Thymic stromal lymphopoietin (tslp)-binding molecules and methods of using the molecules

Abstract

The invention provides molecules, e.g., antibodies or antibody fragments, that specifically bind thymic stromal lymphopoietin (TSLP), compositions comprising these molecules, and methods of using and producing these molecules.

1 . A molecule that specifically binds human thymic stromal lymphopoietin (TSLP) selected from any one of the following:
 a) a molecule that comprises:   a heavy chain complementarity determining region 1 (HCDR1) comprising the amino acid sequence of SEQ ID NO: 4;   a heavy chain complementarity determining region 2 (HCDR2) comprising the amino acid sequence of SEQ ID NO: 2;   a heavy chain complementarity determining region 3 (HCDR3) comprising the amino acid sequence of SEQ ID NO: 3;   a light chain complementarity determining region 1 (LCDR1) comprising the amino acid sequence of SEQ ID NO: 11;   a light chain complementarity determining region 2 (LCDR2) comprising the amino acid sequence of SEQ ID NO: 12; and   a light chain complementarity determining region 3 (LCDR3) comprising the amino acid sequence of SEQ ID NO: 13;   b) a molecule that comprises:   a HCDR1 comprising the amino acid sequence of SEQ ID NO: 5;   a HCDR2 comprising the amino acid sequence of SEQ ID NO: 6;   a HCDR3 comprising the amino acid sequence of SEQ ID NO: 3;   a LCDR1 comprising the amino acid sequence of SEQ ID NO: 14;   a LCDR2 comprising the amino acid sequence of SEQ ID NO: 15; and   a LCDR3 comprising the amino acid sequence of SEQ ID NO: 16;   c) a molecule that comprises a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 7, and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 17;   d) a molecule that comprises a heavy chain comprising the amino acid sequence of SEQ ID NO: 22, and a light chain comprising the amino acid sequence of SEQ ID NO: 25;   e) A molecule that comprises a heavy chain comprising the amino acid sequence of SEQ ID NO: 9, and a light chain comprising the amino acid sequence of SEQ ID NO: 19;   f) A molecule that comprises a paratope comprising at least one of the following residues: Thr28, Asp31, Tyr32, Trp33, Asp56, Glu101, Ile102, Tyr103, Tyr104, Tyr105 of a heavy chain sequence of SEQ ID NO:22 or Gly28, Ser29, Lys30, Tyr31, Tyr48, Asp50, Asn51, Glu52, Asn65, and Trp92 of a light chain sequence of SEQ ID NO:25;   g) An antibody fragment that binds human TSLP and comprises   a HCDR1 comprising the amino acid sequence of SEQ ID NO: 4;   a HCDR2 comprising the amino acid sequence of SEQ ID NO: 2;   a HCDR3 comprising the amino acid sequence of SEQ ID NO: 3;   a LCDR1 comprising the amino acid sequence of SEQ ID NO: 11;   a LCDR2 comprising the amino acid sequence of SEQ ID NO: 12; and   a LCDR3 comprising the amino acid sequence of SEQ ID NO: 13;   
       and
 h) An antibody fragment that binds human TSLP comprises: 
 a HCDR1 comprising the amino acid sequence of SEQ ID NO: 5; 
 a HCDR2 comprising the amino acid sequence of SEQ ID NO: 6; 
 a HCDR3 comprising the amino acid sequence of SEQ ID NO: 3; 
 a LCDR1 comprising the amino acid sequence of SEQ ID NO: 14; 
 a LCDR2 comprising the amino acid sequence of SEQ ID NO: 15; and 
 a LCDR3 comprising the amino acid sequence of SEQ ID NO: 16. 
 
     
     
         2 . A molecule that specifically binds an epitope in human TSLP, wherein the epitope comprises at least one of the following residues: Lys38, Ala41, Leu44, Ser45, Thr46, Ser48, Lys49, Ile52, Thr53, Ser56, Gly57, Thr58, Lys59, Lys101, Gln145, and Arg149 of SEQ ID NO: 38. 
     
     
         3 . The molecule of  claim 2 , wherein the epitope comprises at least one of the following sets of residues of SEQ ID NO: 38:
 (a) Lys49 and Ile52,   (b) Gly57 and Lys59,   (c) Lys101,   (d) Gln145 and Arg149.   
     
     
         4 . The molecule of  claim 1 , wherein the molecule is a monoclonal antibody. 
     
