Biology subjects

Golebiewska, A.

Publications and source records attributed to Golebiewska, A..

9 recordsLinked to original sources

bioRxiv2026

Strength in Unity: a Dual Strategy to Restore NK Cell Cytotoxicity against Pancreatic Ductal Adenocarcinoma

BackgroundPancreatic ductal adenocarcinoma (PDAC), a condition representing 90% of pancreatic cancers, shows one of the lowest 5-year survival rates across all cancer types. Current therapeutic approaches remain largely inefficient, in part due to the presence of a hostile tumor microenvironment (TME), impeding immune cells infiltration and function. Specifically, Natural Killer (NK) cells from PDAC patients exhibit impaired phenotype and cytotoxic functions. NK cell immunotherapy represents a safe and promising approach to restore NK cell cytotoxicity against PDAC. MethodsWe developed a dual strategy based on i) the re-activation of NK cells through Natural Killer activating multimeric immunotherapeutic complexes (NaMiX) composed of IL-15/IL-15R dimers coupled to anti-NKp46 single-chain variable fragments (scFvs) and ii) the crosslinking of activated NK cells to PDAC cells with a Trispecific Killer Engager (TriKE) targeting NKG2D, NKp30 and the tumor-associated antigen CEA. We evaluated the ability of these constructs to stimulate NK cell functions across BxPC-3 PDAC cell line and patient-derived organoid models and in humanized NSG mice bearing PDAC xenografts. ResultsNaMiX stimulated the activation and cytotoxic functions of NK cells towards pancreatic BxPC-3 cells in vitro while TriKE cross-linked NK cells to BxPC-3 cells. The cytotoxic effects of NaMiX were further enhanced when combined with the crosslinking abilities of TriKE for the killing of NK cell-mediated BxPC-3 spheroid and PDAC patient-derived organoids. In humanized mice bearing BxPC-3 xenografts, NaMiX induced cytotoxic lymphocyte expansion, and increased tumor infiltration of NK cells, while TriKE tended to slow tumor progression. ConclusionsThis proof-of-concept study reports for the first time that activating and engaging NK cells with immunoconjugates are a promising therapeutic avenue for PDAC treatment. Efforts should now focus on the optimization of NK cell therapeutic modalities to favor the infiltration of a high number of NK cells into the tumor.

immunology↗

bioRxiv2025

Humanized glioblastoma patient-derived orthotopic xenografts recreate a locally immunosuppressed human immune ecosystem amenable to immunotherapeutic modulation

