Development of Solid-Phase Synthetic Strategies towards Multifunctional Luminescent Lanthanide–Stapled Peptides for Biomedical Applications

Protein-protein interactions (PPIs) are predominant in the underlying mechanism of most cellular processes, and the pathological variants are no exception. Peptides hold tremendous promises as therapeutics as PPI inhibitors due to their larger binding surface areas with target specificity, modular scalable synthesis of structural/functional tunability, and potentially less toxicity. However, their metabolic instability, weak binding affinity affected by conformational flexibility, and poor cell permeability hamper their further direct clinical applications and translations. Recently, several technological approaches to helical peptide stabilization to address the above-mentioned drawbacks via peptide stapling has been developed. Many solution-phase organic synthetic methodologies, e.g. condensations, cross-couplings, C-H activations, and cycloadditions, have been established in both site-specific and combinatorial fashions. In fact, contemporary screenings of stapled peptides of different sequences, lengths, and linkers against specific targets hinge on a trial-and-error basis after each solid-phase peptide synthesis (SPPS), with fluorescent dyes being tediously ligated for bioimaging after multistep transformations and purifications, which is highly inefficient. Bladder cancer (BC) has recently been a global public health concern. Alpha-v and beta-3 (αvβ3) integrin and neurotensin receptor 1 (NTR1), both possessing significant a- helical structural portions, have been reported and recognized as uniquely overexpressed membrane receptors on bladder cancer cells, which can act as attractive specific therapeutic targets against BC. Many of their specific targeting peptide sequences in also a-helical forms have been reported for in vitro/vivo anti-BC studies, serving as good stapling model platforms for enhanced biocompatibilities and bioactivities.

In this proposal, we aim to launch an integrative research program to develop the first proof-of-concept, efficient, and purification-economical solid-phase luminescent lanthanide–stapled BC-targeting peptide (Ln-SP) synthetic protocols by formulating/optimizing various novel stapling reactions in SPPS conditions for incorporating luminescent lanthanide complexes either as the N-terminus/backbone or as the staple linker between the side chain of amino acids. To support our hypothesis, FOUR preliminary experiments have been conducted: we had (i) devised a solid-phase assembly approach for installing NOTA at the N-terminus of peptides for accommodating lanthanides; (ii) & (iii) adopted a lysine-based SPPS stapling strategy to cyclize peptide with cyclen linker via double SN2 reactions and for lanthanide coordination; (iv) conducted molecular docking for the a-helix stability–cyclen stapling length relationship. Successes in the proposal will (i) offer new convenient SPPS procedures for multifunctional stapled lanthanide-peptide bioprobes to understand their PPI modulating mechanisms in vitro/vivo, (ii) set world-first databases/benchmarks for novel Ln-SPs’ properties, and (iii) discover and develop new BC-specific theranostic metallopeptidic drugs.