초록

The present invention provides peptidomimetic macrocycles capable of modulating growth hormone levels and methods of using such macrocycles for the treatment of disease.

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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

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; andii) 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 -L1-L2-;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;R1 and R2 are independently —H, alkyl, alkenyl, alkynyl, arylalkyl, cycloalkyl, cycloalkylalkyl, heteroalkyl, or heterocycloalkyl, unsubstituted or substituted with halo-;R3 is hydrogen, alkyl, alkenyl, alkynyl, arylalkyl, heteroalkyl, cycloalkyl, heterocycloalkyl, cycloalkylalkyl, cycloaryl, or heterocycloaryl, optionally substituted with R5;L1 and L2 are independently alkylene, alkenylene, alkynylene, heteroalkylene, cycloalkylene, heterocycloalkylene, cycloarylene, heterocycloarylene, or [—R4—K—R4—]n, each being optionally substituted with R5;each R4 is alkylene, alkenylene, alkynylene, heteroalkylene, cycloalkylene, heterocycloalkylene, arylene, or heteroarylene;each K is O, S, SO, SO2, CO, CO2, or CONR3;each R5 is independently halogen, alkyl, —OR6, —N(R6)2, —SR6, —SOR6, —SO2R6, —CO2R6, a fluorescent moiety, a radioisotope or a therapeutic agent;each R6 is independently —H, alkyl, alkenyl, alkynyl, arylalkyl, cycloalkylalkyl, heterocycloalkyl, a fluorescent moiety, a radioisotope or a therapeutic agent;R7 is —H, alkyl, alkenyl, alkynyl, arylalkyl, cycloalkyl, heteroalkyl, cycloalkylalkyl, heterocycloalkyl, cycloaryl, or heterocycloaryl, optionally substituted with R5, or part of a cyclic structure with a D residue;R8 is —H, alkyl, alkenyl, alkynyl, arylalkyl, cycloalkyl, heteroalkyl, cycloalkylalkyl, heterocycloalkyl, cycloaryl, or heterocycloaryl, optionally substituted with R5, 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; andn 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 -L1′-L2′-;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;R1′ and R2′ are independently —H, alkyl, alkenyl, alkynyl, arylalkyl, cycloalkyl, cycloalkylalkyl, heteroalkyl, or heterocycloalkyl, unsubstituted or substituted with halo-;L1′ and L2′ are independently alkylene, alkenylene, alkynylene, heteroalkylene, cycloalkylene, heterocycloalkylene, cycloarylene, heterocycloarylene, or [—R4—K—R4—]n, each being optionally substituted with R5;each K is independently O, S, SO, SO2, CO, CO2, or CONR3;R7′ is —H, alkyl, alkenyl, alkynyl, arylalkyl, cycloalkyl, heteroalkyl, cycloalkylalkyl, heterocycloalkyl, cycloaryl, or heterocycloaryl, optionally substituted with R5, or part of a cyclic structure with a D residue;R8′ is —H, alkyl, alkenyl, alkynyl, arylalkyl, cycloalkyl, heteroalkyl, cycloalkylalkyl, heterocycloalkyl, cycloaryl, or heterocycloaryl, optionally substituted with R5, 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; andn 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 R1 and R2 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;R3 is hydrogen, alkyl, alkenyl, alkynyl, arylalkyl, heteroalkyl, cycloalkyl, heterocycloalkyl, cycloalkylalkyl, cycloaryl, or heterocycloaryl, optionally substituted with R5;L1 and L2 are independently alkylene, alkenylene, alkynylene, heteroalkylene, cycloalkylene, heterocycloalkylene, cycloarylene, heterocycloarylene, or [—R4—K—R4—]n, each being optionally substituted with R5;and wherein A, B, C, D, and E, taken together with the crosslinked amino acids connected by the macrocycle-forming linker -L1-L2-, 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 R4 is alkylene, alkenylene, alkynylene, heteroalkylene, cycloalkylene, heterocycloalkylene, arylene, or