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CN117679507A - IgM antibodies and uses thereof - Google Patents

IgM antibodies and uses thereof Download PDF

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Publication number
CN117679507A
CN117679507A CN202311808636.4A CN202311808636A CN117679507A CN 117679507 A CN117679507 A CN 117679507A CN 202311808636 A CN202311808636 A CN 202311808636A CN 117679507 A CN117679507 A CN 117679507A
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nanobody
rhinitis
seq
igm
amino acid
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Inventor
仇思念
陈明键
李澍
杨雨濛
杨继
刘春�
郜培杰
李莹
戴新悦
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Beijing Dongfangluo Biomedical Technology Co ltd
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Beijing Dongfangluo Biomedical Technology Co ltd
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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K16/00—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies
    • C07K16/42—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against immunoglobulins
    • C07K16/4283—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against immunoglobulins against an allotypic or isotypic determinant on Ig
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/50—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates
    • A61K47/51—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent
    • A61K47/62—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being a protein, peptide or polyamino acid
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P11/00—Drugs for disorders of the respiratory system
    • A61P11/02—Nasal agents, e.g. decongestants
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P37/00—Drugs for immunological or allergic disorders
    • A61P37/08—Antiallergic agents
    • 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
    • A61K2039/505—Medicinal preparations containing antigens or antibodies comprising antibodies

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  • Life Sciences & Earth Sciences (AREA)
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  • Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
  • General Chemical & Material Sciences (AREA)
  • Pulmonology (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Otolaryngology (AREA)
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Abstract

The present invention relates to IgM antibodies and uses thereof. In particular, the invention relates to the use of IgM antibodies in the manufacture of a medicament for the prevention and/or treatment of rhinitis.

