Antisense oligonucleotide targeting CRTH2 mRNA and application thereof

Abstract

The invention provides antisense oligonucleotide targeting CRTH2 mRNA and application of the antisense oligonucleotide, and relates to the technical field of biology. The nucleotide sequence of the antisense oligonucleotide targeting CRTH2 mRNA provided by the invention comprises a sequence of which the difference from continuous nucleotides from the (1-3) th site to the (18-20) th site of any sequence shown in SEQ ID NO.1-30 is not more than 7 nucleotides. The antisense oligonucleotide can inhibit CRTH2 mRNA and reduce CRTH2 protein expression of a subject, and is beneficial to treatment or prevention of related diseases caused by CRTH2 of the subject or preparation of drugs for treating the related diseases.

Description

Technical Field

[0001] This invention relates to the field of biotechnology, and in particular to an antisense oligonucleotide targeting CRTH2 mRNA and its applications. Background Technology

[0002] The following statements are provided only as background information in relation to the present invention and do not necessarily constitute prior art.

[0003] The Th2 cell chemotactic receptor homolog (CRTH2) has been shown to mediate chemotaxis in eosinophils, basophils, and Th2 T lymphocytes. The major mast cell product prostaglandin (PG)D2 is considered the major ligand for CRTH2. CRTH2 is the most reliable marker for detecting circulating human type 2 Th and type 2 T cytotoxic cells in both healthy and diseased states. The CRTh2 receptor is expressed on eosinophils, basophils, T helper cells 2, macrophages, and neutrophils, participating in the chemotaxis and activation of these cells, thus forming key events that trigger inflammatory responses in allergic diseases. Studies have shown that CRTH2, as a receptor for prostaglandin D2 (PGD2), plays a crucial role in various allergic and inflammatory diseases. In autoimmune diseases, CRTH2 may exacerbate inflammatory responses by promoting the activation and recruitment of Th2 cells; in atopic dermatitis, CRTH2 activation exacerbates skin inflammation, itching, and epidermal barrier dysfunction, driving disease progression; CRTH2 can promote eosinophil chemotaxis and survival, leading to esophageal and gastrointestinal inflammation; CRTH2 is highly expressed in airway inflammation in asthma and is associated with asthma; in addition, CRTH2 is also associated with allergic rhinitis, conjunctivitis, and the eosinophilic form of chronic obstructive pulmonary disease (COPD).

[0004] Antisense oligonucleotides (ASOs) are short, single-stranded nucleotide sequences complementary to target messenger RNA (mRNA). They bind to specific mRNAs via base pairing principles, thereby regulating gene expression at the post-transcriptional level. Using antisense oligonucleotides that target and bind to CRTH2 mRNA can block or reduce diseases caused by CRTH2 abnormalities. Therefore, developing an antisense oligonucleotide that can effectively target CRTH2 mRNA and thus inhibit CRTH2 activity in subjects is currently in demand in the market.

[0005] In view of this, the present invention is hereby proposed. Summary of the Invention

[0006] The purpose of this invention is to provide an antisense oligonucleotide that targets CRTH2 mRNA in order to inhibit or block CRTH2 in subjects.

[0007] To solve the above-mentioned technical problems, the present invention adopts the following technical solution: In a first aspect, an antisense oligonucleotide targeting CRTH2 mRNA is provided, wherein the nucleotide sequence of the antisense oligonucleotide comprises a sequence that differs from the sequence shown in SEQ ID NO. 1~30 by no more than 7 consecutive nucleotides from position (1~3) to position (18~20).

[0008] In a second aspect, a complex of antisense oligonucleotides targeting CRTH2 mRNA is provided, the complex comprising: (a) the antisense oligonucleotides targeting CRTH2 mRNA as described in the first aspect; and (b) one or more targeting delivery ligands linked to (a).

[0009] Thirdly, the antisense oligonucleotide targeting CRTH2 mRNA as described in the first aspect, or the complex as described in the second aspect, is provided for use in any of the following: (I) Inhibition of CRTH2 mRNA and / or reduction of CRTH2 protein expression in subjects for non-diagnostic and non-therapeutic purposes; (II) Prepare drugs that inhibit CRTH2 mRNA and / or reduce CRTH2 protein expression in subjects; (III) Prepare medicines for the treatment and / or prevention of pathological conditions or diseases caused by CRTH2 in the subject.

