WO2026017005A1 - Aqueous pharmaceutical product for treating axial spondyloarthritis and use - Google Patents

Abstract

The present invention provides an aqueous pharmaceutical product comprising an anti-IL-17A antibody, and a use of the aqueous pharmaceutical product in the preparation of a drug for treating axial spondyloarthritis.

Description

An aqueous pharmaceutical product for treating axial spondyloarthritis and its uses

Related applications

This application claims priority and related interests in Chinese Patent Application No. 202410946890.9, filed on July 15, 2024, the entire contents of which are incorporated herein by reference.

Technical Field

This application belongs to the field of biopharmaceuticals, specifically relating to an aqueous pharmaceutical product and its use for treating axial spondyloarthritis.

Background Technology

Axial spondyloarthritis (axSpA) is a collective term for ankylosing spondylitis (AS) and radiation-negative axial spondyloarthritis. With a deeper understanding of the disease, axSpA has gradually replaced AS in treatment guidelines. axSpA is a common chronic inflammatory autoimmune disease, predominantly affecting men aged 20-30. It is characterized by sacroiliitis, spondylitis, and tendinitis, progressively restricting spinal movement and even causing deformities. It can also cause varying degrees of damage to extra-articular tissues such as the eyes, lungs, and intestines, significantly impacting patients' quality of life and financial burden.

The treatment goals for axial spondyloarthritis (axSpA) are to alleviate symptoms, improve and enhance spinal flexibility, reduce functional limitations, maintain work capacity, and reduce disease-related complications. Currently, axSpA treatment primarily utilizes nonsteroidal anti-inflammatory drugs (NASIDs), tumor necrosis factor-alpha (TNF-α) antagonists, and disease-modifying antirheumatic drugs (DMARDs), but there is no standard treatment to achieve target remission or maintain remission. NASIDs can reduce joint pain and stiffness and increase joint mobility. In 2010, the American College of Rheumatology/European League Against Rheumatism recommended NASIDs as first-line treatment for early-stage axSpA, suggesting that patients try at least two NASIDs before using TNF-α antagonists. However, their side effects limit clinical application, requiring individualized drug selection based on the patient's liver and kidney function, gastrointestinal history, and cardiovascular disease.

The GR1501 antibody molecule has been shown to have good efficacy in treating psoriasis, but there are currently no studies on its efficacy in treating axial spondyloarthritis.

Summary of the Invention

This application aims to provide an aqueous pharmaceutical product and its application in the treatment of axial spondyloarthritis.

The first aspect of this application provides the use of an aqueous pharmaceutical product in the preparation of a medicament for treating axial spondyloarthritis, such as ankylosing spondylitis, wherein the antibody aqueous pharmaceutical product comprises an antibody that specifically binds to IL-17A, the antibody comprising a VH having the following three CDRs:

CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 12, CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 13, and CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 14; and

The antibody comprises a VL having the following three CDRs:

CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 15, CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 16, and CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 17;

Furthermore, the sequence of CDRs is defined according to Chothia.

Based on the above technical solution, the following improvements can be made to this application.

In some embodiments, the antibody comprises VH and VL, wherein VH comprises the amino acid sequence shown in SEQ ID NO: 7, and VL comprises the amino acid sequence shown in SEQ ID NO: 4.

In some embodiments, the antibody comprises a heavy chain and a light chain, the heavy chain comprising an amino acid sequence as shown in SEQ ID NO: 18, and the light chain comprising an amino acid sequence as shown in SEQ ID NO: 19.

In some embodiments, the drug is formulated to be contained in containers, each container having a sufficient quantity of the aqueous drug product to allow delivery of the following unit doses:

a) A total of 6 doses, administered in 2-dose regimens, each dose being a subcutaneous dose of approximately 100 mg to approximately 300 mg, delivered at weeks 0, 2, and 4; and

b) Subsequently, weekly delivery of two doses is initiated after the subcutaneous delivery of the fourth week as described in step a), with each dose being approximately 100 mg to approximately 300 mg.

In some embodiments, the dosage for each administration in step a) and step b) is about 100 mg, or about 200 mg, or about 300 mg.

In some embodiments, the dosage for each administration in step a) and step b) is approximately 200 mg.

In some embodiments, the aqueous pharmaceutical product further comprises trehalose dihydrate, L-histidine, histidine hydrochloride monohydrate, L-methionine, and polysorbate 80.

In some implementations, the antibody content is approximately 100 mg/ml.

In some embodiments, the content of L-histidine is about 1.241 mg/ml, the content of histidine hydrochloride monohydrate is about 2.516 mg/ml, the content of trehalose dihydrate is about 75.67 mg/ml, the content of polysorbate 80 is about 0.2 mg/ml, and the content of L-methionine is about 0.746 mg/ml.

In some implementations, the pH of the aqueous pharmaceutical product is approximately 5.8.

In some embodiments, the aqueous pharmaceutical product is GR1501.

A second aspect of this application provides an aqueous pharmaceutical product comprising an anti-IL-17A antibody and a buffer solution, said antibody comprising a VH having the following three CDRs:

CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 12, CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 13, and CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 14; and

The antibody comprises a VL having the following three CDRs:

CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 15, CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 16, and CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 17.

Based on the above technical solution, the following improvements can be made to this application.

In some embodiments, the antibody comprises VH and VL, wherein VH comprises the amino acid sequence shown in SEQ ID NO: 7, and VL comprises the amino acid sequence shown in SEQ ID NO: 4.

In some embodiments, the antibody comprises a heavy chain and a light chain, the heavy chain comprising an amino acid sequence as shown in SEQ ID NO: 18, and the light chain comprising an amino acid sequence as shown in SEQ ID NO: 19.

In some implementations, the aforementioned aqueous pharmaceutical products also include antioxidants.

In some implementations, the aforementioned aqueous pharmaceutical products also include osmotic pressure regulators.

In some implementations, the aforementioned aqueous pharmaceutical products also include stabilizers.

In some embodiments, the antioxidant is L-methionine.

In some embodiments, the buffer solution is histidine hydrochloride, preferably, the buffer solution is L-histidine and histidine hydrochloride monohydrate.

In some embodiments, the osmotic pressure regulator is trehalose, preferably trehalose dihydrate.

In some embodiments, the stabilizer is polysorbate 80.

In some embodiments, the aqueous pharmaceutical product comprises about 100 mg/ml to about 150 mg/ml of anti-IL-17A antibody, about 70 mg/ml to about 80 mg/ml of trehalose dihydrate, about 1 mg/ml to 2 mg/ml of L-histidine, about 2 mg/ml to about 4 mg/ml of histidine hydrochloride monohydrate, about 0.5 mg/ml to about 1 mg/ml of L-methionine, and about 0.1 mg/ml to about 0.3 mg/ml of polysorbate 80.

In some embodiments, the aqueous pharmaceutical product comprises about 100 mg/ml of anti-IL-17A antibody, about 75.67 mg/ml of trehalose dihydrate, about 1.241 mg/ml of L-histidine, about 2.516 mg/ml of histidine hydrochloride monohydrate, about 0.746 mg/ml of L-methionine, and about 0.2 mg/ml of polysorbate 80.

In some embodiments, the pH of the aqueous pharmaceutical product is about 5.5 to about 6.0, preferably 5.8.

In some embodiments, the product is used to prepare a medicament for treating axial spondyloarthritis.

