Biology subjects

Schueler, J.

Publications and source records attributed to Schueler, J..

10 recordsLinked to original sources

bioRxiv2026

DESI-MS-Based Analysis of Drug Distribution in Human Renal Cystic Tissue Using the Chorioallantoic Membrane (CAM) as a 3D In Vivo Model

The chorioallantoic membrane (CAM) model represents a promising three-dimensional in vivo platform for preclinical drug testing in human tissues. In this study, we investigated whether the tissue penetration and distribution of benzbromarone, a known inhibitor of the Ca2+ activated chloride channel TMEM16A and potential therapeutic agent for autosomal dominant polycystic kidney disease (ADPKD), can be successfully visualized in human renal cyst tissue cultured on the CAM. To this end, desorption electrospray ionization mass spectrometry imaging (DESI-MSI) combined with an ultrahigh-resolution time-of-flight mass spectrometer was employed. We achieved spatially resolved molecular mapping of endogenous metabolites and lipids as well as the applied compound. MSI enabled clear differentiation between CAM and cystic tissue based on their distinct lipid profiles. Benzbromarone was reproducibly detected in the cyst specimens and exhibited selective accumulation along the cyst epithelium, which is considered the principal site of action. These observations were complemented by multivariate analyses including Uniform Manifold Approximation and Projection (UMAP), and sparse multinomial logistic zero-sum classification. The data-driven approach confirmed molecular differences between tissue types and allowed accurate classification of drug-treated and untreated regions. This study demonstrates that topically applied benzbromarone penetrates human renal cyst tissue in the CAM model and localizes to pharmacologically relevant tissue regions, notably the location of the Ca2+ activated chloride channel TMEM16A in the epithelial lining. The integration of high-resolution DESI-MSI with advanced statistical analysis provides a robust and label-free method to study drug distribution in human tissue grafts. Our findings contribute to the advancement of translational research in analytical chemistry and pharmacology.

biochemistry↗

bioRxiv2026

Sensitivity profiling reveals consistent drug responses across preclinical neuroblastoma models

Despite intensive treatment, overall survival for high-risk and relapse neuroblastoma patients remains below 50%. Even though comprehensive molecular profiling enables treatment stratification, druggable alterations have been identified for only a subset of patients. In vitro drug screening offers a complementary approach. Here, we compare the translational potential of three preclinical drug screening methods: ex vivo short-term, in vitro patient-derived organoid and in vivo patient-derived xenograft (PDX) drug testing. In total, 55 screens were performed from 38 neuroblastoma samples and five pediatric non-malignant samples, testing 77-224 drugs per screen. Ex vivo short-term drug screens achieved higher success rates than organoid screens (65% versus 23%) and shorter turnaround times (14 days versus 3-12 months). Matched samples showed consistent drug sensitivities across sample origin (patient versus PDX-derived; mean r = 0.84) and method (ex vivo short-term versus organoids; mean r = 0.87), demonstrating that ex vivo short-term screens recapitulate drug sensitivities found in long-term organoid models. In parallel, as part of the ITCC-P4 consortium, ten compounds were tested in vivo in eight PDX models, with samples matching the ex vivo screens. For seven out of ten clinically available compounds, ex vivo drug responses were comparable with in vivo responses in matched PDX models. These results demonstrate that, while organoids and PDX models remain essential for drug discovery, ex vivo short-term drug screening provides a rapid alternative for functional precision oncology in neuroblastoma.

cell biology↗

bioRxiv2024

Spatial Proteomics by Trapped Ion Mobility supported MALDI MS/MS Imaging: A First Glance into Multiplexed and Spatial Peptide Identification

