Biology subjects

Schweizer, L.

Publications and source records attributed to Schweizer, L..

9 recordsLinked to original sources

bioRxiv2025

AI-powered Deep Visual Proteomics reveals critical molecular transitions in pancreatic cancer precursors

Pancreatic ductal adenocarcinoma (PDAC) evolves through non-invasive precursor lesions, yet its earliest molecular events remain unclear. We established the first spatially resolved proteomic atlas of these lesions using Deep Visual Proteomics (DVP). AI-driven computational pathology classified normal ducts, acinar-ductal metaplasia (ADM), and pancreatic intraepithelial neoplasia (PanIN) from cancer-free organ donors (incidental, "iPanINs") and PDAC patients (cancer-associated, "cPanINs"). Laser microdissection of 96 discrete regions containing as few as 100 phenotypically matched cells and ultrasensitive mass spectrometry quantified a total of 8,512 proteins from formalin-fixed tissues. Distinct molecular signatures stratifying cPanINs from iPanINs, and remarkably, many cancer-associated proteins already marked histologically normal epithelium. Four core programs - stress adaptation, immune engagement, metabolic reprogramming, mitochondrial dysfunction - emerged early and intensified during progression. By integrating DVP with AI-guided tissue annotation, we demonstrate that molecular reprogramming precedes histological transformation, creating opportunities for earlier detection and interception of a near-uniformly lethal cancer. SignificanceOur spatially-resolved proteomics atlas uncovers distinct molecular signatures in pancreatic cancer adjacent precursor lesions, clearly diverging from those in incidental, cancer-free pancreatic lesions. Our deep proteomics dataset offers a valuable resource for identifying novel biomarkers and therapeutic targets, informed by the earliest cancer-associated molecular events in archival pancreatic tissues.

cancer biology↗

bioRxiv2023

Cytosolic retention of the mitochondrial protease HtrA2 during mitochondrial protein import stress (MPIS) triggers the DELE1-HRI pathway

Mitochondrial stress inducers, such as the proton ionophore carbonyl cyanide m-chlorophenyl hydrazone (CCCP) and the ATPase pump inhibitor oligomycin, trigger the DELE1-HRI branch of the integrated stress response (ISR) pathway. Previous studies performed using epitope-tagged forms of DELE1 showed that these stresses induced the accumulation of a cleaved form of DELE1, DELE1-S, which stimulates HRI. Here, we report that mitochondrial protein import stress (MPIS) is an overarching stress that triggers the DELE1-HRI pathway, and that endogenous DELE1 could be cleaved into two forms, DELE1-S and DELE1-VS, the latter accumulating only upon non- depolarizing MPIS. We further showed that DELE1 specifically senses MPIS triggered by the inhibition of the TIM23 complex at the inner mitochondrial membrane (IMM). While MPIS can also cause mitophagy induction through engagement of the NLRX1-RRBP1 pathway, we observed that DELE1-HRI and NLRX1-RRBP1 signaling were engaged independently upon MPIS. Surprisingly, our results suggest that in our cellular model the mitochondrial protease OMA1 was dispensable for DELE1 cleavage upon MPIS. Instead, we identified a key role for another mitochondrial protease, HtrA2, in mediating the cleavage of DELE1 into DELE1-S and DELE1- VS. Our data further suggest that DELE1 is likely cleaved into DELE1-S by HtrA2 in the cytosol, while the DELE1-VS form might be generated during halted translocation of the protein into mitochondria. Together, this study identifies MPIS as the overarching stress detected by DELE1 and identifies HtrA2 as a critical protease involved in DELE1 processing.

cell biology↗

bioRxiv2023

A standardized and reproducible workflow for membrane glass slides in routine histology and spatial proteomics

Defining the molecular phenotype of single cells in-situ is essential for understanding tissue heterogeneity in health and disease. Powerful imaging technologies have recently been joined by spatial omics technologies, promising unparalleled insights into the molecular landscape of biological samples. One approach involves laser microdissection in combination with membrane glass slides for the isolation of single cells from specific anatomical regions for further analysis by spatial omics. However, so far this is not fully compatible with automated staining platforms and routine histology procedures such as heat-induced epitope retrieval, limiting reproducibility, throughput and integration of advanced staining procedures. This study describes a robust workflow for routine use of glass membrane slides, allowing precise extraction of tissue in combination with automated and multicolor immunofluorescence staining. The key advance is the addition of glycerol to standard heat-induced epitope retrieval protocol, preventing membrane distortion while preserving antigen retrieval properties. Importantly, we show that glycerol is fully compatible with mass-spectrometry based proteomics and does not affect proteome depth or quality. Further, we enable single focal plane imaging by removing remaining trapped air pockets with an incision. We demonstrate our workflow using the recently introduced Deep Visual Proteomics technology on the single-cell type analysis of adjacent suprabasal and basal keratinocytes of human skin. Our protocol extends the utility of membrane glass slides and enables much more robust integration with routine histology procedures, high-throughput multiplexed imaging and sophisticated downstream spatial omics technologies.