[0010] Fourthly, a pharmaceutical composition is provided, the pharmaceutical composition comprising the antisense oligonucleotide targeting CRTH2 mRNA as described in the first aspect, or the complex as described in the second aspect.

[0011] Fifthly, a method for inhibiting CRTH2 in a subject for non-diagnostic and therapeutic purposes is provided, the method comprising contacting the subject with an antisense oligonucleotide targeting CRTH2 mRNA as described in the first aspect, or a complex as described in the second aspect, or a pharmaceutical composition as described in the fourth aspect.

[0012] Compared with the prior art, the present invention has the following beneficial effects: The antisense oligonucleotides targeting CRTH2 mRNA provided by this invention can inhibit CRTH2 mRNA, reduce CRTH2 protein expression in subjects, and help treat or prevent diseases caused by CRTH2 in subjects, or prepare drugs for treating related diseases. Detailed Implementation

[0013] The technical solution of the present invention will be clearly and completely described below with reference to the embodiments. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0014] In this document, the terms “comprising” or “including” are open-ended expressions used in this disclosure to mean the phrase “including but not limited to”, and are used interchangeably with it, meaning that they include the contents specified in this disclosure, but do not exclude other contents.

[0015] In this document, the terms “optionally,” “optionally,” or “optionally” generally refer to an event or condition that may or may not occur as described below, and the description includes both cases in which the event or condition occurs and cases in which the event or condition does not occur.

[0016] In this document, “and / or” is used to indicate that one or both of the situations described may occur, for example, A and / or B includes (A and B) and (A or B).

[0017] In this document, unless otherwise stated, any numbering is used to distinguish one entity or behavior from another, and is not required to require or imply any actual relationship, order, or importance between these entities or behaviors, such as numbering i, ii; first, second, etc.

[0018] In this document, the terms “comprising” or “including” mean that the stated elements, integers or steps are included, but do not exclude any other elements, integers or steps.

[0019] In this article, antisense oligonucleotides (ASOs) refer to short nucleic acid fragments that can bind complementary to target nucleic acids. Antisense oligonucleotides include ribonucleotides and / or deoxyribonucleotides, including but not limited to DNA or RNA, DNA-RNA hybrids, or polymers containing purine and pyrimidine bases or other naturally occurring, chemically or biochemically modified, non-natural or derived nucleotide bases. Unless otherwise specified, nucleotides refer to ribonucleotides and / or deoxyribonucleotides.

[0020] In this article, when "connection" refers to the connection of two molecules, the two molecules can be directly connected (e.g., directly connected by chemical bonds) or connected by additional molecules. When two molecules are directly connected, they can be covalently or non-covalently connected. Covalent connection refers to the connection of two molecules through the formation of covalent bonds, which are chemical bonds formed between atoms by sharing electron pairs. Non-covalent connection refers to connection methods other than covalent connection, such as connection through hydrogen bonds, van der Waals forces, electrostatic interactions, hydrophobic interactions, and metal coordination bonds.

[0021] In this document, the terms "subject" or "patient" refer to a mammalian subject or patient, and organs, tissues, or cells derived from them. The mammals include, but are not limited to, humans, rhesus monkeys, rats, mice, guinea pigs, rabbits, dogs, cats, hamsters, nude mice, ferrets, pigs, sheep, goats, cattle, horses, donkeys, dogs, cynomolgus monkeys, macaques, baboons, gibbons, golden monkeys, long-tailed macaques, marmosets, squirrel monkeys, pig-tailed macaques, or chimpanzees. In some embodiments, the subject is a human or a cynomolgus monkey, rhesus monkey, rat, mouse, or guinea pig.

[0022] In this document, the terms “treatment,” “relief,” or “improvement” are used interchangeably. These terms refer to methods of achieving beneficial or desired outcomes, including, but not limited to, treatment benefits. A “treatment benefit” means the eradication or improvement of one or more diseases, conditions, or circumstances associated with the underlying barrier being treated.

[0023] In this article, the terms “prevention” and “avoidance” are used interchangeably to refer to methods for obtaining beneficial or desired outcomes, including but not limited to preventive benefits. To obtain a “preventive benefit,” a drug may be given to a subject at risk of developing a specific disease, or to a subject who reports one or more physiological symptoms of a disease, even if a diagnosis of the disease may not have been made.

