Biology subjects

Bertone, P.

Publications and source records attributed to Bertone, P..

10 recordsLinked to original sources

bioRxiv2025

Enabling CAR-T Cell Immunotherapy in Glioblastoma by Modifying Tumor Microenvironment via Oncolytic Adenovirus Encoding Bispecific T Cell Engager

Recent clinical trials show that CAR-T cell therapies can initially blunt tumor growth in glioblastoma (GBM) patients. However, the tumor microenvironment activates mechanisms that inhibit tumor-killing potential of the CAR-T cells and limit their therapeutic efficacy. To counteract this, we have utilized oncolytic adenovirus (OV) Ad5-{Delta}24-RGD as a platform to overexpress a bispecific T cell engager (BiTE) targeting both T cell marker CD3 and GBM specific tumor associated antigen IL-13R2. We first demonstrated that OV-BiTE could enhance recruitment of T cells to GBM in vitro and in vivo. We then showed that intratumoral injection of OV-BiTE followed by infusion of combined EGFR- and EGFRvIII-CAR-T cells was more effective than OV-BiTE supplemented with either CAR-T therapy alone, and led to significant tumor eradication in a GBM xenograft mouse model. In conclusion, our multimodal OV-BiTE & CAR-T cell immunotherapy is capable of overcoming immunosuppressive tumor microenvironment and GBM resistance to treatment. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=160 SRC="FIGDIR/small/667708v1_ufig1.gif" ALT="Figure 1"> View larger version (47K): org.highwire.dtl.DTLVardef@c55b15org.highwire.dtl.DTLVardef@deffb2org.highwire.dtl.DTLVardef@64ff90org.highwire.dtl.DTLVardef@c65fc2_HPS_FORMAT_FIGEXP M_FIG C_FIG HIGHLIGHTSO_LIOncolytic adenovirus encoding bispecific T cell engager (OV-BiTE) combines two immunotherapeutic agents into one. C_LIO_LIOV-BiTE strategy modifies tumor microenvironment and enhances recruitment of T cells to glioblastoma (GBM) in vitro and in vivo. C_LIO_LIMultimodal OV-BiTE & CAR-T cell immunotherapy effectively reduced tumor mass in a GBM xenograft mouse model and is superior to either immunotherapy alone. C_LI

cancer biology↗

bioRxiv2021

Electrodeformation of White Blood Cells Enriched with Gold Nanoparticles

The elasticity of white blood cells (WBCs) provides valuable insight into the condition of the cells themselves, the presence of some diseases, as well as immune system activity. In this work, we describe a novel process of refined control of WBCs elasticity through a combined use of gold nanoparticles (AuNPs) and the microelectrode array device. The capture and controlled deformation of gold nanoparticles enriched white blood cells in vitro are demonstrated and quantified. Gold nanoparticles enhance the effect of electrically induced deformation and make the DEP related processes more controllable.

biophysics↗

bioRxiv2020

A maternal effect regulates global DNA methylation patterns

Variation in DNA methylation between individuals has been shown to be influenced by both genetic and environmental factors. However, the relative impact of genetic and non-genetic factors on DNA methylation patterns across the mammalian genome has not been systematically studied. We performed whole-genome methylation analysis in two inbred mouse strains, revealing striking differences in the global distribution of DNA methylation. Although global methylation patterns were indistinguishable for most genomic features, a significant increase in the number of unmethylated CpG-island promoters and first exons was observed between strains. Experiments using F1 reciprocal hybrid strains demonstrated that the genotype of the mother dictated global DNA methylation patterns. Cross-fostering experiments ruled out a postnatal maternal effect on these differences and suggested that they were driven by a prenatal maternal effect, possibly via differential deposition of maternal gene products into the oocyte or uterine environment. These data demonstrate that maternal effects have a major impact on global DNA methylation patterns.

