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

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

doi:10.1038/srep43424

Cited by 20 publications

(23 citation statements)

References 33 publications

(36 reference statements)

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“…For more elaborate single-cell molecular characterization (e.g., next-generation sequencing), a novel CellCollector was developed, namely, the Catch & Release CellCollector (Gilupi). The Catch & Release CellCollector allows EpCAM-based capturing of cells and their release by a releasing buffer for subsequent single-cell analysis, but it is not yet cleared for use in patients (35 ). As the CellCollector is based on antiEpCAM antibodies for CTC isolation, it may miss several other CTC subpopulations with downregulated epithelial markers (36 ).…”

Section: Discussionmentioning

confidence: 99%

In Situ Detection and Quantification of AR-V7, AR-FL, PSA, and KRAS Point Mutations in Circulating Tumor Cells

et al. 2018

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BACKGROUND Liquid biopsies can be used in castration-resistant prostate cancer (CRPC) to detect androgen receptor splice variant 7 (AR-V7), a splicing product of the androgen receptor. Patients with AR-V7-positive circulating tumor cells (CTCs) have greater benefit of taxane chemotherapy compared with novel hormonal therapies, indicating a treatment-selection biomarker. Likewise, in those with pancreatic cancer (PaCa), KRAS mutations act as prognostic biomarkers. Thus, there is an urgent need for technology investigating the expression and mutation status of CTCs. Here, we report an approach that adds AR-V7 or KRAS status to CTC enumeration, compatible with multiple CTC-isolation platforms. METHODS We studied 3 independent CTC-isolation devices (CellCollector, Parsortix, CellSearch) for the evaluation of AR-V7 or KRAS status of CTCs with in situ padlock probe technology. Padlock probes allow highly specific detection and visualization of transcripts on a cellular level. We applied padlock probes for detecting AR-V7, androgen receptor full length (AR-FL), and prostate-specific antigen (PSA) in CRPC and KRAS wild-type (wt) and mutant (mut) transcripts in PaCa in CTCs from 46 patients. RESULTS In situ analysis showed that 71% (22 of 31) of CRPC patients had detectable AR-V7 expression ranging from low to high expression [1–76 rolling circle products (RCPs)/CTC]. In PaCa patients, 40% (6 of 15) had KRAS mut expressing CTCs with 1 to 8 RCPs/CTC. In situ padlock probe analysis revealed CTCs with no detectable cytokeratin expression but positivity for AR-V7 or KRAS mut transcripts. CONCLUSIONS Padlock probe technology enables quantification of AR-V7, AR-FL, PSA, and KRAS mut/wt transcripts in CTCs. The technology is easily applicable in routine laboratories and compatible with multiple CTC-isolation devices.

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Section: Discussionmentioning

confidence: 99%

In Situ Detection and Quantification of AR-V7, AR-FL, PSA, and KRAS Point Mutations in Circulating Tumor Cells

et al. 2018

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“…Approximately 75% of single cells subjected to WGA yield high quality WGA products: This is 1) smear of WGA products ranging from 0.1 to > 1 kb ( Figure 4 , top panel) and 2) three or four bands in the 4plex quality control PCR ( Figure 4 , bottom panel). WGA products can be used for 1) array-CGH profiling meeting the quality criteria for single-cell array-CGH profiles 6 7 and 2) targeted NGS after re-amplification WGA 7 .…”

Section: Representative Resultsmentioning

confidence: 99%

“…charging and detaching). The treatment's effects on DNA quality of single cells were previously evaluated after WGA by means of a quality control PCR 4 5 , array-CGH 6 7 and targeted sequencing 7 showing no difference compared to non-treated cells micromanipulated from cell suspensions 7 . The advantage of this method lies in the combination of rare target-cell pre-enrichment and the recovery of cells for single-cell downstream analysis ( Figure 1 ).…”

Section: Introductionmentioning

confidence: 99%

Target Cell Pre-enrichment and Whole Genome Amplification for Single Cell Downstream Characterization

Chen

El‐Heliebi

Schmid

et al. 2018

JoVE

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Rare target cells can be isolated from a high background of non-target cells using antibodies specific for surface proteins of target cells. A recently developed method uses a medical wire functionalized with anti-epithelial cell adhesion molecule (EpCAM) antibodies for in vivo isolation of circulating tumor cells (CTCs)1. A patient-matched cohort in non-metastatic prostate cancer showed that the in vivo isolation technique resulted in a higher percentage of patients positive for CTCs as well as higher CTC counts as compared to the current gold standard in CTC enumeration. As cells cannot be recovered from current medical devices, a new functionalized wire (referred to as Device) was manufactured allowing capture and subsequent detachment of cells by enzymatic treatment. Cells are allowed to attach to the Device, visualized on a microscope and detached using enzymatic treatment. Recovered cells are cytocentrifuged onto membrane-coated slides and harvested individually by means of laser microdissection or micromanipulation. Single-cell samples are then subjected to single-cell whole genome amplification allowing multiple downstream analysis including screening and target-specific approaches. The procedure of isolation and recovery yields high quality DNA from single cells and does not impair subsequent whole genome amplification (WGA). A single cell's amplified DNA can be forwarded to screening and/or targeted analysis such as array comparative genome hybridization (array-CGH) or sequencing. The device allows ex vivo isolation from artificial rare cell samples (i.e. 500 target cells spiked into 5 mL of peripheral blood). Whereas detachment rates of cells are acceptable (50 - 90%), the recovery rate of detached cells onto slides spans a wide range dependent on the cell line used (<10 - >50%) and needs some further attention. This device is not cleared for the use in patients.

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“…However, the volume of blood taken from the patient cannot be substantially increased. In line with this need, CellCollector (GILUPI Nanomedizin) arose as a technology claiming to access large volumes of blood by placing the detection device intravenously . This device is a structured medical guidewire coated with a hydrogel layer which, in turn, is covalently coupled with antibodies against EpCAM.…”

Section: Current Outlook Of Ctcs In Liquid Biopsymentioning

confidence: 99%

Liquid Biopsy Based on Circulating Cancer‐Associated Cells: Bridging the Gap from an Emerging Concept to a Mainstream Tool in Precision Medicine

Jiménez‐Zenteno

Cerf

2019

Adv. Biosys.

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The concept of liquid biopsy and the isolation and analysis of circulating biomarkers from blood samples is proposed as a surrogate to solid biopsies and can have the potential to revolutionize the management of patients with cancer. The relevance of circulating tumor cells (CTCs) and the importance of the information they carry is acknowledged by the medical community. But what are the barriers to clinical adoption? This review draws a panorama of the biological implications of CTCs, their physical and biochemical properties, and the current technological bottlenecks for their analysis in relation with the medical needs. Keys and considerations to bridge the technological and clinical gaps that still need to be overcome to be able to introduce CTCs in clinical routine are finally synthesized.

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

Cited by 20 publications

References 33 publications

In Situ Detection and Quantification of AR-V7, AR-FL, PSA, and KRAS Point Mutations in Circulating Tumor Cells

In Situ Detection and Quantification of AR-V7, AR-FL, PSA, and KRAS Point Mutations in Circulating Tumor Cells

Target Cell Pre-enrichment and Whole Genome Amplification for Single Cell Downstream Characterization

Liquid Biopsy Based on Circulating Cancer‐Associated Cells: Bridging the Gap from an Emerging Concept to a Mainstream Tool in Precision Medicine

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