IP Library › Granted Patent US 12,359,192
Granted Patent B2
US 12,359,192 · App. 16/774,440 · Granted Jul 15, 2025

Methods and compositions for display of macrocyclic peptides

Inventor: Rudi Fasan (Rochester, NY)
Assignee: University of Rochester
C12N15/1037C07K14/005C07K14/245C07K14/395C12N9/93C12N15/11C07K2319/40
View Patent ↗
Loading inventors, assignments & file history…
Monitor This Case
Get email alerts when status or documents change.
Order Certified Copies
Most orders are placed with the USPTO same day — all within 24 business hours.
Order via The Patent Place →
Pre-filled with this patent's details
Quick Facts
Patent No.
US 12,359,192
App. No.
16/774,440
Granted
Jul 15, 2025
Kind
B2
Abstract

Methods and compositions are provided for the display of genetically encoded macrocyclic peptides on a biological surface. Also provided are nucleic acid molecules, polypeptides, and methods for generating combinatorial libraries of macrocyclic peptides displayed on a biological surface. These methods can be used to produce and screen vast libraries of conformationally constrained peptides in a high-throughput manner, from which macrocyclic peptides with a desired property can be selected and identified.

Claims (61)

1. A macrocyclic peptide library display system, comprising at least one artificial nucleic acid molecule encoding for a polypeptide of structure:

(AA) m -Z-(AA) n -Cys-(AA) p   (I)

or

(AA) m -Cys-(AA) n -Z-(AA) p   (II)

or

(AA) m -Cys-(AA) n -Z2-(AA) o -Cys-(AA) p   (V)

wherein:

i. (AA) m is an N-terminal amino acid or peptide sequence,

ii. Z is a non-canonical amino acid carrying a side-chain functional group FG 1 , FG 1 being a functional group selected from the group consisting of —(CH 2 ) n X, where X is F, Cl, Br, or I and n is an integer number from 1 to 10; —C(O)CH 2 X, where X is F, Cl, Br, or I; —CH(R′)X, where X is F, Cl, Br, or I; —C(O)CH(R′)X, where X is F, Cl, Br, or I; —OCH 2 CH 2 X, where X is F, Cl, Br, or I; —C(O)CH═C═C(R′)(R″); —SO 2 C(R′)═C(R′)(R″); —C(O)C(R′)═C(R′)(R″); —C(R′)═C(R′)C(O)OR′; —C(R′)═C(R′)C(O)N(R′)(R″); —C(R′)═C(R′)—CN; —C(R′)═C(R′)—NO 2 ; —C≡C—C(O)OR′; —C≡C—C(O)N(R′)(R″); unsubstituted or substituted oxirane; unsubstituted or substituted aziridine; 1,2-oxathiolane 2,2-dioxide; 4-fluoro-1,2-oxathiolane 2,2-dioxide; and 4,4-difluoro-1,2-oxathiolane 2,2-dioxide, where each R′ and R″ is independently H, an aliphatic, a substituted aliphatic, an aryl, or a substituted aryl group,

iii. Z2 is a non-canonical amino acid carrying two side-chain functional groups FG 1 and FG 2 , wherein each of FG 1 and FG 2 is a functional group independently selected from the group consisting of —(CH 2 ) n X, where X is F, Cl, Br, or I and n is an integer number from 1 to 10; —C(O)CH 2 X, where X is F, Cl, Br, or I; —CH(R′)X, where X is F, Cl, Br, or I; —C(O)CH(R′)X, where X is F, Cl, Br, or I; —OCH 2 CH 2 X, where X is F, Cl, Br, or I; —C(O)CH═C═C(R′)(R″); —SO 2 C(R′)═C(R′)(R″); —C(O)C(R′)═C(R′)(R″); —C(R′)═C(R′)C(O)OR′; —C(R′)═C(R′)C(O)N(R′)(R″); —C(R′)═C(R′)—CN; —C(R′)═C(R′)—NO 2 ; —C≡C—C(O)OR′; —C≡C—C(O)N(R′)(R″); unsubstituted or substituted oxirane; unsubstituted or substituted aziridine; 1,2-oxathiolane 2,2-dioxide; 4-fluoro-1,2-oxathiolane 2,2-dioxide; and 4,4-difluoro-1,2-oxathiolane 2,2-dioxide, where each R′ and R″ is independently H, an aliphatic, a substituted aliphatic, an aryl, or a substituted aryl group,

iv. (AA) n is a target peptide sequence,

v. (AA) o is a second target peptide sequence,

vi. (AA) p is a C-terminal amino acid or peptide sequence; and

vii. wherein at least one of (AA) p and (AA) m comprises an amino acid sequence of a polypeptide for presentation of the macrocyclic peptide on an outer surface of a cell or phage particle,

wherein the functional group FG 1 , and whenever present, FG 2 , react with the side-chain sulfhydryl group (—SH) of the cysteine (Cys) residue(s), thereby producing the macrocyclic peptide, and further wherein the at least one nucleic acid molecule is incorporated into an expression system that allows for the incorporation of the non-canonical amino acid Z or Z2 into an expressed polypeptide.

