Biology subjects

Kovar, H.

Publications and source records attributed to Kovar, H..

9 recordsLinked to original sources

bioRxiv2026

A human lung organoid co-culture model of early bone sarcoma metastasis reveals contact-dependent epithelial remodeling at the metastatic interface

Lung metastasis drives mortality across cancer types, yet how infiltrating tumor cells remodel the lung epithelium to establish metastatic niches remains poorly understood. Here we establish MESCUL (MEtastatic Sarcoma Co-CULture), a co-culture platform combining human lung organoids with patient-derived bone sarcoma cells to model early tumor-lung epithelial interactions in a physiologically relevant 3D system. MESCUL reveals that direct tumor-epithelial contact induces rapid, reproducible lung epithelial remodeling across Ewing sarcoma (ES) and osteosarcoma (OS) models and multiple organoid donor backgrounds, which is contact-dependent and not recapitulated by paracrine signaling. Single-cell RNA sequencing identifies LIMES (Lung Interface Metastasis Signature), a shared transcriptional program encompassing focal adhesion assembly, matrix metalloprotease (MMP) upregulation, and emergence of a damage-associated transitional cell state, in both ES and OS. Mechanistically, tumor-derived fibronectin (FN1) engages epithelial integrin receptors to activate focal adhesion kinase (FAK), driving amphiregulin (AREG) induction and MMP-mediated remodeling; FN1 alone phenocopies this response, and FAK inhibition attenuates it, nominating the FN1-integrin-FAK-AREG axis as a candidate therapeutic vulnerability. The LIMES program, identified through the MESCUL co-culture model, is spatially confined to the tumor-lung interface in patient metastases of both ES and OS, as demonstrated by spatial transcriptomics across nine patients. Massons trichrome staining of matched patient sections reveals pronounced collagen deposition in the peri-tumoral lung parenchyma, consistent with LIMES acting upstream of a wound-healing cascade that proceeds to structural fibrotic remodeling in patient tissue. Together, these findings establish the lung epithelium as an active participant in metastatic colonization, characterize a pharmacologically targetable, spatially restricted epithelial remodeling response at the bone sarcoma-lung interface across OS and ES, and introduce MESCUL as a tractable 3D platform for investigating lung metastasis, with possible implications for tumor types beyond bone sarcomas.

cancer biology↗

bioRxiv2026

The ITCC-P4 PDX platform of pediatric cancers for preclinical testing

Cancer is the leading cause of disease-related deaths among children in high-income countries. Tumor heterogeneity and lack of mechanism-of-action-based therapeutic options are key challenges to overcome in order to improve pediatric cancer patients survival. Here, we report the EU-IMI-2 funded public-private partnership "ITCC-Pediatric Preclinical Proof-of-Concept Platform" (ITCC-P4), which has built a large repertoire of patient-derived xenograft (PDX) models, representing all major solid pediatric cancer types, for in vivo drug testing. Three-hundred-fifty-three PDX models from diagnostic and relapsed pediatric cancers have been established and molecularly characterized, together with matched germline/tumor samples. As proof-of-concept, we present in vivo drug screening data in neuroblastoma and rhabdomyosarcoma models. PDX data, accessible at http://r2platform.com/itcc-p4, allow the selection of models based on oncogenic drivers and/or potential biomarkers for preclinical testing. Operated by a non-profit entity (www.itccp4.com), this sustainable platform aids academic and industrial researchers in developing and prioritizing innovative therapies for pediatric cancer. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=112 SRC="FIGDIR/small/703023v1_ufig1.gif" ALT="Figure 1"> View larger version (41K): org.highwire.dtl.DTLVardef@195ba30org.highwire.dtl.DTLVardef@f2c2d9org.highwire.dtl.DTLVardef@1d63f4dorg.highwire.dtl.DTLVardef@d60027_HPS_FORMAT_FIGEXP M_FIG C_FIG

cancer biology↗

bioRxiv2025

Mevalonate pathway activation in Ewing sarcoma reveals a 3D-specific synergy between statins and BCL-xL inhibition

Bone sarcomas are rare and aggressive pediatric cancers with limited progress in targeted therapy development, in part due to the poor physiological relevance of conventional two-dimensional (2D) culture systems used for preclinical testing. To address this gap, we developed a standardized three-dimensional (3D) culture and drug-testing platform for Ewing sarcoma (ES) and osteosarcoma (OS) that more accurately recapitulates in vivo tumor biology. Across 3D spheroids, bioprinted constructs, and patient-derived xenograft (PDX) cultures, we observed a consistent activation and dependency on the mevalonate pathway in ES. Leveraging this platform, we identified a selective therapeutic synergy between statins, which inhibit mevalonate pathway flux, and BCL-xL inhibitors, a vulnerability that was not detectable in 2D cultures. These findings highlight the mevalonate pathway as a targetable metabolic dependency in ES and demonstrate how physiologically grounded 3D models can uncover clinically actionable treatment strategies that remain hidden in traditional 2D systems. Our findings show that 3D tumor models can expose actionable metabolic vulnerabilities obscured by traditional approaches, supporting their use in rational combination therapy discovery for aggressive pediatric sarcomas.

