A paclitaxel-hyaluronan conjugate (ONCOFID-P-B™) in patients with BCG-unresponsive carcinoma in situ of the bladder: a dynamic assessment of the tumor microenvironment.

BACKGROUND: The intravesical instillation of the paclitaxel-hyaluronan conjugate ONCOFID-P-B™ in patients with bacillus Calmette-Guérin (BCG)-unresponsive bladder carcinoma in situ (CIS; NCT04798703 phase I study), induced 75 and 40% of complete response (CR) after 12 weeks of intensive phase and 12 months of maintenance phase, respectively. The aim of this study was to provide a detailed description of the tumor microenvironment (TME) of ONCOFID-P-B™-treated BCG-unresponsive bladder CIS patients enrolled in the NCT04798703 phase I study, in order to identify predictive biomarkers of response. METHODS: The composition and spatial interactions of tumor-infiltrating immune cells and the expression of the most relevant hyaluronic acid (HA) receptors on cancer cells, were analyzed in biopsies from the 20 patients enrolled in the NCT04798703 phase I study collected before starting ONCOFID-P-B™ therapy (baseline), and after the intensive and the maintenance phases. Clinical data were correlated with cell densities, cell distribution and cell interactions. Associations between immune populations or HA receptors expression and outcome were analyzed using univariate Cox regression and log-rank analysis. RESULTS: In baseline biopsies, patients achieving CR after the intensive phase had a lower density of intra-tumoral CD8+ cytotoxic T lymphocytes (CTL), but also fewer interactions between CTL and macrophages or T-regulatory cells, as compared to non-responders (NR). NR expressed higher levels of the HA receptors CD44v6, ICAM-1 and RHAMM. The intra-tumoral macrophage density was positively correlated with the expression of the pro-metastatic and aggressive variant CD44v6, and the combined score of intra-tumoral macrophage density and CD44v6 expression had an AUC of 0.85 (95% CI 0.68-1.00) for patient response prediction. CONCLUSIONS: The clinical response to ONCOFID-P-B™ in bladder CIS likely relies on several components of the TME, and the combined evaluation of intra-tumoral macrophages density and CD44v6 expression is a potentially new predictive biomarker for patient response. Overall, our data allow to advance a potential rationale for combinatorial treatments targeting the immune infiltrate such as immune checkpoint inhibitors, to make bladder CIS more responsive to ONCOFID-P-B™ treatment.

A multiparameter liquid biopsy approach allows to track melanoma dynamics and identify early treatment resistance.

Melanoma heterogeneity is a hurdle in metastatic disease management. Although the advent of targeted therapy has significantly improved patient outcomes, the occurrence of resistance makes monitoring of the tumor genetic landscape mandatory. Liquid biopsy could represent an important biomarker for the real-time tracing of disease evolution. Thus, we aimed to correlate liquid biopsy dynamics with treatment response and progression by devising a multiplatform approach applied to longitudinal melanoma patient monitoring. We conceived an approach that exploits Next Generation Sequencing (NGS) and droplet digital PCR, as well as the FDA-cleared platform CellSearch, to analyze circulating tumor DNA (ctDNA) trend and circulating melanoma cell (CMC) count, together with their customized genetic and copy number variation analysis. The approach was applied to 17 stage IV melanoma patients treated with BRAF/MEK inhibitors, followed for up to 28 months. BRAF mutations were detected in the plasma of 82% of patients. Single nucleotide variants known or suspected to confer resistance were identified in 70% of patients. Moreover, the amount of ctDNA, both at baseline and during response, correlated with the type and duration of the response itself, and the CMC count was confirmed to be a prognostic biomarker. This work provides proof of principle of the power of this approach and paves the way for a validation study aimed at evaluating early ctDNA-guided treatment decisions in stage IV melanoma. The NGS-based molecular profile complemented the analysis of ctDNA trend and, together with CMC analysis, revealed to be useful in capturing tumor evolution.

Automatic 3D Segmentation and Identification of Anomalous Aortic Origin of the Coronary Arteries Combining Multi-view 2D Convolutional Neural Networks.

