Biology subjects

Wermeling, F.

Publications and source records attributed to Wermeling, F..

9 recordsLinked to original sources

bioRxiv2026

Correlate: A Web Application for Analyzing Gene Sets and Exploring Gene Dependencies Using CRISPR Screen Data

CRISPR screen data provides a valuable resource for understanding gene function and identifying potential drug targets. Here, we present Correlate, a freely accessible web application (https://correlate.cmm.se) that enables exploration of the Cancer Dependency Map (DepMap) CRISPR screen gene effects, hotspot mutations, and translocation/fusion data across more than 1,000 human cancer cell lines. The application supports two main use cases: (i) analysis of user-defined gene sets (e.g. CRISPR screen hits) to identify functionally linked genes based on correlations while providing an overview based on essentiality or user-provided screen statistics; and (ii) exploration of genes of interest in defined biological contexts, such as specific cancer types or mutational backgrounds, to generate hypotheses about gene function and dependencies. Additionally, Correlate supports experimental design by providing rapid overviews of gene essentiality and enabling the identification of cell lines with relevant mutational profiles. In contrast to knowledge-based approaches such as STRING and GSEA, which rely on prior biological annotations and curated interaction networks, Correlate identifies gene connections directly from functional CRISPR screen readouts, offering a complementary and data-driven perspective on gene network analysis. The application runs entirely in the browser, requires no installation or login, and integrates with the Green Listed v2.0 tool family for custom CRISPR screen design. HIGHLIGHTS{blacksquare} Interactive web-based platform for bulk correlation analysis of user-defined gene sets using DepMap CRISPR screen data, requiring no installation or programming expertise. {blacksquare}Identifies functional gene relationships from CRISPR screen readouts rather than curated annotations, offering a data-driven complement to tools such as GSEA and STRING. {blacksquare}Enables contextual exploration of gene dependencies across cancer types and mutational backgrounds, supporting hypothesis generation about gene function and therapeutic targets. {blacksquare}Supports experimental design through gene essentiality overviews, mutation and fusion analysis, and cell line identification, with optional integration of user-provided statistics from CRISPR screens, proteomics, or transcriptomics analyses.

bioinformatics↗

bioRxiv2026

A new humanized TCR transgenic mouse model to study citrullinated tenascin C reactive T cells relevant to rheumatoid arthritis

ObjectivesAutoreactive T cells recognizing citrullinated antigens, although rare and difficult to study, are implicated in rheumatoid arthritis (RA). To allow functional studies and manipulation of such T cells, we have generated and characterized transgenic mice expressing a TCR cloned from an RA patient and reactive with a prominent target, citrullinated tenascin C (citTNC). MethodsA humanized TCR transgenic (hTCR-tg) mouse recognizing the citrullinated TNC22 (citTNC22) antigen in an HLA-DRB1*04:01 (HLA-DR4) restricted manner was developed by genetically engineering a chimeric TCR expressing murine constant domains and human V(D)J sequences. hTCR-tg mice were immune phenotyped in murine H-2b and humanized HLA-DR4 backgrounds using full spectrum flow cytometry, cytokine ELISAs and fluorospot assays at steady-state and after antigen challenge. Additionally, we investigated the presence of antibodies to citTNC and arthritis following citTNC protein immunization. ResultsThymic selection of hTCR T cells differed between the two MHC-II alleles, with a normal CD4+ T cell development observed solely under HLA-DR4 restriction. The chimeric hTCR maintained its citTNC22 specificity in vitro and ex vivo, without cross-reactivity to native TNC22 or other citrullinated autoantigens. hTCR-tg CD4+ T cells responded to antigen challenge in vivo and provided support to IgG class-switch and antigen-specific antibody production. Moreover, protein immunized hTCR-tg mice developed arthritis after periarticular challenge with citTNC. ConclusionshTCR-tg mice expressing an RA patient-derived autoreactive TCR develop functional antigen-specific and HLA-restricted CD4+ T cells. The combination of humanized HLA-DR4 and TNC22 mice will be a valuable tool in the development of antigen-specific therapies translatable to human disease.

immunology↗

bioRxiv2025

A neoantigen-microbead platform for personalized T cell cancer vaccination

Colorectal cancer (CRC) is a leading cause of cancer mortality and is characterized by a high tumor mutational burden, making it responsive to immunotherapy. We developed a bioinformatic and manufacturing pipeline for personalized cancer vaccines and evaluated it in the MC38 colon adenocarcinoma mouse model. Thirty-six high-scoring neoantigens (NAGs) were identified by exome and transcriptome analysis, produced as six purified polypeptides, and coupled to paramagnetic beads. Intralymphatic vaccination of C57BL/6 mice with NAG beads induced robust NAG-specific T cell and antibody responses, resulting in significant inhibition of MC38 tumor growth. Treated tumors displayed increased necrosis and CD8+ T cell infiltration. Compared with soluble peptides, bead-coupled antigens elicited superior protection. Studies in T cell-deficient and antibody-depleted mice confirmed that both CD4+ and CD8+ T cells mediated the antitumor effect. These findings highlight the potential of NAG bead vaccination as an effective immunotherapy for CRC.