Description

IgM antibodies and uses thereof
Technical Field
The invention belongs to the technical field of biological medicine. In particular, the invention relates to IgM antibodies and uses thereof for the prevention and/or treatment of rhinitis.
Background
IgM antibodies are the earliest expressed antibodies after exposure of the immune system to an immunogen, a class of immunoglobulins with the greatest molecular weight, and are predominantly distributed in serum. Classical secretory IgM forms pentamers via the J chain. The J chain in the pentameric IgM structure can act to stabilize the antibody structure.
In recent years, only a single variable domain nanobody has the advantages of small molecular weight, high thermal stability and chemical stability, low production cost, easy expression, simple humanization, high affinity, good solubility and the like, and has been paid attention to in the field of therapeutic drugs, and even has been directly used as an inhalant for treating respiratory tract virus infection. For example, the nanobody drug candidate ALX-0171 is a trivalent form of nanobody for the treatment of pediatric Respiratory Syncytial Virus (RSV) infection, which has entered phase ii in the clinic by aerosolized administration.
Rhinitis is a type of inflammation that occurs in the nasal mucosa caused by viruses, bacteria, allergens, various physicochemical factors, and certain systemic diseases. The existing medicines for treating rhinitis are mostly chemical medicines and have more side effects. The applicant has unexpectedly found that IgM antibodies can be used for treating and/or preventing rhinitis by nasal spray administration, and have good safety.
Disclosure of Invention
In one aspect, the invention provides the use of an IgM antibody in the manufacture of a medicament for the prevention and/or treatment of rhinitis.
In some embodiments, the IgM antibody is an IgM antibody formed from a nanobody fusion protein, which is a fusion protein comprising a nanobody and an Fc fragment, which may optionally be linked by a linker.
In some embodiments, the IgM antibody is an IgM pentamer formed from a nanobody fusion protein having a structure from N-terminus to C-terminus as shown in formula (I):
A-L-B(I)
wherein,
a is a nanobody;
b is an Fc fragment of human IgM;
l is (GGGGS) m, wherein m=0, 1, 2, 3 or 4.
In some embodiments, the nanobody comprises the following CDRs: CDR1 with the amino acid sequence shown as SEQ ID NO. 1, CDR2 with the amino acid sequence shown as SEQ ID NO. 2, and CDR3 with the amino acid sequence shown as SEQ ID NO. 3.
In some embodiments, the nanobody further comprises 4 framework regions FR1-4, the FR1-4 being staggered in sequence with the CDR1, CDR2 and CDR3. In a preferred embodiment, said FR1-4 is shown in SEQ ID NO. 4, 5, 6, 7, respectively.
In some embodiments, the nanobody has an amino acid sequence as set forth in SEQ ID NO. 8.
In some embodiments, the Fc fragment of the humanized IgM has an amino acid sequence as set forth in SEQ ID NO. 10.
In some embodiments, the nanobody fusion protein has an amino acid sequence as set forth in SEQ ID NO. 9.
In some embodiments, the IgM pentamer comprises a J chain having an amino acid sequence as shown in SEQ ID NO. 11.
In one aspect, the invention provides the use of a composition comprising one or more of IgM antibodies of the invention, a citric acid buffer system, sorbitol, naCl, polysorbate 20 in the manufacture of a medicament for the prevention and/or treatment of rhinitis.
In some embodiments, the medicament is in the form of a nasal spray, oral formulation, suppository, or parenteral formulation. In some embodiments, the nasal spray is selected from the group consisting of aerosols, sprays, and powder sprays. In some embodiments, the oral formulation is selected from the group consisting of tablets, powders, pills, powders, granules, fine granules, soft/hard capsules, film coatings, pellets, sublingual tablets, and ointments. In some embodiments, the parenteral formulation is a transdermal agent, an ointment, a plaster, a topical liquid, an injectable or a bolus formulation.
In some embodiments, the rhinitis is allergic rhinitis (allergic rhinitis).
Drawings
Fig. 1: schematic of IgM pentamer structure of nanobody.
Fig. 2: SDS PAGE identification result graph of MR molecule after affinity chromatography.
