrants, chelating agents, preservatives, vitamins, blockers, wetting agents, essential oils, dyes, pigments, hydrophilic active agents, lipophilic active agents or lipid vesicles It may contain adjuvants commonly used in the field of dermatology, such as any other ingredients commonly used in external preparations. The ingredients may also be introduced in amounts generally used in the field of dermatology.
- the pharmaceutical composition of the present invention when provided as an external preparation for skin, it may be a formulation such as, but not limited to, an ointment, a patch, a gel, a cream, or a spray.
- propionibacterium acnes bacteria were cultured to prepare.
- P. acnes is a Gram-positive bacterium, which has a large number of antigens specific to cell membranes.
- scFv-phage 1 ml of a human-derived scFv library phage with a variety of 2.7-10 10 and 2 ml of Staphylococcus epidermidis , an epidermis , were reacted for 2 hours.
- the scFv-phage which is likely to bind, was pretreated (step 1). After removal by reacting the remaining scFv-phage P. acnes and for 2 hours to obtain a poly-scFv-phage to elute the scFv-phage that specifically bind only to P. acnes (step 2).
- the obtained Poly-scFv-phage was infected with XL1-Blue and amplified, and the amplified Poly-scFv-phage was repeated three times in total to repeat the steps 1 and 2 to selectively select only affinity phages. Amplified (panning step). Finally, the obtained poly-scFv-phage was diluted, titration was analyzed, and the amplification level was confirmed.
- the level of the phage candidate group amplified in each panning step is shown in Table 1 below.
- the number of scFv-phage containing a reaction solution of P. acnes and the reaction before 1 ml are each 1? 10 13 1.6? 10, 13, and 5? 10 was 12, obtained by combining react with P. acnes
- the number of one scFv-phage was 2-10 4 , 8-10 4 and 1-10 5 respectively (FIG. 3).
- the reaction solution in all the wells was removed, and the diluted PBST was dispensed twice by washing 200 ⁇ l per well. After the washing was completed, 100 ⁇ l of hFc-HRP diluted 1: 4000 in each well was dispensed and reacted for 10 minutes in the dark at room temperature to induce color development. In order to terminate the blue color reaction, the stop solution was dispensed by 100 ⁇ l per well, and when the color of the reactant turned yellow, the absorbance was measured for each well at 450 nm using a microplate reader.
- step ⁇ 2-1> From the P. acnes binding poly-scFv-phage obtained in step ⁇ 2-1>, 10 single phages showing high affinity were selected. The selected single phage was again subjected to ELISA to confirm the binding to P. acnes (Fig. 5).
- the light and heavy chain CDR region sequences, and the light and heavy chain variable region sequences of the prepared monoclonal antibodies are shown in Table 5 below.
- FIG. 8 and FIG. 9 all four antibodies (3F, 6F, 8F and 10F) were found to be binding-friendly to P. acnes compared to control cells.
- the 6F and 8F antibody is an antibody showing excellent binding to P. acnes .
- the growth rate of P. acnes was about 150% in the untreated control group, while 22.2% and 30.8% in the experimental group added with 6F antibody or 8F antibody alone, respectively. Only it was confirmed that it can show a significant growth rate reduction effect compared to the untreated control.
- the growth rate of P. acnes was 83.5% compared to 0 hours, and it was confirmed that it showed a 16.5% reduced level of survival, and simultaneously treated 6F and 8F antibodies.
- compositions of the present invention are illustrated below.
- the flexible cosmetics containing the monoclonal antibody of the present invention can be prepared according to the preparation method in the conventional cosmetic field by combining as described in the following [Table 7].
- Monoclonal Antibodies of the Invention 0.1-30% 1,3-butylene glycol 3.0 glycerin 5.0 Polyoxyethylene (60) hardening castor oil 0.2 ethanol 8.0 Citric acid 0.02 Sodium citrate 0.06 antiseptic a very small amount Spices a very small amount Purified water To 100
- Monoclonal Antibodies of the Invention 0.1-30% 1,3-butylene glycol 8.0 glycerin 5.0 Squalane 10.0 Monooleic acid polyoxyethylene sorbitan 2.0 Flux oil 0.1-30% 1,3-butylene glycol 3.0 glycerin 5.0 Polyoxyethylene (60) hardening castor oil 0.2 ethanol 8.0 Citric acid 0.02 Sodium citrate 0.06 antiseptic a very small amount Spices a very small amount Purified water To 100
- a cleansing agent comprising the monoclonal antibody of the present invention
- it may be combined as described in the following [Table 11], and may be prepared according to a conventional manufacturing method in the cosmetic field.
- a nourishing cream comprising the monoclonal antibody of the present invention
- it is prepared according to the preparation method in the conventional cosmetic field as described in Table 12 below.
- a massage cream comprising a monoclonal antibody of the present invention it is prepared according to the manufacturing method in the general cosmetic field as described in Table 13 below.
- Monoclonal Antibodies of the Invention 0.1-30% Propylene glycol 6.0 glycerin 4.0 Triethanolamine 0.5 Beeswax 2.0 Tocophenyl Acetate 0.1 Polysorbate 60 3.0 Sorbitan sesquioleate 2.5 Cetearyl Alcohol 2.0 Liquid paraffin 30.0 Xanthan Gum 0.5 Incense, preservative a very small amount Purified water To 100
- a pack comprising the monoclonal antibody of the present invention, it is prepared according to the preparation method in the conventional cosmetic field as described in Table 14 below.
- Monoclonal Antibodies of the Invention 0.1-30% Propylene glycol 2.0 glycerin 4.0 Polyvinyl alcohol 10.0 ethanol 7.0 Fiji-40 Hydrogenated Castor Oil 0.8 Triethanolamine 0.3 Incense, preservative a very small amount Purified water To 100
- the present invention is not limited to the above-described embodiments and preparation examples, and can be variously modified and changed by those skilled in the art, and can be applied to cosmetics of various uses including other color cosmetics. It can be used in the manufacture of a medicament, that is, an ointment that can be applied thinly to the human body depending on its efficacy, which is included in the spirit and scope of the present invention as defined in the appended claims.
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Abstract
The present invention relates to a human antibody which specifically binds to acne bacteria using a phage display technique, and a use thereof and, more specifically, to an antibody which specifically binds to an (antigen), a human antibody comprising the same, a phage display technique using the same, and cosmetic and pharmaceutical compositions using the same.
Description
본 발명은 파지 디스플레이 기술을 이용한 여드름 균에 특이적으로 결합하는 인간 항체 및 이의 용도에 관한 것으로, 보다 상세하게는 프로피오니 박테리움 아크네스(Propioni-bacterium
acnes
: P.
acnes)에 특이적으로 결합하는 항체 및 이를 이용한 화장료 조성물 및 약학적 조성물에 관한 것이다.The present invention relates to a human antibody that specifically binds to acne bacteria using phage display technology, and more particularly, to specifically bound to Propioni-bacterium acnes (P. acnes ). The present invention relates to an antibody and a cosmetic composition and a pharmaceutical composition using the same.
여드름은 아직 발생 원인이 명확하게 밝혀지지 않았으나, 얼굴, 목, 가슴, 등, 어깨 등에서 피지 분비 증가, 모낭 내 이상 각화, 여드름 유발균인 프로피오니 박테리움 아크네스(Propioni
- bacterium
acnes
: P.
acnes)의 군락 형성 등, 복합적 원인에 의한 염증 반응에 의해 형성되는 국한성 만성 염증성 피부 질환이다.Acne has not yet been identified as a cause, but increased sebum secretion in the face, neck, chest, back, and shoulders, abnormal keratinization of hair follicles, acne-causing bacteria Propioni bacterium acnes (P. acnes) It is a chronic chronic inflammatory skin disease that is formed by an inflammatory response due to a complex cause, such as colony formation.
여드름은 10대의 경우 85%, 성인의 경우 11%의 피부에서 관찰될 정도로 흔히 발병하는 피부 질환으로 일반적으로 10대 초반에 발생하고 성인이 되면 감소하지만, 성인이 된 후 발생하거나 오히려 그 증상이 심해지는 경우도 있다. 청소년기의 여드름은 남성에게서 더 빈번하게 발생하지만, 성인 여드름의 경우 여성에게서 더 빈번하게 발생하며, 이는 여성 호르몬의 변화 때문으로 여성은 월경 전 착상과 관련 있는 프로게스테론 분비량이 많아지면서 남성호르몬의 분비 를 자극하게 되고, 이로 인해 성인 여드름의 증상이 악화되는 경우가 많이 있다. 여드름 유발균 P. acnes는 내기성 혐기성 세균으로서, 대부분의 건강한 성인의 피부에서 발견되었으며, P.
acnes는 정상인에게서는 질환을 야기하지 않는 기회감염성 병원균으로, 여드름을 비롯하여 보철 관절 전염, 심내막염, 유육종증, 내안구염, 균혈증 등 다양한 염증 질환과 관련이 있다.Acne is a common skin disease that occurs in 85% of teenagers and 11% of adults.It usually occurs in early teens and decreases in adulthood. In some cases. Acne in adolescence occurs more frequently in men, but adult acne occurs more frequently in women, which is due to changes in female hormones, which stimulates the release of male hormones as women increase the amount of progesterone associated with premenstrual implantation. In many cases, this causes the symptoms of adult acne to worsen. Acne-causing bacteria P. acnes is an air-resistant anaerobic bacterium found in the skin of most healthy adults. P. acnes is an opportunistic pathogen that does not cause disease in normal people. It is associated with various inflammatory diseases, including stomatitis and bacteremia.
P. acnes는 모공 내 혐기적 조건에서 각 종 효소를 분비하여 모낭 속 피지의 지방산(Fatty acid)과 트리글리세리드(Triglyceride)를 짧은 사슬 지방산(Short chain fatty acid)과 프로피온산(Propionic acid)으로 분해하여 양분을 얻으며, P. acnes가 모공 속에서 군락을 형성하면서 주변 세포에 손상을 가하고 피지를 분해하면서 발생되는 각종 부산물들이 모공을 막게 됨으로써 인간의 면역체계에 의해 염증 반응이 야기되면 여드름이 발생하게 된다. P. acnes secretes various enzymes under anaerobic conditions in the pores and breaks down the fatty acid and triglyceride of sebum in hair follicles into short chain fatty acid and propionic acid. As P. acnes colonizes the pores and damages the surrounding cells and breaks down sebum, various by-products block the pores, causing acne to occur when the inflammatory response is caused by the human immune system.
따라서, 여드름을 치료하기 위해 항생제 및 호르몬의 국소 도포(클린다마이신, 에리쓰로마이신 등) 및 경구 투여(레티노이드 계열 등), 외과적 절개, 자외선 치료 등 다양한 방법이 시도되고 있으나, 아직까지 성공적인 치료법은 없으며, 항생제나 호르몬을 장기적으로 사용하면 내성 발생 및 간 독성 유발, 소화성 궤양, 기형아 유발 등의 심각한 부작용을 발생시키기도 하고, 국소 도포제의 경우 피부 자극 및 홍조 발생, 피부염 발생 등의 부작용이 있으며 환부로의 침투가 원활하지 않은 문제점이 있다. 또한, 외과적 절개는 피부 절개에 의한 손상 및 흉터가 남을 수 있다는 문제점이 있고, 자외선 치료의 경우 자외선에 의한 피부 손상 유발 및 특수 장비가 필요하다는 한계점이 있다. Therefore, various methods such as topical application of antibiotics and hormones (Klindamycin, erythromycin, etc.) and oral administration (such as retinoids), surgical incisions, and UV treatments have been tried to treat acne. Long-term use of antibiotics or hormones may cause serious side effects such as resistance and liver toxicity, peptic ulcers, and malformations, and topical coatings may cause skin irritation, flushing, and dermatitis. There is a problem that the penetration is not smooth. In addition, the surgical incision has a problem that the damage due to the skin incision and scars can remain, there is a limitation that the UV treatment causes the skin damage by the ultraviolet rays and special equipment.
이에, 파지 디스플레이 기술은 영국의 Medical Research Council에서 1990년 처음 개발된 것으로, 인간 항체 라이브러리(library)를 제조하여 항체절편(Fab, ScFv)의 형태로 박테리오파지의 표면에 발현시켜 특정 항원에 대한 항체 클론들을 선별하는 기술이다. 항원과 특이적으로 반응하는 거의 모든 종류의 인간 재조합 단일 클론 항체를 단일 pot 항체 라이브러리 시스템으로부터 선별해낼 수 있는 가능성이 제안되었으며, 이는 파지 디스플레이 항체 기술을 활용할 경우 체내 진단이나 치료에 응용할 수 있는 다양한 항체 단편들(Fab 또는 ScFv 형태)을 획득할 수 있다.Thus, phage display technology was first developed in 1990 by the Medical Research Council in the United Kingdom, and produced a human antibody library, expressed in the form of antibody fragments (Fab, ScFv) on the surface of the bacteriophage antibody clones for specific antigens. It is a technique to screen them. The possibility of screening almost any type of human recombinant monoclonal antibody that specifically reacts with an antigen from a single pot antibody library system has been proposed, which can be applied to phage display antibody technology for various diagnostics and treatments in the body. Fragments (in the form of Fab or ScFv) can be obtained.
본 발명의 하나의 목적은 프로피오니 박테리움 아크네스(Propioni
- bacterium acnes: P. acnes)에 특이적으로 결합하는 단일클론항체를 제공하는 것이다.One object of the present invention propionyl sludge tumefaciens arc Ness-: to provide a monoclonal antibody specifically binding to (Propioni bacterium acnes P. acnes).
본 발명의 또다른 목적은, 상기 단일클론항체의 용도를 제공하는 것이다.Another object of the present invention is to provide a use of the monoclonal antibody.
상기 목적을 달성하기 위하여, 본 발명은 프로피오니 박테리움 아크네스(Propioni- bacterium
acnes
: P.
acnes)에 특이적으로 결합하는, 단일클론항체를 제공한다.In order to achieve the above object, the present invention provides a monoclonal antibody that specifically binds to Propioni bacterium acnes (P. acnes ).
또한, 본 발명은 상기 단일클론항체를 암호화하는 폴리뉴클레오티드를 포함하는 발현 벡터를 제공한다.The present invention also provides an expression vector comprising a polynucleotide encoding the monoclonal antibody.
또한, 본 발명은 상기 발현 벡터로 형질전환된 형질전환체를 제공한다.The present invention also provides a transformant transformed with the expression vector.