RATIONALEIn spatial proteomics, matrix-assisted laser desorption/ionization (MALDI) imaging enables rapid and cost-effective peptide measurements. Yet, in situ peptide identification remains challenging. Therefore, this study aims to integrate the trapped ion mobility spectrometry (TIMS)-based parallel accumulation-serial fragmentation (PASEF) into MALDI imaging of tryptic peptides to enable multiplexed MS/MS imaging. METHODSAn initial MALDI TIMS MS1 survey measurement was performed, followed by a manual generation of a precursor list containing mass over charge values and ion mobility windows. Inside the dual TIMS system, submitted precursors were trapped, separately eluted by their ion mobility and analyzed in a quadrupole time-of-flight device, thereby enabling multiplexed MALDI MS/MS imaging. Finally, precursors were identified by peptide to spectrum matching. RESULTSThis study presents the first multiplexed MALDI TIMS MS/MS imaging (iprm-PASEF) of tryptic peptides. Its applicability was showcased on two histomorphologically distinct tissue specimens in a 4-plex and 5-plex setup. Precursors were successfully identified by the search engine MASCOT in one single MALDI imaging experiment for each respective tissue. Peptide identifications were corroborated by liquid-chromatography tandem mass spectrometry experiments and fragment co-localization analyses. CONCLUSIONSIn this study, we present a novel pipeline, based on iprm-PASEF that allows the multiplexed and spatial identification of tryptic peptides in MALDI imaging. Hence, it marks a first step towards the integration of MALDI imaging into the emerging field of spatial proteomics.

biochemistry↗

bioRxiv2024

Proteomic Characterization of Intrahepatic Cholangiocarcinoma Identifies Distinct Subgroups and Proteins Associated with Time-To-Recurrence

Background & AimsIntrahepatic cholangiocarcinoma (ICC) is a poorly understood cancer with dismal survival and high recurrence rates. ICCs are often detected in advanced stages. Surgical resection is the most important first-line treatment but limited to non-advanced cases, whereas chemotherapy provides only a moderate benefit. The proteome biology of ICC has only been scarcely studied and the prognostic value of initial ICCs proteomic features for the time-to-recurrence (TTR) remains unclear. MethodsWe dissected formalin-fixed, paraffin-embedded samples from 80 tumor- and 77 matching adjacent non-malignant (TANM) tissues. All samples were measured via liquid-chromatography mass-spectrometry (LC-MS/MS) in data independent acquisition mode (DIA). ResultsTumor- and TANM tissue showed strongly different biologies and DNA-repair, translation, and matrisomal processes were upregulated in ICC. In a hierarchical clustering analysis, we determined two proteomic subgroups of ICC, which showed significantly diverging TTRs. Cluster 1, which is associated with a beneficial prognosis, was enriched for matrisomal processes and proteolytic processing, while cluster 2 showed increased RNA and protein turnover. In a second, independent Cox proportional hazards model analysis, we identified individual proteins whose expression correlates with TTR distribution. Proteins with a positive hazard ratio were mainly involved in carbon/glucose metabolism and protein turnover. Conversely, proteins associated with a low hazard ratio were mostly linked to the extracellular matrix. Additional proteome profiling of patient-derived xenograft tumor models of ICC successfully distinguished tumor and stromal proteins and provided insights into cell-matrix interactions. ConclusionsWe successfully determine the proteome biology of ICC and present two proteome clusters in ICC patients with significantly different TTR rates and distinct biological motifs. A xenograft model confirmed the importance of tumor-stroma interactions for this cancer.

