Biology subjects

Moreno, S.

Publications and source records attributed to Moreno, S..

At least 19 recordsLinked to original sources

bioRxiv2026

Structural basis of eIF4E recognition of synthetic mRNA 5' cap analogues links cap architecture to affinity and biological activity

Eukaryotic mRNAs carry a 5'-terminal 7-methylguanosine (m7G) cap that is recognized by multiple cellular proteins and is essential for mRNA self-discrimination, stability, and translation. The eukaryotic translation initiation factor 4E (eIF4E) binds the 5' cap and promotes recruitment of mRNAs to the ribosomal translation initiation complex, a critical step in cap-dependent translation. Beyond naturally occurring cap modifications that modulate mRNA interactions and fate, non-natural 5' cap modifications can be used to finely tune diverse aspects of mRNA biology. This has gained particular relevance with the emergence of in vitro transcribed mRNAs as a powerful therapeutic modality for protein replacement therapies and vaccines. Here, we report binding data for a series of synthetic 5' cap analogues interacting with eIF4E, together with X-ray crystal structures of eIF4E-cap complexes, providing detailed insights into the structural determinants of cap recognition by eIF4E. Furthermore, analysis of gene expression from mRNAs carrying different synthetic cap analogues in human cell lines and mice revealed a relationship between cap architecture and protein expression. Taken together, our structural, biochemical, and functional data establish structure-activity relationships for synthetic cap analogues and identify structural features associated with enhanced translational performance.

biochemistry↗

bioRxiv2026

Novelty-seeking and directed exploration scale with dissociable social network outcomes and brain systems in older adults

Social isolation and loneliness are major determinants of well-being in older adults. Shrinking social networks and age-related shifts in social behavior and decision-making may increase vulnerability to loneliness. These changes in social decision-making strategies can be conceptualized within an exploration-exploitation tradeoff framework, in which social options with known outcomes are prioritized over uncertain ones. Yet, no study has characterized how complementary decision-making strategies, undirected novelty seeking and goal-directed exploration, relate to social network constructs or are supported by distinct neural systems. These strategies capture distinct forms of decision-making: undirected novelty seeking is driven by the intrinsic salience of novel options independent of expected informational gain, whereas goal-directed exploration involves deliberately sampling uncertain options to reduce uncertainty and optimize future choices. Here, we used fMRI and a novel social bandit reinforcement task to identify dissociable social constructs and brain systems linked to undirected novelty seeking and goal-directed exploration in older adults. Undirected novelty seeking was associated with social network size and with activity of temporoparietal and mid-cingulate brain areas. Goal-directed exploration predicted subjective feelings of social connectedness and was associated with activity of anterior cingulate and prefrontal brain areas. Crucially, choice probabilities from a non-social version of the three-arm bandit task were not associated with social decision-making nor with social constructs. Our findings suggest that, in later life, dissociable social decision-making strategies, when faced with unknown options, rely on distinct neural systems supporting social network size as well as subjective feelings of social connectedness.

neuroscience↗

bioRxiv2026

Genome-Scale Codon Deoptimization Enables Attenuation of Rift Valley Fever Virus

Rift Valley Fever Virus (RVFV) is a mosquito-borne zoonotic pathogen responsible for severe disease in domestic and wild ungulates as well as humans, representing a major threat to livestock production and human public health. RVFV is endemic in many African countries and has the potential to spread to new geographical regions. Current vaccines have limitations in safety and efficacy, highlighting the need for strategies to develop new vaccines candidates. In this study, we explored the use of codon deoptimization (CD) as a novel attenuation approach for the development of live-attenuated vaccine (LAV) against RVFV. CD exploits the redundancy of the genetic code by replacing frequently used codons with synonymous, less-preferred codons, thereby reducing translational efficiency without altering the amino acid sequence. We recoded parts of the M and S genome segments of RVFV using the least frequently used codons in mammalian cells, ensuring complete preservation of protein functionality and immunogenicity. Using reverse genetics, we rescued a panel of recombinant (r)RVFV encoding codon-deoptimized S-segment NSs gene (rNScd), M-segment Gn/Gc genes (rMcd), or both (rMcd/NScd). These recombinant CD viruses were characterized in vitro in mammalian and insect cell lines and in vivo using wild-type and immunocompromised mice. Results demonstrated varying degrees of attenuation among the three CD rRVFV, with the one deoptimized in both viral segments, rMcd/NScd, as a promising LAV based on the safety profiles. This study provides proof of concept for the use of CD as a rational strategy to generate attenuated RVFV, for the development of next-generation vaccines against this zoonotic threat.