Fig. 3: rhinitis clinical symptom severity score change curve (14.5 days).
Detailed Description
The following description of the technical aspects of the present invention will be clear and complete, and it is apparent that the described embodiments are some embodiments of the present invention, but not all embodiments. All other embodiments, based on the embodiments of the invention, which are apparent to those of ordinary skill in the art without inventive faculty, are intended to be within the scope of the invention. Throughout the specification and claims, unless explicitly stated otherwise, the term "comprise" or variations thereof such as "comprises" or "comprising" or the like will be understood to include the stated element or component without excluding other elements or components.
In addition, numerous specific details are set forth in the following description in order to provide a better illustration of the invention. It will be understood by those skilled in the art that the present invention may be practiced without some of these specific details. In some embodiments, materials, elements, methods, means, etc. well known to those skilled in the art are not described in detail in order to highlight the gist of the present invention.
The present invention will be described in detail below.
Definition of the definition
"nanobodies", i.e. "heavy chain single domain antibodies", comprise only one heavy chain variable region (VHH), and light chains are naturally deleted compared to other antibodies.
Allergic rhinitis (allergic rhinitis, AR) is a allergic inflammation of the nasal mucosa mediated mainly by IgE, with major symptoms of paroxysmal sneezing, runny nose and nasal congestion, after a susceptible individual contacts the allergen. Seasonal Allergic Rhinitis (SAR) and Perennial Allergic Rhinitis (PAR) are classified according to the kind of allergen. Symptom onset of SAR is seasonal, common allergens are pollen, partial fungi, etc.; the symptom onset of PAR is perennial, and common allergens are dust mites, cockroaches, pet dander, etc. Some patients may be allergic to both seasonal and perennial allergens and symptoms may be perennial attacks with seasonal exacerbations.
In one aspect, the invention provides an IgM antibody formed from a nanobody fusion protein, e.g., an IgM antibody formed from a nanobody fusion protein and a J-chain, the nanobody fusion protein being a fusion protein comprising a nanobody and an Fc fragment, the nanobody and Fc fragment optionally being linked by a linker. In some embodiments, the IgM antibody is an IgM pentamer formed from a nanobody fusion protein, e.g., the IgM antibody is an IgM pentamer formed from a nanobody fusion protein and a J chain, the nanobody fusion protein is a fusion protein comprising a nanobody and an Fc fragment selected from the Fc fragment of a human IgM, the nanobody and the Fc fragment optionally being linked by a linker.
In some embodiments, the nanobody comprises the following CDRs:
CDR1 having an amino acid sequence shown in SEQ ID NO. 1 (i.e., GFTLDYYAIG),
CDR2 having the amino acid sequence shown in SEQ ID NO. 2 (i.e., CISSSDGSTSYADSVKG), and
the amino acid sequence is shown as CDR3 of SEQ ID NO. 3 (i.e., TPATYYSGRYYYQCPAGGMDY).
In some embodiments, the nanobody further comprises 4 framework regions FR1-4, the FR1-4 being staggered in sequence with the CDR1, CDR2 and CDR3. In a preferred embodiment, the amino acid sequences of FR1-4 are shown in SEQ ID NO. 4 (i.e., QVQLQESGGGLVQPGGSLRLSCAVS), SEQ ID NO. 5 (i.e., WFRQAPGKEREGVS), SEQ ID NO. 6 (i.e., RFTISRDNAKNTVYLQMNSLKPEDTALYYCAA) and SEQ ID NO. 7 (i.e., WGQGTQVTVSS), respectively.
In some embodiments, the nanobody has an amino acid sequence as set forth in SEQ ID NO. 8, or an amino acid sequence having at least 95%, 96%, 97%, 98% or 99% sequence identity to the amino acid sequence set forth in SEQ ID NO. 8. In a preferred embodiment, the nanobody has an amino acid sequence as shown in SEQ ID NO 8:
wherein the underlined parts are the framework regions FR1-4, respectively, and the darkened parts are CDR1, CDR2 and CDR3, respectively, of the heavy chain variable region.
