Peptidomimetic macrocycles
The present invention provides peptidomimetic macrocycles capable of modulating growth hormone levels and methods of using such macrocycles for the treatment of disease.
1. A peptidomimetic macrocycle comprising an amino acid sequence which is at least about 60% identical to GHRH 1-29, comprising at least two macrocycle-forming linkers, wherein the first of said two macrocycle-forming linkers connects a first amino acid to a second amino acid, and the second of said two macrocycle-forming linkers connects a third amino acid to a fourth amino acid, wherein the peptidomimetic macrocycle comprises an α,α-disubstituted amino acid.
2. The peptidomimetic macrocycle of claim 1 , comprising two macrocycle-forming linkers.
3. The peptidomimetic macrocycle of claim 1 , wherein each of said macrocycle-forming linkers connects a pair of amino acids corresponding to one of the following locations of amino acids: 4 and 8; 5 and 12; 8 and 12; 8 and 15; 9 and 16; 12 and 16; 12 and 19; 15 and 22; 18 and 25; 21 and 25; 21 and 28; 22 and 29; 25 and 29 of GHRH 1-29.
4. The peptidomimetic macrocycle of claim 3 , wherein each of said macrocycle-forming linkers connects a pair of amino acids corresponding to one of the following locations of amino acids: 4 and 8; 5 and 12; 12 and 19; 15 and 22; 18 and 25; 21 and 25; 21 and 28 of GHRH 1-29.
5. The peptidomimetic macrocycle of claim 3 , wherein the first macrocycle-forming linker connects a pair of amino acids corresponding to one of the following locations of amino acids: 4 and 8; 5 and 12; 8 and 12; 8 and 15; 9 and 16; 12 and 16; or 12 and 19; and the second macrocycle-forming linker connects amino acid pairs 15 and 22; 18 and 25; 21 and 25; 21 and 28; 22 and 29; or 25 and 29 of GHRH 1-29.
6. The peptidomimetic macrocycle of claim 3 , wherein the first macrocycle-forming linker connects a pair of amino acids corresponding to one of the following locations of amino acids: 4 and 8; 5 and 12; or 12 and 19; and the second macrocycle-forming linker connects amino acid pairs 15 and 22; 18 and 25; 21 and 25; or 21 and 28 of GHRH 1-29.
7. The peptidomimetic macrocycle of claim 3 , wherein the first macrocycle-forming linker connects a pair of amino acids corresponding to amino acid locations 4 and 8 of GHRH 1-29, and the second macrocycle-forming linker connects a pair of amino acids corresponding to amino acid locations 21 and 25 of GHRH 1-29.
8. A peptidomimetic macrocycle comprising:
i) an amino acid sequence which is at least about 60% identical to GHRH 1-29; and
ii) a macrocycle-forming linker connecting a first amino acid to a second amino acid, wherein the peptidomimetic macrocycle comprises an α,α-disubstituted amino acid, and wherein the first and second amino acids are selected from amino acids corresponding to the following locations of amino acids: 4 and 8; 5 and 12; 8 and 12; 8 and 15; 9 and 16; 12 and 16; 12 and 19; 15 and 22; 18 and 25; 21 and 25; 21 and 28; 22 and 29 of GHRH 1-29.
9. The peptidomimetic macrocycle of claim 8 , wherein the macrocycle-forming linker connects amino acids corresponding to amino acid locations 12 and 19 of GHRH 1-29.
10. A peptidomimetic macrocycle of claim 1 , comprising an amino acid sequence which is at least about 60% identical to an amino acid sequence chosen from the group consisting of the amino acid sequences in Tables 1, 2 or 4.
11. The peptidomimetic macrocycle of claim 1 , wherein the amino acid sequence of said peptidomimetic macrocycle is at least about 80% identical to an amino acid sequence chosen from the group consisting of the amino acid sequences in Tables 1, 2 or 4.
12. The peptidomimetic macrocycle of claim 1 , wherein the amino acid sequence of said peptidomimetic macrocycle is at least about 90% identical to an amino acid sequence chosen from the group consisting of the amino acid sequences in Tables 1, 2 or 4.
