Formulations and uses of fusion proteins

Abstract

The present invention provides solid formulations consisting of particles comprising a fusion protein and at least one carrier, methods of making the solid formulations, and methods of treating allergic rhinitis in a subject, wherein the methods comprise administering to the subject a therapeutically effective amount of a solid formulation of a fusion protein.

Description

Technical Field

[0001] The present invention relates to the field of biomedicine, and in particular to a solid preparation of a fusion protein, a method for preparing the solid preparation, and a method for treating allergic rhinitis in an individual with the solid preparation. Background Art

[0002] The immune system is always dedicated to defending against parasites, fungi, viruses, and bacteria. However, sometimes the immune system treats harmless substances (called allergens) as unwanted invaders and tries to fight against them. This overreaction of the body's immune system to normally harmless substances is called an allergic reaction.

[0003] The number of bioactive macromolecules for treating allergic reactions is limited. Omalizumab is the only antibody approved by the FDA for targeting and blocking immunoglobulin E (IgE).

[0004] Studies have shown that by conjugating the FcεRI receptor and the FcγRII receptor in a fusion protein, the fusion protein can regulate the IgE-mediated signaling pathway, inhibit the release of active mediators, and thus can inhibit allergic reactions (WO2005085291A1).

[0005] However, due to the highly degradable nature of such fusion proteins, the preservation and storage of large proteins such as fusion proteins are of concern, which may be caused by chemical instability (e.g., any process involving modifying the protein by bond formation or cleavage to produce new chemical entities) or physical instability (e.g., changes in the higher-order structure of the protein). Chemical instability may result from racemization, hydrolysis, oxidation, or β-elimination, and physical instability may be due to denaturation, aggregation, precipitation, or adsorption. Commercially available protein preparations must be able to be administered safely and remain physically, chemically, and biologically stable during the recommended shelf life.

[0006] Allergic rhinitis is also known as "hay fever" and is a nasal inflammation that occurs when the immune system overreacts to allergens in the air.

[0007] The present invention surprisingly reveals that the solid preparation of the present application can be applied to mammals by nasal administration, preferably by inhalation, and can effectively inhibit or relieve the symptoms of allergic rhinitis. Summary of the Invention

[0008] One object of the present invention is to provide a solid preparation of a fusion protein that is stable during the recommended shelf life, preferably for up to one year at room temperature. Such a solid preparation comprises substantially spherical fusion protein microparticles dispersed in at least one carrier and can be applied by nasal administration, preferably by inhalation.

[0009] Another object of the present invention is to provide a method for preparing a solid preparation.

[0010] Another object of the present invention is to provide a method for treating allergic rhinitis with a solid preparation.

[0011] In one aspect, the present invention provides a solid preparation composed of particles containing a fusion protein and at least one carrier. Carriers suitable for the present invention include polyols, carbohydrates, and sugars. The carrier is selected from the group consisting of sucrose, trehalose, sorbitol, glycerol, mannitol, lactose, xylitol, arabitol, erythritol, lactitol, maltitol, glucose, raffinose, maltose, dextran, inositol, or a combination thereof.

[0012] In a preferred aspect, the particles have a fusion protein and two or more carriers, and preferably the two carriers are lactose and mannitol.

[0013] The fusion protein is a polypeptide comprising the amino acid sequence of SEQ ID No. 2 or an amino acid sequence containing one or more deletions, substitutions, additions, or insertions of amino acid residues in the amino acid sequence of SEQ ID No. 2.

[0014] In a preferred aspect, the fusion protein comprises one of the following sequences: 1) The DNA sequence of SEQ ID No. 1; 2) The amino acid sequence of SEQ ID No. 2; 3) A DNA sequence having at least 95% identity with SEQ ID No. 2 and encoding the same functional protein sequence.

[0015] In a preferred aspect, the DNA sequence of sequence 1 consists of 1665 base pairs; the reading frame is from the 1st base pair to the 1665th base pair starting from the 5' end.

[0016] Extensions in any fragment of the fusion protein are also covered within the scope of the fusion protein.

[0017] The fusion protein can be those disclosed in WO2005085291A1.

[0018] In a preferred aspect, the fusion protein comprises the amino acid sequence of SEQ ID No. 2 or has at least 60%, 70%, 80%, 90%, 95%, 96%, 97%, 98%, or 99% identity with SEQ ID No. 2.

