Peptide compounds and their use as protease substrates

• US 20040018561A1
• Filed: 05/10/2003
• Published: 01/29/2004
• Est. Priority Date: 05/10/2002
• Status: Abandoned Application

• Current Assignee: Pharmacia Corporation (Pfizer Inc.)
• Original Assignee: Pharmacia Corporation (Pfizer Inc.)
• Inventors: Zhang, Kun, DeCrescenzo, Gary A., Rico, Joseph G., Howard, Carol P.
• Application Number:
  US10/435,486
• Publication Number:
  US 20040018561A1
• Time in Patent Office: Days
• US Class Current: 435/7.1
• CPC Class Codes: C07K 14/78 Connective tissue peptides, e.g. collagen, elastin, laminin, fibronectin, vitronectin or cold insoluble globulin [CIG]
• CPC Class Codes: C07K 7/06 having 5 to 11 amino acids
• CPC Class Codes: C07K 7/08 having 12 to 20 amino acids gastrins C07K14/595; somatostatins C07K14/655; melanotropins C07K14/68
• CPC Class Codes: C12Q 1/37 involving peptidase or proteinase
• CPC Class Codes: C12Q 2337/00 N-linked chromogens for determinations of peptidases and proteinases
• CPC Class Codes: G01N 2333/96486 Metalloendopeptidases (3.4.24)
• CPC Class Codes: G01N 2500/02 Screening involving studying the effect of compounds C on the interaction between interacting molecules A and B (e.g. A = enzyme and B = substrate for A, or A = receptor and B = ligand for the receptor)

Abstract

This invention is directed generally to peptide compounds and salts, and particularly to peptide compounds and salts that are useful as protease substrates, such as matrix metalloprotease (“MMP”) substrates. This invention also is directed to methods for making such compounds and salts, as well as amino acids that may, for example, be used in such methods. This invention is further directed to methods for using such compounds and salts to, for example, evaluate the effectiveness of potential protease inhibitors and to detect or monitor a disease associated with protease activity.

Claims

1. A compound or a salt thereof, wherein:

the compound comprises a peptide and corresponds in structure to Formula (I);

aa_(i)-X—

Y-aa_(j)-Z  

 

(I);

aa_(i) comprises a sequence of i amino acids at the N-terminus of the peptide;

aa_(j) comprises a sequence of j amino acids at the C-terminus of the peptide;

i is an integer from 0 to 5;

j is an integer from 1 to 6;

a fluorophore is covalently attached to the peptide on one side of the X—

Y bond, and at least one of a fluorescence quencher and a ligand is covalently attached to the peptide on the other side of the X—

Y bond;

X comprises an MMP recognition sequence;

X is not Pro-Gln-Gln, Pro-Tyr-Ala, or Pro-Val-Glu;

Y comprises an amino acid; and

Z is;

a hydroxyl group at the C-terminus of the peptide, or

a protecting group at the C-terminus of the peptide.

2. A compound or salt thereof according to claim 1, wherein Y comprises a side chain comprising at least 8 non-hydrogen atoms.

3. A compound or salt thereof according to claim 2, wherein Y comprises a side chain comprising at least 10 non-hydrogen atoms.

4. A compound or salt thereof according to claim 3, wherein Y comprises a side chain comprising at least 11 non-hydrogen atoms.

5. A compound or salt thereof according to claim 4, wherein Y comprises a side chain comprising from 11 to 15 non-hydrogen atoms.

6. A compound or salt thereof according to claim 4, wherein Y comprises a side chain comprising at least 12 non-hydrogen atoms.

7. A compound or salt thereof according to claim 1, wherein the compound or salt is characterized in that MMP-2, MMP-9, or MMP-13 reacts with the compound or salt to cleave the compound or salt at the bond between X and Y when the compound or salt is combined with the MMP-2, MMP-9, or MMP-13.

8. A compound or salt thereof according to claim 7, wherein:

k_cat/K_m of the compound or salt with at least one of MMP-1 and MMP-7 is no greater than about 0.5×

10<sup>−

4</sup>M<sup>−

1</sup>s<sup>−

1</sup>, and

k_cat/K_m of the compound or salt with at least one of MMP-2, MMP-9, and MMP-13 is at least about 5×

10<sup>−

4</sup>M<sup>−

1</sup>s<sup>−

1</sup>.