본 발명의 바람직한 일실시예에서, 상기 단일클론항체는 하기 (a) 내지 (d)로 이루어진 군으로부터 선택되는 어느 하나인 것일 수 있다:In a preferred embodiment of the present invention, the monoclonal antibody may be any one selected from the group consisting of (a) to (d):
(a) 서열번호 1로 기재된 중쇄 CDR1, 서열번호 2로 기재된 중쇄 CDR2, 및 서열번호 3으로 기재된 중쇄 CDR3를 포함하는 중쇄 가변영역과 서열번호 4로 기재된 경쇄 CDR1, 서열번호 5로 기재된 경쇄 CDR2 및 서열번호 6으로 기재된 경쇄 CDR3를 포함하는 경쇄 가변영역을 포함하는 항체;(a) a heavy chain variable region comprising a heavy chain CDR1 as set out in SEQ ID NO: 1, a heavy chain CDR2 as set out in SEQ ID NO: 2, and a heavy chain CDR3 as set out in SEQ ID NO: 3, a light chain CDR1 as set out in SEQ ID NO: 4, a light chain CDR2 as set out in SEQ ID NO: 5, and An antibody comprising a light chain variable region comprising a light chain CDR3 set forth in SEQ ID NO: 6;
(b) 서열번호 7로 기재된 중쇄 CDR1, 서열번호 8로 기재된 중쇄 CDR2, 및 서열번호 9로 기재된 중쇄 CDR3를 포함하는 중쇄 가변영역과 서열번호 10으로 기재된 경쇄 CDR1, 서열번호 11로 기재된 경쇄 CDR2 및 서열번호 12로 기재된 경쇄 CDR3를 포함하는 경쇄 가변영역을 포함하는 항체;(b) a heavy chain variable region comprising a heavy chain CDR1 as set out in SEQ ID NO: 7, a heavy chain CDR2 as set out in SEQ ID NO: 8, and a heavy chain CDR3 as set out in SEQ ID NO: 9, a light chain CDR1 as set out in SEQ ID NO: 10, a light chain CDR2 as set out in SEQ ID NO: 11, and An antibody comprising a light chain variable region comprising a light chain CDR3 set forth in SEQ ID NO: 12;
(c) 서열번호 13으로 기재된 중쇄 CDR1, 서열번호 14로 기재된 중쇄 CDR2, 및 서열번호 15로 기재된 중쇄 CDR3를 포함하는 중쇄 가변영역과 서열번호 16으로 기재된 경쇄 CDR1, 서열번호 17로 기재된 경쇄 CDR2 및 서열번호 18로 기재된 경쇄 CDR3를 포함하는 경쇄 가변영역을 포함하는 항체; 및(c) a heavy chain variable region comprising a heavy chain CDR1 as set out in SEQ ID NO: 13, a heavy chain CDR2 as set out in SEQ ID NO: 14, and a heavy chain CDR3 as set out in SEQ ID NO: 15, a light chain CDR1 as set out in SEQ ID NO: 16, a light chain CDR2 as set out in SEQ ID NO: 17, and An antibody comprising a light chain variable region comprising a light chain CDR3 set forth in SEQ ID NO: 18; And
(d) 서열번호 19로 기재된 중쇄 CDR1, 서열번호 20으로 기재된 중쇄 CDR2, 및 서열번호 21로 기재된 중쇄 CDR3를 포함하는 중쇄 가변영역과 서열번호 22로 기재된 경쇄 CDR1, 서열번호 23으로 기재된 경쇄 CDR2 및 서열번호 24로 기재된 경쇄 CDR3를 포함하는 경쇄 가변영역을 포함하는 항체.(d) a heavy chain variable region comprising a heavy chain CDR1 as set out in SEQ ID NO: 19, a heavy chain CDR2 as set out in SEQ ID NO: 20, and a heavy chain CDR3 as set out in SEQ ID NO: 21, a light chain CDR1 as set out in SEQ ID NO: 22, a light chain CDR2 as set out in SEQ ID NO: 23, and An antibody comprising a light chain variable region comprising the light chain CDR3 set forth in SEQ ID NO: 24.
본 발명의 바람직한 일실시예에서, 상기 단일클론항체는 하기 (a) 내지 (d)로 이루어진 군으로부터 선택되는 어느 하나일 수 있다:In one preferred embodiment of the invention, the monoclonal antibody may be any one selected from the group consisting of (a) to (d):
(a) 서열번호 25로 기재된 중쇄 가변영역 및 서열번호 26으로 기재된 경쇄 가변영역을 포함하는 항체;(a) an antibody comprising a heavy chain variable region as set out in SEQ ID NO: 25 and a light chain variable region as set out in SEQ ID NO: 26;
(b) 서열번호 27로 기재된 중쇄 가변영역 및 서열번호 28로 기재된 경쇄 가변영역을 포함하는 항체;(b) an antibody comprising a heavy chain variable region as set out in SEQ ID NO: 27 and a light chain variable region as set out in SEQ ID NO: 28;
(c) 서열번호 29로 기재된 중쇄 가변영역 및 서열번호 30로 기재된 경쇄 가변영역을 포함하는 항체;(c) an antibody comprising a heavy chain variable region as set out in SEQ ID NO: 29 and a light chain variable region as set out in SEQ ID NO: 30;
(d) 서열번호 31로 기재된 중쇄 가변영역 및 서열번호 32로 기재된 경쇄 가변영역을 포함하는 항체.(d) an antibody comprising a heavy chain variable region as set out in SEQ ID NO: 31 and a light chain variable region as set out in SEQ ID NO: 32.
본 발명의 바람직한 일실시예에서, 상기 단일클론항체는 인간 항체일 수 있다.In a preferred embodiment of the present invention, the monoclonal antibody may be a human antibody.
또한, 본 발명은 상기 단일클론항체를 유효성분으로 포함하는, 여드름에 대한 예방 또는 개선용 화장료 조성물을 제공한다.The present invention also provides a cosmetic composition for preventing or improving acne, comprising the monoclonal antibody as an active ingredient.
아울러, 본 발명은 상기 단일클론항체를 유효성분으로 포함하는, 여드름에 대한 예방 또는 치료용 약학적 조성물을 제공한다.In addition, the present invention provides a pharmaceutical composition for preventing or treating acne, comprising the monoclonal antibody as an active ingredient.
본 발명에 따른 단일클론항체는 여드름 균을 특이적으로 인지하고 결합할 수 있다. 이에, 여드름 균에 대해서 항균활성을 나타내는 화합물 또는 효소와 결합된 형태의 항체인 것을 이용함으로써 부작용 없이 여드름 증세를 완화하거나 제거할 수 있을 것으로 기대된다.The monoclonal antibody according to the present invention can specifically recognize and bind to acne bacteria. Therefore, it is expected that by using a compound having an antimicrobial activity against acne bacteria or an antibody combined with an enzyme, acne symptoms may be alleviated or eliminated without side effects.
따라서, 본 발명의 단일클론항체는 원료로 사용하여 장기적으로 사용하여도 내성 및 부작용이 없는 여드름 개선용 화장료 조성물 및 약학적 조성물을 제공할 수 있는 효과가 있다.Therefore, the monoclonal antibody of the present invention has the effect of providing a cosmetic composition and pharmaceutical composition for improving acne without resistance and side effects even when used as a raw material in the long term.
도 1은 본 발명의 일 구현예에 따른 Reinforced clostridial medium (RCM)에서 배양된 P.
acnes에 관한 사진이다.1 is a photograph of P. acnes cultured in Reinforced clostridial medium (RCM) according to an embodiment of the present invention.
도 2은 본 발명의 일 구현예에 따른 파지 디스플레이 기술에 관한 그림이다.2 is a diagram of a phage display technology according to an embodiment of the present invention.
도 3는 본 발명의 일 구현예에 따른 3차 P.
acnes의 Panning titration에 관한 사진이다.Figure 3 is a photograph of the panning titration of the tertiary P. acnes according to an embodiment of the present invention.
도 4는 본 발명의 일 구현예에 따른 Poly-scFv-파지 ELISA 결과에 관한 그래프이다.Figure 4 is a graph of the results of Poly-scFv-phage ELISA according to an embodiment of the present invention.
도 5은 본 발명의 일 구현예에 따른 Mono-scFv-파지 ELISA 결과에 관한 그래프이다.5 is a graph of the results of Mono-scFv-phage ELISA according to an embodiment of the present invention.
도 6는 본 발명의 일 구현예에 따른 IgG form으로 전환하기 위한 방법에 관한 그림이다.6 is a diagram of a method for converting to an IgG form according to an embodiment of the present invention.
도 7은 본 발명의 일 구현예에 따른 인간 항체 발현 및 정제에 관한 실험 데이터이다.7 is experimental data on human antibody expression and purification according to one embodiment of the present invention.
도 8은 본 발명의 일 구현예에 따른 제작된 인간 항체와 P.
acnes 간 ELISA 결과에 관한 그래프이다.Figure 8 is a graph of the ELISA results between the human antibody prepared according to an embodiment of the present invention and P. acnes .
도 9는 본 발명의 일 구현예에 따른 제작된 인간 항체와 P.
acnes 간 ELISA를 통한 친화도 분석 결과에 관한 그래프이다.9 is a graph showing the results of affinity analysis by ELISA between human antibodies and P. acnes prepared according to an embodiment of the present invention.
도 10은 본 발명의 일 구현예에 따른 제작된 인간 항체의 P.
acnes 생장 억제 효과를 확인한 결과에 관한 그래프이다.10 is a graph showing the results of confirming the growth inhibitory effect of P. acnes of the human antibody prepared according to an embodiment of the present invention.
이하, 본 발명에서 사용한 용어를 설명한다.Hereinafter, the term used by this invention is demonstrated.
본 발명에서 사용되는 용어 "항체"는 면역학적으로 특정 항원과 반응성을 갖는 면역글로불린 분자를 포함하며, 다클론항체 및 단일클론항체를 모두 포함하는 개념이다. 또한, 상기 용어는 키메라 항체(예를 들면, 인간화 뮤린 항체) 및 이종결합항체(예를 들면, 양특이성 항체)와 같은 유전공학에 의해 생산된 형태를 포함한다. 상기 용어는 추가로 FcRn에 대한 결합 기능을 보유한 단쇄 항체, 스캡, 항체 불변영역의 유도체 및 단백질 스캐폴드에 기초한 인공 항체를 포함한다. As used herein, the term “antibody” includes immunoglobulin molecules that are immunologically reactive with specific antigens and is a concept that includes both polyclonal and monoclonal antibodies. The term also includes forms produced by genetic engineering such as chimeric antibodies (eg, humanized murine antibodies) and heterologous antibodies (eg, bispecific antibodies). The term further encompasses single chain antibodies, caps, derivatives of antibody constant regions and artificial antibodies based on protein scaffolds that have a binding function to FcRn.
본 발명에서 사용되는 용어 "단일클론항체"란 당해 분야에 공지된 용어로서 단일 항원성 부위에 대해서 지시되는 고도의 특이적인 항체를 의미한다. 통상적으로, 상이한 에피토프(항원결정기)들에 대해 지시되는 상이한 항체들을 포함하는 다클론 항체와는 다르게, 단일클론항체는 항원상의 단일 결정기에 대해서 지시된다. 단일클론항체는 항원-항체 결합을 이용하는 진단 및 분석학적 분석법의 선택성과 특이성을 개선시키는 장점이 있으며, 또한 하이브리도마 배양에 의해 합성되기 때문에 다른 면역글로불린에 의해 오염되지 않는 또 다른 장점을 갖는다.The term "monoclonal antibody" as used herein refers to a highly specific antibody directed against a single antigenic site as it is known in the art. Typically, unlike polyclonal antibodies that include different antibodies directed against different epitopes (antigenic determinants), monoclonal antibodies are directed against a single determinant on an antigen. Monoclonal antibodies have the advantage of improving the selectivity and specificity of diagnostic and analytical assays using antigen-antibody binding, and also have another advantage of not being contaminated by other immunoglobulins because they are synthesized by hybridoma culture.
전형적으로, 면역글로불린은 중쇄 및 경쇄를 가지며 각각의 중쇄 및 경쇄는 불변 영역 및 가변 영역 (상기 부위는 "도메인"으로 또한 알려져 있음)을 포함한다. 경쇄 및 중쇄의 가변 영역은, "상보성 결정 영역"(complementarity determining region, 이하, "CDR"이라 함)이라고 불리는 3개의 가변 영역 및 4개의 "구조 영역" (framework region)을 포함한다. 상기 CDR은 주로 항원의 에피토프(epitope)에 결합하는 역할을 한다. 각각의 사슬의 CDR은 전형적으로 N-말단으로부터 시작하여 순차적으로 CDR1, CDR2, CDR3로 불리고, 또한 특정 CDR이 위치하고 있는 사슬에 의해서 식별된다.Typically, immunoglobulins have heavy and light chains, each heavy and light chain comprising a constant region and a variable region (the site is also known as a "domain"). The variable regions of the light and heavy chains comprise three variable regions and four "framework regions" called "complementarity determining regions" (hereinafter referred to as "CDRs"). The CDRs mainly serve to bind epitopes of antigens. The CDRs of each chain are typically called CDR1, CDR2, CDR3, starting sequentially from the N-terminus and also identified by the chain in which the particular CDR is located.
본 발명에서 사용되는 용어 "가변"이란 항체들 간에 특정 영역의 서열이 크게 상이하고 특정 항원에 대해 각각의 특정 항체의 결합 특이성을 나타내는데 사용된다는 것을 일컫기 위해 사용된다. 항체의 가변성은 항체의 가변 도메인 전체에 걸쳐 균일하게 분포하는 것이 아니라 CDR에 집중된다. 단클론 항체의 중쇄 및 경쇄는 각각 3개의 CDR을 가지며 이들 영역이 여드름균의 표면 항원를 인식함으로써 항원-항체 복합체를 형성한다. 이러한 CDR은 각각의 단일클론항체마다 특징적인 서열을 가지며 하나의 단일클론항체가 특정 에피토프를 인식하기 위해 이들 6개의 CDR의 일부 또는 모두가 상호작용할 수 있다.As used herein, the term “variable” is used to refer to the fact that the sequence of a particular region differs greatly between antibodies and is used to indicate the binding specificity of each particular antibody for a particular antigen. The variability of the antibody is concentrated in the CDRs rather than uniformly distributed throughout the variable domains of the antibodies. The heavy and light chains of monoclonal antibodies each have three CDRs and these regions form an antigen-antibody complex by recognizing the surface antigens of the acne bacterium. Such CDRs have a characteristic sequence for each monoclonal antibody and some or all of these six CDRs may interact in order for one monoclonal antibody to recognize a particular epitope.
본 발명에서 사용되는 용어 "파지 디스플레이 기술"은 파지 라이브러리로부터 표적 항원과 유의적인 결합능을 나타내는 파지만을 선별하는 것을 통해, 항원과의 결합능을 나타내는 scFv를 선별하는 기술을 나타낸다. 상기 기술에서, "패닝(panning)"은 파지의 외벽(coat)에 펩타이드를 발현(display)하는 파지 라이브러리로부터 표적 분자(항체, 효소, 세포표면 리셉터 등)와 결합하는 성질을 지닌 펩타이드를 표면에 발현하고 있는 파지만을 선택해 내는 과정을 일컫는다. 이러한 과정을 3 내지 10회 반복하여, 표적 항원에 대하여 유의적인 결합능을 나타내는 scFv를 선별할 수 있고, 최종적으로 선별된 scFv를 인간화 단일클론항체로 제조할 수 있다.The term "phage display technique" as used herein refers to a technique for selecting scFv exhibiting binding ability with an antigen by selecting only phage showing significant binding ability with a target antigen from a phage library. In this technique, "panning" refers to a surface of a peptide that has the property of binding to a target molecule (antibody, enzyme, cell surface receptor, etc.) from a phage library that displays the peptide on the outer coat of the phage. It refers to the process of selecting only phage expressing. This procedure can be repeated 3 to 10 times to select scFvs that show significant binding capacity to the target antigen, and finally the selected scFvs can be prepared as humanized monoclonal antibodies.
이하, 본 발명을 상세하게 설명한다.EMBODIMENT OF THE INVENTION Hereinafter, this invention is demonstrated in detail.
