Abstract 5104: Isolation, culture and immunophenotyping of live patient-derived circulating tumor cells.
Authors: Matthew Elizabeth M. , Gallant Jean-Nicolas , Zhou Lanlan , Yang Zhaohai , Drabick Joseph J. , Truica Cristina I. , Lamparella Nicholas E. , Dicker David T. , El-Deiry Wafik S. 
Published: August 2014

Abstract
Metastasis is responsible for the majority of cancer-related deaths. A limiting step in metastasis is access to circulation, thus circulating tumor cells (CTCs) should be present in all patients with metastatic tumors. Tissue biopsy is the standard for diagnosis of carcinoma of unknown primary (CUP) and other metastatic carcinomas but there is a need for improved methodology. Circulating tumor cells not only enable real-time monitoring of cancer progression and response to therapy but may be utilized to provide personalized treatment for patients by evaluating the patients own CTCs in culture with therapeutics prior to systemic administration. Our overall objective is to utilize CTCs to provide insight into systemic cancer progression and approaches to therapy, including a CTC blood test as a tool for diagnosis, prognosis and therapy. We hypothesize that isolation, culture and real-time, single-cell immunophenotype profiling of CTC populations will inform diagnosis of tissue of origin, reveal prognostic markers such as stem-like properties, predict tumor aggressiveness and enable evaluation of potential therapies. We have been developing a post-Veridex CTC analysis method using multiplexed Q-dot conjugated antibodies for detection of multiple markers in each single cell within a CTC population to aid in CUP diagnosis. In parallel, we are developing a flexible micro spring array (FMSA) size-based CTC enrichment protocol and other approaches for capturing, culturing and immunophenotyping live CTCs. We have demonstrated that as few as 20 cancer cells spiked into blood may be captured with the FMSA device and grown into the millions of cells necessary for evaluating potential therapies. Due to the challenges of growing patient-derived tumor cells in culture, minimal progress has been made in this area. Cells captured from a patient with adenocarcinoma of unknown primary that have proliferated in culture on an FMSA device are being be evaluated for CK8,18 expression to identify putative CTCs. Utilizing an alternate approach, we have successfully captured and cultured CK8,18 positive and CD45 negative putative CTCs from the blood of a colorectal cancer patient. In addition to developing our approach for diagnosis of tissue of origin, we will utilize the FMSA device and other approaches for isolating live CTCs to study the biological properties of these cells in the context of metastatic spread. Successful application of our protocol to CTCs in patient samples is expected to provide a noninvasive, relatively quick and inexpensive blood test that can be both diagnostic and prognostic, and provide a platform for personalized therapeutic strategies. Circulating tumor cells may provide insight into the mechanism of systemic cancer progression, the characteristics of metastatic founder cells and changes within these cells as cancer progresses, thereby impacting the efficacy and evaluation of cancer therapy.
Citation Format: Elizabeth M. Matthew, Jean-Nicolas Gallant, Lanlan Zhou, Zhaohai Yang, Joseph J. Drabick, Cristina I. Truica, Nicholas E. Lamparella, David T. Dicker, Wafik S. El-Deiry. Isolation, culture and immunophenotyping of live patient-derived circulating tumor cells. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 5104. doi:10.1158/1538-7445.AM2013-5104