microbiology↗

bioRxiv2026

An approach for single-amino-acid resolution epitope mapping by kinetic affinity screening of antibody drugs against biosensor on-chip library of deep mutationally-scanned target variants

Epitope mapping is central to rational antibody drug design, affinity optimization and the anticipation of therapeutic resistance mechanisms. Here, we demonstrate the use of Sensor Integrated Proteome on Chip (SPOC) technology for single amino acid resolution epitope mapping. By generating high throughput (HTP) binding kinetics data, we identify important residues within the target epitope whose mutations alter drug-target interactions. The SPOC platform integrates simultaneous HTP cell-free production of folded proteins in nanowells from immobilized plasmid DNAs or linear expression cassettes and capture onto biosensor chips for subsequent label-free binding kinetic analysis using surface plasmon resonance (SPR). The model system comprised the extracellular domain (ECD) of CD20, a membrane-spanning 4-domain family protein, screened against its FDA-approved therapeutic monoclonal antibodies (thAbs) - rituximab and ocrelizumab. Using our proprietary POC protein nanofactory system, a partial deep mutationally scanned (DMS) CD20 ECD mutant library of 79 variants was produced on SPOC biosensor chips via rational single amino acid substitutions of the epitope and surrounding residues with alanine, aspartic acid, lysine, and serine, collectively representing four broad classes of amino acid side chain chemistries: nonpolar, acidic, basic, and polar neutral. The SPOC protein biosensor chip was then screened with both thAbs using SPOC SPR to generate kinetic affinity data, evaluate mutations that led to affinity loss or gain, and ultimately identify critical epitope residues that interface with the antibodies. Most mutations within the rituximab and ocrelizumab epitopes - EPANPSEK and YNCEPANPSEKNSPST, respectively - resulted in complete loss of binding or >25% increase in apparent KD. Notably, N171, P172, and S173 mutations, irrespective of side chain substitution, resulted in complete loss of rituximab binding while at least three diverse side chain substitutions at E168, P169, N171, P172, S173, E174, K175, and T180, led to complete loss of binding for ocrelizumab. These outcomes identify the listed residues as the most critical contact points for their respective antibodies. Interestingly, we also found that functional side-chain substitutions at some residues flanking the epitope increased affinity. This indicates that these non-epitope residues contribute to antibody contact, and that polarity at these sites is a tractable lever for affinity modulation by targeting the corresponding contact residues on the antibody CDRs. The proposed SPOC approach of screening drug candidates against on-chip library of mutationally-scanned therapeutic targets is relevant in the early phase of drug development to resolve epitopes at the residue-level to support more informed down-selection of candidates. It facilitates cost-effective improvement of thAbs, enhancing therapeutic efficacy across a wide array of therapeutic targets, including rare variants that might otherwise lead to therapeutic resistance.

immunology↗

bioRxiv2026

Interactions between age and sex in multiscale entropy and spectral power changes across the lifespan

Multiscale entropy (MSE) changes in relation to age, whereby aging is associated with an increasing bias towards fine scale entropy. This change is thought to represent a shift toward localized information processing in the brain as we age. However, this relationship has not been tested in large sample sizes alongside other demographic factors and cognitive behaviours. This study aimed to validate previously reported effects of aging on MSE in a large open access database (Cambridge Centre for Ageing and Neuroscience, N=587) and expand the findings to include an investigation of the effects of sex and a variety of cognitive behaviours. MSE curves and power spectrum densities (PSD) were calculated for each region of interest from the magnetoencephalography data. Multivariate partial least squares analyses were used to assess the relationship between MSE or PSD and 5 behavioural / demographic factors including: age, sex, fluid intelligence, visual short-term memory and a generalized measure of cognitive function. Age was associated with increased fine scale and decreased coarse scale entropy, as well as complementary spectral changes, including slowing of peak alpha rhythms, increased beta-band activity, and reduced gamma-band activity, which replicates prior MSE and PSD findings. In both domains, these age-related patterns differentiated based on sex with advancing age. Importantly, the unique effects of sex diverged between MSE and PSD. This result indicates that entropy-based measures can isolate aspects of temporal organization that are not clearly summarized by spectral structure alone.

