Stapled intracellular-targeting antimicrobial peptides to treat infection
Structurally stabilized, e.g., stapled, peptides with the ability to translocate through microbial cell membranes to the interior of microbial cells and exert a biological activity there are provided, as are methods of designing, making and using such peptides.
1. A stapled peptide or a pharmaceutically acceptable salt thereof comprising the amino acid sequence TRSSRAGLQWPVGRVX 1 RLLX 2 K (SEQ ID NO:38), TRSSRAGLQWPX 1 GRVX 2 RLLRK (SEQ ID NO:42), TRSSRAGLX 1 WPVX 2 RVHRLLRK (SEQ ID NO:43), TRX 1 SRAX 2 LQWPVGRVHRLLRK (SEQ ID NO:47), TRSSRAX 3 LQWPVGX 4 VHRLLRK (SEQ ID NO:55), or TRSSX 3 AGLQWPX 4 GRVHRLLRK (SEQ ID NO:57),
wherein each of X 1 , X 2 , X 3 , and X 4 is a non-natural amino acid,
wherein X 1 and X 2 are cross-linked to each other where present, and
wherein X 3 and X 4 are cross-linked to each other where present.
2. The stapled peptide or pharmaceutically acceptable salt thereof of claim 1 , wherein each of X 1 , X 2 , X 3 , and X 4 is independently an α, α-disubstituted non-natural amino acid comprising an olefinic side chain.
3. The stapled peptide or pharmaceutically acceptable salt thereof of claim 1 , wherein each of X 1 and X 2 , and X 4 is (S)-2-(4′-pentenyl)alanine, and wherein X 3 is (R)-2-(7′-octenyl)alanine.
4. The stapled peptide or pharmaceutically acceptable salt thereof of claim 1 , comprising the amino acid sequence TRSSRAGLX 1 WPVX 2 RVHRLLRK (SEQ ID NO:43), TRX 1 SRAX 2 LQWPVGRVHRLLRK (SEQ ID NO:47), or TRSSRAX 3 LQWPVGX 4 VHRLLRK (SEQ ID NO:55), wherein each of X 1 and X 2 , and X 4 is (S)-2-(4′-pentenyl)alanine, and wherein X 3 is (R)-2-(7′-octenyl)alanine.
5. The stapled peptide or pharmaceutically acceptable salt thereof of claim 1 , which is 21 to 30 amino acids in length.
6. The stapled peptide or pharmaceutically acceptable salt thereof of claim 1 , wherein the pharmaceutically acceptable salt is an acetate, a citrate, a fumarate, a maleate, a succinate, a sulfate, or a malonate.
7. The stapled peptide or pharmaceutically acceptable salt thereof of claim 1 , consisting of the amino acid sequence
TRSSRAGLQWPVGRVX 1 RLLX 2 K (SEQ ID NO:38), TRSSRAGLQWPX 1 GRVX 2 RLLRK (SEQ ID NO:42), TRSSRAGLX 1 WPVX 2 RVHRLLRK (SEQ ID NO:43), TRX 1 SRAX 2 LQWPVGRVHRLLRK (SEQ ID NO:47), TRSSRAX 3 LQWPVGX 4 VHRLLRK (SEQ ID NO:55), or TRSSX 3 AGLQWPX 4 GRVHRLLRK (SEQ ID NO:57).
8. The stapled peptide or pharmaceutically acceptable salt thereof of claim 7 , wherein each of X 1 , X 2 , X 3 , and X 4 is independently an α, α-disubstituted non-natural amino acid comprising an olefinic side chain.
9. The stapled peptide or pharmaceutically acceptable salt thereof of claim 7 , wherein each of X 1 and X 2 , and X 4 is (S)-2-(4′-pentenyl)alanine, and wherein X 3 is (R)-2-(7′-octenyl)alanine.