2. The system of claim 1 wherein Z is an amino acid of structure:

wherein FG 1 is a functional group selected from the group consisting of —(CH 2 ) n X, where X is F, Cl, Br, or I and n is an integer number from 1 to 10; —C(O)CH 2 X, where X is F, Cl, Br, or I; —CH(R′)X, where X is F, Cl, Br, or I; —C(O)CH(R′)X, where X is F, Cl, Br, or I; —OCH 2 CH 2 X, where X is F, Cl, Br, or I; —C(O)CH═C═C(R′)(R″); —SO 2 C(R′)═C(R′)(R″); —C(O)C(R′)═C(R′)(R″); —C(R′)═C(R′)C(O)OR′; —C(R′)═C(R′)C(O)N(R′)(R″); —C(R′)═C(R′) CN; —C(R′)═C(R′)—NO 2 ; —C≡C—C(O)OR′; —C≡C—C(O)N(R′)(R″); unsubstituted or substituted oxirane, unsubstituted or substituted aziridine; 1,2-oxathiolane 2,2-dioxide; 4-fluoro-1,2-oxathiolane 2,2-dioxide; and 4,4-difluoro-1,2-oxathiolane 2,2-dioxide; where each R′ and R″ is independently H, an aliphatic, a substituted aliphatic, an aryl, or a substituted aryl group;

wherein Y is a linker group selected from the group consisting of aliphatic, aryl, substituted aliphatic, substituted aryl, heteroatom-containing aliphatic, heteroatom-containing aryl, substituted heteroatom-containing aliphatic, substituted heteroatom-containing aryl, alkoxy, and aryloxy groups.

3. The system of claim 2 wherein Z is an amino acid of structure (IV) and Y is a linker group selected from the group consisting of C 1 -C 24 alkyl, C 1 -C 24 substituted alkyl, C 1 -C 24 substituted heteroatom-containing alkyl, C 1 -C 24 substituted heteroatom-containing alkyl, C 2 -C 24 alkenyl, C 2 -C 24 substituted alkenyl, C 2 -C 24 substituted heteroatom-containing alkenyl, C 2 -C 24 substituted heteroatom-containing alkenyl, C 5 -C 24 aryl, C 5 -C 24 substituted aryl, C 5 -C 24 substituted heteroatom-containing aryl, C 5 -C 24 substituted heteroatom-containing aryl, C 1 -C 24 alkoxy, and C 5 -C 24 aryloxy groups.

4. The system of claim 3 wherein Y is a linker group selected from the group consisting of —CH 2 —C 6 H 4 —, —CH 2 —C 6 H 4 —O—, —CH 2 —C 6 H 4 NH—, —(CH 2 ) 4 —, —(CH 2 ) 4 NH—, —(CH 2 ) 4 NHC(O)—, and —(CH 2 ) 4 NHC(O)O—.

5. The system of claim 1 wherein the amino acid Z is selected from the group consisting of 4-(2-bromoethoxy)-phenylalanine, 3-(2-bromoethoxy)-phenylalanine, 4-(2-chloroethoxy)-phenylalanine, 4-(4-bromobutoxy)-phenylalanine, 4-(4-chlorobutoxy)-phenylalanine, 3-(4-bromobutoxy)-phenylalanine, 3-(4-bromobutoxy)-phenylalanine, 3-(2-chloroethoxy)-phenylalanine, 4-(1-bromoethyl)-phenylalanine, 3-(1-bromoethyl)-phenylalanine, 4-(aziridin-1-yl)-phenylalanine, 3-(aziridin-1-yl)-phenylalanine, 4-acrylamido-phenylalanine, 3-acrylamido-phenylalanine, 4-(2-fluoro-acetamido)-phenylalanine, 3-(2-fluoro-acetamido)-phenylalanine, 4-(2-chloro-acetamido)-phenylalanine, 3-(2-chloro-acetamido)-phenylalanine, 4-(2-bromo-acetamido)-phenylalanine, 3-(2-bromo-acetamido)-phenylalanine, 4-(acrylamido)-phenylalanine, 3-(acrylamido)-phenylalanine, 4-(vinylsulfonamido)-phenylalanine, 3-(vinylsulfonamido)-phenylalanine, 3-(2-fluoro-acetyl)-phenylalanine, 4-(2-fluoro-acetyl)-phenylalanine, N ε -((2-bromoethoxy)carbonyl)-lysine, N ε -((2-chloroethoxy)carbonyl)-lysine, N-(buta-2,3-dienoyl)-lysine, N ε -acryl-lysine, N ε -crotonyl-lysine, N ε -(2-fluoro-acetyl)-lysine, N ε -(2-chloro-acetyl)-lysine, N ε -(2-bromoacetyl)-lysine, and N ε -vinylsulfonyl-lysine.

