WO2022111547A1

WO2022111547A1 - Inhaled powder formulations for respiratory delivery of antibodies

Info

Publication number: WO2022111547A1
Application number: PCT/CN2021/132924
Authority: WO
Inventors: Ka Wing Lam; Harry Weijie PAN; Chun Kit Jason LO
Original assignee: The University Of Hong Kong
Priority date: 2020-11-24
Filing date: 2021-11-24
Publication date: 2022-06-02

Abstract

Dry powder formulations for inhalation, delivery systems, and methods of making and using thereof have been developed. The dry powder formulation contains particles containing (1) an antibody and/or an antibody derivative; and (2) a β-cyclodextrin and/or a β-cyclodextrin derivative. The total amount of the β-cyclodextrin and/or the β-cyclodextrin derivative is at least 50 wt%of the total amount of the antibody and/or antibody derivative and the β-cyclodextrin and/or β-cyclodextrin derivative. The particles of the dry powder formulation have favorable aerodynamic properties, e.g. mass median aerodynamic diameter < 5 μm, fine particle fraction > 40%in cascade impactor study, and emitted fraction >50%. The dry powder formulation can be prepared using spray-drying or spray-freeze-drying. Each administration of the dry powder formulation can deliver to a subject a unit dose of the antibody and/or antibody derivative to prevent, treat, or ameliorate symptom (s) associated with a respiratory disease in the subject.

Description

INHALED POWDER FORMULATIONS FOR RESPIRATORY DELIVERY OF ANTIBODIES

FIELD OF THE INVENTION

This invention is generally directed to powder formulations for delivery of antibodies and methods for making and using thereof.

BACKGROUND OF THE INVENTION

Antibody therapeutics are exponentially developed in recent years for the treatment of various types of diseases including a range of lung-related diseases such as severe asthma, respiratory infections and lung cancer. The major advantage is their high specificity and affinity for their targets, which facilitates their precise action. Due to the large size of monoclonal antibody (mAb) , tissue penetration is poor. Current antibody therapeutics for the treatment of lung diseases are limited to parenteral injection. For example, biopharmaceutical products approved for the treatment of asthma and lung cancer are confined to parenteral administration. These formulations are associated with issues related to stability, sterility, lung distribution and patient compliance. Antibody fragments with lower molecular weight such as antigen-binding fragment (Fab) have been developed with retained efficacy, but these fragments have short serum half-lives, thus frequent administration is required.

Pulmonary delivery is an attractive non-invasive alternative route of administration for local or even systemic effect. At present, no inhaled mAb or its fragments are commercially available in the market for the treatment of respiratory conditions. The major challenges are to achieve favorable aerosol performance for efficient lung deposition while preserving the structural integrity of antibodies for their proper function.

There remains a need for formulations of antibodies for inhalation therapy.

Therefore, it is the object of the present invention to provide dry powder formulations for delivery of antibodies by inhalation.

It is another object of the present invention to provide methods of making the dry powder formulations.

It is yet another object of the present invention to provide methods of using the dry powder formulations.

SUMMARY OF THE INVENTION

Dry powder formulations for inhalation, delivery systems for the dry powder formulations, and methods of making and using thereof have been developed. The dry powder formulation contains particles containing (1) an antibody or an antibody derivative, or a combination thereof; and (2) a β-cyclodextrin or a β-cyclodextrin derivative, or a combination thereof, where the amount of the β-cyclodextrin or the β-cyclodextrin derivative, or the total amount of the β-cyclodextrin and β-cyclodextrin derivative is at least 50 wt%of the total amount of the antibody and/or antibody derivative and the β-cyclodextrin and/or β-cyclodextrin derivative.

The particles of the dry powder formulation have favorable aerodynamic properties for effective lung deposition. For example, the particles of the dry powder formulation have a mass median aerodynamic diameter < 5 μm, < 4 μm, < 3.5 μm, < 3 μm, < 2.5 μm, or < 2 μm; a geometric standard deviation < 4.5, < 4, < 3.5, < 3, < 2.5, or < 2; and/or a fine particle fraction > 40%, > 45%, > 50%, > 55%, or > 60%in cascade impactor study.

In some aspects, the dry powder formulation contains particles containing an antibody derivative and a β-cyclodextrin derivative. The β-cyclodextrin derivative can have a degree of substitution of from 1 to 21, from 1 to 12, from 2 to 15, from 2 to 12, from 2 to 10, from 4 to 10, from 5 to 8, from 6 to 9, or from 6 to 8. In some aspects, the antibody derivative contained in the particles of the dry powder formulation can be an antigen-binding fragment (Fab) . In some aspects, the β-cyclodextrin derivative contained in the particles of the dry powder formulation can be 2-hydroxypropyl-β-cyclodextrin, methyl-β-cyclodextrin, or sulfobutylether β-cyclodextrin, or a combination thereof. In some aspect, the β-cyclodextrin derivative contained in the particles of the dry powder formulation is 2-hydroxypropyl-β-cyclodextrin.

The dry powder formulation may further contain a pharmaceutically acceptable excipient or an additional active agent, or a combination thereof. The pharmaceutically acceptable excipient can be an amino acid, a peptide, a lipid, a protein, a chelating agent, a salt, a taste masking agent, a cation, or a polymer, or a combination thereof. The amount of the pharmaceutically acceptable excipient in the dry powder formulation can be in a range from 0.1 wt%to 20 wt%, from, from 0.1 wt%to 15 wt%, or from 1 wt%to 10 wt%of the dry powder formulation. The additional active agent can be an anti-inflammatory agent, an anti-viral agent, or a chemotherapeutic agent, or a combination thereof.

In some aspects, the dry powder formulation does not contain any additional pharmaceutically acceptable excipients. In some aspects, the dry powder formulation does not contain any additional carbohydrates, such as mannitol, trehalose, 1, 4 O-linked saccharose or 1, 4 O-linked saccharose derivatives, or dexran, or a combination thereof. In some aspects, the dry powder formulation does not contain mannitol or trehalose, or a combination of mannitol and trehalose. In some aspects, the dry powder formulation does not contain 1, 4 O-linked saccharose or 1, 4 O-linked saccharose derivatives, or a combination thereof. In some aspects, the dry powder formulation does not contain dexran. In some aspects, the dry powder formulation does not contain any one of mannitol, trehalose, 1, 4 O-linked saccharose, 1, 4 O-linked saccharose derivatives, and dexran.

Delivery systems containing an inhaler and the dry powder formulation are provided. In some aspects, the dry powder formulation may be in a unit dosage form that contains a single unit does of the antibody and/or antibody derivative. The total amount of antibody and/or antibody derivative in the unit dosage form of the dry powder formulation can be in a range from about 0.1 mg to about 50 mg, from about 0.5 to about 50 mg, or from about 1 to about 50 mg. In some aspects, the dry powder formulation may contain multiple doses of the antibody and/or antibody derivative. The inhaler for delivering the dry powder formulation can be a dry powder inhaler. The emitted fraction of the powder formulation can be > 50%, > 55%, > 60%, > 65%, > 70%, > 75%, > 80%, > 85%, > 90%, or > 95%.

Also provided are methods of making the dry powder formulation. Generally, the method includes (i) mixing an antibody and/or an antibody derivative and a β-cyclodextrin and/or a β-cyclodextrin derivative, and optionally a pharmaceutically acceptable excipient and/or an additional active agent, in a solvent to form a liquid feed; and (ii) spray-drying or spray-freeze drying the liquid feed to form particles containing the antibody and/or antibody derivative and the β-cyclodextrin and/or β-cyclodextrin derivative, and optionally the pharmaceutically acceptable excipient and/or the additional active agent. Typically, following step (ii) , the production yield of the particles is at least about 50 wt%, at least 55 wt%, at least 60 wt%, at least 65 wt%, at least 70 wt%, at least 75 wt%, at least 80 wt%, at least 85 wt%, or at least 90 wt%, or in a range from about 50 wt%to about 95 wt%, from about 55 wt%to about 95 wt%, from about 55 wt%to about 90 wt%, from about 60 wt%to about 95 wt%, from about 65 wt%to about 95 wt%, from about 70 wt%to about 95 wt%, or from about 75 wt%to about 95 wt%.

Further provided are methods of preventing, treating, or ameliorating symptom (s) associated with a respiratory disease in a subject. The subject being treated can be a mammal having or at the risk of having a respiratory disease, such as an obstructive airway disease, an inflammatory airway disease, or lung cancer, or a combination thereof. Generally, the method includes (i) administering to the subject the dry powder formulation. Optionally, the method further includes a step of loading the dry powder formulation in a dry powder inhaler prior to step (i) .

Typically, following step (i) , a unit dose of the antibody and/or the antibody derivative is delivered to the lung (s) of the subject in an amount from about 1 mg to about 3000 mg, from about 1 mg to about 1500 mg, from about 10 mg to about 1500 mg, from about 10 mg to about 1000 mg, from about 20 mg to about 1000 mg, from about 50 mg to about 1500 mg, from about 50 mg to about 1000 mg, or from about 20 mg to about 500 mg.

The method may further include repeating step (i) administering to the subject the dry powder formulation. In some aspects, step (i) can be repeated every hour, every 2 hours, every 5 hours, every 8 hours, every day, every 2 days, every 3 days, every 5 days, every 7 days, every 10 days, every two weeks, or every month. The period for repeated administration of the dry powder formulation can be between one day and 6 months, between one day and 3 months, between one and thirty days, between one and ten days, between one and three days, between one and two days, or during one day.

BRIEF DESCRIPTION OF THE DRAWINGS

Figures 1A-1B are schematic diagrams of forming particles using spray drying (Figure 1A) or spray freeze drying (Figure 1B) .

Figure 2A is a scanning electron microscopy (SEM) image of particles formed using spray drying, where the particles contain 5 wt%antigen-binding fragment (also referred herein as “Fab” ) and 95 wt%2-hydroxypropyl-β-cyclodextrin (also referred herein as “HPβCD” ) (i.e. formulation SD-B1) . Figure 2B is a SEM image of the particles at a higher magnification.

Figure 3A is a SEM image of particles formed using spray freeze drying, where the particles contain 10 wt%Fab and 90 wt%HPβCD (i.e. formulation SFD-F12) . Figure 3B is a SEM image of the particles at a higher magnification.

Figure 4A is a SEM image of particles formed using spray freeze drying, where the particles contain 2 wt%Fab and 98 wt%HPβCD (i.e. formulation SFD-F14) . Figure 4B is a SEM image of the particles at a higher magnification.

Figure 5 is an image of SDS-PAGE analysis of powder formulations of Fab formed by spray drying without reducing (left) and reduced with dithiothreitol (also referred herein as “DTT” ) (right) . “CD” represents unprocessed cyclodextrin; “Fab up”represents unprocessed Fab; and “B1 and B3” represents formulations SD-B1 and SD-B3, respectively.

Figure 6 is an image of SDS-PAGE analysis of powder formulations of Fab formed by spray freeze drying without reducing (left) and reduced with DTT (right) . “CD” represents unprocessed cyclodextrin; “Fab up” represents unprocessed Fab; and “F12 and F14” represents formulations SFD-F12 and SFD-F14, respectively.

Figure 7 is a graph showing the antibody binding of formulation SD-B1 in comparison with unprocessed Fab, assessed by ELISA.

Figure 8 is a graph showing the antibody binding of formulations SFD-F12 and SFD-F14, respectively, in comparison with unprocessed Fab, assessed by ELISA.

Figure 9 is a graph showing the in vitro toxicity of formulation SD-B3 on TF-l cells in comparison with unprocessed Fab, assessed by a TF-1 cell proliferation assay.

Figure 10 is a graph showing the in vitro toxicity of formulations SFD-F12 and SFD-F14 on TF-l cells, respectively, in comparison with unprocessed Fab, assessed by a TF-1 cell proliferation assay.

DETAILED DESCRIPTION OF THE INVENTION

I. Definitions

As used herein, the term “active agent” refers to a physiologically or pharmacologically active substance that acts locally and/or systemically in the body. An active agent is a substance that is administered to a patient for the treatment (e.g., therapeutic agent) , prevention (e.g., prophylactic agent) , or diagnosis (e.g., diagnostic agent) of a disease or disorder.

As used herein, the term “pharmaceutically acceptable” as used herein refers to those compounds, materials, and/or compositions, which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of human beings and animals without excessive toxicity, irritation, allergic response, or other problems or complications commensurate with a reasonable benefit/risk ratio.

As used herein, the term “treating” includes inhibiting or eliminating one or more symptoms or side effects associated with the disease, condition, or disorder being treated.

As used herein, the term “alleviating” is relative to a control. One of skill in the art would readily identify the appropriate control to use for each experiment. For example, a decreased response in a subject or cell treated with the dry powder formulation containing an antibody and/or antibody derivative is compared to a control, such as to a response in subject or cell that is not treated with the dry powder formulation, or treated with a different formulation. The decrease may be an inhibition or reduction of in activity, expression, or a symptom, or a partial inhibition or reduction. Inhibition can be 1, 2, 5, 10, 20, 30, 50, 75, 80, 90, 95, or higher%reduction in activity, expression, or a symptom relative to the control.