genomics↗

bioRxiv2020

Sox2 modulation increases naive pluripotency plasticity

Induced pluripotency provides a tool to explore the molecular mechanisms underlying the establishment, maintenance and differentiation of naive pluripotent stem cells (nPSCs). Here, we report that self-renewal of nPSCs requires minimal Sox2 expression (Sox2-low). Sox2-low nPSCs do not show impaired neuroectoderm specification and differentiate efficiently in vitro into all embryonic germ lineages. Strikingly, Sox2-low cells also differentiate towards the trophoblast lineage both in vitro and in vivo. At the single-cell level self-renewing Sox2-low nPSCs exhibit a homogeneous naive molecular signature. However, they also display a basal trophoblast molecular signature and decreased ability of Oct4 to bind naive-associated regulatory sequences compared to control cells. These features underlie observed enhanced cell potency upon the removal of self-renewing cues. In sum, this work defines Sox2 as a restrictor of developmental potential and suggests perturbation of the naive pluripotent network as an underlying cause of increased cell potency. HighlightsO_LILow Sox2 expression is sufficient for naive pluripotent stem cell self-renewal C_LIO_LILow Sox2 expression does not impair neurectoderm differentiation in vitro C_LIO_LILow Sox2 expression impairs Oct4 genomic occupancy C_LIO_LILow Sox2 expression increases naive pluripotent cell plasticity in vitro and in vivo C_LI

developmental biology↗

bioRxiv2019

StemBond hydrogels optimise the mechanical microenvironment for embryonic stem cells

Studies of mechanical signalling are typically performed by comparing cells cultured on soft and stiff hydrogel-based substrates. However, it is challenging to independently and robustly control both substrate stiffness and tethering of extracellular matrix (ECM) to substrates, making ECM tethering a potentially confounding variable in mechanical signalling investigations. Moreover, poor ECM tethering can lead to weak cell attachment. To address this, we developed StemBond hydrogels, a hydrogel formulation in which ECM tethering is stable and can be varied independently of stiffness. We show that soft StemBond hydrogels provide an optimal format for culturing embryonic stem (ES) cells. We find that soft StemBond substrates improve the homogeneity of ES cell populations, boost their self-renewal, and increase the efficiency of cellular reprogramming. Our findings underline how soft microenvironments impact mechanosensitive signalling pathways regulating self-renewal and differentiation, indicating that optimising the complete mechanical microenvironment will offer greater control over stem cell fate specification.

cell biology↗

bioRxiv2018

Single-cell transcriptome analysis of human, marmoset and mouse embryos reveals common and divergent features of preimplantation development

The mouse embryo is the canonical model for mammalian preimplantation development. Recent advances in single-cell profiling allow detailed analysis of embryogenesis in other eutherian species, including human, to distinguish conserved from divergent regulatory programs and signalling pathways in the rodent paradigm. Here, we identify and compare transcriptional features of human, marmoset and mouse embryos by single-cell RNA-seq. Zygotic genome activation correlates with the presence of Polycomb Repressive Complexes in all three species, while ribosome biogenesis emerges as a predominant attribute in primate embryos, supporting prolonged translation of maternally deposited RNAs. We find that transposable element expression signatures are species-, stage- and lineage-specific. The pluripotency network in the primate epiblast lacks certain regulators operative in mouse, but encompasses WNT components and genes associated with trophoblast specification. Sequential activation of GATA6, SOX17 and GATA4 markers of primitive endoderm identity is conserved in primates. Unexpectedly, OTX2 is also associated with primitive endoderm specification in human and nonhuman primate blastocysts. Our cross-species analysis demarcates both conserved and primate-specific features of preimplantation development and underscores the molecular adaptability of early mammalian embryogenesis.

developmental biology↗

bioRxiv2017

Integrated analysis of single-cell embryo data yields a unified transcriptome signature for the human preimplantation epiblast

