Biology subjects

Carolis, C.

Publications and source records attributed to Carolis, C..

9 recordsLinked to original sources

bioRxiv2026

A cold-adapted Cas9 nuclease from the deep ocean

The abundance of CRISPR systems in bacteria and archaea fuels the search for Cas nucleases that perform where established enzymes fail. Deep Ocean 1 Cas9 (DO1Cas9) is to our knowledge the first Cas9 nuclease characterized from the deep ocean, recovered from a metagenomic sample at 1,500 m depth in the Atlantic Ocean. DO1Cas9 shows cold-adapted structural flexibility, thermal susceptibility, salt-dependent stability, higher specificity than SpCas9 and cleaves double-stranded DNA efficiently under cold conditions where mesophilic SpCas9 retains only residual activity. DO1Cas9 edits genomes in diverse organisms such as Caenorhabditis elegans, zebrafish, the marine tunicate Oikopleura dioica, and the malaria parasite Plasmodium falciparum. In the marine bacterium Marinomonas mediterranea, DO1Cas9 programmed against an invading phage reduced plaquing efficiency about 10-fold more than SpCas9. These results expand the CRISPR-Cas9 toolbox with a nuclease for cold-chain editing and marine biotechnology.

genetics↗

bioRxiv2026

Protective near-germline antibodies to malaria elicited by RTS,S/AS01E vaccination in African infants

Malaria is a leading cause of childhood mortality, yet infant immune responses to vaccination remain poorly understood. Here, we characterize 88 monoclonal antibodies (mAbs) directed against the Plasmodium falciparum circumsporozoite protein cloned from African infants immunized with the RTS,S/AS01E malaria vaccine, identifying potent antiparasitic mAbs exhibiting molecular features resembling those observed in adults and minimal somatic hypermutation. Importantly, germline-reverted variants of four mAbs inducing sterile protection in vivo maintained high binding affinity and Fab-dependent parasite killing. Structural analyses revealed that a few amino acid replacements drive homotypic helical assemblies, fine-tuning parasite inhibition and protective efficacy. Thus, despite maturational constraints of early life, infants can generate potent antimalarial antibodies in a near-germline configuration, informing next-generation vaccine development for this vulnerable group.

immunology↗

bioRxiv2025

Minimal impact of ivermectin on immune response and transcriptional profiles in naïve adults with mild COVID-19

Ivermectin (IVM), an antiparasitic drug, was repurposed to treat COVID-19 based on its in vitro antiviral effects. However, it was abandoned after multiple clinical trials reported a lack of efficacy. Immunomodulatory effects have been proposed but remain unclear, yet they may be relevant given IVM use for other infections. We assessed the IVM immunomodulatory effect in 24 participants from a clinical trial evaluating its potential to reduce COVID-19 transmission in mild cases within 48 hours of symptoms onset. The IVM-treated patients showed non-significant lower viral loads, and a significantly shorter duration of hyposmia/anosmia. We measured IgG, IgA, and IgM against five SARS-CoV-2 antigens, and 30 cytokines by Luminex, alongside whole blood RNA sequencing, pan-leukocyte immunophenotyping, and SARS-CoV-2-specific T cell analysis by flow cytometry from day 1 to day 28 post-treatment. All antibody responses increased from day 4, while 13 cytokines significantly decreased over time (adjusted p<0.05). IVM-treated patients had only significantly higher anti-nucleocapsid IgG levels at day 4 (adjusted p=0.041) and 7 (adjusted p=0.045) compared to placebo. SARS-CoV-2-specific CD4+ and CD8+ T cells increased over time, with significantly higher effector memory CD4+ T cells at day 7 compared to day 1 (p=0.027) and the only difference between groups was lower frequencies of spike-specific naive CD4+ T cells at day 7 in IVM-treated participants (0.006% vs 0.036% p=0.02). Transcriptomic data showed downregulation of innate and antiviral blood transcriptional modules (BTMs) over time, with an increase in adaptive immune related BTMs. While no differential gene expression was detected, the IVM-treated had upregulated innate and downregulated T cell and cell cycle BTMs compared to placebo. Overall, our comprehensive longitudinal analysis of early immune responses in mild COVID-19 revealed no robust immunological effects of IVM, consistent with clinical trials results and suggesting a lack of efficacy of IVM in COVID-19 treatment. AUTHOR SUMMARYIvermectin (IVM), an antiparasitic drug, was tested in clinical trials as a potential treatment for COVID-19 due to its in vitro antiviral properties and hypothetical immunomodulatory effects. In this study, we explored the immunomodulatory effects of IVM in a clinical trial involving 24 mild COVID-19 patients who received IVM within 48 hours of symptoms onset. The IVM group showed a trend towards lower viral loads post-treatment, and a significantly shorter duration of hyposmia/anosmia. We comprehensively analyzed immune responses by measuring antibody levels, cytokine profiles, immune cell subsets and whole blood gene expression over 28 days. The IVM group only had increased levels of IgG against the SARS-CoV-2 nucleocapsid compared to the control group. IVM did not affect the kinetics of SARS-CoV-2 T cells, despite a slight decrease in naive CD4+ T cells. Additionally, gene expression analysis showed a decrease in innate and antiviral responses and an increase of adaptive responses over time that were slightly stronger in the placebo compared to the IVM group. In conclusion, despite some differences in IVM treated participants, our detailed analyses do not support significant immunomodulatory effects that could benefit disease progression.