As used herein, the term “alkyl” refers to univalent groups derived from alkanes by removal of a hydrogen atom from any carbon atom. Alkanes represent saturated hydrocarbons, including those that are linear, branched, or cyclic (either monocyclic or polycyclic) . An alkyl can be a linear C ₁-C ₃₀ alkyl, a branched C ₄-C ₃₀ alkyl, a cyclic C ₃-C ₃₀ alkyl, a linear C ₁-C ₃₀ alkyl or a branched C ₄-C ₃₀ alkyl, a linear C ₁-C ₃₀ alkyl or a cyclic C ₃-C ₃₀ alkyl, a branched C ₄-C ₃₀ alkyl or a cyclic C ₃-C ₃₀ alkyl. Optionally, alkyl groups have up to 20 carbon atoms. An alkyl can be a linear C ₁-C ₂₀ alkyl, a branched C ₄-C ₂₀ alkyl, a cyclic C ₃-C ₂₀ alkyl, a linear C ₁-C ₂₀ alkyl or a branched C ₄-C ₂₀ alkyl, a branched C ₄-C ₂₀ alkyl or a cyclic C ₃-C ₂₀ alkyl, a linear C ₁-C ₂₀ alkyl or a cyclic C ₃-C ₂₀ alkyl. Optionally, alkyl groups have up to 10 carbon atoms. An alkyl can be a linear C ₁-C ₁₀ alkyl, a branched C ₄-C ₁₀ alkyl, a cyclic C ₃-C ₁₀ alkyl, a linear C ₁-C ₁₀ alkyl or a branched C ₄-C ₁₀ alkyl, a branched C ₄-C ₁₀ alkyl or a cyclic C ₃-C ₁₀ alkyl, a linear C ₁-C ₁₀ alkyl or a cyclic C ₃-C ₁₀ alkyl. Optionally, alkyl groups have up to 6 carbon atoms. An alkyl can be a linear C ₁-C ₆ alkyl, a branched C ₄-C ₆ alkyl, a cyclic C ₃-C ₆ alkyl, a linear C ₁-C ₆ alkyl or a branched C ₄-C ₆ alkyl, a branched C ₄-C ₆ alkyl or a cyclic C ₃-C ₆ alkyl, or a linear C ₁-C ₆ alkyl or a cyclic C ₃-C ₆ alkyl. Optionally, alkyl groups have up to four carbons. An alkyl can be a linear C ₁-C ₄ alkyl, cyclic C ₃-C ₄ alkyl, a linear C ₁-C ₄ alkyl or a cyclic C ₃-C ₄ alkyl. Preferably, the alkyl group is unsubstituted alkyl group. Preferably, the alkyl group is a linear C ₁-C ₅, C ₁-C ₄, C ₁-C ₃, C ₁-C ₂ alkyl group, such as methyl group.

As used herein, the term “heteroalkyl” refers to alkyl groups where one or more carbon atoms are replaced with a heteroatom, such as, O, N, or S. Heteroalkyl group can be linear, branched, or cyclic. A heteroalkyl can be a linear C ₁-C ₃₀ heteroalkyl, a branched C ₃-C ₃₀ heteroalkyl, a cyclic C ₂-C ₃₀ heteroalkyl, a linear C ₁-C ₃₀ heteroalkyl or a branched C ₃-C ₃₀ heteroalkyl, a linear C ₁-C ₃₀ heteroalkyl or a cyclic C ₂-C ₃₀ heteroalkyl, a branched C ₃-C ₃₀ heteroalkyl or a cyclic C ₂-C ₃₀ heteroalkyl. Optionally, heteroalkyl groups have up to 20 carbon atoms. A heteroalkyl can be a linear C ₁-C ₂₀ heteroalkyl, a branched C ₃-C ₂₀ heteroalkyl, a cyclic C ₂-C ₂₀ heteroalkyl, a linear C ₁-C ₂₀ heteroalkyl or a branched C ₃-C ₂₀ heteroalkyl, a branched C ₃-C ₂₀ heteroalkyl or a cyclic C ₂-C ₂₀ heteroalkyl, or a linear C ₁-C ₂₀ heteroalkyl or a cyclic C ₂-C ₂₀ heteroalkyl. Optionally, heteroalkyl groups have up to 10 carbon atoms. A heteroalkyl can be a linear C ₁-C ₁₀ heteroalkyl, a branched C ₃-C ₁₀ heteroalkyl, a cyclic C ₂-C ₁₀ heteroalkyl, a linear C ₁-C ₁₀ heteroalkyl or a branched C ₃-C ₁₀ heteroalkyl, a branched C ₃-C ₁₀ heteroalkyl or a cyclic C ₂-C ₁₀ heteroalkyl, or a linear C ₁-C ₁₀ heteroalkyl or a cyclic C ₂-C ₁₀ heteroalkyl. Optionally, heteroalkyl groups have up to 6 carbon atoms. A heteroalkyl can be a linear C ₁-C ₆ heteroalkyl, a branched C ₃-C ₆ heteroalkyl, a cyclic C ₂-C ₆ heteroalkyl, a linear C ₁-C ₆ heteroalkyl or a branched C ₃-C ₆ heteroalkyl, a branched C ₃-C ₆ heteroalkyl or a cyclic C ₂-C ₆ heteroalkyl, or a linear C ₁-C ₆ heteroalkyl or a cyclic C ₂-C ₆ heteroalkyl. Optionally, heteroalkyl groups have up to four carbons. A heteroalkyl can be a linear C ₁-C ₄ heteroalkyl, a branched C ₃-C ₄ heteroalkyl, a cyclic C ₂-C ₄ heteroalkyl, a linear C ₁-C ₄ heteroalkyl or a branched C ₃-C ₄ heteroalkyl, a branched C ₃-C ₄ heteroalkyl or a cyclic C ₂-C ₄ heteroalkyl, or a linear C ₁-C ₄ heteroalkyl or a cyclic C ₂-C ₄ heteroalkyl.

As used herein, the term “alkenyl” refers to univalent groups derived from alkenes by removal of a hydrogen atom from any carbon atom. Alkenes are unsaturated hydrocarbons that contain at least one carbon-carbon double bond. Alkenyl group can be linear, branched, or cyclic. An alkenyl can be a linear C ₂-C ₃₀ alkenyl, a branched C ₄-C ₃₀ alkenyl, a cyclic C ₃-C ₃₀ alkenyl, a linear C ₂-C ₃₀ alkenyl or a branched C ₄-C ₃₀ alkenyl, a linear C ₂-C ₃₀ alkenyl or a cyclic C ₃-C ₃₀ alkenyl, a branched C ₄-C ₃₀ alkenyl or a cyclic C ₃-C ₃₀ alkenyl. Optionally, alkenyl groups have up to 20 carbon atoms. An alkenyl can be a linear C ₂-C ₂₀ alkenyl, a branched C ₄-C ₂₀ alkenyl, a cyclic C ₃-C ₂₀ alkenyl, a linear C ₂-C ₂₀ alkenyl or a branched C ₄-C ₂₀ alkenyl, a linear C ₂-C ₂₀ alkenyl or a cyclic C ₃-C ₂₀ alkenyl, a branched C ₄-C ₂₀ alkenyl or a cyclic C ₃-C ₂₀ alkenyl. Optionally, alkenyl groups have two to 10 carbon atoms. An alkenyl can be a linear C ₂-C ₁₀ alkenyl, a branched C ₄-C ₁₀ alkenyl, a cyclic C ₃-C ₁₀ alkenyl, a linear C ₂-C ₁₀ alkenyl or a branched C ₄-C ₁₀ alkenyl, a linear C ₂-C ₁₀ alkenyl or a cyclic C ₃-C ₁₀ alkenyl, a branched C ₄-C ₁₀ alkenyl or a cyclic C ₃-C ₁₀ alkenyl. Optionally, alkenyl groups have two to 6 carbon atoms. An alkenyl can be a linear C ₂-C ₆ alkenyl, a branched C ₄-C ₆ alkenyl, a cyclic C ₃-C ₆ alkenyl, a linear C ₂-C ₆ alkenyl or a branched C ₄-C ₆ alkenyl, a linear C ₂-C ₆ alkenyl or a cyclic C ₃-C ₆ alkenyl, a branched C ₄-C ₆ alkenyl or a cyclic C ₃-C ₆ alkenyl. Optionally, alkenyl groups have two to four carbons. An alkenyl can be a linear C ₂-C ₄ alkenyl, a cyclic C ₃-C ₄ alkenyl, a linear C ₂-C ₄ alkenyl or a cyclic C ₃-C4 alkenyl.

As used herein, the term "amino" includes the group NH ₂ (primary amino) , alkylamino (secondary amino) , and dialkylamino (tertiary amino) , where the two alkyl groups in dialkylamino may be the same or different, i.e. alkylalkylamino. Illustratively, amino include methylamino, ethylamino, dimethylamino, methylethylamino, and the like. In addition, it is to be understood that when amino modifies or is modified by another term, such as aminoalkyl, or acylamino, the above variations of the term amino continue to apply. Illustratively, aminoalkyl includes H ²N-alkyl, methylaminoalkyl, ethylaminoalkyl, dimethylaminoalkyl, methylethylaminoalkyl, and the like. Illustratively, acylamino includes acylmethylamino, acylethylamino, and the like.

As used herein, the term "amide" includes the group CONH ₂ (primary amide) , CONHalkyl (secondary amide) , and CONdialkyl (tertiary amide) , where the two alkyl groups in CONdialkyl may be the same or different.

In a substituted functional group, one or more hydrogen atoms in the chemical group or moiety is replaced with one or more substituents. Any substitution is in accordance with permitted valence of the substituted atom and the substituent, and that the substitution results in a stable compound, i.e., a compound that does not spontaneously undergo transformation such as by rearrangement, cyclization, elimination, etc. Suitable substituents include, but are not limited to a halogen atom, an alkyl group, a cycloalkyl group, a heteroalkyl group, a cycloheteroalkyl group, an alkenyl group, a heteroalkenyl group, an alkynyl group, a heteroalkynyl group, an aryl group, a heteroaryl group, a polyaryl group, a polyheteroaryl group, -OH, -SH, -NH ₂, -N ₃, -OCN, -NCO, -ONO ₂, -CN, -NC, -ONO, -CONH ₂, -NO, -NO ₂, -ONH ₂, -SCN, -SNCS, -CF ₃, -CH ₂CF ₃, -CH ₂Cl, -CHCl ₂, -CH ₂NH ₂, -NHCOH, -CHO, -COCl, -COF, -COBr, -COOH, -SO ₃H, -CH ₂SO ₂CH ₃, -PO ₃H ₂, -OPO ₃H ₂, -P (=O) (OR ^T1′) (OR ^T2′) , -OP (=O) (OR ^T1′) (OR ^T2′) , -BR ^T1′ (OR ^T2′) , -B (OR ^T1′) (OR ^T2′) , or -G ^′R ^T1′in which -T′ is -O-, -S-, -NR ^T2′-, -C (=O) -, -S (=O) -, -SO ₂-, -C (=O) O-, -C (=O) NR ^T2′-, -OC (=O) -, -NR ^T2′C (=O) -, -OC (=O) O-, -OC (=O) NR ^T2′-, -NR ^T2′C (=O) O-, -NR ^T2′C (=O) NR ^T3′-, -C (=S) -, -C (=S) S-, -SC (=S) -, -SC (=S) S-, -C (=NR ^T2′) -, -C (=NR ^T2′) O-, -C (=NR ^T2′) NR ^T3′-, -OC (=NR ^T2′) -, -NR ^T2′C (=NR ^T3′) -, -NR ^T2′SO ₂-, -C (=NR ^T2′) NR ^T3′-, -OC (=NR ^T2′) -, -NR ^T2′C (=NR ^T3′) -, -NR ^T2′SO ₂-, -NR ^T2′SO ₂NR ^T3′-, -NR ^T2′C (=S) -, -SC (=S) NR ^T2′-, -NR ^T2′C (=S) S-, -NR ^T2′C (=S) NR ^T3′-, -SC (=NR ^T2′) -, -C (=S) NR ^T2′-, -OC (=S) NR ^T2′-, -NR ^T2′C (=S) O-, -SC (=O) NR ^T2′-, -NR ^T2′C (=O) S-, -C (=O) S-, -SC (=O) -, -SC (=O) S-, -C (=S) O-, -OC (=S) -, -OC (=S) O-, -SO ₂NR ^T2′-, -BR ^T2′-, or –PR ^T2′-; where each occurrence of R ^T1′, R ^T2′, and R ^T3′is, independently, a hydrogen atom, a halogen atom, an alkyl group, a heteroalkyl group, an alkenyl group, a heteroalkenyl group, an alkynyl group, a heteroalkynyl group, an aryl group, or a heteroaryl group.

Recitation of ranges of values herein are merely intended to serve as a shorthand method of referring individually to each separate value falling within the range, unless otherwise indicated herein, and each separate value is incorporated into the specification as if it were individually recited herein.

Use of the term "about" is intended to describe values either above or below the stated value in a range of approx. +/-10%.

II. DRY POWDER FORMULATIONS

Disclosed herein are dry powder formulations of antibodies for inhalation into the lungs of a subject. The dry powder formulation contains particles containing an antibody or an antibody derivative, or a combination thereof, and a β-cyclodextrin or a β-cyclodextrin derivative, or a combination thereof. Generally, the amount of the β-cyclodextrin or β-cyclodextrin derivative, or the total amount of the β-cyclodextrin and β-cyclodextrin derivative is at least 50 wt%of the total amount of the antibody and/or the antibody derivative and the β-cyclodextrin and/or β-cyclodextrin derivative in the dry powder formulation.