In some embodiments, the Fc fragment of the humanized IgM has an amino acid sequence as set forth in SEQ ID NO. 10 (i.e., VIAELPPKVSVFVPPRDGFFGNPRKSKLICQATGFSPRQIQVSWLREGKQVGSGVTTDQVQAEAKESGPTTYKVTSTLTIKESDWLGQSMFTCRVDHRGLTFQQNASSMCVPDQDTAIRVFAIPPSFASIFLTKSTKLTCLVTDLTTYDSVTISWTRQNGEAVKTHTNISESHPNATFSAVGEASICEDDWNSGERFTCTVTHTDLPSPLKQTISRPKGVALHRPDVYLLPPAREQLNLRESATITCLVTGFSPADVFVQWMQRGQPLSPEKYVTSAPMPEPQAPGRYFAHSILTVSEEEWNTGETYTCVVAHEALPNRVTERTVDKSTGKPTLYNVSLVMSDTAGTCY), or an amino acid sequence having at least 95%, 96%, 97%, 98% or 99% sequence identity to the amino acid sequence set forth in SEQ ID NO. 10.
In some embodiments, the nanobody fusion protein has an amino acid sequence as set forth in SEQ ID No. 9 (i.e., QVQLQESGGGLVQPGGSLRLSCAVSGFTLDYYAIGWFRQAPGKEREGVSCISSSDGSTSYADSVKGRFTISRDNAKNTVYLQMNSLKPEDTALYYCAATPATYYSGRYYYQCPAGGMDYWGQGTQVTVSSVIAELPPKVSVFVPPRDGFFGNPRKSKLICQATGFSPRQIQVSWLREGKQVGSGVTTDQVQAEAKESGPTTYKVTSTLTIKESDWLGQSMFTCRVDHRGLTFQQNASSMCVPDQDTAIRVFAIPPSFASIFLTKSTKLTCLVTDLTTYDSVTISWTRQNGEAVKTHTNISESHPNATFSAVGEASICEDDWNSGERFTCTVTHTDLPSPLKQTISRPKGVALHRPDVYLLPPAREQLNLRESATITCLVTGFSPADVFVQWMQRGQPLSPEKYVTSAPMPEPQAPGRYFAHSILTVSEEEWNTGETYTCVVAHEALPNRVTERTVDKSTGKPTLYNVSLVMSDTAGTCY), or an amino acid sequence having at least 95%, 96%, 97%, 98% or 99% sequence identity to the amino acid sequence set forth in SEQ ID No. 9.
In some embodiments, the nanobody and Fc fragment are linked by a linker, wherein the linker is (GGGGS) m, wherein m = 0, 1, 2, 3, or 4. In a preferred embodiment, m=0. In a preferred embodiment, m=1.
In one aspect, the present invention provides an IgM pentamer formed from a nanobody fusion protein having a structure from N-terminus to C-terminus as shown in formula (I):
A-L-B(I)
wherein,
a is a nanobody; in specific embodiments, a is a single nanobody, as defined hereinabove; alternatively, a is a multivalent nanobody comprising two or more, preferably three, of said nanobodies linked by a linker, wherein the linker is (GGGGS) n, wherein n = 1, 2, 3 or 4, preferably n = 2 or 3;
b is an Fc fragment of human IgM; in specific embodiments, the Fc fragment of the humanized IgM has an amino acid sequence as set forth in SEQ ID NO. 10, or an amino acid sequence having at least 95%, 96%, 97%, 98% or 99% sequence identity to the amino acid sequence set forth in SEQ ID NO. 10;
l is (GGGGS) m, wherein m = 0, 1, 2, 3 or 4; in a preferred embodiment, m=0; in a preferred embodiment, m=1.
In some embodiments, the nanobody fusion protein has an amino acid sequence as set forth in SEQ ID NO. 9, or an amino acid sequence having at least 95%, 96%, 97%, 98% or 99% sequence identity to the amino acid sequence set forth in SEQ ID NO. 9.
In some embodiments, the IgM pentamer comprises a J chain having an amino acid sequence as set forth in SEQ ID No. 11, or an amino acid sequence having at least 95%, 96%, 97%, 98% or 99% sequence identity to the amino acid sequence set forth in SEQ ID No. 11.
In one aspect, the invention provides a pharmaceutical composition comprising a nanobody, nanobody fusion protein, or IgM pentamer of the invention, and a pharmaceutically acceptable carrier and/or excipient.
In some embodiments, the pharmaceutical composition may be in the form of a nasal spray, an oral formulation, a suppository, or a parenteral formulation.
In a preferred embodiment, the nasal spray is selected from the group consisting of aerosols, sprays and powder sprays.
In a preferred embodiment, the oral formulation is selected from the group consisting of tablets, powders, pills, powders, granules, fine granules, soft/hard capsules, film coatings, pellets, sublingual tablets and ointments.
In preferred embodiments, the parenteral formulation is a transdermal, ointment, plaster, topical liquid, injectable or bolus formulation.
The amount of the active ingredient to be administered of the pharmaceutical composition of the present invention varies depending on the administration subject, the organ to be administered, the symptoms, the administration method, etc., and can be determined by considering the type of the dosage form, the administration method, the age and weight of the patient, the symptoms of the patient, etc., and the judgment of the doctor.