[0024] In this document, the term “inhibition” may be used interchangeably with “reduction,” “silencing,” “downregulation,” “blocking,” and other similar terms, and includes any level of inhibition. Inhibition can be assessed by a reduction in the absolute or relative level of one or more of these variables compared to a control level. This control level can be any type of control level used in the art, such as a baseline level before administration or a level determined from an untreated or controlled (e.g., a buffer-only control or an inert agent control) subject, cell, or sample.

[0025] In a first aspect, an antisense oligonucleotide targeting CRTH2 mRNA is provided. The nucleotide sequence of the antisense oligonucleotide comprises a sequence whose consecutive nucleotides from position (1-3) to position (18-20) of the sequence shown in SEQ ID NO. 1-30 differ from the sequence shown in SEQ ID NO. 1-30 by no more than 1, 2, 3, 4, 5, 6 or 7 nucleotides. For example, it can be, but is not limited to, a sequence containing no more than 7 nucleotide differences in consecutive nucleotides from position 1 to position 18 of any of the sequences shown in SEQ ID NO. 1 to 30, a sequence containing no more than 7 nucleotide differences in consecutive nucleotides from position 1 to position 19, a sequence containing no more than 7 nucleotide differences in consecutive nucleotides from position 1 to position 20, a sequence containing no more than 7 nucleotide differences in consecutive nucleotides from position 2 to position 18, a sequence containing no more than 7 nucleotide differences in consecutive nucleotides from position 2 to position 19, a sequence containing no more than 7 nucleotide differences in consecutive nucleotides from position 2 to position 20, a sequence containing no more than 7 nucleotide differences in consecutive nucleotides from position 3 to position 18, a sequence containing no more than 7 nucleotide differences in consecutive nucleotides from position 3 to position 19, or a sequence containing no more than 7 nucleotide differences in consecutive nucleotides from position 3 to position 20.

[0026] In an optional embodiment, the differential nucleotides in the antisense oligonucleotides from the (1st to 3rd) to (18th to 20th) consecutive nucleotides of the sequence shown in SEQ ID NO. 1 to 30 are derived from mutations at any one or more sites in the sequence shown in SEQ ID NO. 1 to 30.

[0027] In an optional embodiment, the differential nucleotides in the antisense oligonucleotides that are adjacent to the (1st to 3rd) to (18th to 20th) consecutive nucleotides of the sequence shown in SEQ ID NO. 1 to 30 are from the 1st, 2nd, 3rd, 4th, 5th, 6th or 7th nucleotides adjacent to the 5' end and / or 3' end of the fragment where the CRTH2 mRNA is located, according to any of the sequences shown in SEQ ID NO. 1 to 30.

[0028] In an optional embodiment, positions 1 to 5 of any of the sequences shown in SEQ ID NO. 1 to 30 are all ribonucleotides, positions 6 to 15 are all deoxyribonucleotides, and positions 16 to 20 are all ribonucleotides.

[0029] In an optional embodiment, the nucleotide sequence of the antisense oligonucleotide comprises a sequence that differs from the sequence shown in SEQ ID NO. 8 by no more than 7 consecutive nucleotides from positions (1-3) to (18-20).

[0030] In an optional embodiment, the nucleotide sequence of the antisense oligonucleotide comprises a sequence in which the first to 18th, the first to 19th, the first to 20th, the second to 18th, the second to 19th, the second to 20th, the third to 18th, the third to 19th, or the third to 20th positions of the sequence shown in SEQ ID NO.8 differ from the sequence in which the first to 20th positions by no more than 1, 2, 3, 4, 5, 6, or 7 nucleotides.

[0031] In an optional embodiment, the nucleotide sequence of the antisense oligonucleotide comprises the first to 20th consecutive nucleotides of the sequence shown in SEQ ID NO.8; or the third to 18th consecutive nucleotides.

[0032] In an optional embodiment, positions 1 to 5 of the sequence shown in SEQ ID NO.8 are all ribonucleotides, positions 6 to 15 are all deoxyribonucleotides, and positions 16 to 20 are all ribonucleotides.

[0033] In an optional embodiment, at least one internucleotide bond of the antisense oligonucleotide is a modified internucleotide bond. Further optionally, at least one internucleotide bond is a phosphate thioester internucleotide bond; further optionally, all internucleotide bonds are phosphate thioester internucleotide bonds.

[0034] In an optional embodiment, the antisense oligonucleotide contains at least one modified nucleotide.

