Biology subjects

Hadad, U.

Publications and source records attributed to Hadad, U..

5 recordsLinked to original sources

bioRxiv2026

Spatially enforced engagement of PD-1⁺ T cells with AXL⁺ cells reprograms the tumor microenvironment and overcomes anti-PD-1 resistance

Background: AXL overexpression promotes tumor-cell plasticity, local immune suppression, and resistance to PD-1 blockade. Conventional anti-AXL and anti-PD-1 antibodies combinations act independently and do not ensure physical proximity between PD-1+ T cells and AXL+ cells within the immunosuppressive tumor microenvironment. Methods: To address this limitation, we developed a non-tandem, tetravalent IgG bispecific antibody designed to simultaneously engage PD-1 and AXL (BiTE-AXL/PD-1). Its antitumor activity was evaluated using in vitro functional assays and humanized and immunocompetent head and neck squamous cell carcinoma (HNSCC) models and compared with the combination of monospecific anti-AXL and anti-PD-1 antibodies. Spatial multiplex imaging was used to characterize treatment-associated changes in the tumor microenvironment of an anti-PD-1-resistant murine model. Results: BiTE-AXL/PD-1 enhanced T-cell activation, promoted AXL-dependent tumor-T-cell conjugation, and increased tumor-cell killing in vitro, outperforming the combination of the two monospecific antibodies. In humanized and immunocompetent HNSCC models, BiTE treatment significantly delayed tumor growth compared with the antibody combination and was well tolerated, with no overt toxicity. Spatial multiplex imaging revealed a shift from a fibroblast-dominated microenvironment characterized by exhausted CD8+ T cells toward dendritic-cell- and CD8+ T-cell-enriched niches associated with antitumor immune activity. Conclusion: BiTE-AXL/PD-1 promotes physical proximity between PD-1+ T cells and AXL+ cells, enhances antitumor immune responses, and remodels the immunosuppressive tumor microenvironment. These findings support PD-1/AXL bispecific targeting as a promising therapeutic strategy to overcome resistance to PD-1 blockade in HNSCC.

cancer biology↗

bioRxiv2026

Primordial endocrine cells uncouple sexual differentiation from sex-chromosomes

Sexuality among the animal kingdom is remarkably plastic, spanning from gonochorism to sequential or simultaneous hermaphroditism. Over 70 years ago, Charniaux-Cotton hypothesized that sexual plasticity in crustaceans is mediated by primordial cells present in both sexes, which differentiate into an androgenic gland (AG) only in males. Although AG engraftments can masculinize genetic females, it remained unclear whether this reflects long-term integration of donor cells within the host or activation of host primordial AG progenitors. Using the freshwater prawn as an exceptional case of sexual plasticity, where AG-cell transfer yields functional neo-males bearing only female sex-chromosomes, we show, through cellular-resolution approaches, that the AG arises from primordial cells present in every individual, indicating autosomal machinery of sexual differentiation completely independent from the sex-chromosomes.

developmental biology↗

bioRxiv2024

Relative Distribution of DnaA and DNA in Escherichia coli Cells as a Factor of Their Phenotypic Variability

Phenotypic variability in isogenic bacterial populations is a remarkable feature that helps them cope with external stresses, yet it is incompletely understood. This variability can stem from gene expression noise and/or unequal partitioning of low-copy-number freely diffusing proteins during cell division. Some high-copy-number components are transiently associated with almost immobile large assemblies (hyperstructures) and may be unequally distributed, contributing to bacterial phenotypic variability. We focus on the nucleoid hyperstructure containing numerous DNA-associated proteins, including the replication initiator DnaA. Previously we found an increasing asynchrony in the nucleoid segregation dynamics in growing E. coli cell lineages and suggested that variable replication initiation timing may be the main cause of this phenomenon. Here we support this hypothesis revealing that DnaA/DNA variability represents a key factor leading to the enhanced asynchrony in E. coli. We followed the intra- and intercellular distribution of fluorescently tagged DnaA and histone-like HU chromosomally encoded under their native promoters. The diffusion rate of DnaA is low, corresponding to a diffusion-binding mode of mobility, but still one order faster than that of HU. The intracellular distribution of DnaA concentration is homogeneous in contrast to the significant asymmetry in the distribution of HU to the cell halves, leading to unequal DNA content of nucleoids and DnaA/DNA ratios in future daughter compartments. Accordingly, the intercellular variabilities of HU concentration (CV=26%) and DnaA/DNA ratio (CV=18%) are high. The variable DnaA/DNA may cause a variable replication initiation time (initiation noise). Asynchronous initiation at different replication origins may, in turn, be the mechanism leading to the observed asymmetric intracellular DNA distribution. Our findings indicate that the feature determining the variability of initiation time in E. coli is the DnaA/DNA ratio rather than separately each of them. We provide a likely mechanism for the loss of segregation synchrony phenomenon.

