Biology subjects

Battaglia, G.

Publications and source records attributed to Battaglia, G..

At least 19 recordsLinked to original sources

bioRxiv2025

A computational analysis of the glycoprotein LRP1 structure and the role of glycans as quaternary glue

The low-density lipoprotein receptor-related protein 1 (LRP1) plays a critical role in development and transport across the blood-brain barrier (BBB), yet until now, its molecular architecture remained unresolved due to the absence of an experimentally determined structure. Using homology modeling and neural network-based structure prediction algorithms, complemented with molecular dynamics (MD) simulations, we propose a comprehensive model of LRP1 structures. We observe a natural dimerization mechanism and provide insight into the dynamic behavior of its flexible domains under physiological conditions. We investigated the stability of non-covalent interactions keeping LRP1s and {beta} chains linked together, and found the energy required to break the link is 180{+/-}2 kT. The MD characterization highlights the fundamental role of glycans in the creation of LRP1s quaternary structure, increasing the number of intra-dimeric contacts. This study opens new avenues for targeted drug design strategies, enhancing our molecular understanding of LRP1s receptor-mediated transport in the brain and the key mediation of glycosylation in protein-protein interactions.

biophysics↗

bioRxiv2025

A microRNA-based Therapy against Human Medulloblastoma Validated in a Xenotransplant Model using Wild-Type Mouse Embryos

Medulloblastoma (MB) is the most prevalent malignant pediatric brain tumor. Its poor prognosis is driven by therapy-resistant cancer stem-like cells (CSCs), which are challenging to study in traditional preclinical models. MicroRNAs (miRNAs) modulate tumor-related pathways by repressing oncogenic gene expression, a function lost in cancer. We assessed the therapeutic potential of miRNA restoration in human MB from distinct subgroups, validated in a xenotransplant model using wild-type (WT) mouse embryos. Multi-omics, imaging, and in silico analyses were employed to identify underlying mechanisms and therapeutic targets. Human MB cells formed tumors in embryonic mouse brains that mirrored key MB features, including vascularized CSC niches and metastases. Restoration of underexpressed miRNAs reduced tumor growth and invasiveness in vitro and in vivo. These miRNAs synergistically repressed gene networks related to cell adhesion and RNA metabolism, shared oncogenic pathways across MB subgroups. Our embryonic xenotransplant model provides a clinically relevant platform for evaluating next-generation gene therapies in pediatric brain cancer.

cancer biology↗

bioRxiv2025

Transferrin Receptor 1-targeted polymersomes therapy for Colorectal Cancer

Colorectal cancer (CRC) is a major global health concern, ranking among the most common cancers and the second leading cause of cancer-related deaths. The high mortality associated with CRC is attributed mainly to difficulties in early detection and the lack of effective targeted therapies. The Transferrin receptor 1 (TfR1) is particularly attractive as a therapy target given its notable overexpression in tumor cells, particularly in CRC. This study explored the potential of a polymeric nanoparticle (POs)-based drug delivery system targeting TfR1 to improve the precision and efficacy of CRC treatment. For this study, we used three human colorectal cancer cell lines (SW480, HT-29, and HCT116), a healthy human intestinal epithelial cell line (hIECs), and a murine CRC cell line (MC38). We first engineered POs composed of poly (ethylene glycol) (PEG) and poly (lactic acid) (PLA), functionalized with the T7 peptide to enhance their specificity for TfR1-expressing cells. Targeting efficiency of these POs was assessed across all cell lines by evaluating the cellular uptake using flow cytometry. Upon establishing the optimal formulation for these NPs for TfR1-targeting, we encapsulated doxorubicin (DOX) to evaluate their therapeutic potential. Both in vitro and in vivo studies were performed to assess the efficacy of these DOX-loaded TfR1-targeted POs. In vitro studies demonstrated selective delivery of DOX to CRC cells, suggesting a marked reduction in off-target effects. In vivo studies in a murine model of CRC further supported these findings, showing that DOX-loaded TfR1-targeted POs significantly improved survival rates and reduced tumor growth compared to free DOX or PBS treatments. These results highlight the promise of TfR1-targeted POs as a precise strategy for CRC therapy, offering enhanced treatment efficacy while reducing systemic toxicity. This novel approach could lead to the development of more targeted and less harmful cancer treatments. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=76 SRC="FIGDIR/small/639871v1_ufig1.gif" ALT="Figure 1"> View larger version (26K): org.highwire.dtl.DTLVardef@1714603org.highwire.dtl.DTLVardef@1ab7f15org.highwire.dtl.DTLVardef@245f6eorg.highwire.dtl.DTLVardef@1f56f75_HPS_FORMAT_FIGEXP M_FIG C_FIG HighlightsO_LIT-7 peptide-functionalized polymersomes exhibit high binding affinity to CRC cell lines C_LIO_LIDoxorubicin-loaded T7-peptide polymersomes effectively suppress tumor growth and prolong survival in vivo C_LIO_LITargeted Doxorubicin delivery via T7-peptide polymersomes minimizes toxicity to healthy tissues C_LI