[0035] In optional embodiments, the modified nucleotide includes one or more of the following: 2'-O-methoxyethyl modified nucleotides, 5'-methyl modified nucleotides, 2'-(S)-restricted ethyl modified nucleotides, 2'-O-methyl modified nucleotides, 2'-fluorine modified nucleotides, 2'-deoxy-modified nucleotides, locked nucleotides, non-locked nucleotides, configuration-restricted nucleotides, restricted ethyl nucleotides, base-free nucleotides, 2'-amino-modified nucleotides, 2'-O-allyl-modified nucleotides, 2'-C-alkyl-modified nucleotides, 2'-hydroxy-modified nucleotides, 2'-O-alkyl-modified nucleotides, morpholinyl nucleotides, bridging nucleic acids, peptide nucleic acids, locked nucleic acids, and non-locked nucleic acids. The modified nucleotide can be one or more modified nucleotides.

[0036] In an optional embodiment, the modified nucleotide includes nucleotides modified with 2'-O-methoxyethyl or nucleotides modified with 5'-methyl.

[0037] In an optional embodiment, the uracil ribonucleotide in the antisense oligonucleotide is a 5'-methyl modified nucleotide.

[0038] In an optional embodiment, the cytosine deoxyribonucleotide in the antisense oligonucleotide is a 5'-methyl modified nucleotide; more optionally, only the cytosine deoxyribonucleotide in the antisense oligonucleotide is a 5'-methyl modified nucleotide.

[0039] In an optional embodiment, at least one ribonucleotide is a 2'-O-methoxyethyl modified nucleotide.

[0040] In an optional embodiment, the antisense oligonucleotide is selected from the antisense oligonucleotides listed in Table 1 below.

[0041] In an optional embodiment, the antisense oligonucleotide is CRTH2aso-8: The sequence of CRTH2aso-8 is as follows: mA*mA*mG*mC*mC*dGdCdTdGdGdGdCdGdAdAdC*mA*mU*mG*mU*mU; In this sequence, positions 1 to 5 are all ribonucleotides, positions 6 to 15 are all deoxyribonucleotides, and positions 16 to 20 are all ribonucleotides; "*" indicates that the nucleoside bond between the two adjacent nucleotides to its left and right is a phosphate thioester bond; the lowercase letter "m" indicates that the uppercase letter to its right represents a nucleotide modified with 2'-O-methoxyethyl; the lowercase letter "d" indicates that the uppercase letter to its right represents a nucleotide modified with deoxyribonucleotide; the sequence is in the 5'-3' direction.

[0042] In a second aspect, a complex of an antisense oligonucleotide targeting CRTH2 mRNA is provided, the complex comprising: (a) the antisense oligonucleotide targeting CRTH2 mRNA as claimed in the first aspect; and (b) one or more targeting delivery ligands linked to (a), the targeting delivery ligands being, for example, but not limited to, nucleic acid aptamers, targeting peptides, or compound drugs.

[0043] Thirdly, an antisense oligonucleotide targeting CRTH2 mRNA, or the complex described in the second aspect, is provided for use in any of the following: (I) Inhibition of CRTH2 mRNA and / or reduction of CRTH2 protein expression in subjects for non-diagnostic and non-therapeutic purposes.

[0044] (II) Prepare drugs that inhibit CRTH2 mRNA and / or reduce CRTH2 protein expression in subjects.

[0045] (III) To prepare medicines for the treatment and / or prevention of pathological conditions or diseases caused by CRTH2 in the subject; In an optional implementation, the pathological condition or disease caused by the subject's CRTH2 includes inflammation, respiratory diseases, cardiovascular diseases, hypersensitivity reactions, or skin diseases. Optionally, the pathological conditions or diseases caused by the subject's CRTH2 include asthma (including allergic asthma, eosinophilic asthma, persistent asthma, severe asthma), chronic sinusitis with nasal polyps, nasal polyps, eosinophilia, end-stage renal disease, chronic obstructive pulmonary disease, rhinitis (including allergic rhinitis), conjunctivitis (including allergic conjunctivitis, atopic conjunctivitis), atopic dermatitis, androgenetic alopecia, colds, eosinophilic esophagitis, rhinovirus infection, or chronic urticaria.

[0046] Fourthly, a pharmaceutical composition is provided, the pharmaceutical composition comprising the antisense oligonucleotide targeting CRTH2 mRNA as described in the first aspect, or the complex as described in the second aspect.

