10189-CB-6 Rapid Raman Discrimination of Glioblastoma Edema, Infiltration, and Enhancement via Single-Cell and Spatial Omics

Abstract Introduction In glioblastoma(GBM), extending resection beyond the contrast-enhancing margins on MRI to include infiltrative regions improves patients prognosis. However, objective biomarker to delineate tumor infiltration remain lacking. Raman spectroscopy rapidly provides label-free molecular information, making it a promising tool for intraoperative tumor boundary assessment. We investigated whether Raman spectra could distinguish edematous, infiltrative, and contrast-enhancing GBM regions and integrated single-cell Raman spectroscopy with spatial transcriptomics (Xenium) to elucidate molecular mechanisms of spectral variations. Methods Seventeen GBM patients undergoing resection from June 2023 to March 2025 were enrolled. Navigation guided stereotactic sampling based on MRI yielded specimens from each areas; normal white matter from epilepsy surgery served as control. Tissue samples were analyzed using a Raman microscope with 532nm excitation, and spectral classification was performed via PCA and XGBoost. Single-cell suspensions were also prepared for Raman measurement and clustering. Spatial transcriptomic analysis (Xenium) was performed to validate the spectral findings. Results We obtained 17 enhancing, 8 infiltrative, 7 edematous, and 11 control samples. Tissue-level spectra showed a significant increase in carotenoid peaks(1156, 1514cm-1) and a decrease in lipid peaks(1440, 1654cm-1) along the gradient from edema to infiltration to enhancement (p < 0.05). Five-fold cross-validation yielded classification accuracies of 96.7%(normal), 71.5%(edema), 81.5%(infiltration), and 93.0%(enhancement). Single-cell Raman analyses identified three major clusters: normal cells, tumor cells, and tumor-associated macrophages (TAMs). TAMs exhibited unique spectral profiles and contributed to tissue-level spectral differences. Spatial transcriptomics confirmed upregulation of lipid transport-related genes in TAMs, corroborating the spectroscopic findings. Conclusion Raman spectroscopy enables accurate discrimination of edematous, infiltrative, and contrast-enhancing GBM regions at both tissue and single-cell levels. Integration with spatial transcriptomics revealed that TAM-mediated lipid metabolism underlies the observed spectral changes. This approach may facilitate intraoperative delineation of tumor margins and provides novel insights into the mechanisms of GBM infiltration.