Abstract LB-197: Circulating rare cells enable highly efficient cancer detection

Abstract

Abstract Objectives of the study We intended to develop a protocol combining a novel micro-fluidic enrichment technology and RT-qPCR for the molecular analysis of CTCs. Recently we identified CTC-specific mRNA markers allowing CTC detection in 29% of breast cancer patients and in 24.5% of ovarian cancer patients at diagnosis. However, the detection of cancer cells was hampered by the large number of contaminating leukocytes. The necessary use of cut-off values for positivity reduced the specificity of the down-stream PCR assay. By improving the purity of cancer cells and PCR analysis we sought to increase both sensitivity and specificity of the diagnostic procedure. Methodology For reducing the blood sample volume (up to 20ml) density gradient centrifugation was used when needed. The samples were passed through a micro-fluidic disposable cassette, which captures tumor cells based on their less deformable nature and larger size compared to blood cells. Lysis of the captured cells was done directly in the cassette and total RNA was extracted. After a cDNA pre-amplification step gene expression levels of leukocyte-specific and CTC-related markers were measured using RT-qPCR. The efficiency of the combined protocol was assessed in blood samples taken from patients with primary and recurrent malignant diseases. Results 7 out of 13 pre-selected RNA markers were not detectable in blood samples from 11 healthy volunteers. In cancer patients we observed measurable gene expression of at least one out of these 7 RNA markers. 5 out of 5 breast cancer (primary disease: N = 1; metastatic: N = 4), 9 out of 10 primary and 4 out of 8 relapsed ovarian cancer patients were classified correctly by the test. The most striking finding is the detection rate of 90% for primary ovarian cancer at a specificity of 100%. Conclusions The enrichment of rare cells from blood samples using micro-fluidics results in a highly pure cell population. This enables the application of extremely sensitive methods, like RT-qPCR to specifically detect rare events. By combining a novel micro-fluidic cell enrichment and molecular analysis we have taken a major step forward, which allows the implementation of ′liquid biopsies′ in cancer detection studies and as a companion diagnostic in clinical trials. Citation Format: Eva Obermayr, Elisabeth Maritschnegg, Paul Speiser, Christian Singer, Eva M. Schuster, Sabine Danzinger, Nina Pecha, Robert Zeillinger. Circulating rare cells enable highly efficient cancer detection. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr LB-197. doi:10.1158/1538-7445.AM2015-LB-197