[0047] In an optional embodiment, the pharmaceutical composition further comprises pharmaceutically acceptable optional excipients. The acceptable excipients may be any excipients known in the art and conventionally used. Examples of excipients include, but are not limited to, any physiologically compatible solvents, dispersion media, coatings, antibacterial agents, antifungal agents, pH adjusters, lyophilization protectants, and emulsifiers.

[0048] In optional embodiments, the pharmaceutical composition comprises an antisense oligonucleotide targeting CRTH2 mRNA or a delivery system for delivering the complex. The delivery cells include, but are not limited to, lipid nanoparticles, liposomes, nanoparticles, cationic lipids, cationic polymers, metal nanopolymers, nanorods, micelles, microvesicles, cell-penetrating peptides, viral particles, protein coats, or lipoglobules.

[0049] Fifthly, a method for inhibiting CRTH2 in a subject for non-diagnostic and therapeutic purposes is provided, the method comprising contacting the subject with an antisense oligonucleotide targeting CRTH2 mRNA as described in the first aspect, or a complex as described in the second aspect, or a pharmaceutical composition as described in the fourth aspect.

[0050] In an optional implementation, the subjects are selected from humans or cynomolgus monkeys and rhesus monkeys. Homology comparisons with humans, rats, mice, guinea pigs, cynomolgus monkeys, rhesus monkeys, New Zealand white rabbits, and beagles were performed. The CRTH2aso-8 described in the first aspect was able to match with humans, cynomolgus monkeys, rhesus monkeys, rats, mice, and guinea pigs in cases with ≤3 mismatches.

[0051] In an optional implementation, the subject is a cell, such as HuH-7 cells (human liver cancer cells).

[0052] In an optional implementation, the subject is a cell, and the contact includes delivering the antisense oligonucleotide targeting CRTH2 mRNA or the complex into the cell using any method known in the art.

[0053] In an optional implementation, the subject is a cell, and the working concentration of the antisense oligonucleotide targeting CRTH2 mRNA can be, for example, but is not limited to, 30 nM.

[0054] In an optional implementation, the subject is an individual organism, and the contact includes administering the drug to the subject via any suitable route known in the art, including but not limited to: oral administration, intravenous administration, intramuscular administration, subcutaneous administration, transdermal administration, airway administration (aerosol), pulmonary administration, nasal administration, ocular administration, or rectal administration.

[0055] The present invention will be further illustrated below with specific embodiments. However, it should be understood that these embodiments are merely for the purpose of more detailed illustration and should not be construed as limiting the present invention in any way.

[0056] Preparation Example: CRTH2 ASO Sequence Synthesis Steps 1. Synthesis preparation: Nucleosides, including T, A, C, G and U, were synthesized using standard phosphoramide structural units and solid-phase supports. The phosphoramide solution of all monomers used was a 0.05M acetonitrile solution. 2. Synthesis: A 200 nmol synthesis column made of CPG solid support was packed into an InstrumentPlus 192 synthesizer, and the specified sequence was synthesized using a phosphorusamide coupling method. For the coupling step, the phosphorusamide monomer was added in an amount approximately 70 times greater than the loading on the solid support. A 0.3 M BTT solution in acetonitrile was used as the activator in the coupling step, and phosphorusamide condensation was carried out for 4.5 min. All other steps were performed according to the manufacturer's standard protocol. Dimethoxytriphenylmethyl (DMT) was removed using a 3% trichloroacetic acid solution in dichloromethane. Thiophosphate bonds were introduced by sulfidation for 4 min with 0.05 M DDTT in a pyridine:acetonitrile = 2:3 (v / v). 3. Ammonolysis and Deprotection: After synthesizing the specified sequence, the specified sequence bound to the solid-phase support was placed in a 7M ammonia-methanol solution and ammonolysed at 90°C for 105 min. The solid-phase support was then filtered off, and the ammonia was removed under reduced pressure. If the sample contained TBDMS-protected native RNA, the resulting sample needed to be dissolved in DMSO, and a solution of triethylamine:triethylamine trifluoride = 2:1 (v / v) was added. TBDMS protection was then removed at 65°C for 150 min. After deprotection, 1.8 mL of ethanol was added to the sample, and the mixture was thoroughly vortexed and placed in a -20°C freezer for 2.5 h. The sample was then removed and centrifuged at 4°C and 12500 rpm for 10 min. The supernatant was removed, and the residue was treated with reduced pressure to remove residual ethanol. 4. Purification: Add 0.2 mL of water to the sample, shake thoroughly to dissolve, filter the sample using a 0.22 μm membrane, and then purify by high performance liquid chromatography to obtain the ASO sequence; 5. Quantitative dispensing: The purified sample is quantified using an ultra-micro spectrophotometer and dispensed and dried as required.