Immune-based strategies have so far failed to demonstrate clinical benefit in glioblastoma (GBM), largely due to the profound immunosuppressive tumor microenvironment (TME). To achieve more predictive preclinical insights, advanced in vivo models that faithfully recapitulate the human brain immune landscape are urgently needed. Here, we established GBM patient-derived orthotopic xenografts (PDOXs) across diverse mouse strains, including humanized models. Humanization was achieved through transplantation of CD34+ hematopoietic stem cells (HU-CD34+) or peripheral blood mononuclear cells (HU-PBMC). Both models successfully reconstituted human T-cells systemically, with stronger engraftment in HU-CD34+ mice. We observed selective infiltration and spatial organization to intracranial GBM tumors, including exhausted, memory-like, and regulatory CD4+ T-cell phenotypes, TIM-3+ immunosuppressive-like myeloid cells and intratumoral B cells. Mouse microglia-derived tumor-associated macrophages (TAMs) remained the dominant immunosuppressive immune population. Anti-PD-1 therapy, but not anti-GITR, modestly modulated the infiltration dynamics, demonstrating the susceptibility of the reconstructed adaptive immunity to immunotherapeutic intervention. These findings position humanized GBM PDOXs as a relevant preclinical platform to interrogate tumor-immune interactions and evaluate immunotherapeutic strategies in a human context. Key pointsO_LIGBM PDOXs developed in HU-CD34+ and HU-PBMC mice faithfully reconstitute systemic and local human adaptive immunity. C_LIO_LIHuman immune components undergo selective infiltration, spatial organization and transition towards exhausted CD4+ T-cells and immunosuppressive CD11c+ myeloid cells. C_LIO_LIAnti-PD-1, but not anti-GITR, locally promote human immune infiltration into intracranial GBM tumors, while sparing systemic compartments. C_LIO_LIHumanized GBM PDOXs provide a powerful preclinical platform to test novel immunotherapeutic strategies. C_LI Study importanceImmune checkpoint blockade has shown limited efficacy in GBM, reflecting the highly immunosuppressive and lymphocyte-poor nature of the TME. Conventional syngeneic and GEMM models fail to recapitulate these features, contributing to the translational disconnect between preclinical success and clinical failure. Humanized mice provide a solution to interrogate human-specific immunity in vivo, but their use in GBM has remained limited. Here, we provide the first comparison of GBM PDOX modeling in two complementary modes of humanization based on CD34+ HSCs and PBMCs. We systematically profile systemic and intratumoral compartments, showing that these models faithfully reconstitute human adaptive immunity and capture the interplay with the murine brain TME. Furthermore, we demonstrate clinically-relevant responses upon treatment with checkpoint antibodies targeting PD-1 and GITR, showing modulation of human immune subsets without altering murine TAM immunosuppression, underscoring the translational value of the system. This study establishes humanized GBM PDOXs as a versatile platform for dissecting tumor-immune interactions in the brain and for preclinical evaluation and development of novel immunotherapies. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=200 SRC="FIGDIR/small/689484v2_ufig1.gif" ALT="Figure 1"> View larger version (74K): org.highwire.dtl.DTLVardef@1b684e1org.highwire.dtl.DTLVardef@1cdfbbdorg.highwire.dtl.DTLVardef@4a570borg.highwire.dtl.DTLVardef@98b112_HPS_FORMAT_FIGEXP M_FIG C_FIG

cancer biology↗

bioRxiv2025

Integrative multi-omics combined with functional pharmacological profiling in patient-derived organoids identifies personalized therapeutic vulnerabilities of adult high-grade gliomas

