Catch and Release: rare cell analysis from a functionalised medical wire

Shukun Chen,Amin El-Heliebi,Klaus Pantel,Gerlinde Tauber,Michaela Pötscher,Tanja Langsenlehner,Andra Kuske,Zbigniew T Czyż,Gerd Leitinger,Karl Kashofer,Thomas Kroneis,Peter Sedlmayr,Sabine Riethdorf,Bernhard Polzer

doi:10.1038/srep43424

Abstract

Enumeration and especially molecular characterization of circulating tumour cells (CTCs) holds great promise for cancer management. We tested a modified type of an in vivo enrichment device (Catch&Release) for its ability to bind and detach cancer cells for the purpose of single-cell molecular downstream analysis in vitro. The evaluation showed that single–cell analysis using array comparative genome hybridization (array-CGH) and next generation sequencing (NGS) is feasible. We found array-CGH to be less noisy when whole genome amplification (WGA) was performed with Ampli1 as compared to GenomePlex (DLRS values 0.65 vs. 1.39). Moreover, Ampli1-processed cells allowed detection of smaller aberrations (median 14.0 vs. 49.9 Mb). Single-cell NGS data obtained from Ampli1-processed samples showed the expected non-synonymous mutations (deletion/SNP) according to bulk DNA. We conclude that clinical application of this refined in vivo enrichment device allows CTC enumeration and characterization, thus, representing a promising tool for personalized medicine.

Highlights

The CellCollector CANCER01 (DC01, GILUPI) is a CE-approved medical device that uses antibodies against the epithelial cell adhesion molecule (EpCAM) for isolating circulating tumour cells (CTCs) directly from peripheral blood in vivo[11]
One of the disadvantages of the currently used in vivo enrichment devices is that captured cells firmly attach to the wire preventing CTCs to be recovered for further analysis
We investigated the C&R’s capacity of isolating and detaching pre-stained cells (Fig. 1c) by spiking 1·102, 1·103, and 1·104 HT-29 cells per ml in 5 ml of peripheral blood (n = 3 each)

Summary

Introduction

The CellCollector CANCER01 (DC01, GILUPI) is a CE-approved medical device that uses antibodies against the epithelial cell adhesion molecule (EpCAM) for isolating CTCs directly from peripheral blood in vivo[11]. To enable downstream analysis at the single-cell level we tested a novel device allowing recovery of attached cells [CellCollector CANCER03 (Catch & Release, C&R, GILUPI)] which is not yet CE-certified for clinical application, currently allowing for in vitro use only. We tested if the C&R which is based on cell enrichment by EpCAM capture, allows isolation and recovery of single tumour cells in vitro. For this purpose, we investigated the charging and detachment performance of the CellCollector C&R in vitro using tumour cells suspended at different cell densities either in PBS or peripheral blood. For cell characterisation we amplified single cells recovered from the C&R using two whole genome amplification (WGA) strategies and analysed the amplified single-cell products using comparative genomic hybridization (array-CGH) and generation sequencing (NGS)

Methods

Results

Conclusion