High-Throughput Label-Free Isolation of Heterogeneous Circulating Tumor Cells and CTC Clusters from Non-Small-Cell Lung Cancer Patients

Mina Zeinali; Maggie Lee; Arthi Nadhan; Anvya Mathur; Casey Hedman; Eric Lin; Ramdane Harouaka; Max S Wicha; Lili Zhao; Nallasivam Palanisamy; Mathias Hafner; Rishindra Reddy; Gregory P Kalemkerian; Bryan J Schneider; Khaled A Hassan; Nithya Ramnath; Sunitha Nagrath

doi:10.3390/cancers12010127

High-Throughput Label-Free Isolation of Heterogeneous Circulating Tumor Cells and CTC Clusters from Non-Small-Cell Lung Cancer Patients

Cancers (Basel). 2020 Jan 3;12(1):127. doi: 10.3390/cancers12010127.

Authors

Mina Zeinali^{1

2

3}, Maggie Lee^{1

2}, Arthi Nadhan^{1

2}, Anvya Mathur^{1

2}, Casey Hedman⁴, Eric Lin^{1

2}, Ramdane Harouaka⁵, Max S Wicha⁵, Lili Zhao⁶, Nallasivam Palanisamy⁷, Mathias Hafner³, Rishindra Reddy⁸, Gregory P Kalemkerian⁵, Bryan J Schneider⁵, Khaled A Hassan⁵, Nithya Ramnath^{5

9}, Sunitha Nagrath^{1

2}

Affiliations

¹ Chemical Engineering, University of Michigan, 2800 Plymouth Road, NCRC, Building 20-3rd Floor, Ann Arbor, MI 48109, USA.
² Biointerfaces Institute, University of Michigan, 2800 Plymouth Road, NCRC B10-A184, Ann Arbor, MI 48109, USA.
³ Institute for Medical Technology of Heidelberg University & University of Applied Sciences Mannheim, Paul-Wittsack-Straße 10, 68163 Mannheim, Germany.
⁴ Molecular, Cellular, and Developmental Biology, University of Michigan, 1105 North University Avenue, 2220 Biological Science Building, Ann Arbor, MI 48109, USA.
⁵ Department of Internal Medicine, University of Michigan, 1500 E Medical Center Dr, Ann Arbor, MI 48109, USA.
⁶ Biostatistics Department, University of Michigan, Ann Arbor, MI 48109, USA.
⁷ Department of Urology, Henry Ford Health System, 1 Ford Place, Room 2D26, Detroit, MI 48202, USA.
⁸ Department of Surgery, University of Michigan, 1500 E Medical Center Dr, Ann Arbor, MI 48109, USA.
⁹ Veterans Administration Ann Arbor Healthcare System, 2215 Fuller Road, Ann Arbor, MI 48105, USA.

Abstract

(1) Background: Circulating tumor cell (CTC) clusters are emerging as clinically significant harbingers of metastases in solid organ cancers. Prior to engaging these CTC clusters in animal models of metastases, it is imperative for technology to identify them with high sensitivity. These clusters often present heterogeneous surface markers and current methods for isolation of clusters may fall short. (2) Methods: We applied an inertial microfluidic Labyrinth device for high-throughput, biomarker-independent, size-based isolation of CTCs/CTC clusters from patients with metastatic non-small-cell lung cancer (NSCLC). (3) Results: Using Labyrinth, CTCs (PanCK+/DAPI+/CD45-) were isolated from patients (n = 25). Heterogeneous CTC populations, including CTCs expressing epithelial (EpCAM), mesenchymal (Vimentin) or both markers were detected. CTCs were isolated from 100% of patients (417 ± 1023 CTCs/mL). EpCAM- CTCs were significantly greater than EpCAM+ CTCs. Cell clusters of ≥2 CTCs were observed in 96% of patients-of which, 75% were EpCAM-. CTCs revealed identical genetic aberrations as the primary tumor for RET, ROS1, and ALK genes using fluorescence in situ hybridization (FISH) analysis. (4) Conclusions: The Labyrinth device recovered heterogeneous CTCs in 100% and CTC clusters in 96% of patients with metastatic NSCLC. The majority of recovered CTCs/clusters were EpCAM-, suggesting that these would have been missed using traditional antibody-based capture methods.

Keywords: CTC clusters; circulating tumor cells (CTCs); epithelial-to-mesenchymal transition (EMT); inertial microfluidics; non-small-cell lung cancer (NSCLC).

Abstract

Grants and funding