6. The system of claim 1 wherein Z2 is an amino acid of structure:

wherein each of FG 1 and FG 2 is a functional group independently selected from the group consisting of —(CH 2 ) n X, where X is F, Cl, Br, or I and n is an integer number from 1 to 10; —C(O)CH 2 X, where X is F, Cl, Br, or I; —CH(R′)X, where X is F, Cl, Br, or I; —C(O)CH(R′)X, where X is F, Cl, Br, or I; —OCH 2 CH 2 X, where X is F, Cl, Br, or I; —C(O)CH═C═C(R′)(R″); —SO 2 C(R′)═C(R′)(R″); —C(O)C(R′)═C(R′)(R″); —C(R′)═C(R′)C(O)OR′; —C(R′)═C(R′)C(O)N(R′)(R″); —C(R′)═C(R′)—CN; —C(R′)═C(R′)—NO 2 , —C≡C—C(O)OR′; —C≡C—C(O)N(R′)(R″); unsubstituted or substituted oxirane; unsubstituted or substituted aziridine; 1,2-oxathiolane 2,2-dioxide; 4-fluoro-1,2-oxathiolane 2,2-dioxide; and 4,4-difluoro-1,2-oxathiolane 2,2-dioxide, where each R′ and R″ is independently H, an aliphatic, a substituted aliphatic, an aryl, or a substituted aryl group;

wherein Y 2 , Y 3 , and L are linker groups selected from the group consisting of aliphatic, aryl, substituted aliphatic, substituted aryl, heteroatom-containing aliphatic, heteroatom-containing aryl, substituted heteroatom-containing aliphatic, substituted heteroatom-containing aryl, alkoxy, and aryloxy groups.

7. The system of claim 6 wherein Z2 is an amino acid of structure (VI) and Y 2 is a linker group selected from the group consisting of C 1 -C 24 alkyl, C 1 -C 24 substituted alkyl, C 1 -C 24 substituted heteroatom-containing alkyl, C 1 -C 24 substituted heteroatom-containing alkyl, C 2 -C 24 alkenyl, C 2 -C 24 substituted alkenyl, C 2 -C 24 substituted heteroatom-containing alkenyl, C 2 -C 24 substituted heteroatom-containing alkenyl, C 5 -C 24 aryl, C 5 -C 24 substituted aryl, C 5 -C 24 substituted heteroatom-containing aryl, C 5 -C 24 substituted heteroatom-containing aryl, C 1 -C 24 alkoxy, and C 5 -C 24 aryloxy groups.

8. The system of claim 7 wherein Y is a linker group selected from the group consisting of —CH 2 —C 6 H 4 —, —CH 2 —C 6 H 4 —O—, —CH 2 —C 6 H 4 —NH—, —CH 2 —C 6 H 4 —OCH 2 —, —(CH 2 ) 4 NH—, —(CH 2 ) 4 NHC(O)—, —(CH 2 ) 4 NHC(O)O—, —(CH 2 ) 4 NHC(O)OCH 2 ,

9. The system of claim 1 wherein the amino acid Z2 is selected from the group consisting of 3,5-bis(2-bromoethoxy)-phenylalanine, 3,5-bis(2-chloroethoxy)-phenylalanine, 3,5-bis(4-bromobutoxy)-phenylalanine, 3,5-bis(4-chlorobutoxy)-phenylalanine, 3,5-bis(1-bromoethyl)-phenylalanine, 3,5-bis(4-acrylamido)-phenylalanine, 3,5-bis(2-chloro-acetamido)-phenylalanine, 3,5-bis(2-bromo-acetamido)-phenylalanine, 3,5-bis(vinylsulfonamido)-phenylalanine, 3,5-bis(aziridin-1-yl)-phenylalanine, 3,5-bis-acrylamido-phenylalanine, 3,5-bis(2-fluoro-acetamido)-phenylalanine, 3,5-bis(2-fluoro-acetyl)-phenylalanine, 4-((1,3-dibromopropan-2-yl)oxy)-phenylalanine, 4-((1,3-dichloropropan-2-yl)oxy)-phenylalanine, N ε -(((1,3-dibromopropan-2-yl)oxy)carbonyl)-lysine, N ε -(((1,3-dichloropropan-2-yl)oxy)carbonyl)-lysine, 4-(2,3-dibromopropoxy)-phenylalanine, 3-(2,3-dibromopropoxy)-phenylalanine, 4-(2,3-dichloropropoxy)-phenylalanine, 3-(2,3-dichloropropoxy)-phenylalanine, N ε -((2,3-dibromopropoxy)carbonyl)-lysine, N ε -((2,3-dichloropropoxy)carbonyl)-lysine, N ε -bis-(acryl)-lysine, N ε -bis-(crotonyl)-lysine, N ε -bis-(2-fluoro-acetyl)-lysine, N ε -bis-(2-chloro-acetyl)-lysine, N ε -bis-(2-bromoacetyl)-lysine, and N ε -bis-(vinylsulfonyl)-lysine.