In one aspect, the invention provides the use of IgM antibodies in the manufacture of a medicament for the prevention and/or treatment of rhinitis. In some embodiments, the IgM antibody is an IgM antibody formed from a nanobody fusion protein, which is a fusion protein comprising a nanobody and an Fc fragment, which may optionally be linked by a linker. In some embodiments, the Fc fragment of the IgM antibody is selected from the Fc fragment of a human IgM. In some embodiments, the IgM antibody is in pentameric form, the IgM antibody comprises a J chain of human IgM. In some embodiments, the IgM antibody is in hexamer form.
In one aspect, the present invention provides the use of a nanobody, nanobody fusion protein or IgM pentamer formed from a nanobody fusion protein of the invention or a pharmaceutical composition or formulation or therapeutic agent comprising the same in the manufacture of a medicament for the prevention and/or treatment of rhinitis.
In one aspect, the invention provides the use of a composition comprising one or more of IgM antibodies of the invention, a citric acid buffer system, sorbitol, naCl, polysorbate 20 in the manufacture of a medicament for the prevention and/or treatment of rhinitis. In some embodiments, a bacteriostatic agent, such as benzalkonium chloride, may optionally be included in the composition.
In some embodiments, the invention provides the use of a composition comprising one or more of IgM antibodies of the invention, a citric acid buffer system, sorbitol, naCl, polysorbate 20, benzalkonium chloride, in the manufacture of a medicament for the prevention and/or treatment of rhinitis.
In some embodiments, the composition comprises about 0.1-20mg/mL IgM antibody. In some embodiments, the composition comprises about 10-30mM citric acid buffer system. In some embodiments, the composition comprises about 20-35mg/mL sorbitol. In some embodiments, the composition comprises about 40-65mM NaCl. In some embodiments, the composition comprises about 0.1 to about 0.4mg/mL polysorbate 20. In some embodiments, the composition comprises benzalkonium chloride in the range of about 0.05-0.3mg/mL. In some embodiments, the pH of the composition is about 5.6 to 6.4.
In some embodiments, the composition comprises: igM antibodies about 0.1-20mg/mL; about 10-30mM citric acid buffer system; sorbitol is about 20-35mg/mL; about 40-65mM NaCl; about 0.1 to about 0.4mg/mL of polysorbate 20; and water; and the pH of the composition is about 5.6 to 6.4. In some embodiments, the composition comprises: igM antibodies about 0.1-20mg/mL; about 10-30mM citric acid buffer system; sorbitol is about 20-35mg/mL; about 40-65mM NaCl; about 0.1 to about 0.4mg/mL of polysorbate 20; about 0.1-0.2mg/mL benzalkonium chloride; and water; and the pH of the composition is about 5.6 to 6.4.
In some embodiments, the composition comprises: igM antibodies about 1-20mg/mL; about 10-20mM citric acid buffer system; sorbitol is about 20-35mg/mL; about 40-60mM NaCl; about 0.2 to about 0.4mg/mL polysorbate 20; and water; and the pH of the composition is about 5.6 to 6.4. In some embodiments, the composition comprises: igM antibodies about 1-20mg/mL; about 10-20mM citric acid buffer system; sorbitol is about 20-35mg/mL; about 40-60mM NaCl; about 0.2 to about 0.4mg/mL polysorbate 20; about 0.15-0.2mg/mL benzalkonium chloride; and water; and the pH of the composition is about 5.6 to 6.4.
In some embodiments, the composition comprises: igM antibodies about 1mg/mL or about 5mg/mL; about 20mM citrate buffer system; sorbitol about 30mg/mL; naCl about 60mM (3.5 mg/mL); polysorbate 20 about 0.3 or about 0.4mg/mL; and water; and the pH of the composition is about 6.0. In some embodiments, the composition comprises: igM antibodies about 1mg/mL or about 5mg/mL; about 20mM citrate buffer system; sorbitol about 30mg/mL; naCl about 60mM (3.5 mg/mL); polysorbate 20 about 0.3 or about 0.4mg/mL; about 0.2mg/mL benzalkonium chloride; and water; and the pH of the composition is about 6.0.
In some embodiments, the rhinitis is allergic rhinitis (allergic rhinitis).
In some specific embodiments, rhinitis of the present invention includes various types of rhinitis:
the rhinitis of the present invention is inflammation of nasal mucosa, and includes rhinitis with symptoms such as sneeze, runny nose (nasal discharge), nasal obstruction (nasal obstruction), and nasal itching. The antibody or therapeutic agent of the present invention can be applied to various rhinitis.