[0019] In a preferred aspect, the fusion protein is FP4 having the amino acid sequence of SEQ ID No. 2.

[0020] It has surprisingly been found in the present invention that when the solid preparation is stored at room temperature for one year, its biological activity remains substantially unchanged. In addition, the solid preparation has a substantially spherical shape and a substantially uniform diameter, and is suitable for administration by inhalation.

[0021] In another aspect, the present invention provides a method for treating allergic rhinitis, the method comprising administering to an individual a therapeutically effective amount of a solid preparation comprising a fusion protein.

[0022] In another aspect, the present invention provides the use of a solid preparation comprising a fusion protein in the manufacture of a medicament for treating allergic rhinitis in an individual.

[0023] In a preferred aspect, the fusion protein comprises the amino acid sequence of SEQ ID No.2 or has at least 60%, 70%, 80%, 90%, 95%, 96%, 97%, 98% or 99% identity with SEQ ID No.2.

[0024] In a preferred aspect, the fusion protein is FP4 having the amino acid sequence of SEQ ID No.2.

[0025] In one aspect, the present invention relates to a solid preparation of a fusion protein, wherein the solid preparation consists of particles comprising a fusion protein and at least one carrier, and the fusion protein is a protein comprising the amino acid sequence of SEQ ID No.2 or having at least 60%, 70%, 80%, 90%, 95%, 96%, 97%, 98% or 99% identity with SEQ ID No.2.

[0026] In a preferred aspect, the present invention relates to a solid preparation, wherein the particles comprise a fusion protein and one or more carriers.

[0027] In a preferred aspect, the present invention relates to a solid preparation, wherein the carrier is selected from polyols, carbohydrates and sugars.

[0028] In a preferred aspect, the present invention relates to a solid preparation, wherein the carrier is selected from the group consisting of sucrose, trehalose, sorbitol, glycerol, mannitol, lactose, xylitol, arabitol, erythritol, lactitol, maltitol, glucose, raffinose, maltose, dextran, inositol or a combination thereof.

[0029] In a preferred aspect, the present invention relates to a solid preparation, wherein the particles have a 50% volume median diameter (D50) of 5 μm or less, preferably 4.5 μm or less, more preferably 3.5 to 4.5 μm, and a 90% volume median diameter (D90) of 35 μm or less, preferably 30 μm or less, more preferably 28 to 30 μm.

[0030] In another preferred aspect, the present invention relates to a solid preparation, wherein the particles have a 50% volume median diameter (D50) higher than 5 μm to 12 μm, preferably 8 to 12 μm, still preferably 8 to 10 μm, more preferably about 9 μm or about 10 μm, and most preferably about 10 μm, and a 90% volume median diameter (D90) of 40 μm or lower, preferably 38 μm or lower, and more preferably 34 to 36 μm.

[0031] In a preferred aspect, the present invention relates to a solid preparation, wherein the solid preparation is substantially free of surfactants and is preferably a pure preparation.

[0032] In a preferred aspect, the present invention relates to a solid preparation, wherein the solid preparation has a shelf life of 6 months or longer, preferably one year or longer.

[0033] In a preferred aspect, the present invention relates to a solid preparation, wherein the solid preparation is inhalable.

[0034] In a preferred aspect, the present invention relates to a solid preparation, wherein the solid preparation contains 0.1 - 5 wt%, preferably about 0.5 - 4 wt%, and more preferably 1 - 4 wt% of a fusion protein.

[0035] In another aspect, the present invention relates to a method for preparing the above solid preparation, the method comprising the steps of: spraying a suspension or solution of a fusion protein into a stream of liquid droplets, entraining the stream of liquid droplets within a coolant stream to freeze the droplets into frozen particles, and drying such particles to form the solid preparation.

[0036] In another aspect, the present invention relates to a method for treating allergic rhinitis in an individual, the method comprising administering to the individual a therapeutically effective amount of the above solid preparation.

[0037] In a preferred aspect, the present invention relates to a method for treating allergic rhinitis in an individual, wherein the solid preparation is administered by inhalation, preferably by means of a nasal spray device. BRIEF DESCRIPTION OF THE DRAWINGS

[0038] Figure 1 Showing the particle size of an exemplary solid preparation of the present invention.