10. The system of claim 1 , wherein the codon encoding for Z or Z2 is an amber stop codon TAG, an ochre stop codon TAA, an opal stop codon TGA, or a four base codon.

11. The system of claim 1 , wherein the expression system comprises:

an aminoacyl-tRNA synthetase polypeptide or an engineered variant thereof that is at least 90% identical to SEQ ID NO:77, 78, 79, or 80; and

a transfer RNA molecule encoded by a polynucleotide that is at least 90% identical to SEQ ID NO:101, 105, 109, 113, or 117.

12. The system of claim 1 , wherein the presentation peptide comprised within the N-terminal tail polypeptide, (AA) m , comprises at least one polypeptide sequence selected from the group consisting of a T7 phage protein 10A (SEQ ID NO:138), T7 phage protein 10B (SEQ ID NO:139), E. coli NlpA (SEQ ID NO:140), E. coli OmpC (SEQ ID NO:141), E. coli FadL (SEQ ID NO:142), E. coli Lpp-OmpA (SEQ ID NO:143), E. coli PgsA (SEQ ID NO:144), E. coli EaeA (SEQ ID NO:145), S. cerevisiae Aga2p (SEQ ID NO:146), S. cerevisiae Flo1p (SEQ ID NO:147), S. cerevisiae Cwp1p (SEQ ID NO:217), S. cerevisiae Cwp2p (SEQ ID NO:218), S. cerevisiae Tip1p (SEQ ID NO:219), S. cerevisiae Sed1p (SEQ ID NO:220), S. cerevisiae YCR89w (SEQ ID NO:221), S. cerevisiae Tir1 (SEQ ID NO:222), human NF-κB p50 protein (SEQ ID NO:148), M13 phage coat protein pIII leader sequence (SEQ ID NO:149), M13 phage coat protein pVIII leader sequence (SEQ ID NO:150), M13 phage protein pVI (SEQ ID NO:151), M13 phage protein pIII (SEQ ID NO:154), Snap-tag (SEQ ID NO:152), Clip-Tag (SEQ ID NO:153), a barcode sequence, a pelB leader sequence (SEQ ID NO:216.

13. The system of claim 1 , wherein the presentation peptide comprised within the C-terminal tail polypeptide, (AA) p , comprises at least one polypeptide sequence selected from the group consisting of a M13 phage coat protein pIII (SEQ ID NO:154), M13 phage coat protein pVIII (SEQ ID NO:155), M13 phage coat protein pIX (SEQ ID NO:214), M13 phage coat protein pVII (SEQ ID NO:215), RepA protein (SED ID NO: 156), S. cerevisiae Aga1p (SEQ ID NO:157), Snap-tag (SEQ ID NO:152), Clip-Tag (SEQ ID NO:153), P2A protein (SED ID NO: 158), a barcode sequence.

14. The system of claim 1 , wherein the outer biological surface is selected from a phage surface and a cell surface.

15. The system of claim 14 , wherein the phage is a M13 phage.

16. The system of claim 14 , wherein the cell is selected from the group consisting of a bacterial, a yeast, an insect, and a mammalian cell.

17. The system of claim 1 , wherein at least one of polypeptides (AA) n , (AA) n , (AA) m , or (AA) p , is fully or partially genetically randomized.

18. A method for generating a macrocyclic peptide display library for use in the system of claim 1 , the method comprising

a) providing at least one artificial nucleic acid molecule encoding for a macrocyclic polypeptide of structure:

(AA) m -Z-(AA) n -Cys-(AA) p (I)

or

(AA) m -Cys-(AA) n -Z-(AA) p   (II)

or

(AA) m -Cys-(AA) n -Z2-(AA) o -Cys-(AA) p   (V)

wherein:

i. (AA) m is an N-terminal amino acid or peptide sequence,

ii. Z is a non-canonical amino acid carrying a side-chain functional group FG 1 , FG 1 being a functional group selected from the group consisting of —(CH 2 ) n X, where X is F, Cl, Br, or I and n is an integer number from 1 to 10; —C(O)CH 2 X, where X is F, Cl, Br, or I; —CH(R′)X, where X is F, Cl, Br, or I; —C(O)CH(R′)X, where X is F, Cl, Br, or I; —OCH 2 CH 2 X, where X is F, Cl, Br, or I; —C(O)CH═C═C(R′)(R″); —SO 2 C(R′)═C(R′)(R″); —C(O)C(R′)═C(R′)(R″); —C(R′)═C(R′)C(O)OR′; —C(R′)═C(R′)C(O)N(R′)(R″); —C(R′)═C(R′)—CN; —C(R′)═C(R′)—NO 2 ; —C≡C—C(O)OR′; —C≡C—C(O)N(R′)(R″); unsubstituted or substituted oxirane; unsubstituted or substituted aziridine; 1,2-oxathiolane 2,2-dioxide; 4-fluoro-1,2-oxathiolane 2,2-dioxide; and 4,4-difluoro-1,2-oxathiolane 2,2-dioxide, where each R′ and R″ is independently H, an aliphatic, a substituted aliphatic, an aryl, or a substituted aryl group,