The third aspect of this application provides the aqueous pharmaceutical product described in the second aspect above, for the treatment of axial spondyloarthritis.

The fourth aspect of this application provides a method for treating axial spondyloarthritis, comprising administering a therapeutically effective amount of the aqueous pharmaceutical product described in the second aspect above to an individual in need, such as a patient suffering from axial spondyloarthritis. In some embodiments, the axial spondyloarthritis described above is active axial spondyloarthritis.

In some implementations, the aforementioned axial spondyloarthritis is radiographically positive axial spondyloarthritis, also known as ankylosing spondylitis.

This application has one or more of the following beneficial effects: the aqueous drug product of this application has a more stable formulation and can effectively treat axial spondyloarthritis, bringing good therapeutic effects to patients; the product quality is more stable; and the aqueous drug product can produce better efficacy. The specific container assembly ratio of the aqueous drug product of this application ensures the therapeutic effect during medication.

Attached Figure Description

Figure 1 shows the flowchart of the Phase II clinical trial GR1501-007;

Figure 2 shows the diagnostic diagram of the linear relationship of C _max ;

Figure 3 shows the diagnostic plot of the linear relationship between AUC _inf ;

Figure 4 shows a forest plot (FAS filling) of the proportion of patients who achieved ASAS20 at week 16.

Figure 5 shows a forest plot (PPS) of the proportion of patients who achieved ASAS20 at week 16.

Figure 6 shows the flowchart of the Phase III clinical trial GR1501-007;

Figure 7-1 shows the stability-HIC purity of the first round of formulation screening;

Figure 7-2 shows the stability-SEC monomer purity of the first round of formulation screening;

Detailed Implementation

The principles and features of this application are described below with reference to the accompanying drawings. The examples given are only for explaining this application and are not intended to limit the scope of this application.

The abbreviations in this application are explained in the table below.

Information regarding some of the sequences involved in this application is described in the table below:

Detailed Description of the Invention

Terminology definition:

As used herein, the terms “ankylosing spondylitis,” “AS,” and “vertebral arthropathy” refer to inflammatory arthritis characterized by chronic inflammation of the joints, which may include the sacrum in the spine and pelvis and may lead to eventual fusion of the spine. Patients with AS can be diagnosed using the modified New York criteria for AS or the ASAS axial SPA criteria (2009). In some embodiments of the disclosed methods, protocols, uses, kits, and pharmaceutical compositions, patients have AS.

2009 International Association for the Assessment of Spondyloarthritis Classification (AxSpA)

Patients with onset age <45 years and lower back pain ≥3 months, plus meeting one of the following criteria:

1. Imaging findings suggest sacroiliitis plus ≥1 SpA feature;

2. HLA-B27 positive plus ≥2 SpA features.

SpA characteristics include: (1) inflammatory back pain; (2) arthritis; (3) origin-end-point inflammation (Achilles tendon); (4) uveitis; (5) dactylitis; (6) psoriasis; (7) Crohn's disease/ulcerative colitis; (8) good response to nonsteroidal anti-inflammatory drugs; (9) family history of SpA; (10) HLA-B27 positivity; (11) elevated CRP.

*Imaging findings suggest sacroiliitis refers to MRI showing active (acute) inflammation of the sacroiliac joint, highly suggestive of sacroiliitis associated with SpA; or X-ray findings consistent with the imaging changes of sacroiliitis defined in the 1984 New York criteria: bilateral grade 2-4 or unilateral grade 3-4 sacroiliitis.

In searching for predictive indicators of response to IL-17 binding molecules (e.g., IL-17 antibodies, such as celecoxib) in AS patients, we referenced the EU's "Guideline on the Clinical Investigation of Medicinal Products for the Treatment of Axial Spondyloarthritis" and the 2009 "The Assessment of Spondyloarthritis International Society (ASAS) Handbook: a guide to assess spondyloarthritis." We also incorporated clinical trial designs for other drugs treating ankylosing spondylitis, ultimately setting the primary efficacy endpoint as the ASAS20 response rate, with secondary endpoints including the ASAS40 response rate. The ASAS20 (International Society for the Assessment of Spondyloarthritis 20% response rate) is an important indicator for evaluating the treatment efficacy of ankylosing spondylitis (AS) and axial spondyloarthritis (axSpA). Subjects meeting the following criteria were considered to have achieved an ASAS20 response. Compared with baseline, at least three of the four indicators (including overall patient assessment; spinal pain assessment; BASFI assessment; BASDAI assessment) showed an improvement of >20% with an absolute improvement of ≥1 point, while the remaining indicators did not show a deterioration of ≥20% or an increase of ≥1 point.

The term “comprising” encompasses both “including” and “consisting of”, for example, a composition that “comprising” X may consist of only X or may include other things, such as X+Y.

Unless the context otherwise indicates, the term “about” in relation to the numerical value x means +/- 10%. When used with respect to pharmacokinetic (PK) parameters (e.g., AUC, _Cmax , _tmax , trough level, etc.), the term “about” means that a person skilled in the art would consider a treatment (e.g., dosage and/or dosing regimen) to be bioequivalent to a control treatment. For bioequivalence, the standard method for demonstrating bioequivalence is to statistically demonstrate that the ratio of a given PK parameter (e.g., AUC, _Cmax ) between two treatments (i.e., the control treatment and the test treatment) is between 0.8 and 1.25, indicated by a 90% confidence interval (CI) around that ratio (the lower limit of this CI is above 0.8, and the upper limit is below 1.25). Thus, for example, if a control _Cmax of 10 μg/ml is obtained during an experiment comparing the PK curves of the control and test treatments, the test treatment would be considered “about 10 μg/ml” if a person skilled in the art considers the test treatment to be bioequivalent. As used herein, pharmacokinetic terms such as _tmax , t1 _/2 , AUC, AUC _(0-τ) (AUC up to the end of a given dosing period, hereinafter “ _AUCτ ”), and _Cmax have the meanings accepted in the art.

The term “administration” in relation to compounds (such as IL-17 binding molecules or antirheumatic drugs) is used to refer to the delivery of the compound via any route.

The term "analysis" is used to refer to the act of detecting, identifying, screening, or determining, which can be performed by any conventional method. For example, the presence of a specific marker in a sample can be detected by analyzing the presence of that marker using methods such as ELISA analysis, Northern blot, or imaging.

The term "substantially" does not exclude "completely," for example, a composition that is "substantially free" of Y may be completely free of Y. The term "substantially" may be omitted from the definition in this application if necessary.

The term "mg/kg" as used in this article refers to mg of drug per kg of the patient's body weight to which the drug is administered.

Antibodies that specifically bind to IL-17A

The structure, amino acid sequence, preparation method and biological activity of the antibody against IL-17A described in this application have been described in Chinese Patent Application No. CN201510097117.0. The entire contents of Chinese Patent Application No. CN201510097117.0 are hereby incorporated herein by reference.

In some embodiments, the anti-IL-17 monoclonal antibody of this application contains a light chain variable region polypeptide containing three LCDR sequences, characterized in that: the light chain variable region LCDR1 sequence is RASQNVHNRLT; the light chain variable region LCDR2 sequence is GASNLES; and the light chain variable region LCDR3 sequence is QQYNGSPTT. Furthermore, the LCDRs are defined according to Chothia's definition. Preferably, the anti-IL-17 monoclonal antibody of this application comprises a light chain variable region polypeptide having an amino acid sequence selected from SEQ ID NO:21.

