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Background & Aims: Chronic HBV infection is the leading cause of liver disease and of hepatocellular carcinoma. The improvement of antiviral therapy remains an unmet medical need. Capsid assembly modulators (CAMs) target the HBV core antigen (HBc) and inhibit HBV replication. Although CAM-A compounds are well-known inducers of aberrant viral capsid aggregates, their mechanisms of action in HBV-hepatocyte interactions are poorly understood. Recently, we demonstrated that CAM-A molecules lead to a sustained reduction of HBsAg in the serum of HBV replicating mice and induce HBc aggregation in the nucleus of HBc-expressing cells leading to cell death. Methods: The mechanism of action by which CAM-A compounds induce cell death was investigated using an HBV infection model, HBc-overexpressing HepG2-NTCP cells, primary human hepatocytes, and HBV replicating HepAD38 cells. Results: We first confirmed the decrease in HBsAg levels associated with CAM-A treatment and the induction of cell toxicity in HBV-infected differentiated HepaRG cells. Next, we showed that CAM-A-mediated nuclear aggregation of HBc was associated with cell death through the activation of apoptosis. Transcriptomic analysis was used to investigate the mechanism of action driving this phenotype. CAM-A-induced HBc nuclear aggregation led to the upregulation of ANXA1 expression, a documented driver of apoptosis. Finally, silencing of ANXA1 expression delayed cell death and apoptosis in CAM-A-treated cells, confirming its direct involvement in CAM-A-induced cell death. Conclusions: Our results unravel a previously undiscovered mechanism of action involving CAM-As and open the door to new therapeutic strategies involving CAM to achieve a functional cure in patients with chronic infections. Impact and implications:: Chronic HBV infection is a global health threat. To date, no treatment achieves viral clearance in chronically infected patients. In this study, we characterized a new mechanism of action of an antiviral molecule targeting the assembly of the viral capsid (CAM). The study demonstrated that a CAM subtype, CAM-A-induced formation of aberrant structures from HBV core protein aggregates in the nucleus leading to cell death by ANXA1-driven apoptosis. Thus, CAM-A treatment may lead to the specific elimination of HBV-infected cells by apoptosis, paving the way to novel therapeutic strategies for viral cure.

Abstract The conversion of chemical structures into computer-readable descriptors, able to capture key structural aspects, is of pivotal importance in the field of cheminformatics and computer-aided drug design. Molecular fingerprints represent a widely employed class of descriptors; however, their generation process is time-consuming for large databases and is susceptible to bias. Therefore, descriptors able to accurately detect predefined structural fragments and devoid of lengthy generation procedures would be highly desirable. To meet additional needs, such descriptors should also be interpretable by medicinal chemists, and suitable for indexing databases with trillions of compounds. To this end, we developed—as integral part of EXSCALATE, Dompé’s end-to-end drug discovery platform—the DompeKeys (DK), a new substructure-based descriptor set, which encodes the chemical features that characterize compounds of pharmaceutical interest. DK represent an exhaustive collection of curated SMARTS strings, defining chemical features at different levels of complexity, from specific functional groups and structural patterns to simpler pharmacophoric points, corresponding to a network of hierarchically interconnected substructures. Because of their extended and hierarchical structure, DK can be used, with good performance, in different kinds of applications. In particular, we demonstrate how they are very well suited for effective mapping of chemical space, as well as substructure search and virtual screening. Notably, the incorporation of DK yields highly performing machine learning models for the prediction of both compounds’ activity and metabolic reaction occurrence. The protocol to generate the DK is freely available at https://dompekeys.exscalate.eu and is fully integrated with the Molecular Anatomy protocol for the generation and analysis of hierarchically interconnected molecular scaffolds and frameworks, thus providing a comprehensive and flexible tool for drug design applications.