For example, the weight ratio of the β-cyclodextrin and/or β-cyclodextrin derivative to the antibody and/or antibody derivative is in a range from 1: 1 to 49: 1, from 1: 1 to 45: 1, from 1: 1 to 40: 1, from 1: 1 to 35: 1, from 1: 1 to 30: 1, from 1: 1 to 25: 1, from 1: 1 to 19: 1, from 1: 1 to 15: 1, from 1: 1 to 12: 1, from 1: 1 to 10: 1, from 1: 1 to 8: 1, from 1: 1 to 5: 1, from 1.2: 1 to 5: 1, from 1.5: 1 to 5: 1, from 1.8: 1 to 5: 1, from 2: 1 to 5: 1, from 2: 1 to 8: 1, from 2: 1 to 10: 1, from 2: 1 to 12: 1, from 2: 1 to 15: 1, from 2: 1 to 19: 1, from 2: 1 to 25: 1, from 2: 1 to 30: 1, from 2: 1 to 35: 1, from 2: 1 to 40: 1, from 2: 1 to 45: 1, from 2: 1 to 49: 1, from 2.5: 1 to 5: 1, from 2.5: 1 to 8: 1, from 2.5: 1 to 10: 1, from 2.5: 1 to 12: 1, from 2.5: 1 to 15: 1, from 2.5: 1 to 19: 1, from 2.5: 1 to 25: 1, from 2.5: 1 to 30: 1, from 2.5: 1 to 35: 1, from 2.5: 1 to 40: 1, from 2.5: 1 to 45: 1, from 2.5: 1 to 49: 1, from 3: 1 to 5: 1, from 3: 1 to 8: 1, from 3: 1 to 10: 1, from 3: 1 to 12: 1, from 3: 1 to 15: 1, from 3: 1 to 19: 1, from 3: 1 to 25: 1, from 3: 1 to 30: 1, from 3: 1 to 35: 1, from 3: 1 to 40: 1, from 3: 1 to 45: 1, from 3: 1 to 49: 1, from 4: 1 to 5: 1, from 4: 1 to 8: 1, from 4: 1 to 10: 1, from 4: 1 to 12: 1, from 4: 1 to 15: 1, from 4: 1 to 19: 1, from 4: 1 to 25: 1, from 4: 1 to 30: 1, from 4: 1 to 35: 1, from 4: 1 to 40: 1, from 4: 1 to 45: 1, or from 4: 1 to 49: 1, such as from 1: 1 to 49: 1, from 1: 1 to 19: 1, from 1: 1 to 9: 1, or from 1: 1 to 5: 1.

The dry powder formulation may also contain a pharmaceutically acceptable carrier and/or an additional active agent.

Generally, the particles of the dry powder formulation have favorable aerodynamic properties (e.g. mass median aerodynamic diameter < 5 μm and fine particle fraction > 40%in cascade impactor study) for effective lung deposition, and retain the structural integrity and binding activity of the antibody. For example, the antibody and/or antibody derivative formulated in the dry powder formulation can have a binding activity (which can be evaluated by ELISA, for example) and/or an inhibitory potency (which can be evaluated by cellular assays, for example) that are comparable to those of the same but unprocessed and unformulated antibody and/or antibody derivative. That is, the change of LogEC ₅₀ and/or LogIC ₅₀ value of the antibody and/or antibody derivative against the same binding target following formulation can be less than about 30%compared to those prior to processing and formulation.

Systems for delivering the dry powder formulations are also disclosed. Generally, the system includes an inhaler and the dry powder formulation. The emitted fraction of the powder formulation, i.e., the fraction of powder exited the inhaler, is >50%.

A. Antibodies and Antibody Derivatives

The dry powder formulation contains an antibody or an antibody derivative, or a combination thereof.

Suitable antibodies and antibody derivatives that can be used in the dry powder formulation include, but are not limited to, monoclonal antibodies, polyclonal antibodies, multi-specific antibodies, single-chain antibodies, and their fragments thereof. Preferably, the dry powder formulation contains an antigen-binding fragment ( “Fab” ) .

1. Exemplary Antibodies and Antibody Derivatives

Antibodies suitable for use in the dry powder formulation may be of human or non-human origin. Exemplary antibodies include, but are not limited to, IgA, IgD, IgE, IgG, and IgM with their various subclasses, such as e.g. IgA1, IgA2, IgG1, IgG2, IgG3, and IgG4. In some aspects, the antibodies may be humanized or chimeric antibodies. Additionally, exemplary antibodies include antibodies against various surface antigens (e.g. CD4. CD20, or CD44) or various cytokines (e.g. IL-2, IL-4, or IL-5) , antibodies against specific antibody classes (e.g. anti-IgE antibodies, anti-IgG antibodies, etc. ) , and antibodies against viral proteins (e.g. anti-RSV, anti-CMV antibodies, etc. ) .

Exemplary antibody derivatives for use in the dry powder formulation include, but are not limited to, Fab, Fab’, F (ab’) ₂, Fc, and Fc’ fragments, light (L) and heavy (H) antibody chains and their constant, variable, or hypervariable regions, Fd and Fv fragments, and fusion proteins containing at least an antibody fraction or at least a fragment of a double-or single-chain antibody (see, e.g., Chamov et al., 1999, Antibody Fusion Proteins, Wiley-Liss Inc. ) . In some aspects, the antibody derivatives for use in the dry powder formulation are Fab, i.e., antigen-binding fragments. Fab typically contains the variable regions of both chains that are held together by the adjacent constant regions. For example, the Fab used in the dry powder formulation is the Fab that targets the alpha subunit of the receptor for IL-4 (IL-4Ra) from Shanghai MabGeek BioTech. In some aspects, the antibody derivatives for use in the dry powder formulation are F (ab') , fragments that can be produced by proteolytic digestion with pepsin. In some aspects, the derivatives for use in the dry powder formulation are Fv, shortened antibody fragments that only contain the variable regions of the heavy (V _H) and light (V _L) chain. Such antibody fragments are also known as single-chain FV (scFv) fragments. Examples of scFv fragments are known (see, e.g., Huston, et al., 1988, Proc. Natl. Acad. Sci. USA, 16, 5879ff) . In some aspects, the scFv fragments may be multimeric scFv derivatives, such as dia-, tria-, and pentabodies (see, e.g., Perisicetal, Structure, 1994, 2, 1217ff; Hu et al., 1996, Cancer Research, 56, 3055ff; and Kortt et al., 1997, Protein Engineering, 10, 423ff) and mini-antibodies having a bi, tri-, or tetravalent structures (see, e.g., Pack P. et al., 1993, Biotechnology, 11 1271ff; Lovejoy B. et al., 1993, Science, 259, 1288ff; and Pack P. et al., 1995, J. Mol. Biol., 246, 28ff) .

FDA-approved mAb used in the treatment of respiratory diseases for subcutaneous or intravenous injection may also be used in the dry powder formulation for inhalation therapy, such as Dupilumab (

Sanofi) , Mepolizumab (

GSK) , Omalizumab (

Genentech) , Atezolizumab (

Genentech) , Bevacizumab (

Genentech) , Ipilimumab (

BMS) , and Necitumumab (

Eli Lilly) .

2.Amount of Antibodies and/or Antibody Derivatives

Generally, the amount of the antibody or antibody derivative, or the total amount of the antibody and antibody derivative in the dry powder formulation is in a range from about 1 wt%to about 50 wt%, from about 2 wt%to about 50 wt%, from about 5 wt%to about 50 wt%, from about 8 wt%to about 50 wt%, from about 10 wt%to about 50 wt%, from about 12 wt%to about 50 wt%, from about 15 wt%to about 50 wt%, from about 16 wt%to about 50 wt%, from about 17 wt%to about 50 wt%, from about 18 wt%to about 50 wt%, or from about 20 wt%to about 50 wt%of the total weight of the antibody and/or antibody derivative and the β-cyclodextrin and/or β-cyclodextrin derivative. For example, the amount of the antibody or antibody derivative, or the total amount of the antibody and antibody derivative in the dry powder formulation is in a range from about 5 wt%to about 50 wt%of the total weight of the antibody and/or antibody derivative and the β-cyclodextrin and/or β-cyclodextrin derivative. The term “total amount of the antibody and antibody derivative” refers to the total weight of the antibody and antibody derivative relative to the total weight of the antibody and antibody derivative and the β-cyclodextrin and/or β-cyclodextrin derivative in the dry powder formulation.

In some aspects, the dry powder formulation contains a Fab and the amount of the Fab is in a range from about 1 wt%to about 50 wt%, from about 2 wt%to about 50 wt%, from about 5 wt%to about 50 wt%, from about 8 wt%to about 50 wt%, from about 10 wt%to about 50 wt%, from about 12 wt%to about 50 wt%, from about 15 wt%to about 50 wt%, from about 16 wt%to about 50 wt%, from about 17 wt%to about 50 wt%, from about 18 wt%to about 50 wt%, or from about 20 wt%to about 50 wt%of the total weight of the Fab and the β-cyclodextrin and/or β-cyclodextrin derivative.

In some aspects, the dry powder formulation contains two or more Fabs and the amount of each Fab can be in a suitable range to provide a total amount of the Fabs in a range from about 1 wt%to about 50 wt%, from about 2 wt%to about 50 wt%, from about 5 wt%to about 50 wt%, from about 8 wt%to about 50 wt%, from about 10 wt%to about 50 wt%, from about 12 wt%to about 50 wt%, from about 15 wt%to about 50 wt%, from about 16 wt%to about 50 wt%, from about 17 wt%to about 50 wt%, from about 18 wt%to about 50 wt%, or from about 20 wt%to about 50 wt%of the total weight of the Fabs and the β-cyclodextrin and/or β-cyclodextrin derivative.

B. β-Cyclodextrins and Derivatives Thereof

The dry powder formulation contains a β-cyclodextrin or a β-cyclodextrin derivative, or a combination thereof. In some aspects, the dry powder formulation contains a β-cyclodextrin. In some aspects, the dry powder formulation contains a β-cyclodextrin derivative, such as 2-hydroxypropyl-β-cyclodextrin ( “HPβCD” ) , methyl-β-cyclodextrin ( “MβCD” ) , or sulfobutylether β-cyclodextrin ( “SBEβCD” ) , or a combination thereof. In some aspects, the dry powder formulation contains a combination of a β-cyclodextrin and a β-cyclodextrin derivative.

The β-cyclodextrin and/or β-cyclodextrin derivative in the dry powder formulation can protect the antibody and/or antibody derivative from fragmentation and aggregation during the manufacture process, such as during spray drying or spray freeze drying. For example, the antibody and/or antibody derivative in the dry powder formulation do not show fragments following manufacture (as determined by SDA-PAGE analysis) and shows an aggregation percentage of less than about 1% (as determined by SEC-HPLC) following manufacture. Additionally or alternatively, the β-cyclodextrin and/or β-cyclodextrin derivative in the dry powder formulation can stabilize the antibody and/or antibody derivative to prevent or reduce aggregation during storage following manufacture. For example, the antibody and/or antibody derivative in the dry powder formulation shows an aggregation percentage of less than 1%or less than 0.5% (as determined by SEC-HPLC) 4 weeks post-drying, 8 weeks post-drying, 10 weeks post-drying, 12 weeks post-drying, or 16 weeks post-drying.

Generally, a β-cyclodextrin derivative can be obtained by chemical modification of the hydroxyl group (s) of β-cyclodextrin with an organic functional group, i.e. by substituting the hydrogen (s) of the hydroxyl group (s) with an organic functional group. For example, there are 21 hydroxyl groups on a β-cyclodextrin, which are 21 substitution sites for reaction with the organic functional group. A β-cyclodextrin derivative can have different degrees of substitution. The term “degree of substitution” refers to the number of hydroxyl groups that are modified with the organic functional group. For example, a degree of substitution of 6 means that 6 hydroxyl groups on the β-cyclodextrin are modified with an organic group.

The degree of substitution of the β-cyclodextrin derivative for use in the dry powder formulation can be in a range from 1 to 21, from 1 to 20, from 1 to 19, from 1 to 18, from 1 to 17, from 1 to 16, from 1 to 15, from 1 to 14, from 1 to 13, from 1 to 12, from 1 to 11, from 1 to 10, from 1 to 9, from 1 to 8, from 1 to 7, from 1 to 6, from 2 to 21, from 2 to 20, from 2 to 19, from 2 to 18, from 2 to 17, from 2 to 16, from 2 to 15, from 2 to 14, from 2 to 13, from 2 to 12, from 2 to 11, from 2 to 10, from 2 to 9, from 2 to 8, from 2 to 7, from 2 to 6, from 3 to 21, from 3 to 20, from 3 to 19, from 3 to 18, from 3 to 17, from 3 to 16, from 3 to 15, from 3 to 14, from 3 to 13, from 3 to 12, from 3 to 11, from 3 to 10, from 3 to 9, from 3 to 8, from 3 to 7, from 3 to 6, from 4 to 21, from 4 to 20, from 4 to 19, from 4 to 18, from 4 to 17, from 4 to 16, from 4 to 15, from 4 to 14, from 4 to 13, from 4 to 12, from 4 to 11, from 4 to 10, from 4 to 9, from 4 to 8, from 4 to 7, from 5 to 21, from 5 to 20, from 5 to 19, from 5 to 18, from 5 to 17, from 5 to 16, from 5 to 15, from 5 to 14, from 5 to 13, from 5 to 12, from 5 to 11, from 5 to 10, from 5 to 9, from 5 to 8, or from 5 to 7, such as from 6 to 7.