microbiology↗

bioRxiv2020

Molecular Scale Spatio-Chemical Control of the Activating-Inhibitory Signal Integration in NK Cells

The role of the spatial juxtaposition between activating and inhibitory receptors in cytotoxic lymphocytes has been strongly debated in the context of the inhibition of immune signaling. The challenge in addressing this problem was so far a lack of experimental tools which can simultaneously manipulate different signaling molecules. Here, we circumvent this challenge by introducing a nanoengineered multifunctional cell niche, in which activating and inhibitory ligands are positioned with molecular-scale variability and control, and applied it to elucidate the role of the spatial juxtaposition between ligands for NKG2D and KIR2DL1 - activating and inhibitory receptors in Natural Killer (NK) cells - in KIR2DL1-mediated inhibition of NKG2D signaling. We realized the niche by a nanopatterning of nanodots of different metals with molecular scale registry in one lithographic step, followed by a novel ternary functionalization of the fabricated bi-metallic pattern and its background to with three distinct biochemical moieties. We found, that within the probed range, the 40 nm gap between the activating and inhibitory ligands provided an optimal inhibition condition. Supported by theoretical modeling and simulations we interpret these findings as a consequence of the size and conformational flexibility of the ligands in their spatial interaction. Our findings provide an important insight onto the spatial mechanism of the inhibitory immune checkpoints, whose understanding is both fundamentally important, and essential for the rational design of future immunotherapies. Furthermore, our approach is highly versatile and paves the way to numerous complex molecular platforms aimed at revealing molecular mechanisms through which receptors integrate their signals.

immunology↗

bioRxiv2020

Mechanical Regulation of the Cytotoxic Activity of Natural Killer Cells

Mechanosensing has been recently explored for T cells and B cells and is believed to be part of their activation mechanism. Here, we explore the mechanosensing of the third type of lymphocytes - Natural Killer (NK) cells, by showing that they modulate their immune activity in response to changes in the stiffness of a stimulating surface. Interestingly, we found that this immune response is bell-shaped, and peaks for a stiffness of a few hundreds of kPa. This bell-shape behavior was observed only for surfaces functionalized with the activating ligand MHC class I polypeptide-related sequence A (MICA), but not for control surfaces lacking immunoactive functionalities. We found that stiffness does not affect uniformly all cells but increases the size of a little group of extra-active cells, which in turn contribute to the overall activation effect of the entire cell population. We further imaged the clustering of costimulatory adapter protein DAP10 on NK cell membrane and found that it shows the same bell -shape dependence to surface stiffness. Based on these findings, we propose a catch-bond-based model for the mechanoregulation of NK cell cytotoxic activity, through interaction of NKG2D activating receptors with MICA. Our findings reveal what seems to be "the tip of iceberg" of mechanosensation of NK cells, and provides an important insight on the mechanism of their immune signaling. Statement of SignificanceThe mechanical sensing of immune lymphocytes was recently demonstrated for T cells and B cells, but not for the third type of lymphocytes - Natural Killer (NK) cells. Interestingly, previous reports on lymphocyte mechanosensing were controversial, and showed either positive or negative changes in their immune activity with environmental stiffness, depending on the stiffness range. In this paper, we directly demonstrated that NK cells modulate their response with the stiffness of the stimulating surface, and this modulation has a bell-shape trend. We found that there is a strong correlation between the response to stiffness and clustering of adaptor proteins. Upon this correlation, we proposed a mechanosensing model based on the catch-bond nature of activating ligand-receptor complexes in NK cells.

immunology↗