US 2014/0179593 A1 · METHODS AND COMPOSITIONS FOR INHIBITING NEDDYLATION OF PROTEINS

Abstract

Provided herein is a novel binding pocket within NEDD8 co-E3 proteins that binds NEDD8 E2 enzymes. Particularly at its M-Terminus. Methods are provided for screening for compounds that bind to the disclosed E2-binding pocket in NEDD8 co-E3 proteins. Compounds that bind to the E2-binding pocket and optionally inhibit the activity of NEDD8 co-E3 proteins and pharmaceutical compositions comprising the same are further provided. The NEDD8 co-E3 inhibitors find use, as agents preventing the NEDDylation of a target protein, in inhibiting cell growth and methods for treating cancers, inflammatory disorders, and pathogenic infections. The preferred inhibitors are peptides corresponding to a M-terminal fragment of Dnc1, e.g. MTLASKLKRDD, MLKLRQLQKKKQ, and MIKLFSLKQQKK, which are substituted at the M-Terminus with an uncharged group (e.g. acyl).

Claims35

Dependent claims

An isolated peptide selected from the group consisting of:

a) a peptide having the sequence set forth in SEQ ID NO:
7, 12, 13, 14, 15, or 16 or a fragment thereof;
and
b) a peptide having at least 75% sequence identity to SEQ ID NO:
7, 12, 13, 14, 15, or 16;
wherein the nitrogen atom of the amino terminus of said isolated peptide is covalently bound to at least one —R group, wherein said —R group is uncharged under physiological conditions, provided that said —R group is not an acetyl group, and wherein said isolated peptide binds to an E2-binding pocket of a NEDD8 co-E3 protein.

3 more elements

A method for identifying compounds that bind to the E2-binding pocket in a NEDD8 co-E3 protein, said method comprising comparing the three-dimensional structure of a test compound and a three-dimensional structure of the E2-binding pocket to determine if at least one atom of said test compound will interact with at least one atom of said E2-binding pocket, wherein said three-dimensional structure of the E2-binding pocket is selected from the group consisting of:

a) an E2-binding pocket having the atomic coordinates of Table 2, 3, 8, 9, 10, 11, 18, or 19;
b) a structural variant of the E2-binding pocket of part a), wherein said structural variant comprises a root mean square deviation from the back-bone atoms of the amino acids of Table 2, 3, 8, 9, 10, 11, 18, or 19 of not more than 2 Å;
c) an E2-binding pocket having the atomic coordinates of Tables 2 and 5;
d) a structural variant of the E2-binding pocket of part c), wherein said structural variant comprises a root mean square deviation from the back-bone atoms of the amino acids of Tables 2 and 5 of not more than 2 Å;
e) an E2-binding pocket having the atomic coordinates of Tables 3 and 6;
f) a structural variant of the E2-binding pocket of part e), wherein said structural variant comprises a root mean square deviation from the back-bone atoms of the amino acids of Tables 3 and 6 of not more than 2 Å;
g) an E2-binding pocket having the atomic coordinates of Tables 8 and 13;
h) a structural variant of the E2-binding pocket of part g), wherein said structural variant comprises a root mean square deviation from the back-bone atoms of the amino acids of Tables 8 and 13 of not more than 2 Å;
i) an E2-binding pocket having the atomic coordinates of Tables 9 and 14;
j) a structural variant of the E2-binding pocket of part i), wherein said structural variant comprises a root mean square deviation from the back-bone atoms of the amino acids of Tables 9 and 14 of not more than 2 Å;
k) an E2-binding pocket having the atomic coordinates of Tables 10 and 15;
l) a structural variant of the E2-binding pocket of part k), wherein said structural variant comprises a root mean square deviation from the back-bone atoms of the amino acids of Tables 10 and 15 of not more than 2 Å;
m) an E2-binding pocket having the atomic coordinates of Tables 11 and 16;
n) a structural variant of the E2-binding pocket of part k), wherein said structural variant comprises a root mean square deviation from the back-bone atoms of the amino acids of Tables 11 and 16 of not more than 2 Å;
o) an E2-binding pocket having the atomic coordinates of Tables 18 and 21;
p) a structural variant of the E2-binding pocket of part k), wherein said structural variant comprises a root mean square deviation from the back-bone atoms of the amino acids of Tables 18 and 21 of not more than 2 Å;
q) an E2-binding pocket having the atomic coordinates of Tables 19 and 22;
and
r) a structural variant of the E2-binding pocket of part k), wherein said structural variant comprises a root mean square deviation from the back-bone atoms of the amino acids of Tables 19 and 22 of not more than 2 Å.