cancer biology↗

bioRxiv2024

Dnmt3a mutations limit normal and autoreactive Tfh differentiation

Rheumatoid arthritis (RA) is an autoimmune disease characterized by joint inflammation, strongly associated with the activity of autoreactive CD4+ T cells. DNMT3A mutations are the most common somatic mutations found in the hematopoietic system of patients with rheumatoid arthritis. However, the role of DNMT3A in CD4+ T cells and CD4+ T follicular helper (Tfh) cells is poorly understood. Since somatic mutations are not identified in standard genome-wide association studies, somatic mutations impact on the etiology of diseases could be underestimated. Here, we thoroughly characterized and used the KRN+ splenocyte transfer model of autoimmune joint inflammation and inactivated Dnmt3a using CRISPR-Cas9 and standard Cre/loxP approaches. Experiments with competitive bone marrow (BM) chimeras identified a positive role for Dnmt3a in Tfh differentiation, which was validated by comparing mice with Dnmt3a mutations in CD4+ cells to animals with WT Dnmt3a. In conclusion, We identify that Dnmt3a mutations limit normal and autoreactive Tfh differentiation. Key findings- Dnmt3a mutations limit Tfh differentiation, which could contribute to reduced immune responses in individuals with somatic DNMT3A mutations. - Deep characterization of the KRN+ splenocyte transfer model defines a dynamic process leading to reproducible autoimmune joint inflammation. - The immuno-CRISPR (iCR) methodology can be used to test the role of candidate genes in disease models.

immunology↗

bioRxiv2022

MDM2 inhibitors, nutlin-3a and navtemadelin, retain efficacy in human and mouse cancer cells cultured in hypoxia

Activation of p53 by small molecule MDM2 inhibitors can induce cell cycle arrest or death in p53 wildtype cancer cells. However, cancer cells exposed to hypoxia can develop resistance to other small molecules, such as chemotherapies, that activate p53. Here, we evaluated whether hypoxia could render cancer cells insensitive to two MDM2 inhibitors with different potencies, nutlin-3a and navtemadlin. Inhibitor efficacy and potency were evaluated under short-term hypoxic conditions in human and mouse cancer cells expressing different p53 genotypes (wild-type, mutant, or null). Treatment of wild-type p53 cancer cells with MDM2 inhibitors reduced cell growth by >75% in hypoxia through activation of the p53-p21 signaling pathway; no inhibitor-induced growth reduction was observed in hypoxic mutant or null p53 cells except at very high concentrations. The concentration of inhibitors needed to induce the maximal p53 response was not significantly different in hypoxia compared to normoxia. However, inhibitor efficacy varied by species and by cell line, with stronger effects at lower concentrations observed in human cell lines than in mouse cell lines grown as 2D and 3D cultures. Together, these results indicate that MDM2 inhibitors retain efficacy in hypoxia, suggesting they could be useful for targeting acutely hypoxic cancer cells.

cancer biology↗

bioRxiv2022

Divergent and dominant anti-inflammatory effects of patient-derived anti-citrullinated protein antibodies (ACPA) in arthritis development

About two thirds of rheumatoid arthritis (RA) patients develop anti-citrullinated protein antibodies (ACPA), which are a characteristic of the disease and sparsely observed in other clinical settings or in the healthy population. ACPA are used as a diagnostic criterion and often develop prior to diagnosis. Therefore, ACPA titers are important in the identification of individuals at risk of developing RA. Interestingly, the titers and target cross-reactivity of ACPA increase by time of diagnosis, suggesting a causality between anti-citrulline reactivity and the development of RA. However, only 50% of ACPA-positive at-risk individuals will progress to a clinical RA diagnosis. This observation suggests that there are different types of ACPA and that their relationship with RA development is more complex than presently assumed. To address the biological effect of ACPA in the establishment and development of inflammatory arthritis, we made use of the collagen antibody-induced arthritis (CAIA) model of passive arthritis. Using seven unique patient derived monoclonal ACPA, we observed that ACPA are predominantly anti-inflammatory, with some clones (C03 and BVCA1) completely inhibiting disease development. We have also identified a clone (C04) as having a disease-prone effect, by increasing and sustaining disease prevalence. This clone had previously been reported as being pro-inflammatory in a different model of joint inflammation. The anti-inflammatory effects of C03 were dependent on Fc{gamma}R, since neither F(ab)2 or a mutated Fc-null GRLR-C03 clone could mediate disease protection. Most importantly, mice receiving polyclonal ACPA enriched from RA sera were also partially protected from disease. Looking at the cell and tissue of origin of the ACPA clones, their V(D)J sequences, Fc glycosylation pattern and fine-specificities, we could not identify a common feature between anti-inflammatory ACPA distinguishing them from those without the properties. Together, our data demonstrates that circulating ACPA in RA patients are predominantly anti-inflammatory, emphasizing the need to study ACPA repertoires in order to determine their influence on the clinical outcome of the patient or at-risk individual.

