Rational Design of a Dual-Targeting Natural Toxin-Like Bicyclic Peptide for Selective Bioenergetic Blockage in Cancer Cells

Abstract

Stapled α-helical peptides emerge as one of the attractive peptidomimetics which can efficiently penetrate the cell membrane to access intracellular targets. However, the incorporation of a highly lipophilic cross-link may lead to nonspecific membrane toxicity in certain cases. Here, we report a new class of thioether-tethered bicyclic α-helical peptide to mimic the highly constrained loop-helix structure of natural toxins with the dualtargeting ability for both cell-surface receptors and intracellular targets. The thioether cross-links are introduced to replace the redox-sensitive disulfide bonds in natural toxins via a photoinduced thiol-yne reaction followed by macrolactamization. As a proof of concept, α V β 3 integrin targeting ligand was grafted into one of the macrocycles in the bicyclic scaffold, while a mitochondriatargeting proapoptotic motif was introduced into the other macrocycle stabilized by an i, i + 7 alkyl thioether cross-link to recapitulate its α-helical conformation. The obtained dual-targeting bicyclic α-helical BIRK peptides showed highly stable α-helical conformation in the presence of denaturants or under high temperature. Notably, BIRK peptides could induce selective cell death in α V β 3 integrin-positive B16F10 cells by interfering with the bioenergetic functions of mitochondria. This work provides a new avenue to design and stabilize α-helical peptides in a highly constrained bicyclic loop-helix scaffold with dual functionality.

Bicyclic peptides: Paving the road for therapeutics of the future

“…Recent progress has also been made in designing intramolecular bicyclization reactions. [67][68][69] For this type of bicyclization, a reactive handle is installed as part of the linear peptide chain, as opposed to applying external reagents to the assembled linear peptide. Reymond and co-workers coupled 3,5-bis(chloromethyl)-4-methylbenzoic acid to the peptide N-terminus to later form two thioether linkages with cysteines (Scheme 6, Figure 5a).…”

“…[67] Bicyclization can also be achieved through the synthesis of an amino acid bearing two carboxylic acids during SPPS (Scheme 7). [68] A S C H E M E 3 Peptide bicyclization using S-arylation with Au(III)-complexes or triiodoarenes and Pd-catalysis. [52][53][54] S C H E M E 4 Generation of peptide-bismuth bicycles under biocompatible conditions.…”

“…[68] Orthogonal deprotection of two amines in the peptide allows selective internal amide couplings. [68] A dual-targeted, α-helical bicycle synthesized this way showed potential as a cytotoxin for cancer treatment. [68] An alternative peptide bicyclization method is based on the reaction of 2,6-dicyanopyridine with 1,2-aminothiols.…”

“…[68] A dual-targeted, α-helical bicycle synthesized this way showed potential as a cytotoxin for cancer treatment. [68] An alternative peptide bicyclization method is based on the reaction of 2,6-dicyanopyridine with 1,2-aminothiols. [69] Both functional groups were integrated into SPPS-compatible unnatural amino acids that can be assembled before or during the synthesis of the linear peptide.…”

“…[92] S C H E M E 7 Synthesis of bicyclic peptides with an in situ created dicarboxylic acid amino acid. [68] S C H E M E 8 Synthesis of bicyclic peptides using the reactivity of 2,6-dicyanopyridine and 1,2-aminothiol residues incorporated into a linear peptide during SPPS. [69]…”