Biology subjects

Lux, A.

Publications and source records attributed to Lux, A..

10 recordsLinked to original sources

bioRxiv2026

Targeting neutrophil signaling networks in immune complex-mediated autoimmune disease

Fragment crystallizable gamma receptor (Fc{gamma}R)-induced signaling is a crucial process that determines the cellular response to immune complexes (IC) in autoimmune diseases. In several diseases including pemphigoid diseases (PD), such as epidermolysis bullosa acquisita (EBA), or rheumatoid arthritis (RA), neutrophils are prominently involved as effector cells, while others such as immune thrombocytopenia (ITP) are independent of neutrophils. At the same time, most diseases are commonly treated by broad-range immunosuppression accompanied by severe risk for adverse effects. Signal transduction inhibitors (STIs) have been successfully applied in cancer therapy. However, their use in autoimmune diseases is an emerging, but so far understudied potential treatment avenue. Therefore, we screened a target-selective compound library consisting of 155 STIs in a neutrophil-based assay and conducted a multiplex kinase activity profiling with IC-stimulated neutrophils. Thus, we found novel potential therapeutic targets that were validated both in vitro in functional neutrophil assays and in vivo in murine models of EBA Here, we demonstrate that both systemic and topical treatment with several individual STIs is effective in a prophylactic approach in these models. Furthermore, therapeutic treatment with the BTK inhibitor ibrutinib in the immunization-induced EBA model reduced disease severity by approximately 85 % and showed efficacy in additional experimental models of EBA, arthritis, and ITP. Together, the present study contributes to the elucidation of Fc{gamma}R-dependent signaling in neutrophils and identifies multiple novel promising treatment options including inhibition of PLC, PDK-1, PKC, p38, DNA-PK, KSP, c-Met, TBK-1 and BTK for IC-mediated autoimmune diseases.

immunology↗

bioRxiv2026

A human immune system mouse model for preclinical evaluation of therapies in pemphigoid disease

Pemphigoid diseases (PD) including Epidermolysis bullosa acquisita (EBA) are rare immunoglobulin G (IgG)-driven autoimmune skin blistering diseases with limited therapeutic options. Mechanistically, chronic inflammation in the skin leads to disruption of the dermal-epidermal junction (DEJ) with a crucial contribution of Fc{gamma}R-mediated activation of myeloid immune cells such as neutrophils. Thus, targeting of kinases involved in Fc{gamma}R-dependent activation of myeloid immune cells holds great promise as a therapeutic strategy. In this study, we employ human immune system (HIS) mice featuring all major human leukocytes which allows to investigate the impact of therapeutics on human immune cells in vivo. In a passive transfer approach, repetitive application of collagen VII (COL7c) specific IgG was associated with skin inflammation including infiltration of activated human immune cells and thickening of the epidermis. While application of recombinant human G-CSF boosted myeloid cell maturation and thus disease severity, treatment with Fc{gamma}R-blocking antibodies impaired disease development confirming the crucial role of human cells. Finally, small molecule PDK1 inhibitor BX-795 abrogated development of skin inflammation associated with reduced leukocyte infiltration and activation supporting the role of PDK1 in Fc{gamma}R-driven immune cell activation. This study establishes the first in vivo model of EBA in HIS mice and reveals its suitability for pre-clinical screening and evaluation of therapeutic agents. Importantly, it highlights the potential of kinase inhibition for treatment of EBA.

immunology↗

bioRxiv2025

CDK7, CDK9, or CDK11 Inhibition Reduces Neutrophil-Driven Inflammation and Tissue Damage in Experimental Autoimmune Models