In a more preferred embodiment, the heavy chain variable region sequence of the anti-IL-17 monoclonal antibody of this application is SEQ ID NO: 24, and the light chain variable region sequence is SEQ ID NO: 4; or the heavy chain variable region sequence is SEQ ID NO: 25, and the light chain variable region sequence is SEQ ID NO: 4; or the heavy chain variable region sequence is SEQ ID NO: 7, and the light chain variable region sequence is SEQ ID NO: 4.

The anti-IL-17 monoclonal antibody of this application may comprise or consist of a complete antibody (i.e., full-length), a substantially complete antibody, or its antigen-binding portion, such as a Fab fragment, an F(ab')2 fragment, or a single-chain Fv fragment.

The anti-IL-17 monoclonal antibody of this application includes a variable region and a constant region, wherein the antibody heavy chain constant region may be IgG1 subtype (SEQ ID NO:8) or IgG4 subtype (SEQ ID NO:9), and the light chain constant region may be kappa subtype (SEQ ID NO:10) or Lambda subtype (SEQ ID NO:11).

This application also provides isolated nucleic acid molecules encoding mouse IL-17 (SEQ ID NO:1), macaque IL-17 (SEQ ID NO:2), or cynomolgus monkey IL-17 (SEQ ID NO:3); IL-17 proteins encoded by mouse, macaque, or cynomolgus monkey nucleic acids (SEQ ID NO:1-3, respectively); vectors containing said nucleic acid molecules; host cells containing said vectors; and methods for producing mouse IL-17, macaque IL-17, or cynomolgus monkey IL-17.

"IL-17 binding molecule" refers to any molecule capable of binding to the human IL-17 antigen, alone or in combination with other molecules. The binding reaction can be visualized by standard methods (qualitative analysis) including, for example, binding assays, competitive assays, or bioassays to determine inhibition of IL-17 binding to its receptor, or any type of binding assay, using a negative control test with an antibody of the same isotype but with unrelated specificity (e.g., anti-CD25 antibody) as a reference. Non-limiting examples of IL-17 binding molecules include small molecules, IL-17 receptor decoys and antibodies and chimeric antibodies produced by B cells or hybridomas, antibodies transplanted with CDRs or human antibodies or any fragment thereof (e.g., F(ab')2 and Fab fragments), and single-chain or single-domain antibodies. Preferably, the IL-17 binding molecule antagonizes (e.g., reduces, inhibits, diminishes, blocks, delays) IL-17 function, expression, and/or signal transduction. IL-17 binding molecules are employed in some embodiments of the disclosed methods, protocols, kits, procedures, uses, and compositions.

"IL-17 receptor-binding molecule" refers to any molecule capable of binding to the human IL-17 receptor, alone or in combination with other molecules. The binding reaction can be visualized by standard methods (qualitative analysis), including, for example, binding assays, competitive assays, or bioassays to determine inhibition of IL-17 receptor binding to IL-17, or any type of binding assay, using a negative control test with an antibody of the same isotype but with unrelated specificity (e.g., anti-CD25 antibody) as a reference. Non-limiting examples of IL-17 receptor-binding molecules include small molecules, IL-17 receptor decoys and antibodies and chimeric antibodies against the IL-17 receptor generated by B cells or hybridomas, antibodies transplanted from CDRs or human antibodies or any fragment thereof (e.g., F(ab') ₂ and Fab fragments), and single-chain or single-domain antibodies. Preferably, the IL-17 receptor-binding molecule antagonizes (e.g., reduces, inhibits, diminishes, blocks, delays) IL-17 function, expression, and/or signal transduction. In some embodiments of the disclosed methods, protocols, kits, procedures, uses, and compositions, the IL-17 receptor-binding molecule is employed.

As used herein, the term "antibody" includes whole antibodies and any antigen-binding portion or single chain thereof. Natural antibodies are glycoproteins comprising at least two heavy (H) chains and two light (L) chains linked by disulfide bonds. Each heavy chain contains a heavy chain variable region (VH) and a heavy chain constant region. The heavy chain constant region contains three domains: CH1, CH2, and CH2. Each light chain contains a light chain variable region (VL) and a light chain constant region. The light chain constant region contains one domain, CL. The VH and VL regions can be further subdivided into highly variable regions (called complementarity-determining regions (CDRs)) and more conserved regions (called framework regions (FRs)), interspersed. Each VH and VL consists of three CDRs and four FRs, arranged in the following order from the amino terminus to the carboxyl terminus: FR1, CDR1, FR2, CDR2, FR3, CDR3, FR4. The variable regions of the heavy and light chains contain binding domains that interact with the antigen. The constant region of an antibody can mediate the binding of immunoglobulins to host tissues or factors, including various cells of the immune system (e.g., effector cells) and the first component (Clq) of the classical complement system. In some embodiments of the disclosed methods, protocols, kits, procedures, uses, and compositions, antibodies targeting IL-17 or the IL-17 receptor are employed.

As used herein, the term "antigen-binding moiety" of an antibody refers to an antibody fragment that retains the ability to specifically bind to an antigen (e.g., IL-17). It has been shown that the antigen-binding function of an antibody can be achieved by fragments of a full-length antibody. Examples of binding fragments covered by the term "antigen-binding moiety" of an antibody include Fab fragments, i.e., monovalent fragments consisting of VL, VH, CL, and CH1 domains; F(ab') ₂ fragments, i.e., bivalent fragments containing two Fab fragments linked by disulfide bridging in the hinge region; Fd fragments consisting of VH and CH1 domains; Fv fragments consisting of VL and VH domains of a single antibody arm; dAb fragments (Ward et al., (1989) Nature 341:544-546), which consist of a VH domain; and separated complementarity-determining regions (CDRs). Furthermore, although the two domains (VL and VH) of the Fv fragment are encoded by separate genes, these two domains can be combined using recombination methods by synthesizing linkers, enabling them to form a single protein chain, where the VL and VH regions pair to form a monovalent molecule (referred to as a single-chain Fv (scFv); see, for example, Bird et al., 1988 Science 242:423-426; and Huston et al., 1988 Proc. Natl. Acad. Sci. 85:5879-5883). The single-chain antibody is also intended to be included within the term "antibody." The single-chain antibody and antigen-binding moiety are obtained using conventional techniques known to those skilled in the art. In some embodiments of the disclosed methods, protocols, kits, procedures, uses, and compositions, the antigen-binding moiety of a single-chain antibody or an antibody against IL-17 (e.g., celecoxib) or the IL-17 receptor is employed.

The term "pharmaceutical acceptable" refers to a non-toxic material that does not interfere with the bioactivity and effectiveness of the active ingredient.

As used herein, "isolated antibody" refers to an antibody that is substantially free of other antibodies with different antigen specificities (e.g., an isolated antibody that specifically binds to IL-17 is substantially free of antibodies that specifically bind to antigens other than IL-17). The isolated antibody may be substantially free of other cellular material and/or chemicals. However, isolated antibodies that "specifically bind" to IL-17 may cross-react with other antigens (e.g., IL-17 molecules from other species). In some embodiments of the disclosed methods, protocols, kits, procedures, uses, and compositions, the IL-17 antagonist is the isolated antibody.

As used herein, the term "monoclonal antibody" or "monoclonal antibody composition" refers to an antibody molecular formulation having a single molecular composition. A monoclonal antibody composition exhibits single binding specificity and affinity for a specific epitope. In some embodiments of the disclosed methods, protocols, kits, procedures, uses, and compositions, the IL-17 antagonist is a monoclonal antibody.