[0057] Example 1 1. The ASO sequence (20nt) used in this embodiment is shown in Table 1: Table 1: 30 CRTH2 ASO sequences

[0058] Note: In the table above, "*" indicates that the internucleotide bond between two adjacent nucleotides on its left and right sides is a phosphate thioester bond; using the Gapmer modification method (5nt RNA + 10nt DNA + 5nt RNA): the RNA bases are modified with 2'-O-methoxyethyl, m indicates that the nucleotide represented by the uppercase letter to its right is a nucleotide modified with 2'-O-methoxyethyl, and the lowercase letter "d" indicates that the nucleotide represented by the uppercase letter to its right is a deoxyribonucleotide.

[0059] 2. Experimental Procedure: 2.1 ASO transfection of CRTH2 target gene (30 nM): Cell count: HuH-7 cells; Cell seeding (48-well plate): On the day of transfection, dilute the cell suspension with fresh, pre-warmed complete culture medium and seed the cells into the plate. Transfection: Prepare 300 nM ASO transfection complex, add 25 μL of the prepared ASO transfection complex to each cell well, shake gently to mix, and incubate in a 37℃ cell culture incubator; after 24 h, collect the cells, add 450 μL of lysis buffer for subsequent experiments or store at -80℃.

[0060] 2.2 RNA extraction, reverse transcription to cDNA, and qPCR reaction: RNA was extracted using a fully automated nucleic acid extractor (Wuhan Namag: #S48) and matching nucleic acid extraction reagents (Wuhan Namag: #NMR0211), cDNA was synthesized, and the relative expression level of CRTH2 mRNA in cells of each experimental group was detected by qPCR.

[0061] 3. Experimental Results: The knockdown efficiency of 30 ASOs was verified at a concentration of 30 nM. The experimental results are shown in Table 2. Table 2: Knockdown efficiency statistics of 30 CRTH2 ASOs at a concentration of 30 nM

[0062] The main reagents and consumables used in the above embodiments are shown in Table 3.

[0063] Table 3: Reagents and Consumables

[0064] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, and not to limit them; although the present invention has been described in detail with reference to the foregoing embodiments, those skilled in the art should understand that modifications can still be made to the technical solutions described in the foregoing embodiments, or equivalent substitutions can be made to some or all of the technical features; and these modifications or substitutions do not cause the essence of the corresponding technical solutions to deviate from the scope of the technical solutions of the embodiments of the present invention.

Claims

1. An antisense oligonucleotide targeting CRTH2 mRNA, characterized in that, The nucleotide sequence of the antisense oligonucleotide comprises a sequence that differs from the sequence shown in SEQ ID NO. 1 to 30 by no more than 7 consecutive nucleotides from the (1st to 3rd) to (18th to 20th) positions.

2. The antisense oligonucleotide according to claim 1, characterized in that, The nucleotide sequence of the antisense oligonucleotide comprises a sequence in which the consecutive nucleotides from position (1-3) to position (18-20) of the sequence shown in SEQ ID NO.8 differ from the sequence shown in SEQ ID NO.8 by no more than 7 nucleotides. Optionally, the nucleotide sequence of the antisense oligonucleotide is the 1st to 20th consecutive nucleotides of the sequence shown in SEQ ID NO.8; or, the nucleotide sequence of the antisense oligonucleotide is the 3rd to 18th consecutive nucleotides of the sequence shown in SEQ ID NO.

8.

