Biology subjects

Musico, A.

Publications and source records attributed to Musico, A..

4 recordsLinked to original sources

bioRxiv2026

Enhanced Workflow for Urinary Extracellular Vesicle Isolation Using Membrane-Sensing Peptides

Urinary extracellular vesicles (uEVs) represent a promising source of non-invasive biomarkers; however, their clinical translation is still limited by suboptimal isolation strategies, which often suffer from poor scalability, co-isolation of abundant urinary proteins, and bias toward specific EV subpopulations. Here, we employ a membrane-sensing peptide (MSP)-based affinity approach for uEVs isolation, that exploits the highly lipid membranes curvature of EV as universal target, enabling pan-specific capture independent of surface marker expression. MSP-functionalized beads were applied to minimally processed urine samples and benchmarked against differential ultracentrifugation (dUC) and size-exclusion chromatography (SEC). Comprehensive characterization by nanoparticle tracking analysis, transmission electron microscopy, high-sensitivity flow cytometry, single-molecule array (SiMoA), and fluorescence nanoparticle tracking analysis, demonstrated that MSP-based isolation preserves vesicle integrity and maintains the native distribution of canonical tetraspanins (CD9, CD63, CD81), without evidence of subpopulation bias. Notably, MSP-based isolation significantly reduced co-isolated contaminants, such as uromodulin, resulting in improved sample purity. By combining high recovery, improved purity, and operational simplicity, MSP workflow offers practical advantages, including reduced processing time, scalability, and compatibility with standard laboratory equipment, without the need for extensive pre-processing. These properties characterize MSP-based affinity capture as a robust and versatile alternative to conventional uEVs isolation approaches, with strong potential for translational and clinical applications.

biochemistry↗

bioRxiv2025

Red Blood Cell-derived Extracellular Vesicles enable Cisplatin and Cetuximab Synergistic Therapy against Triple-Negative Breast Cancer

BackgroundTriple-negative breast cancer is an aggressive breast cancer subtype characterized by the absence of human epidermal growth factor receptor 2, estrogen and progesterone receptors, limiting targeted therapy options. Cisplatin, a chemotherapeutic agent, induces DNA damage and exhibits some efficacy against triple-negative breast cancer, but its effectiveness is often reduced by chemoresistance and systemic toxicity. A very promising strategy to augment cisplatin treatment can be based on combining it with the biologic Cetuximab, an epidermal growth factor receptor inhibitor, which boosts cisplatin efficacy by inducing ferroptosis. ResultsTo optimize this strategy in a biocompatible and precise manner, we developed a nanoplatform based on red blood cell-derived extracellular vesicles for the combined delivery of Cetuximab and cisplatin, enabling immune evasion, and the possibility of autologous personalization and GMP-compliant production. Owing to their DNA-free lumen and lack of EGFR, RBC-EVs preserve cisplatin activity and prevent interference with cetuximab. This formulation enhances cisplatins cytotoxicity by up to 50%, as shown in vitro and in patient-derived organoids. It effectively reduces chemoresistance by downregulating hypoxia-related genes and promoting ferroptosis, additionally, it improves cisplatins cytotoxic effects while reducing hemotoxicity compared to the administration of free cisplatin. ConclusionsThese findings highlight the potential of red blood cell-derived extracellular vesicles as a biocompatible delivery system enabling combined therapy and offering a promising strategy to overcome current limitations in TNBC treatment.

cell biology↗

bioRxiv2022

Compositional profiling of extracellular vesicles/lipoproteins mixtures by AFM nanomechanical imaging

The widely overlapping physicochemical properties of lipoproteins (LPs) and extracellular vesicles (EVs) represents one of the main obstacles for the isolation and characterization of these pervasive biogenic lipid nanoparticles. We herein present the application of an atomic force microscopy (AFM)-based quantitative morphometry assay to the rapid nanomechanical screening of mixed LPs and EVs samples. The method can determine the diameter and the mechanical stiffness of hundreds of individual nanometric objects within few hours. The obtained diameters are in quantitative accord with those measured via cryo-electron microscopy (cryo-EM); the assignment of a specific nanomechanical readout to each object enables the simultaneous discrimination of co-isolated EVs and LPs even if they have overlapping size distributions. EVs and all classes of LPs are shown to be characterized by specific combinations of diameter and stiffness, thus making it possible to estimate their relative abundance in EV/LP mixed samples in terms of stoichiometric ratio, surface area and volume. As a side finding, we show how the mechanical behaviour of specific LP classes is correlated to distinctive structural features revealed by cryo-EM. To the best of our knowledge, these results represent the first systematic single-particle mechanical investigation of lipoproteins. The described approach is label-free, single-step and relatively quick to perform. Importantly, it can be used to analyze samples which prove very challenging to assess with several established techniques due to ensemble-averaging, low sensibility to small particles, or both, thus providing a very useful tool for quickly assessing the purity of EV/LP isolates including plasma- and serum-derived preparations.

biophysics↗

bioRxiv2020

SARS-CoV-2 epitope mapping on microarrays highlights strong immune-response to N protein region

A workflow for SARS-CoV-2 epitope discovery on peptide microarrays is herein reported. The process started with a proteome-wide screening of immunoreactivity based on the use of a high-density microarray followed by a refinement and validation phase on a restricted panel of probes using microarrays with tailored peptide immobilization through a click-based strategy. Progressively larger, independent cohorts of Covid-19 positive sera were tested in the refinement processes, leading to the identification of immunodominant regions on SARS-CoV-2 Spike (S), Nucleocapsid (N) protein and Orf1ab polyprotein. A summary study testing 50 serum samples highlighted an epitope of the N protein (region 155-171) providing 92% sensitivity and 100% specificity of IgG detection in Covid-19 samples thus being a promising candidate for rapid implementation in serological tests.

biochemistry↗