The term "IL-17" refers to IL-17A and includes wild-type IL-17A from different species (e.g., humans, mice, and monkeys), polymorphs of IL-17A, and functional equivalents of IL-17A. The term "KD" refers to the dissociation constant, derived from the ratio of Kd to Ka (i.e., Kd/Ka), and is expressed as molar concentration (M). The KD value of an antibody can be determined using methods well-established in the art. Methods for determining the KD of an antibody may employ surface plasmon resonance or a biosensor system (e.g., a system). In some embodiments of this application, an IL-17 antagonist (e.g., an IL-17 binding molecule (e.g., an IL-17 antibody or its antigen-binding fragment, such as celecoxib) or an IL-17 receptor binding molecule (e.g., an IL-17 antibody or its antigen-binding fragment) binds human IL-17 with a KD of about 100 pM to 250 pM.

As used herein, the term "affinity" refers to the strength of the interaction between an antibody and an antigen at a single antigenic site. Within each antigenic site, the variable region of the antibody "arm" interacts with the antigen at multiple sites through weak non-covalent forces; the greater the interaction, the stronger the affinity. Standard analytical methods known in the art for assessing the affinity of antibodies for IL-17 of different species include, for example, ELISA, Western blot, and RIA. The binding kinetics (e.g., binding affinity) of antibodies can also be assessed by standard analytical methods known in the art (e.g., by Biacore assays). The embodiments further describe in detail analytical methods for assessing the effectiveness of antibodies in assessing the functional properties of IL-17 (e.g., receptor binding, prevention or improvement of bone resorption).

The terms “individual” and “patient” as used in this article include any human or non-human animal. The term “non-human animal” includes all vertebrates, such as mammals and non-mammals, such as non-human primates, sheep, dogs, cats, horses, cattle, chickens, amphibians, reptiles, etc.

It should be understood that an antibody that “inhibits” one or more IL-17 functional properties (e.g., biochemical, immunochemical, cellular, physiological, or other biological activities, or the like) as determined by methods known in the art and described herein involves a statistically significant reduction relative to a specific activity observed in the absence of the antibody (or in the presence of a control antibody with unrelated specificity). An antibody that inhibits IL-17 activity statistically significantly reduces the measured parameter, for example, by at least 10%, at least 50%, 80%, or 90%, and in some embodiments, the antibody of this application can inhibit more than 95%, 98%, or 99% of IL-17 functional activity.

"Amino acid" refers to, for example, all naturally occurring L-α-amino acids, including D-amino acids. Amino acids are identified by their well-known single-letter or three-letter names.

The term "amino acid sequence variant" refers to a molecule whose amino acid sequence differs from that of the present application. For example, an amino acid sequence variant of the present application's polypeptide with a specific sequence may still bind to human IL-17 or, for example, inhibit IL-17-induced IL-6 production in human dermal fibroblasts. A substitution variant is a variant in which at least one amino acid residue is removed from, for example, the present application's polypeptide with a specific sequence, and a different amino acid is inserted at the same position. These substitutions may be single substitutions, where only one amino acid in the molecule is substituted, or multiple substitutions, where two or more amino acids in the same molecule are substituted. An insertion variant is a variant in which one or more amino acids are inserted into, for example, the present application's polypeptide with a specific sequence, at an amino acid immediately adjacent to a specific position. An adjacent amino acid means linked to the α-carboxyl or α-amino functional group of the amino acid. A deletion variant is a variant in which one or more amino acids are removed from, for example, the present application's polypeptide with a specific sequence. Deletion variants generally lack one or two amino acids in a specific region of the molecule.

As used herein, “therapeutic effective amount” refers to the amount of an IL-17 antagonist (e.g., an IL-17 binding molecule (e.g., an IL-17 antibody, such as GR1501) or an IL-17 receptor binding molecule (e.g., an IL-17 receptor antibody) that, when administered in single or multiple doses to an individual (such as a human patient), effectively treats, prevents, cures, delays the onset or recurrence of the disease, reduces its severity, improves at least one symptom, or prolongs the individual’s survival beyond what would have been expected in the absence of such treatment. When applied to a single active ingredient (e.g., an IL-17 antagonist, such as an IL-17 binding molecule (e.g., an IL-17 antibody or its antigen-binding fragment)) administered alone, the term refers to the single ingredient. When applied to a combination, the term refers to the combined amount of active ingredients that produce a therapeutic effect (whether administered sequentially or in combination).

The term "treatment" refers to preventative or therapeutic treatment, as well as curative or disease-modifying treatment, including treating patients who are at risk of contracting an infectious disease or suspected of having one, and patients who are ill or have been diagnosed with an infectious disease or medical condition, and includes suppressing clinical relapse. It can treat individuals with or potentially developing a medical condition to prevent, cure, or cure the condition or its recurrence, delay its onset, reduce its severity, or improve one or more symptoms, or extend the individual's survival beyond what would have been expected without such treatment.

As used in this article, "C-reactive protein" and "CRP" refer to serum C-reactive protein, a plasma protein commonly used to indicate the acute phase response to inflammation. Plasma CRP levels can be given at any concentration, such as mg/dL or nmol/L. CRP levels can be measured using various well-known methods, including radioimmunodiffusion, electroimmunoassay, immunoturbidimetry, ELISA, turbidimetry, fluorescence polarization immunoassay, and laser turbidimetry. CRP can be tested using a standard CRP test or a high-sensitivity CRP (hs-CRP) test (i.e., a high-sensitivity test capable of measuring low CRP levels in a sample using laser turbidimetry). Kits for detecting CRP levels are available from various companies, such as Calbiotech, Cayman Chemical, Roche Diagnostics, Abazyme, DADE Behring, Abnova, Aniara, Bio-Quant, and Siemens Healthcare Diagnostics.

As used in this article, “erythrocyte sedimentation rate,” “ESR,” and “sedimentation rate” (“sedimentation rate” and “sedrate”) refer to the sedimentation rate of red blood cells in a patient sample (e.g., a plasma sample). ESR reflects plasma viscosity and the presence of acute-phase proteins and is usually reported in “mm/hr”. ESR is determined by measuring the distance that red blood cells settle over time in a tube. Typical ESR testing methods utilize the Westergren test, the Zeta sedimentation rate (ZSR) test, and the Wintrobe test. (See Moseley and Bull (1982) Clin. Lab Haematol. 4:169-78; Miller et al., (1983) Br Med J (Clin Res Ed) 286 (6361):266; Wetteland P et al. (1996) J. Intern. Med. 240 (3):125-310, all of which are incorporated herein by reference in their entirety.) Commercially available kits for measuring ESR are available from companies such as ARKRAY USA, BD Diagnostic Systems, and Polymedco. ESR instruments are available from companies such as US Patent 6,974,701, and include Steellex Scientific, Nicesound Electronics, Globe Scientific, Alifax, Analysis Instrument AB, Streck Laboratories, PolyMed, and Quantimetrix.

GR1501: This refers to the recombinant fully human anti-IL-17A monoclonal antibody produced by Chongqing Zhixiang Jintai Biopharmaceutical Co., Ltd., also known as celecoxib or celecoxib monoclonal antibody injection. For specific molecular details, please refer to Chinese patent application CN201510097117.0.