[0039] Figure 2 Showing the particle size of another exemplary solid preparation of the present invention.

[0040] Figure 3 Showing the biological activity and stability of the fusion protein FP4 present in the solid preparation of the present invention.

[0041] Figure 4 Showing that the FP4 fusion protein blocks allergic reactions in transgenic mice. Detailed implementation manners

[0042] Definition Unless otherwise defined herein, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art. References to techniques used herein are intended to refer to techniques commonly understood in the art, including those obvious variations or equivalent substitutions of techniques by one of ordinary skill in the art. Although it is believed that the following terms are well understood by one of ordinary skill in the art, the following definitions are set forth to better explain the present invention.

[0043] As used herein, the terms "comprising", "including", "having", "containing", or "involving" and other variants thereof are inclusive or open-ended and do not exclude other unlisted elements or method steps.

[0044] The term "amino acid" refers to any compound containing an amino group and a carboxylic acid group. Although the amino group most commonly occurs adjacent to the carboxyl functional group, the amino group can be located anywhere within the molecule. Amino acids can also contain additional functional groups such as amino, thio, carboxyl, formamide, imidazole, etc. Amino acids can be synthetic or naturally occurring and can be used in racemic or optically active (D- or L-) forms, including various proportions of stereoisomers.

[0045] A "neat" formulation according to the present invention refers to a solid formulation (usually the fusion protein accounts for 0.1-5% by weight of the total mass of the fusion protein and at least one carrier) composed of particles or powder containing a fusion protein and at least one carrier, and the solid formulation is substantially free of additional excipients, that is, it contains less than about 1% by weight of additional excipients.

[0046] A "fast-acting" powder or particle is a powder or particle that does not have controlled or sustained release characteristics when administered by inhalation.

[0047] A formulation composed of particles or powder refers to a powder formulation that usually contains less than about 10% moisture, preferably less than about 7% moisture, more preferably contains less than about 5-6% moisture, even more preferably contains less than about 3% moisture, even more preferably contains less than about 2% moisture, and most preferably contains less than about 1% moisture, depending on the type of active ingredient in the formulation.

[0048] An "inhalable" formulation "suitable for nasal delivery" refers to a formulation containing solid (i.e., non-solution) microparticles that can (i) be easily dispersed in or through an inhalation device and (ii) be inhaled by a subject such that at least most of the microparticles reach the nasal cavity. Such a powder is considered "respirable" or "inhalable".

[0049] In the context of the present invention, "surfactant-free" means a formulation containing less than about 0.1% by weight of surfactant.

[0050] When "about" is followed by a value or ratio, it means such value or ratio ±10%, preferably ±5%, more preferably ±1%.

[0051] "Substantially" has the meaning commonly known in the art. For example, "substantially free of X" means that X is present at 10 wt% or less, preferably 5 wt% or less, more preferably 1 wt% or less; "substantially unchanged" means that the decomposition rate is 10% or less, preferably 5% or less, more preferably 1% or less.

[0052] "50% volume median diameter (D50) of m or less" means that the particles with a diameter of m or less account for 50 volume% of the total particles. A similar meaning applies to 10% volume median diameter (D10), 90% volume median diameter (D90), etc. The particle sizes reported herein are determined by laser diffraction methods, but any common technique can be used to measure the particle size (e.g., electron microscopy, light scattering, centrifugal sedimentation).

[0053] The term "solvate" as used herein is a substance formed by the combination, physical association, and / or solvation of a compound of the present invention with solvent molecules, such as a disolvate, monosolvate, or hemisolvate, wherein the ratio of solvent molecules to the compound of the present invention is about 2:1, about 1:1, or about 1:2, respectively. Such physical bonding involves varying degrees of ionization and covalent bonding (including hydrogen bonding). In some cases (e.g., when one or more solvent molecules are incorporated into the crystal lattice of a crystalline solid), the solvate can be isolated. Thus, solvates include both the solution phase and separable solvates. The compounds of the present invention can be in solvated forms with pharmaceutically acceptable solvents (such as water, methanol, and ethanol), and this application is intended to cover both solvated and non-solvated forms of the compounds of the present invention.

