Botulinum neurotoxin-specific capture agents, compositions, and methods of using and making
The present application provides stable peptide-based Botulinum neurotoxin (BoNT) serotype A capture agents and methods of use as detection and diagnosis agents and in the treatment of diseases and disorders. The application further provides methods of manufacturing BoNT serotype A capture agents using iterative on-bead in situ click chemistry.
1. A capture agent comprising an anchor ligand that comprises a cyclic peptide comprising the formula of Dab(DNP)-R-Lys(N3)-T-Dab-Pra-L-SEQ ID NO:3).
2. The capture agent of claim 1 , wherein the anchor ligand comprises a molecule of formula Inh-1
wherein R 1 is absent or is a linker to the secondary ligand.
3. The capture agent of claim 1 , further comprising a secondary ligand comprises a cyclic peptide comprising the formula of -Pra-NYRWL-Lys(N3) (SEQ ID NO:7).
4. The capture agent of claim 1 , further comprising a secondary ligand comprises a molecule of formula L2
wherein R 2 is absent or is a linker to the anchor ligand.
5. The capture agent of claim 1 further comprising a linker.
6. The capture agent of claim 5 , wherein the linker is a tripeptide.
7. The capture agent of claim 6 , wherein the linker comprises the amino acid sequence of Gly-Aib-Leu or Leu-Aib-Gly.
8. The capture agent of claim 7 , wherein the linker comprises the amino acid sequence of Gly-Aib-Leu.
9. The capture agent of claim 7 , wherein the linker comprises the amino acid sequence of Leu-Aib-Gly.
10. The capture agent of claim 5 , wherein the linker comprises PEG 4 .
11. The capture agent of claim 5 , wherein the anchor ligand and/or the secondary ligand are linked to the linker via a 1,4-substituted-1,2,3-triazole residue (Tz4) or via a 1,5-substituted-1,2,3-triazole residue (Tz5).
12. The capture agent of claim 11 , wherein the anchor ligand is linked to the linker via a Tz4.
13. The capture agent of claim 1 , wherein the capture agent comprises a molecule of formula Inh-2
wherein R 3 is hydrogen, a capping group, or comprises a label or a spontaneously translocating peptide.
14. The capture agent of claim 13 , wherein the label is selected from the group consisting of biotin, copper-DOTA, biotin-PEG3, aminooxyacetate, 19 FB, 18 FB, 5-carboxyfluorescein, and FITC-PEG3.
15. The capture agent of claim 13 , wherein the label is a detectable moiety selected from the group consisting of 64 Cu DOTA, 68 Ga DOTA, 68 Ga NOTA, 18 F, Al 18 F NOTA, 64 Cu, 68 Ga 89 Zr, 124 I, 86 Y, 94m Tc, 110m In, 11 C and 76 Br.
16. The capture agent of claim 13 , wherein the spontaneously translocating peptide comprises the amino acid sequence of pliylrllrGqf (SEQ ID NO:24).