9. A compound or salt thereof according to claim 8, wherein:

k_cat/K_m of the compound or salt with at least one of MMP-1 and MMP-7 is no greater than about 10<sup>−

5</sup> M<sup>−

1</sup>s<sup>−

1</sup>, and

k_cat/K_m of the compound or salt with at least one of MMP-2, MMP-9, and MMP-13 is at least about 50×

10<sup>−

4</sup>M<sup>−

1</sup>s<sup>−

1</sup>.

10. A compound or salt thereof according to claim 7, wherein the compound or salt is characterized in that the rate of reaction of the compound or salt with MMP-1 or MMP-7 is about zero.

11. A compound or salt thereof according to claim 10, wherein the compound or salt is characterized in that the rate of reaction of the compound or salt with MMP-1 and MMP-7 is about zero.

12. A compound or salt thereof according to claim 7, wherein Y is selected from the group consisting of 3-(2-napthyl)-L-alanine, O-benzyl-L-tyrosine, 6-(benzyloxy)-L-norleucine, S-(3-phenylpropyl)-L-cysteine, 6-phenoxy-L-norleucine and S-(4-methoxybenzyl)-L-cysteine.

13. A compound or salt thereof according to claim 7, wherein Y is selected from the group consisting of 6-(benzyloxy)-L-norleucine and 6-phenoxy-L-norleucine.

14. A compound or salt thereof according to claim 7, wherein the recognition sequence of X is selected from the group consisting of Ala-Gln-Gly, Ala-Met-His, Asn-Gln-Gly, Asp-Lys-Glu, Dnp-Gln-Gly, Dnp-Leu-Gly, Ile-Gly-Phe, Lys-Pro-Asn, Pro-Arg-Gly, Pro-Asp-Gly, Pro-Gln -Tyr, Pro-Gln-Ala, Pro-Gln-Glu, Pro-Gln-Gly, Pro-Gln-His, Pro-Gln-Leu, Pro-Gln-Met, Pro-Gln-Phe, Pro-Gln-Pro, Pro-Gln-Val, Pro-Glu-Asn, Pro-Glu-Gly, Pro-His-Gly, Pro-Leu-Ala, Pro-Leu-Gly, Pro-Met-Gly, Pro-Tyr-Gly, Pro-Val-Ala, Pro-Ser-Glu-Asn, Ser-Gly-Asn, Ser-His-Ser, Ser-Ile-Pro, Thr-Glu-Lys, Tyr-Arg-Trp, Tyr-His-Ser, and Pro-Leu-MeCys.

15. A compound or salt thereof according to claim 7, wherein Y comprises a side chain comprising at least 8 non-hydrogen atoms.

16. A compound or salt thereof according to claim 15, wherein Y comprises a side chain comprising 11 non-hydrogen atoms.

17. A compound or salt thereof according to claim 16, wherein the side chain corresponds in structure to Formula (17-1):

18. A compound or salt thereof according to claim 16, wherein the side chain corresponds in structure to Formula (18-1):

19. A compound or salt thereof according to claim 16, wherein the side chain corresponds in structure to Formula (19-1):

20. A compound or salt thereof according to claim 16, wherein the side chain corresponds in structure to Formula (20-1):

21. A compound or salt thereof according to claim 15, wherein Y comprises a side chain comprising at least 12 non-hydrogen atoms.

22. A compound or salt thereof according to claim 21, wherein the side chain corresponds in structure to Formula (22-1):

23. A compound or salt thereof according to claim 22, wherein the compound corresponds in structure to Formula (23-1):

24. A compound or salt thereof according to claim 22, wherein the compound corresponds in structure to Formula (24-1):

25. A compound or salt thereof according to claim 15, wherein Y comprises a side chain comprising 15 non-hydrogen atoms.

26. A compound or salt thereof according to claim 25, wherein the side chain corresponds in structure to Formula (26-1):

27. A compound or salt thereof according to claim 7, wherein i is zero.

28. A compound or salt thereof according to claim 7, wherein i is 1.

29. A compound or salt thereof according to claim 7, wherein j is from 3 to 5.

30. A compound or salt thereof according to claim 29, wherein i is zero or 1.

31. A compound or salt thereof according to claim 7, wherein aa(i) or aa(j) comprises an amino acid selected from the group consisting of arginine, lysine, aspartic acid, and glutamic acid.

32. A compound or salt thereof according to claim 31, wherein:

aa(i) comprises an amino acid selected from the group consisting of arginine, lysine, aspartic acid, and glutamic acid; and

aa(j) comprises an amino acid selected from the group consisting of arginine, lysine, aspartic acid, and glutamic acid.