[0054] One type of solvate is a hydrate. A "hydrate" refers to a specific subset of solvates in which the solvent molecule is water. Solvates generally act in the form of pharmacologically equivalent substances. The preparation of solvates is known in the art, see for example M. Caira et al., J. Pharmaceut. Sci., 93(3): 601-611 (2004), which describes the preparation of a solvate of fluconazole with ethyl acetate and water. Similar methods for the preparation of solvates, hemisolvates, hydrates, etc. are described by van Tonder et al., AAPS Pharm. Sci. Tech., 5(1): Article 12 (2004) and A.L. Bingham et al., Chem. Commun. 603-604 (2001). Representative and non-limiting methods for preparing solvates involve dissolving the compounds of the present invention in the desired solvent (organic solvent, water or a mixture thereof) at a temperature above 20 °C to about 25 °C, then cooling the solution at a rate sufficient to form crystals, and separating the crystals by known methods such as filtration. Analytical techniques such as infrared spectroscopy can be used to confirm the presence of the solvent in the solvate crystals.

[0055] In the context of the present invention, a "pharmaceutically acceptable carrier" or "carrier" or "excipient" refers to a diluent, adjuvant, excipient or vehicle administered together with a therapeutic agent, and which is suitable, within the scope of sound medical judgment, for contact with the tissues of humans and / or other animals and which does not have excessive toxicity, irritation, allergic response, or other problems or complications commensurate with a reasonable benefit / risk ratio. Further, in the context of the present invention, a "pharmaceutically acceptable carrier" or "carrier" or "excipient" refers to an excipient that may optionally be included in the formulations of the present invention and that enters the nasal cavity or lungs without a significant adverse toxicological effect on the subject, particularly the nasal cavity or lungs of the subject.

[0056] Pharmaceutically acceptable carriers or excipients useful in the pharmaceutical formulations of the present invention include, but are not limited to, sterile liquids such as water and oils, including those oils from petroleum, animal, vegetable or synthetic sources, such as peanut oil, soybean oil, mineral oil, sesame oil, etc. Other pharmaceutical carriers or excipients include starch, glucose, lactose, sucrose, gelatin, maltose, chalk, silica, sodium stearate, glycerol monostearate, talc, sodium chloride, skim milk powder, glycerol, propylene glycol, mannitol, water, ethanol, etc. The pharmaceutical formulations may also contain pH buffering agents as required. Examples of suitable pharmaceutically acceptable carriers are described in Remington’s Pharmaceutical Sciences (1990).

[0057] The solid formulations of the present invention can act systemically and / or locally. For this purpose, they can be administered by suitable routes, such as nasally, transmucosally, topically or by inhalation.

[0058] For these administration routes, the solid formulations of the present invention can be administered in suitable dosage forms.

[0059] Dosage forms include, but are not limited to, powders (pulvis) or sprays.

[0060] As used herein, the term "effective amount" refers to the amount of the active ingredient that, upon administration, will, to some extent, alleviate one or more symptoms of the disorder being treated.

[0061] As used herein, "individual" includes human or non-human animals. Exemplary human individuals include human individuals suffering from a disease (such as the diseases described herein) (referred to as patients) or normal individuals. "Non-human animals" in the present invention include all vertebrates, such as non-mammals (such as birds, amphibians, reptiles) and mammals, such as non-human primates, domestic animals and / or domesticated animals (such as sheep, dogs, cats, cows, pigs, etc.).

[0062] Solid preparation For better treatment of nasal diseases, administration by inhalation may be advantageous because in this way the medicament can reach the nasal cavity or the lungs directly, act rapidly, increase bioavailability and reduce the required dose. In addition, administration by inhalation reduces systemic exposure, alleviates side effects by reducing the toxicity of the medicament and is more convenient for the subject.

[0063] However, formulating protein drugs into stable powders or granules remains challenging.

[0064] Freeze-drying and spray-drying are commonly used to formulate protein drugs into powders. Freeze-drying is most widely used today. Freeze-drying is suitable for heat-sensitive proteins, but not suitable for producing uniform powders with diameters ranging from a few micrometers to dozens of micrometers, which can be easily inhaled. Freeze-drying also tends to concentrate the protein between ice crystals upon cooling. This concentration causes rapid changes in the pH and ionic strength around the protein to cause denaturation and precipitation (Schwartz, P.L. et al., Endocrinology, 92(6):1795, 1973; Koseki, T. et al., J. Biochem., 107:389, 1990). In contrast, spray-drying involves spraying a continuous stream of a liquid sample to form micro-dispersed droplets while instantaneously drying them with hot air. Spray-drying has been used to formulate various drugs. Spray-drying has the advantage of producing powders with particle sizes suitable for delivering drugs to the respiratory tract and lungs. Since proteins are usually thermally unstable, there are not many cases of protein drug formulations spray-dried with hot air.