cancer biology↗

bioRxiv2024

10x GENOMICS GENE EXPRESSION FLEX IS A POWERFUL TOOL FOR SINGLE-CELL TRANSCRIPTOMICS OF XENOGRAFT MODELS

The 10x Genomics Gene Expression Flex protocol allows profiling of fixed or frozen material, greatly simplifying the logistics of sample collection, storage and transfer prior to single -cell sequencing. The method makes single-cell transcriptomics possible for existing fresh-frozen or FFPE tissue samples, but also facilitates the logistics of the sampling process, allowing instant preservation of samples. The technology relies on species-specific probes available for human and mouse. Nevertheless, processing of patient-derived (PDX) or cell line (CDX) xenografts, which contain mixed human and mouse cells, is currently not supported by this protocol due to the high degree of homology between the probe sets. Here we show that it is feasible to simultaneously profile populations containing both human and mouse cells by mixing the transcriptome probe sets of both species. Cellranger outputs a count table for each of the species allowing evaluation of the performance of the different probe sets. Cross-reactive probes are greatly outperformed by the specific probe hybridizations leading to a clear difference in the recovery of UMIs and unique genes per cell. Furthermore, we developed a pipeline that removes cross-reactive signal from the data and provides species-specific count tables for further downstream analysis. Hence, the 10x Genomics Gene Expression Flex protocol can be used to process xenograft samples without the need for separation of human and mouse cells by flow sorting and allows analysis of the human and mouse single-cell transcriptome from each sample. We anticipate it will be increasingly used for single-cell sequencing of cancer cell line and patient-derived xenografts, facilitating the preservation of the samples and allowing the interrogation of both the (human) xenograft and the (mouse) tumor microenvironment at single-cell resolution.

genomics↗

bioRxiv2023

Integrative ensemble modelling of cetuximab sensitivity in colorectal cancer PDXs

AO_SCPLOWBSTRACTC_SCPLOWPatient-derived xenografts (PDXs) are tumour fragments engrafted into mice for preclinical studies. PDXs offer clear advantages over simpler in vitro cancer models - such as cancer cell lines (CCLs) and organoids - in terms of structural complexity, heterogeneity, and stromal interactions. We characterised 231 colorectal cancer PDXs at the genomic, transcriptomic, and epigenetic level and measured their response to cetuximab, an EGFR inhibitor in clinical use for metastatic colorectal cancer. After assessing PDXs quality, stability, and molecular concordance with publicly available patient cohorts, we trained, interpreted, and validated an integrated ensemble classifier (CeSta) which takes in input the PDXs multi-omic characterisation and predicts their sensitivity to cetuximab treatment (AUROC > 0.9). Our study shows that large PDX collections can be used to train accurate, interpretable models of drug sensitivity, which 1) better recapitulate patient-derived therapeutic biomarkers than other models trained on CCL data, 2) can be robustly validated across independent PDX cohorts, and 3) can be used for the development of novel therapeutic biomarkers.

bioinformatics↗

bioRxiv2023

Deprivation of arginine and lysine interferes with growth of patient-derived tumor xenografts

In recent years, a novel treatment method for cancer emerged, which is based on the starvation of tumors of amino acids like arginine. The deprivation of arginine in serum is based on enzymatic degradation and can be realized by arginine deaminases like the L-amino acid oxidase found in the ink toxin of the sea hare Aplysia punctata. Previously isolated from the ink, the L-amino acids oxidase was described to oxidate the essential amino acid L-lysine and L-arginine to their corresponding deaminated alpha-keto acids. Here, we present the recombinant production and functionalization of amino acid oxidase Aplysia Punctata ink toxin (APIT). PEGylated APIT (APIT-PEG) increased the blood circulation time. APIT-PEG treatment of patient-derived xenografted mice shows a significant dose dependent reduction of tumor growth over time mediated by amino acid starvation of the tumor. Treatment of mice with APIT-PEG which lead to deprivation of arginine was well tolerated.

cancer biology↗

bioRxiv2020

Docosahexaenoic acid significantly reduces tumor cell growth in preclinical models of ovarian cancer

