María-José Camarasa

Human coronaviruses include both endemic respiratory pathogens and highly pathogenic viruses with pandemic potential, underscoring the need for antiviral agents with activity beyond single viral species. This review focuses on selected, representative antiviral inhibitors directed against viral proteins of human coronaviruses. Accordingly, the review examines representative inhibitors targeting viral proteins essential for coronavirus infection and propagation. The discussion includes inhibitors acting at different functional levels of the viral life cycle, such as entry, genome replication, RNA processing and particle assembly, and reflects the range of molecular entities reported to date. By organizing known inhibitors according to their viral protein targets, this review provides an integrated overview of current antiviral candidates and illustrates the central role of viral proteins as points of intervention. Overall, the review aims to consolidate existing knowledge on antiviral inhibitors across human coronaviruses, offering a coherent framework for appreciating their scope and diversity.

Leishmania infantum trypanothione reductase (Li‐TryR) is an essential enzyme for parasite redox homeostasis and a validated target for antileishmanial therapy. Classical inhibitors acting at the active site have shown limited success due to poor selectivity, suboptimal pharmacological properties, and lack of in‐cell efficacy. This work presents a comprehensive strategy to develop first‐in‐class Li‐TryR dimerization disruptors, targeting the protein–protein interface required for enzymatic activity. Through molecular modeling, mutagenesis, and structural analysis, a key α‐helical hotspot (around residue E436) was identified as a druggable region. Interface‐derived peptides, helix‐stabilized stapled analogs, α/β‐foldamers, and cell‐penetrating conjugates demonstrated that dimer disruption effectively inhibits LiTryR and can translate into antiparasitic activity. Building on these insights, several families of small‐molecule proteomimetics, including imidazole‐phenyl‐thiazoles, triazole analogs, and triazolium salts, were developed, achieving low‐micromolar to submicromolar inhibition and significant activity in parasite cultures and intracellular models. Overall, this work establishes dimer‐interface targeting as a novel therapeutic paradigm for leishmaniasis and provides multiple optimized chemical scaffolds for future drug development.

Cationic ultrashort lipopeptides (USLPs) are promising antimicrobial candidates to combat multidrug-resistant bacteria. Using DICAMs, a newly synthesized family of tripeptides with net charges from −2 to +1 and a fatty amine conjugated to the C-terminus, we demonstrate that anionic and neutral zwitterionic USLPs can possess potent antimicrobial and membrane-disrupting activities against prevalent human pathogens such as Streptococcus pneumoniae and Streptococcus pyogenes. The strongest antimicrobials completely halt bacterial growth at low micromolar concentrations, reduce bacterial survival by several orders of magnitude, and may kill planktonic cells and biofilms. All of them comprise either an anionic or neutral zwitterionic peptide attached to a long fatty amine (16–18 carbon atoms) and show a preference for anionic lipid membranes enriched in phosphatidylglycerol (PG), which excludes electrostatic interactions as the main driving force for DICAM action. Hence, the hydrophobic contacts provided by the long aliphatic chains of their fatty amines are needed for DICAM’s membrane insertion, while negative-charge shielding by salt counterions would reduce electrostatic repulsions. Additionally, we show that other components of the bacterial envelope, including the capsular polysaccharide, can influence the microbicidal activity of DICAMs. Several promising candidates with good-to-tolerable therapeutic ratios are identified as potential agents against S. pneumoniae and S. pyogenes. Structural characteristics that determine the preference for a specific pathogen or decrease DICAM toxicity have also been investigated.

The influenza virus glycoprotein hemagglutinin (HA) participates in critical steps of the attachment of viral particles to the host cell membrane receptor and membrane fusion. Due to its crucial involvement in the initial phases of influenza A infections, HA emerges as a promising target in the search of novel drug-like candidates. Given its pivotal role in the early stages of influenza A infections, intense drug discovery efforts have been undertaken to target HA in the past decades. Drug discovery studies mainly rely on preventing the recognition of sialic acid units by the receptor binding site in the globular head (GH) domain, or the conformational rearrangement required for the fusion of viral and cell membranes. In this work, the aim is to summarize the progress made in HA-targeted development of small molecule fusion inhibitors. To this end, attention will primarily be focused on the analysis of the X-ray crystallographic structures of HA bound to fusion inhibitors. Furthermore, this study also aims to highlight the efforts made in exploiting the structural information in conjunction with molecular modeling techniques to discern the mechanism of action of the fusion inhibitors and to assist the design and interpretation of structure-activity relationships of novel lead compounds will be highlighted. The final section will be dedicated to elucidating novel and promising antiviral strategies proceeding from the transformation of known small molecule antivirals in proteolysis targeting chimera (PROTAC)-based targeted protein degradation. This knowledge will be valuable to assist the exploitation of classical and novel antiviral structure-based strategies, together with a deeper understanding of the mechanism of action and minimization of the impact of drug resistance.

