Dr Miller on Whole Genome Sequencing of CSF ctDNA in High-Grade Glioma

“[W]e were able to see completely different mutational signatures in the CSF and in the tumor, which suggests that maybe what we’re seeing in the CSF is really early evidence of leptomeningeal disease, which has big therapeutic implications.”

Alexandra M. Miller, MD, an associate professor in the Departments of Neurology, Medicine, and Neurosurgery at New York University Grossman School of Medicine, co-director of the Brain and Spine Tumor Center, and chief of the Neuro-Oncology Program at New York University Langone Health, discussed findings on whole genome sequencing of circulating tumor DNA in cerebrospinal fluid from patients with high-grade glioma, presented at the 2026 SNO Annual Meeting.

Targeted next-generation sequencing panels on cerebrospinal fluid (CSF) are a great technique that are often used to track tumor alterations, Miller said. However, she explained that whole genome sequencing (WGS) is better for understanding evolutionary changes and shifts in mutational signatures. For example, patients with glioblastoma treated with temozolomide (Temodar) commonly develop resistance via hypermutation, which is best identified through mutational signature patterns, she explained. WGS has historically also performed better in detecting copy number alterations, many of which are disease defining, according to Miller.

The current analysis included patients with high-grade glioma, of whom most had IDH–wild-type disease. Among the patients with a CSF tumor fraction of at least 10%, most oncogenic CSF mutations were detected by both next-generation sequencing and WGS, and WGS captured several additional mutations, including CDKN2A homozygous deletions and chromosome 7 gain/10 loss.

The samples were collected from patients with no clinical evidence that the tumor had spread into the CSF, Miller noted. Despite this, completely different mutational signatures were seen in the CSF and the tumor, which may reflect early leptomeningeal disease, she said; clonal reconstruction in certain patients suggested that a clone seeded the CSF before temozolomide treatment.

Miller outlined 4 uses of CSF sequencing. The first is diagnosis, identifying a World Health Organization–defining alteration when biopsy is not possible or recommended. The second is monitoring through serial sequencing over time. The third is better understanding tumor biology, including mechanisms of clonal evolution and therapeutic resistance. The fourth is prognosis, as detectable circulating tumor DNA is a negative sign that might warrant escalating therapy or increasing screening frequency, Miller concluded.