3. The antisense oligonucleotide according to claim 1, characterized in that, In any of the sequences shown in SEQ ID NO.1~30, positions 1 to 5 are all ribonucleotides, positions 6 to 15 are all deoxyribonucleotides, and positions 16 to 20 are all ribonucleotides. Optionally, the antisense oligonucleotide contains at least one modified nucleotide; Optionally, the modified nucleotides include one or more of the following: 2'-O-methoxyethyl modified nucleotides, 5'-methyl modified nucleotides, 2'-(S)-restricted ethyl modified nucleotides, 2'-O-methyl modified nucleotides, 2'-fluorine modified nucleotides, 2'-deoxy-modified nucleotides, locked nucleotides, non-locked nucleotides, configuration-restricted nucleotides, restricted ethyl nucleotides, base-free nucleotides, 2'-amino-modified nucleotides, 2'-O-allyl-modified nucleotides, 2'-C-alkyl-modified nucleotides, 2'-hydroxy-modified nucleotides, 2'-O-alkyl-modified nucleotides, morpholinyl nucleotides, bridging nucleic acids, peptide nucleic acids, locked nucleic acids, and non-locked nucleic acids. Optionally, the modified nucleotide includes one or more of 2'-O-methoxyethyl modified nucleotides and 5'-methyl modified nucleotides; Optionally, the uracil ribonucleotide in the antisense oligonucleotide is a 5'-methyl modified nucleotide; Optionally, the cytosine deoxyribonucleotide in the antisense oligonucleotide is a 5'-methyl modified nucleotide; more preferably, only the cytosine deoxyribonucleotide in the antisense oligonucleotide is a 5'-methyl modified nucleotide. Optionally, at least one ribonucleotide is a nucleotide modified with 2'-O-methoxyethyl.

4. The antisense oligonucleotide according to claim 1, characterized in that, At least one internucleotide bond is a modified internucleotide bond; Optionally, at least one nucleoside inter-bond is a phosphate thioside inter-bond; Optionally, all nucleoside inter-bonds are phosphate thioside inter-bonds.

5. The antisense oligonucleotide according to any one of claims 1 to 4, characterized in that, The antisense oligonucleotide is CRTH2aso-8: The sequence of CRTH2aso-8 is as follows: mA*mA*mG*mC*mC*dGdCdTdGdGdGdCdGdAdAdC*mA*mU*mG*mU*mU; In this sequence, positions 1 to 5 are all ribonucleotides, positions 6 to 15 are all deoxyribonucleotides, and positions 16 to 20 are all ribonucleotides; "*" indicates that the nucleoside bond between the two adjacent nucleotides on its left and right sides is a phosphate thioester bond; the lowercase letter "m" indicates that the uppercase letter to its right represents a nucleotide modified with 2'-O-methoxyethyl; the lowercase letter "d" indicates that the uppercase letter to its right represents a nucleotide modified with deoxyribonucleotide; the sequence is in the 5'-3' direction.

6. A complex of antisense oligonucleotides targeting CRTH2 mRNA, characterized in that, The complex contains: (a) the antisense oligonucleotide targeting CRTH2 mRNA as described in any one of claims 1 to 5; and (b) one or more targeted delivery ligands linked to (a).

7. The antisense oligonucleotide targeting CRTH2 mRNA according to any one of claims 1 to 5, or the complex according to claim 6, used in any one of the following: (I) Inhibition of CRTH2 mRNA and / or reduction of CRTH2 protein expression in subjects for non-diagnostic and non-therapeutic purposes; (II) Prepare drugs that inhibit CRTH2 mRNA and / or reduce CRTH2 protein expression in subjects; (III) Prepare medicines for the treatment and / or prevention of pathological conditions or diseases caused by CRTH2 in the subject.

8. The application according to claim 7, characterized in that, The pathological conditions or diseases caused by the subject's CRTH2 include inflammation, respiratory diseases, cardiovascular diseases, hypersensitivity reactions, or skin diseases; Optionally, the pathological conditions or diseases caused by the subject's CRTH2 include asthma, chronic sinusitis with nasal polyps, nasal polyps, eosinophilia, end-stage renal disease, chronic obstructive pulmonary disease, rhinitis, conjunctivitis, atopic dermatitis, androgenetic alopecia, colds, eosinophilic esophagitis, rhinovirus infection, or chronic urticaria.

9. A pharmaceutical composition, characterized in that, It comprises an antisense oligonucleotide targeting CRTH2 mRNA as described in any one of claims 1 to 5, or a complex as described in claim 6.

10. A method for inhibiting CRTH2 in a subject for non-diagnostic and non-therapeutic purposes, characterized in that, This includes contacting the subject with the antisense oligonucleotide targeting CRTH2 mRNA as described in any one of claims 1 to 5, or the complex as described in claim 6, or the pharmaceutical composition as described in claim 9.

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