ATP-binding cassette (ABC) transport systems are crucial for bacteria to ensure sufficient uptake of nutrients that are not produced de novo or improve the energy balance. The cell surface of the pathobiont Streptococcus pneumoniae (pneumococcus) is decorated with a substantial array of ABC transporters, critically influencing nasopharyngeal colonization and invasive infections. Given the auxotrophic nature of pneumococci for certain amino acids, the Ami ABC transporter system, orchestrating oligopeptide uptake, becomes indispensable in host compartments lacking amino acids. The system comprises five exposed Oligopeptide Binding Proteins (OBPs) and four proteins building the ABC transporter channel. Here, we present a structural analysis of all the OBPs in this system. Multiple crystallographic structures, capturing both open and closed conformations along with complexes involving chemically synthesized peptides, have been solved at high resolution providing insights into the molecular basis of their diverse peptide specificities. Mass spectrometry analysis of oligopeptides demonstrates the remarkable promiscuity of some of these proteins when expressed in Escherichia coli , displaying affinity for a wide range of peptides. Finally, a model is proposed for the complete Ami transport system in complex with its various OBPs. We further disclosed some essential structural changes facilitating oligopeptide transport into the cellular cytoplasm. Thus, the structural analysis of the Ami system provides valuable insights into the mechanism and specificity of oligopeptide binding by the different OBPs, shedding light on the intricacies of the uptake mechanism. Author Summary: The uptake of diverse oligopeptides enables pneumococcal growth despite auxotrophies and functions as a critical sensor for assessing the composition of the local environment. The identification of additional OBPs in non-encapsulated S. pneumoniae strains suggests their involvement in sensing a broader spectrum of bacterial competitors coexisting with the highly commensal pneumococcus. This study presents a comprehensive analysis of the initial phase of peptide transport mediated by OBPs within the pneumococcal Ami permease system. We disclose a common mechanism for oligopeptide recognition that is modulated in each OBP to accommodate a diverse array of oligopeptides. Understanding how pneumococcus perceives external stimuli and responds to them is imperative for unraveling the transition from a commensal to a pathogenic state.

AL-471, the leading exponent of a class of potent HIV and enterovirus A71 (EV-A71) entry inhibitors discovered in our research group, contains four l-tryptophan (Trp) units bearing an aromatic isophthalic acid directly attached to the C2 position of each indole ring. Starting from AL-471, we (i) replaced l-Trp with d-Trp, (ii) inserted a flexible linker between C2 and the isophthalic acid, and (iii) substituted a nonaromatic carboxylic acid for the terminal isophthalic acid. Truncated analogues lacking the Trp motif were also synthesized. Our findings indicate that the antiviral activity seems to be largely independent of the stereochemistry (l- or d-) of the Trp fragment and also that both the Trp unit and the distal isophthalic moiety are essential for antiviral activity. The most potent derivative, 23 (AL-534), with the C2 shortest alkyl urea linkage (three methylenes), showed subnanomolar potency against different EV-71 clinical isolates. This finding was only observed before with the early dendrimer prototype AL-385 (12 l-Trp units) but remained unprecedented for the reduced-size prototype AL-471. Molecular modeling showed the feasibility of high-affinity binding of the novel l-Trp-decorated branches of 23 (AL-534) to an alternative site on the VP1 protein that harbors significant sequence variation among EV-71 strains.

There is a clear need for novel antiviral concepts to control SARS-CoV-2 infection. Based on the promising anti-coronavirus activity observed for a class of 1,4,4-trisubstituted piperidines, we here conducted a detailed analysis of the structure–activity relationship of these structurally unique inhibitors. Despite the presence of five points of diversity, the synthesis of an extensive series of analogues was readily achieved by Ugi four-component reaction from commercially available reagents. After evaluating 63 analogues against human coronavirus 229E, four of the best molecules were selected and shown to have micromolar activity against SARS-CoV-2. Since the action point was situated post virus entry and lying at the stage of viral polyprotein processing and the start of RNA synthesis, enzymatic assays were performed with CoV proteins involved in these processes. While no inhibition was observed for SARS-CoV-2 nsp12-nsp7-nsp8 polymerase, nsp14 N7-methyltransferase and nsp16/nsp10 2’-O-methyltransferase, nor the nsp3 papain-like protease, the compounds clearly inhibited the nsp5 main protease (M^pro). Although the inhibitory activity was quite modest, the plausibility of binding to the catalytic site of M^pro was established by in silico studies. Therefore, the 1,4,4-trisubstituted piperidines appear to represent a novel class of non-covalent CoV M^pro inhibitors that warrants further optimization and development.