immunology↗

bioRxiv2024

De novo design of high-affinity single-domain antibodies

Antibody-based therapeutics have become indispensable in modern medicine, but traditional methods of antibody discovery often present with limitations in developability, cross-reactivity, and ethical concerns. While deep learning and generative approaches have shown promise in the design of high affinity protein binders, de novo antibody design remains challenging. Here, we present EvolveX, a structure-based computational pipeline for designing antibody fragments. EvolveX utilizes ModelX and empirical force field FoldX to optimize complementarity- determining regions (CDRs) and TANGO for aggregation analysis. We demonstrate the ability of EvolveX to redesign a single-domain VHH antibody fragment targeting mouse Vsig4 to address two challenges: enhancing stability and affinity for the original target and redesigning it for high affinity to the human ortholog. The redesigned variants of VHH fragments specific to mouse Vsig4 showed improved physicochemical properties, while retaining binding affinities comparable to the original version. Notably, EvolveX improved the binding affinity of VHHs to human Vsig4 by over 1000-fold, transforming low-affinity binders into nanomolar-affinity molecules. Structural analyses by X-ray crystallography and NMR confirmed the accuracy of the designs, which display optimized interactions with the antigen. NGS and re-modelling analysis further demonstrated the efficiency of FoldX-based design pipeline. Collectively, our study highlights EvolveXs potential to overcome current limitations in antibody design, offering a powerful tool for the development of next-generation therapeutics with enhanced specificity, stability, and efficacy.

bioinformatics↗

bioRxiv2023

A Novel Monoclonal Antibody Targeting a Large Surface of the Receptor Binding Motif Shows Pan-neutralizing SARS-CoV-2 Activity Including BQ.1.1 Variant

In the present study we report the functional and structural characterization of 17T2, a new highly potent pan-neutralizing SARS-CoV-2 human monoclonal antibody (mAb) isolated from a convalescent COVID-19 individual infected during the first wave of the COVID-19 pandemic. 17T2 is a class 1 VH1-58/{kappa}3-20 antibody, derived from a receptor binding domain (RBD)-specific IgA memory B cell and developed as a human recombinant IgG1. Functional characterization revealed that 17T2 mAb has a high and exceptionally broad neutralizing activity against all SARS-CoV-2 spike variants tested, including BQ.1.1. Moreover, 17T2 mAb has in vivo prophylactic activity against Omicron BA.1.1 infection in K18-hACE2 transgenic mice. 3D reconstruction from cryogenic-electron microscopy (cryo-EM) showed that 17T2 binds the Omicron BA.1 spike protein with the RBD domains in "up" position and recognizes an epitope overlapping with the receptor binding motif, as it is the case for other structurally similar neutralizing mAbs, including S2E12. Yet, unlike S2E12, 17T2 retained its high neutralizing activity against all Omicron sublineages tested, probably due to a larger contact area with the RBD, which could confer a higher resilience to spike mutations. These results highlight the impact of small structural antibody changes on neutralizing performance and identify 17T2 mAb as a potential candidate for future therapeutic and prophylactic interventions.

immunology↗

bioRxiv2021

Genome editing in animals with minimal PAM CRISPR-Cas9 enzymes

The requirement for Cas nucleases to recognize a specific PAM is a major restriction for genome editing. SpCas9 variants SpG and SpRY, recognizing NGN and NRN PAM, respectively, have contributed to increase the number of editable genomic sites in cell cultures and plants. However, their use has not been demonstrated in animals. We have characterized and optimized the activity of SpG and SpRY in zebrafish and C. elegans. Delivered as mRNA-gRNA or ribonucleoprotein (RNP) complexes, SpG and SpRY were able to induce mutations in vivo, albeit at a lower rate than SpCas9 in equivalent formulations. This lower activity was overcome by optimizing mRNA-gRNA or RNP concentration, leading to efficient mutagenesis at regions inaccessible to SpCas9. We also found that the CRISPRscan algorithm can predict SpG and SpRY activity in vivo. Finally, we applied SpG and SpRY to generate knock-ins by homology-directed repair. Altogether, our results expand the CRISPR-Cas targeting genomic landscape in animals.