10. The stapled peptide or pharmaceutically acceptable salt thereof of claim 1 , consisting of the amino acid sequence TRSSRAGLX 1 WPVX 2 RVHRLLRK (SEQ ID NO:43), TRX 1 SRAX 2 LQWPVGRVHRLLRK (SEQ ID NO:47), or TRSSRAX 3 LQWPVGX 4 VHRLLRK (SEQ ID NO:55), wherein each of X 1 and X 2 , and X 4 is (S)-2-(4′-pentenyl)alanine, and wherein X 3 is (R)-2-(7′-octenyl)alanine.
11. The stapled peptide or pharmaceutically acceptable salt thereof of claim 1 , wherein the stapled peptide or pharmaceutically acceptable salt thereof is dihydroxylated.
12. The stapled peptide or pharmaceutically acceptable salt thereof of claim 1 , which is linked to an antibiotic or methotrexate.
13. The stapled peptide or pharmaceutically acceptable salt thereof of claim 12 , wherein the antibiotic is a Gram-positive antimicrobial.
14. The stapled peptide or pharmaceutically acceptable salt thereof of claim 13 , wherein the Gram-positive antimicrobial is a beta-lactam drug.
15. A pharmaceutical composition comprising the stapled peptide or pharmaceutically acceptable salt thereof of claim 1 , and a pharmaceutically acceptable carrier.
16. A pharmaceutical composition comprising the stapled peptide or pharmaceutically acceptable salt thereof of claim 3 , and a pharmaceutically acceptable carrier.
17. A pharmaceutical composition comprising the stapled peptide or pharmaceutically acceptable salt thereof of claim 4 , and a pharmaceutically acceptable carrier.
18. A pharmaceutical composition comprising the stapled peptide or pharmaceutically acceptable salt thereof of claim 7 , and a pharmaceutically acceptable carrier.
19. A pharmaceutical composition comprising the stapled peptide or pharmaceutically acceptable salt thereof of claim 10 , and a pharmaceutically acceptable carrier.
20. A method of treating a bacterial infection in a human subject in need thereof, the method comprising administering a therapeutically-effective amount of the stapled peptide or pharmaceutically acceptable salt thereof of claim 1 to the human subject.
21. The method of claim 20 , wherein the bacterial infection is caused by a Gram-positive bacterium.
22. The method of claim 20 , wherein the bacterial infection is caused by a Gram-negative bacterium.
23. The method of claim 20 , wherein the bacterial infection is caused by E. coli, B. cereus, P aeruginosa, S. aureus , or methicillin-resistant S. aureus.
24. The method of claim 20 , wherein the stapled peptide or pharmaceutically acceptable salt thereof comprises the amino acid sequence TRSSRAGLX 1 WPVX 2 RVHRLLRK (SEQ ID NO:43), TRX 1 SRAX 2 LQWPVGRVHRLLRK (SEQ ID NO:47), or TRSSRAX 3 LQWPVGX 4 VHRLLRK (SEQ ID NO:55), wherein each of X 1 and X 2 , and X 4 is (S)-2-(4′-pentenyl)alanine, and wherein X 3 is (R)-2-(7′-octenyl)alanine.
25. The method of claim 20 , further comprising administering to the subject a therapeutically-effective amount of an antibiotic.
26. A method of making a stapled peptide or pharmaceutically acceptable salt thereof, the method comprising:
(a) providing a peptide comprising the amino acid sequence:
TRSSRAGLQWPVGRVX 1 RLLX 2 K (SEQ ID NO:38), TRSSRAGLQWPX 1 GRVX 2 RLLRK (SEQ ID NO:42), TRSSRAGLX 1 WPVX 2 RVHRLLRK (SEQ ID NO:43), TRX 1 SRAX 2 LQWPVGRVHRLLRK (SEQ ID NO:47), TRSSRAX 3 LQWPVGX 4 VHRLLRK (SEQ ID NO:55), or TRSSX 3 AGLQWPX 4 GRVHRLLRK (SEQ ID NO:57), wherein each of X 1 , X 2 , X 3 , and X 4 is an α, α-disubstituted non-natural amino acid comprising an olefinic side chain; and
(b) performing a ring-closing metathesis reaction on the peptide, thereby making the stapled peptide.
27. The method of claim 26 , further comprising formulating the stapled peptide as a sterile pharmaceutical composition.