16 more elements

A method for identifying compounds that bind to an E2-binding pocket in a NEDD8 co-E3 protein, said method comprising contacting a NEDD8 co-E3 protein with a test compound and determining if said test compound binds to the E2-binding pocket, wherein said E2-binding pocket comprises amino acid residues selected from the group consisting of:

a) an isoleucine corresponding to position 90 of SEQ ID NO:
48, a leucine corresponding to position 93 of SEQ ID NO:
48, a valine corresponding to position 94 of SEQ ID NO:
48, an isoleucine corresponding to position 97 of SEQ ID NO:
48, a leucine corresponding to position 104 of SEQ ID NO:
48, a glutamic acid corresponding to position 105 of SEQ ID NO:
48, an aspartic acid corresponding to position 106 of SEQ ID NO:
48, a leucine corresponding to position 107 of SEQ ID NO:
48, a threonine corresponding to position 109 of SEQ ID NO:
48, a leucine corresponding to position 110 of SEQ ID NO:
48, an alanine corresponding to position 113 of SEQ ID NO:
48, a leucine corresponding to position 121 of SEQ ID NO:
48, a glutamic acid corresponding to position 122 of SEQ ID NO:
48, a leucine corresponding to position 173 of SEQ ID NO:
48, an isoleucine corresponding to position 174 of SEQ ID NO:
48, a leucine corresponding to position 175 of SEQ ID NO:
48, an aspartic acid corresponding to position 176 of SEQ ID NO:
48, a glutamine corresponding to position 189 of SEQ ID NO:
48, a tyrosine corresponding to position 190 of SEQ ID NO:
48, and a leucine corresponding to position 193 of SEQ ID NO:
48;
b) an isoleucine corresponding to position 90 of SEQ ID NO:
48, a leucine corresponding to position 93 of SEQ ID NO:
48, a valine corresponding to position 94 of SEQ ID NO:
48, an isoleucine corresponding to position 97 of SEQ ID NO:
48, a leucine corresponding to position 104 of SEQ ID NO:
48, a glutamic acid corresponding to position 105 of SEQ ID NO:
48, an aspartic acid corresponding to position 106 of SEQ ID NO:
48, a leucine corresponding to position 107 of SEQ ID NO:
48, a threonine corresponding to position 109 of SEQ ID NO:
48, a leucine corresponding to position 110 of SEQ ID NO:
48, an alanine corresponding to position 113 of SEQ ID NO:
48, a leucine corresponding to position 121 of SEQ ID NO:
48, a glutamic acid corresponding to position 122 of SEQ ID NO:
48, a leucine corresponding to position 173 of SEQ ID NO:
48, an isoleucine corresponding to position 174 of SEQ ID NO:
48, a leucine corresponding to position 175 of SEQ ID NO:
48, an aspartic acid corresponding to position 176 of SEQ ID NO:
48, a glutamine corresponding to position 189 of SEQ ID NO:
48, a tyrosine corresponding to position 190 of SEQ ID NO:
48, a leucine corresponding to position 193 of SEQ ID NO:
48, an asparagine corresponding to position 84 of SEQ ID NO:
48;
an aspartic acid corresponding to position 89 of SEQ ID NO:
48, an aspartic acid corresponding to position 91 of SEQ ID NO:
48, a glycine corresponding to position 101 of SEQ ID NO:
48, a tyrosine corresponding to position 102 of SEQ ID NO:
48, an asparagine corresponding to position 103 of SEQ ID NO:
48, and a glutamic acid corresponding to position 186 of SEQ ID NO:
48;
c) an isoleucine corresponding to position 83 of SEQ ID NO:
33, an isoleucine corresponding to position 86 of SEQ ID NO:
33, a glutamine corresponding to position 87 of SEQ ID NO:
33, a proline corresponding to position 97 of SEQ ID NO:
33, an alanine corresponding to position 98 of SEQ ID NO:
33, a serine corresponding to position 99 of SEQ ID NO:
33, a valine corresponding to position 102 of SEQ ID NO:
33, a leucine corresponding to position 103 of SEQ ID NO:
33, an alanine corresponding to position 106 of SEQ ID NO:
33, a glutamine corresponding to position 114 of SEQ ID NO:
33, a cysteine corresponding to position 115 of SEQ ID NO:
33, a phenylalanine corresponding to position 117 of SEQ ID NO:
33, a phenylalanine corresponding to position 164 of SEQ ID NO:
33, an asparagine corresponding to position 167 of SEQ ID NO:
33, a methionine corresponding to position 177 of SEQ ID NO:
33, an alanine corresponding to position 180 of SEQ ID NO:
33, a tyrosine corresponding to position 181 of SEQ ID NO:
33, and a leucine corresponding to position 184 of SEQ ID NO:
33;
d) an isoleucine corresponding to position 83 of SEQ ID NO:
33, an isoleucine corresponding to position 86 of SEQ ID NO:
33, a glutamine corresponding to position 87 of SEQ ID NO:
33, a proline corresponding to position 97 of SEQ ID NO:
33, an alanine corresponding to position 98 of SEQ ID NO:
33, a serine corresponding to position 99 of SEQ ID NO:
33, a valine corresponding to position 102 of SEQ ID NO:
33, a leucine corresponding to position 103 of SEQ ID NO:
33, an alanine corresponding to position 106 of SEQ ID NO:
33, a glutamine corresponding to position 114 of SEQ ID NO:
33, a cysteine corresponding to position 115 of SEQ ID NO:
33, a phenylalanine corresponding to position 117 of SEQ ID NO:
33, a phenylalanine corresponding to position 164 of SEQ ID NO:
33, an asparagine corresponding to position 167 of SEQ ID NO:
33, a methionine corresponding to position 177 of SEQ ID NO:
33, an alanine corresponding to position 180 of SEQ ID NO:
33, a tyrosine corresponding to position 181 of SEQ ID NO:
33, a leucine corresponding to position 184 of SEQ ID NO:
33, an aspartic acid corresponding to position 84 of SEQ ID NO:
33, an alanine corresponding to position 94 of SEQ ID NO:
33, a leucine corresponding to position 95 of SEQ ID NO:
33, and an aspartic acid corresponding to position 96 of SEQ ID NO:
33;
e) a glutamic acid corresponding to position 108 of SEQ ID NO:
39, a methionine corresponding to position 111 of SEQ ID NO:
39, a glutamic acid corresponding to position 112 of SEQ ID NO:
39, a proline corresponding to position 122 of SEQ ID NO:
39, a threonine corresponding to position 123 of SEQ ID NO:
39, a glutamic acid corresponding to position 124 of SEQ ID NO:
39, a valine corresponding to position 127 of SEQ ID NO:
39, a leucine corresponding to position 128 of SEQ ID NO:
39, an alanine corresponding to position 131 of SEQ ID NO:
39, a methionine corresponding to position 139 of SEQ ID NO:
39, a cysteine corresponding to position 140 of SEQ ID NO:
39, a phenylalanine corresponding to position 142 of SEQ ID NO:
39, a phenylalanine corresponding to position 189 of SEQ ID NO:
39, an isoleucine corresponding to position 204 of SEQ ID NO:
39, an alanine corresponding to position 207 of SEQ ID NO:
39, a leucine corresponding to position 208 of SEQ ID NO:
39, and a leucine corresponding to position 211 of SEQ ID NO:
39;
and
f) a glutamic acid corresponding to position 108 of SEQ ID NO:
39, a methionine corresponding to position 111 of SEQ ID NO:
39, a glutamic acid corresponding to position 112 of SEQ ID NO:
39, a proline corresponding to position 122 of SEQ ID NO:
39, a threonine corresponding to position 123 of SEQ ID NO:
39, a glutamic acid corresponding to position 124 of SEQ ID NO:
39, a valine corresponding to position 127 of SEQ ID NO:
39, a leucine corresponding to position 128 of SEQ ID NO:
39, an alanine corresponding to position 131 of SEQ ID NO:
39, a methionine corresponding to position 139 of SEQ ID NO:
39, a cysteine corresponding to position 140 of SEQ ID NO:
39, a phenylalanine corresponding to position 142 of SEQ ID NO:
39, a phenylalanine corresponding to position 189 of SEQ ID NO:
39, an isoleucine corresponding to position 204 of SEQ ID NO:
39, an alanine corresponding to position 207 of SEQ ID NO:
39, a leucine corresponding to position 208 of SEQ ID NO:
39, a leucine corresponding to position 211 of SEQ ID NO:
39;
a glutamic acid corresponding to position 109 of SEQ ID NO:
39;
a valine corresponding to position 120 of SEQ ID NO:
39;
and an aspartic acid at position 121 of SEQ ID NO:
39.

132 more elements

Dependencies are inferred from the claim text.

Methods and compositions for inhibiting neddylation of proteins

Life of this patent

Methods and compositions for inhibiting neddylation of proteins

Life of this patent

Abstract

Claims35