More specifically, rhinitis to which the antibody or therapeutic agent of the present invention can be applied is infectious rhinitis, allergic non-infectious rhinitis, irritant rhinitis, atrophic rhinitis, or specific granulomatous rhinitis.
The infectious rhinitis may be any of acute rhinitis or chronic rhinitis. By using the antibody or therapeutic agent of the present invention, sneeze, excessive rhinorrhea (nasal discharge, runny nose), nasal obstruction (nasal obstruction), nasal itching, dysosmia, etc. can be effectively and early cured.
For allergic non-infectious rhinitis, it may be combined type (nasal allergy) rhinitis including allergic rhinitis and non-allergic rhinitis, including rhinitis of the nasal leakage type including gustatory rhinitis, cold air inhalation rhinitis and senile rhinitis, congestion type rhinitis including drug rhinitis, cardiac rhinitis, gestational rhinitis, endocrine rhinitis and cold rhinitis, or dry type rhinitis.
The rhinitis of the present invention is preferably allergic rhinitis (allergic rhinitis) or non-allergic rhinitis, and the allergic rhinitis may be perennial allergic rhinitis or seasonal allergic rhinitis. The antibody or therapeutic agent of the present invention shows excellent efficacy and safety as a therapeutic agent for allergic rhinitis which is difficult to cure or alleviate for a long period of time, particularly perennial allergic rhinitis caused by house dust or mites.
The antibody or the drug is also effective as a therapeutic agent for various rhinitis such as sneeze, runny nose (nasal discharge), nasal obstruction (nasal obstruction) and nasal itching, which are caused by irritative rhinitis such as physical rhinitis, chemical rhinitis and radioactive rhinitis, atrophic rhinitis and specific granulomatous rhinitis. Further, the antibody or therapeutic agent of the present invention can be effectively used for full-charge rhinitis, nasal obstruction type rhinitis, or sneeze rhinorrhea type rhinitis.
The sequences involved in the following examples of the present invention were all synthesized by the biological technologies, inc., nanjing gold Style; the related plasmid extraction and agarose gel recovery kit is purchased from OMEGA and is subjected to standard operation according to the specification; the restriction enzymes used were all purchased from Thermo fisher and the T4 DNA ligase from Invitrogen.
Some commonly used biological materials, such as competent cells and cells to be transformed, are also commercially available products, e.g., trans 5. Alpha. Competent available from TransGene Biotech;CHO K1 was purchased from Merck company, usa.
Example 1: igM pentameric (MR) antibodies formed from nanobody fusion proteins
The structure of the nanometer antibody IgM pentamer is shown IN figure 1, and the antibody variable region adopts nanometer antibody R (SEQ IN NO: 8). The IgM pentamer is formed by self-assembly of a nanometer antibody fusion protein R-Fc with an amino acid sequence shown as SEQ ID NO. 9 and a J chain with an amino acid sequence shown as SEQ ID NO. 11. R-Fc and J chains are expressed by the mammalian host cell CHO K1, and self-assemble within the cell to form the pentameric IgM antibody. Expressed cell supernatant proteins were purified and then identified by SDS-PAGE. As shown in FIG. 2, sample1 is cell supernatant and Sample2 is purified IgM pentamer (MR).
Example 2: igM antibodies formed from nanobody fusion proteins for rhinitis study
Treatment and alleviation effects on symptoms of allergic rhinitis were observed in 49 patients with allergic rhinitis using IgM antibody nasal spray (containing about 1mg/mL of MR IgM antibody; about 20mM of citric acid buffer system; about 30mg/mL of sorbitol; about 60mM (3.5 mg/mL) of NaCl; about 0.3mg/mL of polysorbate 20).
Patients with allergic rhinitis should meet the following requirements:
■ The patients with the seasonal aggravated SAR and PAR are identified, the disease course is more than or equal to 2 years, the disease occurs in the attack season, the time of each disease is less than 2 months, and the disease time is less than or equal to 1 week
■ The age is more than or equal to 18 years old and less than or equal to 60 years old
■ At least two main nose symptoms are above moderate (score not less than 2 points)
Nasal spray application method
■ Spraying once every morning and spraying on two sides of nose for 2 weeks