cancer biology↗

bioRxiv2024

A new class of natural anthelmintics targeting lipid metabolism

Parasitic helminths are a major global health threat, infecting nearly one-fifth of the human population and causing significant losses in livestock and crops. Resistance to the few anthelmintic drugs is increasing. Here, we report a set of avocado fatty alcohols/acetates (AFAs) that exhibit nematocidal activity against four veterinary parasitic nematode species: Brugia pahangi, Teladorsagia circumcincta and Heligmosomoides polygyrus, as well as a multidrug resistant strain (UGA) of Haemonchus contortus. AFA shows significant efficacy in H. polygyrus infected mice. In C. elegans, AFA exposure affects all developmental stages, causing paralysis, impaired mitochondrial respiration, increased reactive oxygen species production and mitochondrial damage. In embryos, AFAs penetrate the eggshell and induce rapid developmental arrest. Genetic and biochemical tests reveal that AFAs inhibit POD-2, encoding an acetyl CoA carboxylase, the rate-limiting enzyme in lipid biosynthesis. These results uncover a new anthelmintic class affecting lipid metabolism.

cell biology↗

bioRxiv2024

Multivalent Targeting of Blood-Brain Barrier LRP1 for Neurovascular Recovery Therapy for Alzheimer s Disease

The blood-brain barrier (BBB) is a highly selective permeability barrier that safeguards the central nervous system (CNS) from potentially harmful substances while regulating the transport of essential molecules. Its dysfunction is increasingly recognized as a pivotal factor in the pathogenesis of Alzheimers disease (AD), contributing to the accumulation of amyloid-{beta} (A{beta}) plaques. We propose an AD therapeutic strategy targeting the BBB low-density lipoprotein receptor-related protein 1 (LRP1). We show that a multivalent scaffold with LRP1-specific peptide modulates the A{beta} transport at the BBB. We show via detailed experimentation on AD model mice that this intervention markedly reduces A{beta} deposits and prevents cognitive decline. This study marks a new approach to drug design, combining multivalent targeting with the regulation of membrane trafficking through advanced molecular engineering. Crucially, the therapeutic effects emerge from the multivalent nature of our proposed system. Furthermore, our findings underscore the paramount significance of the BBB in AD pathogenesis, particularly emphasizing the critical role of LRP1-mediated A{beta} clearance in mitigating disease progression.

neuroscience↗

bioRxiv2024

Imaging Abeta aggregation by liquid-phase transmission electron microscopy

The amyloid beta peptide (A{beta}) readily aggregates into amyloid fibrils. This process has been the subject of intense investigations since it is associated with Alzheimers disease. However, it has been highly challenging to observe the microscopic steps in the aggregation reaction directly and to characterize the oligomeric assemblies formed as intermediates. To achieve this goal, we apply liquid-phase transmission electron microscopy (LTEM) in combination with all-atom molecular dynamics simulations. Our results offer an initial visualization of the dynamics of A{beta} oligomers, the formation of A{beta} protofibrils, and the presence of A{beta} oligomers on the surface of A{beta} fibrils. This work illustrates how the application of LTEM to the study of protein aggregation in solution enables the imaging of key molecular events in the aggregation process of A{beta}.

biophysics↗

bioRxiv2024

A 'double-edged' role for type-5 metabotropic glutamate receptors in pain disclosed by light-sensitive drugs

