Amino acid sequences directed against CXCR4 and other GPCRs and compounds comprising the same
The present invention relates to amino acid sequences that are directed against (as defined herein) G-protein coupled receptors (GPCRs) and in particular to CXCR4 and CXCR7, 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 (also referred to herein as “amino acid sequences of the invention”, “compounds of the invention”, and “polypeptides of the invention”, respectively). Furthermore, the invention provides a new method of making amino acid sequences that are directed against transmembrane protein, and in particular for multiple spanning transmembrane proteins for which the native conformation cannot be reproduced in other “in vitro” system (e.g. GPCRs in general).
1. Polypeptide that comprises at least one Nanobody that specifically binds human CXCR4 (SEQ ID NO: 254) and optionally at least one Nanobody that specifically binds to a serum protein, wherein the at least one Nanobody that specifically binds human CXCR4 essentially consists of 4 framework regions (FR1 to FR4, respectively) and 3 complementarity determining regions (CDR1 to CDR3, respectively), in which CDR1 is:
the amino acid sequence of SEQ ID NO: 142;
and
CDR2 is
the amino acid sequence of SEQ ID NO: 174;
and
CDR3 is
the amino acid sequence of SEQ ID NO: 206;
in which
CDR1 is the amino acid sequence of SEQ ID NO: 143;
and
CDR2 is the amino acid sequence of SEQ ID NO: 175;
and
CDR3 is the amino acid sequence of SEQ ID NO: 207.
2. Polypeptide of claim 1 ; wherein the polypeptide comprises two Nanobodies that specifically bind human CXCR4 and wherein the two Nanobodies that specifically bind human CXCR4 each essentially consists of 4 framework regions (FR1 to FR4, respectively) and 3 complementarity determining regions (CDR1 to CDR3, respectively), in which CDR1
the amino acid sequence of SEQ ID NO: 142;
and
CDR2 is
the amino acid sequence of SEQ ID NO: 174;
and
CDR3 is
the amino acid sequence of SEQ ID NO: 206;
or
in which
CDR1 is the amino acid sequence of SEQ ID NO: 143;
and
CDR2 is the amino acid sequence of SEQ ID NO: 175;
and
CDR3 is the amino acid sequence of SEQ ID NO: 207.
3. Polypeptide of claim 2 ;
wherein one Nanobody that specifically binds human CXCR4 essentially consists of 4 framework regions (FR1 to FR4, respectively) and 3 complementarity determining regions (CDR1 to CDR3, respectively), in which CDR1 is
the amino acid sequence of SEQ ID NO: 142;
and
CDR2 is
the amino acid sequence of SEQ ID NO: 174;
and
CDR3 is
the amino acid sequence of SEQ ID NO: 206;
wherein another Nanobody that specifically binds human CXCR4 essentially consists of 4 framework regions (FR1 to FR4, respectively) and 3 complementarity determining regions (CDR1 to CDR3, respectively), in which CDR1 is
the amino acid sequence of SEQ ID NO: 143;
and
CDR2 is
the amino acid sequence of SEQ ID NO: 175;
and
CDR3 is
the amino acid sequence of SEQ ID NO: 207.
4. Nanobody that essentially consists of 4 framework regions (FR1 to FR4, respectively) and 3 complementarity determining regions (CDR1 to CDR3, respectively), in which CDR1 is
the amino acid sequence of SEQ ID NO: 142;
and
CDR2 is
the amino acid sequence of SEQ ID NO: 174;
and
CDR3 is
the amino acid sequence of SEQ ID NO: 206;
in which
CDR1 is the amino acid sequence of SEQ ID NO: 143;
and
CDR2 is the amino acid sequence of SEQ ID NO: 175;
and
CDR3 is the amino acid sequence of SEQ ID NO: 207.
5. Polypeptide of claim 1 wherein the Nanobodies are selected from the group consisting of Nanobodies represented by the amino acid sequences of SEQ ID NO's 238 to 239.
6. Polypeptide of claim 1 , wherein the polypeptide is selected from the group consisting of polypeptides represented by the amino acid sequences of SEQ ID NO's 261 to 264, preferably SEQ ID NO's 263 to 264.
7. Nanobody of claim 4 selected from the group consisting of Nanobodies represented by the amino acid sequences of SEQ ID NO's 238 to 239.
8. Pharmaceutical composition comprising a polypeptide according to claim 1 .
9. Pharmaceutical composition according to claim 8 that further comprises at least one pharmaceutically acceptable carrier, diluent or excipient and/or adjuvant, and that optionally comprises one or more further pharmaceutically active polypeptides and/or compounds.
10. Pharmaceutical composition comprising a Nanobody according to claim 4 .
11. Pharmaceutical composition according to claim 10 that further comprises at least one pharmaceutically acceptable carrier, diluent or excipient and/or adjuvant, and that optionally comprises one or more further pharmaceutically active polypeptides and/or compounds.
12. Polypeptide of claim 6 , wherein the polypeptide is selected from the group consisting of polypeptides represented by the amino acid sequences of SEQ ID NOs: 263 to 264.