The β-cyclodextrin derivative for use in the dry powder formulation can have a molecular weight in a range from about 1000 g/mol to about 2500 g/mol, such as from about 1500 g/mol to about 2500 g/mol or from about 1000 g/mol to about 2000 g/mol. For example, the β-cyclodextrin derivative for use in the dry powder formulation is HPβCD, MβCD, or SBEβCD, or a combination thereof, and each of HPβCD, MβCD, and SBEβCD has a molecular weight in a range from about 1000 g/mol to about 2500 g/mol, such as from about 1500 g/mol to about 2500 g/mol or from about 1000 g/mol to about 2000 g/mol. The β-cyclodextrin derivative can be an anionic, a cationic, or a nonionic molecule.

1. Exemplary β-Cyclodextrin Derivatives

Exemplary β-cyclodextrin derivatives suitable for use in the dry powder formulation can have structures of Formula I.

where each R is independently a hydrogen, an unsubstituted alkyl group, a substituted alkyl group, an unsubstituted heteroalkyl group, a substituted heteroalkyl group, an aldehyde group, or an acyl group optionally containing an unsubstituted alkyl group, a substituted alkyl group, an unsubstituted heteroalkyl group, a substituted heteroalkyl group, a halogen, an unsubstituted alkenyl group, a substituted alkenyl group, a hydroxyl group, an ether group, an amino group, a carboxylate group, or an amide group, and where the degree of substitution can be any one of the ranges described above, such as from 1 to 21, from 2 to 20, from 3 to 19, from 4 to 18, from 5 to 15, from 2 to 10, from 3 to 9, from 5 to 8, from 6 to 9, or from 6 to 8.

In some aspects of Formula I, the substituents for each substituted R are independently a sulfo group, a hydroxyl group, an unsubstituted alkyl group, a substituted alkyl group, an unsubstituted heteroalkyl group, a substituted heteroalkyl group, a halogen, an unsubstituted alkylene group, a substituted alkenyl group, a substituted alkenyl group, an ether group, an amino group, a carboxylate group, or an amide group.

In some aspects of Formula I, each R is independently a hydrogen, an unsubstituted alkyl group, a substituted alkyl group, or an acyl group optionally containing an unsubstituted alkyl group or a substituted alkyl group, and where the substituents for the substituted alky group are independently a sulfo group, a hydroxyl group, a carboxylate group, an amino group, or an amide group.

In some aspects of Formula I, each R is independently a hydrogen, an unsubstituted alkyl group, a substituted alkyl group, or an acyl group optionally containing an unsubstituted alkyl group or a substituted alkyl group (e.g. a formyl group, an acetyl group, a propionyl group, etc. ) , where the substituents for the substituted alkyl group are independently a sulfo group, a hydroxyl group, a carboxylate group, an amino group, or an amide group, and

where the alkyl group (i.e. unsubstituted alkyl group or substituted alkyl group) is a linear C ₁-C ₃₀ alkyl, a branched C ₄-C ₃₀ alkyl, a cyclic C ₃-C ₃₀ alkyl, a linear C ₁-C ₃₀ alkyl or a branched C ₄-C ₃₀ alkyl, a linear C ₁-C ₃₀ alkyl or a cyclic C ₃-C ₃₀ alkyl, a branched C ₄-C ₃₀ alkyl or a cyclic C ₃-C ₃₀ alkyl, a linear C ₁-C ₂₀ alkyl, a branched C ₄-C ₂₀ alkyl, a cyclic C ₃-C ₂₀ alkyl, a linear C ₁-C ₂₀ alkyl or a branched C ₄-C ₂₀ alkyl, a branched C ₄-C ₂₀ alkyl or a cyclic C ₃-C ₂₀ alkyl, a linear C ₁-C ₂₀ alkyl or a cyclic C ₃-C ₂₀ alkyl, a linear C ₁-C ₁₀ alkyl, a branched C ₄-C ₁₀ alkyl, a cyclic C ₃-C ₁₀ alkyl, a linear C ₁-C ₁₀ alkyl or a branched C ₄-C ₁₀ alkyl, a branched C ₄-C ₁₀ alkyl or a cyclic C ₃-C ₁₀ alkyl, a linear C ₁-C ₁₀ alkyl or a cyclic C ₃-C ₁₀ alkyl, a linear C ₁-C ₆ alkyl, a branched C ₄-C ₆ alkyl, a cyclic C ₃-C ₆ alkyl, a linear C ₁-C ₆ alkyl or a branched C ₄-C ₆ alkyl, a branched C ₄-C ₆ alkyl or a cyclic C ₃-C ₆ alkyl, a linear C ₁-C ₆ alkyl or a cyclic C ₃-C ₆ alkyl, a linear C ₁-C ₅ alkyl, a branched C ₄-C ₅ alkyl, a cyclic C ₃-C ₅ alkyl, a linear C ₁-C ₅ alkyl or a branched C ₄-C ₅ alkyl, a branched C ₄-C ₅ alkyl or a cyclic C ₃-C ₅ alkyl, a linear C ₁-C ₅ alkyl or a cyclic C ₃-C ₅ alkyl, a linear C ₁-C ₄ alkyl, a branched C ₄ alkyl, a cyclic C ₃-C ₄ alkyl, a linear C ₁-C ₄ alkyl or a cyclic C ₃-C ₄ alkyl, such as a linear C ₁-C ₅ alkyl group, a C ₁-C ₄ alkyl group, a C ₁-C ₃ alkyl group, or a C ₁-C ₂ alkyl group (e.g. a methyl group, an ethyl group, a propyl group, or a butyl group) .

In some aspects of Formula I, each R is independently a hydrogen, a methyl group, an ethyl group, a propyl group, a butyl group, a hydroxyalkyl group (e.g. a hydroxymethyl group, a 1-hydroxyethyl group, a 2-hydroxyethyl group, a 1-hydroxypropyl group, a 2-hydroxypropyl group, a 3-hydroxypropyl group, a 1-hydroxybutyl group, a 2-hydroxybutyl group, a 3-hydroxybutyl group, a 4-hydroxybutyl group, etc. ) , a sulfoalkyl group (e.g. a sulfomethyl group, a 1-sulfoethyl group, a 2-sulfoethyl group, a 1-sulfopropyl group, a 2-sulfopropyl group, a 3-sulfopropyl group, a 1-sulfobutyl group, a 2-sulfobutyl group, a 3-sulfobutyl group, a 4-sulfobutyl group, etc. ) , a formyl group, an acetyl group, or a propionyl group, and where the degree of substitution is in a range from 1 to 12, from 2 to 15, from 2 to 12, from 2 to 10, from 4 to 10, from 5 to 8, from 6 to 9, or from 6 to 8.

In some aspects of Formula I, the β-cyclodextrin derivative is HPβCD, MβCD, or SBEβCD, and where the degree of substitution is in a range from 2 to 15, from 2 to 12, from 2 to 10, from 4 to 10, from 5 to 8, from 6 to 9, from 6 to 8, or from 6 to 7, such as

and those described in Albers and Muller, “Cyclodextrin Derivatives in Pharmaceutics, ” Critical Reviews in Therapeutic Drug Carrier Systems, 12 (4) : 311-337 (1995) .

In some aspects, the dry powder formulation can contain two or more β-cyclodextrin derivatives of different species. A different species can be a β-cyclodextrin derivative modified with the same functional group (s) with different degrees of substitution, a β-cyclodextrin derivative modified with different functional group (s) with the same degree of substitution, or a β-cyclodextrin derivatives modified with different functional group (s) and with a different degree of substitution.

For example, the dry powder formulation can contain two or more β-cyclodextrin derivatives, where each of the two or more β-cyclodextrin derivatives is modified with the same functional group with a different degree of substitution from the others. For example, the dry powder formulation can contain two or more β-cyclodextrin derivatives, where each of the two or more β-cyclodextrin derivatives is modified with a different functional group from the others and the degree of substitution is the same or different from the others.

2. Amount of β-Cyclodextrins and Derivatives Thereof

Generally, the amount of the β-cyclodextrin or β-cyclodextrin derivative, or the total amount of the β-cyclodextrin and β-cyclodextrin derivative in the formulation is at least 50 wt%, such as in a range from 50 wt%to 99 wt%, from 50 wt%to 98 wt%, from 50 wt%to 95 wt%, from 50 wt%to 90 wt%, from 50 wt%to 85 wt%, from 50 wt%to 84 wt%, from 50 wt%to 80 wt%, from 50 wt%to 75 wt%, from 50 wt%to 70 wt%, from 55 wt%to 99 wt%, from 55 wt%to 98 wt%, from 55 wt%to 95 wt%, from 55 wt%to 90 wt%, from 55 wt%to 85 wt%, from 55 wt%to 80 wt%, from 55 wt%to 75 wt%, from 55 wt%to 70 wt%, from 60 wt%to 99 wt%, from 60 wt%to 98 wt%, from 60 wt%to 95 wt%, from 60 wt%to 90 wt%, from 60 wt%to 85 wt%, from 50 wt%to 80 wt%, from 60 wt%to 75 wt%, or from 60 wt%to 70 wt%of the total weight of the antibody and/or antibody derivative and the β-cyclodextrin and/or β-cyclodextrin derivative. For example, the amount of the β-cyclodextrin or β-cyclodextrin derivative, or the total amount of the β-cyclodextrin and β-cyclodextrin derivative in the formulation can be in a range from 50 wt%to 95 wt%of the total weight of the antibody and/or antibody derivative and the β-cyclodextrin and/or β-cyclodextrin derivative. The term “total amount of the β-cyclodextrin and β-cyclodextrin derivative” refers to the total weight of the β- cyclodextrin and β-cyclodextrin derivative relative to the total weight of the antibody and/or antibody derivative and the β-cyclodextrin and β-cyclodextrin derivative in the dry powder formulation.

In some aspects, the dry powder formulation contains HPβCD with a single degree of substitution or different degrees of substitution and the amount of HPβCD with a single degree of substitution or the total amount of HPβCD with different degrees of substitution is at least 50 wt%, from 50 wt%to 99 wt%, from 50 wt%to 98 wt%, from 50 wt%to 95 wt%, from 50 wt%to 90 wt%, from 50 wt%to 85 wt%, from 50 wt%to 84 wt%, from 50 wt%to 80 wt%, from 50 wt%to 75 wt%, from 50 wt%to 70 wt%of the total weight of the antibody and/or antibody derivative and the HPβCD. When the dry powder formulation contains HPβCD of different degrees of substitution, the amount of each species can be in a suitable range to provide the above-described ranges.

In some aspects, the dry powder formulation contains MβCD with a single degree of substitution or different degrees of substitution and the amount of MβCD with a single degree of substitution or the total amount of MβCD with different degrees of substitution is at least 50 wt%, from 50 wt%to 99 wt%, from 50 wt%to 98 wt%, from 50 wt%to 95 wt%, from 50 wt%to 90 wt%, from 50 wt%to 85 wt%, from 50 wt%to 84 wt%, from 50 wt%to 80 wt%, from 50 wt%to 75 wt%, from 50 wt%to 70 wt%of the total weight of the antibody and/or antibody derivative and the MβCD. When the dry powder formulation contains MβCD of different degrees of substitution, the amount of each species can be in a suitable range to provide the above-described ranges.

In some aspects, the dry powder formulation contains SBEβCD with a single degree of substitution or different degrees of substitution and the amount of SBEβCD with a single degree of substitution or the total amount of SBEβCD with different degrees of substitution is at least 50 wt%, from 50 wt%to 99 wt%, from 50 wt%to 98 wt%, from 50 wt%to 95 wt%, from 50 wt%to 90 wt%, from 50 wt%to 85 wt%, from 50 wt%to 84 wt%, from 50 wt%to 80 wt%, from 50 wt%to 75 wt%, from 50 wt%to 70 wt%of the total weight of the antibody and/or antibody derivative and the SBEβCD. When the dry powder formulation contains SBEβCD of different degrees of substitution, the amount of each species can be in a suitable range to provide the above-described ranges.

C. Pharmaceutically Acceptable Excipients

The dry powder formulation may contain a pharmaceutically acceptable excipient, optionally more than one pharmaceutically acceptable excipient. In some aspects, the dry powder formulation does not contain any additional pharmaceutically acceptable excipients.

Exemplary pharmaceutically acceptable excipients that can be used in the dry powder formulation include, but are not limited to, amino acids, peptides, lipids (e.g. fatty acids, fatty acid esters, steroids) , proteins, chelating agents (e.g. EDTA) , salts, taste masking agents, cations, non-biological or biological polymers, and additional sugars, and a combination thereof. Examples of suitable pharmaceutically acceptable excipients that can be used in the dry powder formulation are described in Kibbe, et al., “Handbook of Pharmaceutical Excipients, ” 3 ^rd edition, 2000. In some aspects, the dry powder formulation does not contain any additional sugars.

Examples of suitable amino acids that may be include in the dry powder formulation include, but are not limited to, alanine, glycine, arginine, histidine, glutamate, asparagine, cysteine, leucine, lysine, isoleucine, valine, tryptophan, methionine, proline, phenylalanine, tyrosine, citrulline, L-aspartyl-L-phenylalanine-methyl ester (aspartame) , trimethylammonium acetate (betaine) , etc.

Examples of suitable protein excipients that may be included in the dry powder formulation include, but are not limited to, albumin (of human or recombinant origin) , gelatin, casein, hemoglobin, etc.