■ If symptoms are not relieved, nasal spray can be added for 2 times per day
Allergic rhinitis clinical symptom severity score
■ Symptoms were scored in 0-3 scores according to severity scoring criteria (table 1) for clinical symptoms of allergic rhinitis (rhinocnesmus, sneeze, runny nose, nasal congestion), control symptoms appearance.
Table 1: clinical symptom severity scoring criteria for allergic rhinitis
■ Symptoms were scored before the first nasal spray, after the morning of the nasal spray, and before sleep (about 10-30 minutes after nasal spray), respectively, and recorded for 2 weeks.
Research results and conclusions
Each patient was continuously recorded for 2 weeks, once in the morning and once in the evening (integer in the morning and 0.5 in the evening) for a total of 14.5 days. The results are shown in FIG. 3.
Wherein, the total nasal symptom score (TNSS, total rhinitis symptom score calculated by summing the symptom scores) before use is 6.9 score on average, TNSS score on 14.5 days after use is 3.2 score, and the reduction is 3.7 score. The total TNSS score showed a downward trend throughout 14 days of continuous use of the nasal spray, with the downward trend being more pronounced for the first four days, with the total TNSS decrease approaching plateau from day 8.
For single symptoms, the average values of nasal itching, sneezing, nasal discharge and nasal obstruction symptoms before treatment are respectively 1.6 minutes, 2.0 minutes and 1.7 minutes, and the average values of the symptoms at 14.5 days after treatment are respectively 0.8 minutes, 0.7 minutes, 1.0 minutes and 0.7 minutes, and the scores of the symptoms are wholly reduced in the process of continuously using the nasal spray for 14 days, and the descending trend among the symptoms is not obviously different.
Particularly, for individual patients with severe rhinitis, the scores of four symptoms of nasal itching, sneezing, nasal discharge and nasal obstruction were reduced from 3, 2, 3 and 3 to 0 respectively from day 9, and then the scores of symptoms on day 14 were 0, 0 and 1 respectively after one administration on day 11, and only mild nasal obstruction symptoms were present. No obvious discomfort after the nasal spray is used.
The IgM antibody nasal spray provided by the invention has a good effect on treating rhinitis patients, can improve symptoms of nasal itching, sneeze, nasal discharge and nasal obstruction, and has good safety and no obvious discomfort.
Sequence listing
CDR1 sequences of VHH chains of SEQ ID NO. 1 nanobody R
GFTLDYYAIG
CDR2 sequences of VHH chains of SEQ ID NO. 2 nanobody R
CISSSDGSTSYADSVKG
CDR3 sequences of VHH chains of SEQ ID NO 3 nanobody R
TPATYYSGRYYYQCPAGGMDY
FR1 sequence of VHH chain of SEQ ID NO. 4 nanobody R
QVQLQESGGGLVQPGGSLRLSCAVS
FR2 sequence of VHH chain of SEQ ID NO. 5 nanobody R
WFRQAPGKEREGVS
FR3 sequence of VHH chain of SEQ ID NO. 6 nanobody R
RFTISRDNAKNTVYLQMNSLKPEDTALYYCAA
FR4 sequence of VHH chain of SEQ ID NO. 7 nanobody R
WGQGTQVTVSS
Amino acid sequence of VHH chain of SEQ ID NO 8 nanobody R
QVQLQESGGGLVQPGGSLRLSCAVSGFTLDYYAIGWFRQAPGKEREGVSCISSSDGSTSYADSVKGRFTISRDNAKNTVYLQMNSLKPEDTALYYCAATPATYYSGRYYYQCPAGGMDYWGQGTQVTVSS
Amino acid sequence of SEQ ID NO 9 nanobody fusion protein R-Fc
QVQLQESGGGLVQPGGSLRLSCAVSGFTLDYYAIGWFRQAPGKEREGVSCISSSDGSTSYADSVKGRFTISRDNAKNTVYLQMNSLKPEDTALYYCAATPATYYSGRYYYQCPAGGMDYWGQGTQVTVSSVIAELPPKVSVFVPPRDGFFGNPRKSKLICQATGFSPRQIQVSWLREGKQVGSGVTTDQVQAEAKESGPTTYKVTSTLTIKESDWLGQSMFTCRVDHRGLTFQQNASSMCVPDQDTAIRVFAIPPSFASIFLTKSTKLTCLVTDLTTYDSVTISWTRQNGEAVKTHTNISESHPNATFSAVGEASICEDDWNSGERFTCTVTHTDLPSPLKQTISRPKGVALHRPDVYLLPPAREQLNLRESATITCLVTGFSPADVFVQWMQRGQPLSPEKYVTSAPMPEPQAPGRYFAHSILTVSEEEWNTGETYTCVVAHEALPNRVTERTVDKSTGKPTLYNVSLVMSDTAGTCY
Fc sequence of human IgM antibody of SEQ ID NO 10
VIAELPPKVSVFVPPRDGFFGNPRKSKLICQATGFSPRQIQVSWLREGKQVGSGVTTDQVQAEAKESGPTTYKVTSTLTIKESDWLGQSMFTCRVDHRGLTFQQNASSMCVPDQDTAIRVFAIPPSFASIFLTKSTKLTCLVTDLTTYDSVTISWTRQNGEAVKTHTNISESHPNATFSAVGEASICEDDWNSGERFTCTVTHTDLPSPLKQTISRPKGVALHRPDVYLLPPAREQLNLRESATITCLVTGFSPADVFVQWMQRGQPLSPEKYVTSAPMPEPQAPGRYFAHSILTVSEEEWNTGETYTCVVAHEALPNRVTERTVDKSTGKPTLYNVSLVMSDTAGTCY
SEQ ID NO. 11 human IgM antibody J chain sequence
QEDERIVLVDNKCKCARITSRIIRSSEDPNEDIVERNIRIIVPLNNRENISDPTSPLRTRFVYHLSDLCKKCDPTEVELDNQIVTATQSNICDEDSATETCYTYDRNKCYTAVVPLVYGGETKMVETALTPDACYPD。