Example 1: Phase II clinical trial of GR1501 for the treatment of axial spondyloarthritis (AxSpA).

Example 1.1: Design of Participants in Phase II Clinical Trials

As shown in Figure 1, the study population consisted of a representative group of 160 patients aged 18 to 60 years (male or non-pregnant, non-lactating women) who met the International Spondyloarthritis Evaluation Task Force (ASAS) criteria for axial spondyloarthritis (axSpA) and had previously failed or were intolerant to one or more nonsteroidal anti-inflammatory drugs (NSAIDs).

Example 1.2: Study drug, dosage, and administration method in Phase II clinical trials (week 16)

1.2.1 The protocol for the Phase II clinical trial is shown in Figure 1, and detailed information is as follows:

Drug Name: GR1501 Injection / GR1501 Injection Placebo;

Specifications: 100mg/1mL/vial; or placebo 0mg/1mL/vial;

Dosage: 100mg, 200mg, 300mg;

Administration method: subcutaneous injection;

Dosing frequency: For the first 4 weeks, administer the drug every 2 weeks (W0, W2, W4), and then every 4 weeks thereafter (W8, W12).

Batch number: 20100101; 20191224 (placebo);

Expiry date: January 12, 2022; December 23, 2021 (placebo);

Provided by: Chongqing Zhixiang Jintai Biopharmaceutical Co., Ltd. / Zhixiang (Shanghai) Pharmaceutical Technology Co., Ltd.

1.2.2 Basis for Dosage and Frequency Selection

The GR1501-001 dose escalation phase trial and the GR1501-003 trial conducted on this drug showed that it was well tolerated and safe within the range of single-dose doses of 10 mg to 300 mg and multiple-dose doses of 60 mg to 300 mg, and no events meeting the trial termination criteria occurred.

Pharmacokinetic studies showed that after a single subcutaneous injection of GR1501 solution ranging from 10 mg to 300 mg, the dose was linear with exposure. See Figures 2 and 3 for details.

After multiple subcutaneous injections, the blood drug concentrations in the 60mg, 100mg, 200mg, and 300mg groups did not reach steady state during 8-12 weeks. The blood drug concentration in the 150mg group may have reached steady state during 8-12 weeks, but due to the small sample size, the confidence interval may be large.

Preliminary results from the completed dose-escalation phase of the "GR1501-001" clinical trial, combined with PK and PD models, indicate that a 200mg dose is expected to achieve satisfactory PASI75 efficacy in patients with plaque psoriasis. Given that this drug is formulated in 100mg/1ml strengths, and considering that this dose-exploration trial will focus on 200mg, this trial plans to use dosages of 100mg, 200mg, and 300mg.

In this trial, the dosing frequency was set at Q4W. In addition, the dosing frequency in the first 4 weeks (0-4 weeks) of this trial was Q2W, in order to increase the drug concentration of GR1501 injection in a short period of time and control the disease activity level of patients with active axial spondyloarthritis as early as possible.

Example 1.3: Statistical Analysis of Phase II Clinical Trial (Week 16)

The primary efficacy endpoints were based on FAS and PPS analyses. Missing data were imputed using "non-respondent". The proportion of patients achieving ASAS20 at week 16 was compared between groups using the chi-square method or Fisher's exact probability method. Two-sided 95% confidence intervals for individual rates were calculated using the Wilson method, and two-sided 95% confidence intervals for differences between rates were calculated using the Wald method. Logistic regression models were used to analyze EDC data-related influencing factors (previous history of biologics treatment for this disease, disease severity, and axial spondyloarthritis classification) and calculate the corrected rate difference. The standard error of the corrected rate difference was estimated using 1000 samplings with the Bootstrap method, thus obtaining the two-sided 95% CI of the corrected rate difference. Stratified analysis was performed on the proportion of patients achieving ASAS20 at week 16 based on EDC data, prior history of biologics treatment for this disease, disease severity, and axial spondyloarthritis classification, and forest plots were generated.

Full analysis set (FAS): The set of all cases that were randomized and received the investigational drug at least once.

The Protocol Compliance Set (PPS) is a dataset comprised of participants who have fully complied with the trial protocol. Compliance includes the treatment received, the availability of the primary endpoint, and no major deviations from the protocol. Cases of participants who withdrew due to ineffectiveness are also included in the PPS.

Example 1.4, Phase II Clinical Results

1.4.1 Evaluation Indicators:

(1) Key endpoint indicators

The proportion of patients who achieved ASAS20 at week 16.

(2) Analysis of key efficacy indicators:

Based on FAS and PPS analysis, missing data were imputed using "unresponding". The proportion of patients achieving ASAS20 at week 16 was compared between groups using the chi-square method or Fisher's exact probability method. Two-sided 95% confidence intervals for single-group rates were calculated using the Wilson method, and two-sided 95% confidence intervals for inter-group rate differences were calculated using the Wald method. Logistic regression models were used to analyze EDC data-related influencing factors (history of previous biologic therapy for this disease, disease severity, and axial spondyloarthritis classification) and calculate the corrected rate difference. The standard error of the corrected rate difference was estimated using 1000 samples with the Bootstrap method, thus obtaining the two-sided 95% CI of the corrected rate difference.

Subgroup analysis: Based on EDC data, the proportion of patients achieving ASAS20 at week 16 of treatment was stratified according to their prior history of biologic therapy for this disease, disease severity, and axial spondyloarthritis classification, and forest plots were generated. The FAS (filled-in) analysis shown in Figure 4 yielded the following results: the proportions of patients achieving ASAS20 at week 16 after treatment were 52.5% in the placebo group (N=40), 77.5% in the 100mg group (N=40), 75.0% in the 200mg group (N=40), and 72.5% in the 300mg group (N=40), respectively. Statistically significant differences were found between the 100mg and placebo groups, and between the 200mg and placebo groups (P<0.05). No statistically significant differences were found between the 300mg and placebo groups, and between the 200mg and 100mg groups, and between the 300mg and 200mg groups (P>0.05).

As shown in Figure 5, PPS analysis revealed that the proportions of patients achieving ASAS20 at week 16 after medication were 53.8%, 81.6%, 78.4%, and 80.6% in the placebo group (N=39), 100mg group (N=38), 200mg group (N=37), and 300mg group (N=36), respectively. Statistically significant differences were found between the 100mg and placebo groups, the 200mg and placebo groups, and the 300mg and placebo groups (P<0.05). No statistically significant differences were found between the 200mg and 100mg groups, the 300mg and 100mg groups, and the 300mg and 200mg groups (P>0.05).

(3) Security data results

A total of 160 subjects were included in the SS study.

Treatment-associated adverse events (TEAEs): 31 cases in the placebo group (N=40), with an incidence rate of 77.5%; 35 cases in the 100 mg group (N=40), with an incidence rate of 87.5%; 31 cases in the 200 mg group (N=40), with an incidence rate of 77.5%; and 36 cases in the 300 mg group (N=40), with an incidence rate of 90.0%.

Adverse reactions (ADRs): 21 cases were observed in the placebo group (N=40), with an incidence rate of 52.5%; 24 cases were observed in the 100mg group (N=40), with an incidence rate of 60.0%; 24 cases were observed in the 200mg group (N=40), with an incidence rate of 60.0%; and 29 cases were observed in the 300mg group (N=40), with an incidence rate of 72.5%.

