Bicyclic peptidyl inhibitors
The present disclosure provides a large combinatorial library of cell-permeable bicyclic peptides. The bicyclic peptides described herein include the first ring consisted of randomized peptide sequences for potential binding to a target of interest while the second ring featured a family of different cell-penetrating motifs, for both cell penetration and target binding. The library was screened against the IκB kinase α/β (IKKα/β)-binding domain of NF-κB essential modulator (NEMO), resulting in the discovery of several cell-permeable bicyclic peptides which inhibited the NEMO-IKKβ interaction, thereby selectively inhibiting canonical NF-κB signaling in mammalian cells and the proliferation of cisplatin-resistant ovarian cancer cells.
1. A bicyclic polypeptide comprising Formula 5A or 5B:
wherein:
CPP comprises a sequence according to Formula 2:
(AA u ) m -AA 1 -AA 2 -AA 3 -AA 4 -(AA z ) n 2
wherein:
each of AA 1 , AA 2 , AA 3 , AA 4 , AA u , and AA, are independently selected from a D or L amino acid,
m and n are independently selected from a number from 0 to 6; and
wherein:
at least two of AA u , AA′, AA 2 , AA 3 , AA 4 , and AA, are independently arginine, and
at least two of AA u , AA 1 , AA 2 , AA 3 , AA 4 , and AA z are independently a hydrophobic amino acid;
Xm is a 4-7 amino acid peptide comprising a sequence selected from any one of GWIY (SEQ ID NO: 1), GWIYA (SEQ ID NO: 2), GWIYa (SEQ ID NO: 50), AGWIY (SEQ ID NO: 3), aGWIY (SEQ ID NO: 51), AWIYA (SEQ ID NO: 4), GAIYA (SEQ ID NO: 5), GWAYA (SEQ ID NO: 6), GWIAA (SEQ ID NO: 7), GWIYA (SEQ ID NO: 8), GAIAA (SEQ ID NO: 9), GAAAA (SEQ ID NO: 10), or the inverse of any one of the aforementioned Xm sequences (SEQ ID NOs: 52-63);
L is a linker moiety; and
each of R 1 , R 2 , and R 3 independently comprise an amide, an ester, a triazole, or a combination thereof.
2. A bicyclic polypeptide comprising Formula 5C or 5D:
wherein:
CPP comprises a sequence according to Formula 2:
(AA u ) m -AA 1 -AA 2 -AA 3 − AA 4 -(AA z ) n
wherein:
each of AA 1 , AA 2 , AA 3 , AA 4 , AA u , and AA z are independently selected from a D or L amino acid, and
m and n are independently selected from a number from 0 to 6; and
wherein:
at least two of AA u , AA 1 , AA 2 , AA 3 , AA 4 , and AA z are independently arginine, and
at least two of AA u , AA′, AA 2 , AA 3 , AA 4 , and AA, are independently a hydrophobic amino acid;
Xm is a 4-7 amino acid peptide comprising a sequence selected from any one of GWIY (SEQ ID NO: 1), GWIYA (SEQ ID NO: 2), GWIYa (SEQ ID NO: 50), AGWIY (SEQ ID NO: 3), aGWIY (SEQ ID NO: 51), AWIYA (SEQ ID NO: 4), GAIYA (SEQ ID NO: 5), GWAYA (SEQ ID NO: 6), GWIAA (SEQ ID NO: 7), GWIYA (SEQ ID NO: 8), GAIAA (SEQ ID NO: 9), GAAAA (SEQ ID NO: 10), or the inverse of any one of the aforementioned Xm sequences (SEQ ID NOs: 52-63);
AA L at each instance is an amino acid;
p is selected from a number from 0 to 3;
L is a linker moiety; and
each of R 1 , R 2 , and R 3 comprise an amide, an ester, a triazole, or a combination thereof.
3. The bicyclic polypeptide of claim 1 , wherein both the CPP sequence and the Xm sequence participate in binding to NEMO.
4. The bicyclic polypeptide of claim 1 , wherein the bicyclic polypeptide binds to the IKKα/β-binding domain on NEMO.
5. The bicyclic polypeptide of claim 1 , wherein the bicyclic polypeptide inhibits NEMO-IKKα/β interaction by at least about 10%.
6. The bicyclic polypeptide of claim 1 , wherein the bicyclic polypeptide has an IC50 of about 10 μM or less when measured for the NEMO-IKKα/β interaction.
7. The bicyclic polypeptide of claim 6 , wherein the bicyclic polypeptide has an IC50 of about 1.0 μM or less when measured for the NEMO-IKKα/β interaction.
8. The bicyclic polypeptide of claim 1 , wherein the CPP comprises a sequence according to Formula 3A-D:
wherein:
each of AA H1 and AA H2 are independently a hydrophobic amino acid;
each of AA U and AA Z are independently any amino acid; and
m and n are independently selected from a number from 0 to 6.
9. A pharmaceutical composition comprising the bicyclic polypeptide of claim 1 and one or more pharmaceutically acceptable excipients.
10. A cell comprising the bicyclic polypeptide of claim 1 .
11. The bicyclic polypeptide of claim 2 , wherein the bicyclic polypeptide has an IC50 of about 10 μM or less when measured for the NEMO-IKKα/β interaction.