263. Nuclease-Activated Oligonucleotide Probes for the Rapid and Robust Detection of Breast Cancer Circulating Tumor Cells (CTCs)
Authors: David D. Dickey, Sven Kruspe, Kevin T. Urak, William H. Thiel, Karen C. Clark, Elliot Burghardt, Justin P. Dassie, Alexandra Thomas, James O. McNamara, Paloma H. Giangrande
Journal: Molecular Therapy (2016)

Metastatic breast cancer is the second leading cause of female cancer deaths in the United States. Despite substantial progress in its treatment, metastatic breast cancer remains incurable. Early identification of breast cancer patients at greatest risk of developing metastatic disease is thus an important goal that would enable oncologists to aggressively treat these patients while the cancer is still vulnerable. In addition, this would spare patients who do not need or would not benefit from further treatments from having to endure the harmful side-effects of chemotherapeutic drug regimens. Circulating tumor cells (CTCs) are rare cancer cells found in the blood circulation of cancer patients that provide a non-invasively accessible cancer cell specimen (liquid biopsy) from patients. The number of circulating tumor cells (CTCs) in cancer patients has recently been shown to be a valuable diagnostic indicator of the state of metastatic breast cancer. In particular, patients with few or no CTCs were found to have a better overall prognosis compared to patients with high numbers of CTCs. Despite the implications of CTCs as diagnostics for advanced breast cancer treatment, a critical challenge for adopting CTC-based diagnostic tests has been the development of methods with sufficient sensitivity to reliably detect the small number of CTCs that are present in the circulation. Furthermore, current tests for CTC detection are expensive, have high false positives and negatives, have high background noise, are time consuming and require a significant level of expertise to conduct. To overcome the limitations of current CTC detection assays and develop more sensitive, rapid and cost effective CTC detection methods, we explored the potential of detecting CTCs by measuring their nuclease activity with nuclease-activated probes1. We present data towards the development of a rapid and highly-sensitive CTC detection assay based on nuclease-activated oligonucleotide probes that are selectively digested (activated) by target nucleases expressed in breast cancer cells. We confirm that these probes are not activated by serum nucleases or nucleases from a lymphoblastic cell line (e.g. K-562). Furthermore, we present extensive data towards the optimization of activity and sensitivity of these probes in cell lysates from various breast cancer cell lines. Data is also presented confirming the detection of CTCs in blood from patients with stage IV breast cancer as well as healthy controls with spiked cancer cells. Future studies will focus on developing similar probes for detection of CTCs in patients with pancreatic and ovarian cancer where early detection would greatly improve survival outcomes. In conclusion, this work describes a robust assay for detection of breast cancer CTCs that will be straightforward to implement in most clinical diagnostic labs. 1. Hernandez FJ, Huang L, Olson ME, Powers KM, Hernandez LI, Meyerholz DK, Thedens DR, Behlke MA, Horswill AR, McNamara JO, 2nd. Noninvasive imaging of staphylococcus aureus infections with a nuclease-activated probe. Nat Med. 2014;20:301-306