Biology subjects

Koschmieder, S.

Publications and source records attributed to Koschmieder, S..

11 recordsLinked to original sources

bioRxiv2026

Nanoparticle size governs engagement with myeloid cells and hitchhiking to hematopoietic organs in myeloproliferative neoplasms

Nanoparticle design principles contribute to desirable in vivo performance, including prolonged circulation, desirable biodistribution profiles, and tuned interactions with the immune system. The importance of these variables, although well-established in solid tumors, remains elusive in the context of hematological malignancies. Here, we investigated the influence of liposome size on biodistribution and cellular uptake within hematopoietic compartments - bone marrow (BM) and spleen - in JAK2V617F myeloproliferative neoplasms (MPN). A milli-fluidic manufacturing platform combined with a Design-of-Experiments (DoE) approach was used to generate small, medium, and large liposomes. Liposome uptake was assessed ex vivo using blood samples from healthy donors and MPN patients, followed by in vivo biodistribution studies in a transgenic JAK2V617F MPN mouse model. Organ-level accumulation was quantified using hybrid fluorescence / computed tomography (FLT/CT) imaging, while cellular uptake was evaluated via flow cytometry. Whole-body imaging revealed that increasing liposome size enhanced delivery to both the spleen and BM, with larger liposomes exhibiting the highest accumulation in both organs. Cellular analysis corroborated the in vivo observations, demonstrating that large liposomes are taken up to a greater extent by monocytes and granulocytes, in both spleen and BM. At late time-points post-injection, the liposomal accumulation was twice as high in the BM in comparison to the spleen and peripheral blood, highlighting the progressively elevated accumulation and retention in the BM, as opposed to the elimination phase nanoparticles undergo in clearance organs and circulation at these time-points. Of note, among BM mature myeloid cells, neutrophils were associated with a higher nanoparticle uptake than monocytes, highlighting their capability to phagocytose material in circulation and hitchhike it to malignant or inflamed regions. These findings demonstrate that continuous flow manufacturing procedures can swiftly produce nanoparticles with desirable characteristics that are essential for tuning the biodistribution towards hematopoietic organs and myeloid immune cells. By providing mechanistic insights into nanoparticle behavior within hematopoietic compartments, this work advances our understanding of nanomedicine in vivo performance and highlights particle size as a critical quality attribute that can be optimized and controlled to improve targeted delivery to myeloid cells in the treatment of hematological malignancies.

cancer biology↗

bioRxiv2026

PDE3A-SLFN12 Molecular Glues Target Multiple KIT D816V Cell Types in Preclinical Models of Mast Cell Malignancies

A drug discovery approach was used to specifically target malignant cells with KIT D816V mutation, which is the predominant disease-causing mutation in clonal mast cell malignancies. To this end, KIT D816V cells derived from induced pluripotent stem cells (iPS cells) of KIT D816V patients were employed to screen a library of FDA approved and experimental drugs for specific killing of KIT D816V cells. We discovered the novel compound LDC 3416, which targets multiple malignant KIT D816V cell types, including hematopoietic stem/progenitor cells and mast cells. Importantly, by exploring the LDC 3416 targeting profile, we identified the phosphodiesterase 3A-Schlafen 12 (PDE3A-SLFN12) molecular glue pathway as a novel approach for specific targeting of malignant KIT D816V cells. We found that the KIT D816V mutant protein leads to increased expression of PDE3A and SLFN12 and thus confers a selective molecular vulnerability to PDE3A-SLFN12 molecular glues. Primary malignant mast cells of KIT D816V patients with indolent and advanced systemic mastocytosis also exhibit increased expression of PDE3A and SLFN12. We extended our study to include additional PDE3A-SLFN12 molecular glues and demonstrate their synergistic action with KIT D816V selective tyrosine kinase inhibitors (TKIs) in killing KIT D816V cells. Furthermore, the PDE3A-SLFN12 molecular glues also target KIT D816V megakaryocytes, a cell type that has been underestimated in malignant mast cell pathophysiology and molecular targeting. The identified molecular glues, along with their synergy with TKIs and their simultaneous targeting of multiple KIT D816V cell types, open novel treatment options for KIT D816V mast cell malignancies and other KIT D816V associated diseases.

