Federico Iacovelli

Molecular dynamics has been widely used in nanofluidics as it provides an atomistic description of fluid transport that cannot be achieved experimentally. The interactions among atoms are modeled by empirical force fields whose reliability for a specific problem must be tested against experimental evidence. Such validation is often difficult to obtain in nanofluidics due to the challenges associated with measuring nanoscale flows. Here, we systematically compare three popular force fields (CHARMM36, Amber-ff14SB, both with TIP3P water, and Amber-ff19SB with the OPC water) in terms of their electrohydrodynamical outputs. As test cases, we used CytK and MspA, two biological nanopores employed in sensing. For each nanopore, we simulated one cation-selective and one anion-selective mutant. Our results show that while, overall, the anion/cation selectivity and the direction of electroosmosis are coherent across the force fields, quantitative differences are observed. These differences cannot be simply explained by the transport properties of the solution, such as the ion conductivity and viscosity. For narrow pores such as MspA, these differences can become so significant as to yield qualitatively different outcomes; for example, one force field may predict no detectable electroosmosis while another shows a clear electroosmotic flow.

The RNA-binding protein HuR (embryonic lethal abnormal vision-like protein 1) regulates mRNA stability and translation. HuR contains three RNA-recognition motifs (RRMs): the RRM1 and RRM2 confer high-affinity mRNA binding, while RRM3 mediates protein oligomerization. Although HuR is predominantly nuclear, cellular stimuli trigger its cytoplasmic translocation via a nucleocytoplasmic shuttling sequence between the RRM2 and RRM3 domains. Despite HuR's critical role in post-transcriptional gene regulation, its full-length three-dimensional (3D) structure remains uncharacterized. In this study, we employed an in silico approach, combining molecular modeling, atomistic, and coarse-grained molecular dynamics simulations to build and validate a 3D model of the full-length HuR in complex with an mRNA fragment. Structural analysis of the model identified a tyrosine residue as a key mediator of HuR-RNA interaction stability and provided novel structural insights into HuR's RNA-binding mechanism, contributing to a deeper understanding of its regulatory functions.

Cancer remains a leading cause of death worldwide, highlighting the urgent need for novel and more effective treatments. Natural products, with their structural diversity, represent a valuable source for the discovery of anticancer compounds. In this study, we screened 750 Antarctic extracts to identify potential inhibitors of human topoisomerase 1 (hTOP1), a key enzyme in DNA replication and repair, and a target of cancer therapies. Bioassay-guided fractionation led to the identification of palmitic acid (PA) as the active compound from the Antarctic sponge Artemisina plumosa, selectively inhibiting hTOP1. Our results demonstrate that PA irreversibly blocks hTOP1-mediated DNA relaxation and specifically inhibits the DNA religation step of the enzyme’s catalytic cycle. Unlike other fatty acids, PA exhibited unique specificity, which we confirmed through comparisons with linoleic acid. Molecular dynamics simulations and binding assays further suggest that PA interacts with hTOP1-DNA complexes, enhancing the inhibitory effect in the presence of camptothecin (CPT). These findings identify PA as a hTOP1 inhibitor with potential therapeutic implications, offering a distinct mechanism of action that could complement existing cancer therapies.

Candida albicans (C. albicans) is the most common pathogen responsible for a wide spectrum of human infections ranging from superficial mucocutaneous mycoses to systemic life-threatening diseases. Its main virulence factors are the morphological transition between yeast and hyphal forms and the ability to produce biofilm. Novel antifungal strategies are required given the severity of systemic candidiasis, especially in immunocompromised patients, and the lack of effective anti-biofilm treatments. We previously demonstrated that all-trans retinoic acid (ATRA), an active metabolite of vitamin A, exerted an inhibitory effect on Candida growth, yeast–hyphal transition and biofilm formation. Here, we further investigated the possible anti-Candida potential of trifarotene and tazarotene, which are the other two molecules belonging to the retinoid family, compared to ATRA. The results indicate that both drugs were able to suppress Candida growth, germination and biofilm production, although trifarotene was proven to be more effective than tazarotene, showing effectiveness comparable to ATRA. In silico studies suggest that all three retinoids may exert antifungal activity through their molecular interactions with the heat shock protein (Hsp) 90 and 14α-demethylase of C. albicans. Moreover, interactions between retinoids and ergosterol have been observed, suggesting that those compounds have great potential against C. albicans infections.

We report here a strategy to control cell-free in vitro transcription of RNA by the rational design of synthetic structure-switching DNA templates. These templates are programmed to undergo a conformational change between an open (OFF) and a closed (ON) state. By controlling the switching equilibrium constant (KS) of such structure-switching templates, we can finely program their transcription efficiency. We have derived a simplified kinetic model that describes our experimental data with high accuracy, supporting the predictability of our approach. Similarly, we have also performed different molecular dynamic simulations that provide an atomistic description of our results. Finally, the introduction of such a conformational change mechanism in target-responsive templates allows rational control of their dynamic range (i.e., the range of target concentration to which they respond) and of their input-output behavior. The structure-switching templates demonstrated in this work are particularly easy to design and provide a highly programmable approach to control cell-free transcription, a feature that can be of great utility in fields like sensing and RNA therapeutics.