본 발명은 프로피오니 박테리움 아크네스(Propioni
- bacterium
acnes
: P. acnes)에 특이적으로 결합하는, 단일클론항체를 제공한다.The present invention provides a monoclonal antibody that specifically binds to Propioni bacterium acnes (P. acnes ).
구체적으로, 상기 단일클론항체는 하기 (a) 내지 (d)로 이루어진 군으로부터 선택되는 어느 하나의 서열을 포함할 수 있다:Specifically, the monoclonal antibody may comprise any one sequence selected from the group consisting of the following (a) to (d):
(a) 서열번호 1로 기재된 중쇄 CDR1, 서열번호 2로 기재된 중쇄 CDR2, 및 서열번호 3으로 기재된 중쇄 CDR3를 포함하는 중쇄 가변영역과 서열번호 4로 기재된 경쇄 CDR1, 서열번호 5로 기재된 경쇄 CDR2 및 서열번호 6으로 기재된 경쇄 CDR3를 포함하는 경쇄 가변영역을 포함하는 항체; (a) a heavy chain variable region comprising a heavy chain CDR1 as set out in SEQ ID NO: 1, a heavy chain CDR2 as set out in SEQ ID NO: 2, and a heavy chain CDR3 as set out in SEQ ID NO: 3, a light chain CDR1 as set out in SEQ ID NO: 4, a light chain CDR2 as set out in SEQ ID NO: 5, and An antibody comprising a light chain variable region comprising a light chain CDR3 set forth in SEQ ID NO: 6;
(b) 서열번호 7로 기재된 중쇄 CDR1, 서열번호 8로 기재된 중쇄 CDR2, 및 서열번호 9로 기재된 중쇄 CDR3를 포함하는 중쇄 가변영역과 서열번호 10으로 기재된 경쇄 CDR1, 서열번호 11로 기재된 경쇄 CDR2 및 서열번호 12로 기재된 경쇄 CDR3를 포함하는 경쇄 가변영역을 포함하는 항체;(b) a heavy chain variable region comprising a heavy chain CDR1 as set out in SEQ ID NO: 7, a heavy chain CDR2 as set out in SEQ ID NO: 8, and a heavy chain CDR3 as set out in SEQ ID NO: 9, a light chain CDR1 as set out in SEQ ID NO: 10, a light chain CDR2 as set out in SEQ ID NO: 11, and An antibody comprising a light chain variable region comprising a light chain CDR3 set forth in SEQ ID NO: 12;
(c) 서열번호 13으로 기재된 중쇄 CDR1, 서열번호 14로 기재된 중쇄 CDR2, 및 서열번호 15로 기재된 중쇄 CDR3를 포함하는 중쇄 가변영역과 서열번호 16으로 기재된 경쇄 CDR1, 서열번호 17로 기재된 경쇄 CDR2 및 서열번호 18로 기재된 경쇄 CDR3를 포함하는 경쇄 가변영역을 포함하는 항체; 및(c) a heavy chain variable region comprising a heavy chain CDR1 as set out in SEQ ID NO: 13, a heavy chain CDR2 as set out in SEQ ID NO: 14, and a heavy chain CDR3 as set out in SEQ ID NO: 15, a light chain CDR1 as set out in SEQ ID NO: 16, a light chain CDR2 as set out in SEQ ID NO: 17, and An antibody comprising a light chain variable region comprising a light chain CDR3 set forth in SEQ ID NO: 18; And
(d) 서열번호 19로 기재된 중쇄 CDR1, 서열번호 20으로 기재된 중쇄 CDR2, 및 서열번호 21로 기재된 중쇄 CDR3를 포함하는 중쇄 가변영역과 서열번호 22로 기재된 경쇄 CDR1, 서열번호 23으로 기재된 경쇄 CDR2 및 서열번호 24로 기재된 경쇄 CDR3를 포함하는 경쇄 가변영역을 포함하는 항체.(d) a heavy chain variable region comprising a heavy chain CDR1 as set out in SEQ ID NO: 19, a heavy chain CDR2 as set out in SEQ ID NO: 20, and a heavy chain CDR3 as set out in SEQ ID NO: 21, a light chain CDR1 as set out in SEQ ID NO: 22, a light chain CDR2 as set out in SEQ ID NO: 23, and An antibody comprising a light chain variable region comprising the light chain CDR3 set forth in SEQ ID NO: 24.
보다 구체적으로, 상기 단일클론항체는 하기 (a) 내지 (d)로 이루어진 군으로부터 선택되는 어느 하나의 항체인 것이 바람직하다.More specifically, the monoclonal antibody is preferably any one antibody selected from the group consisting of (a) to (d).
(a) 서열번호 25로 기재된 중쇄 가변영역 및 서열번호 26으로 기재된 경쇄 가변영역을 포함하는 항체;(a) an antibody comprising a heavy chain variable region as set out in SEQ ID NO: 25 and a light chain variable region as set out in SEQ ID NO: 26;
(b) 서열번호 27로 기재된 중쇄 가변영역 및 서열번호 28로 기재된 경쇄 가변영역을 포함하는 항체;(b) an antibody comprising a heavy chain variable region as set out in SEQ ID NO: 27 and a light chain variable region as set out in SEQ ID NO: 28;
(c) 서열번호 29로 기재된 중쇄 가변영역 및 서열번호 30로 기재된 경쇄 가변영역을 포함하는 항체;(c) an antibody comprising a heavy chain variable region as set out in SEQ ID NO: 29 and a light chain variable region as set out in SEQ ID NO: 30;
(d) 서열번호 31로 기재된 중쇄 가변영역 및 서열번호 32로 기재된 경쇄 가변영역을 포함하는 항체.(d) an antibody comprising a heavy chain variable region as set out in SEQ ID NO: 31 and a light chain variable region as set out in SEQ ID NO: 32.
보다 구체적으로는 단일클론항체의 중쇄 가변부위가 서열번호 25, 27, 29 및 31로 이루어진 군으로부터 선택된 1종 이상의 아미노산 서열을 포함하거나 또는 CDR 영역 내에서 이러한 서열과의 동일성이 80% 이상, 바람직하게는 90% 이상, 가장 바람직하게 95% 이상인 서열을 포함하거나; 또는/ 및 그들의 경쇄 가변부위가 서열번호 26, 28, 30 및 32로 이루어진 군으로부터 선택된 1종 이상의 아미노산 서열을 포함하거나 또는 CDR 영역 내에서 이러한 서열과의 동일성이 80% 이상, 바람직하게는 90% 이상, 가장 바람직하게는 95% 이상인 서열을 포함할 수 있다.More specifically, the heavy chain variable region of the monoclonal antibody comprises at least one amino acid sequence selected from the group consisting of SEQ ID NOs: 25, 27, 29 and 31 or has at least 80% identity with such sequence in the CDR region, preferably Preferably at least 90%, most preferably at least 95%; Or / and their light chain variable regions comprise at least one amino acid sequence selected from the group consisting of SEQ ID NOs: 26, 28, 30 and 32 or have at least 80% identity, preferably 90% identity with such sequence in the CDR region Or more preferably 95% or more.
본 발명의 단일클론항체는 생물학적 시료와 반응시키고 항원-항체 복합체 형성을 검출함으로써 프로피오니 박테리움 아크네스(Propioni
- bacterium
acnes
: P. acnes)를 검출하는 데 사용할 수 있다.Monoclonal antibodies of the invention can be used to detect Propioni -bacterium acnes (P. acnes ) by reacting with biological samples and detecting antigen-antibody complex formation.
상기 "항원-항체 복합체"는 시료 중의 P.
acnes 단백질 항원과 이를 인지하는 본 발명에 따른 단일클론항체의 결합물을 의미하며, 이러한 항원-항체 복합체의 형성은 비색법(colormetric method), 전기화학법(electrochemical method), 형광법(fluorimetric method), 발광법(luminometry), 입자계수법(particle counting method), 육안측정법(visual assessment) 및 섬광계수법(scintillation counting method)으로 이루어진 그룹 중에서 선택되는 임의의 방법으로 검출할 수 있다. 그러나 반드시 이들로만 제한되지 않고 다양한 응용과 적용이 가능하다.The "antigen-antibody complex" means a combination of the P. acnes protein antigen in the sample and the monoclonal antibody according to the present invention which recognizes the same. The formation of such antigen-antibody complex is performed by colormetric method and electrochemical method. detection by any method selected from the group consisting of electrochemical method, fluorimetric method, luminometry, particle counting method, visual assessment and scintillation counting method can do. However, it is not necessarily limited to these and various applications and applications are possible.
본 발명에서는 항원-항체 복합체를 검출하기 위하여 여러 가지 표지체를 사용할 수 있다. 구체적인 예로는 효소, 형광물, 리간드, 발광물, 미소입자, 방사성 동위원소로 이루어진 그룹 중에서 선택될 수 있으며, 반드시 이들로만 국한되지는 않는다. 바람직하게는, 항원-항체 복합체를 효소면역흡착법(ELISA)을 이용하여 검출할 수 있다. 효소면역흡착법(ELISA)에는 고체 지지체에 부착된 항원을 인지하는 표지된 항체를 이용하는 직접적 ELISA, 고체 지지체에 부착된 항원을 인지하는 항체의 복합체에서 포획 항체를 인지하는 표지된 이차 항체를 이용하는 간접적 ELISA, 고체 지지체에 부착된 항체와 항원의 복합체에서 항원을 인지하는 표지된 또 다른 항체를 이용하는 직접적 샌드위치 ELISA, 고체 지지체에 부착된 항체와 항원의 복합체에서 항원을 인지하는 또 다른 항체와 반응시킨 후 이 항체를 인지하는 표지된 2차 항체를 이용하는 간접적 샌드위치 ELISA 등 다양한 ELISA 방법을 포함한다. 상기 단일클론 항체는 검출 표지를 가질 수 있으며, 검출표지를 가지지 않을 경우는 이들 단일클론 항체를 포획할 수 있고 검출 표지를 가지는 또 다른 항체를 처리하여 확인할 수 있다.In the present invention, various labels can be used to detect the antigen-antibody complex. Specific examples may be selected from the group consisting of enzymes, fluorescent materials, ligands, luminescent materials, microparticles, and radioisotopes, but are not limited thereto. Preferably, the antigen-antibody complex can be detected using enzyme immunosorbent adsorption (ELISA). Enzyme immunosorbent methods (ELISA) include direct ELISA using labeled antibodies that recognize antigens attached to a solid support, and indirect ELISAs using labeled secondary antibodies that recognize capture antibodies in a complex of antibodies that recognize antigens attached to a solid support. Direct sandwich ELISA using another labeled antibody that recognizes the antigen in the complex of the antibody and the antibody attached to the solid support, followed by reaction with another antibody that recognizes the antigen in the complex of the antibody and the antigen attached to the solid support. Various ELISA methods include indirect sandwich ELISAs using labeled secondary antibodies that recognize the antibody. The monoclonal antibody may have a detection label, and if it does not have a detection label, the monoclonal antibody may be captured and can be identified by treating another antibody having the detection label.
또한, 본 발명은 본 발명에 따른 단일클론항체를 암호화하는 폴리뉴클레오치드를 포함하는, 발현 벡터를 제공한다.The present invention also provides an expression vector comprising a polynucleotide encoding a monoclonal antibody according to the present invention.
또한, 본 발명은 상기 발현 벡터로 형질전환된 형질전환체를 제공한다.The present invention also provides a transformant transformed with the expression vector.
본 발명의 항체를 암호화하는 폴리뉴클레오티드는 코돈의 축퇴성(degeneracy)으로 인하여 또는 상기 항체를 발현시키고자 하는 생물에서 선호되는 코돈을 고려하여, 코딩영역으로부터 발현되는 항체의 아미노산 서열을 변화시키지 않는 범위 내에서 코딩영역에 다양한 변형이 이루어질 수 있고, 코딩영역을 제외한 부분에서도 유전자의 발현에 영향을 미치지 않는 범위 내에서 다양한 변형 또는 수식이 이루어질 수 있으며, 그러한 변형 유전자 역시 본 발명의 범위에 포함됨을 당업자는 잘 이해할 수 있을 것이다. 즉, 본 발명의 폴리뉴클레오티드는 이와 동등한 활성을 갖는 단백질을 코딩하는 한, 하나 이상의 핵산 염기가 치환, 결실, 삽입 또는 이들의 조합에 의해 변이될 수 있으며, 이들 또한 본 발명의 범위에 포함된다. 이러한 폴리뉴클레오티드의 서열은 단쇄 또는 이중쇄일 수 있으며, DNA 분자 또는 RNA(mRNA)분자일 수 있다.The polynucleotide encoding the antibody of the present invention does not change the amino acid sequence of the antibody expressed from the coding region due to degeneracy of the codon or in consideration of the codon preferred in the organism to express the antibody. Various modifications may be made to the coding region within the region, and various modifications or modifications may be made within a range that does not affect the expression of the gene even in portions other than the coding region, and such modified genes are also included in the scope of the present invention. Will be well understood. That is, as long as the polynucleotide of the present invention encodes a protein having equivalent activity, one or more nucleic acid bases may be mutated by substitution, deletion, insertion, or a combination thereof, which are also included in the scope of the present invention. The sequence of such polynucleotides may be single or double chained and may be DNA molecules or RNA (mRNA) molecules.
상기 발현벡터의 제작 시에는, 상기 항체를 생산하고자 하는 숙주세포의 종류에 따라 프로모터(promoter), 종결자(terminator), 인핸서(inhancer) 등과 같은 발현조절 서열, 막 표적화 또는 분비를 위한 서열 등을 적절히 선택하고 목적에 따라 다양하게 조합할 수 있다.When preparing the expression vector, expression control sequences such as promoters, terminators, enhancers, and the like, sequences for membrane targeting or secretion, etc., depending on the type of host cell in which the antibody is to be produced. It can be appropriately selected and various combinations depending on the purpose.
본 발명의 발현벡터는 플라스미드 벡터, 코즈미드 벡터, 박테리오 파아지 벡터 및 바이러스 벡터 등을 포함하나 이에 제한되지 않는다. 적합한 발현벡터는 프로모터, 오퍼레이터, 개시코돈, 종결코돈, 폴리아데닐화 시그널 및 인핸서 같은 발현 조절 엘리먼트 외에도 막 표적화 또는 분비를 위한 시그널 서열 또는 리더 서열을 포함하며 목적에 따라 다양하게 제조될 수 있다. 발현벡터의 프로모터는 구성적 또는 유도성일 수 있다. 상기 시그널 서열에는 숙주가 에쉐리키아속(Escherichia sp.)균인 경우에는 PhoA 시그널 서열, OmpA 시그널 서열 등이, 숙주가 바실러스속(Bacillus sp.)균인 경우에는 α-아밀라아제 시그널 서열, 서브틸리신 시그널 서열 등이, 숙주가 효모(yeast)인 경우에는 MFα 시그널 서열, SUC2 시그널 서열 등이, 숙주가 동물세포인 경우에는 인슐린 시그널 서열, α-인터페론 시그널 서열, 항체 분자 시그널 서열 등을 이용할 수 있으나, 이에 제한되지 않는다. 또한 발현벡터는 벡터를 함유하는 숙주 세포를 선택하기 위한 선택 마커를 포함할 수 있고, 복제 가능한 발현벡터인 경우 복제 기원을 포함한다.Expression vectors of the present invention include, but are not limited to, plasmid vectors, cosmid vectors, bacteriophage vectors, and viral vectors. Suitable expression vectors include signal sequences or leader sequences for membrane targeting or secretion in addition to expression control elements such as promoters, operators, initiation codons, termination codons, polyadenylation signals and enhancers and can be prepared in various ways depending on the purpose. The promoter of the expression vector may be constitutive or inducible. The signal sequence includes a PhoA signal sequence and an OmpA signal sequence when the host is Escherichia sp., And an α-amylase signal sequence and a subtilisin signal when the host is Bacillus sp. For example, the MFα signal sequence, the SUC2 signal sequence, and the like may be used when the host is a yeast, and the insulin signal sequence, the α-interferon signal sequence, the antibody molecular signal sequence, or the like may be used when the host is an animal cell. This is not restrictive. The expression vector may also include a selection marker for selecting a host cell containing the vector, and in the case of a replicable expression vector, includes the origin of replication.
