Carmine Varricchio

Bexarotene (BEX), a retinoid X receptor (RXR) agonist, has demonstrated therapeutic potential in oncology, neurodegenerative diseases and inflammatory conditions, including multiple sclerosis (MS). However, its broader clinical application is limited by poor aqueous solubility, low bioavailability, and dose-limiting adverse effects, most notably hypertriglyceridaemia. In this study, we investigated whether conjugation of BEX to dextrin, a biodegradable polysaccharide, could improve its physicochemical properties and therapeutic index. A dextrin-BEX conjugate was synthesised using succinoylated dextrin (30 mol%) and characterised by gel permeation chromatography (GPC), Fourier-transform infrared (FTIR) spectroscopy, nuclear magnetic resonance (1H-NMR) and dynamic light scattering (DLS). The effects of conjugation on solubility, aggregation behaviour and biological activity were evaluated using in vitro cytotoxicity assays in leukaemia, liver and fibroblast cell lines, alongside measurements of intracellular triglyceride levels. The impact on lipid metabolism and toxicity was further assessed in an in vivo murine model of hyperlipidaemia. Dextrin conjugation significantly increased the aqueous solubility of BEX (from in vitro potency, with IC50 values reduced by up to 14-fold across multiple cell lines. However, the increased cytotoxicity was not selective for cancer cells and was also observed in normal fibroblast and hepatic cell models. The dextrin-BEX conjugate did not induce intracellular triglyceride accumulation in vitro . In vivo , dextrin conjugation did not mitigate BEX-associated lipid dysregulation, while serum triglyceride levels and markers of liver injury remained unchanged. However, although free BEX led to a significant increase in liver weight after 14 days, the dextrin-BEX conjugate did not produce a statistically significant effect. In conclusion, dextrin conjugation markedly improves the solubility and in vitro potency of BEX but does not enhance its selectivity or reduce its metabolic toxicity in vivo . These findings suggest that while polymer conjugation is an effective strategy for addressing physicochemical limitations, it is insufficient to overcome the inherent dyslipidaemia associated with systemic RXR activation. Future strategies should focus on integrating targeting and controlled release mechanisms, alongside approaches that address the underlying pharmacological drivers of lipid dysregulation. Graphical Dextrin conjugation increased bexarotene aqueous solubility and promoted formation of nanoaggregates, leading to enhanced cellular potency. However, the conjugate lacked tumour selectivity and did not mitigate the lipid dysregulation associated with systemic RXR activation in vivo. Graphical abstract generated with the assistance of Google Gemini.

The covalent reversible modification of proteins is a validated strategy for the development of probes and candidate therapeutics. However, the covalent reversible targeting of noncatalytic lysines is particularly challenging. Herein, we characterize the 2-hydroxy-1-naphthaldehyde (HNA) fragment as a targeted covalent reversible ligand of a noncatalytic lysine (Lys⁷²⁰) of the Krev interaction trapped 1 (KRIT1) protein. We show that the interaction of HNA with KRIT1 is highly specific, results in prolonged residence time of >8 h, and inhibits the Heart of glass 1 (HEG1)-KRIT1 protein-protein interaction (PPI). Screening of HNA derivatives identified analogs exhibiting similar binding modes as the parent fragment but faster target engagement and stronger inhibition activity. These results demonstrate that HNA is an efficient site-directing fragment with promise in developing HEG1-KRIT1 PPI inhibitors. Further, the aldimine chemistry, when coupled with templating effects that promote proximity, can produce a long-lasting reversible covalent modification of noncatalytic lysines.

Ubiquitin phosphorylation by the mitochondrial protein kinase PTEN-induced kinase 1 (PINK1), upon mitochondrial depolarization, is an important intermediate step in the recycling of damaged mitochondria via mitophagy. As mutations in PINK1 can cause early-onset Parkinson's disease (PD), there has been a growing interest in small-molecule activators of PINK1-mediated mitophagy as potential PD treatments. Herein, we show that N⁶-substituted adenosines, such as N⁶-(2-furanylmethyl)adenosine (known as kinetin riboside) and N⁶-benzyladenosine, activate PINK1 in HeLa cells and induce PINK1-dependent mitophagy in primary mouse fibroblasts. Interestingly, pre-treatment of HeLa cells and astrocytes with these compounds inhibited elevated ubiquitin phosphorylation that is induced by established mitochondrial depolarizing agents, carbonyl cyanide m-chlorophenyl-hydrazine and niclosamide. Together, this highlights N⁶-substituted adenosines as progenitor PINK1 activators that could potentially be developed, in the future, as treatments for aged and sporadic PD patients who have elevated phosphorylated ubiquitin levels in the brain.

The oxidative aromatization of aliphatic N-heterocycles is a fundamental organic transformation for the preparation of a diverse array of heteroaromatic compounds. Despite many attempts to improve the efficiency and practicality of this transformation, most synthetic methodologies still require toxic and expensive reagents as well as harsh conditions. Herein, we describe two enzymatic strategies for the oxidation of 1,2,3,4-tetrahydroquinolines (THQs) and N-cyclopropyl-N-alkylanilines into quinolines and 2-quinolones, respectively. Whole cells and purified monoamine oxidase (MAO-N) enzymes were used to effectively catalyze the biotransformation of THQs into the corresponding aromatic quinoline derivatives, while N-cyclopropyl-N-alkylanilines were converted into 2-quinolone compounds through a horseradish peroxidase (HRP)-catalyzed annulation/aromatization reaction followed by Fe-mediated oxidation.