neuroscience↗

bioRxiv2026

IFI16/AIM2 inflammasomes control Gal-9 and PVR in myeloid cells from PWH and their targeting improves immunotherapy against HIV-1

Chronic inflammation in people with HIV-1 (PWH) is associated with low CD4/CD8 T-cell ratios and inflammasome activation, which may impair monocyte (Mo) and monocyte-derived dendritic cell (MDDC) function and limit the efficacy of myeloid-based immunotherapies. Here, we investigated the role of inflammasome sensors in regulating immune ligands for TIM3 and TIGIT immune checkpoints in Mo and MDDC from PWH and their capacity to activate HIV-1-specific CD8+ T cells. Following TLR stimulation, galectin-9 (Gal-9), PVR and IL-1{beta} were preferentially upregulated in cells from PWH with low CD4/CD8 ratios. IFI16 and AIM2 expression was increased basally or inducible in Mo and co-localized with caspase-1 in these myeloid cells present in lymphoid tissues of HIV-1-infected humanized mice. Si-RNA or small molecule-mediated inhibition of IFI16/AIM2 reduced Gal-9 and PVR expression in MDDC and enhanced their ability to activate polyfunctional HIV-1-specific CD8+ T-cells displaying improved elimination of infected CD4+ T cells in vitro and in vivo. These findings identify IFI16 and AIM2 as key regulators of myeloid dysfunction and therapeutic targets to enhance HIV-1 immunotherapies.

immunology↗

bioRxiv2025

AETA peptide drives Alzheimer's disease signature of synapse dysfunction

Alzheimers disease (AD), the leading cause of dementia, is characterized by early synaptic dysfunction that precedes overt cognitive decline. While amyloid-{beta} and Tau remain central to AD pathogenesis, molecular triggers of synapse weakening remain unclear. Here, we investigated AETA, a novel brain-secreted peptide derived from amyloid precursor protein (APP), as a potential mediator of synapse dysfunction in AD. We previously identified AETA as a unique modulator of NMDA receptor activity in the healthy brain; however, its role in AD etiology was yet to be explored. Post-mortem analyses of human hippocampal and prefrontal cortex tissues revealed significantly elevated AETA levels in AD patients, particularly in females. To further explore the contribution of AETA to AD synaptic pathology, we analyzed a new mouse model, the AETA-m mouse, exhibiting chronically increased brain AETA expression. Hippocampi of female AETA-m mice display an increase in the number of astrocyte and microglia, but no overt neuroinflammation. RNA sequencing of female AETA-m hippocampi revealed alterations in synaptic gene expression that closely paralleled those observed in vulnerable human AD brain regions, most notably in the hippocampus. These two phenotypes were absent in males. Functionally, hippocampal neurons from AETA-m mice displayed impaired NMDA receptor signaling, dendritic spine loss, and memory deficits especially in females, mirroring early AD-associated synaptic dysfunction. Together, these findings identify AETA as a novel key contributor of synaptic vulnerability in AD and associated memory processing, especially in females. Targeting AETA signaling may therefore offer new therapeutic avenues for preventing or mitigating synaptic and cognitive decline in AD.

neuroscience↗

bioRxiv2025

Sterols regulate ciliary membrane dynamics and hedgehog signaling in health and disease

The primary cilium is a specialized signaling hub whose function depends on a tightly regulated membrane composition. While its protein content is well-characterized, its lipid identity, particularly regarding sterols, remains poorly defined. Here, we used mass spectrometry-based lipidomics to map the sterol profile of isolated primary cilia from MDCK cells. We found that ciliary membranes are enriched in cholesterol and desmosterol while excluding precursors like 7-lathosterol and limiting others, suggesting a selective sterol barrier. Inhibiting cholesterol biosynthesis at distinct enzymatic steps led to sterol accumulation, altered ciliary membrane fluidity, and impaired Hedgehog signaling, including defective Smoothened (Smo) retention-- even in the presence of a constitutively active form of Smo. These findings link sterol homeostasis to ciliary membrane properties and signaling fidelity. Our work provides a molecular framework for understanding Hedgehog-related phenotypes in disorders like Smith-Lemli-Opitz Syndrome, highlighting the importance of membrane lipid composition in developmental signaling.

cell biology↗

bioRxiv2025

Combination of Cas9 and adeno-associated vectors (AAVs) enables efficient in vivo knockdown of precise miRNAs in the rodent brain