또한, 본 발명은 1) 상기 형질전환체를 배지에 접종하고 배양하는 단계; 및In addition, the present invention comprises the steps of 1) inoculating and transforming the transformant in the medium; And
2) 상기 단계 1)에서 배양한 배양액으로부터 P.
acnes에 특이적으로 결합하는 단일클론항체를 정제하는 단계를 포함하는, 단일클론항체의 제조 방법을 제공한다.2) provides a method for producing a monoclonal antibody comprising the step of purifying a monoclonal antibody that specifically binds to P. acnes from the culture medium cultured in step 1).
본 발명에 따른 상기 발현벡터를 적절한 숙주 세포, 예를 들어, 대장균 또는 효모 세포 등에 형질전환시킨 후, 형질전환된 숙주세포를 배양함으로써 본 발명에 따른 항체를 대량 생산할 수 있다. 숙주세포의 종류에 따른 적절한 배양 방법 및 배지 조건 등은 당해 분야의 통상의 기술자에게 알려진 공지 기술로부터 당업자가 용이하게 선택할 수 있다. 상기 숙주세포는 대장균(E.
coli) 또는 바실러스 서브틸러스(Bacillus subtilis)와 같은 원핵 생물일 수 있다. 또한, 사카로마이세스 세르비시아(Saccharomyces
cerevisiae)와 같은 효모, 곤충 세포, 식물 세포, 동물 세포로부터 유래한 진핵 세포일 수 있다. 더욱 바람직하게는, 상기 동물 세포는 인간 유래의 세포주일 수 있다. 상기 숙주세포로의 발현벡터 도입방법은 당업자에게 공지된 어느 방법을 사용해도 무방하다.The expression vector according to the present invention may be transformed into an appropriate host cell, for example, E. coli or yeast cell, and then cultured in the transformed host cell, thereby producing a large amount of the antibody according to the present invention. Suitable culture methods, media conditions, and the like according to the type of host cell can be easily selected by those skilled in the art from known techniques known to those skilled in the art. The host cell may be a prokaryote such as E. coli or Bacillus subtilis . It may also be eukaryotic cells derived from yeast, insect cells, plant cells, animal cells such as Saccharomyces cerevisiae . More preferably, the animal cell may be a cell line derived from human. The expression vector introduction method into the host cell may be any method known to those skilled in the art.
또한, 본 발명은 본 발명에 따른 P.
acnes에 특이적으로 결합하는 단일클론항체를 유효성분으로 포함하는, 여드름에 대한 예방 또는 개선용 화장료 조성물을 제공한다.The invention also relates to P. It provides a cosmetic composition for preventing or improving acne comprising a monoclonal antibody that specifically binds to acnes as an active ingredient.
또한, 본 발명은 상기 단일클론항체를 여드름에 대한 예방 또는 개선용 화장료 조성물에 사용하기 위한 용도를 제공한다.In addition, the present invention provides a use for the use of the monoclonal antibody in the cosmetic composition for preventing or improving acne.
본 발명의 화장료 조성물은 상기 인간 항체 이외에 화장료 조성물에 통상적으로 이용되는 성분들이 포함되며, 예켠대 황산화제, 안정화제, 용해화제, 비타민, 안료 및 향료와 같은 통상적인 보조제, 그리고 담체를 포함한다. The cosmetic composition of the present invention includes components commonly used in cosmetic compositions in addition to the human antibody, and includes, for example, conventional auxiliaries such as sulfates, stabilizers, solubilizers, vitamins, pigments and flavors, and carriers.
본 발명의 화장료 조성물에 있어서, 통상적으로 함유되는 화장료 조성물에 대한 본 발명의 단일클론항체는 0.1 내지 50 중량%, 바람직하게는 1 내지 10 중량%의 양으로 첨가될 수 있다. In the cosmetic composition of the present invention, the monoclonal antibody of the present invention to the cosmetic composition usually contained may be added in an amount of 0.1 to 50% by weight, preferably 1 to 10% by weight.
본 발명의 화장료 조성물은 당업계에서 통상적으로 제조되는 어떠한 제형으로도 제조될 수 있으며, 예를 들어, 용액, 현탁액, 유탁액, 페이스트, 겔, 크림, 로션, 파우더, 비누, 계면활성제-함유 클렌징, 오일, 분말 파운데이션, 유탁액 파운데이션, 왁스 파운데이션 및 스프레이 등으로 제형화될 수 있으나, 이에 한정되는 것은 아니다. 보다 상세하게는, 유연 화장수(스킨), 영양 화장수(밀크로션), 영양 크림, 맛사지 크림, 에센스, 아이크림, 클렌징 크림, 클렌징 포옴, 클렌징 워터, 팩, 스프레이 또는 파우더의 제형으로 제조될 수 있다. Cosmetic compositions of the present invention may be prepared in any formulation conventionally prepared in the art and include, for example, solutions, suspensions, emulsions, pastes, gels, creams, lotions, powders, soaps, surfactant-containing cleansing , Oils, powder foundations, emulsion foundations, wax foundations and sprays, and the like, but are not limited thereto. More specifically, it may be prepared in the form of a flexible lotion (skin), nutrition lotion (milk lotion), nutrition cream, massage cream, essence, eye cream, cleansing cream, cleansing foam, cleansing water, pack, spray or powder .
본 발명의 제형이 페이스트, 크림 또는 겔인 경우에는 담체 성분으로서 동물성 유, 식물성 유, 왁스, 파라핀, 전분, 트라가칸타, 셀룰로오스 유도체, 폴리에틸렌 글리콜, 실리콘, 벤토나이트, 실리카, 탈크 또는 산화아연 등이 이용될 수 있으며, 상기 제형이 파우더 또는 스프레이인 경우에는 담체 성분으로서 락토스, 탈크, 실리카, 알루미늄 히드록시드, 칼슘 실리케이트 또는 폴리아미드 파우더가 이용될 수 있고, 특히 스프레이인 경우에는 추가적으로 클로로플루오로히드로카본, 프로판/부탄 또는 디메틸 에테르와 같은 추진체를 포함할 수 있다.When the formulation of the present invention is a paste, cream or gel, animal oil, vegetable oil, wax, paraffin, starch, tragacanta, cellulose derivative, polyethylene glycol, silicone, bentonite, silica, talc or zinc oxide are used as carrier components. If the formulation is a powder or spray, lactose, talc, silica, aluminum hydroxide, calcium silicate or polyamide powder may be used as the carrier component, especially in the case of a spray, additionally chlorofluorohydrocarbons. Propellant such as propane / butane or dimethyl ether.
본 발명의 제형이 용액 또는 유탁액인 경우에는 담체 성분으로서 용매, 용해화제 또는 유탁화제가 이용되고, 예컨대 물, 에탄올, 이소프로판올, 에틸 카보네이트, 에틸 아세테이트, 벤질 알코올, 벤질 벤조에이트, 프로필렌글리콜, 1,3-부틸글리콜 오일, 글리세롤 지방족 에스테르, 폴리에틸렌 글리콜 또는 소르비탄의 지방산 에스테르가 있다. When the formulation of the present invention is a solution or emulsion, a solvent, solubilizer or emulsifier is used as the carrier component, such as water, ethanol, isopropanol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene glycol, 1 Fatty acid esters of, 3-butylglycol oil, glycerol aliphatic ester, polyethylene glycol or sorbitan.
본 발명의 제형이 현탁액인 경우에는 담체 성분으로서 물, 에탄올 또는 프로필렌 글리콜과 같은 액상의 희석제, 에톡실화 이소스테아릴 알코올, 폴리옥시에틸렌 소르비톨 에스테르 및 폴리옥시에틸렌 소르비탄 에스테르와 같은 현탁제, 미소 결정성 셀룰로오스, 알루미늄 메타히드록시드, 벤토나이트, 아가 또는 트라가칸타 등이 이용될 수 있다.When the formulation of the present invention is a suspension, liquid carrier diluents such as water, ethanol or propylene glycol, suspending agents such as ethoxylated isostearyl alcohol, polyoxyethylene sorbitol esters and polyoxyethylene sorbitan esters, and microcrystals are used as carrier components. Soluble cellulose, aluminum metahydroxy, bentonite, agar or tragacanta and the like can be used.
본 발명의 제형이 계면-활성제 함유 클렌징인 경우에는 담체 성분으로서 지방족 알코올 설페이트, 지방족 알코올 에테르 설페이트, 설포숙신산 모노에스테르, 이세티오네이트, 이미다졸리늄 유도체, 메틸타우레이트, 사르코시네이트, 지방산 아미드 에테르 설페이트, 알킬아미도베타인, 지방족 알코올, 지방산 글리세리드, 지방산 디에탄올아미드, 식물성 유, 라놀린 유도체 또는 에톡실화 글리세롤 지방산 에스테르 등이 이용될 수 있다.When the formulation of the present invention is a surfactant-containing cleansing, the carrier component is aliphatic alcohol sulfate, aliphatic alcohol ether sulfate, sulfosuccinic acid monoester, isethionate, imidazolinium derivative, methyltaurate, sarcosinate, fatty acid amide. Ether sulfates, alkylamidobetaines, aliphatic alcohols, fatty acid glycerides, fatty acid diethanolamides, vegetable oils, lanolin derivatives or ethoxylated glycerol fatty acid esters and the like can be used.
또한, 본 발명은 본 발명에 따른 P.
acnes에 특이적으로 결합하는 단일클론항체를 유효성분으로 포함하는, 여드름에 대한 예방 또는 치료용 약학적 조성물을 제공한다.The invention also relates to P. It provides a pharmaceutical composition for preventing or treating acne, comprising a monoclonal antibody that specifically binds to acnes as an active ingredient.
또한, 본 발명은 상기 단일클론항체를 포함하는 여드름에 대한 예방 또는 치료용 약학적 조성물의 용도를 제공한다.The present invention also provides the use of a pharmaceutical composition for the prevention or treatment of acne comprising the monoclonal antibody.
본 발명의 단일클론항체는 임상투여시 비경구로 투여가 가능하며 일반적인 의약품 제제의 형태로 사용될 수 있다. 비경구적인 투여방법으로는 이에 제한되는 것은 아니나, 정맥내, 근육내, 동맥내, 골수내, 경막내, 심장내, 경피, 피하, 복강내, 비강내, 장관, 국소, 설하 또는 직장내 투여일 수 있다.The monoclonal antibodies of the present invention can be administered parenterally during clinical administration and can be used in the form of general pharmaceutical formulations. Parenteral methods of administration include, but are not limited to, intravenous, intramuscular, intraarterial, intramedullary, intradural, intracardiac, transdermal, subcutaneous, intraperitoneal, intranasal, intestinal, topical, sublingual or rectal administration. Can be.
비경구적으로 투여하는 경우 본 발명의 약학적 조성물은 적합한 비경구용 담체와 함께 주사제, 경피 투여제 및 비강 흡입제의 형태로 당 업계에 공지된 방법에 따라 제형화될 수 있다. 상기 주사제의 경우에는 반드시 멸균되어야 하며 박테리아 및 진균과 같은 미생물의 오염으로부터 보호되어야 한다. 주사제의 경우 적합한 담체의 예로는 이에 한정되지는 않으나, 물, 에탄올, 폴리올(예를 들어, 글리세롤, 프로필렌 글리콜 및 액체 폴리에틸렌 글리콜 등), 이들의 혼합물 및/또는 식물유를 포함하는 용매 또는 분산매질일 수 있다. 보다 바람직하게는, 적합한 담체로는 행크스 용액, 링거 용액, 트리에탄올 아민이 함유된 phosphate buffered saline (PBS) 또는 주사용 멸균수, 10% 에탄올, 40% 프로필렌 글리콜 및 5% 덱스트로즈와 같은 등장 용액 등을 사용할 수 있다. 상기 주사제를 미생물 오염으로부터 보호하기 위해서는 파라벤, 클로로부탄올, 페놀, 소르빈산, 티메로살 등과 같은 다양한 항균제 및 항진균제를 추가로 포함할 수 있다. 또한, 상기 주사제는 대부분의 경우 당 또는 나트륨 클로라이드와 같은 등장화제를 추가로 포함할 수 있다.When administered parenterally, the pharmaceutical compositions of the present invention may be formulated according to methods known in the art in the form of injections, transdermal and nasal inhalants with suitable parenteral carriers. Such injections must be sterile and protected from contamination of microorganisms such as bacteria and fungi. Examples of suitable carriers for injections include, but are not limited to, solvents or dispersion media comprising water, ethanol, polyols (e.g., glycerol, propylene glycol and liquid polyethylene glycols, etc.), mixtures thereof and / or vegetable oils Can be. More preferably, suitable carriers include Hanks' solution, Ringer's solution, phosphate buffered saline (PBS) containing triethanol amine or sterile water for injection, 10% ethanol, 40% propylene glycol and 5% dextrose Etc. can be used. In order to protect the injection from microbial contamination, various antibacterial and antifungal agents such as parabens, chlorobutanol, phenol, sorbic acid, thimerosal, and the like may be further included. In addition, the injection may in most cases further comprise an isotonic agent such as sugar or sodium chloride.
경피 투여제의 경우 연고제, 크림제, 로션제, 겔제, 외용액제, 파스타제, 리니멘트제, 에어롤제 등의 형태가 포함된다. 상기에서 '경피 투여'는 약학적 조성물을 국소적으로 피부에 투여하여 약학적 조성물에 함유된 유효한 양의 활성성분이 피부 내로 전달되는 것을 의미한다. In the case of transdermal administrations, ointments, creams, lotions, gels, external preparations, pasta preparations, linen preparations, air rolls and the like are included. As used herein, 'transdermal administration' means that the pharmaceutical composition is locally administered to the skin so that an effective amount of the active ingredient contained in the pharmaceutical composition is delivered into the skin.