Desert strains of the genus Chroococcidiopsis are among the most desiccation-resistant cyanobacteria capable of anhydrobiosis. The accumulation of two sugars, sucrose and trehalose, facilitates the entrance of anhydrobiotes into a reversible state of dormancy by stabilizing cellular components upon water removal. This study aimed to evaluate, at the atomistic level, the role of trehalose in desiccation resistance by using as a model system the 30S ribosomal subunit of the desert cyanobacterium Chroococcidiopsis sp. 029. Molecular dynamic simulations provided atomistic evidence regarding its protective role on the 30S molecular structure. Trehalose forms an enveloping shell around the ribosomal subunit and stabilizes the structures through a network of direct interactions. The simulation confirmed that trehalose actively interacts with the 30S ribosomal subunit and that, by replacing water molecules, it ensures ribosomal structural integrity during desiccation, thus enabling protein synthesis to be carried out upon rehydration.

(1) Objective: Keloid and hypertrophic scars are a challenge in clinical management, causing functional and psychological discomfort. These pathological scars are caused by a proliferation of dermal tissue following skin injury. The TGF-β/Smad signal pathway in the fibroblasts and myofibroblasts is involved in the scarring process of skin fibrosis. Today, multiple therapeutic strategies that target the TGF-β/Smad signal pathway are evaluated to attenuate aberrant skin scars that are sometimes difficult to manage. We performed a head-to-head, randomized controlled trial evaluating the appearance of the post-surgical scars of 64 subjects after two times daily topical application to compare the effect of a class I pullulan-based medical device containing Allium cepa extract 5% and hyaluronic acid 5% gel versus a class I medical device silicone gel on new post-surgical wounds. (2) Methods: Objective scar assessment using the Vancouver Scar Scale (VSS), POSAS, and other scales were performed after 4, 8, and 12 weeks of treatment and statistical analyses were performed. The trial was registered in clinicalTrials.gov ( NCT05412745). In parallel, molecular docking simulations have been performed to investigate the role of Allium cepa in TGF-β/Smad signal pathway. (3) Results: We showed that VSS, POSAS scale, itching, and redness reduced significantly at week 4 and 8 in the subjects using devices containing Allium cepa and HA. No statistically significant differences in evaluated scores were noted at 12 weeks of treatment. Safety was also evaluated by gathering adverse events related to the application of the gel. Subject compliance and safety with the assigned gel were similar between the two study groups. Molecular docking simulations have shown how Allium cepa could inhibit fibroblasts proliferation and contraction via TGF-β/Smad signal pathway. (4) Conclusions: The topical application of a pullulan-based medical device containing Allium cepa and HA showed a clear reduction in the local inflammation, which might lead to a reduced probability of developing hypertrophic scars or keloids.

Tea Tree Oil (TTO) is an essential oil obtained from the distillation of Melaleuca alternifolia leaves and branches. Due to its beneficial properties, TTO is widely used as an active ingredient in antimicrobial preparations for topical use or in cosmetic products and contains about 100 different compounds, with terpinen-4-ol, γ-terpinene and 1,8-cineole (or eucalyptol) being the molecules most responsible for its biological activities. In this work, the antimicrobial activity of whole TTO and these three major components was evaluated in vitro against fungi, bacteria and viruses. Molecular dynamics simulations were carried out on a bacterial membrane model and a Coxsackievirus B4 viral capsid, to propose an atomistic explanation of their mechanism of action. The obtained results indicate that the strong antimicrobial activity of TTO is attributable to the induction of an altered membrane functionality, mediated by the incorporation of its components within the lipid bilayer, and to a possible ability of the compounds to bind and alter the structural properties of the viral capsid.

Human topoisomerase 1B regulates the topological state of supercoiled DNA enabling all fundamental cell processes. This enzyme, which is the unique molecular target of the natural anticancer compound camptothecin, acts by nicking one DNA strand and forming a transient protein–DNA covalent complex. The interaction of human topoisomerase 1B and dimethylmyricacene, a compound prepared semisynthetically from myricanol extracted from Myrica cerifera root bark, was investigated using enzymatic activity assays and molecular docking procedures. Dimethylmyricacene was shown to inhibit both the cleavage and the religation steps of the enzymatic reaction, and cell viability of A-253, FaDu, MCF-7, HeLa and HCT-116 tumor cell lines.

SARS-CoV-2 causes COVID-19, a predominantly pulmonary disease characterized by a burst of pro-inflammatory cytokines and an increase in free iron. The viral glycoprotein Spike mediates fusion to the host cell membrane, but its role as a virulence factor is largely unknown. Recently, the antiviral activity of lactoferrin against SARS-CoV-2 was demonstrated in vitro and shown to occur via binding to cell surface receptors, and its putative interaction with Spike was suggested by in silico analyses. We investigated the anti-SARS-CoV-2 activity of bovine and human lactoferrins in epithelial and macrophagic cells using a Spike-decorated pseudovirus. Lactoferrin inhibited pseudoviral fusion and counteracted the deleterious effects of Spike on iron and inflammatory homeostasis by restoring basal levels of iron-handling proteins and of proinflammatory cytokines IL-1β and IL-6. Using pull-down assays, we experimentally proved for the first time that lactoferrin binds to Spike, immediately suggesting a mechanism for the observed effects. The contribution of transferrin receptor 1 to Spike-mediated cell fusion was also experimentally demonstrated. In silico analyses showed that lactoferrin interacts with transferrin receptor 1, suggesting a multifaceted mechanism of action for lactoferrin. Our results give hope for the use of bovine lactoferrin, already available as a nutraceutical, as an adjuvant to standard therapies in COVID-19.