Chimeric fibroblast growth factor 19 proteins and methods of use
The present invention relates to a chimeric protein that includes an N-terminus coupled to a C-terminus, where the N-terminus includes a portion of a paracrine fibroblast growth factor (“FGF”) and the C-terminus includes a C-terminal portion of an FGF19 molecule. The portion of the paracrine FGF is modified to decrease binding affinity for heparin and/or heparan sulfate compared to the portion without the modification. The present invention also relates to pharmaceutical compositions including chimeric proteins according to the present invention, methods for treating a subject suffering from diabetes, obesity, or metabolic syndrome, and methods of screening for compounds with enhanced binding affinity for the βKlotho-FGF receptor complex involving the use of chimeric proteins of the present invention.
1. A chimeric protein comprising:
an N-terminus coupled to a C-terminus,
wherein the N-terminus comprises an FGF2 portion beginning at any one of residues 1 to 25 and ending at any one of residues 151-155 of SEQ ID NO: 121,
wherein the FGF2 amino acid positions corresponding to those selected from the group consisting of N36, K128, R129, K134, K138, Q143, K144, and combinations thereof are substituted to decrease binding affinity for heparin and/or heparan sulfate compared to FGF2 without the substitution, and
wherein the C-terminus comprises a portion of an FGF19 comprising amino acid residues 169 to 216 of SEQ ID NO: 233.
2. The chimeric protein according to claim 1 , wherein the FGF2 portion is amino acid residues 1-151 of SEQ ID NO: 121.
3. The chimeric protein according to claim 1 , wherein the FGF2 portion is amino acid residues 25-151 of SEQ ID NO: 121.
4. The chimeric protein according to claim 1 , wherein the FGF2 portion is amino acid residues 1-152, 1-153, 1-154, 1-155, 2-151, 2-152, 2-153, 2-154, 2-155, 3-151, 3-152, 3-153, 3-154, 3-155, 4-151, 4-152, 4-153, 4-154, 4-155, 5-151, 5-152, 5-153, 5-154, 5-155, 6-151, 6-152, 6-153, 6-154, 6-155, 7-151, 7-152, 7-153, 7-154, 7-155, 8-151, 8-152, 8-153, 8-154, 8-155, 9-151, 9-152, 9-153, 9-154, 9-155, 10-151, 10-152, 10-153, 10-154, 10-155, 11-151, 11-152, 11-153, 11-154, 11-155, 12-151, 12-152, 12-153, 12-154, 12-155, 13-151, 13-152, 13-153, 13-154, 13-155, 14-151, 14-152, 14-153, 14-154, 14-155, 15-151, 15-152, 15-153, 15-154, 15-155, 16-151, 16-152, 16-153, 16-154, 16-155, 17-151, 17-152, 17-153, 17-154, 17-155, 18-151, 18-152, 18-153, 18-154, 18-155, 19-151, 19-152, 19-153, 19-154, 19-155, 20-151, 20-152, 20-153, 20-154, 20-155, 21-151, 21-152, 21-153, 21-154, 21-155, 22-151, 22-152, 22-153, 22-154, 22-155, 23-151, 23-152, 23-153, 23-154, 23-155, 24-151, 24-152, 24-153, 24-154, 24-155, 25-152, 25-153, 25-154, or 25-155 of SEQ ID NO: 121.
5. The chimeric protein according to claim 1 , wherein the one or more substitutions are selected from the group consisting of N36; K128D; R129Q; K134V; K138H; Q143M; K144T, K144L, or K144I; and combinations thereof.
6. A pharmaceutical composition comprising the chimeric protein according to claim 1 and a pharmaceutically-acceptable carrier.
7. The pharmaceutical composition according to claim 6 further comprising:
one or more agents selected from the group consisting of an anti-inflammatory agent, an antifibrotic agent, an antihypertensive agent, an antidiabetic agent, a triglyceride-lowering agent, and a cholesterol-lowering agent.
8. The chimeric protein according to claim 1 , wherein the chimeric protein comprises the amino acid sequence of SEQ ID NO:335 or SEQ ID NO:336.
9. The chimeric protein according to claim 1 , wherein the one or more amino acid substitutions comprises a substitution at amino acid residue N36.
10. The chimeric protein according to claim 9 , wherein the substitution is N36T.
11. The chimeric protein according to claim 1 , wherein the one or more amino acid substitutions comprises a substitution at amino acid residue K128.
12. The chimeric protein according to claim 11 , wherein the substitution is K128D.
13. The chimeric protein according to claim 1 , wherein the one or more amino acid substitutions comprises a substitution at amino acid residue R129.
14. The chimeric protein according to claim 13 , wherein the substitution is R129Q.
15. The chimeric protein according to claim 1 , wherein the one or more amino acid substitutions comprises a substitution at amino acid residue K134.
16. The chimeric protein according to claim 15 , wherein the substitution is K134V.
17. The chimeric protein according to claim 1 , wherein the one or more amino acid substitutions comprises a substitution at amino acid residue K138.
18. The chimeric protein according to claim 17 , wherein the substitution is K138H.
19. The chimeric protein according to claim 1 , wherein the one or more amino acid substitutions comprises a substitution at amino acid residue Q143.
20. The chimeric protein according to claim 19 , wherein the substitution is Q143M.
21. The chimeric protein according to claim 1 , wherein the one or more amino acid substitutions comprises a substitution at amino acid residue K144.
22. The chimeric protein according to claim 21 , wherein the substitution is K144T.
23. The chimeric protein according to claim 21 , wherein the substitution is K144L.
24. The chimeric protein according to claim 21 , wherein the substitution is K144I.
25. The chimeric protein of claim 1 , wherein the one or more amino acid substitutions comprises substitutions at amino acid residues K128, R129, and K134.
26. The chimeric protein of claim 25 , wherein the substitutions are K128D, R129Q, and K134V.