[0065] Although some powder or particulate solid formulations of proteins in the prior art remain in a physical state unchanged (do not form lumps) at room temperature for a certain period of time, such solid formulations are most difficult to maintain biological activity for a long time. Alternatively, such solid formulations may not be in particulate form and may not be suitable for inhalation.

[0066] It has been further found in the present invention that a solid formulation having a D50 particle size of about 10 μm is most suitable for nasal administration to treat allergic rhinitis. The particle size of the solid formulation is about 10 μm, such as 8 to 12 μm, 5 to 15 μm or 2 to 20 μm, which is the optimal particle size for adsorption on the nasal mucosa to increase the bioavailability of the fusion protein.

[0067] The present invention surprisingly finds that the fusion protein can be well dispersed in lactose and mannitol to form substantially uniform particles, which are stable at room temperature, and the biological activity of the fusion protein contained therein remains substantially unchanged for up to one year, and is particularly suitable for administration by inhalation. The mass percentages of the fusion protein and carriers such as lactose and mannitol can vary as long as the fusion protein remains stable in the solid formulation.

[0068] In a preferred aspect, the solid formulation of the present invention is substantially free of surfactants and is more preferably a pure formulation.

[0069] In a preferred aspect, the solid formulation of the present invention is prepared by an atmospheric pressure freeze-drying process.

[0070] On the other hand, the present invention provides a method for preparing a solid preparation of a fusion protein, the method comprising spraying a suspension or solution of the fusion protein into a stream of liquid droplets, entraining the stream of liquid droplets within a coolant stream to freeze the droplets into frozen particles, and drying such particles to form the solid preparation.

[0071] Therapeutic methods and uses The present invention discloses that solid preparations comprising a fusion protein formed by conjugating the FcεRI receptor and the FcγRII receptor (preferably comprising the amino acid sequence of SEQ ID No. 2 or having at least 60%, 70%, 80%, 90%, 95%, 96%, 97%, 98% or 99% identity with SEQ ID No. 2, and more preferably the fusion protein is FP4 having the amino acid sequence of SEQ ID No. 2) are particularly effective for treating allergic rhinitis. Preferably, the solid preparations of the present invention can be applied to mammals by nasal administration, preferably by inhalation, and can effectively inhibit or relieve the symptoms of allergic rhinitis.

[0072] Examples In order to make the objectives and technical solutions of the present invention clearer, the present invention will be further described below in conjunction with specific examples. It should be understood that these examples are not intended to limit the scope of the present invention. In addition, specific experimental methods not mentioned in the following examples are carried out according to conventional experimental methods.

[0073] Materials and Methods A fusion protein having the amino acid sequence of SEQ ID No. 2 (referred to as FP4) was prepared and purified according to WO2005085291A1.

[0074] Other reagents including mannitol, lactose, PBS buffer and Evans blue were purchased from Fisher Scientific.

[0075] FcεRIα-transfected CHO3D10 cells were purchased from ATCC.

[0076] Transgenic mice were purchased from Jackson Laboratory.

[0077] Example 1: Preparation and Characterization of Solid Preparations FP4 was thoroughly mixed with mannitol and lactose in water, and the mixture was subjected to a lyophilization process at atmospheric pressure to obtain a solid preparation. Specifically, the mixture was sprayed into a stream of liquid particles, the stream of liquid particles was entrained within a coolant stream to form frozen particles at a reduced temperature sufficient to form frozen particles, and such frozen particles were dried at a relatively low temperature (which may be higher than the reduced temperature) to form the solid preparation.

[0078] The particle size of such particles was measured using a SympaTEC HELOS / BR device, and exemplary results are as Figure 1 shown. It can be seen that particles with a diameter of 1.09 μm or less accounted for 10% by volume, particles with a diameter of 1.44 μm or less accounted for 16% by volume, particles with a diameter of 4.02 μm or less accounted for 50% by volume, particles with a diameter of 13.41 μm or less accounted for 84% by volume, particles with a diameter of 29.05 μm or less accounted for 90% by volume, and particles with a diameter of 134.82 μm or less accounted for 99% by volume.