흡입 투여제의 경우, 본 발명에 따라 사용되는 화합물은 적합한 추진제, 예를 들면, 디클로로플루오로메탄, 트리클로로플루오로메탄, 디클로로테트라플루오로에탄, 이산화탄소 또는 다른 적합한 기체를 사용하여, 가압 팩 또는 연무기로부터 에어로졸 스프레이 형태로 편리하게 전달 할 수 있다. 가압 에어로졸의 경우, 투약 단위는 계량된 양을 전달하는 밸브를 제공하여 결정할 수 있다. 예를 들면, 흡입기 또는 취입기에 사용되는 젤라틴 캡슐 및 카트리지는 화합물, 및 락토즈 또는 전분과 같은 적합한 분말 기제의 분말 혼합물을 함유하도록 제형화할 수 있다. 비경구 투여용 제형은 모든 제약 화학에 일반적으로 공지된 처방서인 문헌(Remington's Pharmaceutical Science, 15th Edition, 1975. Mack Publishing Company, Easton, Pennsylvania 18042, Chapter 87: Bla?g, Seymour)에 기재되어 있다.In the case of inhaled dosages, the compounds used according to the invention may be pressurized packs or by means of suitable propellants, for example dichlorofluoromethane, trichlorofluoromethane, dichlorotetrafluoroethane, carbon dioxide or other suitable gas. It can be delivered conveniently from the nebulizer in the form of an aerosol spray. In the case of a pressurized aerosol, the dosage unit can be determined by providing a valve to deliver a metered amount. For example, gelatin capsules and cartridges for use in inhalers or blowers can be formulated to contain a mixture of the compound and a suitable powder base such as lactose or starch. Formulations for parenteral administration are described in Remington's Pharmaceutical Science, 15th Edition, 1975. Mack Publishing Company, Easton, Pennsylvania 18042, Chapter 87: Blag, Seymour, a prescription generally known in all pharmaceutical chemistries.
본 발명의 약학적 조성물은 본 발명에 따른 단일클론항체를 유효량으로 포함할 때 바람직한 여드름 예방 또는 치료의 효과를 제공할 수 있다. 본 명세서에서, '유효량'이라 함은 음성 대조군에 비해 그 이상의 반응을 나타내는 양을 말하며 바람직하게는 P.
acnes 균의 사멸 또는 여드름 감소의 효과를 나타내기에 충분한 양을 말한다. 본 발명의 약학적 조성물에 본 발명에 따른 단일클론항체가 0.01 내지 99.99% 포함될 수 있으며, 잔량은 약학적으로 허용 가능한 담체가 차지할 수 있다. 본 발명의 약학적 조성물에 포함되는 본 발명에 따른 단일클론항체의 유효량은 조성물이 제품화되는 형태 등에 따라 달라질 것이다.The pharmaceutical composition of the present invention may provide the desired effect of preventing or treating acne when it contains an effective amount of a monoclonal antibody according to the present invention. In the present specification, the 'effective amount' refers to an amount that exhibits a higher response than a negative control, and preferably refers to an amount sufficient to exhibit an effect of killing P. acnes or reducing acne. The pharmaceutical composition of the present invention may contain 0.01 to 99.99% monoclonal antibody according to the present invention, the remaining amount may be occupied by a pharmaceutically acceptable carrier. The effective amount of the monoclonal antibody according to the present invention included in the pharmaceutical composition of the present invention will vary depending on the form in which the composition is commercialized and the like.
본 발명의 약학적 조성물의 총 유효량은 단일 투여량(single dose)으로 환자에게 투여될 수 있으며, 다중 투여량(multiple dose)으로 장기간 투여되는 분할 치료 방법(fractionated treatment protocol)에 의해 투여될 수 있다. 본 발명의 약학적 조성물은 질환의 정도에 따라 유효성분의 함량을 달리할 수 있다. 비경구 투여시는 상기 본 발명에 따른 단일클론항체를 기준으로 하루에 체중 1 kg당 바람직하게 0.01 내지 50 mg, 더 바람직하게는 0.1 내지 30 mg의 양으로 투여되도록, 그리고 경구 투여시는 본 발명에 따른 단일클론항체를 기준으로 하루에 체중 1 kg당 바람직하게 0.01 내지 100 mg, 더 바람직하게는 0.01 내지 10 mg의 양으로 투여되도록 1 내지 수회에 나누어 투여할 수 있다. 그러나 상기 본 발명에 따른 단일클론항체의 용량은 약학적 조성물의 투여 경로 및 치료 횟수뿐만 아니라 환자의 연령, 체중, 건강 상태, 성별, 질환의 중증도, 식이 및 배설율 등 다양한 요인들을 고려하여 환자에 대한 유효 투여량이 결정되는 것이므로, 이러한 점을 고려할 때 당 분야의 통상적인 지식을 가진 자라면 본 발명에 따른 단일클론항체를 여드름 예방 또는 치료를 위한 특정한 용도에 따른 적절한 유효 투여량을 결정할 수 있을 것이다. 본 발명에 따른 약학적 조성물은 본 발명의 효과를 보이는 한 그 제형, 투여 경로 및 투여 방법에 특별히 제한되지 아니한다.The total effective amount of the pharmaceutical composition of the present invention may be administered to a patient in a single dose and may be administered by a fractionated treatment protocol which is administered in multiple doses for a long time. . The pharmaceutical composition of the present invention may vary the content of the active ingredient depending on the extent of the disease. When administered parenterally, the present invention is administered in an amount of preferably 0.01 to 50 mg, more preferably 0.1 to 30 mg per kg of body weight per day, based on the monoclonal antibody according to the present invention, and when administered orally. Based on the monoclonal antibody according to the present invention may be administered in several divided doses so as to be administered in an amount of preferably 0.01 to 100 mg, more preferably 0.01 to 10 mg per kg body weight per day. However, the dose of the monoclonal antibody according to the present invention may be adjusted to the patient in consideration of various factors such as the age, weight, health condition, sex, severity of the disease, diet and excretion rate, as well as the route of administration and the frequency of treatment of the pharmaceutical composition. As the effective dosage is determined, one of ordinary skill in the art will be able to determine the appropriate effective dosage for the specific use of the monoclonal antibody according to the invention for the prevention or treatment of acne. . The pharmaceutical composition according to the present invention is not particularly limited to its formulation, route of administration and method of administration as long as the effect of the present invention is shown.
본 발명의 약학적 조성물은 단독으로, 또는 수술, 방사선 치료, 호르몬 치료, 화학 치료 또는 생물학적 반응조절제를 사용하는 방법들과 병용하여 사용할 수 있다.The pharmaceutical composition of the present invention may be used alone or in combination with methods using surgery, radiation therapy, hormone therapy, chemotherapy or biological response modifiers.
본 발명의 약학적 조성물은 또한 본 발명에 따른 단일클론항체를 유효성분으로 포함하는 외용제의 제형으로 제공할 수 있다.The pharmaceutical composition of the present invention may also be provided in the formulation of an external preparation containing the monoclonal antibody according to the present invention as an active ingredient.
본 발명의 약학적 조성물을 피부외용제로 사용하는 경우, 추가로 지방 물질, 유기 용매, 용해제, 농축제 및 겔화제, 연화제, 항산화제, 현탁화제, 안정화제, 발포제(foaming agent), 방향제, 계면활성제, 물, 이온형 유화제, 비이온형 유화제, 충전제, 금속이온봉쇄제, 킬레이트화제, 보존제, 비타민, 차단제, 습윤화제, 필수 오일, 염료, 안료, 친수성 활성제, 친유성 활성제 또는 지질 소낭 등 피부 외용제에 통상적으로 사용되는 임의의 다른 성분과 같은 피부 과학 분야에서 통상적으로 사용되는 보조제를 함유할 수 있다. 또한 상기 성분들은 피부 과학 분야에서 일반적으로 사용되는 양으로 도입될 수 있다.When the pharmaceutical composition of the present invention is used as an external preparation for skin, it is additionally used for fatty substances, organic solvents, solubilizers, thickening and gelling agents, emollients, antioxidants, suspending agents, stabilizers, foaming agents, fragrances, interfaces. Skin such as active agents, water, ionic emulsifiers, nonionic emulsifiers, fillers, metal ion sequestrants, chelating agents, preservatives, vitamins, blockers, wetting agents, essential oils, dyes, pigments, hydrophilic active agents, lipophilic active agents or lipid vesicles It may contain adjuvants commonly used in the field of dermatology, such as any other ingredients commonly used in external preparations. The ingredients may also be introduced in amounts generally used in the field of dermatology.
본 발명의 약학적 조성물이 피부 외용제로 제공될 경우, 이에 제한되는 것은 아니나, 연고, 패취, 겔, 크림 또는 분무제 등의 제형일 수 있다.When the pharmaceutical composition of the present invention is provided as an external preparation for skin, it may be a formulation such as, but not limited to, an ointment, a patch, a gel, a cream, or a spray.
이하, 실시예를 통하여 본 발명을 더욱 상세히 설명하고자 한다. 이들 실시예는 오로지 본 발명을 예시하기 위한 것으로서, 본 발명의 범위가 이들 실시예에 의해 제한되는 것으로 해석되지 않는 것은 당업계에서 통상의 지식을 가진 자에 있어서 자명할 것이다.Hereinafter, the present invention will be described in more detail with reference to Examples. These examples are only for illustrating the present invention, it will be apparent to those skilled in the art that the scope of the present invention is not to be construed as limited by these examples.
[[
실시예Example
1] One]
프로피오니Propioni
박테리움Bacterium
아크네스( Acnes
PropioniPropioni
- bacterium -bacterium
acnesacnes
: :
PP
..
acnesacnes
)의 배양Culture
본 발명의 단일클론항체를 생산하기 위한 항원균으로서, 여드름을 유발하는 프로피오니박테리움 아크네스(P.
acnes
) 균을 배양하여 준비하였다. P.
acnes는 그람 양성의 세균으로 세포막에 특이적인 항원이 다수 존재하여, 병원균 자체를 특이적으로 인지하고 결합하는 인간 항체를 제조하고자 하였다.As an antigen bacterium for producing a monoclonal antibody of the present invention, propionibacterium acnes ( P. acnes ) bacteria were cultured to prepare. P. acnes is a Gram-positive bacterium, which has a large number of antigens specific to cell membranes.
고체배지에서 계대배양하기 위해, Reinforced Clostridial Medium(RCM 배지)에 P.
acnes를 도말하고, 혐기 배양기 또는 AnaeroPackTM-Anaero(A-04, MGC)를 넣고 밀봉한 박스에 넣고, 37에서 5일 동안 배양하였다(도 1a).For passage in solid medium, P. in Reinforced Clostridial Medium (RCM medium). The acnes were plated, placed in an anaerobic incubator or AnaeroPackTM-Anaero (A-04, MGC) in a sealed box and incubated for 37 to 5 days (FIG. 1A).
액체배지에서 배양하기 위해, Reinforced Clostridial broth(RCM 액체배지)를 튜브에 1.5 ㎖ 가한 다음 계대 배양한 P.
acnes의 콜로니를 접종한 후, 37에서 5일간 정치배양하였다. 배양한 배양액은 원심분리하여 균주를 수확하고, 50㎖ RCM 액체배지를 채운 원심분리기 튜브(centrifuge tube)에 수확한 균주를 풀어, 다시 37에서 5일간 정치배양 하였다(도 1b).To incubate in liquid medium, 1.5 ml of Reinforced Clostridial broth (RCM liquid medium) was added to the tube, followed by passage P. After inoculation with acnes colonies, 37-day incubation was performed. Cultured culture was centrifuged to harvest the strain, the strain was harvested in a centrifuge tube filled with 50ml RCM liquid medium, and further cultured for 37 to 5 days (FIG. 1b).
[[
실시예Example
2] 2]
PP
..
acnesacnes
에on
특이적으로 결합하는 Specifically bound
모노클론항체의Monoclonal antibody
제조 Produce
<2-1> 파지 디스플레이 기술을 이용한 <2-1> Using phage display technology
PP
..
acnes acnes
결합 항체 후보군 제작 Binding Antibody Candidate Construction
P. acnes에 대하여 특이적으로 결합하는 단일클론항체를 제조하기 위하여, 먼저 도 2의 모식도에서 나타내는 파지 디스플레이(phage display) 기술을 통하여 P. acnes 결합 항체 후보군을 제조하였다. P. In order for the acnes producing a monoclonal antibody that specifically binds to, first, even through the display phage (phage display) technology shown in the schematic diagram of P 2. acnes binding antibody candidates were prepared.
구체적으로, 2.7?1010의 다양성을 갖는 인간 유래 scFv 라이브러리 파지(library phage) 1 ㎖과 표피포도구균인 스타필로코커스 에피더미디스(Staphylococcus
epidermidis) 2 ㎖을 2 시간 동안 반응시켜 피부에 무해한 균에도 결합할 가능성이 있는 scFv-phage를 선 제거하였다(단계 1). 제거 후 남은 scFv-phage를 P.
acnes과 2 시간 동안 반응시켜 P.
acnes에만 특이적으로 결합하는 scFv-phage를 용출하여 poly-scFv-phage로서 확보하였다(단계 2). 확보한 Poly-scFv-파지를 XL1-Blue에 감염시켜 증폭하고, 증폭된 Poly-scFv-파지를 사용하여 상기 단계 1 및 단계 2의 과정을 총 3회 반복 수행하여 친화력이 좋은 파지만을 선택적으로 증폭시켰다(패닝 단계:panning step). 최종적으로 확보한 poly-scFv-phage를 희석하고 적정(titration)을 분석하고 증폭 수준을 확인하였다.Specifically, 1 ml of a human-derived scFv library phage with a variety of 2.7-10 10 and 2 ml of Staphylococcus epidermidis , an epidermis , were reacted for 2 hours. The scFv-phage, which is likely to bind, was pretreated (step 1). After removal by reacting the remaining scFv-phage P. acnes and for 2 hours to obtain a poly-scFv-phage to elute the scFv-phage that specifically bind only to P. acnes (step 2). The obtained Poly-scFv-phage was infected with XL1-Blue and amplified, and the amplified Poly-scFv-phage was repeated three times in total to repeat the steps 1 and 2 to selectively select only affinity phages. Amplified (panning step). Finally, the obtained poly-scFv-phage was diluted, titration was analyzed, and the amplification level was confirmed.
각각의 패닝 단계에서 증폭된 파지 후보군의 수준은 하기 [표 1]과 같다. P. acnes와 반응 전 1 ㎖의 반응액에 포함된 scFv-phage의 수는 각각 1?1013 개, 1.6?1013 개 및 5?1012 개였으며, P.
acnes과 반응하여 결합하는 것으로 수득한 scFv-phage의 수는 각각 2?104 개, 8?104 개 및 1?105 개였다(도 3).The level of the phage candidate group amplified in each panning step is shown in Table 1 below. The number of scFv-phage containing a reaction solution of P. acnes and the reaction before 1 ㎖ are each 1? 10 13 1.6? 10, 13, and 5? 10 was 12, obtained by combining react with P. acnes The number of one scFv-phage was 2-10 4 , 8-10 4 and 1-10 5 respectively (FIG. 3).
| 여드름 유발균Acne-causing bacteria | 1차Primary | 2차Secondary |
3차 |
| InputInput | 1?1013/ml1? 10 13 / ml |
1.6?1013/ml1.6? 10 13 / |
5?1012/ml5? 10 12 / |
| OutputOutput |
2?104/ml2? 10 4 / |
8?104/ml8? 10 4 / |
1?105/ml1? 10 5 / ml |
선별한 poly-scFv-phage가 P.
acnes와 친화력을 나타내지 확인하기 위하여, 각 패닝 단계에서의 poly-scFv-phage를 대상으로 ELISA 분석을 수행하였다.In order to confirm that the selected poly-scFv-phage showed affinity with P. acnes , ELISA analysis was performed on the poly-scFv-phage at each panning step.