Knowing the site of drug action is important to optimize effectiveness and address any side effects. We used light-sensitive drugs to identify the brain region-specific role of mGlu5 metabotropic glutamate receptors in the control of pain. Optical activation of systemic JF-NP-26, a caged, normally inactive, negative allosteric modulator (NAM) of mGlu5 receptors, in cingulate, prelimbic and infralimbic cortices and thalamus inhibited neuropathic pain hypersensitivity. Systemic treatment of alloswitch-1, an intrinsically active mGlu5 receptor NAM, caused analgesia, and the effect was reversed by light-induced drug inactivation in in the prelimbic and infralimbic cortices, and thalamus. This demonstrates that mGlu5 receptor blockade in the medial prefrontal cortex and thalamus is both sufficient and necessary for the analgesic activity of mGlu5 receptor antagonists. Surprisingly, when light was delivered in the basolateral amygdala, local activation of systemic JF-NP-26 reduced pain thresholds, whereas inactivation of alloswitch-1 enhanced analgesia. Electrophysiological analysis showed that alloswitch-1 increased excitatory synaptic responses in prelimbic pyramidal neurons evoked by stimulation of BLA input, and decreased feedforward inhibition of amygdala output neurons by BLA. Both effects were reversed by optical silencing and reinstated by optical reactivation of alloswitch-1. These findings demonstrate for the first time that the action of mGlu5 receptors in the pain neuraxis is not homogenous, and suggest that blockade of mGlu5 receptors in the BLA may limit the overall analgesic activity of mGlu5 receptor antagonists. This could explain the suboptimal effect of mGlu5 NAMs on pain in human studies and validate photopharmacology as an important tool to determine ideal target sites for systemic drugs.

neuroscience↗

bioRxiv2023

SimpliPyTEM: An open-source Python library and app to simplify Transmission Electron Microscopy and in situ-TEM image analysis.

Introducing SimpliPyTEM, a Python library and accompanying GUI that simplifies the post-acquisition evaluation of transmission electron microscopy (TEM) images, helping streamline the workflow. After an imaging session, a folder of image and/or video files, typically containing low contrast and large file size 32-bit images, can be quickly processed via SimpliPyTEM into high-quality, high-contrast .jpg images with suitably sized scale-bars. The app can also generate HTML or PDF files containing the processed images for easy viewing and sharing. Additionally, SimpliPyTEM has a specific focus on in situ TEM videos, an emerging field of EM, allowing for fast data processing into preview movies, averages, image series, or motion corrected averages using MotionCor2. The accompanying Python library offers many standard image processing methods, all simplified to a single command, plus a module to analyse particle morphology and population. This latter application is particularly useful for life sciences investigations. User-friendly tutorials and clear documentation are included to help guide users through the processing and analysis. We invite the EM community to contribute to and further develop this open-source package.

biophysics↗

bioRxiv2023

Kinetic stabilization of translation-repression condensates by a neuron-specific microexon

The inclusion of microexons by alternative splicing is frequent in neuronal proteins. The roles of these sequences are in most cases unknown, but changes in their degree of inclusion are associated with neurodevelopmental diseases. We recently found that the decreased inclusion of a 24-nucleotide neuron-specific microexon in CPEB4, an RNA-binding protein that regulates translation through cytoplasmic changes in poly(A) tail length, is linked to idiopathic autism spectrum disorder (ASD). Why this microexon is required and how small changes in its degree of inclusion generate a dominant-negative effect on the expression of ASD-linked genes is not clear. Here we show that neuronal CPEB4 forms condensates that dissolve upon depolarization, a transition associated with a switch from translational repression to activation. Heterotypic intermolecular interactions between the microexon and a cluster of histidine residues kinetically stabilize the condensates by competing with homotypic interactions between clusters, that otherwise lead to the irreversible aggregation of CPEB4. We conclude that the microexon is required in neuronal CPEB4 to preserve the reversible regulation of CPEB4-mediated gene expression in response to neuronal stimulation.

biophysics↗

bioRxiv2022

The Role of Glycocalyx Steric Effects on Viral Endocytosis

Understanding the mechanisms of viral entry is key to aiding the development of effective anti-viral treatments. In this work, we produce a simple thermodynamic model of viral entry, which is able to predict the differences between viruses. We also show that the glycocalyx, which is often neglected in studies of cell-entry, plays a key role, and the density and structure of the glycocalyx can determine whether or not a virus can enter the cell. We further find that co-receptors play not only a signalling role, but an important thermodynamic role in viral entry. We then show that this model can be used to calculate the cell-entry probabilities of a given virus, which can shine a light on the clinical observations associated with a virus.