Although the advent of Cas9 technology has expanded our ability to precisely edit the genome, manipulating microRNAs in vivo has been shown to be particularly challenging, especially in the brain. Here, we sought to generate novel tools aiming at targeting and efficiently downregulating defined microRNAs species in a cell-specific manner so that their function in discrete neuronal networks could be investigated. Focusing on miR-124, a microRNA highly expressed in the mammalian brain and transcribed from three independent chromosomal loci, we designed and validated different guide RNAs directed against this miRNA. In vitro, our Cas9 designs show not only a significant reduction in miR-124 levels but also a functional effect on miR-124 silencing. Similarly, when packed into AAV vectors and injected into the mouse cortex, miR-124-Cas9 vectors strongly downregulate miR-124 levels without affecting the expression of other miRNAs. In parallel, levels of endogenous miR-124 targets exhibit a significant increase supporting the release of its silencing activity. To functionally validate our tools, we provide evidences that deletion of miR-124 in the subventricular zone altered migration of newly generated neurons into the olfactory bulb. Finally, we also showed that our vectors modified the Ca2+ permeability of AMPA receptors, a robust functional output downstream of miR-124. These tools are expected to help elucidating miRNA function in complex experimental settings such as brain networks in vivo.

neuroscience↗

bioRxiv2025

Generation and characterization of an ovine cell line derived from peripheral blood and its potential use to study livestock and zoonotic viral infections

BackgroundMesenchymal stromal cells (MSCs) are a population of undifferentiated non-hematopoietic fibroblast-like cells isolated from several tissues with multipotent differentiation capacity in vitro. This study focused on the establishment and characterization of a mesenchymal stromal cell line derived from ovine peripheral blood mononuclear cells (PBMCs). MethodsMSC were isolated from ovine blood and used to develop a cell line. The characterization of the cell line to confirm its mesenchymal origin was carried out by different assays. First at all, cells were characterized by flow cytometry (FACS) using monoclonal antibodies specific for MSC and hematopoietic non-stromal cell surface antigens. Real-time quantitative PCR (RT-qPCR) was performed using gene primer pairs specific for hematopoietic and mesenchymal cell surface markers. In addition, we assayed the potential of the cell line to differentiate in vitro to other lineages such as osteoblasts and neurons and confirmed that by specific staining with Alizarin Red (AR) and RT-qPCR and immunofluorescence (IFA). Finally, we assayed the susceptibility of the cell line to various livestock, animal and human viruses. ResultsFACS and RT-qPCR analyses revealed that the ovine cell line expressed mesenchymal markers and was negative for hematopoietic markers assayed. MSC were able to differentiate to osteoblasts and neurons, these results were confirmed by quantitative analyses of AR staining and RT-qPCR and IFA assays. The cell line showed susceptibility to infection with all viruses assayed confirmed by IFA asssays. ConclusionsIn this work, we used isolated MSCs from peripheral blood to establish a new ovine cell line. The characterization and the osteogenic and neuronal differentiation carried out confirmed that the cell line had a mesenchymal stromal origin. In fact, the new established ovine cell line was permissive to all viruses studied in this work. The new sheep cell line could be a useful tool for isolating and characterizing viruses and studying virus-host interactions.

cell biology↗

bioRxiv2025

The hyper-attenuated RVFV 40Fp8 strain can be safely administered to pregnant ewes to protect them from a virulent challenge

In the present study, we evaluated the immunogenicity, safety and protective efficacy of the attenuated RVFV-40Fp8 strain in natural hosts (non-pregnant ewes) and in a highly susceptible host infection model such as pregnant ewes in the first third of pregnancy. Our results confirm the immunogenicity of 40Fp8 administration in non-pregnant and pregnant ewes, as well as the absence of foetal damage even after a high-dose vaccination regime in pregnant ewes. In addition, the ewes and their foetuses were protected against a virulent RVFV-56/74 strain challenge, as shown by comparative histopathological evaluation of tissue samples from vaccinated and non-vaccinated pregnant ewes. These results confirm the potential use of 40Fp8 as a RVF live-attenuated vaccine candidate and pave the way for further clinical developments.

microbiology↗

bioRxiv2025

An approach to produce thousands of single-chain antibody variants on a SPR biosensor chip for measuring target binding kinetics and for deep characterization of antibody paratopes

