Biology subjects

Drake, R. R.

Publications and source records attributed to Drake, R. R..

7 recordsLinked to original sources

bioRxiv2026

Different spatial profiles of aberrant N-glycans in pediatric and adult MOGHE brain tissue

Mild malformation of cortical development with oligodendroglial hyperplasia in epilepsy (MOGHE) is a recently recognized epilepsy-associated lesion frequently linked to brain-restricted somatic variants in SLC35A2, a gene encoding the Golgi UDP-galactose transporter. Although previous studies demonstrated altered glycosylation in SLC35A2-mutated MOGHE tissue, the spatial relationship between glycosylation defects and histopathological abnormalities remains poorly understood. We applied matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI) using formalin-fixed paraffin-embedded brain tissue from six histologically confirmed MOGHE cases (three pediatric and three adult) and three temporal lobe epilepsy with hippocampal sclerosis (TLE-HS). We spatially evaluated N-glycan profiles across diagnostic tissue groups, with particular attention to molecular differences between lesional and perilesional regions and to recurrent abundance trends. All MOGHE cases harboured somatic SLC35A2 variants. Histologically, oligodendroglial hyperplasia and heterotopic neurons were present in all cases, while patchy hypomyelination was restricted to pediatric cases. Unsupervised spatial segmentation, integrated with neuropathological evaluation, revealed marked molecular heterogeneity in pediatric MOGHE. In these cases, lesional and perilesional regions were clearly distinguishable in both white matter (WM) and overlying grey matter (GM) boundaries patterns, whereas adult MOGHE and TLE-HS mainly showed a clearcut separation between WM and GM. Spatial analysis confirmed enrichment of the previously reported aberrant N-glycan species m/z 2094 and, to a lesser extent, m/z 2297 within MOGHE tissue, particularly in pediatric lesional WM. Notably, the distribution of m/z 2094 closely overlapped with areas of hypomyelination. Quantitative trajectory analysis of 151 detected N-glycan ions identified recurrent abundance profiles. Three representative spatial patterns emerged: pediatric lesion-enriched, pediatric perilesion-enriched, and TLE-HS-enriched profiles. Pediatric lesions were characterized by increased abundance of multiantennary glycans lacking terminal galactose residues and reduced abundance of galactosylated biantennary and multiantennary structures, consistent with defective UDP-galactose transport. In contrast, adult lesional and perilesional tissues exhibited largely overlapping glycomic profiles. These findings provide the first spatially resolved evidence that glycosylation abnormalities in SLC35A2-mutated MOGHE are closely associated with lesional pathology, particularly hypomyelination, and are substantially more pronounced in pediatric than adult cases. Spatial glycomics may therefore offer new insights into MOGHE pathophysiology and support the development of targeted therapeutic approaches aimed at correcting galactosylation defects.

neuroscience↗

bioRxiv2026

Extracellular Matrix Proteomic Signatures Associate with Disease-Free Survival in Later Events of Ductal Carcinoma In Situ or Invasive Breast Cancer

BackgroundDuctal carcinoma in situ (DCIS) is a noninvasive breast lesion with variable risk of progression to invasive breast cancer (IBC). Current transcription and cell marker investigations suggest ECM decreases in later events but are limited in details of ECM proteomic composition, including post-translational modifications. We investigated whether the extracellular matrix (ECM) proteome alters with later breast events of DCIS or IBC. MethodsECM-targeted mass spectrometry imaging and liquid chromatography-tandem mass spectrometry (LC-MS/MS) were applied to ten tissue microarrays from the Resource of Archival Human Breast Tissue cohort (RAHBT). Primary DCIS specimens (n=136) were analyzed in relation to later events of DCIS (n=40) or IBC(n=30), with a mean follow-up of 192.1 months 95% CI [179.1,205.1]. Statistical modeling, survival analyses, and exploratory machine learning approaches were used to identify ECM peptide signatures associated with later events. ResultsDistinct ECM peptide profiles were associated with later events of DCIS or IBC. Fifteen peptides derived from fibrillar collagens (COL1A1, COL1A2, COL3A1) and elastin, showed significantly reduced abundance in patients who developed IBC. Lower expression of specific collagen peptides associated with overall 19.9% 95% CI [17.92, 21.81] decreased disease-free survival for IBC. Lower expression of these peptides was significantly associated with reduced disease-free survival (age-adjusted hazard ratio [HR] = 2.45, 95% CI: 2.33-2.57; P < 0.05). Patient-matched samples of primary DCIS, later DCIS, and later invasive breast cancer further demonstrated reduction in ECM peptide detection. Exploratory predictive modeling from patient-matched samples achieved high performance (AUROC >0.98, accuracy >93%) in distinguishing primary from later events. Following prior work in the RAHBT cohort, reduction of certain collagen peptides was also observed in primary DCIS samples from higher risk patient groups. ConclusionsECM proteomic remodeling, particularly decreases of specific collagen domains, is strongly associated with later events of DCIS and IBC. These findings highlight ECM proteome as a critical regulator of breast cancer emergence with potential as a prognosticator of risk stratification to guide clinical management of DCIS.