iii. Z2 is a non-canonical amino acid carrying two side-chain functional groups FG 1 and FG 2 , wherein each of FG 1 and FG 2 is a functional group independently selected from the group consisting of —(CH 2 ) n X, where X is F, Cl, Br, or I and n is an integer number from 1 to 10; —C(O)CH 2 X, where X is F, Cl, Br, or I; —CH(R′)X, where X is F, Cl, Br, or I; —C(O)CH(R′)X, where X is F, Cl, Br, or I; —OCH 2 CH 2 X, where X is F, Cl, Br, or I; —C(O)CH═C═C(R′)(R″); —SO 2 C(R′)═C(R′)(R″); —C(O)C(R′)═C(R′)(R″); —C(R′)═C(R′)C(O)OR′; —C(R′)═C(R′)C(O)N(R′)(R″); —C(R′)═C(R′)—CN; —C(R′)═C(R′)—NO 2 ; —C≡C—C(O)OR′; —C≡C—C(O)N(R′)(R″); unsubstituted or substituted oxirane; unsubstituted or substituted aziridine; 1,2-oxathiolane 2,2-dioxide; 4-fluoro-1,2-oxathiolane 2,2-dioxide; and 4,4-difluoro-1,2-oxathiolane 2,2-dioxide, where each R′ and R″ is independently H, an aliphatic, a substituted aliphatic, an aryl, or a substituted aryl group,

iv. (AA) n is a target peptide sequence,

v. (AA) o is a second target peptide sequence,

vi. (AA) p is a C-terminal amino acid or peptide sequence, and

vii. wherein at least one of (AA) p and (AA) m comprises an amino acid sequence of a polypeptide, for presentation of the macrocyclic peptide on an outer surface of a cell or phage particle;

b) fully or partially randomizing at least one of polypeptides (AA) n , (AA) o , (AA) m , and (AA) p , to generate a plurality of unique macrocyclic peptide encoding nucleic acid molecules;

c) introducing the plurality of nucleic acid molecules into an expression system that allows for the incorporation of the non-canonical amino acid Z or Z2 into the polypeptide; and

d) expressing the nucleic acid molecule in said expression system, thereby producing the polypeptide; and allowing the functional group FG 1 , and whenever present, FG 2 , to react with the side-chain sulfhydryl group (—SH) of the cysteine (Cys) residue(s), thereby producing a plurality of display macrocyclic peptides anchored on an outer biological surface of a host display organism, wherein each host display organism contains a nucleic acid molecule encoding for the macrocyclic peptide displayed on its outer biological surface.

19. A method for displaying a macrocyclic peptide on an outer biological surface, the method comprising:

a) expressing at least one nucleic acid molecule of the system of claim 1 , thereby producing the polypeptide; and

b) allowing the functional group FG 1 , and whenever present, FG 2 , to react with the side-chain sulfhydryl group (—SH) of the cysteine (Cys) residue(s), thereby producing the macrocyclic peptide anchored on the outer biological surface.

20. A method for screening a macrocyclic peptide display library, the method comprising:

a) contacting the macrocyclic peptide library display system of claim 1 with a target molecule; and

b) selecting macrocyclic peptides that have a desired property based on interaction or lack thereof with the target molecule.

