Abstract 3709: Single cell analysis from CSF of patients with suspected leptomeningeal metastasis
Authors: Koestler Caecilia , Feliciello Giancarlo , Lueke Florian , Weidele Kathrin , Zoubaa Saida , Hau Peter , Pukrop Tobias , Riemenschneider Markus J. , Klein Christoph A. , Polzer Bernhard 
Published: March 2024

Introduction: Leptomeningeal metastases (LM) are diagnosed in up to 10% of metastatic cancer patients, the most frequent cancers being carcinomas of breast and lung or melanoma. Due to a dismal prognosis and an overall median survival of only four months new approaches for molecular analysis could help to identify individual treatment options. Material and Methods: In this study, we collected 28 cerebrospinal fluid (CSF) samples from patients with different primary tumors but suspected LM. CSF was divided in two parts for standard neuropathologic cytomorphology and detection of circulating tumor cells (CTCs) by using the CellSearch® CTC Kit, respectively. Subsequently, single Cytokeratin-positive CTCs were isolated for whole genome amplification by Ampli1TM and molecular analysis. Additionally, we were able to isolate cell-free DNA (cfDNA) from CSF in a subset of patients which we amplified by an adapted Ampli1TM library protocol. In a small subset of patients, we collected slices of formalin-fixed paraffin embedded (FFPE) primary tumors and systemic metastases for dissociation and isolation of pure subpopulations by DEPArrayTM technology. Isolated CTCs, cfDNA and subpopulations of tumor and stromal cells were analyzed for copy number variations (CNV) applying the Ampli1TM LowPass protocol on the Illumina MiSeqTM platform. Results: By comparing results from classical cytomorphology and CellSearch® workflow, we could find high concordance (24/28 samples) with a slightly higher tumor cell detection rate by CellSearch® (11 positive cases vs. 9 by cytomorphology). Importantly, we could confirm the malignant origin of isolated cells by CNV analysis in all patients positive in CellSearch® assay. Moreover, by comparing single cell data with cfDNA analysis of a subset of 20 samples, we could also detect a higher sensitivity for tumor detection in CTC-based analysis (8/20 cases positive) in comparison to cfDNA-based analysis (5/20 cases positive). In a comprehensive analysis of multiple samples of an individual patient, including CTCs and cfDNA from CSF at different time points, CTCs from blood and biopsies from multiple visceral metastasis, we could identify divergent genomic evolution between tumor cells from the central nervous syste Conclusion: In this proof-of-concept study we show that a liquid biopsy-based approach for single cell analysis from CSF holds potential for innovative diagnostic assays for patients with suspected LM. Combining CTC quantification with molecular single cell analysis could enable us to define predictive tests to select novel treatment options for patients with LM in the future.

Citation Format: Caecilia Koestler, Giancarlo Feliciello, Florian Lueke, Kathrin Weidele, Saida Zoubaa, Peter Hau, Tobias Pukrop, Markus J. Riemenschneider, Christoph A. Klein, Bernhard Polzer. Single cell analysis from CSF of patients with suspected leptomeningeal metastasis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 3709.