33. A compound or salt thereof according to claim 7, wherein the compound or salt has a water solubility that is greater than a K_m of MMP-1, MMP-2, MMP-3, MMP-7, MMP-8, MMP-9, MMP-10, MMP-13, MMP-14, or MMP-26.

34. A compound or salt thereof according to claim 7, wherein the compound or salt has a water solubility of at least about 20 μ

M in 1% DMSO at pH 7.5.

35. A compound or salt thereof according to claim 34, wherein the compound or salt has a water solubility of at least about 100 μ

M in 1% DMSO at pH 7.5.

36. A compound or salt thereof according to claim 7, wherein the compound or salt comprises at least one amino acid having a log D contribution of less than about zero.

37. A compound or salt thereof according to claim 7, wherein the amino acid having a log D contribution of less than about zero is selected from the group consisting of D-Arginine, L-Arginine, D-Asparagine, L-Asparagine, D-Glutamic Acid, L-Glutamic acid, D-Histidine, L-Histidine, D-Glutamine, L-Glutamine, D-Lysine, L-Lysine, D-Serine, L-Serine, D-Threonine, L-Threonine, Glycine, D-Alanine, L-Alanine, D-citrulline, L-citrulline, D-ornithine, and L-ornithine.

38. A compound or salt thereof according to claim 7, wherein the fluorophore is a coumarin.

39. A compound or salt thereof according to claim 38, wherein the coumarin is substituted N-2-[(7-methoxy-2-oxo-2H-chromen-4-yl)acetyl].

40. A compound or salt thereof according to claim 38, wherein the coumarin is N-2-[(7-methoxy-2-oxo-2H-chromen-4-yl)acetyl].

41. A compound or salt thereof according to claim 7, wherein the fluorophore is a fluorescein.

42. A compound or salt thereof according to claim 41, wherein the fluorescein is substituted N-6-(4-{[3-carboxy-4-(6-hydroxy-3-oxo-3H-xanthen-9-yl)phenyl]amino}-6-chloro-1,3,5-triazin-2-yl).

43. A compound or salt thereof according to claim 41, wherein the fluorescein is N-6-(4-{[3-carboxy-4-(6-hydroxy-3-oxo-3H-xanthen-9-yl)phenyl]amino}-6-chloro-1,3,5-triazin-2-yl).

44. A compound or salt thereof according to claim 7, wherein a fluorophore is covalently attached to the peptide on one side of the X—

Y bond, and a fluorescence quencher is covalently attached to the peptide on the other side of the X—

Y bond.

45. A compound or salt thereof according to claim 44, wherein the fluorescence quencher is substituted (2,4-dinitrophenyl)amino.

46. A compound or salt thereof according to claim 44, wherein the fluorescence quencher is (2,4-dinitrophenyl)amino.

47. A compound or salt thereof according to claim 7, wherein a fluorophore is covalently attached to the peptide on one side of the X—

Y bond, and a ligand is covalently attached to the peptide on the other side of the X—

Y bond.

48. A compound or salt thereof according to claim 47, wherein the ligand is bound non-covalently to a solid support.

49. A compound or salt thereof according to claim 47, wherein the ligand is substituted biotin.

50. A compound or salt thereof according to claim 47, wherein the ligand is biotin.

51. A compound or salt thereof according to claim 47, wherein the ligand is substituted N-2-[(5-(2-oxohexahydro-1H-thieno[3,4-d]imidazol-4-yl)pentanoyl].

52. A compound or salt thereof according to claim 47, wherein the ligand is N-2-[(5-(2-oxohexahydro-1H-thieno[3,4-d]imidazol-4-yl)pentanoyl].

53. A compound or salt thereof according to claim 47, wherein the ligand is substituted epsilon amino caproic acid.

54. A compound or salt thereof according to claim 47, wherein the ligand is epsilon amino caproic acid.

55. A compound or salt thereof according to claim 7, wherein the compound or salt comprises at least one D-amino acid.

56. A compound or salt thereof according to claim 7, wherein the compound or salt comprises at least one amino acid selected from the group consisting of a beta-amino acid and a gamma-amino acid.

57. A compound or salt thereof according to claim 7, wherein Z is a protecting group.

58. A compound or salt thereof according to claim 57, wherein Z is an optionally-substituted amine.

59. A compound or salt thereof according to claim 1, wherein the compound corresponds in structure to a formula selected from group consisting of:

60. A compound or salt thereof according to claim 1, wherein the compound is selected from the group consisting of, SEQ ID NO:

8, SEQ ID NO;

20, SEQ ID NO;

21, SEQ ID NO;

22, SEQ ID NO;

23, SEQ ID NO;

24, and SEQ ID NO;

25.