pathology↗

bioRxiv2022

Spatial single-cell mass spectrometry defines zonation of the hepatocyte proteome

Single-cell proteomics by mass spectrometry (MS) is emerging as a powerful and unbiased method for the characterization of biological heterogeneity. So far, it has been limited to cultured cells, whereas an expansion of the method to complex tissues would greatly enhance biological insights. Here we describe single-cell Deep Visual Proteomics (scDVP), a technology that integrates high-content imaging, laser microdissection and multiplexed MS. scDVP resolves the context-dependent, spatial proteome of murine hepatocytes at a current depth of 1,700 proteins from a slice of a cell. Half of the proteome was differentially regulated in a spatial manner, with protein levels changing dramatically in proximity to the central vein. We applied machine learning to proteome classes and images, which subsequently inferred the spatial proteome from imaging data alone. scDVP is applicable to healthy and diseased tissues and complements other spatial proteomics or spatial omics technologies.

systems biology↗

bioRxiv2021

Integrated phospho-proteogenomic and single-cell transcriptomic analysis of meningiomas establishes robust subtyping and reveals subtype-specific immune invasion

Meningiomas are the most frequent primary intracranial tumors. They can follow a wide clinical spectrum from benign to highly aggressive clinical course. No specific therapy exists for refractory cases or cases not amenable to resection and radiotherapy. Identification of risk of recurrence and malignant transformation for the individual patients is challenging. However, promising molecular markers and prognostic subgrouping by DNA methylation are emerging. Still, the biological underpinnings of these diagnostic subgroups are elusive, and, consequently, no novel therapeutic options arise thereof. Here we establish robust subgroups across the full landscape of meningiomas, consistent through DNA methylation, mutations, the transcriptomic, proteomic and phospho-proteomic level. Pronounced proliferative stress and DNA damage repair signals in malignant cells and in clusters exclusive to recurrent tumors are in line with their higher mitotic activity, but also provide an explanation for the accumulation of genomic instability in anaplastic meningiomas. Although homozygous deletion of CDKN2A/B is a diagnostic marker of high-grade meningioma, the expression of its gene product increased from low to non-deleted high-grade cases. Differences between subgroups in lymphocyte and myeloid cell infiltration, representing a majority of tumor mass in low-grade NF2 tumors, could be assigned to cluster-specific interaction with tumor cells. Activation to a more proinflammatory phenotype and decreased infiltration of myeloid cells in high-grade cases correlated with lower expression of CSF1, located on chromosome arm 1p, whose deletion is known as prognostic marker, with no proposed mechanism before. Our results demonstrate a robust molecular subclassification of a tumor type across multiple layers, provide insight into heterogeneous growth dynamics despite shared pathognomonic mutations, and highlight immune infiltration modulation as a novel target for meningioma therapy.

genomics↗

bioRxiv2021

Recurrent fusions in PLAGL1 define a distinct subset of pediatric-type supratentorial ependymoma

Ependymomas encompass a heterogeneous group of central nervous system (CNS) neoplasms that occur along the entire neuroaxis. In recent years, extensive (epi-)genomic profiling efforts have identified several molecular groups of ependymoma that are characterized by distinct molecular alterations and/or patterns. Based on unsupervised visualization of a large cohort of genome-wide DNA methylation data, we identified a highly distinct group of pediatric-type tumors (n = 40) forming a cluster separate from all established CNS tumor types, of which a high proportion were histopathologically diagnosed as ependymoma. RNA sequencing revealed recurrent fusions involving the pleomorphic adenoma gene-like 1 (PLAGL1) gene in 19 of 20 of the samples analyzed, with the most common fusion being EWSR1:PLAGL1 (n = 13). Five tumors showed a PLAGL1:FOXO1 fusion and one a PLAGL1:EP300 fusion. High transcript levels of PLAGL1 were noted in these tumors, with concurrent overexpression of the imprinted genes H19 and IGF2, which are regulated by PLAGL1. Histopathological review of cases with sufficient material (n = 16) demonstrated a broad morphological spectrum of largely ependymoma-like tumors. Immunohistochemically, tumors were GFAP-positive and OLIG2- and SOX10-negative. In 3/16 of the cases, a dot-like positivity for EMA was detected. Consistent with other fusion-positive ependymal groups, all tumors in our series were located in the supratentorial compartment. Median age of the patients at the time of diagnosis was 6.2 years. Analysis of time to progression or recurrence revealed survival times comparable to those of patients with ZFTA:RELA-fused ependymoma. In summary, our findings suggest the existence of a novel group of supratentorial ependymomas that are characterized by recurrent PLAGL1 fusions and enriched for pediatric patients.