This work aimed to automatically segment and classify the coronary arteries with either normal or anomalous origin from the aorta (AAOCA) using convolutional neural networks (CNNs), seeking to enhance and fasten clinician diagnosis. We implemented three single-view 2D Attention U-Nets with 3D view integration and trained them to automatically segment the aortic root and coronary arteries of 124 computed tomography angiographies (CTAs), with normal coronaries or AAOCA. Furthermore, we automatically classified the segmented geometries as normal or AAOCA using a decision tree model. For CTAs in the test set (n = 13), we obtained median Dice score coefficients of 0.95 and 0.84 for the aortic root and the coronary arteries, respectively. Moreover, the classification between normal and AAOCA showed excellent performance with accuracy, precision, and recall all equal to 1 in the test set. We developed a deep learning-based method to automatically segment and classify normal coronary and AAOCA. Our results represent a step towards an automatic screening and risk profiling of patients with AAOCA, based on CTA.

Identification of druggable host dependency factors shared by multiple SARS-CoV-2 variants of concern.

The high mutation rate of SARS-CoV-2 leads to the emergence of multiple variants, some of which are resistant to vaccines and drugs targeting viral elements. Targeting host dependency factors, e.g. cellular proteins required for viral replication, would help prevent resistance. However, it remains unclear whether different SARS-CoV-2 variants induce conserved cellular responses and exploit the same core host factors. To this end, we compared three variants of concern and found that the host transcriptional response was conserved, differing only in kinetics and magnitude. Through CRISPR screening, we identified host genes required for infection by each variant. Most of the genes were shared by multiple variants. We validated our hits with small molecules and repurposed Food and Drug Administration-approved drugs. All the drugs were highly active against all the variants tested, including new variants that emerged during the study (Delta and Omicron). Mechanistically, we identified reactive oxygen species production as a key step in early virus replication. Antioxidants such as N-acetyl cysteine (NAC) were effective against all the variants in both human lung cells and a humanised mouse model. Our study supports the use of available antioxidant drugs, such as NAC, as a general and effective anti-COVID-19 approach.

A scalable route to quaternary ammonium-functionalized AgCl colloidal antimicrobials inhibiting food pathogenic bacteria and biofilms.

This study explores how a simple argentometric titration-like approach could be evolved into a versatile, scalable, fast, and robust strategy for the production of AgCl/quaternary ammonium compounds (QACs) colloidal nanoantimicrobials (NAMs). These systems, which are green, stable, cost-effective, and reproducible are found to be effective against a wide range of food pathogenic bacteria and biofilms. The option of a large-scale production for such colloidal suspensions was explored via the use of a peristaltic pump. The utilization of various types of biosafe QACs and a wide range of solvents including aqueous and organic ones renders this system green and versatile. Nanocolloids (NCs) were characterized using UV-Vis, X-ray photoelectron and Fourier transform infrared (FTIR) spectroscopies. Their morphology and crystalline nature were investigated by transmission electron microscopy (TEM) and selected area diffraction pattern (SAED). Nanoparticle (NP) size distribution and hydrodynamic radius were measured by dynamic light scattering (DLS), while the ζ-potential was found to be highly positive, thus indicating significant colloidal stability and antimicrobial activity. In fact, the higher the NP surface charge, the stronger was their bioactivity. Furthermore, the antibacterial and antibiofilm effects of the as-prepared NCs were tested against Gram-positive bacteria, such as Staphylococcus aureus (ATCC 29213) and Listeria monocytogenes 46, and Gram-negative bacteria, such as Escherichia coli (ATCC 25922) and Pseudomonas aeruginosa (ATCC 27853). The results clearly indicate that AgCl/QACs provide pronounced antibiofilm activity with long-term bacteriostatic effects against foodborne pathogenic bacteria rendering them an ideal choice for active food packaging systems.

Cytokine-induced killer (CIK) cells, successes and challenges: report on the first international conference dedicated to the clinical translation of this unique adoptive cell immunotherapy.

