Unlocking the potential of protein-derived peptides to target G-quadruplex DNA: from recognition to anticancer activity.

Noncanonical nucleic acid structures, particularly G-quadruplexes, have garnered significant attention as potential therapeutic targets in cancer treatment. Here, the recognition of G-quadruplex DNA by peptides derived from the Rap1 protein is explored, with the aim of developing novel peptide-based G-quadruplex ligands with enhanced selectivity and anticancer activity. Biophysical techniques were employed to assess the interaction of a peptide derived from the G-quadruplex-binding domain of the protein with various biologically relevant G-quadruplex structures. Through alanine scanning mutagenesis, key amino acids crucial for G-quadruplex recognition were identified, leading to the discovery of two peptides with improved G-quadruplex-binding properties. However, despite their in vitro efficacy, these peptides showed limited cell penetration and anticancer activity. To overcome this challenge, cell-penetrating peptide (CPP)-conjugated derivatives were designed, some of which exhibited significant cytotoxic effects on cancer cells. Interestingly, selected CPP-conjugated peptides exerted potent anticancer activity across various tumour types via a G-quadruplex-dependent mechanism. These findings underscore the potential of peptide-based G-quadruplex ligands in cancer therapy and pave the way for the development of novel therapeutic strategies targeting these DNA structures.

Expanding Structure-Activity Relationships of Human Urotensin II Peptide Analogues: A Proposed Key Role of the N-Terminal Region for Novel Urotensin II Receptor Modulators.

While the urotensinergic system plays a role in influencing various pathologies, its potential remains untapped because of the absence of therapeutically effective urotensin II receptor (UTR) modulators. Herein, we developed analogues of human urotensin II (hU-II) peptide in which, along with well-known antagonist-oriented modifications, the Glu1 residue was subjected to single-point mutations. The generated library was tested by a calcium mobilization assay and ex vivo experiments, also in competition with selected ligands. Interestingly, many derivatives showed noncompetitive modulation that was rationalized by the lateral allostery concept applied to a G protein-coupled receptor (GPCR) multimeric model. UPG-108 showed an unprecedented ability to double the efficacy of hU-II, while UPG-109 and UPG-111 turned out to be negative allosteric modulators of UTR. Overall, our investigation will serve to explore and highlight the expanding possibilities of modulating the UTR system through N-terminally modified hU-II analogues and, furthermore, will aim to elucidate the intricate nature of such a GPCR system.

Unveiling the interaction between DNA G-quadruplexes and RG-rich peptides.

G-quadruplexes are non-canonical DNA secondary structures formed within guanine-rich strands that play important roles in various biological processes, including gene regulation, telomere maintenance and DNA replication. The biological functions and formation of these DNA structures are strictly controlled by several proteins that bind and stabilize or resolve them. Many G-quadruplex-binding proteins feature an arginine and glycine-rich motif known as the RGG or RG-rich motif. Although this motif plays a crucial role in the recognition of such non-canonical structures, their interaction is still poorly understood. Here, we employed a combination of several biophysical techniques to provide valuable insights into the interaction between a peptide containing an RGG motif shared by numerous human G-quadruplex-binding proteins (NIQI) and various biologically relevant G-quadruplex DNA structures with different topologies. We also shed light on the key amino acids involved in the binding process. Our findings contribute to lay the basis for the development of a new class of peptide-based G-quadruplex ligands as an alternative to small molecules. These ligands may serve as valid tools for interfering in DNA-protein interactions, with potential therapeutic applications.