BackgroundPrecision medicine has transformed cancer treatment by tailoring therapies to specific molecular aberrations. Integrating high-resolution multi-omics with high-throughput functional profiling in patient-derived organoids of-fers a powerful strategy to further refine patient stratification. While (epi)genetic profiling has drastically improved the classification in diffuse adult gliomas, these advances have not yet translated into effective therapeutic interventions and precision medicine approaches remain to be established. Material and MethodsWe investigated a panel of 48 patient-derived organoid and orthotopic xenograft models of adult high-grade gliomas, comprehensively characterized at genomic, epigenomic and transcriptomic levels. A functional drug screen was performed on 27 organoid models using a 202-compound library targeting cancer-related pathways and epigenetic regulators. Unsupervised multi-omics factor analysis was employed to identify patient-specific therapeutic vulnerabilities. Validation included dose-dependent drug efficacy assessments, as well as biomarker assessment in patient tumors across molecular subgroups. ResultsMulti-omics analysis revealed a broad spectrum of molecular profiles capturing the genetic, epigenetic, and transcriptomic diversity of high-grade gliomas. Multi-omics factor analysis, integrating multi-omics and drug response profiles, identified distinct subgroups associated with IDH1 mutation and MYCN amplification. IDH1 mutant grade 4 astrocytomas showed selective sensitivity to histone deacetylase 3 inhibitors, while a MYCN-amplified glioblastoma responded preferentially to histone methyltransferase inhibitors. The differential drug responses were linked to specific (epi)genetic and transcriptomic biomarkers. While other glioblastomas exhibited heterogeneous treatment responses, no robust biomarker-defined responder subgroups were identified. ConclusionOur findings highlight the value of integrating multi-omics and functional profiling to inform precision medicine strategies. This approach enables the stratification of distinct patient subgroups in preclinical models, paving the way for tailored therapeutic interventions. While we observed distinct pharmacogenomic profiles in IDH1 mutant grade 4 astrocytomas and a MYCN-amplified glioblastoma, implementing precision medicine in other glioblastoma subtypes remains a substantial challenge. Key pointsO_LIIntegrating drug screening in a panel of patient-derived organoids with multi-omics enables pharmacogenomic profiling in adult diffuse high-grade gliomas C_LIO_LIIDH1 mutant grade 4 astrocytomas are sensitive to histone deacetylase 3 inhibitors C_LIO_LIMYCN-amplified glioblastoma exhibits distinct DNA methylation pattern and drug responses C_LI Study importanceTo date, attempts to develop effective precision medicine in adult high-grade gliomas failed. Here, we provide a preclinical framework for identifying personalized therapeutic by integrating multi-omics profiling with functional drug screening in patient-derived organoids. We show that IDH1 mutant high-grade astrocytomas present distinct therapeutic vulnerabilities compared to glioblastomas, linked to sensitivity to histone deacetylase 3 inhibitors. Within glioblastomas, we identified a distinct MYCN-amplified tumor, sensitive to histone methyltransferase inhibitors. Applying pharmacogenomic approaches using novel drug libraries holds promise for uncovering additional clinically relevant patient subgroups in the future. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=104 SRC="FIGDIR/small/675145v1_ufig1.gif" ALT="Figure 1"> View larger version (28K): org.highwire.dtl.DTLVardef@12c04f9org.highwire.dtl.DTLVardef@fa8872org.highwire.dtl.DTLVardef@141de6org.highwire.dtl.DTLVardef@b72086_HPS_FORMAT_FIGEXP M_FIG C_FIG

cancer biology↗

bioRxiv2025

Tumor-initiating genetics and therapy drive divergent molecular evolution in IDH-mutant gliomas

Astrocytomas and oligodendrogliomas are slow-growing and treatment-sensitive IDH-mutant gliomas diagnosed at ages 30-50. Local tumor regrowth and treatment resistance is inevitable resulting in 3-10 year astrocytoma and up to >20 years oligodendroglioma survival. We sought to identify genetic changes associated with tumor evolution in response to therapy through multi-timepoint whole-genome/whole-exome sequencing of 206 IDH-mutant glioma patient samples collected through the Glioma Longitudinal Analysis (GLASS) Consortium. We validated known genomic markers of tumor progression, including hypermutation and CDKN2A homozygous deletion, and discovered novel genetic alterations that distinguish the response to treatment in astrocytomas compared to oligodendrogliomas. Point mutations in PIK3CA, PIK3R1, and NOTCH1 were newly acquired in recurrent oligodendrogliomas and associated with increased mutation rates. Focal oncogene amplifications, together with CDKN2A homozygous deletions, were associated with an increase in recurrence-specific chromosomal imbalances in astrocytomas. Mutational signature analysis revealed additional differences and detected enrichment for the SBS11, and SBS119 mutational signatures after temozolomide treatment in both IDH-glioma subtypes, whereas astrocytomas showed increased ID8 signatures after radiotherapy. These signatures suggest that the genomes of oligodendroglioma and astrocytoma adapt to the selective pressures of tumor progression and treatment in different ways. However, in both IDH-mutant glioma subtypes we observed a convergence of acquired driver gene alterations with genome-wide changes and worse patient outcomes, signaling selection of treatment-refractory clones. By identifying new prognostic markers and delineating the genomic divergence of oligodendrogliomas and astrocytomas after diagnosis, our results suggest that different DNA damage response mechanisms are engaged following chemo- and radiation therapy.

cancer biology↗

bioRxiv2024

Adenoviral Delivery of the CIITA Transgene Induces T-Cell-Mediated Killing in Glioblastoma Organoids

