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Attilio Varricchio,1 Giorgio Ciprandi,2 Study Group on respiratory infection management in clinical practice included:Francesco Paolo Brunese, Luigi Costanzo, Lorenzo Cresta, Maurizio Daccò, Donatella Del Gaizo, Carla Dominici, Cosimo Lanza, Michele Lisco, Anastasia Matteucig, Carlotta Moretti On behalf of the Study Group on respiratory infection management in clinical practice1Department of Otolaryngology, University of Molise, Campobasso, Italy; 2Allergy Clinic, Casa di Cura Villa Montallegro, Genoa, ItalyCorrespondence: Giorgio Ciprandi, Casa di Cura Villa Montallegro, Genoa, Italy, Email gio.cip@libero.itPurpose: Acute upper respiratory infections (AURI) represent a daily challenge in primary care practice. Mucus production may impair during AURI. Sobrerol is a muco-active agent that improves rheological characteristics and exerts other ancillary activities. The aim of this retrospective case–series study was to compare the efficacy and safety of different uses of sobrerol (only oral, only nebulized, and combined or standard treatment for infections alone) in patients with AURIs.Patients and Methods: The present clinical experience retrospectively collected clinical data of patients with AURIs visited by ten primary care doctors (pediatricians and general practitioners) over a long period. Patients could take standard therapy for infections, or as add-on: oral sobrerol, nebulized sobrerol, or combined oral and nebulized aerosol during the infection for 3 days.Results: Patients treated with combined oral and nebulized sobrerol experienced less intense symptoms, mainly concerning cough (p < 0.001) and nasal complaints (p = 0.043). In addition, the patients taking the combined therapy reported a more rapid disappearance of cough and nasal symptoms at day 7 than patients treated with the other options (OR 4.47 and 3.16, respectively).Conclusion: The current retrospective and observational study showed that a three-day combined (oral and nebulized) sobrerol course may represent a valuable add-on option in patients with AURIs.Keywords: acute upper respiratory infections, combined therapy, nebulization, oral formulation, primary care, sobrerol

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.

Tubulin and microtubules (MTs) are potential protein targets to treat parasitic infections and our previous studies have shown that the triazolopyrimidine (TPD) class of MT-active compounds hold promise as antitrypanosomal agents. MT-targeting TPDs include structurally related but functionally diverse congeners that interact with mammalian tubulin at either one or two distinct interfacial binding sites; namely, the seventh and vinca sites, which are found within or between α,β-tubulin heterodimers, respectively. Evaluation of the activity of 123 TPD congeners against cultured Trypanosoma brucei enabled a robust quantitative structure-activity relationship (QSAR) model and the prioritization of two congeners for in vivo pharmacokinetics (PK), tolerability and efficacy studies. Treatment of T. brucei-infected mice with tolerable doses of TPDs significantly decreased blood parasitemia within 24 h. Further, two once-weekly doses at 10 mg/kg of a candidate TPD significantly extended the survival of infected mice relative to infected animals treated with vehicle. Further optimization of dosing and/or the dosing schedule of these CNS-active TPDs may provide alternative treatments for human African trypanosomiasis.

Purpose Anti-programmed cell death 1 (PD1) is the first-choice treatment in patients with advanced cutaneous squamous cell carcinoma (cSCC), when curative options are unavailable. However, reliable biomarkers for patient selection are still lacking.Experimental design In this translational study, clinical annotations, tissue and liquid biopsies were acquired to investigate the association between sustained objective responses and transcriptional profiles, immune cell dynamics in tumor tissue and peripheral blood samples, as well as circulating cytokine levels.Results First, we investigated the baseline characteristics of the immune landscape of cSCC biopsies. Gene Set Enrichment Analysis showed upregulation of interleukin (IL)2/STAT5 pathways and downregulation of Interferon signatures in non-responder patients compared with responders. Next, we studied the early changes induced by cemiplimab in tissue biopsies. Notably, after only three weeks, cemiplimab treatment induced an increase in B cells and CD8+ T cells in responders, whereas their abundance decreased in non-responder patients. Moreover, analyzing differentially expressed genes modulated early during treatment, compared with baseline biopsies, we found that IL1β and IL8 exhibited early downregulation in responder patients’ tumor specimens. We assessed whether changes in the local tumor microenvironment were mirrored in peripheral blood. Similar to tissue findings, no changes were observed in the whole T regulatory (Treg) population, although PD1+ Tregs, which were downregulated in responder patients (vs T0), showed a rebound enrichment in non-responders after three cycles of cemiplimab. Finally, IL8 mirrored the tissue results, unlike IL1β, with early (T1) and then sustained (T3) downregulation of its levels in responder patients, while increased in non-responders.Conclusions Taken together, these findings shed light on the significance of early transcriptomic and immune cell modulation in predicting responses to cemiplimab therapy. Additionally, our data suggest that IL8 levels in peripheral blood offer promising avenues for personalized treatment selection and response assessment in patients with cSCC receiving cemiplimab, while PD1+Tregs can be followed longitudinally to monitor response to therapy.