Drug discovery continues to face a staggering 90% failure rate, with many setbacks occurring during late-stage clinical trials. To address this challenge, there is an increasing focus on developing and evaluating new technologies to enhance the "design" and "test" phases of antibody-based drugs (e.g., monoclonal antibodies, bispecifics, CAR-T therapies, ADCs) and biologics during early preclinical development, with the goal of identifying lead molecules with a higher likelihood of clinical success. Artificial intelligence (AI) is becoming an indispensable tool in this domain, both for improving molecules identified through traditional approaches and for the de novo design of novel therapeutics. However, critical bottlenecks persist in the "build" and "test" phases of AI-designed antibodies and protein binders, impeding early preclinical evaluation. While AI models can rapidly generate thousands to millions of putative drug designs, technological and cost limitations mean that only a few dozen candidates are typically produced and tested. Drug developers often face a tradeoff between ultra-high-throughput wet lab methods that provide binary yes/no binding data and biophysical methods that offer detailed characterization of a limited number of drug-target pairs. To address these bottlenecks, we previously reported the development of the Sensor-integrated Proteome On Chip (SPOC(R)) platform, which enables the production and capture-purification of 1,000 - 2,400 folded proteins directly onto a surface plasmon resonance (SPR) biosensor chip for measuring kinetic binding rates with picomolar affinity resolution. In this study, we extend the SPOC technology to the expression of single-chain antibodies (sc-antibodies), specifically scFv and VHH, and dual-chain Fab constructs. We demonstrate that these proteins are capture-purified at high levels on SPR biosensors and retain functionality as shown by the binding specificity to their respective target antigens, with affinities comparable to those reported in the literature. SPOC outputs comprehensive kinetic data including quantitative binding (Rmax), on-rate (ka), off-rate (kd), affinity (KD), and half-life (t1/2), for each of thousands of on-chip sc-antibodies. Additionally, we present a case study showcasing single amino acid mutational scan of the complementarity-determining regions (CDRs) of a HER2 VHH (nanobody) paratope. Using 92 unique mutated variants from four different amino acid substitutions, we pinpoint critical residues within the paratope that could further enhance binding affinity. This study serves as a demonstration of a novel high-throughput approach for biophysical screening of hundreds to thousands of single chain antibody sequences in a single assay, generating high affinity resolution kinetic data to support antibody discovery and AI-enabled pipelines.

immunology↗

bioRxiv2024

p75NTR modulation by LM11A-31 counteracts oxidative stress and cholesterol dysmetabolism in a rotenone-induced cell model of Parkinson's disease

Neurotrophins play pivotal roles in the development and proper functioning of the nervous system. The effects of these growth factors are mediated through the binding of high-affinity receptors, known as Trks, as well as the low-affinity receptor p75NTR. The latter has the capacity to induce complex signal pathways, as it favors both pro-survival and pro-apoptotic cascades depending on the physiopathological condition. Recent findings have indicated that p75NTR expression is increased in post-mortem Parkinsons disease (PD) brains, and this upregulation is associated with a significant reduction in neuroprotection. Given its double-edged sword nature, p75NTR has recently been identified as a promising therapeutic target to counteract neurodegenerative events. The present study aims to assess the neuroprotective effects of p75NTR modulation in a rotenone-induced neuronal model of PD. To this end, differentiated SH-SY5Y cells were exposed to rotenone to mimic the PD phenotype, and the small molecule LM11A-31 was used to modulate p75NTR activity. The main results revealed that LM11A-31 significantly mitigated the hallmarks of PD, including cell death, neuromorphological aberrations, and -synuclein accumulation. Pharmacological manipulation of p75NTR also reduced oxidative damage by increasing the expression of transcriptional factors that regulate the antioxidant response and by decreasing the expression of the pro-oxidant NADPH-oxidase modulatory subunits. Furthermore, LM11A-31 hampered cholesterol buildup induced by rotenone, normalizing the expression of proteins involved in cholesterol biosynthesis, uptake and intracellular trafficking. Taken together, these findings suggest that p75NTR modulation may represent a novel approach to counteracting PD abnormalities of redox and cholesterol metabolism.

neuroscience↗

bioRxiv2024

Novel replication-competent reporter-expressing Rift Valley fever viruses for molecular studies