13. The peptidomimetic macrocycle of claim 1 , wherein the amino acid sequence of said peptidomimetic macrocycle is chosen from the group consisting of the amino acid sequences in Tables 1, 2 or 4.
14. The peptidomimetic macrocycle of claim 1 , wherein the peptidomimetic macrocycle comprises a helix.
15. The peptidomimetic macrocycle of claim 1 , wherein the peptidomimetic macrocycle comprises an α-helix.
16. The peptidomimetic macrocycle of claim 1 , wherein each amino acid connected by the macrocycle-forming linkers is an α,α-disubstituted amino acid.
17. The peptidomimetic macrocycle of any preceding claim, having the formula:
wherein:
each A, C, D, and E is independently an amino acid;
B is an amino acid, or
L is a macrocycle-forming linker of the formula -L 1 -L 2 -;
and wherein A, B, C, D, and E, taken together with the crosslinked amino acids connected by the macrocycle-forming linker L, form the amino acid sequence of the peptidomimetic macrocycle;
R 1 and R 2 are independently —H, alkyl, alkenyl, alkynyl, arylalkyl, cycloalkyl, cycloalkylalkyl, heteroalkyl, or heterocycloalkyl, unsubstituted or substituted with halo-;
R 3 is hydrogen, alkyl, alkenyl, alkynyl, arylalkyl, heteroalkyl, cycloalkyl, heterocycloalkyl, cycloalkylalkyl, cycloaryl, or heterocycloaryl, optionally substituted with R 5 ;
L 1 and L 2 are independently alkylene, alkenylene, alkynylene, heteroalkylene, cycloalkylene, heterocycloalkylene, cycloarylene, heterocycloarylene, or [—R 4 —K—R 4 —] n , each being optionally substituted with R 5 ;
each R 4 is alkylene, alkenylene, alkynylene, heteroalkylene, cycloalkylene, heterocycloalkylene, arylene, or heteroarylene;
each K is O, S, SO, SO 2 , CO, CO 2 , or CONR 3 ;
each R 5 is independently halogen, alkyl, —OR 6 , —N(R 6 ) 2 , —SR 6 , —SOR 6 , —SO 2 R 6 , —CO 2 R 6 , a fluorescent moiety, a radioisotope or a therapeutic agent;
each R 6 is independently —H, alkyl, alkenyl, alkynyl, arylalkyl, cycloalkylalkyl, heterocycloalkyl, a fluorescent moiety, a radioisotope or a therapeutic agent;
R 7 is —H, alkyl, alkenyl, alkynyl, arylalkyl, cycloalkyl, heteroalkyl, cycloalkylalkyl, heterocycloalkyl, cycloaryl, or heterocycloaryl, optionally substituted with R 5 , or part of a cyclic structure with a D residue;
R 8 is —H, alkyl, alkenyl, alkynyl, arylalkyl, cycloalkyl, heteroalkyl, cycloalkylalkyl, heterocycloalkyl, cycloaryl, or heterocycloaryl, optionally substituted with R 5 , or part of a cyclic structure with an E residue;
v and w are independently integers from 1-100;
u is an integer from 1 to 3;
x, y and z are independently integers from 0-10; and
n is an integer from 1-5.