On August 30, 2023, experts from Germany and abroad met to discuss the successes and challenges of cytokine-induced killer cell (CIK) therapy, that recently celebrated its 30th anniversary providing treatment for cancer. This first virtual conference was hosted by CIO Bonn, a certified Comprehensive Cancer Center (CCC) funded by German Cancer Aid (DKH). In addition to keynote speakers involved in CIK cell clinical trials or optimized preclinical models to improve this adoptive cell immunotherapy, more than 100 attendees from around the world also participated in this event. Initiatives to establish the International Society of CIK Cells (ISCC) and a stronger CIK cell network guiding preclinical research and future clinical trials were also announced.

Prolonging the stability of cetuximab (Erbitux®) and panitumumab (Vectibix®): An orthogonal assessment of physicochemical, biological and microbiological properties of original opened glass vials and diluted saline preparations.

The two anti-epidermal growth factor receptor monoclonal antibodies (mAbs) cetuximab and panitumumab are the pillars for the treatment of EGFR-positive, KRAS wild-type metastatic colorectal cancers. However, stability data of these mAbs are generally missing or incomplete. Here, we report for the first time an orthogonal analysis of the stability of cetuximab (Erbitux®) and panitumumab (Vectibix®), either undiluted vial leftovers or saline dilutions in polyolefin/polyamide infusion bags. All samples were stored at 2-8 °C protected from light, according to their summary of product characteristics (SmPCs). Alternatively, opened vials and preparations were maintained at 25 °C for 15 h, and then stored again at 2-8 °C protected from light to mimic a temporary interruption of the cold chain. Vial leftovers proved stable up to 180 days when stored according to their SmPCs, while compounded preparations in infusion bags maintained their physiochemical, biological and microbiological stability up to 30 days. Additionally, no changes were detected up to 30 days for the same samples undergoing a thermal excursion. Our results provide additional rationale to the SmPCs, crucial especially in the case of reassignment and pre-preparation of bags. This information will allow hospitals to achieve significant cost savings, and better organization of the entire therapeutic process.

Benzothiazole-Phthalimide Hybrids as Anti-Breast Cancer and Antimicrobial Agents.

The benzothiazole nucleus is a major heterocyclic scaffold whose therapeutic potential has been thoroughly explored due to its structural simplicity and ease of synthesis. In fact, several benzothiazole derivatives have been synthesized over time, demonstrating numerous pharmacological properties such as anticancer, antimicrobial, anti-inflammatory, and antioxidant activities. Herein, we propose a new series of benzothiazole-phthalimide hybrids obtained by linking the phthalimide moiety to differently substituted benzothiazole nuclei through the N atom. These compounds have been screened for their anticancer properties against two human breast cancer cell lines. Furthermore, we delved into the mechanism of action of the most active hybrid, compound 3h, by assessing its capability to damage the nuclear DNA, trigger the apoptotic process in the high metastatic MDA-MB-231 cells, and prevent cellular migration. Moreover, in view of the documented antimicrobial activities of the two scaffolds involved, we explored the antibacterial and antifungal effects of the studied compounds by means of the broth microdilution method. Among the studied compounds, 3h showed the highest antimicrobial activity, both against gram-positive and gram-negative bacterial strains belonging to the ESKAPE pathogens (Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, and Enterobacter species) and against fungal strains of the Candida species with MICs values ranging from 16 to 32 µg/mL.

Antifungal Biofilm Inhibitory Effects of Combinations of Diclofenac and Essential Oils.