biophysics↗

bioRxiv2022

A Multiscale study of phosphorylcholine driven cellular phenotypic targeting

Phenotypic targeting requires the ability of the drug delivery system to discriminate over cell populations expressing a particular receptor combination. Such selectivity control can be achieved using multiplexed-multivalent carriers often decorated with multiple ligands. Here, we demonstrate that the promiscuity of a single ligand can be leveraged to create multiplexed-multivalent carriers achieving phenotypic targeting. We show how the cellular uptake of poly(2-methacryloyloxyethyl phosphorylcholine)-poly(2- (diisopropylamino)ethyl methacrylate) (PMPC-PDPA) polymersomes varies depending on the receptor expression among different cells. We investigate the PMPC-PDPA polymersome insertion at the single chain/receptor level using all-atom molecular modelling. We propose a theoretical statistical mechanics-based model for polymersome-cell association that explicitly considers the interaction of the polymersome with the cell glycocalyx shedding light on its effect on the polymersome binding. We validate our model experimentally and show that the binding energy is a non-linear function, allowing us to tune interaction by varying the radius and degrees of polymerisation. Finally, we show that PMPC-PDPA polymersomes can be used to target monocytes in vivo due to their promiscuous interaction with SRB1, CD36 and CD81.

biochemistry↗

bioRxiv2021

Targeting activated macrophages intracellular milieu to augment anti-inflammatory drug potency

We present pH-responsive phosphorylcholine polymersomes ability to target activated macrophages via scavenger receptors, enter them via endocytosis, and escape from early endosomes enabling the intra-cellular drug delivery. Using an arthritis experimental model and the gold standard disease modifying anti-rheumatic drug, methotrexate, we prove that polymersomes augments therapeutic efficacy, while minimizing the off-target effect. First, we demonstrate the selective accumulation of polymersomes within the inflamed synovial tissues and cells, including macrophages. Second, we show the beneficial therapeutic effect of methotrexate loaded polymersomes in preventing both joint inflammation and further damage. Hence, we prove the therapeutic potential of polymersomes in enhancing the complete prevention of arthritis progression, which makes it a promising nanotherapy for arthritis treatment as well as other inflammatory disorders. TeaserWe show that the effective targeting and delivery of drugs to the main inflammation actors, the macrophages, enhances arthritis therapy.

immunology↗

bioRxiv2021

The role of host cell glycans on virus infectivity: The SARS-CoV-2 case

Long and complex chains of sugars, called glycans, often coat both the cell and protein surface. Glycans both modulate specific interactions and protect cells. On the cell surface, these sugars form a cushion known as the glycocalyx. Here, we show that Heparan Sulfate (HS) chains - part of the glycocalyx - and other glycans - expressed on the surface of both host and virus proteins - have a critical role in modulating both attractive and repulsive potentials during viral infection. We analyse the SARS-CoV-2 virus, modelling its spike proteins binding to HS chains and two key entry receptors, ACE2 and TMPRSS2. We include the volume exclusion effect imposed on the HS chains impose during virus insertion into glycocalyx and the steric repulsion caused by changes in the conformation of the ACE2 glycans involved in binding to the spike. We then combine all these interactions, showing that the interplay of all these components is critical to the behaviour of the virus. We show that the virus tropism depends on the combinatorial expression of both HS chains and receptors. Finally, we demonstrate that when both HS chains and entry receptors express at high density, steric effects dominate the interaction, preventing infection.

biophysics↗

bioRxiv2021

Imaging protein conformational space in liquid water

Withdrawal StatementThe authors have withdrawn their manuscript owing to a mistake in one of the proteins used for the data used to generate Fig.1, where we believed we used the apoferritin but instead used the ferritin proteins. Therefore, the authors do not wish this work to be cited as a reference for the project. If you have any questions, please get in touch with the corresponding author.

biophysics↗

bioRxiv2021

4D imaging of soft matter in liquid water

Water is a critical component for both function and structure of soft matter and it is what bestows the adjective soft. Imaging samples in liquid state is thus paramount to gathering structural and dynamical information of any soft materials. Herein we propose the use of liquid phase electron microscopy to expand ultrastructural analysis into dynamical investigations. We imaged two soft matter examples: a polymer micelle and a protein in liquid phase using transmission electron microscopy and demonstrate that the inherent Brownian motion associated with the liquid state can be exploited to gather three-dimensional information of the materials in their natural state. We call such an approach brownian tomography (BT). We combine BT with single particle analysis (Brownian particle analysis BPA) to image protein structures with a spatial resolution close that achievable using cryogenic TEM. We show that BPA allows sub-nanometer resolution of soft materials and enables to gather information on conformational changes, hydration dynamics, and the effect of thermal fluctuations. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=109 SRC="FIGDIR/small/427613v1_ufig1.gif" ALT="Figure 1"> View larger version (58K): org.highwire.dtl.DTLVardef@1a54a32org.highwire.dtl.DTLVardef@aa49acorg.highwire.dtl.DTLVardef@1caeabborg.highwire.dtl.DTLVardef@f8b380_HPS_FORMAT_FIGEXP M_FIG C_FIG