cancer biology↗

bioRxiv2021

Single-cell immune profiling with TCR clonotype barcoding identifies biomarker signatures that predict response to immune checkpoint blockade

The identification of predictive biomarkers for patient treatment response is urgently needed to increase the probability of success of existing and novel experimental therapies. Single-cell profiling has provided novel biological insights into drug responses in the tumor microenvironment, but its potential for biomarker discovery has not been fully explored for therapeutic purposes. We describe a novel approach to discover predictive response biomarkers from single-cell data from a small patient cohort using the T cell receptor sequence intrinsic to each T cell to match clonotypes between pre- and post-treatment tumor samples. As a result, we have identified a predictive gene expression signature for immune checkpoint blockade and validated its predictive performance using data from three larger clinical studies. Our results demonstrated that applying clonotyping with single-cell genomic profiling is a promising novel approach for biomarker identification that does not require data collected from large patient cohorts. This could increase success rates, reduce clinical trial size, and significantly impact future clinical developments of immunomodulatory therapeutics.

immunology↗

bioRxiv2021

A new parallel high-pressure packing system enables rapid multiplexed production of capillary columns

Reversed-phase high performance liquid chromatography (HPLC) is the most commonly applied peptide separation technique in mass spectrometry (MS)-based proteomics. Particle-packed capillary columns are predominantly used in nano-flow HPLC systems. Despite being the broadly applied standard for many years capillary columns are still expensive and suffer from short lifetimes, particularly in combination with ultra-high-pressure chromatography systems. For this reason, and to achieve maximum performance, many laboratories produce their own in-house packed columns. This typically requires a considerable amount of time and trained personnel. Here, we present a new packing system for capillary columns enabling rapid, multiplexed column production with pressures reaching up to 3000 bar. Requiring only a conventional gas pressure supply and methanol as driving fluid, our system replaces the traditional setup of helium pressured packing bombs. By using 10x multiplexing, we have reduced the production time to just under 2 minutes for several 50 cm columns with 1.9 {micro}m particle size, speeding up the process of column production 40 to 800 times. We compare capillary columns with various inner diameters (ID) and length packed under different pressure conditions with our newly designed, broadly accessible high-pressure packing station. One sentence summaryA newly constructed parallel high-pressure packing system enables the rapid multiplexed production of capillary columns. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=162 SRC="FIGDIR/small/433033v2_ufig1.gif" ALT="Figure 1"> View larger version (50K): org.highwire.dtl.DTLVardef@4d224forg.highwire.dtl.DTLVardef@79dd3org.highwire.dtl.DTLVardef@aa047org.highwire.dtl.DTLVardef@1a0b837_HPS_FORMAT_FIGEXP M_FIG C_FIG

biochemistry↗

bioRxiv2020

High throughput functional screening for next generation cancer immunotherapy using droplet-based microfluidics

Currently high throughput approaches are lagged for isolation of antibodies whose function goes beyond simple binding, which have prevented the next generation cancer immunotherapeutics, such as bispecific T cell engager antibodies or agonist antibody of costimulatory receptor, from reaching their full potential. Here we developed a highly efficient droplet-based microfluidics platform combining with lentivirus transduction system that enables functional screening of millions of antibodies. To showcase the capacity of the system, functional antibodies for CD40 agonism with low frequency (<0.02%) were identified with 2 rounds of screening. To demonstrate its versatility, an anti-Her2/anti-CD3 bispecific antibody library was established using bispecific T cell Engager (BiTE) platform and functional screening enabled efficient identification of potent anti-Her2/anti-CD3 BiTE antibodies. The platform could revolutionize the next generation cancer immunotherapy drug development and research world.

cancer biology↗