Amino acid sequences directed against rank-l and polypeptides comprising the same for the treatment of bone diseases and disorders

Abstract

The present invention relates to amino acid sequences that are directed against RANK-L, as well as to compounds or constructs, and in particular proteins and polypeptides, that comprise or essentially consist of one or more such amino acid sequences. The invention also relates to nucleic acids encoding such amino acid sequences and polypeptides; to methods for preparing such amino acid sequences and polypeptides; to host cells expressing or capable of expressing such amino acid sequences or polypeptides; to compositions, and in particular to pharmaceutical compositions, that comprise such amino acid sequences, polypeptides, nucleic acids and/or host cells; and to uses of such amino acid sequences or polypeptides, nucleic acids, host cells and/or compositions, in particular for prophylactic, therapeutic or diagnostic purposes.

1 .- 61 . (canceled) 
     
     
         62 . A method for the prevention and/or treatment of at least one disease or disorder that is associated with RANK-L, with its biological or pharmacological activity, and/or with the biological pathways or signalling in which RANK-L is involved, said method comprising administering, to a subject in need thereof, a pharmaceutically active amount of at least one multivalent polypeptide, compound or construct, wherein the multivalent polypeptide, compound or construct comprises or essentially consists of at least two amino acid sequences and/or Nanobodies that specifically bind to RANKL, wherein the at least two amino acid sequences and/or Nanobodies essentially consist of 4 framework regions (FR1 to FR4 respectively) and 3 complementary determining regions (CDR1 to CDR3 respectively), in which:
 CDR1 is chosen from the group consisting of:   a) the amino acid sequences of SEQ ID NOs: 188-249;   b) amino acid sequences that have at least 80% amino acid identity with at least one of the amino acid sequences of SEQ ID NOs: 188-249; or   c) amino acid sequences that have 3, 2 or 1 amino acid difference with at least one of the amino acid sequences of SEQ ID NOs: 188-249;   
       and/or
 CDR2 is chosen from the group consisting of: 
 d) the amino acid sequences of SEQ ID NOs: 312-373 and 758; 
 e) amino acid sequences that have at least 80% amino acid identity with at least one of the amino acid sequences of SEQ ID NOs: 312-373 and 758; or 
 f) amino acid sequences that have 3, 2 or 1 amino acid difference with at least one of the amino acid sequences of SEQ ID NOs: 312-373 and 758; 
 
       and/or
 CDR3 is chosen from the group consisting of: 
 g) the amino acid sequences of SEQ ID NOs: 436-497; 
 h) amino acid sequences that have at least 80% amino acid identity with at least one of the amino acid sequences of SEQ ID NOs: 436-497; or 
 i) amino acid sequences that have 3, 2 or 1 amino acid difference with at least one of the amino acid sequences of SEQ ID NOs: 436-497. 
 
     
     
         63 .- 66 . (canceled) 
     
     
         67 . The method according to  claim 62 , wherein the at least two amino acid sequences and/or Nanobodies essentially consist of 4 framework regions (FR1 to FR4 respectively) and 3 complementary determining regions (CDR1 to CDR3 respectively), in which:
 CDR1 is chosen from the group consisting of:   a) the amino acid sequences of SEQ ID NOs: 188-249;   b) amino acid sequences that have at least 80% amino acid identity with at least one of the amino acid sequences of SEQ ID NOs: 188-249; or   c) amino acid sequences that have 3, 2 or 1 amino acid difference with at least one of the amino acid sequences of SEQ ID NOs: 188-249;   
       and
 CDR2 is chosen from the group consisting of: 
 d) the amino acid sequences of SEQ ID NOs: 312-373 and 758; 
 e) amino acid sequences that have at least 80% amino acid identity with at least one of the amino acid sequences of SEQ ID NOs: 312-373 and 758; or 
 f) amino acid sequences that have 3, 2 or 1 amino acid difference with at least one of the amino acid sequences of SEQ ID NOs: 312-373 and 758; 
 
       and
 CDR3 is chosen from the group consisting of: 
 g) the amino acid sequences of SEQ ID NOs: 436-497; 
 h) amino acid sequences that have at least 80% amino acid identity with at least one of the amino acid sequences of SEQ ID NOs: 436-497; or 
 i) amino acid sequences that have 3, 2 or 1 amino acid difference with at least one of the amino acid sequences of SEQ ID NOs: 436-497. 
 
     
     
         68 . The method according to  claim 62 , wherein the at least two amino acid sequences and/or Nanobodies essentially consist of 4 framework regions (FR1 to FR4 respectively) and 3 complementary determining regions (CDR1 to CDR3 respectively), in which the CDR sequences of said amino acid sequences and/or Nanobodies have at least 70% amino acid identity, preferably at least 80% amino acid identity, more preferably at least 90% amino acid identity, such as 95% amino acid identity or more or even essentially 100% amino acid identity with the CDR sequences of at least one of the amino acid sequences of SEQ ID NOs: 560-621. 
     
     
         69 . The method according to  claim 62 , wherein the at least two amino acid sequences essentially consist of a domain antibody (or an amino acid sequence that is suitable for use as a domain antibody), of a single domain antibody (or an amino acid sequence that is suitable for use as a single domain antibody), of a “dAb” (or an amino acid sequence that is suitable for use as a dAb), a Nanobody (including but not limited to a V HH  sequence), a partially humanized Nanobody or a fully humanized Nanobody. 
     