BackgroundCyclin-dependent kinases (CDKs) are involved in basic cellular processes like regulation of cell-cycle progression and transcription. However, recent data also indicate a specific role in terminally differentiated neutrophils by promoting reactive oxygen species (ROS) release, degranulation, neutrophil extracellular trap formation, or apoptosis. Since these mechanisms are implicated in multiple autoimmune diseases, we aimed to delineate the role of CDKs in IC-mediated autoimmune diseases both in vitro and in vivo. MethodsWe analyzed CDK gene expression in unstimulated and immune complex (IC)-stimulated neutrophils. Subsequently, we investigated the effect of pharmacological CDK inhibition on IC-activated neutrophil functions. To analyze the inhibitors in a more translational approach, we proceeded with the systemic and topical application of the effective inhibitors in a murine antibody transfer-induced local epidermolysis bullosa acquisita (EBA) model. The most efficient inhibitor, MC180295, was validated in two other IC-mediated models of autoimmune disease: Serum-transfer arthritis (STA), which also, but not exclusively, depends on neutrophils and immune thrombocytopenia (ITP), which is considered less neutrophil-dependent. ResultsWe found 14 CDKs expressed in unstimulated cells, while the IC-stimulation showed an upregulation of CDK2 and CDK4 expression. Inhibitors selectively targeting CDK1, CDK2, CDK4/6, CDK7, CDK9, CDK11, and CDK12 showed effects on different neutrophil functions (surface activation marker expression, ROS release, adhesion, apoptosis) in vitro. In the predominantly neutrophil-driven EBA model, we observed a reduction of disease severity upon treatment with CDK7, CDK9, or CDK11 inhibitors. Inhibiting these CDKs with topical THZ2, MC180295, or OTS964, respectively, also improved the clinical phenotype. In line with our hypothesis, MC180295 impaired the development of STA, but not ITP. ConclusionsCDK7, CDK11, and especially CDK9 inhibition show therapeutic potential in IC-driven neutrophil-mediated diseases such as rheumatoid arthritis and EBA.

immunology↗

bioRxiv2025

ERK5 is required for neutrophil-mediated ROS release and essential in epidermolysis bullosa acquisita

Neutrophils are key effector cells in antibody-mediated autoimmune diseases, contributing to inflammation via the release of reactive oxygen species (ROS). Extracellular signal-regulated kinase 5 (ERK5), a member of the MAPK family, is expressed in neutrophils but its role in autoimmune disease pathogenesis remains unclear. We investigated the functional relevance of ERK5 in antibody-mediated autoimmune diseases by comparing neutrophil-dependent (epidermolysis bullosa acquisita, EBA; serum transfer arthritis, STA) and neutrophil-independent (immune thrombocytopenia, ITP) murine models using the small-molecule ERK5 inhibitor XMD8-92. In vitro, pharmacological ERK5 inhibition specifically reduced neutrophil ROS release and CD62L shedding without affecting adhesion, chemotaxis, or CD18 expression. No major effects on viability were observed. In vivo, ERK5 inhibition with XMD8-92 significantly ameliorated antibody transfer-induced EBA, supporting a critical role of neutrophil-derived ROS in disease pathogenesis. In STA and ITP, ERK5 inhibition did not affect clinical outcomes. Together, these findings highlight ERK5 as a regulator of neutrophil ROS release and a potential therapeutic target in ROS-driven autoimmune diseases such as EBA.

immunology↗

bioRxiv2025

A novel Fc-optimized antibody drug conjugate targeting CD7 for the therapy of T-cell acute lymphoblastic leukemia

While treatment for patients with T-cell acute lymphoblastic leukemia (T-ALL) has improved in the last decades, therapeutic options for patients refractory to standard therapy or with relapsing disease are limited. In particular, no immunotherapy option has been approved in T-ALL yet. Here, a novel dual antibody engineering approach for targeting CD7 was evaluated. The chimeric CD7 antibody chimTH69 was modified by Fc engineering to improve antibody-dependent cell-mediated cytotoxicity (ADCC) and antibody-dependent cellular phagocytosis (ADCP). In addition, it was conjugated to monomethyl auristatin E (MMAE), a microtubule-disrupting agent. The resulting Fc-optimized antibody-drug conjugate (ADC), designated chimTH69-DE-vcMMAE, showed a unique set of effector functions in vitro. It triggered ADCC by mononuclear cells at picomolar concentrations, mediated ADCP by macrophages and directly inhibited the growth of a panel of T-ALL cell lines by delivering the cytotoxic compound to induce G2 cell cycle arrest and apoptosis. In addition, due to its specific linker design, chimTH69-DE-vcMMAE demonstrated bystander killing activity against CD7-negative leukemia cells. In mice, CD7-directed therapy with chimTH69-DE-vcMMAE inhibited the growth of subcutaneous CCRF-CEM T-ALL xenografts. Moreover, chimTH69-DE-vcMMAE exerted strong antileukemic effects in a phase II-like patient-derived xenograft preclinical trial in pediatric and adult patients when applied in an experimental overt leukemia setting. ChimTH69-DE-vcMMAE induced minimal residual disease-negativity in one PDX model. These findings indicate that targeting CD7 with the novel Fc-optimized ADC is a potent strategy to trigger anti-leukemia responses and may open a novel therapeutic avenue for T-ALL treatment. Key PointA novel antibody drug conjugate targeting CD7 showed efficient anti-leukemia activity in preclinical models of T-ALL.