cancer biology↗

bioRxiv2026

Oncodevelopmental plasticity of the skeleton in myeloid neoplasms

Myelofibrosis in patients with myeloproliferative neoplasms (MPNs) is traditionally characterized by bone marrow fibrosis and osteosclerosis, with de novo bone formation commonly attributed to impaired osteoclast-mediated resorption. Here, we challenge this paradigm by demonstrating that a solitary clonal driver mutation simultaneously induces pathological bone formation and resorption, with osteosclerosis acting to conceal localized and active bone destruction rather than inhibiting it. Through population analysis; clinical imaging; patient-derived multi-tissue sequencing; murine models and organ-on-a-chip systems, we demonstrate that spatial and ontogeny-dependent remodeling in mesoderm- and neural crest-derived bones is mechanistically interconnected via a previously unidentified osteochondral stromal injury program. Neural crest-derived stromal cells suppress osteogenic programs and undergo injury-induced lineage plasticity with ectopic chondrogenesis, mirroring pathological remodeling in mesoderm-derived growth plate regions. This shared injury response promotes osteoclastogenesis and is mediated by a conserved Thrombospondin 1+ (THBS1+) stromal population that links fibrotic remodeling to bone loss. Combined pharmacological inhibition of THBS1 and JAK signaling reduces myeloproliferation, halts fibrosis progression, and restores two developmentally distinct bones, establishing THBS1 as a unifying therapeutic target in myelofibrosis.

cancer biology↗

bioRxiv2025

The V617F mutation in JAK2 renders myeloid cells more sensitive to IL-6-mediated gp130 signaling

The somatic V617F mutation in the pseudokinase domain of JAK2 (JAK2VF) causes various phenotypes of myeloproliferative neoplasms (MPN). By interacting with cytokine receptors such as those for erythropoietin (EPO) or thrombopoietin (TPO), JAK2VF induces ligand-independent dimerization and activation, leading to deregulated blood cell production, cytokine hypersensitivity, and inflammatory cytokine release. Interleukin-6 (IL-6), a key mediator of inflammatory symptoms in MPN, signals via homodimers of the gp130 receptor. We investigated whether JAK2VF alters gp130 dimerization and IL-6 sensitivity. Molecular dynamics simulations demonstrated that the JAK2VF pseudokinase domain forms more stable dimers than wild-type (WT) JAK2, potentially supporting gp130 tetramerization. In cell-based assays, IL-6 stimulation of JAK2VF+ cells induced stronger STAT3 activation than in JAK2-WT cells, reflecting enhanced IL-6 sensitivity. Moreover, JAK2VF expression elevated gp130 surface levels, dependent on the JAK2-binding motif in gp130. These findings indicate that JAK2VF promotes gp130 expression and dimerization, sensitizing mutant cells to IL-6. Thus, JAK2VF-driven amplification of IL-6/gp130 signaling may foster chronic inflammation and disease progression in MPN, representing a potential therapeutic target.

cancer biology↗

bioRxiv2024

IFNα Inhibits JAK2V617F-Driven Neoplastic Vasculogenesis and Endothelial-to-Mesenchymal Transition in a 3D iPSC-Based Bone Marrow Niche Model

The vascular niche is a critical regulator of hematopoiesis and disease progression in myeloproliferative neoplasms (MPN). The presence of JAK2V617F+ endothelial cells (EC) in MPN patients and their association with cardiovascular complications highlight the need to understand and therapeutically target this compartment. Using patient-specific induced pluripotent stem cells (iPSC) harboring JAK2WT or the MPN-driver JAK2V617F (heterozygous, JAK2V617FHET, or homozygous, JAK2V617FHOM), we identified zygosity-dependent transcriptional profiles in iPSC-derived EC (iEC) at baseline and following interferon-alpha (IFN) treatment. JAK2V617FHET iEC exhibited an endothelial-to-mesenchymal transition (EndMT) signature, while JAK2V617FHOM iEC showed suppression of translation and ribosome biogenesis. Leveraging iPSC-based 3D assembloids that mimic the bone marrow (BM) niche, we showed that JAK2V617F-driven EndMT is inhibited by tyrosine kinase inhibitors and IFN. In both JAK2V617F-driven polycythemia vera and TPO-driven myelofibrosis murine models, scRNA-seq analysis of the BM vascular niche consistently revealed inflammatory and EndMT-associated signatures in arterial and arteriolar EC. Notably, dysregulation of ribosome- and translation-related pathways emerged in the myelofibrosis model and at advanced disease stages in JAK2V617F-driven polycythemia vera, indicating progressive vascular remodeling with disease evolution. Chronic pegylated IFN treatment in vivo effectively reversed these pathological changes. IFNs anti-EndMT activity was further validated in BM biopsies from MPN patients undergoing IFN therapy. This is the first study to define MPN stage-dependent vascular remodeling and zygosity-specific endothelial effects of JAK2V617F, and to directly link IFN-mediated EndMT inhibition as a novel antifibrotic mechanism. Our 3D assembloids provide a translational platform for mechanistic studies and therapeutic targeting of the BM microenvironment in MPN. Bullet PointsO_LIArterial vascular remodeling emerges as a novel hallmark of MPN, characterized by TNF-inflammation, ribosomal dysregulation and EndMT. C_LIO_LIIFN restores neoplastic endothelial dysfunction, highlighting its role as a vascular niche-modulating and anti-fibrotic agent in MPN. C_LI

cancer biology↗

bioRxiv2024

SIngle cell level Genotyping Using scRna Data (SIGURD)