     
         5 . The molecule of  claim 1 , wherein the molecule is an antibody fragment selected from a Fab, Fab′, F(ab′)2, scFv, minibody, or diabody. 
     
     
         6 . The molecule of  claim 5 , wherein the molecule is a Fab. 
     
     
         7 . The molecule of  claim 6 , wherein the molecule is a human or humanized Fab. 
     
     
         8 . The molecule of  claim 1 , wherein the molecule is a human immunoglobulin. 
     
     
         9 . The molecule of  claim 1 , wherein the molecule binds human TSLP with a dissociation constant (K D ) of less than 100 pM. 
     
     
         10 . The molecule of  claim 1 , wherein the molecule binds human TSLP with a dissociation constant (K D ) of less than 10 pM. 
     
     
         11 . A pharmaceutical composition comprising the molecule of  claim 1  and at least one pharmaceutically acceptable excipient. 
     
     
         12 . The pharmaceutical composition of  claim 11 , wherein the molecule is about 5% to about 95%, or about 10% to about 90%, or about 15% to about 85%, or about 20% to about 80%, or about 25% to about 75%, or about 30% to about 70%, or about 40% to about 60%, or about 40-50% (w/w) of the composition. 
     
     
         13 . The pharmaceutical composition of  claim 11 , wherein the composition comprises a shell-forming agent. 
     
     
         14 . The pharmaceutical composition of  claim 13 , wherein the shell-forming agent is trileucine or leucine. 
     
     
         15 . The pharmaceutical composition of  claim 14 , wherein the trileucine or leucine is about 10-75% (w/w) of the composition. 
     
     
         16 . The pharmaceutical composition of  claim 15 , wherein the trileucine is about 10-30% (w/w) of the composition, or wherein the leucine is about 50-75% (w/w) of the composition. 
     
     
         17 . The pharmaceutical composition of  claim 11 , wherein the composition comprises at least one glass-forming excipient. 
     
     
         18 . The pharmaceutical composition of  claim 17 , wherein the glass-forming excipient is selected from histidine, trehalose, mannitol, sucrose, or sodium citrate. 
     
     
         19 . The pharmaceutical composition of  claim 18 , wherein the at least one glass-forming excipient is selected from trehalose or a mixture of trehalose and mannitol. 
     
     
         20 . The pharmaceutical composition of  claim 17 , wherein the glass-forming excipient is about 15-35% (w/w) of the composition. 
     
     
         21 . The pharmaceutical composition of  claim 11 , wherein the composition comprises a buffer. 
     
     
         22 . The pharmaceutical composition of  claim 21 , wherein the buffer is selected from a histidine, glycine, acetate, or phosphate buffer. 
     
     
         23 . The pharmaceutical composition of  claim 21 , wherein the buffer is about 5-13% of the composition. 
     
     
         24 . The pharmaceutical composition of  claim 11 , wherein the composition is formulated as a dry powder formulation. 
     
     
         25 . The pharmaceutical composition of  claim 24 , wherein the composition is formulated as a dry powder formulation suitable for inhalation. 
     
     
         26 . The pharmaceutical composition of  claim 11 , wherein the composition comprises: spray-dried particles comprising a shell and a core, wherein the shell comprises trileucine or leucine and the core comprises:
 i) the molecule, trehalose, mannitol, and a buffer; or   ii) the molecule, trehalose, buffer, and HCl.   
     
     
         27 . The pharmaceutical composition of  claim 26 , wherein the buffer is selected from a histidine, glycine, acetate, or phosphate buffer. 
     
     
         28 . A pharmaceutical composition comprising spray-dried particles comprising:
 i) a core comprising trehalose, mannitol, histidine, and a TSLP-binding molecule, or a core comprising trehalose, histidine, HCl, and a TSLP-binding molecule, wherein the TSLP-binding molecule is an antibody Fab fragment comprising:   a) a HCDR1 comprising the amino acid sequence of SEQ ID NO: 4;
 a HCDR2 comprising the amino acid sequence of SEQ ID NO: 2; 
 a HCDR3 comprising the amino acid sequence of SEQ ID NO: 3; 
 a LCDR1 comprising the amino acid sequence of SEQ ID NO: 11; 
 a LCDR2 comprising the amino acid sequence of SEQ ID NO: 12; and 
 a LCDR3 comprising the amino acid sequence of SEQ ID NO: 13; or 
   b) a HCDR1 comprising the amino acid sequence of SEQ ID NO: 5;
 a HCDR2 comprising the amino acid sequence of SEQ ID NO: 6; 
 a HCDR3 comprising the amino acid sequence of SEQ ID NO: 3; 
 a LCDR1 comprising the amino acid sequence of SEQ ID NO: 14; 
 a LCDR2 comprising the amino acid sequence of SEQ ID NO: 15; and 
 a LCDR3 comprising the amino acid sequence of SEQ ID NO: 16; and 
   ii) a shell comprising trileucine or leucine.   
     
