Viviana De Luca

Malaria remains a major global health burden, compounded by increasing resistance to current therapies. Plasmodium falciparum carbonic anhydrase ( Pfa CA), a η ‐class metalloenzyme essential for the parasite metabolism and survival, has recently emerged as a promising antimalarial target. In this study, benzoxaborole 4 and its 6‐substituted triazolyl derivatives ( 5–18 ) were evaluated for Pfa CA inhibition and in vitro antiplasmodial activity against the P. falciparum 3D7 strain. Compounds 13 and 14 emerged as the most potent inhibitors ( K _I = 0.45 and 0.43 µM, respectively) of the enzyme, coupled with notable antiplasmodial activity (IC ₅₀ = 34 and 47 µM). Among the tested compounds, the ethyl ester 7 demonstrated the strongest antiplasmodial activity (IC ₅₀ = 2.5 µM). Molecular docking studies further supported the favorable binding of active derivatives within the Pfa CA active site. These findings highlight the potential of benzoxaborole‐based scaffolds as a new chemotype for antimalarial drug development.

Background/Objectives: Fusobacterium nucleatum and Akkermansia muciniphila are key components of the human microbiota, influencing health and disease. F. nucleatum is associated with colorectal cancer (CRC) and poor prognosis through its pro-inflammatory and pro-tumorigenic activity, whereas A. muciniphila is linked to metabolic benefits and anti-inflammatory effects. This study aimed to evaluate the immunomodulatory impact of protein extracts from these bacteria on peripheral T cell responses in healthy individuals and CRC patients. Methods: Peripheral blood mononuclear cells (PBMCs) were exposed to bacterial extracts, individually or in combination, and T cell subsets were analyzed by polychromatic flow cytometry. Results: In healthy donors, F. nucleatum increased Th0, Th2, and Tc9 cell frequencies while reducing Th1, Th1/Th17, and Treg cells. Conversely, A. muciniphila promoted a pro-inflammatory-associated T cell phenotype characterized by higher Th0, Th2, Th17, and Tc17 cells. Combined exposure enhanced Th0, Th17, and Tc17 cells while decreasing Th9 cells. In CRC patients, bacterial extracts induced no significant changes in T cell subsets. Conclusions: These findings indicate that F. nucleatum skews immune responses toward humoral and mucosal defense, whereas A. muciniphila enhances T cell polarization toward subsets usually associated with pro-inflammatory immune responses in healthy subjects. Further studies are needed to clarify their systemic immunological roles and interactions within the tumor microenvironment of CRC.

Bacterial carbonic anhydrases (CAs) are essential for intracellular pH regulation, bicarbonate homeostasis, and energy metabolism, making them attractive antimicrobial targets. Here, building on evidence that acetazolamide (AZA) delivered via hyaluronic acid–palmitate (HA-PA) nanocarriers impairs Escherichia coli growth and its glucose uptake, we investigated the physiological roles of β- and γ-class CAs using sulphonamide inhibitors with distinct selectivity encapsulated in HA-PA nanomicelles to ensure intracellular delivery. AZA, a potent dual β/γ-CA inhibitor, ethoxzolamide (EZA), a selective β-CA inhibitor, and hydrochlorothiazide (HCT), a weaker inhibitor of both classes, were tested for effects on bacterial physiology. The nanoparticles reduced growth in a dose- and class-dependent manner, with AZA exerting the strongest activity, EZA intermediate inhibition, and HCT only modest effects at higher concentrations. Early metabolic responses assessed via intracellular ATP after three hours of exposure revealed an unexpected and reproducible ATP increase for all inhibitors relative to untreated cells, suggesting reduced ATP consumption in bicarbonate-dependent pathways. These findings provide indirect yet compelling evidence that β- and γ-class CAs influence bacterial energy homeostasis and support the rationale for CA inhibition as an antimicrobial strategy, while highlighting HA-PA carriers as effective systems for delivering CA inhibitors intracellularly and enhancing their functional activity in bacterial cells.

This study investigates derivatives including a tetrazole zinc‐binding group as inhibitors of the α‐, β‐, and γ‐class carbonic anhydrase (CA) isoforms from pathogenic bacteria such as Vibrio cholerae , Escherichia coli , and Streptococcus mutans . A library of 22 previously synthesized compounds was evaluated for their inhibitory activity against bacterial CAs using a stopped‐flow CO ₂ hydrase assay. The tetrazole‐based inhibitors displayed marked activity toward bacterial α‐ and β‐class CAs, with inhibition constants ( K _I s) ranging from 0.43 to 18.7 µM. Among them, 2i and 2p emerged as two of the most potent inhibitors. Selectivity studies showed that only a few compounds exhibited moderate (10‐ to 20‐fold) selectivity for bacterial β‐CAs over human CA I, with limited selectivity against human CA II. Molecular modelling studies of the most representative compounds elucidated the key interactions responsible for their potency and selectivity, in agreement with experimental findings. Overall, these results highlight tetrazole derivatives as promising scaffolds for the development of novel antibacterial CA inhibitors.