Systemic fungal infections have risen in recent decades and most of them are caused by Candida species, which are becoming increasingly resistant to conventional antifungal drugs. Biofilm production has been considered the most common growth form of Candida cells and is associated with a high level of antifungal resistance. At present, international research reports on the antifungal activity of non-traditional antimicrobial drugs and their potential use against life-threatening resistant fungal infections. Indeed, drug repurposing has led to the consideration of well-known compounds as a last-line therapy. The goal of this work is to evaluate the potential synergistic antifungal biofilm activity of new combinations between diclofenac sodium salt (DSS), a widely used non-steroidal anti-inflammatory drug (NSAID), with the essential oils (EOs) of Mentha piperita, Pelargonium graveolens, and Melaleuca alternifolia, whose antifungal activity has been well documented over the years. The in vitro antifungal activity of DSS and EOs was determined on different Candida strains. Susceptibility testing and the synergism of DSS and EOs versus biofilm cells was performed by using the broth microdilution assay and checkerboard methods. Minimum inhibitory concentrations (sMIC50) of DSS alone ranged from 1.25 to 2.05 mg/mL for all the strains considered. These values significantly decreased when the drug was used in combination with the EOs. The fractional inhibitory concentration index (FICI) was lower than 0.5 for almost all the associations, thus indicating a significant synergism, particularly for the DSS-Pelargonium graveolens combination towards the Candida strains examined. These preliminary results show that the combination of the EOs with DSS improves the antifungal activity on all the tested Candida strains, significantly lowering the concentrations of the components used and thus allowing any toxic effects to be overcome.

"Be sustainable": EOSC-Life recommendations for implementation of FAIR principles in life science data handling.

The main goals and challenges for the life science communities in the Open Science framework are to increase reuse and sustainability of data resources, software tools, and workflows, especially in large-scale data-driven research and computational analyses. Here, we present key findings, procedures, effective measures and recommendations for generating and establishing sustainable life science resources based on the collaborative, cross-disciplinary work done within the EOSC-Life (European Open Science Cloud for Life Sciences) consortium. Bringing together 13 European life science research infrastructures, it has laid the foundation for an open, digital space to support biological and medical research. Using lessons learned from 27 selected projects, we describe the organisational, technical, financial and legal/ethical challenges that represent the main barriers to sustainability in the life sciences. We show how EOSC-Life provides a model for sustainable data management according to FAIR (findability, accessibility, interoperability, and reusability) principles, including solutions for sensitive- and industry-related resources, by means of cross-disciplinary training and best practices sharing. Finally, we illustrate how data harmonisation and collaborative work facilitate interoperability of tools, data, solutions and lead to a better understanding of concepts, semantics and functionalities in the life sciences.

3D patient-specific modeling and structural finite element analysis of atherosclerotic carotid artery based on computed tomography angiography.

The assessment of carotid plaque vulnerability is a relevant clinical information that can help prevent adverse cerebrovascular events. To this aim, in this study, we propose a patient-specific computational workflow to quantify the stress distribution in an atherosclerotic carotid artery, by means of geometric modeling and structural simulation of the plaque and vessel wall. Ten patients were involved in our study. Starting with segmentation of the lumen, calcific and lipid plaque components from computed tomography angiography images, the fibrous component and the vessel wall were semi-automatically reconstructed with an ad-hoc procedure. Finite element analyses were performed using local pressure values derived from ultrasound imaging. Simulation outputs were analyzed to assess how mechanical factors influence the stresses within the atherosclerotic wall. The developed reconstruction method was first evaluated by comparing the results obtained using the automatically generated fibrous component model and the one derived from image segmentation. The high-stress regions in the carotid artery wall around plaques suggest areas of possible rupture. In mostly lipidic and heterogeneous plaques, the highest stresses are localized at the interface between the lipidic components and the lumen, in the fibrous cap.

Transglutaminase Type 2-MITF axis regulates phenotype switching in skin cutaneous melanoma.

Skin cutaneous melanoma (SKCM) is the deadliest form of skin cancer due to its high heterogeneity that drives tumor aggressiveness. Melanoma plasticity consists of two distinct phenotypic states that co-exist in the tumor niche, the proliferative and the invasive, respectively associated with a high and low expression of MITF, the master regulator of melanocyte lineage. However, despite efforts, melanoma research is still far from exhaustively dissecting this phenomenon. Here, we discovered a key function of Transglutaminase Type-2 (TG2) in regulating melanogenesis by modulating MITF transcription factor expression and its transcriptional activity. Importantly, we demonstrated that TG2 expression affects melanoma invasiveness, highlighting its positive value in SKCM. These results suggest that TG2 may have implications in the regulation of the phenotype switching by promoting melanoma differentiation and impairing its metastatic potential. Our findings offer potential perspectives to unravel melanoma vulnerabilities via tuning intra-tumor heterogeneity.