[0079] Multiple experiments were repeated. The particle size distribution of another exemplary result is as Figure 2 shown. It can be seen that particles with a diameter of 1.33 μm or less accounted for 10% by volume, particles with a diameter of 2.03 μm or less accounted for 16% by volume, particles with a diameter of 8.87 μm or less accounted for 50% by volume, particles with a diameter of 24.56 μm or less accounted for 84% by volume, particles with a diameter of 34.76 μm or less accounted for 90% by volume, particles with a diameter of 48.21 μm or less accounted for 95% by volume, particles with a diameter of 61.42 μm or less accounted for 98% by volume, and particles with a diameter of 70.04 μm or less accounted for 99% by volume. Compared with the Figure 1 results shown therein, the median particle size shifted to a relatively higher value and the distribution range was relatively narrow.

[0080] Example 2: Study on the activity and stability of solid preparations The solid preparation was stored at room temperature for one month. The biological activity of the solid preparation was measured according to the following procedure.

[0081] The solid preparation prepared in Example 1 with a D50 of 8.87 μm and stored as above was dissolved in PBS buffer and then incubated with CHO3D10 cells transfected with human FcεRIα. After 1 hour, the cells were washed and stained with PE-conjugated anti-human IgE. The binding ability was analyzed by flow cytometry (see the "Sample" row in Figure 3 ). Purified FP4 protein was used as a positive control (see the "PC" row in Figure 3 ). Only cells and only antibody ("Ab") were used as negative controls (see the "NC" row in Figure 3 ).

[0082] Figure 3 It was shown that when the solid preparation containing FP4 was stored at room temperature for one month, the biological activity of FP4 remained substantially unchanged.

[0083] After storing the solid preparation at room temperature for a longer period of time, such as two months, three months... up to one year, the biological activity of the solid preparation was measured according to the above procedure, and the test results showed that when the solid preparation containing FP4 was stored at room temperature up to one year, the biological activity of FP4 remained substantially unchanged.

[0084] Example 3: Biological study of solid preparation Transgenic mice expressing the human FcεRIα chain and knocking out the murine FcεRIα chain were primed by intradermal injection of 250 ng of NP-specific recombinant human IgE in 50 μl of saline. Different doses of FP4 fusion protein were injected simultaneously at each site. Four hours later, the mice were subjected to intravenous challenge with 1.5 mg / ml NP-BSA in 300 μl of saline solution plus 1% Evans blue. Skin allergic reactions were visually evaluated by the blue dye exuded from the blood vessels into the skin. If an allergic reaction occurred, the local skin color turned blue due to the exudation of the dye from the blood vessels. As Figure 4 shown, when the FP4 fusion protein was added, for 0.1 μg and 1 μg of the FP4 fusion protein, the allergic reaction was blocked to some extent, and for 10 μg of the FP4 fusion protein, it was completely blocked (the local skin color did not turn blue). The experimental results further demonstrated that the FP4 fusion protein inhibited allergic reactions in vivo.

[0085] Example 4: Treatment of allergic rhinitis with solid preparation Volunteers diagnosed with allergic rhinitis participated in testing the therapeutic effect of the solid preparation of the present invention. Fifty volunteer subjects with different demographic characteristics (such as gender, age, weight) participated in the test. The participants self-evaluated their nasal symptoms considering the symptoms of sneezing, runny nose, nasal congestion, and nasal itching according to the following criteria in Table 1 below. Before and after treatment, each participant gave themselves a score of 0, 1, 2, or 3.

[0086] Table 1. Criteria for allergic rhinitis symptoms

[0087] During the treatment, each participant was treated by inhaling into each nostril 2 mg of the solid preparation prepared in Example 1 with a D50 of 8.87 μm (the FP4 fusion protein accounted for about 1 wt% of the total mass, i.e., a dose of about 20 μg) twice a day for 3 days. The scores reported by the participants before and after treatment are listed in Table 2 below.