Rift Valley fever virus (RVFV) is a mosquito-borne zoonotic disease that causes severe disease in both domestic and wild ungulates and humans, making it a significant threat to livestock and public health. The RVFV genome consists of three single-stranded, negative-sense RNA segments differing in size: Small (S), Medium (M) and Large (L). Segment S encodes the virus nucleoprotein N and the virulence-associated factor non-structural (NSs) protein in opposite orientations, separated by an intergenic region (IGR). To overcome the current need of using secondary techniques to detect the presence of RVFV in infected cells, we used T7-driven polymerase plasmid-based reverse genetics to generate replication-competent recombinant (r)RVFV expressing Nanoluciferase (Nluc) or Venus fluorescent proteins. These reporter genes were used as valid surrogates to track the presence of RVFV in mammalian and insect cells. Notably, we explored the genome plasticity of RVFV and compared four different strategies by modifying the viral segment S in order to introduce the reporter gene foreign sequences. The reporter-expressing rRVFV were stable and able to replicate in cultured mammalian and insect cells, although to a lesser extent than the recombinant wild-type (WT) counterpart. Moreover, rRVFV expressing reporter genes were validated to identify neutralizing antibodies or compounds with antiviral activity. In vivo, all mice infected with the reporter-expressing rRVFV displayed an attenuated phenotype, although at different levels. These rRVFV expressing reporter genes provide a novel approach to better understand the biology and pathogenesis of RVFV, and represent an excellent biotechnological tool for developing new therapeutics against RVFV infections. IMPORTANCERift Valley fever virus (RVFV) is a mosquito-borne virus and zoonotic agent threat that can be deadly to domestic or wild ungulates, and humans. In this work, we used reverse genetics approaches to explore the genome plasticity of RVFV by generating a set of recombinant rRVFV that express fluorescent or luminescent proteins to track viral infection. All the generated reporter-expressing rRVFV were able to propagate in mammalian or insect cells, and in a mouse model of infection. Our studies may contribute to advances in research on RVFV and other bunyaviruses and pave the way for the development of novel vaccines and the identification of new antivirals for the prophylactic and therapeutic treatment, respectively, of RVFV infections.

microbiology↗

bioRxiv2024

Viral overexpression of human alpha-synuclein in mouse substantia nigra dopamine neurons results in hyperdopaminergia but no neurodegeneration

Loss of select neuronal populations such as midbrain dopamine (DA) neurons is a pathological hallmark of Parkinsons disease (PD). The small neuronal protein -synuclein has been related both genetically and neuropathologically to PD, yet how it contributes to selective vulnerability remains elusive. Here, we describe the generation of a novel adeno-associated viral vector (AAV) for Cre-dependent overexpression of wild-type human -synuclein. Our strategy allows us to restrict -synuclein to select neuronal populations and hence investigate the cell-autonomous effects of elevated -synuclein in genetically-defined cell types. Since DA neurons in the substantia nigra pars compacta (SNc) are particularly vulnerable in PD, we investigated in more detail the effects of increased -synuclein in these cells. AAV-mediated overexpression of wildtype human -synuclein in SNc DA neurons increased the levels of -synuclein within these cells and augmented phosphorylation of -synuclein at serine-129, which is considered a pathological feature of PD and other synucleinopathies. However, despite abundant -synuclein overexpression and hyperphosphorylation we did not observe any DA neurodegeneration up to 90 days post virus infusion. In contrast, we noticed that overexpression of -synuclein resulted in increased locomotor activity and elevated striatal DA levels suggesting that -synuclein enhanced dopaminergic activity. We therefore conclude that cell-autonomous effects of elevated -synuclein are not sufficient to trigger acute DA neurodegeneration.

neuroscience↗

bioRxiv2024

Identification of Fibrinogen as a Plasma Protein Binding Partner for Lecanemab Biosimilar IgG: Implications for Alzheimer's Disease Therapy

ObjectiveRecombinant monoclonal therapeutic antibodies like lecanemab, which target amyloid beta in Alzheimers disease, offer a promising approach for modifying the disease progression. Due to its relatively short half-life, Lecanemab, administered as a bi-monthly infusion (typically 10mg/kg) has a relatively brief half-life. Interaction with abundant plasma proteins binder in the bloodstream can affect pharmacokinetics of drugs, including their half-life. In this study we investigated potential plasma protein binding interaction to lecanemab using lecanemab biosimilar. MethodsLecanemab biosimilar used in this study was based on publicly available sequences. ELISA and Western blotting were used to assess lecanemab biosimilar immunoreactivity in the fractions human plasma sample obtained through size exclusion chromatography. The binding of lecanemab biosimilar to candidate binders was confirmed by Western blotting, ELISA, and surface plasmon resonance analysis. ResultsUsing a combination of equilibrium dialysis, ELISA, and Western blotting in human plasma, we first describe the presence of likely plasma protein binding partner to lecanemab biosimilar, and then identify fibrinogen as one of them. Utilizing surface plasmon resonance, we confirmed that lecanemab biosimilar does bind to fibrinogen, although with lower affinity than to monomeric amyloid beta. InterpretationIn the context of lecanemab therapy, these results imply that fibrinogen levels could impact the levels of free antibodies in the bloodstream and that fibrinogen might serve as a reservoir for lecanemab. More broadly, these results indicate that plasma protein binding may be an important consideration when clinically utilizing therapeutic antibodies in neurodegenerative disease.