Single-cell profiling techniques create opportunities to delineate cell fate progression in mammalian development. Recent studies provide transcriptome data from human preimplantation embryos, in total comprising nearly 2000 individual cells. Interpretation of these data is confounded by biological factors such as variable embryo staging and cell-type ambiguity, as well as technical challenges in the collective analysis of datasets produced with different sample preparation and sequencing protocols. Here we address these issues to assemble a complete gene expression time course spanning human preimplantation embryogenesis. We identify key transcriptional features over developmental time and elucidate lineage-specific regulatory networks. We resolve post hoc cell-type assignment in the blastocyst, and define robust transcriptional prototypes that capture epiblast and primitive endoderm lineages. Examination of human pluripotent stem cell transcriptomes in this framework identifies culture conditions that sustain a naive state pertaining to the inner cell mass. Our approach thus clarifies understanding both of lineage segregation in the early human embryo and of in vitro stem cell identity, and provides an analytical resource for comparative molecular embryology.

developmental biology↗

bioRxiv2017

Epigenetic resetting of human pluripotency

Much attention has focussed on conversion of human pluripotent stem cells (PSC) to a more naive developmental status. Here we provide a method for resetting via transient histone deacetylase inhibition. The protocol is effective across multiple PSC lines and can proceed without karyotype change. Reset cells can be expanded without feeders with a doubling time of around 24 hours. WNT inhibition stabilises the resetting process. The transcriptome of reset cells diverges markedly from primed PSC and shares features with human inner cell mass (ICM). Reset cells activate expression of primate-specific transposable elements. DNA methylation is globally reduced to the level in the ICM but is non-random, with gain of methylation at specific loci. Methylation imprints are mostly lost, however. Reset cells can be re-primed to undergo tri-lineage differentiation and germline specification. In female reset cells, appearance of bi-allelic X-linked gene transcription indicates re-activation of the silenced X chromosome. On re-conversion to primed status, XIST-induced silencing restores monoallelic gene expression. The facile and robust conversion routine with accompanying data resources will enable widespread utilisation, interrogation, and refinement of candidate naive cells.

developmental biology↗

bioRxiv2017

The Nucleosome Remodelling and Deacetylation complex restricts Mediator access to enhancers to control transcription

A number of different chromatin remodelling complexes in mammalian cells are implicated in the control of gene expression. The genetic requirements for many such complex components have been described, and the biochemical activities of complex components characterised in vitro, yet the molecular mechanisms by which these biochemical activities impact transcriptional regulation in vivo remain ill-defined. Using an inducible system with fine temporal resolution, we show that the Nucleosome Remodelling and Deacetylation (NuRD) complex directly regulates chromatin architecture at enhancer regions in ES cells, in turn influencing the activity of RNA polymerase II via Mediator. Through this mechanism NuRD restricts Mediator access to enhancer chromatin during lineage commitment, thereby enabling appropriate transcriptional regulation. In contrast, acetylation levels of histone H3 lysine 27 are not immediately impacted by NuRD activity, correlating with transcriptional response only after expression levels have changed. These findings provide a detailed, molecular picture of genome-wide modulation of lineage-specific transcription by an abundant chromatin remodelling complex.

molecular biology↗

bioRxiv2016

Tracking the embryonic stem cell transition from ground state pluripotency

Mouse embryonic stem (ES) cells are locked into self-renewal by shielding from inductive cues. Release from this ground state in minimal conditions offers a system for delineating developmental progression from naive pluripotency. Here we examined the initial transition of ES cells. The population behaves asynchronously. We therefore exploited a short-half-life Rex1::GFP reporter to isolate cells either side of exit from naive status. Extinction of ES cell identity in single cells is acute. It occurs only after near-complete elimination of naive pluripotency factors, but precedes appearance of lineage specification markers. Cells newly departed from the ES cell state exhibit global transcriptome features consistent with features of early post-implantation epiblast and distinct from primed epiblast. They also exhibit a genome-wide increase in DNA methylation, intermediate between early and late epiblast. These findings are consistent with the proposition that naive cells transition to a discrete formative phase of pluripotency preparatory to lineage priming.\n\nHighlightsO_LIThe Rex1 destabilized GFP reporter demarcates naive pluripotency.\nC_LIO_LIExit from the naive state is asynchronous in the population.\nC_LIO_LITransition is relatively acute in individual cells and precedes lineage priming.\nC_LIO_LITranscriptome and DNA methylome reflect events in the pre-gastrulation embryo.\nC_LI

developmental biology↗