Despite recent advances in diagnosis and treatment, ovarian cancer (OC) is the most lethal gynecological malignancy and improving the efficacy of chemotherapy is of great interest. This study increases our understanding of how dietary intervention with docosahexaenoic acid (DHA)-a supplement proven safe for human consumption - enhances the anti-cancer effects of conventional chemotherapy. Our results demonstrated synergistic cell killing by DHA and carboplatin in OC cell lines. Furthermore, DHA supplementation alone and in combination with carboplatin significantly reduced OC growth in a high-grade serous OC patient-derived xenograft mouse model. Carboplatin administered intraperitoneally significantly reduced tumor growth in DHA-fed mice compared to mice on the control diet. Intravenous carboplatin administration in combination with DHA reduced tumor growth similarly to carboplatin or DHA monotherapies. The DHA-induced reduction in tumor growth in this model was associated with increased tumor necrosis and improved survival. As such, our findings provide a strong rationale to move to clinical trials that will determine whether DHA supplementation enhances the efficacy and tolerance of cytotoxic chemotherapy in patients with OC.

cancer biology↗

bioRxiv2020

ADAM protease inhibition overcomes resistance of breast cancer stem-like cells to γδ T cell immunotherapy

Breast cancer stem cells (BCSC) are highly resistant to current therapies, and are responsible for metastatic burden and relapse. Gamma delta T cells ({gamma}{delta}Tc) are immunosurveillance cells with tremendous anti-tumoral activity, and a growing number of clinical trials have confirmed the safety of {gamma}{delta}Tc immunotherapy for various malignancies. Herein, we demonstrate that {gamma}{delta}Tc can kill BCSC, but to a lesser extent than non-cancer stem cells (NSC). Immune evasion was orchestrated by several mechanisms. The BCSC secretome rendered {gamma}{delta}Tc hypo-responsive by reducing proliferation, cytotoxicity and IFN-{gamma} production, while increasing expression of co-inhibitory receptors on {gamma}{delta}Tc. BCSC and target cells surviving {gamma}{delta}Tc cytotoxicity had higher PD-L1 co-inhibitory ligand expression, and blocking PD-1 on {gamma}{delta}Tc significantly overcame BCSC resistance to {gamma}{delta}Tc killing. Fas/FasL signaling was dysfunctional in BCSC due to upregulation of the anti-apoptotic protein MCL-1, which could be partially overcome using dMCL1-2, an MCL-1 degrader. Moreover, the BCSC fraction shed higher levels of the NKG2D ligand MICA compared to NSC. Inhibiting MICA shedding using the ADAM inhibitor GW280264X overcame BCSC resistance to {gamma}{delta}Tc killing, rendering BCSC as sensitive to {gamma}{delta}Tc cytotoxicity as NSC. Collectively, our data unravel multiple mechanisms exploited by BCSC to evade {gamma}{delta}Tc killing, which may also come into play in BCSC resistance to other cytotoxic lymphocytes. Developing strategies to overcome this resistance will increase the efficacy of cancer immunotherapy and lead to improved outcomes for cancer patients. One Sentence SummaryBreast cancer stem-like cells are resistant to {gamma}{delta} T cell targeting, which can be overcome by inhibiting ADAM proteases that facilitate MICA/B shedding.

immunology↗

bioRxiv2019

Translational control of breast cancer plasticity

Plasticity of neoplasia, whereby cancer cells attain stem-cell-like properties, is required for disease progression and represents a major therapeutic challenge. We report that in breast cancer cells NANOG, SNAIL and NODAL transcripts manifest multiple isoforms characterized by different 5 Untranslated Regions (5UTRs), whereby translation of a subset of these isoforms is stimulated under hypoxia. This leads to accumulation of corresponding proteins which induce plasticity and \"fate-switching\" toward stem-cell like phenotypes. Surprisingly, we observed that mTOR inhibitors and chemotherapeutics induce translational activation of a subset of NANOG, SNAIL and NODAL mRNA isoforms akin to hypoxia, engendering stem cell-like phenotypes. Strikingly, these effects can be overcome with drugs that antagonize translational reprogramming caused by eIF2 phosphorylation (e.g. ISRIB). Collectively, our findings unravel a hitherto unappreciated mechanism of induction of plasticity of breast cancer cells, and provide a molecular basis for therapeutic strategies aimed at overcoming drug resistance and abrogating metastasis.

cancer biology↗