72. A method for determining the activity of a matrix metalloprotease, wherein the method comprises combining the matrix metalloprotease with a compound or salt recited in claim 1.

73. A method according to claim 72, wherein the compound or salt is a compound or salt recited in claim 59.

74. A method according to claim 72, wherein the matrix metalloprotease is MMP-2.

75. A method according to claim 72, wherein the matrix metalloprotease is MMP-9.

76. A method according to claim 72, wherein the matrix metalloprotease is MMP-13.

77. A method according to claim 72, wherein the method comprises combining:

one or more of MMP-2, MMP-9, and MMP-13;

one or more of MMP-1 and MMP-7; and

the compound or salt recited in claim 1.

78. A method for ex-vivo detection or monitoring of a disease associated with a pathological matrix metalloprotease level, wherein the method comprises measuring a cleavage of a compound or salt recited in claim 1 at the bond between X and Y.

79. A method according to claim 78, wherein the compound or salt is a compound or salt recited in claim 59.

80. A method according to claim 78, wherein the matrix metalloprotease is MMP-2.

81. A method according to claim 78, wherein the matrix metalloprotease is MMP-9.

82. A method according to claim 78, wherein the matrix metalloprotease is MMP-13.

83. A method for determining the activity of a matrix metalloprotease in a biological sample, wherein the method comprises:

combining the biological sample with a compound or salt recited in claim 1 to form a mixture, and

analyzing the mixture for the presence of a reaction product of the compound or salt with the matrix metalloprotease.

84. A method according to claim 83, wherein the compound or salt is a compound or salt recited in claim 59.

85. A method according to claim 83, wherein the matrix metalloprotease is MMP-2.

86. A method according to claim 83, wherein the matrix metalloprotease is MMP-9.

87. A method according to claim 83, wherein the matrix metalloprotease is MMP-13.

88. A method for measuring inhibitory activity of a prospective inhibitor of a matrix metalloprotease, wherein the method comprises:

combining the following to form a mixture;

a compound or salt recited in claim 1,

the prospective inhibitor, and

the matrix metalloprotease; and

analyzing the mixture for the presence of a reaction product of the compound or salt with the matrix metalloprotease.

89. A method according to claim 88, wherein the compound or salt is a compound or salt recited in claim 59.

90. A method according to claim 88, wherein the matrix metalloprotease is MMP-2.

91. A method according to claim 88, wherein the matrix metalloprotease is MMP-9.

92. A method according to claim 88, wherein the matrix metalloprotease is MMP-13.

139. A kit for detecting or monitoring a disease associated with pathological activity of a matrix metalloprotease, wherein the kit comprises a compound or salt recited in claim 1.

140. A kit according to claim 139, wherein the compound or salt is a compound or salt recited in claim 59.

145. A kit for evaluating the effectiveness of a prospective inhibitor of a matrix metalloprotease, wherein the kit comprises a compound or salt recited in claim 1.

61. A compound or a salt thereof, wherein:

the compound comprises a peptide and corresponds in structure to Formula (I);

aa_(i)-X—

Y-aa_(j)-Z  

 

(I);

aa_(i) comprises a sequence of i amino acids at the N-terminus of the peptide;

aa_(j) comprises a sequence of j amino acids at the C-terminus of the peptide;

the peptide comprises a D-amino acid;

i is an integer from 0 to 5;

j is an integer from 1 to 6;

a fluorophore is covalently attached to the peptide on one side of the X—

Y bond, and at least one of a fluorescence quencher and a ligand is covalently attached to the peptide on the other side of the X—

Y bond;

X comprises an MMP recognition sequence;

Y comprises a bond or amino acid; and

Z is;

a hydroxyl group at the C-terminus of the peptide, or

a protecting group at the C-terminus of the peptide.