Assignments (1)
ASSIGNMENT OF ASSIGNOR'S INTEREST Recorded Apr 4, 2024
From: FASAN, RUDI
To: UNIVERSITY OF ROCHESTER
Reel/Frame 067001/0081 →
Continuity (3)
Continuation In Part 15107387
Provisional Application 61920181 · Dec 23, 2013
Related Publication 20200299675A1 · Sep 24, 2020
References Cited (75)
US 5525488A · Mason · 1996 [cited by applicant]
US 6906176B2 · Ley · 2005 [cited by examiner]
US 7105341B2 · Kinsella · 2006 [cited by applicant]
US 7235626B1 · Cochran · 2007 [cited by applicant]
US 7252952B2 · Lorens · 2007 [cited by applicant]
US 7354756B1 · Benkovic · 2008 [cited by applicant]
US 7378263B2 · Schultz · 2008 [cited by examiner]
US 8986953B2 · Fasan · 2015 [cited by applicant]
US 20060166319A1 · Chan · 2006 [cited by examiner]
US 20150064207A1 · Titball · 2015 [cited by examiner]
US 20160355552A1 · Fasan · 2016 [cited by applicant]
EP 2141175A1 · 2010 [cited by applicant]
EP 2647721A1 · 2013 [cited by applicant]
Smith et al., “Modular Assembly of Macrocyclic Organo-Peptide Hybrids Using Synthetic and Genetically Encoded Precursors” 50 Angewandte Chemie International Edition 5075-5080 (Year: 2011). [cited by examiner]
Frost et al., “Design, synthesis, and diversfication of riobsomally derived peptide macrocycles” 23 Current Opinion in Structural Biology 571-580 (Year: 2013). [cited by examiner]
Tang YQ, Yuan J, Osapay G, Osapay K, Tran D, Miller CJ, Ouellette AJ, Selsted Me. A cyclic antimicrobial peptide produced in primate leukocytes by the ligation of two truncated alpha-defensins. Science. Oct. 15, 1999;28… [cited by applicant]
Tian F, Tsao ML, Schultz PG. A phage display system with unnatural amino acids. J Am Chem Soc. Dec. 15, 2004;126 (49):15962-3. [cited by applicant]
Touati J, Angelini A, Hinner MJ, Heinis C. Enzymatic cyclisation of peptides with a transglutaminase. Chembiochem. Jan. 3, 2011;12(1):38-42. [cited by applicant]
Walensky LD, Kung AL, Escher I, Malia TJ, Barbuto S, Wright RD, Wagner G, Verdine GL, Korsmeyer SJ. Activation of apoptosis in vivo by a hydrocarbon-stapled BH3 helix. Science. Sep. 3, 2004;305(5689):1466-70. [cited by applicant]
Wang D, Liao W, Arora PS. Enhanced metabolic stability and protein-binding properties of artificial alpha helices derived from a hydrogen-bond surrogate: application to Bcl-xL. Angew Chem Int Ed Engl. Oct. 14, 2005;44(4… [cited by applicant]
Wang L, Xie J, Schultz PG. Expanding the genetic code. Annu Rev Biophys Biomol Struct. 2006;35:225-49. [cited by applicant]
White CJ, Yudin AK. Contemporary strategies for peptide macrocyclization. Nat Chem. Jun. 23, 2011;3(7):509-24. [cited by applicant]
Young DD, Young TS, Jahnz M, Ahmad I, Spraggon G, Schultz PG. An evolved aminoacyl-tRNA synthetase with atypical polysubstrate specificity. Biochemistry. Mar. 22, 2011;50(11):1894-900. [cited by applicant]
Young TS, Ahmad I, Yin JA, Schultz PG. An enhanced system for unnatural amino acid mutagenesis in [cited by applicant]
Young TS, Young DD, Ahmad I, Louis JM, Benkovic SJ, Schultz PG. Evolution of cyclic peptide protease inhibitors. Proc Natl Acad Sci U S A. Jul. 5, 2011;108(27):11052-6. [cited by applicant]
Abbas A, Xing B, Loh TP. Allenamides as orthogonal handles for selective modification of cysteine in peptides and proteins. Angew Chem Int Ed Engl. Jul. 14, 2014;53(29):7491-4. [cited by applicant]
Angelini A, Heinis C. Post-translational modification of genetically encoded polypeptide libraries. Curr Opin Chem Biol. Jun. 2011;15(3):355-61. [cited by applicant]
Bionda N, Cryan AL, Fasan R. Bioinspired strategy for the ribosomal synthesis of thioether-bridged macrocyclic peptides in bacteria. ACS Chem Biol. Sep. 19, 2014;9(9):2008-13. [cited by applicant]
Boder ET, Raeeszadeh-Sarmazdeh M, Price JV. Engineering antibodies by yeast display. Arch Biochem Biophys. Oct. 15, 2012;526(2):99-106. [cited by applicant]
Bosma T, Rink R, Moosmeier MA, Moll GN. Genetically Encoded Libraries of Constrained Peptides. Chembiochem. Jul. 15, 2019;20(14):1754-1758. [cited by applicant]
Broders O, Breitling F, Dübel S. Hyperphage. Improving antibody presentation in phage display. Methods Mol Biol. 2003;205:295-302. [cited by applicant]