ELISA 분석을 위해서, 먼저 P.
acnes 또는 대조군 세포(S.
epidermidis)를 배양하여 PBS로 세척하여 준비한 후, 흡광도가 0.5가 되도록 희석하여 96-웰 플레이트의 각 웰당 100 ㎕씩 분주하고, 4 ℃에서 16 시간 이상 배양하여 코팅하였다. 그런 다음, 모든 웰에 있는 반응액을 제거하고, 각 웰당 PBSA를 100 ㎕ 씩 분주한 후, 4℃에서 2 시간동안 배양하여 차단(blocking)한 후, PBSA를 제거하였다. 제거 후 웰에 poly-scFv-phage 포함 용액을 100 ㎕씩 분주하고 4 ℃에서 1 시강 동안 반응하였다. 그런 다음, 모든 웰에 있는 반응액을 제거하고, 희석한 PBST를 웰당 200 ㎕씩 분주하여 세척하는 단계를 2회 반복하였다. 세척이 완료된 후, 각 웰에 1:4000으로 희석한 hFc-HRP를 100 ㎕씩 분주하고 실온의 암실에서 10 분간 반응시켜 발색을 유도하였다. 푸른색으로 변하는 발색 반응을 종료시키기 위해 stop solution을 각 웰당 100㎕씩 분주하고, 반응물의 색이 노란색으로 변하면 마이크로플레이트 리더기를 이용하여 450 ㎚ 파장에서 각 웰별로 흡광도를 측정하였다.For ELISA analysis, first P. Acnes or control cells ( S. epidermidis ) were cultured and prepared by washing with PBS, diluted with an absorbance of 0.5, divided into 100 μl of each well of a 96-well plate, and incubated at 4 ° C. for at least 16 hours for coating. Then, the reaction solution in all the wells were removed, 100 μl of PBSA was dispensed for each well, and then incubated at 4 ° C. for 2 hours to block, and then PBSA was removed. After removal, 100 μl of a poly-scFv-phage-containing solution was dispensed into the wells and reacted at 4 ° C. for 1 hour. Then, the reaction solution in all the wells was removed, and the diluted PBST was dispensed twice by washing 200 μl per well. After the washing was completed, 100 μl of hFc-HRP diluted 1: 4000 in each well was dispensed and reacted for 10 minutes in the dark at room temperature to induce color development. In order to terminate the blue color reaction, the stop solution was dispensed by 100 μl per well, and when the color of the reactant turned yellow, the absorbance was measured for each well at 450 nm using a microplate reader.
그 결과, 하기 [표 2] 및 도 4에서 나타난 바와 같이, 패닝 수행의 횟수가 증가할수록 poly-scFv-phage와 P.
acnes의 결합력이 증가하는 것을 확인하였고, 반면 Control cell과는 결합하지 않는 것을 확인하였다.As a result, as shown in [Table 2] and FIG. 4, it was confirmed that as the number of panning increases, the binding force of poly-scFv-phage and P. acnes increases, whereas it does not bind to the control cell. Confirmed.
| 성분ingredient | Poly-scFv-파지Poly-scFv-phage | ||
| 1차Primary | 2차Secondary | 3차3rd | |
| 여드름 유발균Acne-causing bacteria | 1.75771.7577 | 1.96881.9688 | 2.30012.3001 |
| 대조군 세포Control cells | 0.00070.0007 | 0.00300.0030 | 0.00060.0006 |
<2-2> <2-2>
PP
..
acnes acnes
결합 항체 후보군으로부터 높은 결합력의 단일파지 선별 Single binding phage selection from binding antibody candidates
상기 <2-1> 단계에서 수득한 P.
acnes 결합 poly-scFv-phage으로부터, 높은 친화력을 나타내는 10 개의 단일 파지를 선별하였다. 선별한 단일 파지는 다시 ELISA를 수행하여 P. acnes 과의 결합력을 확인하였다(도 5).From the P. acnes binding poly-scFv-phage obtained in step <2-1>, 10 single phages showing high affinity were selected. The selected single phage was again subjected to ELISA to confirm the binding to P. acnes (Fig. 5).
| 번호number | 여드름 유발균Acne-causing bacteria (( O.DO.D .450㎚).450 nm) | 대조군 세포Control cells (( O.DO.D .450㎚).450 nm) |
| 1One | 0.64520.6452 | 0.00050.0005 |
| 22 | 0.5040.504 | 0.00050.0005 |
| 33 | 3.35633.3563 | 0.00020.0002 |
| 44 | 0.33940.3394 | 0.00050.0005 |
| 55 | 0.95090.9509 | 00 |
| 66 | 1.95201.9520 | 0.00410.0041 |
| 77 | 1.38531.3853 | 0.00130.0013 |
| 88 | 1.81531.8153 | 0.00440.0044 |
| 99 | 1.73361.7336 | 0.00120.0012 |
| 1010 | 1.80621.8062 | 0.00230.0023 |
<2-3> 선별한 단일파지의 서열 분석<2-3> Sequence Analysis of Selected Single Phage
확보된 10 개의 Mono-scFv-phage의 서열을 분석하여, 서열 상동성에 따라 군으로 나누었다. 그 결과, 하기 [표 4]에서 나타난 바와 같이 3, 6, 7, 8, 9, 10번 클론이 중복없이 서로 다른 CDR3을 갖는 것으로 확인하였다. 이들이 P.
acnes에 대하여 나타내는 결합력을 ELISA 분석으로 확인한 결과, 3, 6, 8 및 10 번의 총 4 개 클론이 가장 우수한 결합력을 나타내는 것으로 확인하였다.The sequences of the 10 Mono-scFv-phages obtained were analyzed and divided into groups according to sequence homology. As a result, as shown in Table 4, it was confirmed that clones 3, 6, 7, 8, 9 and 10 have different CDR3s without duplication. As a result of confirming the binding force to the P. acnes by ELISA analysis, it was confirmed that a total of four clones of 3, 6, 8 and 10 shows the best binding force.
| 번호number | VHVH | 동정값Identification | VLVL | 동정값Identification | VHVH (( CDR3CDR3 -a.a seq)-a.a seq) | VLVL (( CDR3CDR3 -a.a seq)-a.a seq) |
ELISA O.D. 값 |
| 33 | VH1-2VH1-2 | 272/295 (92.2%)272/295 (92.2%) | V3-4V3-4 | 281/293 (95.9%)281/293 (95.9%) | VKGLEHAAGSAIFDRVKGLEHAAGSAIFDR | ALSMGSGIWVALSMGSGIWV | 3.62343.6234 |
| 66 | VH3-20VH3-20 | 273/387 (95.1%)273/387 (95.1%) | A23A23 | 291/299 (97.3%)291/299 (97.3%) | STRHLHHSTRHLHH | VQAKQFPLTVQAKQFPLT | 2.21912.2191 |
| 77 | VH1-69VH1-69 | 267/304 (87.8%)267/304 (87.8%) | L5L5 | 257/286 (89.9%)257/286 (89.9%) | ARAVDTAMVGDSARAVDTAMVGDS | QQVDSYPLTQQVDSYPLT | 1.65241.6524 |
| 88 | VH3-9VH3-9 | 267/294 (90.8%)267/294 (90.8%) | L5L5 | 268/294 (94.7%)268/294 (94.7%) | TTDLGVVPAAIYAFDITTDLGVVPAAIYAFDI | QQTATFQITQQTATFQIT | 2.08242.0824 |
| 99 | VH3-74VH3-74 | 276/295 (93.6%)276/295 (93.6%) | A23A23 | 281/299 (94.0%)281/299 (94.0%) | ARDDGATWLHDYARDDGATWLHDY | ARDDGATWLHDYARDDGATWLHDY | 2.00072.0007 |
| 1010 | VH3-53VH3-53 | 262/284 (92.3%)262/284 (92.3%) | V2-14V2-14 | 252/289 (87.2%)252/289 (87.2%) | SCEGKAVSGSRDLHFEFSCEGKAVSGSRDLHFEF | QVWDSSSDHLIQVWDSSSDHLI | 2.07332.0733 |
[실시예 3] Example 3
P. acnesP. acnes
특이적인 인간 단일클론항체의 발현 및 정제 Expression and Purification of Specific Human Monoclonal Antibodies
P. acnes에 대하여 높은 친화력을 나타내는 것으로 수득한 4 개의 클론을 scFv 형태에서 인간 항체의 IgG 형태로 전환하기 위해 중쇄 및 경쇄를 각각 동물세포 발현 벡터인 pNATVH 와 pNATVL에 클로닝하였다(도 6). 클로닝한 발현 벡터는 대장균(E.
coli)에 형질전환하여, 벡터를 증폭하였다. 증폭된 8종의 플라스미드를 각각 HEK293F 세포에 공동-형질감염하고 6 일간 배양한 후, 친화도 크로마토그래피(Protein A affinity chromatography)를 수행하여 발현된 항체를 정제 회수하였다. 정제한 항체 단백질은 SDS-PAGE를 통해 분자량 및 순수 분리도를 확인하였다(도 7a). 그런 다음, 항-Fc-HRP 항체를 2차 항체로 사용하여 웨스턴 블럿을 수행하여, 인간화 단일클론항체의 제조를 검증하였다(도 7b).Four clones obtained with high affinity for P. acnes were cloned into the animal cell expression vectors pNATVH and pNATVL, respectively, to convert from the scFv form to the IgG form of the human antibody (FIG. 6). The cloned expression vector was transformed into E. coli to amplify the vector. Each of the eight amplified plasmids was co-transfected into HEK293F cells and incubated for 6 days, followed by affinity chromatography to recover and express the expressed antibody. Purified antibody protein was confirmed molecular weight and pure separation through SDS-PAGE (Fig. 7a). Then, Western blot was performed using anti-Fc-HRP antibody as the secondary antibody to verify the preparation of the humanized monoclonal antibody (FIG. 7B).
제조한 단일클론항체의 경쇄 및 중쇄 CDR 영역 서열, 및 경쇄 및 중쇄 가변영역 서열은 하기 [표 5]에 나타난 바와 같다.The light and heavy chain CDR region sequences, and the light and heavy chain variable region sequences of the prepared monoclonal antibodies are shown in Table 5 below.
| 항체명Antibody Name | 서열번호SEQ ID NO: | 서열명Sequence name |
아미노산 서열 |
|
| 3F3F |
1 | CDRH1CDRH1 | GYTFTDYYGYTFTDYY | |
|
2 | CDRH2CDRH2 | INPNSGAPINPNSGAP | ||
|
3 | CDRH3CDRH3 | VKGLEHAAGSAIFDRVKGLEHAAGSAIFDR | ||
|
4 | CDRL1CDRL1 | SGSVSTSHFSGSVSTSHF | ||
|
5 | CDRL2CDRL2 | FKDFKD | ||
|
6 |
CDRL3 | ALSMGSGIWVALSMGSGIWV | ||
| 6F6F | 77 |
CDRH1 | GFTFDDHGGFTFDDHG | |
| 88 | CDRH2CDRH2 | INLNGGSTINLNGGST | ||
|
9 | CDRH3CDRH3 | STRHLHHSTRHLHH | ||
| 1010 |
CDRL1 | QSLVHSNGNTYQSLVHSNGNTY | ||
| 1111 | CDRL2CDRL2 | KISKIS | ||
| 1212 |
CDRL3 | VQAKQFPLTVQAKQFPLT | ||
| 8F8F | 1313 | CDRH1CDRH1 | GFSFNDYAGFSFNDYA | |
| 1414 |
CDRH2 | ISWNSRSTISWNSRST | ||
| 1515 | CDRH3CDRH3 | TTDLGVVPAAIYAFDITTDLGVVPAAIYAFDI | ||
| 1616 | CDRL1CDRL1 | QGITNWQGITNW | ||
| 1717 | CDRL2CDRL2 | AASAAS | ||
| 1818 |
CDRL3 | QQTATFQITQQTATFQIT | ||
| 10F10F | 1919 |
CDRH1 | GFTVSSSFGFTVSSSF | |
| 2020 | CDRH2CDRH2 | AYSGGNTAYSGGNT | ||
| 2121 | CDRH3CDRH3 | SCEGKAVSGSRDLHFEFSCEGKAVSGSRDLHFEF | ||
| 2222 | CDRL1CDRL1 | NLRTKYNLRTKY | ||
| 2323 |
CDRL2 | NDNNDN | ||
| 2424 |
CDRL3 | QVWDSSSDHLIQVWDSSSDHLI | ||
| 3F3F | 2525 | VH 중쇄가변영역VH heavy chain variable region | QMQLVQSGAEVKKPGASVKVSCKASGYTFTDYYIHWVRQAPGQGLEWMGWINPNSGAPEFAQRFQGRVSMTRDASINTTYMELSGLRSEDTAVYYCVKGLEHAAGSAIFDRWGQGTMVTVSSQMQLVQSGAEVKKPGASVKVSCKASGYTFTDYYIHWVRQAPGQGLEWMGWINPNSGAPEFAQRFQGRVSMTRDASINTTYMELSGLRSEDTAVYYCVKGLEHAAGSAIFDRWGQGTMVTVSS | |
| 2626 |
VL 경쇄가변영역VL light chain | QTVVTQEPSFSVSPGGTVTLTCGLTSGSVSTSHFPSWYQQTPGQAPRTLIYFKDTRSSGVPDRFSGSILGNKAALTITGA | QADDESDYYCALSMGSGIWVFGGGTKLTVLQTVVTQEPSFSVSPGGTVTLTCGLTSGSVSTSHFPSWYQQTPGQAPRTLIYFKDTRSSGVPDRFSGSILGNKAALTITGAQADDESDYYCALSMGSGIWVFGGGTKLTVL | |
| 6F6F | 2727 | VH 중쇄가변영역VH heavy chain variable region | QVQLVESGGGVVRPGGSLRLSCTASGFTFDDHGMSWVRQAPGKGLEWVSTINLNGGSTAYADSVKGRFTISRDNAKNSLYLQMNSLRAEDTAVYYCSTRHLHHWGQGTLVTVSSQVQLVESGGGVVRPGGSLRLSCTASGFTFDDHGMSWVRQAPGKGLEWVSTINLNGGSTAYADSVKGRFTISRDNAKNSLYLQMNSLRAEDTAVYYCSTRHLHHWGQGTLVTVSS | |
| 2828 |
VL경쇄가변영역VL light chain | DIVMTQTPLSSPVTLGQPASISCRSSQSLVHSNGNTYLTWLQQRPGQPPRLLIHKISNRFSGVPDRFSGSGAGTDFTLKISR | VEAEDVGVYYCVQAKQFPLTFGQGTRLEIKDIVMTQTPLSSPVTLGQPASISCRSSQSLVHSNGNTYLTWLQQRPGQPPRLLIHKISNRFSGVPDRFSGSGAGTDFTLKISRVEAEDVGVYYCVQAKQFPLTFGQGTRLEIK | |
| 8F8F | 2929 | VH 중쇄가변영역VH heavy chain variable region | QVQLVQSGGGVVQPGGSLRLSCAASGFSFNDYAMHWVRQVPGKGLEWVSSISWNSRSTVYAASVEGRFSISRDNSKNSLYLQMNSLRAEDAAVYYCTTDLGVVPAAIYAFDIWGQGTMVTVSSQVQLVQSGGGVVQPGGSLRLSCAASGFSFNDYAMHWVRQVPGKGLEWVSSISWNSRSTVYAASVEGRFSISRDNSKNSLYLQMNSLRAEDAAVYYCTTDLGVVPAAIYAFDIWGQGTMVTVSS | |
| 3030 |
VL경쇄가변영역VL light chain | DIQMTQSPSVMSASVGDRVNITCRASQGITNWLAWYQQKPGKAPKLLISAASSLQSGVPSRFSGSGSGTDFTLTISS | LQPDDFATYYCQQTATFQITFGQGTRLDIKDIQMTQSPSVMSASVGDRVNITCRASQGITNWLAWYQQKPGKAPKLLISAASSLQSGVPSRFSGSGSGTDFTLTISSLQPDDFATYYCQQTATFQITFGQGTRLDIK | |
| 10F10F | 3131 | VH중쇄가변영역VH heavy chain variable region | QVQLVESGGGLIQPGGSLRLSCVASGFTVSSSFMSWVRLAPGKGLEWVALAYSGGNTYYADSVKGRFTVSRDDSSNTLYLQMNSLRAEDTAVYYCSCEGKAVSGSRDLHFEFWSPGTLVTVSSQVQLVESGGGLIQPGGSLRLSCVASGFTVSSSFMSWVRLAPGKGLEWVALAYSGGNTYYADSVKGRFTVSRDDSSNTLYLQMNSLRAEDTAVYYCSCEGKAVSGSRDLHFEFWSPGTLVTVSS | |
| 3232 | VL경쇄가변영역VL light chain variable region | SYELTQAPSLSVSPGQTANIICSGDNLRTKYVSWYQQKPGQAPVLVIYNDNDRPSGIPERFSGTNSGNTAALTISRVEAGDEADYYCQVWDSSSDHLIFGGGTKLTVLSYELTQAPSLSVSPGQTANIICSGDNLRTKYVSWYQQKPGQAPVLVIYNDNDRPSGIPERFSGTNSGNTAALTISRVEAGDEADYYCQVWDSSSDHLIFGGGTKLTVL |
[[
실시예Example
4] 4]
P. P.
acnesacnes
에on
대한 인간 For human
단일클론항체의Monoclonal antibody
친화력 확인 Affinity Check
본 발명에서 제조한 단일클론항체가 P.
acnes에 대하여 나타내는 친화력을 ELISA 분석으로 확인하였다.The affinity of the monoclonal antibody prepared in the present invention against P. acnes was confirmed by ELISA analysis.