Examples of suitable polymers that may be included in the dry powder formulation include, but are not limited to, polyvinyl pyrrolidone, derivatized celluloses, such as hydroxymethyl, hydroxyethyl, hydroxypropyl ethylcellulose, polyethylene glycol, and polypropylene glycol.

Examples of additional sugars that may be included in the dry powder formulation can be a mono-, di-, oligo-, or polysaccharide, or a combination thereof. Examples of monosaccharides are fructose, maltose, galactose, glucose, D-mannose, sorbose, and the like. Examples of disaccharides are lactose, saccharose, trehalose, cellobiose, etc. Examples of sugar alcohols are mannitol, xylitol, maltitol, galactitol, arabinitol, adonitol, lactitol, sorbitol (glucitol) , pyranosylsorbitol, inositol, myoinositol, etc. Examples of oligosaccharide are other types of cyclodextrin (e.g. α-cyclodextrin, γ-cyclodextrin, and their derivatives thereof) , maltodextrin, and pectins.

In some aspects, the dry powder formulation does not contain any additional sugars, such as mannitol, trehalose, 1, 4 O-linked saccharose or 1, 4 O-linked saccharose derivatives, or dexran, or a combination thereof. In some aspects, the dry powder formulation does not contain mannitol or trehalose, or a combination of mannitol and trehalose. In some aspects, the dry powder formulation does not contain 1, 4 O-linked saccharose or 1, 4 O-linked saccharose derivatives, or a combination thereof. In some aspects, the dry powder formulation does not contain dexran. In some aspects, the dry powder formulation does not contain any one of mannitol, trehalose, 1, 4 O-linked saccharose, 1, 4 O-linked saccharose derivatives, and dexran.

Examples of salts that may be included in the dry powder formulation are inorganic salts such as chloride, sulphate, phosphate, diphosphate, hydrobromide, and nitrate salts and organic salts such as malate, maleate, fumarate, tartrate. Succinate, ethylsuccinate, citrate, acetate, lactate, methaneSulphonate, benzoate, ascorbate, para-toluenesulphonate, palmoate, salicylate, stearate, estolate, gluceptate, and lactobionate salts. The salts may simultaneously contain pharmaceutically acceptable cations, such as sodium, potassium, calcium, aluminium, lithium, and ammonium.

The amount of the pharmaceutical acceptable excipient, or the total amount of two or more pharmaceutically acceptable excipients may be in a range from 0.1 wt%to 20 wt%, from, from 0.1 wt%to 15 wt%, from 1 wt%to 12 wt%, from 1 wt%to 10 wt%, from 1 wt%and 15 wt%, from 2 wt%to 20 wt%, from 2 wt%to 15 wt%, from 2 wt%to 10 wt%, from 3 wt%to 20 wt%, from 3 wt%to 15 wt%, or from 3 wt%to 10 wt%of the dry powder formulation. The term “total amount of the two or more pharmaceutically acceptable excipients” refers to the total weight of the two or more pharmaceutically acceptable excipients relative to the total weight of the dry powder formulation.

D. Additional Active Agents

The dry powder formulation may contain an additional active agent, optionally more than one additional active agent. The additional active agents that can be included in the dry powder formulation may be therapeutic, nutritional, prophylactic, or diagnostic agents, or a combination thereof, such as anti-inflammatory agents, anti-viral agents, and chemotherapeutic agents.

Examples of anti-inflammatory and chemotherapeutic agents that can be included in the dry powder formulation include, but are not limited to, immunosuppressants cyclosporin (including cyclosporin A, cyclosporin B, cyclosporin C, cyclosporin D, cyclosporin E, cyclosporin F, cyclosporin G, cyclosporin H, cyclosporin I) , methotrexate, azathioprine, steroids, corticosteroid (e.g., fluticasone, budesonide, mometasone, beclomethasone, ciclesonide, triamcinolone acetonide or dexamethasone, cyclophosphamide) , non-steroidal drug (e.g., ketorolac or diclofenac) , tacrolimus (FR-900506, FK506, or fujimycin) , ridaforolimus, sirolimus (rapamycin) , and their derivatives, such as temsirolimus and everolimus.

Antiviral agents that can be included in the dry powder formulation are known, such as Ribavirin and Favipiravir and those described in Erik De Clercq, Medmicro, Chapter 52 (2000) .

The amount of the additional active agents needed will vary from subject to subject according to their need.

E. Delivery Systems

Systems for delivering the dry powder formulations are disclosed. Generally, the system includes an inhaler and the dry powder formulation. The dry powder formulation may be in a unit dosage form that contains a single unit does of the antibody and/or antibody derivative. Alternatively, the dry powder formulation may contain multiple doses of the antibody and/or antibody derivative.

Typically, suitable inhalers that can be used with the dry powder formulation are dry powder inhalers ( “DPIs” ) . The dry powder formulation may be pre-packaged in a capsule or a replaceable set and then loaded in the inhaler. Alternatively, the dry powder formulation may be directly loaded in the reservoir of the inhaler.

1. Unit Dosage Form

In some aspects, the dry powder formulation prepackaged in a capsule or replaceable set and then loaded in the inhaler is in a unit dosage form, i.e. containing a single unit does of the antibody and/or antibody derivatives to be delivered to the lung (s) of a subject.

A suitable unit dose of the antibody and/or antibody derivatives in the unit dosage form of the dry powder formulation can range from about 0.1 mg to about 50 mg, from about 0.5 to about 50 mg, or from about 1 to about 50 mg. For example, a suitable unit dose of the antibody and/or antibody derivatives prepackaged in a capsule is in a range from about 0.1 mg to about 50 mg, from about 0.5 to about 50 mg, or from about 1 to about 50 mg.

Alternatively, the dry powder formulation can contain multiple unit doses of the antibody and/or antibody derivative that can be loaded in the reservoir of a metered inhaler such that a single dose may be delivered to the subject per administration.

Treatment regimens utilizing antibodies and/or antibody derivative can include administration of from about 1 mg to about 3000 mg of the antibodies and/or antibody derivatives per kilogram body weight of the recipient per day in a single unit dose or multiple unit doses (such as two, three, four, five, or six or more unit doses at appropriate intervals throughout the day) .

2. Inhalers

a. Dry Powder Inhalers

DPIs are breath actuated, thus the problem of coordinated inspiration with actuation, as in the case of pMDIs, is avoided. Exemplary dry powder inhaler types include single capsule unit dose in an inhaler, single disposable unit dose in the inhaler, multiple unit doses in a replaceable set in an inhaler, and multiple unit doses in a reservoir in an inhaler.

The dry powder inhaler may carry one or more capsules, each capsule containing a unit dose of the dry powder formulation in unit dosage form. For example, the DPI may be a single unit dose DPI containing one capsule loaded with one unit dose of the dry powder formulation in unit dosage form, where the capsule may be repeatedly loaded or disposable. Alternatively, the DPI may be a multiple unit doses DPI containing two or more capsules where each of the two or more capsules is loaded with one unit dose of the dry powder formulation in unit dosage form and the capsules may be repeatedly loaded or disposable.

The dry powder inhalers may contain a replaceable set with multiple doses of the dry powder formulation in unit dosage form, such as a replaceable blister package, cartridge, strip, or wheel. For example, the DPI may be a multiple unit doses DPI containing a foil-foil blister prepackaged with several unit doses of the dry powder formulation in unit dosage form, where each unit dose is spatially separated from the other unit doses (i.e. discrete unit doses) .

The dry powder inhalers may contain a reservoir with a capacity for holding multiple unit doses of the dry powder formulation in unit dosage form. For example, the DPI may be a multiple unit doses DPI containing a pre-metered cartridge containing multiple unit doses of the dry powder formulation in unit dosage form. In some aspects, the DPI may be a multiple unit doses DPI containing a dry powder formulation that contains multiple unit doses of the antibody and/or antibody derivative in a reservoir. For these DPIs, it may include a pre-metered valve such that one unit dose can be delivered to the subject per administration.

Suitable DPIs for use with the dry powder formulations may be those described in U.S. Patent No. 7,305,986 and U.S. Patent Application Publication No. 2004/0182387. Exemplary commercially available multi-dose DPIs suitable for use with the dry powder formulation include, but are not limited to,

(Glaxo Group Limited Corp, Brentford, Middlesex United Kingdom) ,

(GSK, also described in WO 05/14089) ,

(Vectura, also described in WO 05/37353) ,

(Valois, also described in WO 03/77979) and

(Merck, also described in WO 93/00123, WO 94/14492, and WO 97/30743) . Exemplary commercially available single dose DPIs suitable for use with the dry powder formulation include, but are not limited to,

(Novartis Ag Corporation Switzerland, Basel, Switzerland) and

(Novartis Ag Corporation Switzerland, Basel, Switzerland) . Other exemplary DPIs suitable for use with the dry powder formulation include, but are not limited to high-resistance osmohale ^TM inhaler,

(Boehringer Ingelheim Pharma KG, Ingelheim am Rhein, Fed Rep Germany) ,

(Direct-Haler A/SCorp Denmark, Odense Sv Denmark) ,

(Glaxo Group Limited Corp, Brentford, Middlesex United Kingdom) ,

(Astra Aktiebolag Corp., Sodertalie Sweden) ,

(Orion Corporation, Espoo Finland) , and Nexthaler (Lavorini et al. Multidisciplinary Respiratory Medicine, 12: 11 (2017) ) .

F. Particle Properties

The aerosolisation performance of the dry powder formulation can be evaluated by the properties of particles of the dry powder formulation, such as mass median aerodynamic diameter ( “MMAD” ) , geometric standard deviation ( “GSD” ) , fine particle fraction ( “FPF” ) , and emitted fraction ( “EF” ) . The term “fine particle fraction” or “FPF” refers to the weight percentage of particles with MMAD less than 5 μm relative to the total weight of particles in the formulation. The term “EF” refers to the fraction of powder that exits the inhale after a dispersion event, expressed as the ratio of the dose delivered by an inhaler to the nominal dose, i.e. the mass of powder per unit dose placed into the inhaler prior to dispersion. Additional particle properties, such as solid-state form (crystalline/amorphous) and thermal behavior (dehydration, glass transition, degradation, etc. ) , can be determined by differential scanning calorimetry (DSC) .

MMAD, GSD, and FPF of the particles in the dry powder formulation may be determined using methods known in the art. These include dynamic light scattering, aerodynamic particle sizing, light microscopy, and/or cryo-transmission electron microscopy ( “cryo-TEM” ) . For example, MMAD may be measure by dispersing powder by a high-resistance osmohale ^TM inhaler and the powder is evaluated by a Next Generation Impactor (NGI) operated at 60 L/min for 4 s. For example, MMAD may be measure by dispersing powder by a

and the powder is evaluated by a Next Generation Impactor (NGI) operated at 90 L/min for 2.7 s.

Generally, the particles of the dry powder formulation have favorable aerodynamic properties for effective lung deposition. For example, the particles of the dry powder formulation have a MMAD < 5 μm, < 4 μm, < 3.5 μm, < 3 μm, < 2.5 μm, or < 2 μm; a GSD < 4.5, < 4, < 3.5, < 3, < 2.5, or < 2; a FPF > 40%, > 45%, > 50%, > 55%, or > 60%in cascade impactor study; and/or an EF > 50%, > 55%, > 60%, > 65%, > 70%, > 75%, > 80%, > 85%, > 90%, or > 95%.

In some aspects, the particles of the dry powder formulation are spray-dried particles and the particles have a MMAD < 2.5 μm or < 2 μm; a GSD < 3, < 2.5, or < 2; a FPF > 40%, > 45%, > 50%, > 55%, or > 60%in cascade impactor study; and an EF > 50%, > 55%, > 60%, > 65%, or > 70%.

In some aspects, the particles of the dry powder formulation are spray-freeze-dried particles and the particles have a MMAD < 2 μm or < 1.5 μm; a GSD < 4.5 or < 4; a FPF > 60%, > 65%, > 70%, or > 75%in cascade impactor study; and an EF > 90%or > 95%.

The antibody integrity in the particles of the dry powder formulation following manufacturing process, such as spray drying or spray freeze drying, can be assessed by SDS-PAGE and/or SEC. Generally, the antibody and/or antibody derivative formulated in the dry powder formulation do not show fragments following manufacture (as determined by SDA-PAGE analysis) and/or shows an aggregation percentage of less than about 1% (as determined by SEC-HPLC) immediately following manufacture and/or an aggregation percentage of less than 1%or less than 0.5% (as determined by SEC-HPLC) 4 weeks post-drying, 8 weeks post-drying, 10 weeks post-drying, 12 weeks post-drying, or 16 weeks post-drying.

The antigen-binding affinity and inhibitory potency of the antibody and/or antibody derivative of the dry powder formulation following manufacturing process, such as spray drying or spray freeze drying, can be assessed by ELISA and cellular assays, respectively. Generally, the antibody and/or antibody derivative formulated in the dry powder formulation have a binding activity and/or an inhibitory potency that are comparable to those of unprocessed and unformulated antibody and/or antibody derivative. That is, the change of LogEC ₅₀ and/or LogIC ₅₀ value of the antibody and/or antibody derivative against the same binding target following formulation is less than about 30%compared to those prior to processing and formulation.

G. Exemplary Formulations

An exemplary dry powder formulation contains spray-dried or spray-freeze-dried particles containing a β-cyclodextrin derivative and a Fab, where the weight ratio of the β-cyclodextrin derivative to the Fab is in a range from 1: 1 to 49: 1, from 1: 1 to 19: 1, from 1: 1 to 9: 1, or from 1: 1 to 5: 1.