In terms of safety, the investigational drug showed good overall safety and tolerability. However, the overall incidence of adverse events/reactions was higher in the 300mg group than in the 100mg and 200mg groups.

In summary, under the existing conditions, GR1501 injection at therapeutic doses of 100 mg and 200 mg has proven to be safe and effective in patients with active axial spondyloarthritis.

Example 2: Phase III clinical trial of GR1501 for the treatment of axial spondyloarthritis (AxSpA).

Example 2.1, Participant Design

As shown in Figure 6, the study population consisted of a representative group of 465 patients aged 18 years and older (male or non-pregnant, non-lactating women) who met the diagnostic criteria for axial spondyloarthritis (axSpA) of the International Task Force on Evaluation of Spondyloarthritis (ASAS) and whose imaging changes of sacroiliitis met the New York modified criteria: bilateral sacroiliitis grade ≥2 or unilateral sacroiliitis grade 3-4; poor response to previous treatment with nonsteroidal anti-inflammatory drugs (NSAIDs); or contraindications to or intolerance to NSAIDs.

Example 2.2: Study drug, dosage, and administration method in Phase III clinical trials.

The dosing regimen for phase III clinical trials is as follows:

Example 2.3: Statistical Analysis of Phase III Clinical Trials

After all participants completed the efficacy and safety assessment at week 32, the study was unblinded for treatment-period data analysis.

General considerations:

SAS 9.4 software was used for analysis. All statistical tests were two-tailed, and a p-value less than 0.05 was considered statistically significant. Continuous variables were described using mean, standard deviation, four-digit values, minimum, and maximum values, while count and ordinal data were described using frequency and percentage.

Drug exposure and concomitant medications, efficacy, safety, immunogenicity, other indicators, and pharmacokinetic analysis results are presented separately for the 0-16 week and 0-32 week periods. The 0-16 week analysis is divided into placebo, 100 mg, and 200 mg groups; the safety analysis will also show the total results for all experimental groups. The 0-32 week analysis is based on subjects who have used GR1501 at least once. For the placebo group, only data after the first use of GR1501 are considered, divided into 100 mg, 200 mg, placebo → 100 mg (16-32 week), and placebo → 200 mg (16-32 week). Safety analysis also included the combined analysis of the 100 mg group and the placebo → 100 mg group (16-32 weeks), the 200 mg group and the placebo → 200 mg group (16-32 weeks), and the analysis of the GR1501 group (i.e., those who had used GR1501 at least once), and the 100 mg (16-32 weeks) and 200 mg (16-32 weeks) groups (i.e., those who had been given at least once during the maintenance therapy period in the trial).

Example 2.4, Phase III Clinical Results

2.4.1 Evaluation Indicators:

(1) Key endpoint indicators

The proportion of patients who achieved ASAS20 at week 16.

(2) Analysis of key efficacy indicators:

At week 16, both the 100mg and 200mg groups met their primary endpoints and showed significant improvement compared to the placebo group. Durational analysis showed that the 200mg group had a data advantage over the 100mg group.

1) Concurrent events

The percentage of subjects who experienced at least one comorbid event during the 0-16 week period was: 14 (9.0%) in the placebo group, 5 (3.2%) in the 100 mg group, and 8 (5.2%) in the 200 mg group.

The percentage of subjects who experienced at least one comorbid event during the 16-32 week period was: 2 (1.3%) in the 100 mg group, 8 (5.2%) in the 200 mg group, 0 in the placebo → 100 mg group, and 2 (2.8%) in the placebo → 200 mg group.

2) Main target estimation analysis:

Master estimation method: At week 16, the ASAS20 response rates in the 100 mg and 200 mg dose groups were significantly higher than those in the placebo group (65.8% vs 35.9%, P < 0.001; 74.0% vs 35.9%, P < 0.001). Sensitivity analyses were consistent with the conclusions of the master estimation method.

ASAS20 response rate in EDC subjects who had previously received biologic therapy for this disease ("non-responder" filled in):

The ASAS20 response rate was 35.9% in the placebo group (N=64), 71.4% in the 100mg group (N=63) (rate difference 35.5%), and 83.3% in the 200mg group (N=60) (rate difference 47.4%). For subjects who had not previously received biologics for this disease, the ASAS20 response rate (filled in "non-respondent") was 35.9% in the placebo group (N=92), 62.0% in the 100mg group (N=92) (rate difference 26.1%), and 68.1% in the 200mg group (N=94) (rate difference 32.2%).

ASAS20 response rates ("non-response" filled) for subjects with EDC weight ≥70kg: 41.4% in placebo group (N=70), 66.7% in 100mg group (N=72) (rate difference 25.2%), and 80.3% in 200mg group (N=71) (rate difference 38.9%).

3) Secondary estimation target analysis

At week 16, the response rates of ASAS40 in the 100 mg and 200 mg dose groups were significantly higher than those in the placebo group (40.0% vs 18.6%, P < 0.001; 41.6% vs 18.6%, P < 0.001) (missing data were filled with non-responders, and differences between groups were compared using the χ² test or Fisher's exact test).

At week 16, the response rates of ASAS 5/6 in the 100 mg and 200 mg dose groups were significantly higher than those in the placebo group (49.0% vs 15.0%, P < 0.001; 55.2% vs 15.4%, P < 0.001) (missing data were filled with non-responders, and differences between groups were compared using the χ² test or Fisher's exact test).

(3) Security Analysis Results

No new risk signals were observed in the 100 mg and 200 mg groups in patients with radiographically positive axial spondyloarthritis, indicating a good safety profile. Common adverse events included various infectious diseases, most of which were mild to moderate and resolved or improved.

(4) Pharmacokinetic results

Based on the population pharmacokinetic model simulation results over 48 weeks, the half-life of celecoxib injection is approximately 28.3 days. In the Phase III clinical trial of radiologically positive axial spondyloarthritis, under the clinically recommended dosing regimen, the peak time of celecoxib injection was 4 days, and steady state was reached after approximately 20 weeks of multiple dosing.

(5) Conclusion

After 16 weeks of treatment with celecoxib injection, both the 100mg and 200mg groups showed better efficacy than the placebo group. Administered once every two weeks for the first four weeks, it has a rapid onset of action, quickly relieving pain, reducing spinal stiffness, improving physical function, and decreasing disease activity.

Subsequent administration every 4 weeks resulted in sustained improvement in symptoms and signs by week 16, with significant improvements in disease activity, inflammatory markers, and quality of life. Stratified analysis indicated that the 200mg group was significantly superior to the 100mg group in individuals weighing ≥70kg. Long-term administration of celecoxib injection demonstrated continuously improving efficacy, showing potential advantages in controlling clinical symptoms, alleviating disease progression, improving physical function, and enhancing quality of life.

In summary, celecoxib injection 200 mg (administered at weeks 0, 2, and 4, followed by maintenance doses of 200 mg every 4 weeks) demonstrates clear efficacy and good safety in treating patients with radiographically positive axial spondyloarthritis. Since studies have shown that the aqueous version of celecoxib (GR1501) at 100 mg has the best stability, a twice-dose regimen was designed with a corresponding container.

Example 3: Determination of the formulation of the first round of aqueous drug products.

3.1 Prescriptions to be screened

Table 1-1 Formulations of Samples from the First Round of Formulation Screening

Note: √: indicates that the prescription contains the corresponding excipients;
- indicates that the prescription does not contain the corresponding excipients.