Chitosan-based nanoparticles were prepared using an eco-friendly chemical procedure that conjugates natural fatty acids to the backbone of chitosan. This consists of reacting two molecules in the absence of a solvent and using microwaves to promote the chemical transformation. Both conditions make the whole chemical process more eco-compatible in terms of reagents and energy consumption. The chemical structure and the self-association behavior of chitosan–fatty acid conjugates were characterized by FT-IR, NMR, and dynamic light scattering. The conjugates displayed an enhanced solubility and efficient self-assembly in aqueous solution. The antimicrobial activity of the resulting nanoparticles was evaluated against Escherichia coli (Gram-negative) and Bacillus subtilis (Gram-positive). The micelles significantly inhibited E. coli growth (35–60%), even at relatively low concentrations, whereas negligible activity was observed against B. subtilis. The antibacterial efficacy appears to arise primarily from the ability of the developed nanostructured conjugates to perturb bacterial membranes. These results support the potential of chitosan–fatty acid conjugates as sustainable nanomaterials for biomedical applications, particularly as eco-friendly antimicrobial agents. Future work will evaluate their activity against other Gram-positive pathogens and explore their use in drug delivery.

Acetazolamide (AZA) is a validated carbonic anhydrase inhibitor (CAI) that has the potential for use in various therapeutic applications. Herein, we report a novel AZA-loaded biodegradable nanodelivery system that was proven to enhance the antibacterial efficacy of the drug against Gram-negative bacteria, such as Escherichia coli. Carbonic anhydrases (CA, EC 4.2.1.1) in E. coli play a crucial role in bacterial metabolism and CO₂/HCO₃⁻ balance; therefore, they represent a suitable target for antimicrobial strategies. The nanoparticles were obtained using a green synthetic protocol that allowed conjugation of a natural fatty acid to hyaluronic acid (HA) under solvent-free conditions. Full characterization of the micellar aggregates was performed (diameter of the micelles, zeta potential, and drug release study). In vitro studies demonstrated that AZA loaded in HA-based nanoparticles significantly inhibited E. coli growth at concentrations as low as 0.5 µg/mL, whereas higher concentrations of free AZA were required, as previously reported. Additionally, encapsulated AZA disrupted glucose consumption in E. coli, indicating its profound impact on bacterial metabolism. These findings suggest that the HA–palmitate nanoparticle not only enhances the delivery and efficacy of AZA but also offers a strategy to affect bacterial metabolism.

Toxoplasma gondii, the causative agent of toxoplasmosis, is a protozoan parasite capable of infecting a wide range of hosts, posing significant health risks, particularly to immunocompromised individuals and congenital transmission. Current therapeutic options primarily target the active tachyzoite stage but are limited by issues such as toxicity and incomplete efficacy. As a result, there is an urgent need for alternative therapies that can selectively target parasite-specific mechanisms critical for metabolic processes and host–parasite interactions. In this context, α-carbonic anhydrase (Tg_CA), an enzyme essential for T. gondii survival has emerged as a promising drug target. Tg_CA was successfully expressed and purified to evaluate its susceptibility to sulfonamide-based inhibitors, represented by compounds 1–24 and the AAZ–HCT series. These inhibitors demonstrated a broad spectrum of activity, with KI values ranging from 17.8 to 8450 nM. Several compounds exhibited moderate to high potency against Tg_CA; however, concerns regarding selectivity arose because of the inhibition of human isoforms, particularly CA I and CA II. Thus, although some inhibitors showed strong activity against Tg_CA, optimizing selectivity remains crucial for minimizing off-target effects and improving therapeutic efficacy. Further structural modifications may enhance selectivity and advance the development of effective treatments for toxoplasmosis.

Acinetobacter baumannii is a Gram-negative opportunistic pathogen responsible for severe hospital-associated infections. Owing to its ability to develop resistance to a wide range of antibiotics, novel therapeutic strategies are urgently needed. One promising approach is to target bacterial carbonic anhydrases (CAs; EC 4.2.1.1), which are enzymes critical for various metabolic processes. The genome of A. baumannii encodes a β-CA (βAbauCA), which is essential for producing bicarbonate ions required in the early stages of uridine triphosphate (UTP) synthesis, a precursor for the synthesis of peptidoglycans, which are vital components of the bacterial cell wall. This study aimed to inhibit βAbauCA in vitro, with the potential to impair the vitality of the pathogen in vivo. We conducted sequence and structural analyses of βAbauCA to explore its differences from those of human CAs. Additionally, kinetic and inhibition studies were performed to investigate the catalytic efficiency of βAbauCAβ and its interactions with sulfonamides and their bioisosteres, classical CA inhibitors. Our results showed that βAbauCA has a turnover rate higher than that of hCA I but lower than that of hCA II and displays distinct inhibition profiles compared to human α-CAs. Based on the obtained data, there are notable differences between the inhibition profiles of the human isoforms CA I and CA II and bacterial βAbauCA. This could open the door to designing inhibitors that selectively target bacterial β-CAs without affecting human α-CAs, as well as offer a novel strategy to weaken A. baumannii and other multidrug-resistant pathogens.