genetics↗

bioRxiv2021

Antibody Conversion rates to SARS-CoV-2 in Saliva from Children Attending Summer Schools in Barcelona, Spain

Surveillance tools to estimate infection rates in young populations are essential to guide recommendations for school reopening and management during viral epidemics. Ideally, field-deployable non-invasive, sensitive techniques are required to detect low viral load exposures among asymptomatic children. We determined SARS-CoV-2 antibody conversion by high-throughput Luminex assays in saliva samples collected weekly in 1,509 children and 396 adults in 22 Summer schools and 2 pre-schools in 27 venues in Barcelona, Spain, from June 29th to July 31st 2020, between the first and second COVID-19 pandemic waves. Saliva antibody conversion defined as [≥]4-fold increase in IgM, IgA and/or IgG levels to SARS-CoV-2 antigens between two visits over a 5-week period was 3.22% (49/1518), or 2.36% if accounting for potentially cross-reactive antibodies, six times higher than the cumulative infection rate (0.53%) by weekly saliva RT-PCR screening. IgG conversion was higher in adults (2.94%, 11/374) than children (1.31%, 15/1144) (p=0.035), IgG and IgA levels moderately increased with age, and antibodies were higher in females. Most antibody converters increased both IgG and IgA antibodies but some augmented either IgG or IgA, with a faster decay over time for IgA than IgG. Nucleocapsid rather than spike was the main antigen target. Anti-spike antibodies were significantly higher in individuals not reporting symptoms than symptomatic individuals, suggesting a protective role against COVID-19. To conclude, saliva antibody profiling including three isotypes and multiplexing antigens is a useful and more user-friendly tool for screening pediatric populations to determine SARS-CoV-2 exposure and guide public health policies during pandemics.

immunology↗

bioRxiv2020

Highly sensitive and specific multiplex antibody assays to quantify immunoglobulins M, A and G against SARS-CoV-2 antigens

Reliable serological tests are required to determine the prevalence of antibodies against SARS-CoV-2 antigens and to characterise immunity to the disease in order to address key knowledge gaps in the context of the COVID-19 pandemic. Quantitative suspension array technology (qSAT) assays based on the xMAP Luminex platform overcome the limitations of rapid diagnostic tests and ELISA with their higher precision, dynamic range, throughput, miniaturization, cost-efficacy and multiplexing capacity. We developed three qSAT assays to detect IgM, IgA and IgG to a panel of eight SARS-CoV-2 antigens including spike (S), nucleoprotein (N) and membrane (M) protein constructs. The assays were optimized to minimize processing time and maximize signal to noise ratio. We evaluated the performance of the assays using 128 plasmas obtained before the COVID-19 pandemic (negative controls) and 115 plasmas from individuals with SARS-CoV-2 diagnosis (positive controls), of whom 8 were asymptomatic, 58 had mild symptoms and 49 were hospitalized. Pre-existing IgG antibodies recognizing N, M and S2 proteins were detected in negative controls suggestive of cross-reactive to common cold coronaviruses. The best performing antibody isotype/antigen signatures had specificities of 100% and sensitivities of 94.94% at [≥]14 days since the onset of symptoms and 96.08% at [≥]21 days since the onset of symptoms, with AUC of 0.992 and 0.999, respectively. Combining multiple antibody markers as assessed by qSAT assays has the highest efficiency, breadth and versatility to accurately detect low-level antibody responses for obtaining reliable data on prevalence of exposure to novel pathogens in a population. Our assays will allow gaining insights into antibody correlates of immunity required for vaccine development to combat pandemics like the COVID-19.

immunology↗

bioRxiv2017

Probing tissue-scale deformation by in vivo force application reveals a fast tissue softening during early embryogenesis

During development, cell-generated forces induce tissue-scale deformations to shape the organism. Here, we present a method that allows to quantitatively relate such tissue-scale deformations to spatially localized forces and measure mechanical properties of epithelia in vivo. Our approach is based on the application of controlled forces on microparticles embedded in individual cells of an embryo. Combining measurements of the bead displacement with the analysis of induced deformation fields in a continuum mechanics framework, we can quantify tissue material properties and follow their change over time. In particular, we uncover a rapid change in tissue response occurring during Drosophila cellularization, resulting from a softening of the blastoderm and an increase of external friction. Pharmacological treatments reveal that in addition to actomyosin, the microtubule cytoskeleton is a major contributor to epithelial mechanics at that stage. Overall, our method allows for measuring essential mechanical parameters governing tissue-scale deformations and flows occurring during morphogenesis.

bioinformatics↗