pathology↗

bioRxiv2025

Identification of sialic acid linkage profiles in primary liver cancers

Cholangiocarcinoma (CCA), also known as bile duct cancer, is the second most common form of primary liver cancer after hepatocellular carcinoma (HCC). Subtypes of CCA are diagnosed according to the anatomic location within the biliary tree, such as intrahepatic (iCCA) or extrahepatic (eCCA). Early diagnosis remains difficult due to the low sensitivity and specificity of current biomarkers. There has been significant interest in finding new biomarkers of CCA and much work has been performed to both discover and validate these new markers. In our previous work, we identified specific alterations in N-linked glycosylation that were associated with iCCA, which differed from non-malignant cholestasis disease and HCC. However, that analysis focused only on N-glycan structures devoid of sialic acid, as these N-glycan residues are not stable and were intentionally removed to minimize variability. Here, we explore the specificity of sialic acid residues in liver cancers, since sialic acid residues have been heavily linked to roles in tumor progression and metastasis by helping evade immunological surveillance. To circumvent the instability of sialic acid, we utilize a novel sialic acid stabilization method coupled with Matrix-Assisted Laser Desorption Ionization (MALDI) Imaging Mass Spectrometry (IMS) for detailed analysis of sialic acid linkages in iCCA tissue. We report a clear association between the linkages type of sialic acid residues with a HCC or CCA, including differentiation between CCA subtypes. Overall, we stabilize N-glycans with sialic acid residues to determine their linkages and elucidate their importance in different types of primary liver cancers and their subtypes by MALDI-IMS.

cancer biology↗

bioRxiv2025

Spatial Proteomics of the Normal Breast Collagen Stroma: Links to BI-RADS Categories and Body Mass Index

Collagen breast stroma is the basis of increased breast density and a well-established breast cancer risk factor, yet proteomic regulation of normal breast stroma remains poorly defined. This study reports spatial regulation of the collagen proteome in normal breast tissue sections annotated by clinical characteristics. Normal breast samples from the Susan G. Komen tissue bank included data on genetic ancestry (n=40 total; n=20 African ancestry; n=20 European ancestry), body-mass-index (BMI), age, and mammogram density by the Breast Imaging Reporting and Data System (BI-RADS). Multiplexed cell marker staining showed CD44 and COL1A1 markers modulated with BMI. Collagen fiber widths by second harmonic generation (SHG) showed potential contrasts in BMI categories by genetic ancestry. Targeted extracellular matrix proteomics mass spectrometry imaging showed collagen alpha-1(I) chain domain proteome was spatially heterogenous across the normal breast microenvironment with site specific post-translational modification of proline hydroxylation. Signatures computationally extracted from breast stroma reported that 47 collagen peptides distinguished BI-RADS categories (area under the receiver operating curve>0.7; p-value>0.05). Proteomic alterations were found between overweight to obese categories with strong positive associations to BMI by multivariate analysis. This study provides the first spatial analysis of the collagen proteome in normal breast within contexts of cellular markers and clinical characteristics.

cancer biology↗

bioRxiv2025

Application of post glycosylation modifying enzymes for mass spectrometry imaging of modified N-glycans in situ.