biophysics↗

bioRxiv2020

Combinatorial Intracellular Delivery Screening (CIDS) of Anticancer Drugs

Conventional drug solubilisation strategies limit the understanding of the full potential of poorly water-soluble drugs during drug screening. Here, we propose a screening approach in which poorly water-soluble drugs are entrapped in poly (2-(methacryloyloxyethyl phosphorylcholine)-poly(2-(diisopropylaminoethyl methacryate) (PMPC-PDPA) polymersomes (POs) to enhance drug solubility and facilitate intracellular delivery. By using a human paediatric glioma cell model, we demonstrated that PMPC-PDPA POs mediated intracellular delivery of cytotoxic and epigenetic drugs by receptor-mediated endocytosis. Additionally, when delivered in combination, drug-loaded PMPC-PDPA POs triggered both an enhanced drug efficacy and synergy compared to that of a conventional combinatorial screening. Hence, our comprehensive synergy analysis illustrates that our screening methodology, in which PMPC-PDPA POs are used for intracellular co-delivery of drugs, allows us to identify potent synergistic profiles of anticancer drugs.

bioengineering↗

bioRxiv2020

Syndapin-2 mediates amyloid-β transcytosis at the brain endothelium: implications in Alzheimer's disease

A deficient transport of amyloid-(A{beta}) across the blood-brain barrier (BBB), and its diminished clearance from the brain, contributes to neurodegenerative and vascular pathologies, including Alzheimers (AD) and cerebral angiopathy, respectively. At the BBB, A{beta} efflux transport is associated with the low-density receptor-related protein 1 (LRP1). However, the precise mechanisms governing A{beta} transport across the BBB, in health and disease, remain to be fully understood. Recent evidence indicates that the LRP1 transcytosis occurs through a tubulation-mediated mechanism stabilised by syndapin-2. Here, we show that syndapin-2 is associated with A{beta} clearance via LRP1 across the BBB. We further demonstrate that risk factors for AD, A{beta} expression and ageing, are associated with a decline in the native expression of syndapin-2 within brain endothelium. Our data reveal that the syndapin-2-mediated pathway, and its balance with the endosomal sorting, are important for A{beta} clearance proposing a measure to evaluate AD and ageing, as well as a target for counteracting A{beta} build-up. Moreover, we provide evidence for the impact of the avidity of A{beta} assemblies in their trafficking across the brain endothelium and in LRP1 expression levels, which may affect the overall clearance of A{beta} across the BBB.

neuroscience↗

bioRxiv2020

On the shuttling across the blood-brain barrier via tubules formation: mechanism and cargo avidity bias

The blood-brain barrier is made of polarised brain endothelial cells (BECs) phenotypically conditioned by the central nervous system (CNS). Transport across BECs is of paramount importance for nutrient uptake as well as to rid the brain of waste products. Nevertheless, currently we do not understand how large macromolecular cargo shuttles across and how BECs discriminate between the brain-bound and own nutrients. Here, we study the low-density lipoprotein receptor-related protein 1 (LRP1) an essential regulator of BEC transport, and show that it is associated with endocytic effectors, endo-lysosomal compartments as well as syndapin-2, a member of the Bin/Amphiphysin/Rvs (BAR) domain superfamily known to stabilise tubular carriers. We employed synthetic self-assembled vesicles, polymersomes, as a multivalent system with tunable avidity as a tool to investigate the mechanism of transport across BECs. We used a combination of conventional and super-resolution microscopy, both in vivo and in vitro, accompanied with biophysical modelling of transport kinetics and membrane-bound interactions. Our results demonstrate that the avidity of the ligand-receptor interaction (the overall cargo binding energy) determines the mechanism of sorting during the early stages of endocytosis and consequent trafficking. We show that high avidity cargo biases the LRP1 towards internalisation and fast degradation in BECs, while mid avidity augments the formation of syndapin-2 stabilised tubular carriers and promotes fast shuttling across BECs. Thus, we map out a very detailed mechanism where clathrin, actin, syndapin-2, dynamin and SNARE act synergistically to enable fast shuttling across BECs.

neuroscience↗