     
         70 . The method according to  claim 62 , wherein the at least two amino acid sequences and/or Nanobodies are a V HH  sequence, a partially humanized V HH  sequence, a fully humanized V HH  sequence, a camelized heavy chain variable domain or a Nanobody that has been obtained by affinity maturation. 
     
     
         71 . The method according to  claim 62 , wherein the at least two amino acid sequences essentially consist of a Nanobody that
 i) has at least 80% amino acid identity with at least one of the amino acid sequences chosen from the group consisting of SEQ ID NOs: 1-22, 560-621, 730-757 and 765, in which for the purposes of determining the degree of amino acid identity, the amino acid residues that form the CDR sequences are disregarded;   
       and in which:
 ii) preferably one or more of the amino acid residues at positions 11, 37, 44, 45, 47, 83, 84, 103, 104 and 108 according to the Kabat numbering are chosen from the Hallmark residues mentioned in Table A-3. 
 
     
     
         72 . The method according to  claim 62 , wherein the multivalent polypeptide, compound or construct comprises at least two Nanobodies that are chosen from the group consisting of SEQ ID NOs: 560-621 or SEQ ID NOs: 730-757 and 765 or from the group consisting of amino acid sequences that have more than 80%, preferably more than 90%, more preferably more than 95%, such as 99% or more sequence identity (as defined herein) with at least one of the amino acid sequences of SEQ ID NOs: 560-621 or SEQ ID NOs: 730-757 and 765. 
     
     
         73 . The method according to  claim 62 , wherein the multivalent polypeptide, compound or construct is a bivalent polypeptide, compound or construct. 
     
     
         74 . The method according to  claim 62 , wherein the multivalent polypeptide, compound or construct comprises or essentially consists of an amino acid sequence chosen from SEQ ID NOs: 622-729, 759-762 and 766-773. 
     
     
         75 . The method according to  claim 62 , wherein the multivalent polypeptide, compound or construct essentially consists of a polypeptide that
 i) has at least 80% amino acid identity with at least one of the amino acid sequences of SEQ ID NOs: 622-729, 759-762 and 766-773, in which for the purposes of determining the degree of amino acid identity, the amino acid residues that form the CDR sequences are disregarded;   and in which:   ii) preferably one or more of the amino acid residues at positions 11, 37, 44, 45, 47, 83, 84, 103, 104 and 108 according to the Kabat numbering are chosen from the Hallmark residues mentioned in Table A-3.   
     
     
         76 . The method according to  claim 62 , wherein the multivalent polypeptide, compound or construct is a multispecific construct. 
     
     
         77 . The method according to  claim 62 , wherein the multivalent polypeptide, compound or construct further comprises one or more other groups, residues, moieties or binding units, optionally linked via one or more linkers. 
     
     
         78 . The method according to  claim 77 , in which said one or more other groups, residues, moieties or binding units provide the multivalent polypeptide, compound or construct with increased half-life. 
     
     
         79 . The method according to  claim 78 , in which said one or more other groups, residues, moieties or binding units that provide the multivalent polypeptide, compound or construct with increased half-life is chosen from the group consisting of serum proteins or fragments thereof, binding units that can bind to serum proteins, an Fc portion, and small proteins or peptides that can bind to serum proteins. 
     
     
         80 . The method according to  claim 78 , in which said one or more other groups, residues, moieties or binding units that provides the multivalent polypeptide, compound or construct with increased half-life are chosen from the group consisting of binding units that can bind to serum albumin (such as human serum albumin) or a serum immunoglobulin (such as IgG). 
     
     
         81 . The method according to  claim 78 , wherein said one or more other groups, residues, moieties or binding units that provides the multivalent polypeptide, compound or construct with increased half-life are chosen from the group consisting of domain antibodies, amino acid sequences that are suitable for use as a domain antibody, single domain antibodies, amino acid sequences that are suitable for use as a single domain antibody, “dAb”'s, amino acid sequences that are suitable for use as a dAb, or Nanobodies that can bind to serum albumin (such as human serum albumin) or a serum immunoglobulin (such as IgG). 
     
     
         82 . The method according to  claim 78 , in which said one or more other binding units that provides the multivalent polypeptide, compound or construct with increased half-life is chosen from SEQ ID NOs: 790-791. 
     
     
         83 . The method according to  claim 62 , wherein the at least one disease or disorder that is associated with RANK-L, with its biological or pharmacological activity, and/or with the biological pathways or signalling in which RANK-L is involved, is a bone disease or disorder. 
     
     
         84 . The method according to  claim 83 , wherein said bone disease or disorder is chosen from the group consisting of osteoporosis, Paget's disease, osteomyelitis, hypercalcemia, osteonecrosis, osteopenic disorders, arthritic disorders and periprosthetic osteolysis. 
     
     
         85 . A multivalent polypeptide, compound or construct comprising or essentially consisting of at least two amino acid sequences and/or Nanobodies that are directed against and/or that can specifically bind to RANK-L, in which the CDR sequences of said amino acid sequences and/or Nanobodies have at least 70% amino acid identity, preferably at least 80% amino acid identity, more preferably at least 90% amino acid identity, such as 95% amino acid identity or more or even essentially 100% amino acid identity with the CDR sequences of the amino acid sequence of SEQ ID NOs: 560-621 and SEQ ID NOs: 730-757 and 765.