cancer biology↗

bioRxiv2025

Dynamic modelling of EWS::FLI1 fluctuations reveals molecular determinants of phenotypic tumor plasticity and prognosis in Ewing sarcoma

The mechanisms underlying tumor cell plasticity driving drug resistance and disease progression remain poorly understood. In Ewing sarcoma (EwS), variations in EWS::FLI1 (EF) activity have been associated with epithelial-mesenchymal plasticity (EMP). Using degron technology, we titrated endogenous EF in an EwS cell line and linked phenotypic states to distinct EF thresholds. Strikingly, modest EF depletion promoted a pro-metastatic phenotype, that diminished upon near-complete EF loss. Nascent RNA sequencing revealed distinct gene clusters with heterogenous response patterns to varying EF dosage. Target genes most sensitive to subtle EF depletion contained GGAA microsatellites in EF-bound enhancers. Furthermore, we identified Kruppel-like zinc-finger transcription factors associated with EF-repressed EMP genes. Transient EF depletion followed by rapid restoration to simulate oncoprotein fluctuations identified persistently dysregulated genes associated with poor prognosis. This study underscores the therapeutic challenge of insufficient EF inhibition and provides a foundation for exploiting oncoprotein dynamics to uncover therapeutic vulnerabilities in fusion-driven cancers. Beyond EwS, our results underscore the broader impact of oncoprotein dosage dynamics in cancers with otherwise quiet genomes. SIGNIFICANCEWe report EwS as a paradigm for the importance of oncogene fluctuations in tumor cell plasticity and disease progression. Effective therapeutic strategies must ensure complete EF depletion to prevent inadvertent metastasis.

cancer biology↗

bioRxiv2025

Small Particles, Big Problems: Polystyrene nanoparticles induce DNA damage, oxidative stress, migration, and mitogenic pathways predominantly in non-malignant lung cells

Polystyrene micro-and nanoplastics (PS-MNPs) are emerging environmental pollutants with potential implications for human health. In this study, we used two different sizes of PS-MNPs (0.25 {micro}m and 1 {micro}m) on non-small cell lung cancer (A549, H460), small cell lung cancer (DMS53, H372), and normal lung epithelial (BEAS-2B) cells, as well as on human-derived lung organoids, to investigate the cytotoxic effects of PS particles. At lower concentrations (< 30 {micro}g/cm2, equivalent to 50 {micro}g/ml), neither PS-MPs nor PS-NPs did not interfere with cell viability or proliferation. Intracellular kinetic assays revealed that non-malignant (BEAS-2B) lung cells showed the strongest turnover of PS-NPs compared to malignant cells. Since PS-NPs exhibited more pronounced cellular effects, we focused further analyses on their impact. Furthermore, we observed significantly increased migration, prolonged S-phase arrest along with induced DNA damage, and oxidative stress in non-malignant (BEAS-2B) lung cells. Thus, our data suggest that BEAS-2B cells exhibit the highest sensitivity to PS-NPs. We also demonstrate that after PS-NP treatment, these cells displayed decreased base excision repair capacity and increased activation of survival pathways, including AKT and ERK phosphorylation. PS-NP internalization and increase of signal pathways were validated in a more physiological lung organoid setting. Altogether, our findings suggest that PS-NPs do not significantly affect the malignant behavior of cancer cells. However, they could promote tumor-like features in normal lung cells by inducing survival pathways, migration, and alterations in stress response mechanisms. Environmental ImplicationsThis study investigates the effects of polystyrene micro-and nanoplastics (PS-MNPs) at environmentally relevant concentrations. The tested concentrations of PS-MNPs (0, 15, 30, and 60 {micro}g/cm2, equivalent to 0, 25, 50, and 100 {micro}g/ml) are commonly studied in the literature in lung cells. While these findings provide insights into cellular responses, the overall environmental impact of PS-MNPs remains limited at realistic exposure levels. HighlightsO_LIPS-MNPs are internalized into lung cells, with higher uptake in non-malignant cells. C_LIO_LIPS-NPs lead to increased migration, DNA damage, oxidative stress, and disturbed cell cycle progression. C_LIO_LIPS-NPs promote activation of survival pathways in non-malignant cells and lung organoids. C_LIO_LIExposure of PS-NPs may cause potential implications for lung cancer development and progression. C_LI

molecular biology↗

bioRxiv2024

YAP1 is a key regulator of EWS::FLI1-dependent malignant transformation upon IGF-1 mediated reprogramming of bone mesenchymal stem cells