그 결과, 하기 [표 6], 도 8 및 도 9에서 나타난 바와 같이, 총 4종의 항체(3F, 6F, 8F 및 10 F)는 모두 대조군 세포에 비해 P.
acnes에 결합 친화적인 것을 확인하였으며, 특히 6F 및 8F 항체가 P.
acnes에 대한 우수한 결합력을 나타내는 항체임을 확인하였다. ELISA 측정값을 이용하여 각 항체의 해리상수(KD)을 계산하였으며, 6F의 KD는 2.6?10-10, 8F의 KD는 1.0?10-9로서, KD =10-6 이하보다 월등히 낮은 해리상수 값을 가지므로 이는 P.
acnes에 대한 6F와 8F 항체가 높은 친화도를 가짐을 확인하였다(도 9).As a result, as shown in [Table 6], FIG. 8 and FIG. 9, all four antibodies (3F, 6F, 8F and 10F) were found to be binding-friendly to P. acnes compared to control cells. In particular, it was confirmed that the 6F and 8F antibody is an antibody showing excellent binding to P. acnes . The dissociation constant (K D ) of each antibody was calculated using ELISA measurements, and K D of 6F was 2.6-10 -10 , K D of 8F was 1.0-10 -9 , and K D = 10 -6 or less. Since the dissociation constant value is significantly lower, it was confirmed that the 6F and 8F antibodies to P. acnes has a high affinity (Fig. 9).
| 항체명Antibody Name | 여드름 유발균Acne-causing bacteria (( O.DO.D .450㎚).450 nm) | 대조군 세포Control cells (( O.DO.D .450㎚).450 nm) |
| 3F3F | 0.014950.01495 | -0.01050-0.01050 |
| 6F6F | 2.362152.36215 | 0.011400.01140 |
| 8F8F | 2.320652.32065 | -0.02680-0.02680 |
| 10F10F | 0.006050.00605 | -0.02475-0.02475 |
[[
실시예Example
5] 인간 5] human
단일클론항체의Monoclonal antibody
P. P.
acnesacnes
생장 억제 효과 확인 Confirmation of growth inhibitory effect
본 발명에서 제조한 인간 항체가 유의적으로 여드름 균에 대하여 사멸 효과를 나타낼 수 있는지 확인하였다.It was confirmed whether the human antibody prepared in the present invention can show a significant killing effect against acne bacteria.
먼저, P.
acnes를 액체 배지에 접종하여 하루밤 동안 배양한 다음, 제조한 6F 항체 또는 8F 항체를 30 ㎍/㎖의 농도로 첨가하거나, 6F 항체 및 8F 항체 동일 비율로 혼합하고 30 ㎍/㎖의 농도로 첨가하여 24 시간 동안 추가 배양하여 반응시켰다. 그런 다음, 고체 배지에 반응한 배지를 도말하고 다시 하룻밤 동안 배양하여 형성된 콜로니의 수를 계수하였다. 대조군으로 사용하기 위해, 동일 조건에서 항체 대신 PBS만을 첨가하여 24시간을 배양한 미처리 대조군을 준비하였다. 콜로니 수를 계수한 후, 항체 처리 전(0 시간)의 콜로니 수 대비 24 시간 처리군의 상대적인 콜로니 수를 계산하여 P. acnes 생장률을 계산하였다.First, inoculate P. acnes in a liquid medium and incubate overnight, and then add the prepared 6F antibody or 8F antibody at a concentration of 30 μg / ml, or mix 6F antibody and 8F antibody at the same ratio and The reaction was added at a concentration and further incubated for 24 hours. Then, the medium reacted with the solid medium was plated and incubated again overnight to count the number of colonies formed. For use as a control, an untreated control was prepared in which 24 hours of incubation was performed using only PBS instead of antibodies under the same conditions. After counting the colonies, P. acnes growth rate was calculated by counting the relative colonies in the 24 hour treatment group relative to the number of colonies before antibody treatment (0 hour).
그 결과, 하기 [표 7] 및 도 9에서 나타난 바와 같이, 미처리 대조군에서는 P.
acnes의 생장률이 약 150%를 나타내는 반면, 6F 항체 또는 8F 항체를 단독으로 첨가한 실험군에서 각각 22.2%, 30.8% 만을 나타내어, 미처리 대조군에 비해 유의적인 생장률 감소 효과를 나타낼 수 있음을 확인하였다. 또한, 6F 항체 및 8F 항체를 동시에 처리한 실험군에서는 0 시간 대비 P.
acnes의 생장률이 83.5%를 나타내므로, 16.5% 감소된 수준의 생존율을 나타내는 것으로 확인하여, 6F 및 8F 항체를 동시에 처리하는 것을 통해 P.
acnes의 생장 억제의 시너지 효과가 나타나 유의적으로 여드름균을 사멸하는 효과를 나타낼 수 있는 것으로 확인하였다.As a result, as shown in [Table 7] and FIG. 9, the growth rate of P. acnes was about 150% in the untreated control group, while 22.2% and 30.8% in the experimental group added with 6F antibody or 8F antibody alone, respectively. Only it was confirmed that it can show a significant growth rate reduction effect compared to the untreated control. In addition, in the experimental group treated with 6F antibody and 8F antibody at the same time, the growth rate of P. acnes was 83.5% compared to 0 hours, and it was confirmed that it showed a 16.5% reduced level of survival, and simultaneously treated 6F and 8F antibodies. Through the synergistic effect of growth inhibition of P. acnes appeared to be able to significantly kill the acne bacteria.
| 구분division |
반응시간 대비 생장률Growth rate compared to reaction | 증감율Change rate | ||
|
0 시간0 |
24 시간24 hours | |||
|
미처리 대조군 |
100 %100% | 148.5 %148.5% | + 48.5 %+ 48.5% | |
| 6F6F | 100 %100% | 122.2 %122.2% | + 22.2 %+ 22.2% | |
| 8F8F | 100 %100% | 130.8 %130.8% | + 30.8 %+ 30.8% | |
|
6F+8F6F + |
100 %100% | 83.5 %83.5% | - 16.5 %-16.5% | |
전술한 본 발명의 설명은 예시를 위한 것이며, 본 발명이 속하는 기술분야의 통상의 지식을 가진 자는 본 발명의 기술적 사상이나 필수적인 특징을 변경하지 않고서 다른 구체적인 형태로 쉽게 변형이 가능하다는 것을 이해할 수 있을 것이다. 그러므로 이상에서 기술한 실시예들은 모든 면에서 예시적인 것이며 한정적이 아닌 것으로 이해해야만 한다. The foregoing description of the present invention is intended for illustration, and it will be understood by those skilled in the art that the present invention may be easily modified in other specific forms without changing the technical spirit or essential features of the present invention. will be. Therefore, it should be understood that the embodiments described above are exemplary in all respects and not restrictive.
하기에 본 발명의 조성물을 위한 제조예를 예시한다.The preparation examples for the compositions of the present invention are illustrated below.
[[
제조예Production Example
1] 화장품의 제조 1] Manufacture of Cosmetics
<1-1> 유연화장수(스킨)<1-1> Softener (skin)
본 발명의 단일클론항체를 포함하는 유연화장수를 제조하기 위해 하기 [표 7]에 기재된 것처럼 배합하여 통상적인 화장품 분야에서의 제조 방법에 따라 제조할 수 있다.In order to manufacture the flexible cosmetics containing the monoclonal antibody of the present invention can be prepared according to the preparation method in the conventional cosmetic field by combining as described in the following [Table 7].
| 성분ingredient | 함량(중량%)Content (% by weight) |
| 본 발명의 단일클론항체Monoclonal Antibodies of the Invention | 0.1 ~ 30 %0.1-30% |
| 1,3-부틸렌글리콜1,3-butylene glycol | 3.03.0 |
| 글리세린glycerin | 5.05.0 |
| 폴리옥시에틸렌(60) 경화피마자유Polyoxyethylene (60) hardening castor oil | 0.20.2 |
| 에탄올ethanol | 8.08.0 |
| 구연산Citric acid | 0.020.02 |
| 구연산나트륨Sodium citrate | 0.060.06 |
| 방부제antiseptic | 미량a very small amount |
| 향료Spices | 미량a very small amount |
| 정제수Purified water | To 100To 100 |
<1-2> 영양화장수(로션)<1-2> nutrient lotion (lotion)
본 발명의 단일클론항체를 포함하는 영양화장수를 제조하기 위해 하기 [표 9]에 기재된 것처럼 배합하여 통상적인 화장품 분야에서의 제조 방법에 따라 제조할 수 있다.In order to produce nutritive cosmetics containing the monoclonal antibody of the present invention, it can be formulated according to the preparation method in the general cosmetic field by combining as described in Table 9 below.
| 성분ingredient | 함량(중량%)Content (% by weight) |
| 본 발명의 단일클론항체Monoclonal Antibodies of the Invention | 0.1 ~ 30 %0.1-30% |
| 1,3-부틸렌글리콜1,3-butylene glycol | 8.08.0 |
| 글리세린glycerin | 5.05.0 |
| 스쿠알란Squalane | 10.010.0 |
| 모노올레인산폴리옥시에틸렌소르비탄Monooleic acid polyoxyethylene sorbitan | 2.02.0 |
| 유창목오일Flux oil | 0.1 ~ 30 %0.1-30% |
| 1,3-부틸렌글리콜1,3-butylene glycol | 3.03.0 |
| 글리세린glycerin | 5.05.0 |
| 폴리옥시에틸렌(60) 경화피마자유Polyoxyethylene (60) hardening castor oil | 0.20.2 |
| 에탄올ethanol | 8.08.0 |
| 구연산Citric acid | 0.020.02 |
| 구연산나트륨Sodium citrate | 0.060.06 |
| 방부제antiseptic | 미량a very small amount |
| 향료Spices | 미량a very small amount |
| 정제수Purified water | To 100To 100 |
<1-3> 에센스<1-3> essence
본 발명의 단일클론항체를 포함하는 에센스를 제조하기 위해 하기 [표 10]에 기재된 것처럼 배합하여 통상적인 화장품 분야에서의 제조 방법에 따라 제조할 수 있다.To prepare an essence comprising the monoclonal antibody of the present invention, it can be combined according to the preparation method in the conventional cosmetic field as described in Table 10 below.
| 성분ingredient | 함량(중량%)Content (% by weight) |
| 본 발명의 단일클론항체Monoclonal Antibodies of the Invention | 0.1 ~ 30 %0.1-30% |
| 시토스테롤Sitosterol | 1.71.7 |
| 플리글리세릴2-올레이트Polyglyceryl 2-oleate | 1.51.5 |
| 세라마이드Ceramide | 0.70.7 |
| 세테아레스-4Ceteares-4 | 1.21.2 |
| 콜레스테롤cholesterol | 1.51.5 |
| 디세틸포스페이트Dicetylphosphate | 0.40.4 |
| 농글리세린Concentrated glycerin | 5.05.0 |
| 카르복시비닐폴리머Carboxy Vinyl Polymer | 0.20.2 |
| 산탄검Xanthan Gum | 0.20.2 |
| 방부제antiseptic | 미량a very small amount |
| 향료Spices | 미량a very small amount |
| 정제수Purified water | To 100To 100 |
<1-4> 세안제(클렌징폼)<1-4> Face wash (cleansing foam)
본 발명의 단일클론항체를 포함하는 세안제(클렌징폼)를 제조하기 위해 하기 [표 11]에 기재된 것처럼 배합하여 통상적인 화장품 분야에서의 제조방법에 따라 제조할 수 있다.To prepare a cleansing agent (cleansing foam) comprising the monoclonal antibody of the present invention, it may be combined as described in the following [Table 11], and may be prepared according to a conventional manufacturing method in the cosmetic field.
| 성분ingredient | 함량(중량%)Content (% by weight) |
| 본 발명의 인간항체Human Antibodies of the Invention | 0.1 ~ 30 %0.1-30% |
| N-아실글루타민산나트륨N-acyl glutamate | 20.020.0 |
| 글리세린glycerin | 10.010.0 |
| PEG-400PEG-400 | 15.015.0 |
| 프로필렌글리콜Propylene glycol | 10.010.0 |
| POE(15) 올레일알코올에테르POE (15) oleyl alcohol ether | 3.03.0 |
| 라우린유도체Laurin derivative | 2.02.0 |
| 메틸파라벤Methylparaben | 0.20.2 |
| EDTA-4NaEDTA-4Na | 0.030.03 |
| 향료Spices | 0.20.2 |
| 정제수Purified water | To 100To 100 |
<1-5> 영양크림<1-5> Nutrition Cream
본 발명의 단일클론항체를 포함하는 영양크림을 제조하기 위해 하기 [표 12]에 기재된 것처럼 통상적인 화장품 분야에서의 제조 방법에 따라 제조한다.To prepare a nourishing cream comprising the monoclonal antibody of the present invention, it is prepared according to the preparation method in the conventional cosmetic field as described in Table 12 below.
