Abstract Medulloblastoma is a rapidly growing embryonal-CNS tumor with a 5-year survival-rate of nearly 0% in recurrent-cases. Current non-invasive monitoring relies primarily on imaging, which lacks the necessary sensitivity and fails to capture molecular features associated with tumor progression. Liquid-biopsies have emerged as a less invasive alternative that studies serially collectible sources of tumor-derived biomolecules. Relapsed Group 4 medulloblastomas are particularly challenging due to their poor prognosis and lack-of-molecular-biomarkers to predict relapse. This study aims to establish robust-assays for monitoring tumor progression, recurrence, and treatment response using liquid-biopsies to guide rational therapeutic interventions. We used patient-derived cerebrospinal-fluid (CSF) and plasma to analyze longitudinal genetic, immunological, and metabolic-profiles with nanopore-sequencing, Mesoscale-Discovery (MSD), and LC-MS-based amino-acid profiling. Nanopore-sequencing assay detected copy-number-variation (CNVs) and distinct methylation patterns in group 4 medulloblastoma. Optimized protocols yielded high-purity cfDNA (64-97%; 20-376 ng) from just 500µL CSF, enabling consistent detection of tumor-derived-CNVs and methylation profiles, even at shallow-sequencing depths. Comparison against the gold-standard Illumina-EPICarray validated the effectiveness of nanopore-sequencing in generating similar CNV profiles using only 1/10th of cfDNA input. Matched-tumor-tissue vs CSF-CNV analysis highlighted the assay’s sensitivity in capturing further tumor heterogeneity. Additionally, we optimized MSD assays for immune-response-monitoring and characterized amino-acid changes during tumor-transformation and treatment. The standardized MSD assay demonstrated a low pg/mL detection-limit and a 7-fold-dynamic detection-range, using only 25µL plasma. Longitudinal-immune- profiling in five medulloblastoma patients demonstrated an expected elevation in proinflammatory-cytokines in Group 3/4 patients undergoing immunotherapy. Meanwhile, metabolomic analysis indicated glutamic-acid and s-sulfocysteine as potential biomarkers of disease recurrence which influence tumor cell survival and proliferation. This integrated-approach highlights the utility of such assays for monitoring progression and treatment-response using serial-liquid-biopsies, paving-the-way for precision-oncology in medulloblastoma-care. However, further investigation on a larger cohort will help to establish statistical significance
Singh et al. (Fri,) studied this question.