MotivationBy accounting for variants within measured transcripts, it is possible to evaluate the status of somatic variants using single-cell RNA-sequencing (scRNA-seq) and to characterize their clonality. However, the sparsity (very few reads per transcript) or bias in protocols (favoring 3 ends of the transcripts) makes the chance of capturing somatic variants very unlikely. This can be overcome by targeted sequencing or the use of mitochondrial variants as natural barcodes for clone identification. Currently, available computational tools focus on genotyping, but do not provide functionality for combined analysis of somatic and mitochondrial variants and functional analysis such as characterization of gene expression changes in detected clones. ResultsHere, we propose SIGURD, which is an R-based pipeline for the clonal analysis of single-cell RNA-seq data. This allows the quantification of clones by leveraging both somatic and mitochondrial variants. SIGURD also allows for functional analysis after clonal detection: association of clones with cell populations, detection of differentially expressed genes across clones and association of somatic and mitochondrial variants. Here, we demonstrate the power of SIGURD by analyzing single-cell data of colony-forming cells derived from patients with myeloproliferative neoplasms. AvailabilityCode and tutorial of SIGURD are available at GitHub https://github.com/CostaLab/sigurd. Contactivan.costa@rwth-aachen.de, skoschmieder@ukaachen.de Supplementary InformationSupplementary data are available online.

bioinformatics↗

bioRxiv2024

NLRP3-induced systemic inflammation controls the development of JAK2V617F mutant myeloproliferative neoplasms

The development of Philadelphia chromosome-negative classical myeloproliferative neoplasms (MPN) involves an inflammatory process that facilitates outgrowth of the malignant clone and correlates with clinical outcome measures. This raises the question to which extent inflammatory circuits in MPN depend on activation of innate immune sensors. Here, we investigated whether NLRP3, which precipitates inflammasome assembly upon detection of cellular stress, drives murine JAK2V617F mutant MPN. Deletion of Nlrp3 within the hematopoietic compartment completely prevented increased IL-1{beta} and IL-18 release in MPN. NLRP3 in JAK2V617F hematopoietic cells, but not in JAK2 wild type radioresistant cells, promoted excessive platelet production via stimulation of the direct thrombopoiesis differentiation pathway, as well as granulocytosis. It also promoted expansion of the hematopoietic stem and progenitor cell compartment despite inducing pyroptosis at the same time. Importantly, NLRP3 inflammasome activation enhanced bone marrow fibrosis and splenomegaly. Pharmacological blockade of NLRP3 in fully established disease led to regression of thrombocytosis and splenomegaly. These findings suggest that NLRP3 is critical for MPN development and its inhibition represents a new therapeutic intervention for MPN patients. Key pointsO_LIThe increased IL-1{beta} and IL-18 release in JAK2V617F mutant MPN depends on NLRP3 inflammasome activation C_LIO_LINLRP3 in MPN promotes excess platelet production, granulocytosis, HSPC compartment expansion, splenomegaly and bone marrow fibrosis C_LI

cancer biology↗

bioRxiv2021

CALR frameshift mutations in MPN patient-derived iPS cells accelerate maturation of megakaryocytes

Calreticulin (CALR) mutations are driver mutations in myeloproliferative neoplasms (MPNs), leading to activation of the thrombopoietin receptor, and causing abnormal megakaryopoiesis. Here, we generated patient-derived CALRins5- or CALRdel52-positive induced pluripotent stem (iPS) cells to establish a MPN disease model for molecular and mechanistic studies. We demonstrated myeloperoxidase deficiency in CD15+ granulocytic cells derived from homozygous CALR-mutant iPS cells, rescued by repairing the mutation using CRISPR/Cas9. iPS cell-derived megakaryocytes showed characteristics of primary megakaryocytes such as formation of demarcation membrane system and cytoplasmic pro-platelets protrusions. Importantly, CALR mutations led to enhanced megakaryopoiesis and accelerated megakaryocytic development in a thrombopoietin-independent manner. Mechanistically, our study identified differentially regulated pathways in mutated vs. unmutated megakaryocytes, such as hypoxia signaling, which represents a potential target for therapeutic intervention. Altogether, we demonstrate key aspects of mutated CALR-driven pathogenesis, dependent on its zygosity and found known and novel therapeutic targets, making our model a valuable tool for clinical drug screening in MPNs.