Hematologic Neoplasms Associated with Down Syndrome: Cellular and Molecular Heterogeneity of the Diseases.

The molecular basis of Down syndrome (DS) predisposition to leukemia is not fully understood but involves various factors such as chromosomal abnormalities, oncogenic mutations, epigenetic alterations, and changes in selection dynamics. Myeloid leukemia associated with DS (ML-DS) is preceded by a preleukemic phase called transient abnormal myelopoiesis driven by GATA1 gene mutations and progresses to ML-DS via additional mutations in cohesin genes, CTCF, RAS, or JAK/STAT pathway genes. DS-related ALL (ALL-DS) differs from non-DS ALL in terms of cytogenetic subgroups and genetic driver events, and the aberrant expression of CRLF2, JAK2 mutations, and RAS pathway-activating mutations are frequent in ALL-DS. Recent advancements in single-cell multi-omics technologies have provided unprecedented insights into the cellular and molecular heterogeneity of DS-associated hematologic neoplasms. Single-cell RNA sequencing and digital spatial profiling enable the identification of rare cell subpopulations, characterization of clonal evolution dynamics, and exploration of the tumor microenvironment's role. These approaches may help identify new druggable targets and tailor therapeutic interventions based on distinct molecular profiles, ultimately improving patient outcomes with the potential to guide personalized medicine approaches and the development of targeted therapies.

Hunting down zinc(II)-binding sites in proteins with distance matrices.

MOTIVATION: In recent years, high-throughput sequencing technologies have made available the genome sequences of a huge variety of organisms. However, the functional annotation of the encoded proteins often still relies on low-throughput and costly experimental studies. Bioinformatics approaches offer a promising alternative to accelerate this process. In this work, we focus on the binding of zinc(II) ions, which is needed for 5%-10% of any organism's proteins to achieve their physiologically relevant form. RESULTS: To implement a predictor of zinc(II)-binding sites in the 3D structures of proteins, we used a neural network, followed by a filter of the network output against the local structure of all known sites. The latter was implemented as a function comparing the distance matrices of the Cα and Cß atoms of the sites. We called the resulting tool Master of Metals (MOM). The structural models for the entire proteome of an organism generated by AlphaFold can be used as input to our tool in order to achieve annotation at the whole organism level within a few hours. To demonstrate this, we applied MOM to the yeast proteome, obtaining a precision of about 76%, based on data for homologous proteins. AVAILABILITY AND IMPLEMENTATION: Master of Metals has been implemented in Python and is available at https://github.com/cerm-cirmmp/Master-of-metals.

Acute myeloid leukemia: from NGS, through scRNA-seq, to CAR-T. dissect cancer heterogeneity and tailor the treatment.

Acute myeloid leukemia (AML) is a malignant blood cancer with marked cellular heterogeneity due to altered maturation and differentiation of myeloid blasts, the possible causes of which are transcriptional or epigenetic alterations, impaired apoptosis, and excessive cell proliferation. This neoplasm has a high rate of resistance to anticancer therapies and thus a high risk of relapse and mortality because of both the biological diversity of the patient and intratumoral heterogeneity due to the acquisition of new somatic changes. For more than 40 years, the old gold standard "one size fits all" treatment approach included intensive chemotherapy treatment with anthracyclines and cytarabine.The manuscript first traces the evolution of the understanding of the pathology from the 1970s to the present. The enormous strides made in its categorization prove to be crucial for risk stratification, enabling an increasingly personalized diagnosis and treatment approach.Subsequently, we highlight how, over the past 15 years, technological advances enabling single cell RNA sequencing and T-cell modification based on the genomic tools are affecting the classification and treatment of AML. At the dawn of the new millennium, the advent of high-throughput next-generation sequencing technologies has enabled the profiling of patients evidencing different facets of the same disease, stratifying risk, and identifying new possible therapeutic targets that have subsequently been validated. Currently, the possibility of investigating tumor heterogeneity at the single cell level, profiling the tumor at the time of diagnosis or after treatments exist. This would allow the identification of underrepresented cellular subclones or clones resistant to therapeutic approaches and thus responsible for post-treatment relapse that would otherwise be difficult to detect with bulk investigations on the tumor biopsy. Single-cell investigation will then allow even greater personalization of therapy to the genetic and transcriptional profile of the tumor, saving valuable time and dangerous side effects. The era of personalized medicine will take a huge step forward through the disclosure of each individual piece of the complex puzzle that is cancer pathology, to implement a "tailored" therapeutic approach based also on engineered CAR-T cells.