For these dry powder formulations, the β-cyclodextrin derivative can be a HPβCD, MβCD, or SBEβCD, and where the degree of substitution is in a range from 2 to 15, from 2 to 12, from 2 to 10, from 4 to 10, from 5 to 8, from 6 to 9, from 6 to 8, or from 6 to 7.

For these dry powder formulations, the Fab do not show fragments following manufacture (as determined by SDA-PAGE analysis) and shows an aggregation percentage of less than about 1% (as determined by SEC-HPLC) following manufacture and less than 1%or less than 0.5% (as determined by SEC-HPLC) up to 16 weeks post-drying.

Additionally, the Fab of these dry powder formulations has comparable binding activity against the target antigen and comparable inhibitory potency against TF-1 cells. The term “comparable antibody binding activity against the target antigen” means that the change of LogEC ₅₀ value of the Fab formulated in these formulations against a particular target antigen is less than about 30%compared to the LogEC ₅₀ value of the same but unformulated Fab. The term “comparable inhibitory potency against TF-1 cells” means that the change of LogIC ₅₀ value of the Fab formulated in these formulations against TF-1 cells is less than about 30%compared to the LogIC ₅₀ value of the same but unformulated Fab.

These dry powder formulations may be in a unit dosage form prepackaged in a capsule or a replaceable set and then loaded in a dry powder inhaler. Alternatively, these dry powder formulation may contain multiple doses of antibody and/or antibody derivative and can be loaded in the reservoir of a dry powder inhaler.

The particles of these dry powder formulations can have a MMAD < 5 μm, < 4 μm, < 3.5 μm, < 3 μm, < 2.5 μm, or < 2 μm; a GSD < 4.5, < 4, < 3.5, < 3, < 2.5, or < 2; a FPF > 40%, > 45%, > 50%, > 55%, or > 60%in cascade impactor study; and an EF > 50%, > 55%, > 60%, > 65%, > 70%, > 75%, > 80%, > 85%, > 90%, or > 95%.

More specific exemplary dry powder formulations are described in the Examples.

III. METHODS OF MAKING THE DRY POWDER FORMULATIONS

Methods of making the dry powder formulations are disclosed. Generally, the method includes (i) mixing an antibody and/or an antibody derivative and a β-cyclodextrin and/or a β-cyclodextrin derivative, and optionally a pharmaceutically acceptable excipient and/or an additional active agent, in a solvent to form a liquid feed; and (ii) spray-drying or spray-freeze drying the liquid feed to form particles containing the antibody and/or antibody derivative and the β-cyclodextrin and/or β-cyclodextrin derivative, and optionally the pharmaceutically acceptable excipient and/or the additional active agent.

Typically, the production yield of particles following step (ii) is at least about 50 wt%, at least 55 wt%, at least 60 wt%, at least 65 wt%, at least 70 wt%, at least 75 wt%, at least 80 wt%, at least 85 wt%, or at least 90 wt%, or in a range from about 50 wt%to about 95 wt%, from about 55 wt%to about 95 wt%, from about 55 wt%to about 90 wt%, from about 60 wt%to about 95 wt%, from about 65 wt%to about 95 wt%, from about 70 wt%to about 95 wt%, or from about 75 wt%to about 95 wt%. The yield of particles is the weight of particles produced relative to the total weight of ingredients in the liquid feed. The “total weight of ingredients” refers to the total weight of the antibody and/or antibody derivative and the β-cyclodextrin and/or β-cyclodextrin derivative, and optionally also includes the weight of the pharmaceutically acceptable excipient, additional active agent, and/or salts in the solvent.

The particles formed in step (ii) by spray-drying or spray-freeze drying have favorable aerodynamic properties for effective lung deposition and retained antibody structural integrity and binding activity, as described above.

A. Forming Liquid Feed

Generally, prior to spray-drying or spray-freeze-drying, a liquid feed is prepared by mixing the antibody and/or antibody derivative and the β-cyclodextrin and/or β-cyclodextrin derivative, and optionally a pharmaceutically acceptable excipient and/or an additional active agent in a solvent. Typically, the solvent is an aqueous solvent. In some aspects, the liquid feed only contains the antibody and/or antibody derivative and the β-cyclodextrin and/or β-cyclodextrin derivative dissolved in the aqueous solvent.

Typically, the antibody and/or antibody derivative and the β-cyclodextrin and/or β-cyclodextrin derivative are soluble in the aqueous solvent. For example, the antibody and/or antibody derivative and the β-cyclodextrin and/or β-cyclodextrin derivative have a solubility of at least about 1.5 g/100 mL of the aqueous solvent at room temperature (R. T. ) , i.e. a temperature between about 20 ℃ and about 25 ℃ under atmospheric pressure. For example, the antibody and/or antibody derivative can have a solubility of > 1.5 g/100 mL of the aqueous solvent at room temperature, and/or the β-cyclodextrin and/or β-cyclodextrin derivative have a solubility of at least about 45 g/100mL of the aqueous solvent at room temperature. When a pharmaceutically acceptable excipient and/or an additional active agent are mixed in the solvent, they may be soluble or suspended in the solvent. For example, the pharmaceutically acceptable excipient and/or additional active agent can be at least as soluble as the antibody and/or antibody derivative in the solvent or insoluble in the solvent.

Suitable solvents for preparing the liquid feed include, but are not limited to, water and aqueous buffers, such as sodium phosphate, potassium phosphate, sodium acetate, potassium acetate, sodium citrate, potassium citrate, sodium succinate, potassium succinate, and ammonium bicarbonate and carbonate, and a combination thereof. Generally, aqueous buffers that are suitable for preparing the liquid feed have a molarity in a range from about 1 mM to about 2 M, from about 2 mM to about 1 M, from about 10 mM to about 0.5 M, or from 50 to 200 mM and have a pH in a range from about 1 to about 10, from about 3 to about 8, or from about 5 to about 7.

The concentration of solute in the solvent, i.e. the total amount of antibody and/or antibody derivative and the β-cyclodextrin and/or β-cyclodextrin derivative in the solvent, is in a range from about 0.5% (w/v) to about 10% (w/v) , from about 1% (w/v) to about 10% (w/v) , from about 1% (w/v) to about 8% (w/v) , from about 1% (w/v) to about 6% (w/v) , from about 1% (w/v) to about 5% (w/v) , or from about 2% (w/v) to about 5% (w/v) , such as about 2% (w/v) for spray-drying or about 5% (w/v) for spray-freeze-drying.

The liquid feed prepared in step (i) is then subject to spray-drying or spray-freeze drying process.

B. Spray-Drying

Spray-drying is a process of producing a dry powder containing particles from a liquid or a dispersion in a liquid by rapidly drying with a hot gas. An exemplary set up is depicted in FIG. 1A. This process can rapidly produce particles for inhalation (i.e., on the order of seconds for laboratory-scale spray dryers and minutes on industrial scale) with controlled particle size, morphology, density, and surface composition.

Generally, in step (ii) , the liquid feed prepared in step (i) may be spray-dried to form the dry powder formulation disclosed herein. The spray-drying process may be carried out using conventional equipment used to prepare spray dried particles for use in pharmaceuticals that are administered by inhalation. Exemplary commercially available spray-dryers include those manufactured by Buchi Ltd., Niro Corp, Bichi, Niro Yamato, Okawara, Kakoki.

Typically, the liquid feed is sprayed into a current of warm filtered air that evaporates the solvent and conveys the dried product to a collector. The spent air is then exhausted with the solvent. Operating conditions of the spray-dryer such as inlet temperature, feed rate, aspirator rate, flow rate of the drying air, and nozzle configuration can be adjusted in order to produce the required particle size, moisture content, and production yield of the resulting dry particles. The selection of appropriate apparatus and processing conditions are within the purview of a skilled artisan in view of the teachings herein.

Exemplary settings for the spray-drying step are as follows: an air inlet temperature between about 60℃ and about 220℃, between about 80℃ and about 220℃, between about 60℃ and about 200℃, between about 60℃ and about 180℃, between about 80℃ and about 200℃, between about 80℃ and about 180℃, between about 80℃ and 150℃, or between about 90℃ and 120℃; an air outlet temperature between about 40℃ to about 120℃, between about 50℃ and 100℃, or between about 50℃ and 80℃; a feed rate between about 0.1 mL/min to about 30 mL/min, between about 0.1 mL/min to about 25 mL/min, between about 0.1 mL/min to about 20 mL/min, between about 0.1 mL/min to about 15 mL/min, between about 0.5 mL/min to about 30 mL/min, between about 0.5 mL/min to about 25 mL/min, between about 0.5 mL/min to about 20 mL/min, between about 0.5 mL/min to about 15 mL/min, between about 1 mL/min to about 30 mL/min, between about 1 mL/min to about 25 mL/min, between about 1 mL/min to about 20 mL/min, between about 1 mL/min to about 15 mL/min, between about 1.5 mL/min to about 15 mL/min, between about 1 mL/min to about 10 mL/min, or between about 1 mL/min to about 5 mL/min; an air flow rate between about 100 L/h to about 1000 L/h, between about 200 L/h to about 1000 L/h, between about 300 L/h to about 1000 L/h, between about 400 L/h to about 1000 L/h, between about 500 L/h to about 1000 L/h, between about 500 L/h to about 900 L/h, between about 600 L/h to about 800 L/h, or between about 650 L/h to about 800 L/h; and an aspiration rate of about 100%. These settings will, however, vary depending on the type of equipment used, and the nature of the solvent system employed. In any event, the use of these and similar methods allow formation of particles with diameters appropriate for aerosol deposition into the lung.

An exemplary spray-drying process uses a Büchi B-290 spray drier and have the following settings: an air inlet temperature between about 90℃ and 120℃, such as about 100℃; an air outlet temperature between about 50℃ and 80℃, such as between 62℃ and 67℃; a feed rate between about 0.1 mL/min and 30 mL/min, such as about 0.9 mL/min, an air flow rate between about 650 L/h to about 800 L/h, such as about 742 L/h, and an aspiration rate of 100%, where the production yield of particles following spray drying is at least 55 wt%, such as between about 55 wt%and about 90 wt%.

C. Spray-Freeze-Drying

Spray-freeze-drying is a process similar to spray drying in that a liquid feed containing the antibody and/or antibody derivative and the β-cyclodextrin and/or β-cyclodextrin derivative, and optionally the pharmaceutically acceptable excipient and/or additional active agent is introduced via a nozzle (e.g. a two-fluid nozzle) , or a spinning disk into a cold fluid to atomize the liquid feed to form fine droplets. The cold fluid, either a liquid or a gas, is at a temperature below the freezing point of the solvent of the liquid feed. Spraying the liquid feed into the cold fluid results in rapid freezing of the atomized droplets to form solid particles. The particles are collected, and then the solvent is removed, generally through sublimation (lyophilization) in a vacuum. Any known technique, such as those described by Mumenthaler et al, Int. J. Pharmaceutics (1991) 72: 97–110 (1991) and Maa et al., Phar. Res., 16: 249 (1999) , may be used to carry out the spray-freeze-drying step. Exemplary commercially available freeze-dryers that can be used in the spray-freeze-drying process include those manufactured by Labconco Corporation, Biolab Scientific, and AAPPTec.

For example, the spray-freeze-drying process is performed in a manner similar to spray-drying, except that instead of spraying into hot air or gas, the liquid feed is sprayed into a cold liquid or cold gas to form liquid fine droplets. An exemplary set up is depicted in FIG. 1B. Generally, the liquid feed is atomized using known technique, for example, via a two-fluid nozzle or ultrasonic nozzle using filtered pressurized air, into the cold fluid. The cold fluid may be a liquid such as liquid nitrogen, liquid argon, or any other gas that results in the immediate freezing of the atomized droplets of the liquid feed. The cold fluid can have a temperature in a range from about -200℃ to -100℃, from about -200℃ to about -80℃, such as about -200℃ (liquid nitrogen at -196℃) . In some aspects, the cold liquid may be under stirring as the atomization process occurs.

The atomization conditions, including atomization air flow rate, liquid flow rate (i.e. the feed rate using a syringe pump) , feed rate and nozzle configuration, can be controlled as described above to produce liquid droplets having a suitable size.

The frozen droplets of the liquid feed are then freeze dried to remove frozen water, leaving particles containing the antibody and/or antibody derivative and the β-cyclodextrin and/or β-cyclodextrin derivative, and optionally the pharmaceutically acceptable excipient and/or additional active agent. This may be done using techniques known for lyophilization, i.e. freezing as a cake rather than as droplets. A vacuum can be applied during the second drying step. For example, the frozen droplets are freeze dried by a two-stage vacuum drying (i.e. primary drying stage and secondary drying stage) optionally under a pressure in a range from about 20 mT to about 500 mT (i.e. about 0.02666 mBar to about 0.6666 mBar) . The primary drying stage may be performed at a temperature in a range from about -50℃ to 0℃, from about -40℃ to 0℃, or from about -40℃ to -10℃, such as -25℃, for a period from about 4 hours to about 24 hours. Frozen water is removed by ice sublimation. In the secondary drying stage, drying is normally performed at a temperature in a range from about 5℃ to 50℃, from about 10℃ to about 40℃, from about 10℃ to about 30℃, such as about 20℃ at a pressure of less than 100 mT or less than 0.15 mbar, such as from about 1 mT to about 100 mT, from about 5 mT to about 100 mT, or from about 0.001 mbar to about 0.15 mbar, from about 0.01 mbar to about 0.1 mbar, or from about 0.005 mbar to about 0.5 mbar, for a period from about 5 hours to about 48 hours. The specific spray-freeze-drying conditions used may be adjusted according to the desired properties of the particles to be produced. The resulting particles can then be collected using conventional techniques and optionally with bulking agents.