The formulation instructions are as follows:

(1) A1 is a histidine buffer system containing trehalose and polysorbate 80, pH 5.8.

(2) The pH of A2 is 5.5, the pH of B2 is 6.0, and other conditions are the same as A1. Compare A2 and B2 with A1 to examine the effect of pH.

(3) A3 does not contain L-methionine, and other conditions are the same as A1. A3 is compared with A1 to examine the effect of L-methionine.

(4) Formulations A3 does not contain L-methionine, Formulation B3 does not contain L-methionine and the osmotic pressure regulator is changed to mannitol, and Formulation B4 does not contain L-methionine and the osmotic pressure regulator is changed to sodium chloride. Formulations B3 and B4 are compared with Formulation A3 to examine the effects of different osmotic pressure regulators.

(5) Buffer salts A4 and B1 are citric acid/sodium citrate, without L-methionine, and the osmotic pressure regulators are trehalose and sodium chloride, respectively. Compare A4 with A3 to examine the effect of citrate buffering.

The L-histidine/histidine hydrochloride is L-histidine and histidine hydrochloride monohydrate; the trehalose is trehalose dihydrate; and the surfactant polysorbate 80, also known as Tween 80, is used as a stabilizer in pharmaceuticals.

The excipients and their concentrations are the same in this embodiment as in the formulation with the same number in Example 4 below.

3.2 Screening Method

Initial Screening: Formulations from the first round of formulation screening were first screened and eliminated using DSF (Differential Scanning Fluorescence) assays to remove those with low Tm values. The remaining formulations were then subjected to accelerated stability testing. DSF is a method used to evaluate the effect of formulation components on protein conformational stability. It records the fluorescence emission wavelength signal as the solution temperature gradually increases (from 25°C to 95°C), monitoring the temperature Tm (Temperature Middle) at which the fluorescence signal jumps. DSF uses SYPRO Orange dye, which binds to the hydrophobic surface of proteins through hydrophobic interactions. As the temperature increases, the higher-order structure of the protein molecule is disrupted, exposing its internal hydrophobic groups. SYPRO Orange binds to these hydrophobic groups and emits a fluorescence signal. The greater the degree of conformational disruption, the stronger the fluorescence signal. A higher Tm value indicates that the protein is more difficult to unfold, indicating better conformational stability. The remaining results after the initial screening were then tested as follows:

(1) HIC purity (chromatographic column: Thermo MabPac™ HIC-10; high performance liquid chromatograph: Thermo Ultimate 3000): The HIC method can separate the oxidized variants and polymers of GR1501 from the main protein peak. HIC purity is greatly affected by the degree of protein oxidation. By comparing HIC purity, the protein oxidation situation under the formulation conditions can be reflected to a certain extent.

(2) SEC-HPLC purity (chromatographic column: TSKgel G3000SWXL; high performance liquid chromatograph: Thermo Ultimate 3000): mainly examines the proportion of soluble polymers, which reflects the aggregation of proteins.

(3) rCE-SDS purity and NrCE-SDS purity (capillary: Micro solv; capillary electrophoresis apparatus: ABSciex PA800 plus): mainly reflect the degree of protein hydrolysis and fragmentation.

(4) CEX-HPLC purity (chromatographic column: Thermo ProPac™ WCX-10 BioLC™ Analytical; high performance liquid chromatograph: Thermo Ultimate 3000): mainly reflects the degree to which amino acid residues of a protein are modified to produce various charge variants under stable conditions.

3.3 Screening Results

(1) DSF test

The results of the first round of formulation screening DSF trials are shown in Table 1-2.

Table 1-2 Results of the first round of formulation screening DSF test

A2 and B4 had low Tm values, so these two formulations were excluded first, and other formulations were used for the next step of stability testing.

The formulation of B4 is basically the same as that of A3 (the osmolarity regulator is replaced by sodium chloride instead of trehalose), but the Tm values are different. It is speculated that the conformational stability of GR1501 protein is relatively poor in formulations with sodium chloride as the osmolarity regulator compared to trehalose.

(2) HIC purity

The stability and purity results of the first round of formulation screening are shown in Table 1-3 and Figure 7-1.

Table 1-3 Results of Stability-HIC Purity in the First Round of Formulation Screening

Results: Under accelerated high-temperature (37℃±2℃) conditions, there was no significant difference in HIC purity among the formulations during weeks 0–4. During weeks 4–8, with increased light exposure (5000 lx), under the combined effects of high temperature and light: formulations A1 and B2, containing the antioxidant L-methionine, exhibited the highest HIC purity at week 8; formulations A3 and B3, lacking L-methionine but using histidine as a buffer, showed the second highest HIC purity at week 8; formulations A4 and B1 exhibited the lowest HIC purity at week 8, with citric acid/sodium citrate as their buffer and lacking L-methionine.

The HIC results show that the L-methionine and histidine buffer system has a good photoprotective effect on GR1501 protein. Among the formulations A1 and B2, GR1501 protein has the best stability, while the A1 formulation has the best effect due to differences in pH and other factors.

(3) SEC-HPLC purity

The stability and purity results of the first round of formulation screening by SEC-HPLC are shown in Table 1-4 and Figure 7-2.

Table 1-4 Results of Stability-SEC Monomer Purity in the First Round of Formulation Screening

Results: Under accelerated conditions (37℃±2℃), there was no significant difference in SEC purity among the formulations during weeks 0–4. During weeks 4–8, increased light exposure (5000 lx) caused protein aggregation in the GR1501 sample under the combined effects of high temperature and light. By week 8, the SEC purity of the formulations was, in descending order: A1 > B2 > A3 > B3 > A4 > B1.

The SEC-HPLC results show that the GR1501 protein in the A1 formulation has the best stability.

In summary, a comparison of the protein stability of formulations A2, B2, and A1 shows that the GR1501 protein formulation exhibits the best conformational stability at a pH of 5.8.

A comparison of the protein stability of formulations B1, B4 and A1 shows that when sodium chloride is used as the osmotic pressure regulator in the GR1501 protein formulation, the conformational stability deteriorates.

A comparison of the protein stability of formulations A3, A4, B3, B4 and A1 shows that L-methionine does not affect the stability of GR1501 under high temperature conditions, but L-methionine inhibits photodegradation.

A comparison of the protein stability of formulations A4, B1, and A1 shows that the histidine buffer system can inhibit degradation caused by light and oxidation to a certain extent, while citrate does not have this effect.

Through comprehensive comparison and analysis of multiple formulations, the type of sugar, the type of buffer salt, and the presence or absence of L-methionine have little impact on the stability of GR1501 protein under high-temperature conditions. However, under light conditions, the purity of each formulation varies considerably, which is related to the antioxidant components contained in each formulation. Among them, the buffer system of formulation A1 is histidine (which has a certain antioxidant capacity), and it also contains a certain amount of the antioxidant L-methionine. Therefore, its degradation rate is the slowest under light conditions.

Therefore, formulation A1 (3.103 mg/ml L-histidine/histidine hydrochloride, 75.67 mg/ml trehalose, 0.2 mg/ml polysorbate 80, 0.746 mg/ml L-methionine, pH 5.8) is the optimal formulation for the first round of formulation screening.

Example 4: Screening of Formulations for the Second Round of Aqueous Drug Products

4.1 Prescriptions to be screened

The formulations of the samples selected for the second round of formulation stability screening are shown in Table 2-1. The second round of formulation screening mainly investigated the effects of trehalose and sucrose on stability, the stability of formulations at different concentrations, and the differences between histidine and acetate buffer systems.

