Antibodies specific for lox1 and use in treatment of cardiovascular disorders

Abstract

This disclosure provides LOX1 (LOX1) binding proteins such as anti-LOX1 antibodies, and compositions and methods for making these binding proteins. In certain aspects the LOX1-binding proteins provided herein, inhibit, or antagonize LOX1 activity. In addition, the disclosure provides compositions and methods for diagnosing and treating conditions associated with atherosclerosis, thrombosis, coronary artery disease (CAD), ischemia (e.g., myocardial ischemia), infarction (e.g., myocardial infarction), acute coronary syndrome (ACS), stroke, reperfusion injury, restenosis, peripheral vascular disease, hypertension, heart failure, inflammation (e.g., chronic inflammation), angiogenesis, preeclampsia, cancer and other LOX1-mediated diseases and conditions.

1 . A method of making a Lectin-like oxidized low density lipoprotein receptor-1 (LOX1) binding protein comprising culturing a host cell under suitable conditions for producing the LOX-1 binding protein, wherein the host cell comprises a nucleic acid molecule or set of nucleic acid molecules encoding the LOX1-binding protein, wherein the LOX1-binding protein comprises a set of complementary determining regions (CDRs): heavy chain variable region (VH)-CDR1, VH-CDR2, VH-CDR3, and light chain variable region (VL)-CDR1, VL-CDR2 and VL-CDR3, wherein:
 (a) (i) VH-CDR1 has the amino acid sequence of SEQ ID NO:1;
 (ii) VH-CDR2 has the amino acid sequence of SEQ ID NO:2; 
 (iii) VH-CDR3 has the amino acid sequence of SEQ ID NO:3; 
 (iv) VL-CDR1 has the amino acid sequence of SEQ ID NO:30; 
 (v) VL-CDR2 has the amino acid sequence of SEQ ID NO:31; and 
 (vi) VL-CDR3 has the amino acid sequence of SEQ ID NO:32; 
   (b) (i) VH-CDR1 has the amino acid sequence of SEQ ID NO:1;
 (ii) VH-CDR2 has the amino acid sequence of SEQ ID NO:5; 
 (iii) VH-CDR3 has the amino acid sequence of SEQ ID NO:14; 
 (iv) VL-CDR1 has the amino acid sequence of SEQ ID NO:30; 
 (v) VL-CDR2 has the amino acid sequence of SEQ ID NO:31; and 
 (vi) VL-CDR3 has the amino acid sequence of SEQ ID NO:32; 
   (c) (i) VH-CDR1 has the amino acid sequence of SEQ ID NO:38;
 (ii) VH-CDR2 has the amino acid sequence of SEQ ID NO:39; 
 (iii) VH-CDR3 has the amino acid sequence of SEQ ID NO:44; 
 (iv) VL-CDR1 has the amino acid sequence of SEQ ID NO:55; 
 (v) VL-CDR2 has the amino acid sequence of SEQ ID NO:60; and 
 (vi) VL-CDR3 has the amino acid sequence of SEQ ID NO:61; 
   or   (d) (i) VH-CDR1 has the amino acid sequence of SEQ ID NO:38;
 (ii) VH-CDR2 has the amino acid sequence of SEQ ID NO:39; 
 (iii) VH-CDR3 has the amino acid sequence of SEQ ID NO:40; 
 (iv) VL-CDR1 has the amino acid sequence of SEQ ID NO:55; 
 (v) VL-CDR2 has the amino acid sequence of SEQ ID NO:56; and 
 (vi) VL-CDR3 has the amino acid sequence of SEQ ID NO:57. 
   
     
     
         2 . The method of  claim 1 , wherein the LOX1-binding protein comprises a heavy chain variable region (VH) and a light chain variable region (VL) selected from the group consisting of:
 (a) a VH having the amino acid sequence of SEQ ID NO:4 and a VL having the amino acid sequence of SEQ ID NO:33;   (b) a VH having the amino acid sequence of SEQ ID NO:29 and a VL having the amino acid sequence of SEQ ID NO:33;   (c) a VH having the amino acid sequence of SEQ ID NO:41 and a VL having the amino acid sequence of SEQ ID NO:58 and   (d) a VH having the amino acid sequence of SEQ ID NO:54 and a VL having the amino acid sequence of SEQ ID NO:70.   
     