18. The peptidomimetic macrocycle of claim 17 , wherein u is 2.
19. The peptidomimetic macrocycle of claim 18 , having the Formula:
wherein each A, C, D, and E is independently an amino acid;
B is an amino acid, or
L′ is a macrocycle-forming linker of the formula -L 1 ′-L 2 ′-;
and wherein A, B, C, D, and E, taken together with the crosslinked amino acids connected by the macrocycle-forming linkers L and L′, form the amino acid sequence of the peptidomimetic macrocycle;
R 1 ′ and R 2 ′ are independently —H, alkyl, alkenyl, alkynyl, arylalkyl, cycloalkyl, cycloalkylalkyl, heteroalkyl, or heterocycloalkyl, unsubstituted or substituted with halo-;
L 1 ′ and L 2 ′ are independently alkylene, alkenylene, alkynylene, heteroalkylene, cycloalkylene, heterocycloalkylene, cycloarylene, heterocycloarylene, or [—R 4 —K—R 4 —] n , each being optionally substituted with R 5 ;
each K is independently O, S, SO, SO 2 , CO, CO 2 , or CONR 3 ;
R 7 ′ is —H, alkyl, alkenyl, alkynyl, arylalkyl, cycloalkyl, heteroalkyl, cycloalkylalkyl, heterocycloalkyl, cycloaryl, or heterocycloaryl, optionally substituted with R 5 , or part of a cyclic structure with a D residue;
R 8 ′ is —H, alkyl, alkenyl, alkynyl, arylalkyl, cycloalkyl, heteroalkyl, cycloalkylalkyl, heterocycloalkyl, cycloaryl, or heterocycloaryl, optionally substituted with R 5 , or part of a cyclic structure with an E residue;
v′ and w′ are independently integers from 1-100;
x′, y′ and z′ are independently integers from 0-10; and
n is an integer from 1-5.
20. The peptidomimetic macrocycle of claim 19 , wherein the sum of x+y+z is 2, 3 or 6.
21. The peptidomimetic macrocycle of claim 19 , wherein the sum of x′+y′+z′ is 2, 3 or 6.
22. The peptidomimetic macrocycle of claim 17 , wherein each of v and w is independently an integer from 1 to 10, 1 to 15, 1 to 20, or 1 to 25.
23. The peptidomimetic macrocycle of claim 1 , wherein the peptidomimetic macrocycle comprises a crosslinker linking a backbone amino group of a first amino acid to a second amino acid within the peptidomimetic macrocycle.
24. The peptidomimetic macrocycle of claim 23 , wherein the peptidomimetic macrocycle has the formula (II) or (IIa):
wherein:
each A, C, D, and E is independently an amino acid;
B is an amino acid, or
R 1 and R 2 are independently —H, alkyl, alkenyl, alkynyl, arylalkyl, cycloalkyl, cycloalkylalkyl, heteroalkyl, or heterocycloalkyl, unsubstituted or substituted with halo-, or part of a cyclic structure with an E residue;
R 3 is hydrogen, alkyl, alkenyl, alkynyl, arylalkyl, heteroalkyl, cycloalkyl, heterocycloalkyl, cycloalkylalkyl, cycloaryl, or heterocycloaryl, optionally substituted with R 5 ;
L 1 and L 2 are independently alkylene, alkenylene, alkynylene, heteroalkylene, cycloalkylene, heterocycloalkylene, cycloarylene, heterocycloarylene, or [—R 4 —K—R 4 —] n , each being optionally substituted with R 5 ;
and wherein A, B, C, D, and E, taken together with the crosslinked amino acids connected by the macrocycle-forming linker -L 1 -L 2 -, form the amino acid sequence of the peptidomimetic macrocycle which is at least about 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identical to GHRH 1-44, GHRH 1-29 and/or to an amino acid sequence chosen from the group consisting of the amino acid sequences in Table 1, 2 or 4;
each R 4 is alkylene, alkenylene, alkynylene, heteroalkylene, cycloalkylene, heterocycloalkylene, arylene, or heteroarylene;
each K is O, S, SO, SO 2 , CO, CO 2 , or CONR 3 ;
each R 5 is independently halogen, alkyl, —OR 6 , —N(R 6 ) 2 , —SR 6 , —SOR 6 , —SO 2 R 6 , —CO 2 R 6 , a fluorescent moiety, a radioisotope or a therapeutic agent;
each R 6 is independently —H, alkyl, alkenyl, alkynyl, arylalkyl, cycloalkylalkyl, heterocycloalkyl, a fluorescent moiety, a radioisotope or a therapeutic agent;
R 7 is —H, alkyl, alkenyl, alkynyl, arylalkyl, cycloalkyl, heteroalkyl, cycloalkylalkyl, heterocycloalkyl, cycloaryl, or heterocycloaryl, optionally substituted with R 5 ;
v and w are independently integers from 1-100;
u is an integer from 1 to 3;
x, y and z are independently integers from 0-10; and
n is an integer from 1-5.