Chen S, Rentero Rebollo I, Buth SA, Morales-Sanfrutos J, Touati J, Leiman PG, Heinis C. Bicyclic peptide ligands pulled out of cysteine-rich peptide libraries. J Am Chem Soc. May 1, 2013;135(17):6562-9. [cited by applicant]
Cheng L, Naumann TA, Horswill AR, Hong SJ, Venters BJ, Tomsho JW, Benkovic SJ, Keiler KC. Discovery of antibacterial cyclic peptides that inhibit the ClpXP protease. Protein Sci. Aug. 2007;16(8):1535-42. [cited by applicant]
Deiters A, Schultz PG. In vivo incorporation of an alkyne into proteins in [cited by applicant]
Dias RL, Fasan R, Moehle K, Renard A, Obrecht D, Robinson JA. Protein ligand design: from phage display to synthetic protein epitope mimetics in human antibody Fc-binding peptidomimetics. J Am Chem Soc. Mar. 1, 2006;128… [cited by applicant]
Driggers EM, Hale SP, Lee J, Terrett NK. The exploration of macrocycles for drug discovery—an underexploited structural class. Nat Rev Drug Discov. Jul. 2008;7(7):608-24. [cited by applicant]
Fairlie DP, Tyndall JD, Reid RC, Wong AK, Abbenante G, Scanlon MJ, March DR, Bergman DA, Chai CL, Burkett BA. Conformational selection of inhibitors and substrates by proteolytic enzymes: implications for drug design an… [cited by applicant]
Frost JR, Smith JM, Fasan R. Design, synthesis, and diversification of ribosomally derived peptide macrocycles. Curr Opin Struct Biol. Aug. 2013;23(4):571-80. [cited by applicant]
Frost JR, Vitali F, Jacob NT, Brown MD, Fasan R. Macrocyclization of organo-peptide hybrids through a dual bio-orthogonal ligation: insights from structure-reactivity studies. Chembiochem. Jan. 2, 2013;14(1): 147-60. [cited by applicant]
Hamamoto T, Sisido M, Ohtsuki T, Taki M. Synthesis of a cyclic peptide/protein using the NEXT—A reaction followed by cyclization. Chem Commun (Camb). Aug. 28, 2011;47(32):9116-8. [cited by applicant]
Heinis C, Rutherford T, Freund S, Winter G. Phage-encoded combinatorial chemical libraries based on bicyclic peptides. Nat Chem Biol. Jul. 2009;5(7):502-7. [cited by applicant]
Henchey LK, Porter JR, Ghosh I, Arora PS. High specificity in protein recognition by hydrogen-bond-surrogate α-helices: selective inhibition of the p53/MDM2 complex. Chembiochem. Oct. 18, 2010;11(15):2104-7. [cited by applicant]
Horswill AR, Savinov SN, Benkovic SJ. A systematic method for identifying small-molecule modulators of protein-protein interactions. Proc Natl Acad Sci U S A. Nov. 2, 2004;101(44):15591-6. [cited by applicant]
Katsara M, Tselios T, Deraos S, Deraos G, Matsoukas MT, Lazoura E, Matsoukas J, Apostolopoulos V. Round and round we go: cyclic peptides in disease. Curr Med Chem. 2006;13(19):2221-32. [cited by applicant]
Katz BA. Binding to protein targets of peptidic leads discovered by phage display: crystal structures of streptavidin-bound linear and cyclic peptide ligands containing the HPQ sequence. Biochemistry. Nov. 28, 1995;34(4… [cited by applicant]
Klabunde T, Sharma S, Telenti A, Jacobs WR Jr., Sacchettini JC. Crystal structure of GyrA intein from [cited by applicant]
Kobayashi T, Nureki O, Ishitani R, Yaremchuk A, Tukalo M, Cusack S, Sakamoto K, Yokoyama S. Structural basis for orthogonal tRNA specificities of tyrosyl-tRNA synthetases for genetic code expansion. Nat Struct Biol. Jun… [cited by applicant]
Ladner RC, Sato AK, Gorzelany J, de Souza M. Phage display-derived peptides as therapeutic alternatives to antibodies. Drug Discov Today. Jun. 15, 2004;9(12):525-9. [cited by applicant]
Linciano S, Pluda S, Bacchin A, Angelini A. Molecular evolution of peptides by yeast surface display technology. Medchemcomm. Jul. 10, 2019;10(9):1569-1580. [cited by applicant]
Liu CC, Mack AV, Tsao ML, Mills JH, Lee HS, Choe H, Farzan M, Schultz PG, Smider VV. Protein evolution with an expanded genetic code. Proc Natl Acad Sci U S A. Nov. 18, 2008;105(46):17688-93. [cited by applicant]
Liu CC, Schultz PG. Adding new chemistries to the genetic code. Annu Rev Biochem. 2010;79:413-44. [cited by applicant]
Löfblom J. Bacterial display in combinatorial protein engineering. Biotechnol J. Sep. 2011;6(9):1115-29. [cited by applicant]
Marsault E, Peterson ML. Macrocycles are great cycles: applications, opportunities, and challenges of synthetic macrocycles in drug discovery. J Med Chem. Apr. 14, 2011;54(7):1961-2004. [cited by applicant]