Lumped-parameter model as a non-invasive tool to assess coronary blood flow in AAOCA patients.

Anomalous aortic origin of the coronary artery (AAOCA) is a rare disease associated with sudden cardiac death, usually related to physical effort in young people. Clinical routine tests fail to assess the ischemic risk, calling for novel diagnostic approaches. To this aim, some recent studies propose to assess the coronary blood flow (CBF) in AAOCA by computational simulations but they are limited by the use of data from literature retrieved from normal subjects. To overcome this limitation and obtain a reliable assessment of CBF, we developed a fully patient-specific lumped parameter model based on clinical imaging and in-vivo data retrieved during invasive coronary functional assessment of subjects with AAOCA. In such a way, we can estimate the CBF replicating the two hemodynamic conditions in-vivo analyzed. The model can mimic the effective coronary behavior with high accuracy and could be a valuable tool to quantify CBF in AAOCA. It represents the first step required to move toward a future clinical application with the aim of improving patient care. The study was registered at Clinicaltrial.gov with (ID: NCT05159791, date 2021-12-16).

How can Cytokine-induced killer cells overcome CAR-T cell limits.

The successful treatment of patients affected by B-cell malignancies with Chimeric Antigen Receptor (CAR)-T cells represented a breakthrough in the field of adoptive cell therapy (ACT). However, CAR-T therapy is not an option for every patient, and several needs remain unmet. In particular, the production of CAR-T cells is expensive, labor-intensive and logistically challenging; additionally, the toxicities deriving from CAR-T cells infusion, such as cytokine release syndrome (CRS) and immune effector cell-associated neurotoxicity syndrome (ICANS), have been documented extensively. Alternative cellular therapy products such as Cytokine-induced killer (CIK) cells have the potential to overcome some of these obstacles. CIK cells are a heterogeneous population of polyclonal CD3+CD56+ T cells with phenotypic and functional properties of NK cells. CIK cell cytotoxicity is exerted in a major histocompatibility complex (MHC)-unrestricted manner through the engagement of natural killer group 2 member D (NKG2D) molecules, against a wide range of hematological and solid tumors without the need for prior antigen exposure or priming. The foremost potential of CIK cells lies in the very limited ability to induce graft-versus-host disease (GvHD) reactions in the allogeneic setting. CIK cells are produced with a simple and extremely efficient expansion protocol, which leads to a massive expansion of effector cells and requires a lower financial commitment compared to CAR-T cells. Indeed, CAR-T manufacturing involves the engineering with expensive GMP-grade viral vectors in centralized manufacturing facilities, whereas CIK cell production is successfully performed in local academic GMP facilities, and CIK cell treatment is now licensed in many countries. Moreover, the toxicities observed for CAR-T cells are not present in CIK cell-treated patients, thus further reducing the costs associated with hospitalization and post-infusion monitoring of patients, and ultimately encouraging the delivery of cell therapies in the outpatient setting. This review aims to give an overview of the limitations of CAR-T cell therapy and outline how the use of CIK cells could overcome such drawbacks thanks to their unique features. We highlight the undeniable advantages of using CIK cells as a therapeutic product, underlying the opportunity for further research on the topic.

Antimicrobial Synergistic Effects of Linezolid and Vancomycin with a Small Synthesized 2-Mercaptobenzothiazole Derivative: A Challenge for MRSA Solving.