25. The peptidomimetic macrocycle of claim 1 , having the formula:
wherein:
each A, C, D, and E is independently an amino acid;
B is an amino acid, or
L is a macrocycle-forming linker of the formula -L 1 -L 2 -;
and wherein A, B, C, D, and E, taken together with the crosslinked amino acids connected by the macrocycle-forming linker L, form an amino acid sequence which is at least about 60% identical to GHRH 1-29;
R 1 and R 2 are independently —H, alkyl, alkenyl, alkynyl, arylalkyl, cycloalkyl, cycloalkylalkyl, heteroalkyl, or heterocycloalkyl, unsubstituted or substituted with halo-;
R 3 is hydrogen, alkyl, alkenyl, alkynyl, arylalkyl, heteroalkyl, cycloalkyl, heterocycloalkyl, cycloalkylalkyl, cycloaryl, or heterocycloaryl, optionally substituted with R 5 ;
L 1 and L 2 are independently alkylene, alkenylene, alkynylene, heteroalkylene, cycloalkylene, heterocycloalkylene, cycloarylene, heterocycloarylene, or [—R 4 —K—R 4 —] n , each being optionally substituted with R 5 ;
each R 4 is alkylene, alkenylene, alkynylene, heteroalkylene, cycloalkylene, heterocycloalkylene, arylene, or heteroarylene;
each K is O, S, SO, SO 2 , CO, or CO 2 ;
each R 5 is independently halogen, alkyl, —OR 6 , —N(R 6 ) 2 , —SR 6 , —SOR 6 , —SO 2 R 6 , —CO 2 R 6 , a fluorescent moiety, a radioisotope or a therapeutic agent;
each R 6 is independently —H, alkyl, alkenyl, alkynyl, arylalkyl, cycloalkylalkyl, heterocycloalkyl, a fluorescent moiety, a radioisotope or a therapeutic agent;
R 7 is —H, alkyl, alkenyl, alkynyl, arylalkyl, cycloalkyl, heteroalkyl, cycloalkylalkyl, heterocycloalkyl, cycloaryl, or heterocycloaryl, optionally substituted with R 5 , or part of a cyclic structure with a D residue;
R 8 is —H, alkyl, alkenyl, alkynyl, arylalkyl, cycloalkyl, heteroalkyl, cycloalkylalkyl, heterocycloalkyl, cycloaryl, or heterocycloaryl, optionally substituted with R 5 , or part of a cyclic structure with an E residue;
v and w are independently integers from 1-100;
u is an integer from 1 to 3;
x, y and z are independently integers from 0-10; and
n is an integer from 1-5.
26. The peptidomimetic macrocycle of claim 25 , wherein A, B, C, D, and E, taken together with the crosslinked amino acids connected by the macrocycle-forming linker L, form an amino acid sequence which is at least about 60% identical to an amino acid sequence of Table 1, 2, or 4.