The phosphorylation of ubiquitin by the mitochondrial protein kinase PINK1, upon mitochondrial depolarization, is an important step in the repair and recycling of the mitochondria via mitophagy. As mutations in PINK1 can cause early-onset Parkinson’s disease (PD), there has been a growing interest in small molecule modulators of PINK1-mediated mitophagy as potential PD treatments. In this work, we show that in HeLa cells, primary mouse fibroblasts and astrocytes, N6-substituted adenosines, such as N6-(2- furanylmethyl)adenosine (known as kinetin riboside) and N6-benzyladenosine, inhibit ubiquitin phosphorylation that is induced by established mitochondrial depolarizing agents, CCCP and niclosamide. Although these nucleoside analogues inhibited niclosamide- and CCCP-induced ubiquitin phosphorylation, they did not prevent the mitochondrial membrane depolarization. Notably, treatment of cells with these nucleoside analogues alone induced low level mitophagy and did not cause mitochondrial fragmentation. Together, this work presents N6-substituted adenosines as new inhibitors of CCCP- and niclosamide-induced PINK1-mediated ubiquitin phosphorylation and highlights their potential utility in treating aged and sporadic PD as well as Lewy body dementia patients who have elevated levels of phosphorylated ubiquitin.

Fungal keratitis, a disease in which the cornea becomes inflamed due to an invasive fungal infection, remains difficult to treat due in part to limited choices of available treatments. Topical eye drops are first-line treatment, but can be ineffective as low levels of drug reach the target site due to precorneal losses and the impenetrability of the cornea. The aim of this study was to determine the corneal delivery of econazole using a novel topical enhancement approach using a composite delivery system based upon cyclodextrins and soft hydrogel contact lenses. Excess econazole nitrate was added to hydroxypropyl-α-cyclodextrin (HP-α-CD) and hydroxypropyl-β-cyclodextrin (HP-β-CD) solutions, and the solubility determined using HPLC. Proprietary soft hydrogel contact lenses were then impregnated with saturated solutions and applied to freshly enucleated porcine eyeballs. Econazole nitrate ‘eye drops’ at the same concentrations served as the control. After 6 h, the corneas were excised and drug-extracted, prior to quantification using HPLC. Molecular dynamic simulations were performed to examine econazole–HP-β-CD inclusion complexation and dissociation. The minimum inhibitory concentration (MIC) of econazole was determined against four fungal species associated with keratitis, and these data were then related to the amount of drug delivered to the cornea, using an average corneal volume of 0.19 mL. The solubility of econazole increased greatly in the presence of HP-β-CD and more so with HP-α-CD (p < 0.001), with ratios >> 2. Hydrogel contact lenses delivered ×2.8 more drug across the corneas in comparison to eye drops alone, and ×5 more drug delivered to the cornea when cyclodextrin was present. Molecular graphics demonstrated dynamic econazole release, which would create transient enhanced drug concentration at the cornea surface. The solution-only drops achieved the least satisfactory result, producing sub-MIC levels with factors of ×0.81 for both Fusarium semitectum and Fusarium solani and ×0.40 for both Scolecobasidium tshawytschae and Bipolaris hawaiiensis. All other treatments delivered econazole at > MIC for all four fungal species. The efficacies of the delivery platforms evaluated were ranked: HP-α-CD contact lens > HP-β-CD contact lens > contact lens = HP-α-CD drops > HP-β-CD drops > solution-only drops. In summary, the results in this study have demonstrated that a composite drug delivery system based upon econazole–HP-β-CD inclusion complexes loaded into contact lenses can achieve significantly greater corneal drug delivery with the potential for improved clinical responses.

Summary: SARS-CoV-2 is currently causing an unprecedented pandemic. While vaccines are massively deployed, we still lack effective large-scale antiviral therapies. In the quest for antivirals targeting conserved structures, we focused on molecules able to bind viral RNA secondary structures. Aminoglycosides are a class of antibiotics known to interact with the ribosomal RNA of both prokaryotes and eukaryotes and have previously been shown to exert antiviral activities by interacting with viral RNA. Here we show that the aminoglycoside geneticin is endowed with antiviral activity against all tested variants of SARS-CoV-2, in different cell lines and in a respiratory tissue model at non-toxic concentrations. The mechanism of action is an early inhibition of RNA replication and protein expression related to a decrease in the efficiency of the -1 programmed ribosomal frameshift (PRF) signal of SARS-CoV-2. Using in silico modelling, we have identified a potential binding site of geneticin in the pseudoknot of frameshift RNA motif. Moreover, we have selected, through virtual screening, additional RNA binding compounds, interacting with the same site with increased potency.

There is a need for new antimicrobial systems due to increased global resistance to current antimicrobials. Pomegranate rind extract (PRE) and Zn (II) ions both possess a level of antimicrobial activity and work has previously shown that PRE/Zn (II) in combination possesses synergistic activity against Herpes simplex virus and Micrococcus luteus. Here, we determined whether such synergistic activity extended to other, more pathogenic, bacteria. Reference strains of methicillin-resistant Staphylococcus aureus (MRSA), Staphylococcus epidermidis, Escherichia coli, and Pseudomonas aeruginosa were cultured and subjected to challenge by PRE, Zn (II), or PRE + Zn (II), in time-kill assays. Data were obtained independently by two researchers using different PRE preparations. Statistically significant synergistic activity for PRE + Zn (II) was shown for all four bacterial strains tested compared to untreated controls, although the extent of efficacy and timescales varied. Zn (II) exerted activity and at 1 h, it was not possible to distinguish with PRE + Zn (II) combination treatment in all cases. PRE alone showed low activity against all four bacteria. Reproducible synergistic bactericidal activity involving PRE and Zn (II) has been confirmed. Potential mechanisms are discussed. The development of a therapeutic system that possesses demonstrable antimicrobial activity is supported which lends itself particularly to topical delivery applications, for example MRSA infections.