ABSTRACT Tumor suppressor p53 and its related proteins, p63 and p73, can be synthesized as multiple isoforms lacking part of the N- or C-terminal regions. Specifically, high expression of the ΔNp73α isoform is notoriously associated with various human malignancies characterized by poor prognosis. This isoform is also accumulated by oncogenic viruses, such as Epstein–Barr virus (EBV), as well as genus beta human papillomaviruses (HPV) that appear to be involved in carcinogenesis. To gain additional insight into ΔNp73α mechanisms, we have performed proteomics analyses using human keratinocytes transformed by the E6 and E7 proteins of the beta-HPV type 38 virus as an experimental model (38HK). We find that ΔNp73α associates with the E2F4/p130 repressor complex through a direct interaction with E2F4. This interaction is favored by the N-terminal truncation of p73 characteristic of ΔNp73 isoforms. Moreover, it is independent of the C-terminal splicing status, suggesting that it could represent a general feature of ΔNp73 isoforms (α, β, γ, δ, ε, ζ, θ, η, and η1). We show that the ΔNp73α-E2F4/p130 complex inhibits the expression of specific genes, including genes encoding for negative regulators of proliferation, both in 38HK and in HPV-negative cancer-derived cell lines. Such genes are not inhibited by E2F4/p130 in primary keratinocytes lacking ΔNp73α, indicating that the interaction with ΔNp73α rewires the E2F4 transcriptional program. In conclusion, we have identified and characterized a novel transcriptional regulatory complex with potential implications in oncogenesis. IMPORTANCE The TP53 gene is mutated in about 50% of human cancers. In contrast, the TP63 and TP73 genes are rarely mutated but rather expressed as ΔNp63 and ΔNp73 isoforms in a wide range of malignancies, where they act as p53 antagonists. Accumulation of ΔNp63 and ΔNp73, which is associated with chemoresistance, can result from infection by oncogenic viruses such as EBV or HPV. Our study focuses on the highly carcinogenic ΔNp73α isoform and uses a viral model of cellular transformation. We unveil a physical interaction between ΔNp73α and the E2F4/p130 complex involved in cell cycle control, which rewires the E2F4/p130 transcriptional program. Our work shows that ΔNp73 isoforms can establish interactions with proteins that do not bind to the TAp73α tumor suppressor. This situation is analogous to the gain-of-function interactions of p53 mutants supporting cellular proliferation.

Abstract Background Despite the limited number of articles dedicated to its use, augmented reality (AR) is an emerging technology that has shown to have increasing applications in multiple different medical sectors. These include, but are not limited to, the Maxillo-facial and Dentistry disciplines of medicine. In these medical specialties, the focus of AR technology is to achieve a more visible surgical field during an operation. Currently, this goal is brought about by an accurate display of either static or dynamic diagnostic images via the use of a visor or specific glasses. The objective of this study is to evaluate the feasibility of using a virtual display for dynamic navigation via AR. The secondary outcome is to evaluate if the use of this technology could affect the accuracy of dynamic navigation. Case presentation Two patients, both needing implant rehabilitation in the upper premolar area, were treated with flapless surgery. Prior to the procedure itself, the position of the implant was virtually planned and placed for each of the patients using their previous scans. This placement preparation contributed to a dynamic navigation system that was displayed on AR glasses. This, in turn, allowed for the use of a computer-aided/image-guided procedure to occur. Dedicated software for surface superimposition was then used to match the planned position of the implant and the real one obtained from the postoperative scan. Accuracies, using this procedure were evaluated by way of measuring the deviation between real and planned positions of the implants. For both surgeries it was possible to proceed using the AR technology as planned. The deviations for the first implant were 0.53 mm at the entry point and 0.50 mm at the apical point and for the second implant were 0.46 mm at the entry point and 0.48 mm at the apical point. The angular deviations were respectively 3.05° and 2.19°. Conclusions From the results of this pilot study, it seems that AR can be useful in dental implantology for displaying dynamic navigation systems. While this technology did not seem to noticeably affect the accuracy of the procedure, specific software applications should further optimize the results.

