Fmoc-(4S)-4-Azido-D-Proline

Alloviroidin is a cyclic heptapeptide, produced by several species of Amanita mushrooms, that demonstrates high affinity for F-actin as is characteristic of virotoxins and phallotoxins. Alloviroidin was synthesized via a [3 + 4] fragment condensation of Fmoc-d-Thr(OTBS)-d-Ser(OTBS)-(2 S,3 R,4 R)-DHPro(OTBS)2-OH and H-Ala-Trp(2-SO2Me)-(2 S,4 S)-DHLeu(5-OTBS)-Val-OMe to form bond A. The linear heptapeptide favored a turn conformation, facilitating cyclization between Val1 and d-Thr2 (position B). Global deprotection and HPLC purification afforded alloviroidin with NMR spectra in excellent agreement with the natural product.

As a key cytokine mediator of inflammation, interleukin-1β (IL-1β) binds to the IL-1 receptor (IL-1R) and activates various downstream signaling mediators, including NF-κB, which is required for immune vigilance and cellular protection. Toward the development of IL-1-targeting therapeutics which exhibit functional selectivity, the all-D-amino acid peptide 1 (101.10, H-D-Arg-D-Tyr-D-Thr-D-Val-D-Glu-D-Leu-D-Ala-NH2) was conceived as an allosteric IL-1R modulator that conserves NF-κB signaling while inhibiting other IL-1-activated pathways. Employing β-hydroxy-α-amino-γ-lactam (Hgl) stereoisomers to study the conformation about the Thr3 residue in 1, [(3R,4S)-Hgl3]-1 (2b), among all possible diastereomers, was found to exhibit identical in vitro and in vivo activity as the parent peptide and superior activity to the α-amino-γ-lactam (Agl) counterpart. Noting the relevance of the β-hydroxyl substituent and configuration for the activity of (3R,4S)-2b, fifteen different β-substituted-Agl3 analogs of 1 (e.g., 2c-q) have now been synthesized by a combination of solution- and solid-phase methods employing N-Fmoc-β-substituted-Agl3-Val-OH dipeptide building blocks. Introduction of a β-azido-Agl3 residue into the resin bound peptide and subsequent reduction and CuAAC chemistry gave access to a series of amine and triazole derivatives (e.g., 2h-q). β-Substituted-[Agl3]-1 analogs 2c-q exhibited generally similar circular dichroism (CD) spectra as that of Hgl analog 2b in water, presenting curve shapes indicative of β-turn structures. The relevance of the β-substituent was indicated in rodent models of preterm labor and retinopathy of prematurity (ROP), in which certain analogs inhibited preterm birth and vaso-obliteration, respectively, with activity similar to 1 and 2b. The β-substituted-[Agl3]-1 analogs exhibited functional selectivity on IL-1-induced signaling pathways. The described solid-phase method has provided discerning probes for exploring peptide structure-activity relationships and valuable leads for developing prototypes to treat inflammatory events leading to prematurity and retinopathy of prematurity, which are leading causes of infant morbidity and blindness respectively.

(2S,4R)- and (2S,4S)-perfluoro-tert-butyl 4-hydroxyproline were synthesized (as Fmoc-, Boc-, and free amino acids) in 2-5 steps. The key step of each synthesis was a Mitsunobu reaction with perfluoro-tert-butanol, which incorporated a perfluoro-tert-butyl group, with nine chemically equivalent fluorines. Both amino acids were incorporated in model α-helical and polyproline helix peptides. Each amino acid exhibited distinct conformational preferences, with (2S,4R)-perfluoro-tert-butyl 4-hydroxyproline promoting polyproline helix. Peptides containing these amino acids were sensitively detected by (19)F NMR, suggesting their use in probes and medicinal chemistry.