27. A peptidomimetic macrocycle comprising an amino acid sequence of formula:
(SEQ ID NO: 1)
X1-X2-X3-X4-X5-X6-X7-X8-X9-X10-X11-X12-X13-X14-
X15-X16-X17-X18-X19-X20-X21-X22-X23-X24-X25-X26-
X27-X28-X29
wherein:
X1 is Tyr or His;
X2 is Ala, D-Ala, or Val;
X3 is Asp;
X4 is Ala or a crosslinked amino acid;
X5 is Ile;
X6 is Phe;
X7 is Thr;
X8 is Gln, Asn, or a crosslinked amino acid;
X9 is Ser or a crosslinked amino acid;
X10 is Tyr;
X11 is Arg, Ala or Gln;
X12 is Lys, Ala, Gln or a crosslinked amino acid;
X13 is Val or Ile;
X14 is Leu;
X15 is Gly, Ala or a crosslinked amino acid;
X16 is Gln, Glu or a crosslinked amino acid;
X17 is Leu;
X18 is Ser, Tyr or a crosslinked amino acid;
X19 is Ala or a crosslinked amino acid;
X20 is Arg or Gln;
X21 is Lys, Gln or a crosslinked amino acid;
X22 is Leu, Ala, or a crosslinked amino acid;
X23 is Leu;
X24 is Gln, Glu or His;
X25 is Asp, Glu or a crosslinked amino acid;
X26 is Ile;
X27 is Met, Ile, Leu or Nle;
X28 is Ser or a crosslinked amino acid;
X29 is Arg, Ala, Gln or a crosslinked amino acid;
wherein at least one of the crosslinked amino acids is an α,α-disubstituted amino acid;
wherein the peptidomimetic macrocycle comprises at least two macrocycle-forming linkers, each independently connecting at least one pair of amino acids selected from X1-X29;
wherein each macrocycle-forming linker is independently of the formula -L 1 -L 2 -;
L 1 and L 2 are independently alkylene, alkenylene, alkynylene, heteroalkylene, cycloalkylene, heterocycloalkylene, cycloarylene, heterocycloarylene, or [—R 4 —K—R 4 —] n , each being optionally substituted with R 5 ;
each R 4 is alkylene, alkenylene, alkynylene, heteroalkylene, cycloalkylene, heterocycloalkylene, arylene, or heteroarylene;
each K is O, S, SO, SO 2 , CO, or CO 2 ;
each R 5 is independently halogen, alkyl, —OR 6 , —N(R 6 ) 2 , —SR 6 , —SOR 6 , —SO 2 R 6 , —CO 2 R 6 , a fluorescent moiety, a radioisotope or a therapeutic agent; and
each R 6 is independently —H, alkyl, alkenyl, alkynyl, arylalkyl, cycloalkylalkyl, heterocycloalkyl, a fluorescent moiety, a radioisotope or a therapeutic agent.
28. The peptidomimetic macrocycle of claim 27 , wherein each macrocycle-forming linker connects one of the following pairs of amino acids: X4 and X8; X5 and X12; X8 and X12; X8 and X15; X9 and X16; X12 and X16; X12 and X19; X15 and X22; X18 and X25; X21 and X25; X21 and X28; X22 and X29; X25 and X29.
29. The peptidomimetic macrocycle of claim 28 , wherein each macrocycle-forming linker connects one of the following pairs of amino acids: X4 and X8; X5 and X12; X12 and X19; X15 and X22; X18 and X25; X21 and X25; X21 and X28.
30. The peptidomimetic macrocycle of claim, wherein L 1 and L 2 are independently alkylene, alkenylene or alkynylene.
31. The peptidomimetic macrocycle of claim 30 , wherein L 1 and L 2 are independently C 3 -C 10 alkylene or alkenylene.
32. The peptidomimetic macrocycle of claim 30 , wherein L 1 and L 2 are independently C 3 -C 6 alkylene or alkenylene.
33. The peptidomimetic macrocycle of claim 17 , wherein R 1 and R 2 are independently H.
34. The peptidomimetic macrocycle of claim 17 , wherein R 1 and R 2 are independently alkyl.
35. The peptidomimetic macrocycle of claim 17 , wherein R 1 and R 2 are independently methyl.
36. The peptidomimetic macrocycle of claim 17 , wherein R 1 and R 2 are independently alkyl, alkenyl, alkynyl, arylalkyl, cycloalkyl, cycloalkylalkyl, heteroalkyl, or heterocycloalkyl, unsubstituted or substituted with halo-.
37. The peptidomimetic macrocycle of claim 17 , wherein one of R 1 and R 2 is —H, and the other of R 1 and R 2 is alkyl.
38. The peptidomimetic macrocycle of claim 1 , wherein at least one of the first, second, third, and fourth amino acids is the α,α-disubstituted amino acid.