62. A compound or salt thereof according to claim 61, wherein the recognition sequence of X is selected from the group consisting of Ala-Gln-Gly, Ala-Met-His, Asn-Gln-Gly, Asp-Lys-Glu, Dnp-Gln-Gly, Dnp-Leu-Gly, Ile-Gly-Phe, Lys-Pro-Asn, Pro-Arg-Gly, Pro-Asp-Gly, Pro-Gln -Tyr, Pro-Gln-Ala, Pro-Gln-Gln, Pro-Gln-Glu, Pro-Gln-Gly, Pro-Gln-His, Pro-Gln-Leu, Pro-Gln-Met, Pro-Gln-Phe, Pro-Gln-Pro, Pro-Gln-Val, Pro-Glu-Asn, Pro-Glu-Gly, Pro-His-Gly, Pro-Leu-Ala, Pro-Leu-Gly, Pro-Met-Gly, Pro-Tyr-Ala, Pro-Tyr-Gly, Pro-Val-Ala, Pro-Val-Glu, Pro-Ser-Glu-Asn, Ser-Gly-Asn, Ser-His-Ser, Ser-Ile-Pro, Thr-Glu-Lys, Tyr-Arg-Trp, Tyr-His-Ser, and Pro-Leu-MeCys.

63. A compound or salt thereof according to claim 61, wherein the compound corresponds in structure to a formula selected from group consisting of:

93. A method for determining the activity of a matrix metalloprotease, wherein the method comprises combining the matrix metalloprotease with a compound or salt recited in claim 61.

94. A method according to claim 93, wherein the compound or salt is a compound or salt recited in claim 63.

95. A method according to claim 93, wherein the matrix metalloprotease is MMP-2.

96. A method according to claim 93, wherein the matrix metalloprotease is MMP-9.

97. A method according to claim 93, wherein the matrix metalloprotease is MMP-13.

98. A method according to claim 93, wherein the method comprises combining:

one or more of MMP-2, MMP-9, and MMP-13;

one or more of MMP-1 or MMP-7; and

the compound or salt recited in claim 61.

99. A method for ex-vivo detection or monitoring of a disease associated with a pathological matrix metalloprotease level, wherein the method comprises measuring a cleavage of a compound or salt recited in claim 61 at the bond between X and Y.

100. A method according to claim 99, wherein the compound or salt is a compound or salt recited in claim 63.

101. A method according to claim 99, wherein the matrix metalloprotease is MMP-2.

102. A method according to claim 99, wherein the matrix metalloprotease is MMP-9.

103. A method according to claim 99, wherein the matrix metalloprotease is MMP-13.

104. A method for determining the activity of a matrix metalloprotease in a biological sample, wherein the method comprises:

combining the biological sample with a compound or salt recited in claim 61 to form a mixture, and

analyzing the mixture for the presence of a reaction product of the compound or salt with the matrix metalloprotease.

105. A method according to claim 104, wherein the compound or salt is a compound or salt recited in claim 63.

106. A method according to claim 104, wherein the matrix metalloprotease is MMP-2.

107. A method according to claim 104, wherein the matrix metalloprotease is MMP-9.

108. A method according to claim 104, wherein the matrix metalloprotease is MMP-13.

109. A method for measuring inhibitory activity of a prospective inhibitor of a matrix metalloprotease, wherein the method comprises:

combining the following to form a mixture;

a compound or salt recited in claim 61,

the prospective inhibitor, and

the matrix metalloprotease; and

analyzing the mixture for the presence of a reaction product of the compound or salt with the matrix metalloprotease.

110. A method according to claim 109, wherein the compound or salt is a compound or salt recited in claim 63.

111. A method according to claim 109, wherein the matrix metalloprotease is MMP-2.

112. A method according to claim 109, wherein the matrix metalloprotease is MMP-9.

113. A method according to claim 109, wherein the matrix metalloprotease is MMP-13.

141. A kit for detecting or monitoring a disease associated with pathological activity of a matrix metalloprotease, wherein the kit comprises a compound or salt recited in claim 61.

142. A kit according to claim 141, wherein the compound or salt is a compound or salt recited in claim 63.

146. A kit for evaluating the effectiveness of a prospective inhibitor of a matrix metalloprotease, wherein the kit comprises a compound or salt recited in claim 61.

64. A compound or a salt thereof, wherein:

the compound comprises a peptide, and

the peptide comprises an amino acid selected from the group consisting of phenyloxynorleucine and benzyloxynorleucine.

65. A compound or salt thereof according to claim 64, wherein the compound or salt is characterized in that MMP-2, MMP-9, or MMP-13 reacts with the compound or salt to cleave the compound or salt at a bond between two amino acids when the compound or salt is combined with the MMP-2, MMP-9, or MMP-13.

66. A compound or salt thereof according to claim 65, wherein the compound or salt comprises phenyloxynorleucine.

67. A compound or salt thereof according to claim 66, wherein the compound or salt is characterized in that MMP-2, MMP-9, or MMP-13 reacts with the compound or salt to cleave the compound or salt at a bond between the phenyloxynorleucine and another amino acid when the compound or salt is combined with the MMP-2, MMP-9, or MMP-13.

