**Preface to "Synthetic Peptides and Peptidomimetics: From Basic Science to Biomedical Applications"**

Synthetic peptides are a very appealing class of compounds as both basic and applied science research tools. Furthermore, the high affinity and specificity of peptides towards biological targets, together with their advantageous pharmacological properties, such as poor immunogenicity and low toxicity, has greatly encouraged their development as therapeutic and diagnostic agents in the clinical setting. More than 60 peptide drugs have reached the market and several hundred new therapeutic peptides are in preclinical and clinical development. However, to date, this class of molecules represents only 2% of the world drug market due to their low in vivo stability, short half-lives, poor cell permeability and low oral bioavailability. Many approaches have been developed and are continually being explored to improve peptide stability and pharmacological properties, while maintaining biological potency and selectivity and avoiding toxicity. For example, cyclization, the protection of N- and C-termini, side chain or backbone modifications of natural structures, replacement with D or non-natural amino acids, and the addition of membrane permeability elements are commonly used strategies to improve the in vivo stability and cellular permeability of this class of molecules. Furthermore, the conjugation of peptides with large polymers, fusion with long-lived plasma proteins and lipidation have also been exploited to prolong plasma half-life and increase oral bioavailability, significantly expanding the applicability of peptides as effective drugs.

Trying to cover all the progresses made in this field, this Special Issue, entitled "Synthetic Peptides and Peptidomimetics: From Basic Science to Biomedical Applications", has included both reviews and original research contributions focused on the chemical design and biomedical applications of structurally modified bioactive peptides. In this framework, Doti et al. provide a wide overview on the applications of retro-inverso-modified peptides in anticancer therapies, in immunology, in neurodegenerative diseases, and as antimicrobials, highlighting the benefits and limits of this interesting subclass of molecules as bioactive compounds. Tarvirdipour et al. review the properties of peptides that promote the site-specific localization of nucleic acids and peptide-based nano-assemblies. Romanowska et al. show the chemical design of a set of DNA-binding peptide-based polymers composed of *N*-substituted L-2,3-diaminopropionic acid (DAPEG) residues, and investigate their relative cellular permeability and localization, cytotoxicity and DNA-binding capacity. The experimental results of this study pave the way for the further development of peptide-based nanocarriers. Pandey et al. summarize the current status on the use of peptides as probes both in non-imaging and imaging diagnostic platforms. Moreover, they discuss the applicability of peptide-based diagnostics in deadly diseases, mainly COVID-19 and cancers. In the same research field, Krajcovicova et al. report on the preparation of a new RGD-based radiolabeled peptide targeting αvβ3 integrin. The binding properties of [68Ga]Ga-DFO-c(RGDyK) towards the αvβ3 integrin were studied in vitro and in vivo with various techniques, including PET/CT imaging in a mouse tumor model. In addition, Siepe et al. describe the characterization of a set of bioactive peptides conjugated with environment-sensitive labels, such as luciferin and aminoluciferin, which are used to study their interactions with model membranes, SDS micelles, lipopolysaccharide micelles and bacterial cells. The results demonstrate that luciferin and aminoluciferin are environment-sensitive labels with widespread potential applications in the study of peptides interacting with membranes.

The contributions included in this book also provide several examples of peptides employed as protein–protein interaction (PPI) modulators, both to elucidate the molecular mechanisms underlying diseases and to use them as a starting template for developing new potential therapeutics. In this framework, Dvorak et al. focus their attention on the interaction between a voltage-gated Na+ channel (Nav1.6) and fibroblast growth factor 14 (FGF14), which plays a role in the regulation of neuron excitability in the central nervous system. Notably, using an FGF14-derived synthetic peptide, they demonstrate that pharmacologically targeting the FGF14 interaction site on the C-terminal domain (CDT) of Nav1.6 results in a powerful strategy to achieve the selective modulation of the isoform activity. The data also show that, more generally, the interaction of the CTDs of Nav channels with auxiliary proteins are a target candidate for developing new therapies. Conte et al. focus their attention on the interaction between the Apoptosis-Inducing Factor and the Cyclophilic A (AIF/CypA), which mediates neuronal cell death in vivo and in vitro. Using AIF(370-394) as a prototypical inhibitor, they elucidate the role of the complex in SH-SY5Y cells treated with high concentrations of staurosporine, which is a well-known cell model to study Parkinson's Disease (PD). The results obtained highlight the role of the AIF/CypA complex in the pathophysiological mechanisms leading to PD, suggesting the complex as a promising target for developing first-in-class therapeutics to treat this currently incurable disease. Levi et al. propose the targeting of the integrin αvβ3, which is involved in different stages of cancer progression, metastasis, invasion, and angiogenesis, with a cyclic non-RGD synthetic peptide (ALOS4). This peptide, nine residues, was tested in a subcutaneous xenograft model of A375 human melanoma to evaluate tumor growth, tumor tissue development and the expression of downstream targets of αvβ3. The stability and toxicity of ALOS4 in mice together with the blood cell profile in healthy mice were also evaluated. The results suggest that ALOS4 is stable in the proposed formulations, presents no overt toxicity risks and is effective in melanoma tumor shrinkage by a mechanism related to αvβ3 and possibly other mechanisms. Di Micco et al. focus on the main proteases (Mpro) of SARS-CoV2, a protein essential for viral replication, modifying and repurposing the active peptide AT1001 (Larazotide acetate). AT1001 and five derivatives were designed and assayed in vitro for their ability to interfere with Mpro catalytic activity. The data provide useful information for the development of new generations of antiviral agents for treating SARS-CoV-2, which so far lacks selective therapeutic treatments. The review by Vanzolini et al. provides an overview of the peptides that are currently used as antimicrobials and their mechanism of action.

Highly ordered secondary structure motifs, α-helix, β-sheets and turns, are scaffolds for key amino acid residues in protein–protein hot-spots. The development of peptides that adopt conformations suitable for their biological activity is one of the most important goals of protein chemists, not only in relation to the design of PPI inhibitors. In the article of Makura et al., this is addressed by stapling peptides at *i,i*+1 positions using hydrocarbon linkers introduced by ring-closing metathesis reactions and analyzing their structures through X-ray crystallography. The authors show how their approach is valid for short oligopeptides where stapling is achieved using two adjacent residues. In this field, Kovacevi ˇ c et al. investigated the conformational ´ behavior and antiproliferative activity of peptidomimetics obtained by the conjugation of methyl-1'-aminoferrocene-1-carboxylate with homo- and heterochiral Pro-Ala dipeptides. The results show a promising outcome which could serve for further research and the development of compounds with antitumor activity.

The papers on peptides collected in this Special Issue, which have been proposed and published by groups operating in various parts of the world, show how successful this class of molecules still is, both as model molecules for studying the structure of proteins, and as potential therapeutics and diagnostics, and also as laboratory tools for advanced basic and applied studies. The large scientific community working in this field is very active and productive, and is making the most of the potential and versatility of these molecules to generate increasingly interesting and innovative molecules of therapeutic interest and to understand the fundamental molecular mechanisms of life.

#### **Nunzianna Doti and Menotti Ruvo**

*Editors*