cancer biology↗

bioRxiv2023

Role of lipid nanodomains for inhibitory FcγRIIb function

The inhibitory Fc{gamma} receptor Fc{gamma}RIIb is involved in immune regulation and is known to localize to specific regions of the plasma membrane called lipid rafts. Previous studies suggested a link between the altered lateral receptor localization within the plasma membrane and the functional impairment of the Fc{gamma}RIIb-I232T variant that is associated with systemic lupus erythematosus. Here, we conducted microsecond all-atom molecular dynamics simulations and IgG binding assays to investigate the lipid nano-environment of Fc{gamma}RIIb monomers and of the Fc{gamma}RIIb-I232T mutant within a plasma membrane model, the orientation of the Fc{gamma}RIIb ectodomain, and its accessibility to IgG ligands. In contrast to previously proposed models, our simulations indicated that Fc{gamma}RIIb does not favor a cholesterol-or a sphingolipid-enriched lipid environment. Interestingly, cholesterol was depleted for all studied Fc{gamma}RIIb variants within a 2-3 nm environment of the receptor, counteracting the usage of raft terminology for models on receptor functionality. Instead, the receptor interacts with lipids that have poly-unsaturated fatty acyl chains and with (poly-) anionic lipids within the cytosolic membrane leaflet. We also found that Fc{gamma}RIIb monomers adopt a conformation that is not suitable for binding to its IgG ligand, consistent with a lack of detectable binding of monomeric IgG in experiments on primary immune cells. However, our results propose that multivalent IgG complexes might stabilize Fc{gamma}RIIb in a binding-competent conformation. We suggest differences in receptor complex formation within the membrane as a plausible cause of the altered membrane localization or clustering and the altered suppressive function of the Fc{gamma}RIIb-I232T variant. Significance StatementOur study sheds new light on the molecular mechanisms underlying the regulation of immune signaling mediated by the inhibitory Fc{gamma} receptor (Fc{gamma}RIIb). By utilizing atomistic simulations and experimental assays, we demonstrate that Fc{gamma}RIIb interacts with specific lipids in the plasma membrane. Notably, our findings challenge the current view of membrane heterogeneity in immune cells, as Fc{gamma}RIIb is not localized in specialized membrane domains known as rafts. Rather, we propose that receptor complex formation modulates receptor localization and conformation, thereby enabling ligand binding. Our findings have important implications for understanding how immune receptors function and communicate with each other, and may provide new opportunities for developing therapeutic strategies targeting Fc{gamma}RIIb in diseases such as autoimmunity and cancer.

immunology↗

bioRxiv2023

Immune cells employ traction forces to overcome steric hindrance in 3D biopolymer networks

To reach targets outside the bloodstream, immune cells can extravasate and migrate through connective tissue. During tissue infiltration, immune cells migrate in an amoeboid fashion, characterized by weak matrix adhesions and low traction forces, that allows them to achieve high migration speeds of up to 10 {micro}m/min. How immune cells reconcile amoeboid migration with the need to overcome steric hindrance in dense matrices is currently not understood. Here we show that NK92 (natural killer) cells can switch from their default amoeboid migration mode to a contractile, mesenchymal-like migration mode when moving through fibrous human amniotic membrane (HAM) tissue. We subsequently study immune cell migration in reconstituted 3D collagen networks with known mechanical properties and pore sizes and apply time-lapse confocal reflection microscopy to obtain simultaneous measurements of migration speed, directional persistence, and cell contractility. We find that NK92 cells are highly mechanoresponsive and exert substantial acto-myosin driven, integrin-mediated contractile forces of up to 100 nN on the extracellular matrix during short contractile phases. This burst-like contractile behavior is also found in primary B, T, NK cells, neutrophils, and monocytes, and is tightly related to the fraction of cells that appear to become stuck in narrow pores of the surrounding matrix. Our results demonstrate that steric hindrance guides the rapid regulation of integrin-mediated adhesion to the ECM in a large number of immune cell subtypes.

biophysics↗

bioRxiv2023

Mixed IgG Fc immune complexes exhibit blended binding profiles and refine FcR affinity estimates