An exemplary spray-freeze-drying process uses a Labconco freeze drier with a two fluid nozzle and have the following settings: an air flow rate between about 600 L/h to about 750 L/h, such as about 670 L/h; a primary drying temperature in a range from about -40℃ to -10℃, such as -25℃; a secondary drying temperature in a range from about 10℃ to 40℃, such as 20℃; a feed rate between about 0.1 mL/min and 30 mL/min, such as about 2 mL/min; an air flow rate between about 600 L/h to about 800 L/h, such as about 670 L/h; and a drying pressure between about 0.001 mbar to about 0.15 mbar, such as about 0.012 mbar, where the production yield of particles following spray drying is at least 75 wt%, such as between about 75 wt%and about 95 wt%.

IV. Methods of Using the Dry Powder Formulations

A. Preventing, Treating, or Ameliorating Symptom (s) Associated with a Respiratory Disease

Methods of using the dry powder formulation or a delivery system containing the dry powder formulation loaded in an inhaler for preventing, treating, or ameliorating symptoms associated with a respiratory disease in a subject are disclosed.

Generally, the method includes (i) administering to the subject the dry powder formulation. Typically, following step (i) , a unit dose of the antibody and/or antibody derivative is delivered to the lung (s) of the subject to prevent, treat, or ameliorate one or more symptoms associated with the respiratory disease of the subject. The dry powder formulation can be administered using an inhaler, such as a dry powder inhaler, by a medical professional or the subject being treated (i.e. self-administration) .

1. Subject being Treated

The subject being treated using the disclosed method can be a mammal. The subject being treated typically has or at the risk of having a respiratory disease, such as an obstructive or inflammatory airways disease or lung cancer. For example, the subject to be treated has asthma, acute/adult respiratory distress syndrome (ARDS) , chronic obstructive pulmonary or airways disease (COPD or COAD) , bronchiectasis, respiratory infections, bronchitis (e.g., acute, arachidic, catarrhal, croupus, chronic or phthinoid bronchitis) or dyspnea associated therewith, emphysema, exacerbation of airways hyperreactivity consequent to other drug therapy (e.g. other inhaled drug therapy) , or pneumoconiosis (e.g. aluminosis, anthracosis, asbestosis, chalicosis, ptilosis, siderosis, silicosis, tabacosis, and byssinosis) , or a combination thereof.

2. Administered Dose

Typically, the administration step (i) is performed to deliver a unit dose of the antibody and/or antibody derivative to the lung (s) of the subject to prevent, treat, or ameliorate symptoms associated with any one or more of the above-described respiratory diseases of the subject.

Suitable unit dose of the antibody and/or antibody derivative in a dry powder formulation can be in a range from about 1 mg to about 3000 mg, from about 1 mg to about 1500 mg, from about 10 mg to about 1500 mg, from about 10 mg to about 1000 mg, from about 20 mg to about 1000 mg, from about 50 mg to about 1500 mg, from about 50 mg to about 1000 mg, or from about 20 mg to about 500 mg.

Step (i) may be repeated to deliver multiple unit doses of the antibody and/or antibody derivative to the subject, as described below.

3. Optional Steps

In addition to step (i) administering the dry powder formulation to the subject described above, the method may include one or more additional steps. The additional steps can occur prior to step (i) and/or subsequent to step (i) .

a. Loading the Dry Powder Formulation into an Inhaler

The method may include a step of loading the dry powder formulation into the inhaler prior to step (i) .

For example, a user, such as a medical professional or the subject being treated, can load the dry powder formulation into the reservoir of an inhaler for delivering the dry powder formulation to the subject. The dry powder formulation loaded in the reservoir of the inhaler may contain one unit dose of the antibody and/or antibody derivative (i.e. dry powder formulation in unit dosage form) or multiple unit doses of the antibody and/or antibody derivative. When the dry formulation containing multiple unit doses of the antibody and/or antibody derivative is loaded in the reservoir of the inhaler, a metered valve is typically included in the inhaler such that each administration delivers one unit dose of the antibody and/or antibody derivative.

Optionally, a capsule or replaceable set prepackaged with the dry powder formulation may be provided. Typically, the dry powder formulation prepackaged in the capsule or replaceable set is in a unit dosage form, i.e. contain a unit dose of the antibody and/or antibody derivative. For example, the capsule is prepackaged with one dry powder formulation in unit dosage form. The user can load the one prepackaged capsule or two or more of the prepackaged capsules in the inhaler.

For example, the replaceable set is a foil-foil blister prepackaged with several unit doses of the dry powder formulations in unit dosage form, where each unit dose is spatially separated from the other unit doses (i.e. discrete unit doses) . The user can load the prepackaged foil-foil blister in the inhaler and optionally replace with another prepackaged foil-foil blister after all doses are delivered to the subject.

b. Repeating Administration of Formulation

The method may include a step of repeating step (i) administering to the subject the dry powder formulation to deliver multiple unit doses of the antibody and/or antibody derivative to the subject.

Step (i) may be repeated at least one time, at least two times, at least three times, at least five times, at least ten times, at least twenty times, up to thirty times, or more than thirty times. For example, the step (i) is repeated one time, two times, three times, five times, ten times, fifteen times, twenty times, or thirty times.

The period for repeated administration of the dry powder formulation can be between one day and 6 months, between one day and 3 months, between one and thirty days, between one and ten days, between one and three days, between one and two days, or during one day.

For example, the step (i) is repeated one time, two times, three times, five times, ten times, fifteen times, twenty times, or thirty times or more for a period between one day and 6 months, between one day and 3 months, between one and thirty days, between one and ten days, between one and three days, between one and two days, or during one day.

The repeating step may be performed consecutively following the first administration. For example, the administration is repeated within 10 minutes, within 8 minutes, within 5 minutes, within 3 minutes, within 2 minutes, within 1 minute, or within 30 seconds following the previous administration.

Optionally, the repeating step is performed regularly at a different time. For example, the administration may be performed at a frequency, such as every hour, every 2 hours, every 5 hours, every 8 hours, every day, every 2 days, every 3 days, every 5 days, every 7 days, every 10 days, every two weeks, or every month. For example, the step (i) is repeated every hour, every 2 hours, every 5 hours, every 8 hours, every day, every 2 days, every 3 days, every 5 days, every 7 days, every 10 days, every two weeks, or every month for a period between one day and 6 months, between one day and 3 months, between one and thirty days, between one and ten days, between one and three days, between one and two days, or during one day.

Alternatively, the administration may be repeated irregularly, for example, repeating the administration 1 day after the first administration, then 2 days after the second administration, then 5 days after the third administration, then 7 day after the fourth administration, and then 30 days after the fifth administration. The time interval between administrations are determined based on the patient’s needs.

The disclosed compositions and methods can be further understood through the following numbered paragraphs.

1. A dry powder formulation for inhalation comprising particles comprising an antibody or an antibody derivative, or a combination thereof; and a β-cyclodextrin or a β-cyclodextrin derivative, or a combination thereof, wherein the amount of the β-cyclodextrin or the β-cyclodextrin derivative, or the total amount of the β-cyclodextrin and β-cyclodextrin derivative is at least 50 wt%of the total amount of the antibody and/or antibody derivative and the β-cyclodextrin and/or β-cyclodextrin derivative.

2. The dry powder formulation of paragraph 1, wherein the particles have a mass median aerodynamic diameter < 5 μm, < 4 μm, < 3.5 μm, < 3 μm, < 2.5 μm, or < 2 μm.

3. The dry powder formulation of paragraph 1 or paragraph 2, wherein the particles have a geometric standard deviation < 4.5, < 4, < 3.5, < 3, < 2.5, or < 2.

4. The dry powder formulation of any one of paragraphs 1-3, wherein the particles have a fine particle fraction > 40%, > 45%, > 50%, > 55%, or > 60%in cascade impactor study.

5. The dry powder formulation of any one of paragraphs 1-4, wherein the particles comprise an antibody derivative and a β-cyclodextrin derivative.

6. The dry powder formulation of paragraph 5, wherein the antibody derivative is an antigen-binding fragment (Fab) .

7. The dry powder formulation of paragraph 5 or paragraph 6, wherein the β-cyclodextrin derivative is 2-hydroxypropyl-β-cyclodextrin, methyl-β-cyclodextrin, or sulfobutylether β-cyclodextrin, or a combination thereof.

8. The dry powder formulation of any one of paragraphs 1-7, wherein the β-cyclodextrin derivative is a 2-hydroxypropyl-β-cyclodextrin.

9. The dry powder formulation of any one of paragraphs 1-8 further comprising a pharmaceutically acceptable excipient, an additional active agent, or a combination thereof.

10. The dry powder formulation of paragraph 9, wherein the pharmaceutically acceptable excipient is an amino acid, a peptide, a lipid, a protein, a chelating agent, a salt, a taste masking agent, a cation, or a polymer, or a combination thereof.

11. The dry powder formulation of paragraph 9 or paragraph 10, wherein the amount of the pharmaceutically acceptable excipient is in a range from 0.1 wt%to 20 wt%, from, from 0.1 wt%to 15 wt%, from 1 wt%to 12 wt%, from 1 wt%to 10 wt%, from 1 wt%and 15 wt%, from 2 wt%to 20 wt%, from 2 wt%to 15 wt%, from 2 wt%to 10 wt%, from 3 wt%to 20 wt%, from 3 wt%to 15 wt%, or from 3 wt%to 10 wt%of the dry powder formulation.

12. The dry powder formulation of paragraph 11, wherein the additional active agent is an anti-inflammatory agent, an anti-viral agent, or a chemotherapeutic agent, or a combination thereof.

13. A delivery system comprising an inhaler and the dry powder formulation of any one of paragraphs 1-12 in a unit dosage form.

14. The delivery system of paragraph 13, wherein the inhaler is a dry powder inhaler.

15. The delivery system of paragraph 13 or 14, wherein the total amount of antibody and/or antibody derivative in the unit dosage form is in a range from about 0.1 mg to about 50 mg, from about 0.5 to about 50 mg, or from about 1 to about 50 mg.

16. The delivery system of any one of paragraphs 13-15, wherein the emitted fraction of the powder formulation is > 50%, > 55%, > 60%, > 65%, > 70%, > 75%, > 80%, > 85%, > 90%, or > 95%.

17. A method of making the dry powder formulation of any one of paragraphs 1-12 comprising

(i) mixing an antibody and/or an antibody derivative and a β-cyclodextrin and/or a β-cyclodextrin derivative, and optionally a pharmaceutically acceptable excipient and/or an additional active agent, in a solvent to form a liquid feed; and

(ii) spray-drying or spray-freeze drying the liquid feed to form particles containing the antibody and/or antibody derivative and the β-cyclodextrin and/or β-cyclodextrin derivative, and optionally the pharmaceutically acceptable excipient and/or the additional active agent.

18. The method of paragraph 17, wherein following step (ii) , the production yield of the particles is at least about 50 wt%, at least 55 wt%, at least 60 wt%, at least 65 wt%, at least 70 wt%, at least 75 wt%, at least 80 wt%, at least 85 wt%, or at least 90 wt%, or in a range from about 50 wt%to about 95 wt%, from about 55 wt%to about 95 wt%, from about 55 wt%to about 90 wt%, from about 60 wt%to about 95 wt%, from about 65 wt%to about 95 wt%, from about 70 wt%to about 95 wt%, or from about 75 wt%to about 95 wt%.

19. A method of preventing, treating, or ameliorating symptom (s) associated with a respiratory disease in a subject, the method comprising (i) administering to the subject the dry powder formulation of any one of paragraphs 1-12.

20. The method paragraph 19, wherein the subject is a mammal having or at the risk of having an obstructive airway disease, an inflammatory airway disease, or lung cancer, or a combination thereof.

21. The method of paragraph 19 or 20 further comprising loading the dry powder formulation in a dry powder inhaler prior to step (i) .

22. The method of any one of paragraphs 19-21, wherein, following step (i) , a unit dose of the antibody and/or the antibody derivative is delivered to the lungs of the subject in an amount from about 0.1 mg to about 50 mg, from about 0.5 to about 50 mg, or from about 1 to about 50 mg.

23. The method of any one of paragraphs 19-22 further comprising repeating step (i) .

24. The method of paragraph 23 further comprising repeating step (i) every hour, every 2 hours, every 5 hours, every 8 hours, every day, every 2 days, every 3 days, every 5 days, every 7 days, every 10 days, every two weeks, or every month.

25. The method of paragraph 23 or paragraph 24, wherein step (i) is repeated for a period between one day and 6 months, between one day and 3 months, between one and thirty days, between one and ten days, between one and three days, or during one day.

26. A method of preventing, treating, or ameliorating symptom (s) associated with a respiratory disease in a subject, the method comprising (i) administering to the subject the dry powder formulation of any one of paragraphs 13-16.

The present invention will be further understood by reference to the following non-limiting examples.

Examples

Example 1. Preparation of inhaled powder formulations of Fab

Materials and Methods

The spray drying and spray freeze drying techniques were used to produce inhalable powder formulation of Fab (see FIG. 1A and FIG. 1B) . Spray drying involved the atomization of liquid followed by drying with hot gas in the drying chamber (see FIG. 1A) . Spray freeze drying, which is a multiple-step process, involved spray freezing (atomisation of liquid into a cryogen) followed by freeze drying of the frozen particles (see FIG. 1B) .