Gut microbiota plays an important role in maintaining respiratory health while providing protection against respiratory illnesses. However, the clinical role of probiotic strains in prevention and treatment of early-onset asthma is still debated. The present study aims to systematically review the available literature on the efficacy of probiotic supplementation, either alone or in combination with other treatments, in managing children with asthma. An electronic search was conducted across multiple databases including PubMed, Web of Science, and EMBASE. The articles reported in English, regardless of year of publication, were reviewed by independent authors for inclusion assessment. Randomized controlled studies evaluating the efficacy of probiotics in children with asthma were included in the study. Of the 203 articles reviewed, 16 were selected and summarized to provide a consistent review, and of these, eight studies reported the anti-asthmatic effects of the treatment with single and multi-strain probiotics alone, seven reported the use of probiotic strains in combination with other treatments, and one was a follow-up study. While some of the studies reported are well conducted and with promising results, no cumulative significant impact was seen for probiotic supplementation on disease parameters of asthma in children. However, the available literature is not sufficient to draw any solid conclusion about the possible use of probiotics in children with asthma, and it is necessary to expand this literature by conducting more well-designed randomized clinical trials to assess the effects of probiotic strains in children with asthma or carrying a high-risk of asthma.

In the current study, we introduced a unique method for identifying and segmenting oral squamous cell carcinoma (OSCC) nuclei, concentrating on those predicted to have significant CAF-1/p60 protein expression. Our suggested model uses the StarDist architecture, a deep-learning framework designed for biomedical image segmentation tasks. The training dataset comprises painstakingly annotated masks created from tissue sections previously stained with hematoxylin and eosin (H&E) and then restained with immunohistochemistry (IHC) for p60 protein. Our algorithm uses subtle morphological and colorimetric H&E cellular characteristics to predict CAF-1/p60 IHC expression in OSCC nuclei. The StarDist-based architecture performs exceptionally well in localizing and segmenting H&E nuclei, previously identified by IHC-based ground truth. In summary, our innovative approach harnesses deep learning and multimodal information to advance the automated analysis of OSCC nuclei exhibiting specific protein expression patterns. This methodology holds promise for expediting accurate pathological assessment and gaining deeper insights into the role of CAF-1/p60 protein within the context of oral cancer progression.

Anatomical pathology is undergoing its third revolution, transitioning from analogical to digital pathology and incorporating new artificial intelligence technologies into clinical practice. Aside from classification, detection, and segmentation models, predictive models are gaining traction since they can impact diagnostic processes and laboratory activity, lowering consumable usage and turnaround time. Our research aimed to create a deep-learning model to generate synthetic Ki-67 immunohistochemistry from Haematoxylin and Eosin (H&E) stained images. We used 175 oral squamous cell carcinoma (OSCC) from the University Federico II’s Pathology Unit’s archives to train our model to generate 4 Tissue Micro Arrays (TMAs). We sectioned one slide from each TMA, first stained with H&E and then re-stained with anti-Ki-67 immunohistochemistry (IHC). In digitised slides, cores were disarrayed, and the matching cores of the 2 stained were aligned to construct a dataset to train a Pix2Pix algorithm to convert H&E images to IHC. Pathologists could recognise the synthetic images in only half of the cases in a specially designed likelihood test. Hence, our model produced realistic synthetic images. We next used QuPath to quantify IHC positivity, achieving remarkable levels of agreement between genuine and synthetic IHC.Furthermore, a categorical analysis employing 3 Ki-67 positivity cut-offs (5%, 10%, and 15%) revealed high positive-predictive values. Our model is a promising tool for collecting Ki-67 positivity information directly on H&E slides, reducing laboratory demand and improving patient management. It is also a valuable option for smaller laboratories to easily and quickly screen bioptic samples and prioritise them in a digital pathology workflow.

