Development of Low-Molecular-Weight Compounds Targeting the Cancer-Associated KLF5 Transcription Factor.

Krüppel-like factor 5 (KLF5) is a potential target for anticancer drugs. However, as an intrinsically disordered protein (IDP) whose tertiary structure cannot be solved, innovative strategies are needed. We focused on its hydrophobic α-helix structure, defined as an induced helical motif (IHM), which is a possible interface for protein-protein interaction. Using mathematical analyses predicting the α-helix's structure and hydrophobicity, a 4-amino-acid site (V-A-I-F) was identified as an IHM. Low-molecular-weight compounds that mimic the main chain conformation of the α-helix with the four side chains of V-A-I-F were synthesized using bicyclic pyrazinooxadiazine-4,7-dione. These compounds selectively suppressed the proliferation and survival of cancer cells but not noncancer cells and decreased the protein but not mRNA levels of KLF5 in addition to reducing proteins of Wnt signaling. The compounds further suppressed transplanted colorectal cancer cells in vivo without side effects. Our approach appears promising for developing drugs against key IDPs.

Visualized and Quantitative Conformational Analysis of Peptidomimetics.

Protein-protein interactions (PPIs) are fundamentally important and challenging drug targets. Peptidomimetic molecules of various types have been developed to modulate PPIs. A particularly promising drug discovery strategy, structural peptidomimetics, was designed based on special mimicking of side-chain Cα-Cß bonds. It is simple and versatile. Nevertheless, no quantitative method has been established to evaluate its similarity to a target peptide motif. We developed two methods that enable visual, comprehensive, and quantitative analysis of peptidomimetics: peptide conformation distribution (PCD) plot and peptidomimetic analysis (PMA) map. These methods specifically examine multiple side-chain Cα-Cß bonds of a peptide fragment motif and their corresponding bonds (pseudo-Cα-Cß bonds) in a mimetic molecule instead of φ and ψ angles of a single amino acid in the traditional Ramachandran plot. The PCD plot is an alignment-free method, whereas the PMA map is an alignment-based method providing distinctive and complementary analysis. Results obtained from analysis using these two methods indicate our multifacial α-helix mimetic scaffold 12 as an excellent peptidomimetic that can precisely mimic the spatial positioning of side-chain functional groups of α-helix. These methods are useful for visualized and quantified evaluation of peptidomimetics and for the rational design of new mimetic scaffolds.