immunology↗

bioRxiv2021

CRISPR enriches for cells with mutations in a p53-related interactome, and this can be inhibited.

CRISPR/Cas9 can be used to inactivate or modify genes by inducing double-stranded DNA breaks1-3. As a protective cellular response, DNA breaks result in p53-mediated cell cycle arrest and activation of cell death programs4,5. Inactivating p53 mutations are the most commonly found genetic alterations in cancer, highlighting the important role of the gene6-8. Here, we show that cells deficient in p53, as well as in genes of a core CRISPR-p53 tumor suppressor interactome, are enriched in a cell population when CRISPR is applied. Such enrichment could pose a challenge for clinical CRISPR use. Importantly, we identify that transient p53 inhibition suppresses the enrichment of cells with these mutations. Furthermore, in a data set of >800 human cancer cell lines, we identify parameters influencing the enrichment of p53 mutated cells, including strong baseline CDKN1A expression as a predictor for an active CRISPR-p53 axis. Taken together, our data identify strategies enabling safe CRISPR use.

cancer biology↗

bioRxiv2021

A rapid CRISPR competitive assay for in vitro and in vivo discovery of potential drug targets affecting the hematopoietic system

CRISPR/Cas9 can be used as an experimental tool to inactivate genes in cells. However, a CRISPR-targeted cell population will not show a uniform genotype of the targeted gene. Instead, a mix of genotypes is generated - from wild type to different forms of insertions and deletions. Such mixed genotypes complicate analyzing the role of the targeted gene in the studied cell population. Here, we present a rapid experimental approach to functionally analyze a CRISPR-targeted cell population that does not involve generating clonal cell lines. As a simple readout, we leverage the CRISPR-induced genetic heterogeneity and use sequencing to identify how different genotypes are enriched or depleted related to the studied cellular behavior or phenotype. The approach uses standard PCR, Sanger sequencing, and a simple sequence deconvoluting software, enabling laboratories without specific in-depth knowledge to also perform these experiments. As proof of principle, we present examples studying the role of different genes for various aspects related to hematopoietic cells (T cell development in vivo and activation in vitro, macrophage phagocytosis, and a leukemia-like phenotype induced by overexpressing a proto-oncogene). In conclusion, we present a rapid experimental approach to identify potential drug targets related to mature immune cells, as well as normal and malignant hematopoiesis. Highlights- CRISPR generates genetic heterogeneity at the targeted site. - Genetic heterogeneity complicates identifying the role of a targeted gene. - Heterogeneity can be quantified by Sanger sequencing with sufficient sensitivity. - Enrichment of specific genotypes can be used to identify roles for targeted genes. - Competitive experiments show the potential of genotype enrichment as a discovery tool. Graphical representation O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=155 SRC="FIGDIR/small/434360v1_ufig1.gif" ALT="Figure 1"> View larger version (36K): org.highwire.dtl.DTLVardef@185bff7org.highwire.dtl.DTLVardef@1fe12bcorg.highwire.dtl.DTLVardef@1800f4eorg.highwire.dtl.DTLVardef@6d020b_HPS_FORMAT_FIGEXP M_FIG C_FIG

immunology↗

bioRxiv2020

Identifying novel B-cell targets for chronic inflammatory autoimmune disease by screening of chemical probes in a patient-derived cell assay

B-cell secretion of autoantibodies drives autoimmune diseases, including systemic lupus erythematosus and idiopathic inflammatory myositis. Few therapies are presently available for treatment of these patients, often resulting in unsatisfactory effects and helping only some of patients. We developed a screening assay for evaluation of novel targets suspending B-cell maturation into antibody secreting cells, which could contribute to future drug development. The assay was employed for testing 43 high quality chemical probes and compounds inhibiting under-explored protein targets, using primary cells from patients with autoimmune disease. Probes inhibiting bromodomain family proteins and histone methyl transferases demonstrated abrogation of B-cell functions to a degree comparable to a positive control, the JAK inhibitor tofacitinib. Inhibition of each target rendered a specific functional cell and potential disease modifying effect, indicating specific epigenetic protein targets as potential new intervention points for future drug discovery and development efforts.

immunology↗