[0088] Table 2. Comparison of nasal symptom scores before and after treatment with the fusion protein solid preparation

[0089] As can be seen from the above results, after treatment, it was observed that the volunteers had significantly alleviated symptoms of nasal congestion, sneezing and / or rhinorrhea, or these symptoms had completely recovered.

[0090] Abbreviations and technical terms

[0091] Sequence Listing SEQ ID NO: 1 SEQ ID NO: 2 FTPPTVKILQSSCDGGGHFPPTIQLLCLVSGYTPGTINITWLEDGQVMDVDLSTASTTQEGELASTQSELTLSQKHWLSDRTYTCQVTYQGHTFEDSTKKCADSNPRGVSAYLSRPSPFDLFIRKSPTITCLVVDLAPSKGTVNLTWSRASGKPVNHSTRKEEKQRNGTLTVTSTLPVGTRDWIEGETYQCRVTHPHLPRALMRSTTKTSGPRAAPEVYAFATPEWPGSRDKRTLACLIQNFMPEDISVQWLHNEVQLPDARHSTTQPRKTKGSGFFVFSRLEVTRAEWEQKDEFICRAVHEAASPSQTVQRAVSVNPGKGSEPKSCDKTHTCPPCPAPELLGGPSVPLPPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKPNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGPYPSDIAVEWESNGQPENNYKTTPPVLDSDGSPPLYSKLTVDKSRWQQGNVPSCSVMHEALHNHYTQKSLSLSPGK

Claims

1. A solid preparation of a fusion protein, wherein the solid preparation consists of particles comprising the fusion protein and at least one carrier, and the fusion protein is a protein comprising the amino acid sequence of SEQ ID No. 2 or having at least 60%, 70%, 80%, 90%, 95%, 96%, 97%, 98% or 99% identity with SEQ ID No.

2.

2. The solid preparation according to claim 1, wherein the fusion protein is FP4 having the amino acid sequence of SEQ ID No.

2.

3. The solid preparation according to claim 1, wherein the particles comprise the fusion protein and one or more carriers.

4. The solid preparation according to any one of claims 1 to 3, wherein the carrier is selected from polyols, carbohydrates and sugars.

5. The solid preparation according to any one of claims 1 to 4, wherein the carrier is selected from the group consisting of sucrose, trehalose, sorbitol, glycerol, mannitol, lactose, xylitol, arabinitol, erythritol, lactitol, maltitol, glucose, raffinose, maltose, dextran, inositol or a combination thereof.

6. The solid preparation according to any one of claims 1 to 5, wherein the particles have a 50% volume median diameter (D50) of 5 μm or less, preferably 4.5 μm or less, more preferably 3.5 to 4.5 μm, and a 90% volume median diameter (D90) of 35 μm or less, preferably 30 μm or less, more preferably 28 to 30 μm.

7. The solid preparation according to any one of claims 1 to 5, wherein the particles have a 50% volume median diameter (D50) higher than 5 μm to 12 μm, preferably 8 to 12 μm, still preferably 8 to 10 μm, more preferably about 9 μm to about 10 μm, most preferably about 10 μm, and a 90% volume median diameter (D90) of 40 μm or less, preferably 38 μm or less, more preferably 34 to 36 μm.

8. The solid preparation according to any one of claims 1 to 7, wherein the solid preparation is substantially free of surfactants and is preferably a pure preparation.

9. The solid preparation according to any one of claims 1 to 8, wherein the solid preparation has a shelf life of 6 months or longer, preferably 1 year or longer.

10. The solid preparation according to any one of claims 1 to 9, wherein the solid preparation is inhalable.

11. The solid preparation according to any one of claims 1 to 10, wherein the solid preparation comprises 0.1 - 5 wt%, preferably about 0.5 - 4 wt%, more preferably 1 - 4 wt% of the fusion protein.

12. A method for preparing a solid preparation according to any one of claims 1 to 11, the method comprising the following steps: Spraying a suspension or solution of the fusion protein into a stream of droplets, entraining the stream of droplets within a coolant stream to freeze the droplets into frozen particles, and drying such particles to form the solid preparation.

13. A method of treating allergic rhinitis in an individual, the method comprising administering to the individual a therapeutically effective amount of the solid preparation according to any one of claims 1 to 11.

14. The method according to claim 13, wherein the solid preparation is administered by inhalation, preferably by means of a nasal spray device.

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