68. A compound or salt thereof according to claim 66, wherein the compound corresponds in structure to Formula (68-1):

69. A compound or salt thereof according to claim 65, wherein the compound or salt comprises benzyloxynorleucine.

70. A compound or salt thereof according to claim 69, wherein the compound or salt is characterized in that MMP-2, MMP-9, or MMP-13 reacts with the compound or salt to cleave the compound or salt at a bond between the benzyloxynorleucine and another amino acid when the compound or salt is combined with the MMP-2, MMP-9, or MMP-13.

71. A compound or salt thereof according to claim 69, wherein the compound corresponds in structure to a formula selected from group consisting of:

114. A method for determining the activity of a matrix metalloprotease, wherein the method comprises combining the matrix metalloprotease with a compound or salt recited in claim 64.

115. A method according to claim 114, wherein the compound or salt is a compound or salt recited in claim 68 or 71.

116. A method according to claim 114, wherein the matrix metalloprotease is MMP-2.

117. A method according to claim 114, wherein the matrix metalloprotease is MMP-9.

118. A method according to claim 114, wherein the matrix metalloprotease is MMP-13.

119. A method according to claim 114, wherein the method comprises combining:

one or more of MMP-2, MMP-9, and MMP-13;

one or more of MMP-1 and MMP-7; and

the compound or salt recited in claim 64.

120. A method for ex-vivo detection or monitoring of a disease associated with a pathological matrix metalloprotease level, wherein the method comprises measuring a cleavage of a compound or salt recited in claim 64.

121. A method according to claim 120, wherein the compound or salt is a compound or salt recited in claim 68 or 71.

122. A method according to claim 120, wherein the matrix metalloprotease is MMP-2.

123. A method according to claim 120, wherein the matrix metalloprotease is MMP-9.

124. A method according to claim 123, wherein the disease is an eye disease.

125. A method according to claim 124, wherein the disease is glaucoma.

126. A method according to claim 124, wherein the disease is selected from the group consisting of macular degeneration and diabetic macular edema.

127. A method according to claim 120, wherein the matrix metalloprotease is MMP-13.

128. A method for determining the activity of a matrix metalloprotease in a biological sample, wherein the method comprises:

combining the biological sample with a compound or salt recited in claim 64 to form a mixture, and

analyzing the mixture for the presence of a reaction product of the compound or salt with the matrix metalloprotease.

129. A method according to claim 128, wherein the compound or salt is a compound or salt recited in claim 68 or 71.

130. A method according to claim 128, wherein the matrix metalloprotease is MMP-2.

131. A method according to claim 128, wherein the matrix metalloprotease is MMP-9.

132. A method according to claim 131, wherein the biological sample comprises a fluid or tissue from an eye.

133. A method according to claim 128, wherein the matrix metalloprotease is MMP-13.

134. A method for measuring inhibitory activity of a prospective inhibitor of a matrix metalloprotease, wherein the method comprises:

combining the following to form a mixture;

a compound or salt recited in claim 64,

the prospective inhibitor, and

the matrix metalloprotease; and

analyzing the mixture for the presence of a reaction product of the compound or salt with the matrix metalloprotease.

135. A method according to claim 134, wherein the compound or salt is a compound or salt recited in claim 68 or 71.

136. A method according to claim 134, wherein the matrix metalloprotease is MMP-2.

137. A method according to claim 134, wherein the matrix metalloprotease is MMP-9.

138. A method according to claim 134, wherein the matrix metalloprotease is MMP-13.

143. A kit for detecting or monitoring a disease associated with pathological activity of a matrix metalloprotease, wherein the kit comprises a compound or salt recited in claim 64.

144. A kit according to claim 143, wherein the compound or salt is a compound or salt recited in claim 68 or 71.

147. A kit for evaluating the effectiveness of a prospective inhibitor of a matrix metalloprotease, wherein the kit comprises a compound or salt recited in claim 64.

148. A compound or a salt thereof, wherein:

the compound corresponds in structure to Formula (II);

n is zero or 1, and

R¹ and R² are independently selected from the group consisting of hydrogen and a nitrogen protecting group.

149. A compound or salt thereof according to claim 148, wherein the compound corresponds in structure to Formula (IIA):

150. A compound or salt thereof according to claim 149, wherein the compound corresponds in structure to a formula selected from the group consisting of: