Discovery of Macrocyclic Peptides Armed with a Mechanism‐Based Warhead: Isoform‐Selective Inhibition of Human Deacetylase SIRT2

Abstract

Designed to inhibit: by using the random nonstandard peptide integrated discovery (RaPID) system, highly potent isoform-selective inhibitors can be identified from a library of nonstandard macrocyclic peptides. These inhibitors, which contain a mechanism-based warhead residue, are active against the human deacetylase SIRT2, with IC(50) values in the low nanomolar region.

Nonstandard Peptide Expression under the Genetic Code Consisting of Reprogrammed Dual Sense Codons

“…Three N-chloroacetylated amino acids with different configurations and side chain structures ( ClAc D F, ClAc L F, and ClAc D Y) were assigned to the table ini . In addition, three nonproteinogenic amino acids (amh, K ac , and K tf ), which are known as mechanism-based warhead residues that target the active site of the deacetylase sirtuins, 18,19 and 17 proteinogenic amino acids were assigned to table elon ( Figure 3A and Scheme 1). When the coexpressed products of the three mRNAs were subjected to mass analysis, three peaks were successfully identified for macrocyclic peptides for which the mass values were consistent with those of the expected peptides ( Figure 3B).…”

Synthesis of fused tricyclic peptides using a reprogrammed translation system and chemical modification

“…The incorporation of an L-lactic acid ( HO A) at a designated position can be achieved by genetic code reprogramming. Since we utilized the Met-deficient FIT system for the reassignment of ClAc-D W, the elongator AUG codon is left vacant; therefore, this vacant codon can be utilized for the assignment for HO A, similar to the strategy reported elsewhere [18,24]. We thus designed a DNA template (D1e) based on the sequence of D1 to include an elongator AUG codon between Cys2 and Cys3, which would be suppressed with HO A by the use of HO A-tRNA enAsn CAU .…”

“…These methods have been coupled with in vitro display technologies to rapidly select for cyclic peptides with exceptional bioactivity [13][14][15][16][17][18]. A very robust method of generating macrocyclic peptides by means of genetic code reprogramming via the FIT (Flexizyme-assisted in vitro translation) system [19,20] is to aminoacylate the initiator tRNA fMet CAU with an amino acid containing an N-chloroacetyl (ClAc) group.…”

Encoded libraries of chemically modified peptides

“…Cyclic peptides from such libraries were isolated to a range of targets including an ubiquitin ligase (K d = 0.6 nM) [27 ], Akt2 kinase (IC 50 = 92 nM) [28], deacetylase SIRT2 (K d = 3.7 nM) [29], a MATE transporter [30] and VEGF receptor 2 (K d = 33 nM) [31]. Suga et al developed several other cyclization strategies with unnatural amino acids that might also be applied to mRNA-encoded peptide libraries in the future.…”