Fab targeting IL-4Rα was used in this study (supplied by MabGeek, Shanghai) . The composition of each of the inhaled powder formulations, preparation parameters, and production yield are summarized in Table 1.

Quantification of monomer in each formulation was performed using HPLC coupled to a DAD (Agilent Technologies 1260 Infinity) with a Yarra ^TM 3 μm SEC-3000 column (

00H-4513-K0) at 25 ℃. The detection UV wavelength was set at 214 nm. The flow rate of the mobile phase, 150 mM trisodium phosphate (pH 6.8) , was 0.8 mL/min. 50 μL of sample solutions, adjusted to a concentration of 200 μg/mL antibody using the mobile phase, was injected with a stoptime of 16 minutes. The formulations were tested in triplicate to determine the protein stability in the formulation. The monomer peaks were integrated using Agilent OpenLab CDS ChemStation Edition (version C. 01.03) . The monomer content (%) was calculated as AUC _monomer/AUC _total.

Table 1. Summary of Fab formulations

Results

The in vitro aerosolisation performance of the spray-dried (also referred herein as “SD” ) powder formulations were evaluated and the aerodynamic properties and partial size distribution of each of the SD formulations are summarized in Table 2.

The emitted fraction (EF) is the fraction of powder exited the inhaler. The fine particle fraction (FPF) is the fraction of powder with aerodynamic diameter < 5 μm, i.e. respirable fraction, expressed as percentage by mass of cyclodextrin relative to the recovered mass. Data is presented as mean ± standard deviation (n=3) . Mass median aerodynamic diameter (MMAD) and geometric standard deviation (GSD) were also calculated.

Table 2. Aerodynamic properties and particle size distribution of SD formulations

Formulation	EF (%)	FPF (%)	MMAD (μm)	GSD
SD-B1 ^#	56.9 ± 1.5	44.9 ± 6.6	2.29	1.92
SD-B3 ^#	58.7 ± 2.5	46.6 ± 7.8	2.44	1.88
SD-B3*	72.1 ± 1.1	50.5 ± 3.8	2.00	2.59

^#The formulations were evaluated by Next Generation Impactor (NGI) operated at 60 L/min for 4 s, and the powder were dispersed by high-resistance osmohale ^TM inhaler.

*The formulations were evaluated by Next Generation Impactor (NGI) operated at 90 L/min for 2.7 s, and the powder were dispersed by

The in vitro aerosolisation performance of the spray-freeze-dried (also referred herein as “SFD” ) powder formulations were evaluated and the aerodynamic properties and partial size distribution of each of the SD formulations are summarized in Table 3.

Table 3. Aerodynamic properties and particle size distribution of SFD formulations

Formulation	EF (%)	FPF (%)	MMAD (μm)	GSD
SFD-F12^	97.9 ± 0.5	66.2 ± 1.3	1.63	3.93
SFD-F14^	96.5 ± 0.8	78.3 ± 2.7	1.07	4.08

^The formulations were evaluated by Next Generation Impactor (NGI) operated at 60 L/min for 4 s, and the powder were dispersed by

The morphology of the particles of the SD-B1 formulation is shown in FIGs. 2A and 2B. The morphologies of the particles of the SFD-F12 formulation and the SFD-F14 formulation are shown in FIGs. 3A and 3B and FIGs. 4A and 4B, respectively.

The SDS-PAGE analysis of powder formulations of Fab prepared by spray drying is shown in Figure 5. The SDS-PAGE analysis of powder formulations of Fab prepared by spray freeze drying is shown in Figure 6.

The monomer content of each of inhaled formulation SD-B1, SD-B3, SFD-F12, and SFD-F14 were evaluated by size-exclusion chromatography ( “SEC” ) to determine the protein stability in each of these formulations. The results are summarized in Table 4.

Table 4. Antibody aggregation analysis by SEC

Formulation	Monomer (%) , n=3	No. of weeks post-drying
SD-B1 (5%Fab)	99.7 ± 0.15	16
SD-B3 (10%Fab)	99.2 ± 0.15	16
SD-F12 (10%Fab)	99.1 ± 0.07	8
SD-F14 (2%Fab)	99.2 ± 0.05	8

Example 2. In vitro binding activity of Fab formulated as inhaled powder formulation is retained.

Materials and Methods

The binding activity of unprocessed Fab ( “upFab” ) and Fab formulated as inhaled powder formulations was assessed by ELISA.

The in vitro toxicity of unprocessed Fab and Fab formulated as inhaled powder formulations was also assessed by a TF-1 cell proliferation assay.

Results

The antibody binding results for upFab and Fab formulations prepared by spray drying process are shown in FIG. 7 and summarized in Table 5. The results show that the SD-B3 formulation has a binding potency comparable to that of the upFab.

Table 5. Antibody binding assessed by ELISA for SD-B3 formulation and unprocessed Fab

Sample	Log EC ₅₀ (μg/mL)	CV (%)
upFab	-1.37 ± 0.22	16%
SD-B3	-1.19 ± 0.12	10%

P values (summary)

Paired t test, upFab vs. B3 = 0.3415 (ns)

CV=coefficient of variation and was calculated as standard deviation divided by mean.

The antibody binding results for upFab and Fab formulations prepared by spray freeze drying process are shown in FIG. 8 and summarized in Table 6. The results show that each of the SFD-F12 and SFD-F14 formulations has a binding potency comparable to that of the upFab.

Table 6. Antibody binding assessed by ELISA for SFD-F12 formulation, SFD-F14 formulation, and unprocessed Fab

Sample	Log EC ₅₀ (μg/mL)	CV (%)
upFab	-1.43 ± 0.07	5%
SFD-F12	-1.39 ± 0.05	3%
SFD-F14	-1.35 ± 0.04	3%

P values (summary)

Paired t test, upFab vs. F12 = 0.0860 (ns)

Paired t test, upFab vs. F14 = 0.2566 (ns)

Paired t test, F12 vs. F14 = 0.4327 (ns)

The in vitro TF-1 cell toxicity results for upFab and Fab formulations prepared by spray drying process are shown in FIG. 9 and summarized in Table 7. The results show that the SD-B3 formulation has an inhibitory potency comparable to that of the upFab.

Table 7. In vitro TF-1 cell toxicity results by TF-1 cell proliferation assay for SD-B3 formulation and unprocessed Fab

Sample	Log IC ₅₀ (μg/mL)	CV (%)
upFab	-1.23 ± 0.25	21%
SD-B3	-1.60 ± 0.24	15%

P values (summary)

Paired t test, upFab vs. B3 = 0.0657 (ns)

The in vitro TF-1 cell toxicity results for upFab and Fab formulations prepared by spray freeze drying process are shown in FIG. 10 and summarized in Table 8. The results show that each of the SFD-F12 and SFD-F14 formulations has an inhibitory potency comparable to that of the upFab.

Table 8. In vitro TF-1 cell toxicity results by TF-1 cell proliferation assay for SFD-F12 formulation, SFD-F14 formulation, and unprocessed Fab

P values (summary)

Paired t test, upFab vs. F12 = 0.4467 (ns)

Paired t test, upFab vs. F14 = 0.2178 (ns)

Paired t test, F12 vs. F14 = 0.6102 (ns)

Unless defined otherwise, all technical and scientific terms used herein have the same meanings as commonly understood by one of skill in the art to which the disclosed invention belongs. Publications cited herein and the materials for which they are cited are specifically incorporated by reference.

Those skilled in the art will recognize, or be able to ascertain using no more than routine experimentation, many equivalents to the specific embodiments of the invention described herein. Such equivalents are intended to be encompassed by the following claims.

Claims

A dry powder formulation for inhalation comprising particles comprising

an antibody or an antibody derivative, or a combination thereof; and

a β-cyclodextrin or a β-cyclodextrin derivative, or a combination thereof,

wherein the amount of the β-cyclodextrin or the β-cyclodextrin derivative, or the total amount of the β-cyclodextrin and β-cyclodextrin derivative is at least 50 wt%of the total amount of the antibody and/or antibody derivative and the β-cyclodextrin and/or β-cyclodextrin derivative.
The dry powder formulation of claim 1, wherein the particles have a mass median aerodynamic diameter < 5 μm, < 4 μm, < 3.5 μm, < 3 μm, < 2.5 μm, or < 2 μm.
The dry powder formulation of claim 1 or claim 2, wherein the particles have a geometric standard deviation < 4.5, < 4, < 3.5, < 3, < 2.5, or < 2.
The dry powder formulation of any one of claims 1-3, wherein the particles have a fine particle fraction > 40%, > 45%, > 50%, > 55%, or > 60%in cascade impactor study.
The dry powder formulation of any one of claims 1-4, wherein the particles comprise an antibody derivative and a β-cyclodextrin derivative.
The dry powder formulation of claim 5, wherein the antibody derivative is an antigen-binding fragment (Fab) .
The dry powder formulation of claim 5 or claim 6, wherein the β-cyclodextrin derivative is 2-hydroxypropyl-β-cyclodextrin, methyl-β-cyclodextrin, or sulfobutylether β-cyclodextrin, or a combination thereof.
The dry powder formulation of any one of claims 1-7, wherein the β-cyclodextrin derivative is a 2-hydroxypropyl-β-cyclodextrin.
The dry powder formulation of any one of claims 1-8 further comprising a pharmaceutically acceptable excipient, an additional active agent, or a combination thereof.
The dry powder formulation of claim 9, wherein the pharmaceutically acceptable excipient is an amino acid, a peptide, a lipid, a protein, a chelating agent, a salt, a taste masking agent, a cation, or a polymer, or a combination thereof.
The dry powder formulation of claim 9 or claim 10, wherein the amount of the pharmaceutically acceptable excipient is in a range from 0.1 wt%to 20 wt%, from, from 0.1 wt%to 15 wt%, from 1 wt%to 12 wt%, from 1 wt%to 10 wt%, from 1 wt%and 15 wt%, from 2 wt%to 20 wt%, from 2 wt%to 15 wt%, from 2 wt%to 10 wt%, from 3 wt%to 20 wt%, from 3 wt%to 15 wt%, or from 3 wt%to 10 wt%of the dry powder formulation.
The dry powder formulation of claim 11, wherein the additional active agent is an anti-inflammatory agent, an anti-viral agent, or a chemotherapeutic agent, or a combination thereof.
A delivery system comprising an inhaler and the dry powder formulation of any one of claims 1-12 in a unit dosage form.
The delivery system of claim 13, wherein the inhaler is a dry powder inhaler.
The delivery system of claim 13 or 14, wherein the total amount of antibody and/or antibody derivative in the unit dosage form is in a range from about 0.1 mg to about 50 mg, from about 0.5 to about 50 mg, or from about 1 to about 50 mg.
The delivery system of any one of claims 13-15, wherein the emitted fraction of the powder formulation is > 50%, > 55%, > 60%, > 65%, > 70%, > 75%, > 80%, > 85%, > 90%, or > 95%.
A method of making the dry powder formulation of any one of claims 1-12 comprising

(i) mixing an antibody and/or an antibody derivative and a β-cyclodextrin and/or a β-cyclodextrin derivative, and optionally a pharmaceutically acceptable excipient and/or an additional active agent, in a solvent to form a liquid feed; and

(ii) spray-drying or spray-freeze drying the liquid feed to form particles containing the antibody and/or antibody derivative and the β-cyclodextrin and/or β-cyclodextrin derivative, and optionally the pharmaceutically acceptable excipient and/or the additional active agent.
The method of claim 17, wherein following step (ii) , the production yield of the particles is at least about 50 wt%, at least 55 wt%, at least 60 wt%, at least 65 wt%, at least 70 wt%, at least 75 wt%, at least 80 wt%, at least 85 wt%, or at least 90 wt%, or in a range from about 50 wt%to about 95 wt%, from about 55 wt%to about 95 wt%, from about 55 wt%to about 90 wt%, from about 60 wt%to about 95 wt%, from about 65 wt%to about 95 wt%, from about 70 wt%to about 95 wt%, or from about 75 wt%to about 95 wt%.
A method of preventing, treating, or ameliorating symptom (s) associated with a respiratory disease in a subject, the method comprising (i) administering to the subject the dry powder formulation of any one of claims 1-12.
The method claim 19, wherein the subject is a mammal having or at the risk of having an obstructive airway disease, an inflammatory airway disease, or lung cancer, or a combination thereof.
The method of claim 19 or 20 further comprising loading the dry powder formulation in a dry powder inhaler prior to step (i) .
The method of any one of claims 19-21, wherein, following step (i) , a unit dose of the antibody and/or the antibody derivative is delivered to the lungs of the subject in an amount from about 0.1 mg to about 50 mg, from about 0.5 to about 50 mg, or from about 1 to about 50 mg.
The method of any one of claims 19-22 further comprising repeating step (i) .
The method of claim 23 further comprising repeating step (i) every hour, every 2 hours, every 5 hours, every 8 hours, every day, every 2 days, every 3 days, every 5 days, every 7 days, every 10 days, every two weeks, or every month.
The method of claim 23 or claim 24, wherein step (i) is repeated for a period between one day and 6 months, between one day and 3 months, between one and thirty days, between one and ten days, between one and three days, or during one day.
A method of preventing, treating, or ameliorating symptom (s) associated with a respiratory disease in a subject, the method comprising (i) administering to the subject the dry powder formulation of any one of claims 1-12.