Complementary Sequential Circulating Tumor Cell (CTC) and Cell-Free Tumor DNA (ctDNA) Profiling Reveals Metastatic Heterogeneity and Genomic Changes in Lung Cancer and Breast Cancer.

Say Li Kong,Wan-Teck Lim,Chwee Teck Lim,Joyce A Tai,Quan Sing Ng,Trifanny Yeo,Xingliang Liu,Axel M Hillmer,Yoon Sim Yap,Chee Keong Toh,Daniel Shao-Weng Tan,Huay Mei Poh,Yong Wei Chua ,L.y Phua ,Wen Huan Ling ,Kiley Wei Jen Loh ,Mei Kim Ang ,Tira J Tan ,Yu Xuan Haw ,Tony Kiat Hon Lim ,Raymond Ng ,Swee Jin Tan ,Yi Fang Lee

doi:10.3389/fonc.2021.698551

Abstract

IntroductionCirculating tumor cells (CTCs) and cell-free tumor DNA (ctDNA) are tumor components present in circulation. Due to the limited access to both CTC enrichment platforms and ctDNA sequencing in most laboratories, they are rarely analyzed together.MethodsConcurrent isolation of ctDNA and single CTCs were isolated from lung cancer and breast cancer patients using the combination of size-based and CD45-negative selection method via DropCell platform. We performed targeted amplicon sequencing to evaluate the genomic heterogeneity of CTCs and ctDNA in lung cancer and breast cancer patients.ResultsHigher degrees of genomic heterogeneity were observed in CTCs as compared to ctDNA. Several shared alterations present in CTCs and ctDNA were undetected in the primary tumor, highlighting the intra-tumoral heterogeneity of tumor components that were shed into systemic circulation. Accordingly, CTCs and ctDNA displayed higher degree of concordance with the metastatic tumor than the primary tumor. The alterations detected in circulation correlated with worse survival outcome for both lung and breast cancer patients emphasizing the impact of the metastatic phenotype. Notably, evolving genetic signatures were detected in the CTCs and ctDNA samples during the course of treatment and disease progression.ConclusionsA standardized sample processing and data analysis workflow for concurrent analysis of CTCs and ctDNA successfully dissected the heterogeneity of metastatic tumor in circulation as well as the progressive genomic changes that may potentially guide the selection of appropriate therapy against evolving tumor clonality.

Highlights

Circulating tumor cells (CTCs) and cell-free tumor DNA are tumor components present in circulation
We utilized a combination of size-based and CD45-negative selection method to enrich for CTCs In order to ensure sample comparison validity, CTCs and cell-free tumor DNA (ctDNA) were isolated from the same blood tube
A total of 116 CTCs and 41 plasma were collected from lung cancer patients while 159 CTCs and 53 plasma were obtained from breast cancer patients (Supplementary Table 5)

Summary

Introduction

Circulating tumor cells (CTCs) and cell-free tumor DNA (ctDNA) are tumor components present in circulation. Tissue biopsies and radiological imaging are routinely utilized by clinicians to monitor treatment efficacy and disease progression in patients. Tissue biopsy may be limited by sampling error as single site biopsy may not reflect the profile of the whole tumor. Minimally invasive liquid biopsy allows repetitive non-invasive sample collection and offers a broader tumor genomic profile based on shed DNA or cells from the tumor, with potential for better realtime monitoring of treatment efficacy and disease progression. The metastatic spread of cancer is largely due to the shedding of circulating tumor cells (CTCs) from tumors into the blood stream and invasion of distant organs. The number of CTCs detected in the blood of cancer patients correlates with inferior treatment response and survival outcomes [3], suggesting that the assessment of CTCs have prognostic and predictive importance in monitoring treatment efficacy [4]. Beyond the quantification of CTCs, molecular analysis of CTCs is essential for improving our understanding of the tumor biology which may in turn have therapeutic implications

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Conclusion