     
         29 . The pharmaceutical composition of  claim 28  comprising:
 a) 40% (w/w) TSLP-binding molecule, 25% (w/w) trileucine, 30% (w/w) combined weight of trehalose and mannitol, and 5% (w/w) histidine; 
 b) 50% (w/w) TSLP-binding molecule, 15% (w/w) trileucine, 2.6% (w/w) HCl, 5.6% (w/w) histidine, and 26.8% (w/w) combined weight of trehalose and a base; or 
 c) 50% (w/w) TSLP-binding molecule, 15% (w/w) trileucine, 19.4% (w/w) trehalose, 13.04% (w/w) histidine, and 2.56% (w/w) HCl. 
 
     
     
         30 . A kit comprising the molecule of  claim 1 , or the pharmaceutical composition of  claim 11 , and a device for delivering the molecule or pharmaceutical composition to a subject. 
     
     
         31 . The kit of  claim 30 , wherein the device delivers the molecule or pharmaceutical composition in an aerosolized form. 
     
     
         32 . The kit of  claim 30 , wherein the device is a dry powder inhaler. 
     
     
         33 . A method of treating a TSLP-related condition in a subject in need thereof, the method comprising administering to the subject a therapeutically effective amount of the molecule of  claim 1 , or a pharmaceutical composition comprising the molecule of  claim 1  and a pharmaceutically acceptable excipient. 
     
     
         34 . The method of  claim 33 , wherein the TSLP-related inflammatory condition is selected from asthma, chronic obstructive pulmonary disease, allergic rhinitis, allergic rhinosinusitis, allergic conjunctivitis, eosinophilic esophagitis, and atopic dermatitis. 
     
     
         35 . The method of  claim 33 , wherein the TSLP-related inflammatory condition is asthma. 
     
     
         36 . The method of  claim 33 , wherein the molecule is formulated as a dry powder formulation suitable for inhalation. 
     
     
         37 . The method of  claim 33 , wherein the molecule is administered to the subject orally or intranasally. 
     
     
         38 . The method of  claim 33 , wherein the molecule is administered to the subject in an aerosolized form. 
     
     
         39 . The method of  claim 33 , wherein the molecule is administered to the subject by a dry powder inhaler. 
     
     
         40 . The method of  claim 33 , wherein the subject is a human. 
     
     
         41 . The method of  claim 33 , further comprising administering a second agent to the subject. 
     
     
         42 . The method of  claim 41 , wherein the second agent is selected from the group consisting of corticosteroids, bronchodilators, antihistamines, antileukotrienes, and PDE-4 inhibitors. 
     
     
         43 . A nucleic acid encoding the molecule of  claim 1 . 
     
     
         44 . A vector comprising the nucleic acid of  claim 43 . 
     
     
         45 . A host cell comprising the nucleic acid of  claim 43  or a vector comprising the nucleic acid of  claim 43 . 
     
     
         46 . A method of producing the molecule of  claim 1 , the method comprising:
 culturing a host cell expressing a nucleic acid encoding the molecule; and   collecting the molecule from the culture medium.   
     
     
         47 . A method for making a dry powder formulation comprising the molecule of  claim 1 , the method comprising:
 (a) providing an aqueous solution comprising the molecule of  claim 1 , trileucine or leucine, a glass forming excipient, and a buffer;   (b) spray drying the aqueous solution of step (a) at a temperature between about 120° C. to about 200° C. (inlet) range and 55° C. to about 75° C. (outlet) range to produce dry powder particles; and   (c) collecting the dry power particles.   
     
     
         48 . The method of  claim 47 , wherein the buffer is selected from a histidine, glycine, acetate, or phosphate buffer. 
     
     
         49 . The method of  claim 47 , wherein the glass forming excipient is selected from histidine, histidine HCl, trehalose, mannitol, sucrose, or sodium citrate. 
     
     
         50 . The pharmaceutical composition of  claim 11 , wherein the excipient:molecule mass ratio is greater than 0.5.