Drug transporters play an important role in the maintenance of chemical balance and homeostasis in different tissues. In addition to their physiological functions, they are crucial for the absorption, distribution, and elimination of many clinically important drugs, thereby impacting therapeutic efficacy and toxicity. Increasing evidence has demonstrated that infectious, metabolic, inflammatory, and neurodegenerative diseases alter the expression and function of drug transporters. However, the current knowledge on transporter regulation in critical protective barriers, such as the brain and placenta, is still limited and requires more research. For instance, while many studies have examined P-glycoprotein, it is evident that research on the regulation of highly expressed transporters in the blood–brain barrier and blood–placental barrier are lacking. The aim of this review is to summarize the currently available literature in order to better understand transporter regulation in these critical barriers.

Tumor suppressors can exert pro-proliferation functions in specific contexts. In the beta human papillomavirus type 38 (HPV38) experimental model, the viral proteins E6 and E7 promote accumulation of a wild-type (WT) p53 form in human keratinocytes (HKs), promoting cellular proliferation. Inactivation of p53 by different means strongly decreases the proliferation of HPV38 E6/E7 HKs. This p53 form is phosphorylated at S392 by the double-stranded RNA-dependent protein kinase PKR, which is highly activated by HPV38. PKR-mediated S392 p53 phosphorylation promotes the formation of a p53/DNMT1 complex, which inhibits expression of integrin alpha 1 (ITGA1), a repressor of epidermal growth factor receptor (EGFR) signaling. Ectopic expression of ITGA1 in HPV38 E6/E7 HKs promotes EGFR degradation, inhibition of cellular proliferation, and cellular death. Itga1 expression was also inhibited in the skin of HPV38 transgenic mice that have an elevated susceptibility to UV-induced skin carcinogenesis. In summary, these findings reveal the existence of a specific WT p53 form that displays pro-proliferation properties.

The synthesis of a new dithiolethione-cysteine ethyl ester hybrid, ACS94, its metabolites, and its effect on GSH levels in rat tissues and on the concentration of circulating H2S is described. ACS94 rapidly enters the cells, where it is metabolised to cysteine and the dithiolethione moiety ACS48. Experiments performed through the oral administration of ACS94 to healthy rats showed that it is capable of increasing the GSH levels in most of the analysed organs and the concentration of circulating H2S. Although the increase in GSH concentration was similar to that obtained by ACS48 and N-acetylcysteine ethyl ester, the H2S increase was long-lasting and more evident with respect to the parent molecules. Moreover, a decrease of homocysteine in several rat organs and in plasma was noted. This effect may represent a potential therapeutic use of ACS94, as hyperhomocysteinaemia is considered a risk factor for cardiovascular diseases. Lastly, ACS94 was more efficient than N-acetylcysteine in protecting the liver and kidneys against acute acetaminophen toxicity.

Animal hides are one of man’s earliest and mostly used materials; many rawhide products, primarily leather, have for centuries been used for several purposes. The peculiar mechanical properties of leather depend on the hide composition, a dense collagen feltwork. Unfortunately, due to their proteic composition, rawhides may undergo microbial attack and biodeterioration. Over centuries, different processes and treatments (brining, vegetal or chrome tanning, tawing, etc.) were set up to face the biological attack and modify/stabilise the hide’s mechanical properties. Nevertheless, even present-day rawhides are subjected to biological colonisation, and traces of this colonisation are clearly shown in Chrome(III) tanned leathers (in the wet blue stage), with obvious economic damages. The colonisation traces on tanned leathers consist of isolated or coalescent red patches, known as red heat deterioration. Parchments are rawhide products, too; they derive from another manufacturing procedure. Even parchments undergo microbial attack; the parchment biodeterioration seems comparable to leather red heat deterioration and is known as purple spots. Recently, an ecological succession model explained the process of historical parchment purple spot deterioration; the haloarchaea Halobacterium salinarum is the pioneer organism triggering this attack. The marine salt used to prevent rawhide rotting is the carrier of haloarchaea colonisers (Migliore et al., 2019). The aim of this study was to investigate the dynamics of biodeterioration on Chrome(III) tanned leathers and its effects on the stability/integrity of collagen structure. To this end, standard cultivation methods were integrated with three updated technologies, Next-Generation Sequencing (NGS), Raman spectroscopy, and Light Transmitted Analysis (LTA). A bioinformatic comparison between chrome tanned leather vs. historical parchment colonisers was performed to evaluate if leather and parchment share common culprits; furthermore, the effect of the biodeterioration on the physical properties of the hide product was evaluated.