heteroarylene;each K is O, S, SO, SO2, CO, CO2, or CONR3;each R5 is independently halogen, alkyl, —OR6, —N(R6)2, —SR6, —SOR6, —SO2R6, —CO2R6, a fluorescent moiety, a radioisotope or a therapeutic agent;each R6 is independently —H, alkyl, alkenyl, alkynyl, arylalkyl, cycloalkylalkyl, heterocycloalkyl, a fluorescent moiety, a radioisotope or a therapeutic agent;R7 is —H, alkyl, alkenyl, alkynyl, arylalkyl, cycloalkyl, heteroalkyl, cycloalkylalkyl, heterocycloalkyl, cycloaryl, or heterocycloaryl, optionally substituted with R5;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; andn 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 -L1-L2-;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;R1 and R2 are independently —H, alkyl, alkenyl, alkynyl, arylalkyl, cycloalkyl, cycloalkylalkyl, heteroalkyl, or heterocycloalkyl, unsubstituted or substituted with halo-;R3 is hydrogen, alkyl, alkenyl, alkynyl, arylalkyl, heteroalkyl, cycloalkyl, heterocycloalkyl, cycloalkylalkyl, cycloaryl, or heterocycloaryl, optionally substituted with R5;L1 and L2 are independently alkylene, alkenylene, alkynylene, heteroalkylene, cycloalkylene, heterocycloalkylene, cycloarylene, heterocycloarylene, or [—R4—K—R4—]n, each being optionally substituted with R5;each R4 is alkylene, alkenylene, alkynylene, heteroalkylene, cycloalkylene, heterocycloalkylene, arylene, or heteroarylene;each K is O, S, SO, SO2, CO, or CO2;each R5 is independently halogen, alkyl, —OR6, —N(R6)2, —SR6, —SOR6, —SO2R6, —CO2R6, a fluorescent moiety, a radioisotope or a therapeutic agent;each R6 is independently —H, alkyl, alkenyl, alkynyl, arylalkyl, cycloalkylalkyl, heterocycloalkyl, a fluorescent moiety, a radioisotope or a therapeutic agent;R7 is —H, alkyl, alkenyl, alkynyl, arylalkyl, cycloalkyl, heteroalkyl, cycloalkylalkyl, heterocycloalkyl, cycloaryl, or heterocycloaryl, optionally substituted with R5, or part of a cyclic structure with a D residue;R8 is —H, alkyl, alkenyl, alkynyl, arylalkyl, cycloalkyl, heteroalkyl, cycloalkylalkyl, heterocycloalkyl, cycloaryl, or heterocycloaryl, optionally substituted with R5, 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; andn 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-X29wherein: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 -L1-L2-;L1 and L2 are independently alkylene, alkenylene, alkynylene, heteroalkylene, cycloalkylene, heterocycloalkylene, cycloarylene, heterocycloarylene, or [—R4—K—R4—]n, each being optionally substituted with R5;each R4 is alkylene, alkenylene, alkynylene, heteroalkylene, cycloalkylene, heterocycloalkylene, arylene, or heteroarylene;each K is O, S, SO, SO2, CO, or CO2;each R5 is independently halogen, alkyl, —OR6, —N(R6)2, —SR6, —SOR6, —SO2R6, —CO2R6, a fluorescent moiety, a radioisotope or a therapeutic agent; andeach R6 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 L1 and L2 are independently alkylene, alkenylene or alkynylene. 31. The peptidomimetic macrocycle of claim 30, wherein L1 and L2 are independently C3-C10 alkylene or alkenylene. 32. The peptidomimetic macrocycle of claim 30, wherein L1 and L2 are independently C3-C6 alkylene or alkenylene. 33. The peptidomimetic macrocycle of claim 17, wherein R1 and R2 are independently H. 34. The peptidomimetic macrocycle of claim 17, wherein R1 and R2 are independently alkyl. 35. The peptidomimetic macrocycle of claim 17, wherein R1 and R2 are independently methyl. 36. The peptidomimetic macrocycle of claim 17, wherein R1 and R2 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 R1 and R2 is —H, and the other of R1 and R2 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.