The immunosuppressive nature of the tumor microenvironment poses a significant challenge to effective immunotherapies against glioblastoma (GB). Boosting the immune response is critical for a successful therapy. Here, we adopted a cancer gene therapy approach to induce T-cell mediated killing of the tumor through increased activation of the immune system. Patient-based 3D GB models were infected with a replication-deficient adenovirus (AdV) armed with the Class II Major Histocompatibility Complex (MHC-II) Transactivator CIITA gene (Ad-CIITA). Successful induction of surface MHC-II was achieved in infected GB cell lines and primary human GB organoids. Infection with an AdV carrying a mutant form of CIITA with a single amino acid substitution resulted in cytoplasmic accumulation of CIITA without subsequent MHC-II expression. Co-culture of infected tumor cells with either PBMCs or isolated T-cells led to dramatic breakdown of GB organoids. Intriguingly, both wild-type and mutant Ad-CIITA but not unarmed AdV, triggered immune-mediated tumor cell death in the co-culture system, suggesting an at least partially MHC-II-independent process. We further show that the observed cancer cell killing requires the presence of either CD8+ or CD4+ T-cells and the direct contact between GB and immune cells. We did not however detect evidence of activation of canonical T-cell mediated cell death pathways. While the precise mechanism remains to be determined, these findings highlight the potential of AdV-mediated CIITA delivery to enhance T-cell-mediated immunity against GB.

cancer biology↗

bioRxiv2023

Deep intravital brain tumor imaging enabled by tailored three-photon microscopy and analysis

Intravital two-photon microscopy has emerged as a powerful technology to study brain tumor biology and its temporal dynamics, including invasion, proliferation and therapeutic resistance in the superficial layers of the mouse cortex. However, intravital microscopy of deeper cortical layers and especially the subcortical white matter, an important route of glioblastoma invasion and recurrence, has not yet been feasible due to low signal-to-noise ratios, missing spatiotemporal resolution and the inability to delineate myelinated axonal tracts. Here, we present a tailored intravital microscopy and artificial intelligence-based analysis methodology and workflow that enables routine deep imaging of glioblastoma over extended time periods, named Deep3P. We show that three-photon microscopy, adaptive optics, as well as customized deep learning-based denoising and machine learning segmentation together allow for deep brain intravital investigation of tumor biology up to 1.2 mm depth. Leveraging this approach, we find that perivascular invasion is a preferred invasion route into the corpus callosum as compared to intracortical glioblastoma invasion and uncover two vascular mechanisms of glioblastoma migration in the white matter. Furthermore, we can define an imaging biomarker of white matter disruption during early glioblastoma colonization. Taken together, Deep3P allows for an efficient and non-invasive investigation of brain tumor biology and its tumor microenvironment in unprecedented deep white and gray matter of the living mouse, opening up novel opportunities for studying the neuroscience of brain tumors and other model systems.

cancer biology↗

bioRxiv2023

Glioblastoma-instructed microglia transit to heterogeneous phenotypic states with phagocytic and dendritic cell-like features in patient tumors and patient-derived orthotopic xenografts

