SARS-CoV-2 is a respiratory pathogen and primarily infects the airway epithelium. As our knowledge about innate immune factors of the respiratory tract against SARS-CoV-2 is limited, we generated and screened a peptide/protein library derived from bronchoalveolar lavage for inhibitors of SARS-CoV-2 spike-driven entry. Analysis of antiviral fractions revealed the presence of α₁-antitrypsin (α₁AT), a highly abundant circulating serine protease inhibitor. Here, we report that α₁AT inhibits SARS-CoV-2 entry at physiological concentrations and suppresses viral replication in cell lines and primary cells including human airway epithelial cultures. We further demonstrate that α₁AT binds and inactivates the serine protease TMPRSS2, which enzymatically primes the SARS-CoV-2 spike protein for membrane fusion. Thus, the acute phase protein α₁AT is an inhibitor of TMPRSS2 and SARS-CoV-2 entry, and may play an important role in the innate immune defense against the novel coronavirus. Our findings suggest that repurposing of α₁AT-containing drugs has prospects for the therapy of COVID-19.

Antimicrobial peptides (AMPs) are widespread in multicellular organisms. These structurally diverse molecules are produced as the first line of defense against pathogens such as bacteria, viruses, fungi, and parasites. Also known as host defense peptides in higher eukaryotic organisms, AMPs display immunomodulatory and anticancer activities. During the last 30 years, technological advances have boosted the research on antimicrobial peptides, which have also attracted great interest as an alternative to tackling the antimicrobial resistance scenario mainly provoked by some bacterial and fungal pathogens. However, the introduction of natural AMPs in clinical trials faces challenges such as proteolytic digestion, short half-lives, and cytotoxicity upon systemic and oral application. Therefore, some strategies have been implemented to improve the properties of AMPs aiming to be used as effective therapeutic agents. In the present review, we summarize the discovery path of AMPs, focusing on preclinical development, recent advances in chemical optimization and peptide delivery systems, and their introduction into the market.

Growth in biofilms as a fascinating and complex microbial lifestyle has become widely accepted as one of the key features of pathogenic microbes, to successfully express their full virulence potential and environmental persistence. This also increases the threat posed by Candida auris, which has a high intrinsic ability to persist on abiotic surfaces including those of surgical instruments and medical tubing. In a previous study, cyclic and helical-stabilized analogues of the antifungal peptide Cm-p5 were designed and synthetized, and proved to have increased activities against C. albicans and C. parapsilosis, but not against planktonic C. auris cells cultivated in suspension cultures. Here, we demonstrate, initially, that these derivatives, however, exhibited semi-inhibitory concentrations between 10–21 µg/mL toward C. auris biofilms. Maturated biofilms were also arrested between 71–97%. These novel biofilm inhibitors may open urgently needed new routes for the development of novel drugs and treatments for the next stage of fight against C. auris.

Following the information obtained by a rational design study, a cyclic and helical-stabilized analogue of the peptide Cm-p5 was synthetized. The cyclic monomer showed an increased activity in vitro against Candida albicans and Candida parapsilosis, compared to Cm-p5. Initially, 14 mutants of Cm-p5 were synthesized following a rational design to improve the antifungal activity and pharmacological properties. Antimicrobial testing showed that the activity was lost in each of these 14 analogues, suggesting, as a main conclusion, that a Glu–His salt bridge could stabilize Cm-p5 helical conformation during the interaction with the plasma membrane. A derivative, obtained by substitution of Glu and His for Cys, was synthesized and oxidized with the generation of a cyclic monomer with improved antifungal activity. In addition, two dimers were generated during the oxidation procedure, a parallel and antiparallel one. The dimers showed a helical secondary structure in water, whereas the cyclic monomer only showed this conformation in SDS. Molecular dynamic simulations confirmed the helical stabilizations for all of them, therefore indicating the possible essential role of the Glu–His salt bridge. In addition, the antiparallel dimer showed a moderate activity against Pseudomonas aeruginosa and a significant activity against Listeria monocytogenes. Neither the cyclic monomer nor the dimers were toxic against macrophages or THP-1 human cells. Due to its increased capacity for fungal control compared to fluconazole, its low cytotoxicity, together with a stabilized α-helix and disulfide bridges, that may advance its metabolic stability, and in vivo activity, the new cyclic Cm-p5 monomer represents a potential systemic antifungal therapeutic candidate.

Antimicrobial peptides (AMPs) are biomolecules with antimicrobial activity against a broad group of pathogens. In the past few decades, AMPs have represented an important alternative for the treatment of infectious diseases. Their isolation from natural sources has been widely investigated. In this sense, mollusks are promising organisms for the identification of AMPs given that their immune system mainly relies on innate response. In this report, we characterized the peptide fraction of the Cuban freshwater snail Pomacea poeyana (Pilsbry, 1927) and identified 37 different peptides by nanoLC-ESI-MS-MS technology. From these peptide sequences, using bioinformatic prediction tools, we discovered two potential antimicrobial peptides named Pom-1 (KCAGSIAWAIGSGLFGGAKLIKIKKYIAELGGLQ) and Pom-2 (KEIERAGQRIRDAIISAAPAVETLAQAQKIIKGG). Database search revealed that Pom-1 is a fragment of Closticin 574 previously isolated from the bacteria Clostridium tyrobutyrium, and Pom-2 is a fragment of cecropin D-like peptide first isolated from Galleria mellonella hemolymph. These sequences were chemically synthesized and evaluated against different human pathogens. Interestingly, structural predictions of both peptides in the presence of micelles showed models that comprise two alpha helices joined by a short loop. The CD spectra analysis of Pom-1 and Pom-2 in water showed for both structures a high random coil content, a certain content of α-helix and a low β-sheet content. Like other described AMPs displaying a disordered structure in water, the peptides may adopt a helical conformation in presence of bacterial membranes. In antimicrobial assays, Pom-1 demonstrated high activity against the Gram-negative bacteria Pseudomonas aeruginosa and moderate activity against Klebsiella pneumoniae and Listeria monocytogenes. Neither of the two peptides showed antifungal action. Pom-1 moderately inhibits Zika Virus infection but slightly enhances HIV-1 infectivion in vitro. The evaluation of cell toxicity on primary human macrophages did not show toxicity on THP-1 cells, although slight overall toxicity was observed in high concentrations of Pom-1. We assume that both peptides may play a key role in innate defense of P. poeyana and represent promising antimicrobial candidates for humans.

EPI-X4, a 16-mer fragment of albumin, is a specific endogenous antagonist and inverse agonist of the CXC-motif-chemokine receptor 4 (CXCR4) and thus a key regulator of CXCR4 function. Accordingly, activity-optimized synthetic derivatives of EPI-X4 are promising leads for the therapy of CXCR4-linked disorders such as cancer or inflammatory diseases. We investigated the binding of EPI-X4 to CXCR4, which so far remained unclear, by means of biomolecular simulations combined with experimental mutagenesis and activity studies. We found that EPI-X4 interacts through its N-terminal residues with CXCR4 and identified its key interaction motifs, explaining receptor antagonization. Using this model, we developed shortened EPI-X4 derivatives (7-mers) with optimized receptor antagonizing properties as new leads for the development of CXCR4 inhibitors. Our work reveals the molecular details and mechanism by which the first endogenous peptide antagonist of CXCR4 interacts with its receptor and provides a foundation for the rational design of improved EPI-X4 derivatives.