Michal Hendrych

A malignant peripheral nerve sheath tumor (MPNST) is a rare, aggressive spindle-cell sarcoma that exceptionally occurs intracranially. Herein, we report a case of intracranial spindle-cell neoplasm with abundant myxoid stroma. The immunophenotype of the primary tumor did not align with any known peripheral nerve sheath tumor and the lesion did not correspond to any methylation category defined by the Heidelberg classifier with a sufficiently high classification score, although MPNST was the highest-scoring category. Over the course of repeated recurrences, the tumor acquired increased cellularity, mitotic activity and loss of H3K27me3 expression. Subsequent methylation profiling supported a diagnosis of MPNST. The tumor harbored a gain-of-function variant in NRAS p.Gly13Arg and TP53 p.Arg273Cys from the initial tumor onwards, consistent with previous reports describing activation of Ras/mitogen-activated protein kinase as an early event in MPNST tumorigenesis. These findings illustrate the diagnostic complexity of rare intracranial sarcomas with myxoid morphology and suggest that some may represent early MPNST.

Objective: Focal cortical dysplasia (FCD) is a common cause of drug‐resistant epilepsy in children and adults and exhibits substantial clinical and histopathological heterogeneity. However, morphological classification alone does not fully explain variability in epileptogenicity, disease severity, or treatment response. Methods: We aimed to define molecular mechanisms underlying FCD heterogeneity using integrated transcriptomic and regulatory analyses. RNA and small RNA sequencing (RNA‐seq) were conducted on cortical tissue resected during epilepsy surgery from 66 patients with histopathologically confirmed FCD subtypes IA ( n = 17), IIA ( n = 25), and IIB ( n = 24), along with 15 postmortem cortical control samples. Differential expression and weighted gene coexpression network analysis were used to identify coordinated transcriptional programs, and microRNA (miRNA) profiling was integrated to assess posttranscriptional regulatory mechanisms. Integrative multivariate analyses (DIABLO and sparse partial least squares discriminant analysis) were used to evaluate subtype discrimination, and findings were validated using an independent RNA‐seq dataset. Results: FCD subtypes showed distinct but partially overlapping molecular profiles. FCD IA showed a comparatively subtle transcriptional profile, dominated by reduced expression of genes involved in cortical development, neuronal signaling, ion regulation, and metabolic stress. FCD IIA displayed a modest yet coherent molecular signature involving developmental and neuronal disruption, together with metabolic stress and autophagy. FCD IIB exhibited the most extensive transcriptional remodeling, including activation of antigen presentation pathways, complement activation, cytoskeleton reorganization, synaptic alterations, and extracellular matrix organization. Differential miRNA profiling identified shared miRNAs, including miR‐320 family, miR‐27a‐5p, miR‐142‐5p, and miR‐21‐5p, alongside subtype‐specific regulatory patterns. Integration of messenger RNA and miRNA data revealed concordant alterations, indicating that miRNA dysregulation may contribute to stabilization of subtype‐specific transcriptional states. Significance: These findings demonstrate that FCD is characterized by coordinated transcriptional and posttranscriptional network alterations that extend beyond morphological classification and may contribute to variability in epileptogenicity across subtypes.

Patient‐derived tumor organoids are widely used in cancer research, yet the biological impact of tissue processing during model generation remains unclear. Fragment‐based and dissociation‐based (DIS) approaches are commonly assumed to trade fidelity for uniformity, but their molecular consequences remain incompletely defined. To address this gap, we performed a proteome‐wide comparison of fragment‐based (cut‐and‐culture [CUT]) and DIS glioblastoma organoid protocols using quantitative mass spectrometry. Organoids from multiple patient tumors were cultured under growth factor‐free or growth factor‐supplemented conditions and compared with matched primary tissue. Results show that both protocols produced technically robust glioblastoma organoids when maintained in their native media. However, CUT organoids matched the reproducibility of DIS cultures while preserving a broader extracellular matrix (ECM) repertoire and networks linked to collagen assembly, vascular support, and cell–matrix signaling. DIS cultures were biased toward exogenous basement membrane components and proliferative, growth factor‐responsive states. Across tumors, CUT organoids consistently showed greater proteomic similarity to matched primary tissue, retaining neural, glial, stromal, and extracellular features largely absent from DIS models. Taken together, fragment‐based glioblastoma organoids can be both reproducible and biologically faithful. Tissue dissociation acts as a major perturbation that reshapes ECM organization, cellular states, and tumor identity, making protocol choice a critical determinant of model fidelity and translational relevance.

Background: Patient-derived tumor organoids are widely used in cancer research, yet the biological impact of tissue processing during model generation remains unclear. Fragment-based and dissociation-based approaches are commonly assumed to trade fidelity for uniformity, but their molecular consequences remain incompletely defined. Methods: We performed a proteome-wide comparison of fragment-based (CUT) and dissociation-based (DIS) glioblastoma organoid protocols using quantitative mass spectrometry. Organoids from multiple patient tumors were cultured under growth factor–free or growth factor–supplemented conditions and compared with matched primary tissue. Results: Both protocols produced technically robust glioblastoma organoids when maintained in their native media. However, CUT organoids matched the reproducibility of DIS cultures while preserving a broader extracellular matrix repertoire and networks linked to collagen assembly, vascular support, and cell–matrix signaling. DIS cultures were biased toward exogenous basement membrane components and proliferative, growth factor–responsive states. Across tumors, CUT organoids consistently showed greater proteomic similarity to matched primary tissue, retaining neural, glial, stromal, and extracellular features largely absent from DIS models. Conclusions: Fragment-based glioblastoma organoids can be both reproducible and biologically faithful. Tissue dissociation acts as a major perturbation that reshapes extracellular matrix organization, cellular states, and tumor identity, making protocol choice a critical determinant of model fidelity and translational relevance.

Background/Objectives : Tumor–stroma ratio (TSR) and tumor-infiltrating lymphocytes (TILs) were suggested as prognostic markers in oral squamous cell carcinoma (OSCC). Identification of markers assessable in preoperative biopsies that could guide treatment planning is of great importance. This study aimed to evaluate the concordance and prognostic impact of TSR and TILs in preoperative biopsies and matched resection specimens of OSCC. Methods: This study included 100 patients with OSCC. TSR and stromal TILs were evaluated on hematoxylin and eosin-stained slides of biopsies and paired resection specimens and categorized (into low TSR and high TSR; high TILs and low TILs). The agreement between resections and biopsies, and the prognostic significance and clinicopathological correlations of TSR and TILs, were investigated. Results: For TSR, substantial agreement between preoperative biopsies and surgical specimens (kappa correlation coefficient 0.713) was demonstrated. The assessment of TILs showed poor concordance between biopsies and resections (kappa correlation coefficient 0.372). For both biopsies and resections, Cox regression showed an independent negative prognostic impact of low TSR on disease-free, disease-specific, and overall survival. Independent prognostic value of TILs evaluated in biopsies was not found, and the negative prognostic impact of low TILs on disease-free and overall survival was observed only in the main resection specimens. Conclusions: TSR evaluated in preoperative biopsies was highly concordant with results in main resection specimens and may provide significant information for OSCC prognostication, risk stratification, and treatment decisions. In contrast, TILs evaluated in biopsies showed poor concordance with main resection specimens and failed to demonstrate prognostic significance.