Millward SW, Takahashi TT, Roberts RW. A general route for post-translational cyclization of mRNA display libraries. J Am Chem Soc. Oct. 19, 2005;127(41):14142-3. [cited by applicant]
Naumann TA, Savinov SN, Benkovic SJ. Engineering an affinity tag for genetically encoded cyclic peptides. Biotechnol Bioeng. Dec. 30, 2005;92(7):820-30. [cited by applicant]
Naumann TA, Tavassoli A, Benkovic SJ. Genetic selection of cyclic peptide Dam methyltransferase inhibitors. Chembiochem. Jan. 25, 2008;9(2):194-7. [cited by applicant]
Obrecht D, Robinson JA, Bernardini F, Bisang C, DeMarco SJ, Moehle K, Gombert FO. Recent progress in the discovery of macrocyclic compounds as potential anti-infective therapeutics. Curr Med Chem. 2009;16(1):42-65. [cited by applicant]
Owens AE, de Paola I, Hansen WA, Liu YW, Khare SD, Fasan R. Design and Evolution of a Macrocyclic Peptide Inhibitor of the Sonic Hedgehog/Patched Interaction. J Am Chem Soc. Sep. 13, 2017;139(36):12559-12568. [cited by applicant]
Passioura T, Katoh T, Goto Y, Suga H. Selection-based discovery of druglike macrocyclic peptides. Annu Rev Biochem. 2014;83:727-52. [cited by applicant]
Rezai T, Bock JE, Zhou MV, Kalyanaraman C, Lokey RS, Jacobson MP. Conformational flexibility, internal hydrogen bonding, and passive membrane permeability: successful in silico prediction of the relative permeabilities … [cited by applicant]
Rezai T, Yu B, Millhauser GL, Jacobson MP, Lokey RS. Testing the conformational hypothesis of passive membrane permeability using synthetic cyclic peptide diastereomers. J Am Chem Soc. Mar. 1, 2006;128(8):2510-1. [cited by applicant]
Rondot S, Koch J, Breitling F, Dübel S. A helper phage to improve single-chain antibody presentation in phage display. Nat Biotechnol. Jan. 2001;19(1):75-8. [cited by applicant]
Rülker T, Voß L, Thullier P, O' Brien LM, Pelat T, Perkins SD, Langermann C, Schirrmann T, Dübel S, Marschall HJ, Hust M, Hülseweh B. Isolation and characterisation of a human-like antibody fragment (scFv) that inactiva… [cited by applicant]
Samuelson P, Gunneriusson E, Nygren PA, Ståhl S. Display of proteins on bacteria. J Biotechnol. Jun. 26, 2002;96 (2):129-54. [cited by applicant]
Sandberg M, Patil J, D'Angelo B, Weber SG, Mallard C. NRF2-regulation in brain health and disease: implication of cerebral inflammation. Neuropharmacology. Apr. 2014;79:298-306. [cited by applicant]
Schlippe YV, Hartman MC, Josephson K, Szostak JW. In vitro selection of highly modified cyclic peptides that act as tight binding inhibitors. J Am Chem Soc. Jun. 27, 2012;134(25):10469-77. [cited by applicant]
Scott CP, Abel-Santos E, Jones AD, Benkovic SJ. Structural requirements for the biosynthesis of backbone cyclic peptide libraries. 2001, p. 801-815. [cited by applicant]
Search Report of International Application No. PCT/US2014/072016 dated Jun. 30, 2017 (9 pages). [cited by applicant]
Seebeck FP, Szostak JW. Ribosomal synthesis of dehydroalanine-containing peptides. J Am Chem Soc. Jun. 7, 2006;128(22):7150-1. [cited by applicant]
Shivange AV, Daugherty PS. De novo discovery of bioactive cyclic peptides using bacterial display and flow cytometry. Methods Mol Biol. 2015; 1248:139-53. [cited by applicant]
Sidhu SS, Lowman HB, Cunningham BC, Wells JA. Phage display for selection of novel binding peptides. Methods Enzymol. 2000;328:333-63. [cited by applicant]
Smith GP, Petrenko VA. Phage Display. Chem Rev. Apr. 1, 1997;97(2):391-410. [cited by applicant]
Smith JM, Frost JR, Fasan R. Emerging strategies to access peptide macrocycles from genetically encoded polypeptides. J Org Chem. Apr. 19, 2013;78(8):3525-31. [cited by applicant]
Smith JM, Vitali F, Archer SA, Fasan R. Modular assembly of macrocyclic organo-peptide hybrids using synthetic and genetically encoded precursors. Angew Chem Int Ed Engl. May 23, 2011;50(22):5075-80. [cited by applicant]
Steel R, Cowan J, Payerne E, O'Connell MA, Searcey M. Anti-inflammatory Effect of a Cell-Penetrating Peptide Targeting the Nrf2/Keap1 Interaction. ACS Med Chem Lett. May 10, 2012;3(5):407-410. [cited by applicant]