Methicillin-resistant Staphylococcus aureus (MRSA) emerged as one of the leading causes of persistent human infections and makes it difficult to treat bacteremia, especially with biofilm formation. In this work, we investigated the in vitro synergism between Linezolid (LNZ) and Vancomycin (VAN) with a 2-mercaptobenzothiazole derivative, resulting in a new small-molecule antibacterial compound that we named BTZ2e, on several clinical MRSA, MRSE (methicillin-resistant Staphylococcus epidermidis) and control (ATCC Collection) strains in their planktonic and biofilms cultures. The broth microdilution method evaluated the susceptibility of planktonic cells to each investigated antibiotic combined with BTZ2e. The biofilm's metabolic activity was studied with the XTT reduction assay. As a result, in this study, biofilm formation was significantly suppressed by the BTZ2e treatment. In terms of minimal biofilm inhibitory concentration (MBIC), BTZ2e revealed an MBIC50 value of 32 µg/mL against methicillin-susceptible S. aureus (MSSA) and 16 µg/mL against methicillin-resistant S. aureus ATCC 43300 biofilms. An inhibition range of 32 µg/mL and 256 µg/mL was registered for the clinical isolates. Interestingly, a synergistic effect (FICI ≤ 0.5) was encountered for the combination of BTZ2e with LNZ and VAN on several planktonic and sessile strains. In particular, the best result against planktonic cells emerged as a result of the synergistic association between LNZ and BTZ2e, while against sessile cells, the best synergistic association resulted from VAN and BTZ2e. The consistent results indicate BTZ2e as a promising adjuvant against multi-resistant strains such as MRSA and MRSE.

Antiviral mechanisms of two broad-spectrum monoclonal antibodies for rabies prophylaxis and therapy.

Rabies is an acute and lethal encephalomyelitis caused by lyssaviruses, among which rabies virus (RABV) is the most prevalent and important for public health. Although preventable through the post-exposure administration of rabies vaccine and immunoglobulins (RIGs), the disease is almost invariably fatal since the onset of clinical signs. Two human neutralizing monoclonal antibodies (mAbs), RVC20 and RVC58, have been shown to be effective in treating symptomatic rabies. To better understand how these mAbs work, we conducted structural modeling and in vitro assays to analyze their mechanisms of action, including their ability to mediate Fc-dependent effector functions. Our results indicate that both RVC20 and RVC58 recognize and lock the RABV-G protein in its pre-fusion conformation. RVC58 was shown to neutralize more potently the extra-cellular virus, while RVC20 mainly acts by reducing viral spreading from infected cells. Importantly, RVC20 was more effective in promoting effector functions compared to RVC58 and 17C7-RAB1 mAbs, the latter of which is approved for human rabies post-exposure treatment. These results provide valuable insights into the multiple mechanisms of action of RVC20 and RVC58 mAbs, offering relevant information for the development of these mAbs as treatment for human rabies.

Molecular Modeling Unveils the Effective Interaction of B-RAF Inhibitors with Rare B-RAF Insertion Variants.

The Food and Drug Administration (FDA) has approved MAPK inhibitors as a treatment for melanoma patients carrying a mutation in codon V600 of the BRAF gene exclusively. However, BRAF mutations outside the V600 codon may occur in a small percentage of melanomas. Although these rare variants may cause B-RAF activation, their predictive response to B-RAF inhibitor treatments is still poorly understood. We exploited an integrated approach for mutation detection, tumor evolution tracking, and assessment of response to treatment in a metastatic melanoma patient carrying the rare p.T599dup B-RAF mutation. He was addressed to Dabrafenib/Trametinib targeted therapy, showing an initial dramatic response. In parallel, in-silico ligand-based homology modeling was set up and performed on this and an additional B-RAF rare variant (p.A598_T599insV) to unveil and justify the success of the B-RAF inhibitory activity of Dabrafenib, showing that it could adeptly bind both these variants in a similar manner to how it binds and inhibits the V600E mutant. These findings open up the possibility of broadening the spectrum of BRAF inhibitor-sensitive mutations beyond mutations at codon V600, suggesting that B-RAF V600 WT melanomas should undergo more specific investigations before ruling out the possibility of targeted therapy.