BackgroundA major contributing factor to glioblastoma (GBM) development and progression is its ability to evade the immune system by creating an immune-suppressive environment, where GBM-associated myeloid cells, including resident microglia and peripheral monocyte-derived macrophages, play critical pro-tumoral roles. However, it is unclear whether recruited myeloid cells are phenotypically and functionally identical in GBM patients and whether this heterogeneity is recapitulated in patient-derived orthotopic xenografts (PDOXs). A thorough understanding of the GBM ecosystem and its recapitulation in preclinical models is currently missing, leading to inaccurate results and failures of clinical trials. MethodsHere, we report systematic characterization of the tumor microenvironment (TME) in GBM PDOXs and patient tumors at the single-cell and spatial levels. We applied single-cell RNA-sequencing, spatial transcriptomics, multicolor flow cytometry, immunohistochemistry and functional studies to examine the heterogeneous TME instructed by GBM cells. GBM PDOXs representing different tumor phenotypes were compared to glioma mouse GL261 syngeneic model and patient tumors. ResultsWe show that GBM tumor cells reciprocally interact with host cells to create a GBM patient-specific TME in PDOXs. We detected the most prominent transcriptomic adaptations in myeloid cells, with brain-resident microglia representing the main population in the cellular tumor, while peripheral-derived myeloid cells infiltrated the brain at sites of blood-brain barrier disruption. More specifically, we show that GBM-educated microglia undergo transition to diverse phenotypic states across distinct GBM landscapes and tumor niches. GBM-educated microglia subsets display phagocytic and dendritic cell-like gene expression programs. Additionally, we found novel microglial states expressing cell cycle programs, astrocytic or endothelial markers. Lastly, we show that temozolomide treatment leads to transcriptomic plasticity and altered crosstalk between GBM tumor cells and adjacent TME components. ConclusionOur data provide novel insights into the phenotypic adaptation of the heterogeneous TME instructed by GBM tumors. We show the key role of microglial phenotypic states in supporting GBM tumor growth and response to treatment. Our data place PDOXs as relevant models to assess the functionality of the TME and changes in the GBM ecosystem upon treatment. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=127 SRC="FIGDIR/small/531162v2_ufig1.gif" ALT="Figure 1"> View larger version (49K): org.highwire.dtl.DTLVardef@1549a3corg.highwire.dtl.DTLVardef@159f16aorg.highwire.dtl.DTLVardef@1f89500org.highwire.dtl.DTLVardef@fe6b67_HPS_FORMAT_FIGEXP M_FIG C_FIG

cancer biology↗

bioRxiv2020

Primary and recurrent glioma patient-derived orthotopic xenografts (PDOX) represent relevant patient avatars for precision medicine

Patient-derived cancer models are essential tools for studying tumor biology and preclinical interventions. Here, we show that glioma patient-derived orthotopic xenografts (PDOXs) enable long-term propagation of patient tumors and represent clinically relevant patient avatars. We created a large collection of PDOXs from primary and recurrent gliomas with and without mutations in IDH1, which retained histopathological, genetic, epigenetic and transcriptomic features of patient tumors with no mouse-specific clonal evolution. Longitudinal PDOX models recapitulate the limited genetic evolution of gliomas observed in patient tumors following treatment. PDOX-derived standardized tumor organoid cultures enabled assessment of drug responses, which were validated in mice. PDOXs showed clinically relevant responses to Temozolomide and to targeted treatments such as EGFR and CDK4/6 inhibitors in (epi)genetically defined groups, according to MGMT promoter and EGFR/CDK status respectively. Dianhydrogalactitol, a bifunctional alkylating agent, showed promising potential against glioblastoma. Our study underlines the clinical relevance of glioma PDOX models for translational research and personalized treatment studies.

cancer biology↗

bioRxiv2017

Dr.Paso: Drug response prediction and analysis system for oncology research

The prediction of anticancer drug response is crucial for achieving a more effective and precise treatment of patients. Models based on the analysis of large cell line collections have shown potential for investigating drug efficacy in a clinically-meaningful, cost-effective manner. Using data from thousands of cancer cell lines and drug response experiments, we propose a drug sensitivity prediction system based on a 47-gene expression profile, which was derived from an unbiased transcriptomic network analysis approach. The profile reflects the molecular activity of a diverse range of cancer-relevant processes and pathways. We validated our model using independent datasets and comparisons with published models. A high concordance between predicted and observed drug sensitivities was obtained, including additional validated predictions for four glioblastoma cell lines and four drugs. Our approach can accurately predict anti-cancer drug sensitivity and will enable further pre-clinical research. In the longer-term, it may benefit patient-oriented investigations and interventions.

bioinformatics↗