Claims (10)

  1. The use of IgM antibodies in the manufacture of a medicament for the prevention and/or treatment of rhinitis,
    preferably, the IgM antibody is an IgM antibody formed by a nanobody fusion protein and a J chain, the nanobody fusion protein being a fusion protein comprising a nanobody and an Fc fragment, the nanobody and Fc fragment optionally being linked by a linker; and is also provided with
    Preferably, the rhinitis is allergic rhinitis (allergic rhinitis).
  2. 2. The use of claim 1, wherein the IgM antibody is an IgM pentamer formed from a nanobody fusion protein having the structure from N-terminus to C-terminus as shown in formula (I):
    A-L-B(I)
    wherein,
    a is a nanobody;
    b is an Fc fragment of human IgM;
    l is (GGGGS) m, wherein m=0, 1, 2, 3 or 4.
  3. 3. The use of claim 1 or 2, wherein the nanobody comprises the following CDRs: CDR1 with the amino acid sequence shown as SEQ ID NO. 1, CDR2 with the amino acid sequence shown as SEQ ID NO. 2, and CDR3 with the amino acid sequence shown as SEQ ID NO. 3.
  4. 4. The use of any one of claims 1-3, wherein the nanobody further comprises 4 framework regions FR1-4, the FR1-4 being staggered in sequence with the CDR1, CDR2 and CDR 3; preferably, the FR1-4 is shown in SEQ ID NO. 4, 5, 6, 7, respectively.
  5. 5. The use of any one of claims 1-4, wherein the nanobody has an amino acid sequence as set forth in SEQ ID No. 8.
  6. 6. The use according to any one of claims 1 to 5, wherein the Fc fragment of human IgM has the amino acid sequence shown in SEQ ID No. 10.
  7. 7. The use of any one of claims 1-6, wherein the nanobody fusion protein has an amino acid sequence as set forth in SEQ ID No. 9.
  8. 8. The use according to any one of claims 1 to 7, wherein the IgM pentamer comprises J chains having the amino acid sequence as shown in SEQ ID No. 11.
  9. 9. Use of a composition comprising one or more of IgM antibodies as claimed in any one of claims 1 to 8, citric acid buffer system, sorbitol, naCl, polysorbate 20 in the manufacture of a medicament for the prevention and/or treatment of rhinitis; and is also provided with
    Preferably, the rhinitis is allergic rhinitis (allergic rhinitis).
  10. 10. The use of any one of claims 1-9, wherein the medicament is in the form of a nasal spray, an oral formulation, a suppository or a parenteral formulation;
    preferably, the nasal spray is selected from the group consisting of aerosols, sprays and powder sprays;
    preferably, the oral formulation is selected from the group consisting of tablets, powders, pills, powders, granules, fine granules, soft/hard capsules, film coatings, pellets, sublingual tablets and ointments;
    preferably, the parenteral formulation is a transdermal agent, an ointment, a plaster, a topical liquid, an injectable or a bolus formulation.
CN202311808636.4A 2023-01-05 2023-12-26 IgM antibodies and uses thereof Pending CN117679507A (en)

Applications Claiming Priority (4)

Application Number Priority Date Filing Date Title
CN2023100137126 2023-01-05
CN202310013712 2023-01-05
CN2023107391965 2023-06-21
CN202310739196 2023-06-21

Publications (1)

Publication Number Publication Date
CN117679507A true CN117679507A (en) 2024-03-12

Family

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Application Number Title Priority Date Filing Date
CN202311808636.4A Pending CN117679507A (en) 2023-01-05 2023-12-26 IgM antibodies and uses thereof

Country Status (1)

Country Link
CN (1) CN117679507A (en)

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