AbstractEwing sarcoma (EwS) is an aggressive cancer of adolescents in need of effective treatments. Insulin like growth factor (IGF) 1 was previously reported an autocrine growth factor for EwS, but only 10% of patients responded to IGF-1 receptor blockade. Although presumed to originate from mesenchymal progenitors during bone development, targeting of the EwS driver oncogene EWS::FLI1 to the mesenchymal lineage in a conditional mouse model did not result in tumor formation but led to skeletal malformations and perinatal death. We report that transient exposure to IGF-1 concentrations mimicking serum levels during puberty reprogrammed limb-derived mesenchymal cells of EWS::FLI1-mutant mice to stable transformation and tumorigenicity. We identified a modular mechanism of IGF-1-driven tumor promotion in the early steps of EwS pathogenesis, in which Yap1 plays a central role. Pharmacologic Yap1/Tead inhibition reversed the transformed phenotype of EWS::FLI1 expressing cells. Our data provide a rationale for combined IGF-1R and YAP/TEAD inhibition in the treatment of EwS patients. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=185 SRC="FIGDIR/small/603565v1_ufig1.gif" ALT="Figure 1"> View larger version (87K): org.highwire.dtl.DTLVardef@a6a298org.highwire.dtl.DTLVardef@1fcab8aorg.highwire.dtl.DTLVardef@1621e27org.highwire.dtl.DTLVardef@a9994e_HPS_FORMAT_FIGEXP M_FIG C_FIG

cancer biology↗

bioRxiv2021

Automated compound testing in zebrafish xenografts identifies combined MCL-1 and BCL-XL inhibition to be effective against Ewing sarcoma

Ewing sarcoma is a pediatric bone and soft tissue cancer for which new therapies to improve disease outcome and to reduce adverse effects of current standard treatments are urgently needed. To identify new and effective drugs, phenotypic drug screening has proven to be a powerful method and a cancer model ideally suited for this approach is the larval zebrafish xenograft system. Complementing mouse xenografts, zebrafish offer high-througput screening possibilities in an intact complex vertebrate organism. Here, we generated Ewing sarcoma xenografts in zebrafish larvae and established a workflow for automated imaging of xenografts, tumor cell recognition within transplanted zebrafish and quantitative tumor size analysis over consecutive days by high-content imaging. The increased throughput of our in vivo screening setup allowed us to identify combination therapies effective against Ewing sarcoma cells. Especially, combined inhibition of MCL-1 and BCL-XL, two anti-apoptotic proteins, was highly efficient at eradicating tumor cells in our zebrafish xenograft assays with two Ewing sarcoma cell lines and with patient-derived cells. Transcriptional analysis across Ewing sarcoma cell lines and tumors revealed that MCL-1 and BCL2L1, coding for BCL-XL, are the most abundantly expressed anti-apoptotic genes, suggesting that combined MCL-1/BCL-XL inhibition might be a broadly applicable strategy for Ewing sarcoma treatment.

cancer biology↗

bioRxiv2017

Synthetic lethality between the cohesin subunits STAG1 and STAG2 in diverse cancer contexts

Recent genome analyses have identified recurrent mutations in the cohesin complex in a wide range of human cancers. Here we demonstrate that the most frequently mutated subunit of the cohesin complex, STAG2, displays a strong synthetic lethal interaction with its paralog STAG1. Mechanistically, STAG1 loss abrogates sister chromatid cohesion in STAG2 mutated but not in wild-type cells leading to mitotic catastrophe, defective cell division and apoptosis. STAG1 inactivation inhibits the proliferation of STAG2 mutated but not wild-type bladder cancer and Ewing sarcoma cell lines. Restoration of STAG2 expression in a mutated bladder cancer model alleviates the dependency on STAG1. Thus, STAG1 and STAG2 support sister chromatid cohesion to redundantly ensure cell survival. STAG1 represents a vulnerability of cancer cells carrying mutations in the major emerging tumor suppressor STAG2 across different cancer contexts. Exploiting synthetic lethal interactions to target recurrent cohesin mutations in cancer, e.g. by inhibiting STAG1, holds the promise for the development of selective therapeutics.

cancer biology↗

bioRxiv2017

Hardwired synthetic lethality within the cohesin complex in human cancer cells

Recent genome analyses have identified recurrent mutations in the cohesin complex in a wide range of human cancers. Here we demonstrate that the most frequently mutated subunit of the cohesin complex, STAG2, displays a strong synthetic lethal interaction with its paralog STAG1. Mechanistically, STAG1 loss abrogates sister chromatid cohesion in STAG2 mutated but not in wild-type cells leading to mitotic catastrophe, defective cell division and apoptosis. STAG1 inactivation inhibits the proliferation of STAG2 mutated but not wild-type bladder cancer and Ewing sarcoma cell lines. Restoration of STAG2 expression in a mutated bladder cancer model alleviates the dependency on STAG1. Thus, STAG1 and STAG2 act redundantly to support sister chromatid cohesion and cell survival. STAG1 represents a hardwired, context independent vulnerability of cancer cells carrying mutations in the major emerging tumor suppressor STAG2. Exploiting synthetic lethal interactions to target recurrent cohesin mutations in cancer, e.g. by inhibiting STAG1, holds the promise for the development of selective therapeutics.

cell biology↗