# Targeting CTC Heterogeneity: Aptamer-Based Liquid Biopsy Predicts Outcome in Lung Cancer

## TL;DR

A new liquid biopsy method using DNA aptamers improves detection of lung cancer cells in blood, offering better monitoring and prognosis.

## Contribution

Aptamers LC-17 and LC-18 enable CTC detection independent of EpCAM, addressing tumor heterogeneity in lung cancer.

## Key findings

- Aptamers LC-17 and LC-18 bind to CTC-specific proteins DEFA1 and PRDX2, absent in healthy donors.
- Higher CTC counts correlate with advanced tumor stage and shorter survival in lung cancer patients.
- The method shows strong potential for clinical use in monitoring and stratifying NSCLC patients.

## Abstract

Liquid biopsy, particularly the detection of circulating tumor cells (CTCs), represents a convenient non-invasive approach for cancer diagnosis and monitoring. However, the clinical utility of current methods is limited by their reliance on epithelial markers like EpCAM, which fails to capture all heterogenic blood CTCs due to epithelial–mesenchymal transition. We address this critical limitation by presenting a novel, aptamer-based detection strategy that overcomes the dependency on a single marker. DNA aptamers LC-17 and LC-18, which have high affinity and specificity to different cancer markers, effectively capture tumor cells. We found that these aptamers successfully identified CTCs in lung cancer patients by binding to two specific proteins. The number of CTCs depends on the T-stage and patient survival. Our findings suggest that this aptamer-based liquid biopsy has strong potential for integration into clinical practice.

Background: The detection of circulating tumor cells (CTCs) holds significant promise for the diagnosis and monitoring of lung cancer (LC). However, the clinical utility of CTCs is limited by the heterogeneity of their phenotypes and the shortcomings of existing detection methods, which often rely on epithelial markers like EpCAM. DNA aptamers offer a promising alternative due to their high affinity, stability, and ability to recognize diverse cancer-specific biomarkers. Methods: This study utilized DNA aptamers LC-17 and LC-18, previously selected against primary lung tumor tissue, to isolate and detect CTCs in the peripheral blood of 43 non-small cell lung cancer (NSCLC) patients. Mass spectrometry (LC-MS/MS) was employed to identify the target proteins of aptamer LC-17. CTCs from patients’ blood and healthy donors were isolated via filtration after erythrocyte and lymphocyte lysis and stained with FAM-labeled LC-17 and LC-18 aptamers for detection using fluorescence and light microscopy. Results: Mass spectrometry identified neutrophil defensin 1 (DEFA1) and peroxiredoxin-2 (PRDX2) as the primary protein targets of aptamer LC-17 in CTCs, both of which were absent in healthy donor samples. CTC enumeration revealed statistically significant correlations between elevated CTC counts (>3 cells/4 mL blood) and advanced primary tumor size (T4 vs. T1–T3, p = 0.012), extensive regional lymph node metastasis (N3 vs. N1–N2, p = 0.014), and shorter overall survival (median 24 vs. 32 months, p < 0.05). Conclusions: The developed aptamer-based liquid biopsy method effectively captures heterogeneous CTC populations independent of EpCAM expression. The strong correlation of CTC counts with disease progression and survival underscores their clinical relevance as a prognostic biomarker in NSCLC. This approach presents a viable, non-invasive tool for disease monitoring and stratification of NSCLC patients, with potential for integration into clinical practice.