Priority Research Papers:

A multiplexed marker-based algorithm for diagnosis of carcinoma of unknown primary using circulating tumor cells

Elizabeth M. Matthew, Lanlan Zhou, Zhaohai Yang, David T. Dicker, Sheldon L. Holder, Bora Lim, Ramdane Harouaka, Si-Yang Zheng, Joseph J. Drabick, Nicholas E. Lamparella, Cristina I. Truica and Wafik S. El-Deiry _

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Oncotarget. 2016; 7:3662-3676. https://doi.org/10.18632/oncotarget.6657

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Elizabeth M. Matthew1,2,*, Lanlan Zhou1,2,*, Zhaohai Yang3, David T. Dicker1,2, Sheldon L. Holder4, Bora Lim1,4,6, Ramdane Harouaka5, Si-Yang Zheng5, Joseph J. Drabick4, Nicholas E. Lamparella4, Cristina I. Truica4 and Wafik S. El-Deiry1,2

1 Laboratory of Translational Oncology and Experimental Cancer Therapeutics, Division of Hematology-Oncology, Penn State Hershey Cancer Institute, Hershey, PA, USA

2 Laboratory of Translational Oncology and Experimental Cancer Therapeutics, Department of Medical Oncology and Molecular Therapeutics Program, Fox Chase Cancer Center, Philadelphia, PA, USA

3 Department of Pathology, Penn State Milton S. Hershey Medical Center, Hershey, PA, USA

4 Division of Hematology-Oncology, Penn State Milton S. Hershey Medical Center and Penn State Hershey Cancer Institute, Hershey, PA, USA

5 Department of Biomedical Engineering, Penn State University, University Park, PA, USA

6 The University of Texas MD Anderson Cancer Center, Houston, TX, USA

* These authors have contributed equally to this work

Correspondence to:

Wafik S. El-Deiry, email:

Keywords: circulating tumor cells (CTCs), carcinoma of unknown primary (CUP), immunofluorescence, diagnosis

Received: November 14, 2015 Accepted: November 23, 2015Published: December 18, 2015


Real-time, single-cell multiplex immunophenotyping of circulating tumor cells (CTCs) is hypothesized to inform diagnosis of tissue of origin in patients with carcinoma of unknown primary (CUP). In 20 to 50% of CUP patients, the primary site remains unidentified, presenting a challenge for clinicians in diagnosis and treatment. We developed a post-CellSearch CTC assay using multiplexed Q-dot or DyLight conjugated antibodies with the goal of detecting multiple markers in single cells within a CTC population. We adapted our approach to size-based CTC enrichment protocols for capturing CTCs and subsequent immunofluorescence (IF) using a minimal set of markers to predict the primary sites for common metastatic tumors. The carcinomas are characterized with cytokeratin 7 (CK7), cytokeratin 20 (CK20), thyroid transcription factor 1 (TTF-1), estrogen receptor (ER) or prostate-specific antigen (PSA. IF has been optimized in cultured tumor cells with individual antibodies, then with conjugated antibodies to form a multiplex antibody set. With IF, we evaluated antibodies specific to these 5 markers in lung, breast, colorectal, and prostate cancer cell lines and blood from metastatic prostate and breast cancer patients. This advanced technology provides a noninvasive, diagnostic blood test as an adjunct to routine tissue biopsy. Its further implementation requires prospective clinical testing.

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