neuroscience↗

bioRxiv2024

The Rift Valley fever (RVF) vaccine candidate 40Fp8 shows an extreme attenuation in IFNARKO mice following intranasal inoculation

Rift Valley fever (RVF) is an important zoonotic viral disease affecting several species of domestic and wild ruminants, causing major economic losses and dozens of human deaths in various geographical areas of Africa, where it is endemic. Although it is not present in Europe, there is a risk of its introduction and spread linked to globalisation and climate change. At present, the only measure that could help to prevent the disease is vaccination of flocks in areas at risk of RVF. Available live attenuated vaccines are an effective means of controlling the disease, but their use is often questioned due to residual virulence, particularly in susceptible hosts such as pregnant sheep. On the other hand, no vaccine is currently licensed for use in humans. The development of safe and effective vaccines is therefore a major area of research. In previous studies, we selected under selective mutagenic pressure a highly attenuated RVFV 56/74 virus variant called 40Fp8. This virus showed an extremely attenuated phenotype in both wild-type and immunodeficient A129 (IFNARKO) mice, yet was still able to induce protective immunity after a single inoculation, thus supporting its use as a safe, live attenuated vaccine. To further investigate its safety, in this work we have analysed the attenuation level of 40Fp8 in immunosuppressed mice (A129) when administered by the intranasal route, and compared it with other attenuated RVF viruses that are the basis of vaccines in use or in development. Our results show that 40Fp8 has a much higher attenuated level than these other viruses and confirm its potential as a candidate for safe RVF vaccine development. Author SummaryVaccines that use attenuated viruses are highly effective in terms of providing protection, duration, breadth, and quality of the immune response. However, they may pose a risk to immunosuppressed individuals or in certain situations, such as during pregnancy. It is therefore important to analyze the residual virulence of attenuated vaccines to ensure their safety. In this work, we analysed the safety of the RVF virus vaccine variant 40Fp8 developed in our laboratory. We studied its effect on immunodeficient mice and used a more aggressive route of administration such as the intranasal delivery route. To assess the degree of attenuation of 40Fp8, we compared it with other attenuated prototype RVF vaccines. The results clearly demonstrate that 40Fp8 is highly attenuated, and its residual virulence level is extremely low compared to other vaccine viruses tested.

pathology↗

bioRxiv2023

The Azotobacter vinelandii AlgU Regulon During Vegetative Growth and Encysting Conditions: A Proteomic Approach

In the Pseduomonadacea family, the extracytoplasmic function sigma factor AlgU is crucial to withstand adverse conditions. Azotobacter vinelandii, a closed relative of Pseudomonas aeruginosa, has been a model for cellular differentiation in Gram-negative bacteria since it forms desiccation-resistant cysts. Previous work demonstrated the essential role of AlgU to withstand oxidative stress and on A. vinelandii differentiation, particularly for the positive control of alginate production. In this study, the AlgU regulon was dissected by a proteomic approach under vegetative growing conditions and upon encystment induction. Our results revealed several molecular targets that explained the requirement of this sigma factor during oxidative stress and extended its role in alginate production. Furthermore, we demonstrate that AlgU was necessary to produce alkyl resorcinols, a type of aromatic lipids that conform the cell membrane of the differentiated cell. AlgU was also found to positively regulate stress resistance proteins such as OsmC, LEA-1, or proteins involved in trehalose synthesis. A position-specific scoring-matrix (PSSM) was generated based on the consensus sequence recognized by AlgU in P. aeruginosa, which allowed the identification of direct AlgU targets in the A. vinelandii genome. This work further expands our knowledge about the function of the ECF sigma factor AlgU in A. vinelandii and contributes to explains its key regulatory role under adverse conditions.

microbiology↗