Background Telomeres maintain chromosome stability, while telomerase counteracts their progressive shortening. Telomere length varies between cell types, with leukocyte telomere length (LTL) decreasing with age. Reduced telomerase activity has been linked to reproductive issues in females, such as low pregnancy rates and premature ovarian failure, with recent studies indicating correlations between telomere length in granulosa cells and IVF outcomes.Objectives The study aims to explore the relationship between telomere length, telomerase activity, and euploid blastocyst rate in infertile women undergoing IVF/ICSI PGT-A cycles.Methods This prospective study involves 108 patients undergoing controlled ovarian stimulation and PGT-A. Telomere length and telomerase activity were measured in peripheral mononuclear cells and granulosa cells (GC), respectively.Results The telomere repeat copy number to single gene copy number ratio (T/S) results respectively 0.6 ± 0.8 in leukocytes and 0.7 ± 0.9 in GC. An inverse relationship was found between LTL and the patient’s age (p

Respiratory infections are a relevant issue during infancy and childhood, constituting a major source of morbidity and imposing a substantial burden on healthcare systems. Conditions such as bronchiolitis, pneumonia, and influenza exemplify the gravity of these infections among pediatric populations. In the context of this health challenge, resveratrol, a naturally occurring polyphenolic antioxidant has emerged as a focal point of extensive research, known for its antioxidant, anti-inflammatory, and antimicrobial properties. This comprehensive review delves into the compound's extensive antimicrobial activities against a diverse range of bacteria and viruses, both as a standalone agent and in combination with conventional antibiotics. This compound exhibits notable antiviral activity against viruses implicated in severe respiratory infections, offering a potential therapeutic avenue. By exploring the antiviral mechanisms and highlighting the anti-inflammatory potential of resveratrol, this comprehensive overview contributes to a deeper understanding of its applicability in mitigating the complexities associated with pediatric respiratory infections.

Microtubule (MT)-stabilizing 1,2,4-triazolo[1,5-a]pyrimidines (TPDs) hold promise as candidate therapeutics for Alzheimer's disease (AD) and other neurodegenerative conditions. However, depending on the choice of substituents around the TPD core, these compounds can elicit markedly different cellular phenotypes that likely arise from the interaction of TPD congeners with either one or two spatially distinct binding sites within tubulin heterodimers (i.e., the seventh site and the vinca site). In the present study, we report the design, synthesis, and evaluation of a series of new TPD congeners, as well as matched molecular pair analyses and computational studies, that further elucidate the structure-activity relationships of MT-active TPDs. These studies led to the identification of novel MT-normalizing TPD candidates that exhibit favorable ADME-PK, including brain penetration and oral bioavailability, as well as brain pharmacodynamic activity.

Abstract Resveratrol is a nonflavonoid polyphenolic compound with a chemical structure consisting of two aromatic rings linked by a methylene bridge. It exists in two primary isomers and has a broad range of desirable biological activities, including antioxidant, anti‐inflammatory, antidiabetic, cardioprotective, and antitumor activities. Some antioxidant properties of resveratrol are known with certainty, such as its potential to positively impact cardiovascular health, inflammation, and the metabolism. On the other hand, many uncertainties and controversies plague its efficacy, including issues related to its bioavailability, dosing, human clinical trial results, interactions with other food components and drugs, and individual variability. In brief, although promising results have been observed in in vitro and in vivo studies, the translation of these findings to human health remains uncertain. Many human clinical trials on resveratrol are ongoing or have proven inconclusive, making it challenging to definitively determine its efficacy for specific health conditions and its dose and duration of treatment. Resveratrol may interact with medications and have varying effects on individuals. In conclusion, it is essential to approach resveratrol with a balanced perspective, consulting with healthcare professionals, and considering the evolving scientific evidence when making decisions regarding its clinical use.

Background: Despite advances in uveal melanoma (UM) diagnosis and treatment, about 50% of patients develop distant metastases, thereby displaying poor overall survival. Molecular profiling has identified several genetic alterations that can stratify patients with UM into different risk categories. However, these genetic alterations are currently dispersed over multiple studies and several methodologies, emphasizing the need for a defined workflow that will allow standardized and reproducible molecular analyses. Methods: Following the findings published by “The Cancer Genome Atlas–UM” (TCGA-UM) study, we developed an NGS-based gene panel (called the UMpanel) that classifies mutation sets in four categories: initiating alterations (CYSLTR2, GNA11, GNAQ and PLCB4), prognostic alterations (BAP1, EIF1AX, SF3B1 and SRSF2), emergent biomarkers (CDKN2A, CENPE, FOXO1, HIF1A, RPL5 and TP53) and chromosomal abnormalities (imbalances in chromosomes 1, 3 and 8). Results: Employing commercial gene panels, reference mutated DNAs and Sanger sequencing, we performed a comparative analysis and found that our methodological approach successfully predicted survival with great specificity and sensitivity compared to the TCGA-UM cohort that was used as a validation group. Conclusions: Our results demonstrate that a reproducible NGS-based workflow translates into a reliable tool for the clinical stratification of patients with UM.