cancer biology↗

bioRxiv2021

Human DC3 Antigen Presenting Dendritic Cells from Induced Pluripotent Stem Cells

Dendritic cells (DC) are professional antigen-presenting cells that develop from hematopoietic stem cells. Different DC subsets exist based on ontogeny, location and function, including the recently identified proinflammatory DC3 subset. DC3 have the prominent activity to polarize CD8+ T cells into CD8+ CD103+ tissue resident T cells. Here we describe human DC3 differentiated from induced pluripotent stem cells (iPS cells). iPS cell-derived DC3 have the gene expression and surface marker make-up of blood DC3 and polarize CD8+ T cells into CD8+ CD103+ tissue-resident memory T cells in vitro. To test the impact of malignant JAK2 V617F mutation on DC3, we differentiated patient-specific iPS cells with JAK2 V617Fhet and JAK2 V617Fhom mutations into JAK2 V617Fhet and JAK2 V617Fhom DC3. The JAK2 V617F mutation enhanced DC3 production and caused a bias towards erythrocytes and megakaryocytes. The patient-specific iPS cell-derived DC3 are expected to allow studying DC3 in human diseases and developing novel therapeutics. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=69 SRC="FIGDIR/small/449451v1_ufig1.gif" ALT="Figure 1"> View larger version (18K): org.highwire.dtl.DTLVardef@9e4ee3org.highwire.dtl.DTLVardef@14d3b40org.highwire.dtl.DTLVardef@16cfae8org.highwire.dtl.DTLVardef@1ccb5e0_HPS_FORMAT_FIGEXP M_FIG C_FIG

immunology↗

bioRxiv2020

Nintedanib Targets KIT D816V Neoplastic Cells Derived from Induced Pluripotent Stem cells of Systemic Mastocytosis

The KIT D816V mutation is found in more than 80% of patients with systemic mastocytosis (SM) and is key to neoplastic mast cell (MC) expansion and accumulation in affected organs. KIT D816V therefore represents a prime therapeutic target for SM. Here we generated a panel of patient-specific KIT D816V induced pluripotent stem cells (iPSCs) from patients with aggressive SM (ASM) and mast cell leukemia (MCL) to develop a patient-specific SM disease model for mechanistic and drug discovery studies. KIT D816V iPSCs differentiated into neoplastic hematopoietic progenitor cells and MCs with patient-specific phenotypic features, thereby reflecting the heterogeneity of the disease. CRISPR/Cas9n-engineered KIT D816V human embryonic stem cells (ESCs), when differentiated into hematopoietic cells, recapitulated the phenotype observed for KIT D816V iPSC hematopoiesis. KIT D816V causes constitutive activation of the KIT tyrosine kinase receptor and we exploited our iPSCs and ESCs to investigate new tyrosine kinase inhibitors targeting KIT D816V. Our study identified nintedanib as a novel KIT D816V inhibitor. Nintedanib selectively reduced the viability of iPSC-derived KIT D816V hematopoietic progenitor cells and MCs in the nanomolar range. Nintedanib was also active on primary samples of KIT D816V SM patients. Molecular docking studies show that nintedanib binds to the ATP binding pocket of inactive KIT D816V. Our results suggest nintedanib as a new drug candidate for KIT D816V targeted therapy of advanced SM.

cancer biology↗

bioRxiv2017

Leukocyte counts based on site-specific DNA methylation analysis

The composition of white blood cells is usually assessed by histomorphological parameters or flow cytometric measurements. Alternatively, leukocyte differential counts (LDCs) can be estimated by deconvolution algorithms for genome-wide DNA methylation (DNAm) profiles. We identified cell-type specific CG dinucleotides (CpGs) that facilitate relative quantification of leukocyte subsets. Site-specific analysis of DNAm levels by pyrosequencing provides similar precision of LDCs as conventional methods, whereas it is also applicable to frozen samples and requires only very small volumes of blood. Furthermore, we describe a new approach for absolute quantification of cell numbers based on a non-methylated reference DNA. Our \"Epi-Blood-Count\" facilitates robust and cost effective analysis of blood counts for clinical application.

molecular biology↗