| 배합성분Ingredient | 함량(중량%)Content (% by weight) |
| 본 발명의 펩타이드Peptides of the Invention | 0.1 ~ 30 %0.1-30% |
| 바셀린vaseline | 7.07.0 |
| 유동파라핀Liquid paraffin | 10.010.0 |
| 밀납Beeswax | 2.02.0 |
| 폴리솔베이트60Polysorbate 60 | 2.52.5 |
| 솔비탄세스퀴올레이트Sorbitan sesquioleate | 1.51.5 |
| 스쿠알란Squalane | 3.03.0 |
| 프로필렌글리콜Propylene glycol | 6.06.0 |
| 글리세린glycerin | 4.04.0 |
| 트리에탄올아민Triethanolamine | 0.50.5 |
| 산탄검Xanthan Gum | 0.50.5 |
| 토코페닐아세테이트Tocophenyl Acetate | 0.10.1 |
| 향, 방부제Incense, preservative | 미량a very small amount |
| 정제수Purified water | To 100To 100 |
<1-6> 마사지크림<1-6> massage cream
본 발명의 단일클론항체를 포함하는 마사지크림을 제조하기 위해 하기 [표 13]에 기재된 것처럼 통상적인 화장품 분야에서의 제조방법에 따라 제조한다.To prepare a massage cream comprising a monoclonal antibody of the present invention it is prepared according to the manufacturing method in the general cosmetic field as described in Table 13 below.
| 배합성분Ingredient | 함량(중량%)Content (% by weight) |
| 본 발명의 단일클론항체Monoclonal Antibodies of the Invention | 0.1 ~ 30 %0.1-30% |
| 프로필렌글리콜Propylene glycol | 6.06.0 |
| 글리세린glycerin | 4.04.0 |
| 트리에탄올아민Triethanolamine | 0.50.5 |
| 밀납Beeswax | 2.02.0 |
| 토코페닐아세테이트Tocophenyl Acetate | 0.10.1 |
| 폴리솔베이트60Polysorbate 60 | 3.03.0 |
| 솔비탄세스퀴올레이트Sorbitan sesquioleate | 2.52.5 |
| 세테아릴알코올Cetearyl Alcohol | 2.02.0 |
| 유동파라핀Liquid paraffin | 30.030.0 |
| 산탄검Xanthan Gum | 0.50.5 |
| 향, 방부제Incense, preservative | 미량a very small amount |
| 정제수Purified water | To 100To 100 |
<1-7> 팩<1-7> pack
본 발명의 단일클론항체를 포함하는 팩을 제조하기 위해 하기 [표 14]에 기재된 것처럼 통상적인 화장품 분야에서의 제조 방법에 따라 제조한다.To prepare a pack comprising the monoclonal antibody of the present invention, it is prepared according to the preparation method in the conventional cosmetic field as described in Table 14 below.
| 배합성분Ingredient | 함량(중량%)Content (% by weight) |
| 본 발명의 단일클론항체Monoclonal Antibodies of the Invention | 0.1 ~ 30 %0.1-30% |
| 프로필렌글리콜Propylene glycol | 2.02.0 |
| 글리세린glycerin | 4.04.0 |
| 폴리비닐알코올Polyvinyl alcohol | 10.010.0 |
| 에탄올ethanol | 7.07.0 |
| 파이지-40 히드로게비이티드캐스터오일Fiji-40 Hydrogenated Castor Oil | 0.80.8 |
| 트리에탄올아민Triethanolamine | 0.30.3 |
| 향, 방부제Incense, preservative | 미량a very small amount |
| 정제수Purified water | To 100To 100 |
이상의 본 발명은 상기에 기술된 실시예 및 제조예들에 의해 한정되지 않고, 통상의 기술자들에 의해 다양한 변형 및 변경을 가져올 수 있으며, 그외의 색채 화장품을 포함하는 다양한 용도의 화장품에 적용될 수 있는 것이고, 그 효능에 따라 인체에 얇게 도포하여 바를 수 있는 약제 즉, 연고로 제조에 이용될 수 있고, 이는 첨부된 청구항에서 정의되는 본 발명의 취지와 범위에 포함된다.The present invention is not limited to the above-described embodiments and preparation examples, and can be variously modified and changed by those skilled in the art, and can be applied to cosmetics of various uses including other color cosmetics. It can be used in the manufacture of a medicament, that is, an ointment that can be applied thinly to the human body depending on its efficacy, which is included in the spirit and scope of the present invention as defined in the appended claims.
Claims (11)
- 프로피오니 박테리움 아크네스(Propioni - bacterium acnes : P. acnes)에 특이적으로 결합하는, 단일클론항체.A monoclonal antibody that specifically binds to Propioni bacterium acnes (P. acnes ).
- 제 1항에 있어서,The method of claim 1,상기 단일클론항체는 하기 (a) 내지 (d)로 이루어진 군으로부터 선택되는 어느 하나인 것을 특징으로 하는, 단일클론항체:The monoclonal antibody is characterized in that any one selected from the group consisting of (a) to (d), monoclonal antibody:(a) 서열번호 1로 기재된 중쇄 CDR1, 서열번호 2로 기재된 중쇄 CDR2, 및 서열번호 3으로 기재된 중쇄 CDR3를 포함하는 중쇄 가변영역과 서열번호 4로 기재된 경쇄 CDR1, 서열번호 5로 기재된 경쇄 CDR2 및 서열번호 6으로 기재된 경쇄 CDR3를 포함하는 경쇄 가변영역을 포함하는 항체;(a) a heavy chain variable region comprising a heavy chain CDR1 as set out in SEQ ID NO: 1, a heavy chain CDR2 as set out in SEQ ID NO: 2, and a heavy chain CDR3 as set out in SEQ ID NO: 3, a light chain CDR1 as set out in SEQ ID NO: 4, a light chain CDR2 as set out in SEQ ID NO: 5, and An antibody comprising a light chain variable region comprising a light chain CDR3 set forth in SEQ ID NO: 6;(b) 서열번호 7로 기재된 중쇄 CDR1, 서열번호 8로 기재된 중쇄 CDR2, 및 서열번호 9로 기재된 중쇄 CDR3를 포함하는 중쇄 가변영역과 서열번호 10으로 기재된 경쇄 CDR1, 서열번호 11로 기재된 경쇄 CDR2 및 서열번호 12로 기재된 경쇄 CDR3를 포함하는 경쇄 가변영역을 포함하는 항체;(b) a heavy chain variable region comprising a heavy chain CDR1 as set out in SEQ ID NO: 7, a heavy chain CDR2 as set out in SEQ ID NO: 8, and a heavy chain CDR3 as set out in SEQ ID NO: 9, a light chain CDR1 as set out in SEQ ID NO: 10, a light chain CDR2 as set out in SEQ ID NO: 11, and An antibody comprising a light chain variable region comprising a light chain CDR3 set forth in SEQ ID NO: 12;(c) 서열번호 13으로 기재된 중쇄 CDR1, 서열번호 14로 기재된 중쇄 CDR2, 및 서열번호 15로 기재된 중쇄 CDR3를 포함하는 중쇄 가변영역과 서열번호 16으로 기재된 경쇄 CDR1, 서열번호 17로 기재된 경쇄 CDR2 및 서열번호 18로 기재된 경쇄 CDR3를 포함하는 경쇄 가변영역을 포함하는 항체; 및(c) a heavy chain variable region comprising a heavy chain CDR1 as set out in SEQ ID NO: 13, a heavy chain CDR2 as set out in SEQ ID NO: 14, and a heavy chain CDR3 as set out in SEQ ID NO: 15, a light chain CDR1 as set out in SEQ ID NO: 16, a light chain CDR2 as set out in SEQ ID NO: 17, and An antibody comprising a light chain variable region comprising a light chain CDR3 set forth in SEQ ID NO: 18; And(d) 서열번호 19로 기재된 중쇄 CDR1, 서열번호 20으로 기재된 중쇄 CDR2, 및 서열번호 21로 기재된 중쇄 CDR3를 포함하는 중쇄 가변영역과 서열번호 22로 기재된 경쇄 CDR1, 서열번호 23으로 기재된 경쇄 CDR2 및 서열번호 24로 기재된 경쇄 CDR3를 포함하는 경쇄 가변영역을 포함하는 항체.(d) a heavy chain variable region comprising a heavy chain CDR1 as set out in SEQ ID NO: 19, a heavy chain CDR2 as set out in SEQ ID NO: 20, and a heavy chain CDR3 as set out in SEQ ID NO: 21, a light chain CDR1 as set out in SEQ ID NO: 22, a light chain CDR2 as set out in SEQ ID NO: 23, and An antibody comprising a light chain variable region comprising the light chain CDR3 set forth in SEQ ID NO: 24.
- 제 1항에 있어서, 상기 단일클론항체는 하기 (a) 내지 (d)로 이루어진 군으로부터 선택되는 어느 하나인 것을 특징으로 하는, 단일클론항체:According to claim 1, wherein the monoclonal antibody is any one selected from the group consisting of (a) to (d), monoclonal antibody:(a) 서열번호 25로 기재된 중쇄 가변영역 및 서열번호 26으로 기재된 경쇄 가변영역을 포함하는 항체;(a) an antibody comprising a heavy chain variable region as set out in SEQ ID NO: 25 and a light chain variable region as set out in SEQ ID NO: 26;(b) 서열번호 27로 기재된 중쇄 가변영역 및 서열번호 28로 기재된 경쇄 가변영역을 포함하는 항체;(b) an antibody comprising a heavy chain variable region as set out in SEQ ID NO: 27 and a light chain variable region as set out in SEQ ID NO: 28;(c) 서열번호 29로 기재된 중쇄 가변영역 및 서열번호 30로 기재된 경쇄 가변영역을 포함하는 항체;(c) an antibody comprising a heavy chain variable region as set out in SEQ ID NO: 29 and a light chain variable region as set out in SEQ ID NO: 30;(d) 서열번호 31로 기재된 중쇄 가변영역 및 서열번호 32로 기재된 경쇄 가변영역을 포함하는 항체.(d) an antibody comprising a heavy chain variable region as set out in SEQ ID NO: 31 and a light chain variable region as set out in SEQ ID NO: 32.
- 제 1항 내지 제 4항 중 어느 한 항에 있어서, 상기 단일클론항체는 인간 항체인 것을 특징으로 하는, 단일클론항체.The monoclonal antibody according to any one of claims 1 to 4, wherein the monoclonal antibody is a human antibody.
- 제 1항 내지 제 4항 중 어느 한 항의 단일클론항체를 암호화하는 폴리뉴클레오티드를 포함하는, 발현 벡터.An expression vector comprising a polynucleotide encoding the monoclonal antibody of any one of claims 1 to 4.
- 제 5항의 발현 벡터로 형질전환된, 형질전환체.A transformant transformed with the expression vector of claim 5.
- 제 1항 내지 제 4항 중 어느 한 항의 단일클론항체를 유효성분으로 포함하는, 여드름에 대한 예방 또는 개선용 화장료 조성물.Claim 1 to 4, wherein the monoclonal antibody of any one comprising as an active ingredient, preventing or improving cosmetic composition for acne.
- 제 1항 내지 제 4항 중 어느 한 항의 단일클론항체를 유효성분으로 포함하는, 여드름에 대한 예방 또는 치료용 약학적 조성물.A pharmaceutical composition for preventing or treating acne, comprising the monoclonal antibody of any one of claims 1 to 4 as an active ingredient.
- 제 1항 내지 제 4항 중 어느 한 항의 단일클론항체를 여드름에 대한 예방 또는 개선용 화장료 조성물에 사용하기 위한 용도.Use of the monoclonal antibody of any one of claims 1 to 4 in a cosmetic composition for preventing or improving acne.
- 제 1항 내지 제 4항 중 어느 한 항의 단일클론항체를 포함하는 여드름에 대한 예방 또는 치료용 약학적 조성물의 용도.Use of a pharmaceutical composition for preventing or treating acne comprising the monoclonal antibody of any one of claims 1 to 4.
- 제 1항 내지 제 4항 중 어느 한 항의 단일클론항체를 약학적으로 유효한량으로 필요한 개체에 투여하는 단계를 포함하는, 여드름 치료 방법.A method of treating acne comprising administering to a subject in need thereof a pharmaceutically effective amount of the monoclonal antibody of claim 1.
Priority Applications (5)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| US16/309,582 US11141369B2 (en) | 2016-06-13 | 2017-06-13 | Human antibody specifically binding to acne bacteria using phage display technique, and use thereof |
| JP2018566417A JP6916216B2 (en) | 2016-06-13 | 2017-06-13 | Human antibodies that specifically bind to acne using phage display technology and their uses |
| EP17813568.7A EP3476862A4 (en) | 2016-06-13 | 2017-06-13 | Human antibody specifically binding to acne bacteria using phage display technique, and use thereof |
| CN201780050100.7A CN109963867B (en) | 2016-06-13 | 2017-06-13 | Human antibodies specifically binding to acne bacteria by phage display technology and uses thereof |
| BR112018076013A BR112018076013A2 (en) | 2016-06-13 | 2017-06-13 | monoclonal antibody and its use, expression vector, transformant, cosmetic composition, pharmaceutical composition and method for treating acne |
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| KR10-2016-0073186 | 2016-06-13 | ||
| KR10-2017-0074010 | 2017-06-13 | ||
| KR1020170074010A KR101992292B1 (en) | 2016-06-13 | 2017-06-13 | Human antibodies specifically binding to acne using phage display technologies and uses thereof |
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| KR102084163B1 (en) * | 2019-12-27 | 2020-05-15 | 주식회사 인투앱 | Novel antibodies specifically binding to lipase from the propionibacterium acnes and uses thereof |
Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| KR950002765A (en) * | 1993-07-13 | 1995-02-16 | 안기영 | Acne treatment and manufacturing method |
| KR20100003753A (en) * | 2008-07-02 | 2010-01-12 | 김종배 | Mehod of production antibody against propionibacterium acnes and cosmetic composition for treatment of disease using the same |
| US20130123132A1 (en) * | 2011-11-15 | 2013-05-16 | Ingen Biosciences | Method for Diagnosing Propionibacterium Bacterial Infections |
| US20140234337A1 (en) * | 2011-08-19 | 2014-08-21 | Immortal Spirit Limited | Antibody and antibody-containing composition |
-
2017
- 2017-06-13 WO PCT/KR2017/006156 patent/WO2017217744A1/en unknown
Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| KR950002765A (en) * | 1993-07-13 | 1995-02-16 | 안기영 | Acne treatment and manufacturing method |
| KR20100003753A (en) * | 2008-07-02 | 2010-01-12 | 김종배 | Mehod of production antibody against propionibacterium acnes and cosmetic composition for treatment of disease using the same |
| US20140234337A1 (en) * | 2011-08-19 | 2014-08-21 | Immortal Spirit Limited | Antibody and antibody-containing composition |
| US20130123132A1 (en) * | 2011-11-15 | 2013-05-16 | Ingen Biosciences | Method for Diagnosing Propionibacterium Bacterial Infections |
Non-Patent Citations (1)
| Title |
|---|
| JUNG ET AL.: "Generation of Human Monoclonal Antibodies against Propionilbacterium Acnes by Applying the Phage Display Method to Human Peripheral Blood Mononuclear Cells Immunized in Vitro", CYTOTECHNOLOGY, vol. 57, 2008, pages 169 - 175, XP019602508 * |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| KR102084163B1 (en) * | 2019-12-27 | 2020-05-15 | 주식회사 인투앱 | Novel antibodies specifically binding to lipase from the propionibacterium acnes and uses thereof |
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