Eukaryotic mRNA decay is a highly regulated process allowing cells to rapidly modulate protein production in response to internal and environmental cues. Mature translatable eukaryotic mRNAs are protected from fast and uncontrolled degradation in the cytoplasm by two cis-acting stability determinants: a methylguanosine (m(7)G) cap and a poly(A) tail at their 5' and 3' extremities, respectively. The hydrolysis of the m(7)G cap structure, known as decapping, is performed by the complex composed of the Dcp2 catalytic subunit and its partner Dcp1. The Dcp1-Dcp2 decapping complex has a low intrinsic activity and requires accessory factors to be fully active. Among these factors, Pat1 is considered to be a central scaffolding protein involved in Dcp2 activation but also in inhibition of translation initiation. Here, we present the structural and functional study of the C-terminal domain from S. cerevisiae Pat1 protein. We have identified two conserved and functionally important regions located at both extremities of the domain. The first region is involved in binding to Lsm1-7 complex. The second patch is specific for fungal proteins and is responsible for Pat1 interaction with Edc3. These observations support the plasticity of the protein interaction network involved in mRNA decay and show that evolution has extended the C-terminal alpha-helical domain from fungal Pat1 proteins to generate a new binding platform for protein partners.

Parkinson's disease (PD), characterized by loss of dopaminergic neurons in the substantia nigra, is a neurodegenerative disorder of central nervous system. The present study was designed to investigate the therapeutic effect of ACS84, a hydrogen sulfide-releasing-L-Dopa derivative compound, in a 6-hydroxydopamine (6-OHDA)-induced PD model. ACS84 protected the SH-SY5Y cells against 6-OHDA-induced cell injury and oxidative stress. The protective effect resulted from stimulation of Nrf-2 nuclear translocation and promotion of anti-oxidant enzymes expression. In the 6-OHDA-induced PD rat model, intragastric administration of ACS84 relieved the movement dysfunction of the model animals. Immunofluorescence staining and High-performance liquid chromatography analysis showed that ACS84 alleviated the loss of tyrosine-hydroxylase positive neurons in the substantia nigra and the declined dopamine concentration in the injured striatums of the 6-OHDA-induced PD model. Moreover, ACS84 reversed the elevated malondialdehyde level and the decreased glutathione level in vivo. In conclusion, ACS84 may prevent neurodegeneration via the anti-oxidative mechanism and has potential therapeutic values for Parkinson's disease.

Objectives: Neonatal sepsis is a complex syndrome involving an uncontrolled systemic inflammatory response associated with an infection. It may result in the dysfunction or failure of one or more organs or even death. Given its high incidence in premature neonates, the identification of prognostic factors to optimize the early diagnosis and therapeutic interventions are highly desirable. This objective study determine the relationship between inflammatory markers and oxidative parameters as prognostic factors in early neonatal sepsis., Methods: We conducted a prospective observational study by collecting data from 120 patients in the maternity unit of a university hospital. Preterm (<37 weeks of pregnancy) infants with at least one additional risk factor for neonatal sepsis were included. The levels of interleukin (IL)-6, IL-10, thiobarbituric acid reactive species (TBARS) and protein carbonyls and their association with sepsis severity were determined in the cord blood., Results: Levels of IL-6 and TBARS, but not IL-10 and protein carbonyls, demonstrated a mild to moderate correlation with the SNAPPE-II severity score (r=0.435, p=0.02 and r = 0.385, p = 0.017, respectively). No correlations were found between these markers and mortality., Conclusion: TBARS and IL-6 have a mild to moderate correlation with SNAPPE-II, but none of the studied markers were able to predict mortality in our sample.