Ensign, Shannon Fortin

Abstract Background Leptomeningeal disease (LMD) remains a challenging complication of both solid tumors and hematologic malignancies, with diagnosis traditionally relying on cerebrospinal fluid (CSF) cytology, which has limited sensitivity. Circulating tumor DNA (ctDNA) analysis in CSF has emerged as a promising diagnostic and molecular profiling tool that may improve detection and guide treatment decisions. We will report real-world clinical experience using a CSF ctDNA assay (Menarini Silicon Biosystems) in routine neuro-oncology practice. Methods Retrospective analysis of over 100 CSF specimens obtained from patients with suspected or confirmed LMD will be conducted. Clinical data to be collected: patient demographics, tumor histology, indication for testing, and prior treatments. ctDNA assay results will be compared with conventional CSF cytology. Molecular alterations identified through ctDNA testing will be recorded, along with subsequent treatment decisions, including use of targeted therapies, systemic therapy modifications, radiation therapy, or intrathecal approaches. Preliminary clinical outcomes, including radiographic response, progression patterns, and survival trends, will be explored. Descriptive statistical methods will be used to summarize the findings. Results We anticipate that among analyzed specimens, ctDNA testing will provide clinically meaningful diagnostic yield across multiple tumor histologies. Concordance and discordance patterns between ctDNA results and conventional cytology will be evaluated, including cases in which ctDNA identified molecular alterations despite negative cytology. Actionable genomic findings in a subset of patients will be analyzed and presented. Early outcome observations that ctDNA-informed management may facilitate more personalized care will be reported. Conclusions CSF ctDNA analysis represents a feasible and informative adjunct to conventional cytology in the evaluation of patients with suspected LMD in real-world clinical practice. Beyond diagnostic support, molecular profiling from ctDNA may influence treatment selection and enable precision oncology approaches in this high-risk population. Prospective studies are warranted to define sensitivity, specificity, and the impact of ctDNA-guided interventions on clinical outcomes.

Abstract Background Tumor heterogeneity underlies resistance and disease progression in glioblastoma (GBM), and tumors most commonly recur adjacent to the surgical resection margins in contrast non-enhancing (NE) regions. To date, no targeted therapies have meaningfully altered overall patient survival in the up-front setting. The aim of this study was to characterize intratumoral heterogeneity in recurrent GBM using bulk samples from primary resection and recurrent samples taken from contrast-enhancing (EN) and contrast NE regions. Methods Whole exome and RNA sequencing were performed on matched bulk primary and multiple recurrent EN and NE tumor samples from 16 GBM patients who received standard of care treatment alone or in combination with investigational clinical trial regimens. Results Private mutations emerge across multi-region sampling in recurrent tumors. Genomic clonal analysis revealed increased enrichment in gene alterations regulating the G2M checkpoint, Kras signaling, Wnt signaling, and DNA repair in recurrent disease. Subsequent functional studies identified augmented PI3K/AKT transcriptional and protein activity throughout progression, validated by phospho-protein levels. Moreover, a mesenchymal transcriptional signature was observed in recurrent EN regions, which differed from the proneural signature in recurrent NE regions. Conclusions Subclonal populations observed within bulk resected primary GBMs transcriptionally evolve across tumor recurrence (EN and NE regions) and exhibit aberrant gene expression of common signaling pathways that persist despite standard or targeted therapy. Our findings provide evidence that there are both adaptive and clonally mediated dependencies of GBM on key pathways, such as the PI3K/AKT axis, for survival across recurrences.