Glycans are essential components of cells and are involved in innumerable biological processes. Their structural diversity and complexity present unique analytical challenges. Glycans are comprised of various types of monosaccharides that are linked together at different positions and with varied stereochemistry. In addition, glycans are frequently decorated with a diverse set of chemical modifications, termed post-glycosylation modifications (PGMs). Characterization of PGMs is essential for a thorough understanding of glycans, however, the technical challenges and low throughput of current methodologies have limited our understanding of these modifications. Here we demonstrate a novel approach for rapid visualization of specific PGMs present in tissue N-glycans by applying PGM-targeting enzymes to mass spectrometry imaging (MSI). The method enables in situ investigation of glycans with PGMs en masse, identifying the sugar residue and position modified, as well as visualizing the spatial distribution of each modified N-glycan in tissues. As the repertoire of PGM-targeting enzymes expands, we anticipate this approach will enable a better understanding of PGM distribution within a dynamic N-glycome. This may yield both new biological insights and the potential for identification of novel disease biomarkers.

molecular biology↗

bioRxiv2025

Multi-omic landscape of human gliomas from diagnosis to treatment and recurrence

Gliomas are among the most lethal cancers, with limited treatment options. To uncover hallmarks of therapeutic escape and tumor microenvironment (TME) evolution, we applied spatial proteomics, transcriptomics, and glycomics to 670 lesions from 310 adult and pediatric patients. Single-cell analysis shows high B7H3+ tumor cell prevalence in glioblastoma (GBM) and pleomorphic xanthoastrocytoma (PXA), while most gliomas, including pediatric cases, express targetable tumor antigens in less than 50% of tumor cells, potentially explaining trial failures. Longitudinal samples of isocitrate dehydrogenase (IDH)-mutant gliomas reveal recurrence driven by tumor-immune spatial reorganization, shifting from T-cell and vasculature-associated myeloid cell-enriched niches to microglia and CD206+ macrophage-dominated tumors. Multi-omic integration identified N-glycosylation as the best classifier of grade, while the immune transcriptome best predicted GBM survival. Provided as a community resource, this study opens new avenues for glioma targeting, classification, outcome prediction, and a baseline of TME composition across all stages.

cancer biology↗

bioRxiv2024

Glycosite Mapping and in situ Mass Spectrometry Imaging of MUC2 Glycopeptides via On-slide Digestion with Mucinase StcE

Many cancers are characterized by altered mucin expression and glycosylation, although the mechanistic relationship between tumor glycosylation and disease progression is not well-defined. Herein, our goal was to map specific mucin glycoforms in diseased tissue, enabling correlation of the tumor glycan profile with malignant features. To this end, we developed a workflow implementing on-tissue digestion with mucinase StcE, followed by matrix-assisted laser desorption ionization mass spectrometry imaging (MALDI-MSI) and liquid chromatography coupled to mass spectrometry (LC-MS). To optimize our workflow, we analyzed four different mucinous carcinomas derived from colon, esophageal, and salivary gland tissue. Using this technique, we deduced the spatial distribution of StcE-generated O-glycopeptides within mucinous tumors using MALDI-IMS. Subsequent LC-MS analyses revealed the identity of different species detected in imaging experiments, in addition to comprehensively characterizing the mucinome and proteome of each tissue. Our coupled MS approach unveiled a striking mucin 2 (MUC2) expression pattern in two colorectal mucinous adenocarcinomas, in which different glycoforms clearly stratified regions within the tumor. Notably, our LC-MS experiments obtained near-complete sequence coverage over the mucin domains of MUC2, enabling glycoproteomic mapping of this canonical mucin in unprecedented depth. MUC2 glycosylation was dominated by the T and Tn antigens, with surprisingly little sialylation detected. However, O-glycans containing mono- and di-O-acetylated sialic acid were detected in low abundance. Finally, we obtained spectral evidence for an endogenous O-acetylated GalNAc, an O-glycan structure not previously reported in the literature. Overall, this proof-of-concept work underscores the potential of this technique to generate new research avenues in oncology and beyond. Significance StatementAberrant mucin expression and glycosylation are hallmarks of cancer, but how these changes promote malignant processes are not well understood. Solid tumors are highly heterogeneous in their cellular and molecular composition, and many advanced spatial techniques have emerged in recent years to study the tumor microenvironment (TME) for better understanding disease progression. Spatially resolved glycoprotein analyses typically detect either the protein or glycan components, but not both. We developed a workflow using a dual mass spectrometry approach to map the location of intact glycopeptides in mucinous tumors, enabled by on-tissue digestion with the mucin-specific protease StcE. Future applications of this method on larger patient cohorts will enhance our understanding of glycans in malignancy, identify disease biomarkers, and define therapeutic targets.

cancer biology↗