     
         3 . The method of  claim 1 , wherein the LOX1-binding protein comprises a heavy chain variable region (VH) comprising SEQ ID NO:4 and a light chain variable region (VL) comprising SEQ ID NO:33. 
     
     
         4 . The method of  claim 1 , wherein the LOX1-binding protein reduces, inhibits or antagonizes LOX1 activity. 
     
     
         5 . The method of  claim 1 , wherein the LOX1-binding protein is an antibody. 
     
     
         6 . The method of  claim 5 , wherein the antibody is a monoclonal antibody, a recombinant antibody, a human antibody, a humanized antibody, a chimeric antibody, a bi-specific antibody, a multi-specific antibody, or a LOX1-binding antibody fragment. 
     
     
         7 . The method of  claim 6 , wherein the LOX1-binding antibody fragment is selected from the group consisting of: a Fab fragment, a Fab′ fragment, a F(ab′)2 fragment, a Fv fragment, a diabody, and a single chain antibody molecule. 
     
     
         8 . The method of  claim 1 , wherein the LOX1-binding protein further comprises a heavy chain immunoglobulin constant domain selected from the group consisting of:
 (a) a human IgA constant domain   (b) a human IgD constant domain;   (c) a human IgE constant domain;   (d) a human IgG1 constant domain;   (e) a human IgG2 constant domain;   (f) a human IgG3 constant domain;   (g) a human IgG4 constant domain; and   (h) a human IgM constant domain.   
     
     
         9 . The method of  claim 1 , wherein the LOX1-binding protein further comprises a light chain immunoglobulin constant domain selected from the group consisting of:
 (a) a human Ig kappa constant domain; and   (b) a human Ig lambda constant domain.   
     
     
         10 . The method of  claim 1 , wherein the LOX1-binding protein further comprises a human IgG1 heavy chain constant domain and a human lambda light chain constant domain. 
     
     
         11 . The method of  claim 10 , wherein the IgG1 heavy chain constant domain contains a mutation at positions 234, 235 and 331, wherein the position numbering is according to the EU index as in Kabat. 
     
     
         12 . The method of  claim 11 , wherein the IgG1 heavy chain constant domain contains the mutations L234F, L235E and P331S, wherein the position numbering is according to the EU index as in Kabat. 
     
     
         13 . The method of  claim 1 , wherein the nucleic acid molecule is operably linked to a control sequence. 
     
     
         14 . The method of  claim 1 , wherein the host cell comprises a vector comprising the nucleic acid molecule according to  claim 1 . 
     
     
         15 . The method of  claim 1 , wherein the host cell is a mammalian host cell. 
     
     
         16 . The method of  claim 15 , wherein the host cell is a NSO murine myeloma cell, a PER.C6® human cell, or a Chinese hamster ovary (CHO) cell. 
     
     
         17 . The method of  claim 1  further comprising isolating the LOX1-binding protein secreted from the host cell. 
     
     
         18 . An isolated LOX1-binding protein that binds the same epitope as a LOX1-binding protein produced according to  claim 1 . 
     
     
         19 . A method of making a Lectin-like oxidized low density lipoprotein receptor-1 (LOX1) binding protein comprising culturing a host cell under suitable conditions for producing the LOX-1 binding protein, wherein the host cell comprises a nucleic acid molecule or set of nucleic acid molecules encoding the LOX1-binding protein, wherein the LOX1-binding protein comprises a set of complementary determining regions (CDRs): heavy chain variable region (VH)-CDR1, VH-CDR2, VH-CDR3, and light chain variable region (VL)-CDR1, VL-CDR2 and VL-CDR3, wherein the set of CDRs has a total of 18 or fewer amino acid substitutions, deletions, and/or insertions from a reference set of CDRs in which:
 (i) VH-CDR1 has the amino acid sequence of SEQ ID NO:1;   (ii) VH-CDR2 has the amino acid sequence of SEQ ID NO:5;   (iii) VH-CDR3 has the amino acid sequence of SEQ ID NO:14;   (iv) VL-CDR1 has the amino acid sequence of SEQ ID NO:30;   (v) VL-CDR2 has the amino acid sequence of SEQ ID NO:31; and   (vi) VL-CDR3 has the amino acid sequence of SEQ ID NO:32.