Immunoglobulin (Ig)G antibodies coordinate immune effector responses by selectively binding to target antigens and then interacting with various effector cells via the Fc{gamma} receptors. The Fc domain of IgG can promote or inhibit distinct effector responses across several different immune cell types through variation based on subclass and Fc domain glycosylation. Extensive characterization of these interactions has revealed how the inclusion of certain Fc subclasses or glycans results in distinct immune responses. During an immune response, however, IgG is produced with mixtures of Fc domain properties, so antigen-IgG immune complexes are likely to almost always be comprised of a combination of Fc forms. Whether and how this mixed composition influences immune effector responses has not been examined. Here, we measured Fc{gamma} receptor binding to immune complexes of mixed Fc domain composition. We found that the binding properties of the mixed-composition immune complexes fell along a continuum between those of the corresponding pure cases. Binding quantitatively matched a mechanistic binding model, except for several low-affinity interactions mostly involving IgG2. We found that the affinities of these interactions are different than previously reported, and that the binding model could be used to provide refined estimates of these affinities. Finally, we demonstrated that the binding model can predict effector-cell elicited platelet depletion in humanized mice, with the model inferring the relevant effector cell populations. Contrary to the previous view in which IgG2 poorly engages with effector populations, we observe appreciable binding through avidity, but insufficient amounts to observe immune effector responses. Overall, this work demonstrates a quantitative framework for reasoning about effector response regulation arising from IgG of mixed Fc composition. Summary pointsO_LIThe binding behavior of mixed Fc immune complexes is a blend of the binding properties for each constituent IgG species. C_LIO_LIAn equilibrium, multivalent binding model can be generalized to incorporate immune complexes of mixed Fc composition. C_LIO_LIParticularly for low-affinity IgG-Fc{gamma} receptor interactions, immune complexes provide better estimates of affinities. C_LIO_LIThe Fc{gamma}R binding model predicts effector-elicited cell clearance in humanized mice. C_LI

immunology↗

bioRxiv2022

Staphylococcal protein A inhibits IgG-mediated phagocytosis by blocking the interaction of IgGs with FcγRs and FcRn

Immunoglobulin G molecules are crucial for the human immune response against bacterial infections. IgGs can trigger phagocytosis by innate immune cells, like neutrophils. To do so, IgGs should bind to the bacterial surface via their variable Fab regions and interact with Fc{gamma} receptors (Fc{gamma}Rs) and complement C1 via the constant Fc domain. C1 binding to IgG-labeled bacteria activates the complement cascade, which results in bacterial decoration with C3-derived molecules that are recognized by complement receptors (CRs) on neutrophils. Next to Fc{gamma}Rs and CRs on the membrane, neutrophils also express the intracellular neonatal Fc receptor (FcRn). We previously reported that staphylococcal protein A (SpA), a key immune evasion protein of Staphylococcus aureus, potently blocks IgG-mediated complement activation and killing of S. aureus by interfering with IgG hexamer formation. SpA is also known to block IgG-mediated phagocytosis in absence of complement but the mechanism behind it remains unclear. Here we demonstrate that SpA blocks IgG-mediated phagocytosis and killing of S. aureus through inhibition of the interaction of IgGs with Fc{gamma}Rs (Fc{gamma}RIIa and Fc{gamma}RIIIb, but not Fc{gamma}RI) and FcRn. Furthermore, our data show that multiple SpA domains are needed to effectively block IgG1-mediated phagocytosis. This provides a rationale for the fact that SpA from S. aureus contains four to five repeats. Taken together, our study elucidates the molecular mechanism by which SpA blocks IgG-mediated phagocytosis and supports the idea that next to Fc{gamma}Rs, also the intracellular FcRn receptor is essential for efficient phagocytosis and killing of bacteria by neutrophils.

microbiology↗

bioRxiv2017

Dissecting FcγR Regulation Through a Multivalent Binding Model

Many immune receptors transduce activation across the plasma membrane through their clustering. With Fc{gamma} receptors, this clustering is driven by binding to antibodies of differing affinities that are in turn bound to multivalent antigen. As a consequence of this activation mechanism, accounting for and rationally manipulating IgG effector function is complicated by, among other factors, differing affinities between Fc{gamma}R species and changes in the valency of antigen binding. In this study, we show that a model of multivalent receptor-ligand binding can effectively account for the contribution of IgG-Fc{gamma}R affinity and immune complex valency. This model in turn enables us to make specific predictions about the effect of immune complexes of defined composition. In total, these results enable both rational immune complex design for a desired IgG effector function and the deconvolution of effector function by immune complexes.\n\nSummary pointsO_LIAvidity most prominently modulates low-affinity Fc{gamma}R-immune complex binding\nC_LIO_LIA multivalent binding model can quantitatively predict Fc{gamma}R-immune complex binding\nC_LIO_LIImmune complex avidity has an outsized contribution to Fc{gamma}R multimerizationas compared to binding\nC_LIO_LIA binding model deconvoles and predicts the influence of interventions modulating in vivo Fc{gamma}R-driven effector function\nC_LI

immunology↗