Table 2-1 Formulation samples from the second round of formulation screening

√: Indicates that the prescription contains the corresponding excipient;
- indicates that the prescription does not contain the corresponding excipients.

3. The 103 mg/ml L-histidine/histidine hydrochloride specifically includes 1.241 mg/ml L-histidine and 2.516 mg/ml histidine hydrochloride monohydrate;

Trehalose selected is trehalose dihydrate.

4.1 Screening Scheme

The experimental protocol for the second round of formulation stability screening is shown in Table 2-2.

Table 2-2 Accelerated stability tests of formulation samples

4.3 Screening Experimental Results

The results of the second round of formulation screening for stability and HIC purity are shown in Table 2-3.

Table 2-3 Results of Accelerated Stability-HIC Purity in the Third Round of Formulation Screening

NA: Indicates untested

At a protein concentration of 100 mg/ml, under high temperature conditions, there was no significant difference in the purity changes of formulations A1, A2, A3, and A4; under light conditions, formulation A4, which does not contain antioxidants, was easily oxidized and degraded. The protein concentration measurements of formulations A2 and A3 were abnormal under both high temperature and light conditions.

At a protein concentration of 150 mg/ml, under high temperature conditions, formulations B2 and B3, which contain sucrose, exhibited poor stability; under light conditions, formulation B4, which lacks antioxidants, was easily oxidized and degraded. B1 decreased by 16.7% under light conditions, while A1 decreased by 8.9%. Therefore, A1 is significantly superior to B1.

This round of formulation screening showed that excessively high protein concentrations (150 mg/ml) are detrimental to the stability of GR1501 protein; meanwhile, histidine buffer system and trehalose osmotic regulator are more effective in maintaining the stability of GR1501 protein than acetate buffer system and sucrose as an osmotic regulator.

Based on the results of the comprehensive formulation stability study and the administration method, the optimal subcutaneous dose of GR1501 was selected as 200 mg per administration. To ensure the efficacy of GR1501, two doses of 100 mg each were administered per administration.

In the description of this specification, the terms "one embodiment," "some embodiments," "specific embodiment," etc., refer to a specific feature, structure, material, or characteristic described in connection with that embodiment or example, which is included in at least one embodiment or example of this application. In this specification, the illustrative expressions of the above terms do not necessarily refer to the same embodiment or example. Furthermore, the specific features, structures, materials, or characteristics described may be combined in any suitable manner in one or more embodiments or examples.

The above description is only a preferred embodiment of this application and is not intended to limit this application. Any modifications, equivalent substitutions, improvements, etc., made within the spirit and principles of this application should be included within the protection scope of this application.

Claims (24)

Use of an aqueous pharmaceutical product in the preparation of a medicament for treating axial spondyloarthritis, wherein the antibody aqueous pharmaceutical product comprises an antibody that specifically binds to IL-17A, the antibody comprising a VH having the following three CDRs: CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 12, CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 13, and CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 14; and The antibody comprises a VL having the following three CDRs: CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 15, CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 16, and CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 17; And the sequence of CDRs is defined according to Chothia. According to the use of claim 1, the antibody comprises a heavy chain and a light chain, the heavy chain comprising the amino acid sequence shown in SEQ ID NO: 18, and the light chain comprising the amino acid sequence shown in SEQ ID NO: 19. The use according to claim 1 or 2, wherein the drug is formulated to be contained in a container, each container having sufficient quantity of the aqueous drug product to allow delivery of the following unit doses: a) A total of 6 doses, administered in 2-dose regimens, each dose being a subcutaneous dose of approximately 100 mg to approximately 300 mg, delivered at weeks 0, 2, and 4; and b) Subsequently, weekly delivery of two doses is initiated after the subcutaneous delivery of the fourth week as described in step a), with each dose being approximately 100 mg to approximately 300 mg. According to the use of claim 3, the dosage of each administration in step a) and the dosage of each administration in step b) are about 100 mg, or about 200 mg, or about 300 mg. According to any one of claims 3 to 4, the dosage for each administration in step a) and step b) is about 200 mg. The use according to any one of claims 1 to 5, wherein the aqueous pharmaceutical product further comprises trehalose dihydrate, L-histidine, histidine hydrochloride monohydrate, L-methionine, and polysorbate 80. The use according to any one of claims 1 to 6, wherein the antibody content is about 100 mg/ml. According to any one of claims 6 or 7, the content of L-histidine is about 1.241 mg/ml, the content of histidine hydrochloride monohydrate is about 2.516 mg/ml, the content of trehalose dihydrate is about 75.67 mg/ml, the content of polysorbate 80 is about 0.2 mg/ml, and the content of L-methionine is about 0.746 mg/ml. The use according to any one of claims 1 to 8, wherein the pH of the aqueous pharmaceutical product is about 5.8. An aqueous pharmaceutical product comprising an anti-IL-17A antibody and a buffer solution, said antibody comprising a VH having the following three CDRs: CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 12, CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 13, and CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 14; and The antibody comprises a VL having the following three CDRs: CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 15, CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 16, and CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 17. The product of claim 10, wherein the antibody comprises a heavy chain and a light chain, the heavy chain comprising an amino acid sequence as shown in SEQ ID NO: 18, and the light chain comprising an amino acid sequence as shown in SEQ ID NO: 19. The product according to any one of claims 10 to 11 further includes an antioxidant. The product according to any one of claims 10 to 12 further includes an osmotic pressure regulator. The product according to any one of claims 10 to 13 further includes a stabilizer. The product according to any one of claims 12 to 14, wherein the antioxidant is L-methionine. The product according to any one of claims 10 to 15, wherein the buffer solution is histidine hydrochloride, preferably, the buffer solution is L-histidine and histidine hydrochloride monohydrate. The product according to any one of claims 13 to 16, wherein the osmotic pressure regulator is trehalose, preferably trehalose dihydrate. The product according to any one of claims 14 to 17, wherein the stabilizer is polysorbate 80. The product according to any one of claims 10 to 18, wherein the aqueous pharmaceutical product comprises about 100 mg/ml to about 150 mg/ml of anti-IL-17A antibody, about 70 mg/ml to about 80 mg/ml of trehalose dihydrate, about 1 mg/ml to about 2 mg/ml of L-histidine, about 2 mg/ml to about 4 mg/ml of histidine hydrochloride monohydrate, about 0.5 mg/ml to about 1 mg/ml of L-methionine, and about 0.1 mg/ml to about 0.3 mg/ml of polysorbate 80. The product according to claim 19, wherein the aqueous pharmaceutical product comprises about 100 mg/ml of anti-IL-17A antibody, about 75.67 mg/ml of trehalose dihydrate, about 1.241 mg/ml of L-histidine, about 2.516 mg/ml of histidine hydrochloride monohydrate, about 0.746 mg/ml of L-methionine and about 0.2 mg/ml of polysorbate 80. According to any one of claims 10 to 20, the pH of the aqueous pharmaceutical product is about 5.5 to about 6.0, preferably 5.8. Use of the product according to any one of claims 10 to 21 in the preparation of a medicament for treating axial spondyloarthritis. The product according to any one of claims 10 to 21 is used to treat axial spondyloarthritis. A method for treating axial spondyloarthritis, comprising administering to an individual in need a therapeutically effective amount of the product as described in any one of claims 10 to 21.