Thymic epithelial tumors are rare malignancies with an incidence of 1.7 cases per million people per year. They pose significant management challenges due to their association with autoimmune disorders. In this case report, we present the 21-year history of a patient diagnosed with advanced B2/B3 thymoma and Good’s syndrome. The patient achieved a complete and durable response after receiving only two cycles of the immune checkpoint inhibitor Nivolumab. However, this positive outcome was accompanied by the development of severe immune-related myocarditis complicated by reactivation of cytomegalovirus. Moreover, the patient developed a highly uncommon subdiaphragmatic pararectal dissemination of the thymic tumor, which is a condition rarely described in the literature. Despite the success in achieving complete and durable response with immune checkpoint inhibitors, the emergence of immune-related adverse events highlights the potential challenges associated with these treatments, emphasizing the need for careful monitoring and a comprehensive understanding of the intricate interplay between cancer, immune system dysregulations and immunotherapy.

Since the start of the pandemic, scientists have directed their research towards identifying COVID-19 risk factors and predictive elements. Numerous clinical studies have established a strong connection between hypoalbuminemia and an unfavorable prognosis for COVID-19. Here, we aim to explore the impact of human serum albumin (HSA) on SARS-CoV-2 infection. Our findings indicate that HSA plays a role in reducing the replication rate of SARS-CoV-2 in Vero E6 cells. This protective effect is due to HSA ability to bind to the S1 domain of the spike protein, effectively competing with ACE2. Moreover, we show that the protective role of HSA is dependent also on its ability to activate the protective axis within the RAS system pathway, which is responsible for inducing vasodilation and promoting anti-inflammatory, anti-fibrotic, and anti-apoptotic responses. In summary, the data presented in this study support the idea that reduced levels of circulating HSA in hypoalbuminemic patients may heighten their susceptibility to SARS-CoV-2 infection, as the spike protein is unhindered in its ability to bind to ACE2 and penetrate human cells. Besides, hypoalbuminemia exacerbates the imbalance of the RAS pathway towards the classical “detrimental” axis. This could potentially contribute to the increased severity and elevated mortality rate observed in individuals with low levels of circulating albumin.

Objectives: Elderly patients with hematologic malignancies face the highest risk of severe COVID-19 outcomes. The infection's impact on different age groups remains unstudied in detail. Methods: We analyzed elderly patients (age groups: 65-70, 71-75, 76-80, and >80 years old) with hematologic malignancies included in the EPICOVIDEHA registry between January 2020 and July 2022. Univariable and multivariable Cox regression models were conducted to identify factors influencing death in COVID-19 patients with hematological malignancy. Results: The study included data from 3,603 elderly patients (aged 65 or older) with hematological malignancy, with a majority being male (58.1%) and a significant proportion having comorbidities. The patients were divided into four age groups, and the analysis assessed COVID-19 outcomes, vaccination status, and other variables in relation to age and pandemic waves. The 90-day survival rate for patients with COVID-19 was 71.2%, with significant differences between groups. The pandemic waves had varying impacts, with the first wave affecting patients over 80 years old, the second being more severe in 65-70, and the third being the least severe in all age groups. Factors contributing to 90-day mortality included age, comorbidities, lymphopenia, active malignancy, acute leukemia, less than three vaccine doses, severe COVID-19, and using only corticosteroids as treatment. Conclusion: These data underscore the heterogeneity of elderly hematological patients, highlight the different impacts of COVID-19 waves and the pivotal importance of vaccination, and may help in planning future healthcare efforts.

SARS-CoV-2 proteases Mpro and PLpro are promising targets for antiviral drug development. In this study, we present an antiviral screening strategy involving a novel in-cell protease assay, antiviral and biochemical activity assessments, as well as structural determinations for rapid identification of protease inhibitors with low cytotoxicity. We identified eight compounds with anti-SARS-CoV-2 activity from a library of 64 repurposed drugs and modeled at protease active sites by in silico docking. We demonstrate that Sitagliptin and Daclatasvir inhibit PLpro, and MG-101, Lycorine HCl, and Nelfinavir mesylate inhibit Mpro of SARS-CoV-2. The X-ray crystal structure of Mpro in complex with MG-101 shows a covalent bond formation between the inhibitor and the active site Cys145 residue indicating its mechanism of inhibition is by blocking the substrate binding at the active site. Thus, we provide methods for rapid and effective screening and development of inhibitors for blocking virus polyprotein processing as SARS-CoV-2 antivirals. Additionally, we show that the combined inhibition of Mpro and PLpro is more effective in inhibiting SARS-CoV-2 and the delta variant.