Abstract 5242: Obtaining a transcriptomic profile of a single prostate cancer cell

Abstract

Abstract There have been considerable advances in the detection, isolation and characterization of circulating tumor cells (CTC) and disseminated tumor cells (DTC) in prostate cancer (PCa) patients. However, the ability to interrogate these cells is restricted by the small number detected and isolated (typically <10), and current technologies available to characterize individual cells. We set out to determine the limit of commercially available technologies to obtain a transcriptomic profile of a single PCa cell. To do this we isolated clonally selected Aphidicolin synchronized C4-2B PCa cells. Ten sets of C4-2B cells were isolated individually or in pools of 5 or 10 cells. All thirty samples were amplified using WT-Ovation™ One-Direct (NuGEN), hybridized on a 44K Whole Human Gene Expression Microarray (Agilent Technologies) normalized for batch effects, and normalized within and between arrays. The amplified material was also assessed by realtime-PCR. Using a high stringency cut off (mean signal intensity of 300) on the oligonucleotide arrays 22,365 probes were Cy3 positive for 10 cells, 19,393 for 5 cells and 15,441 for 1 cell. Furthermore, using the same high stringency cut off, the sensitivity and specificity between 1 and 10 cells was 0.640 and 0.946 respectively, and between 1 and 5 cells was 0.713 and 0.932, demonstrating a very low false positive rate. Ten-thousand randomly selected pairs of commonly expressed genes had a Pearson correlation coefficient of 0.862 and 0.908 for 1 vs. 10 cells and 1 vs. 5 cells respectively, indicating that the vast majority of detected probes maintained consistent relative abundance. The cycle threshold values for single cell samples by realtime-PCR were well within the detectable range for abundant transcripts (e.g. AR and KLK3). However, other transcripts (e.g. FKBP5 and TMPRSS2) were not detected in the single cell samples and had limited detection in 5 cell samples when compared to 10 cell samples. These data suggest that currently commercially available technologies allow for the detection of a limited but significant number of genes from a single PCa cell. We are using the methodology described herein, to determine the heterogeneity of single DTC isolated from the bone marrow of patients with PCa. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 5242. doi:10.1158/1538-7445.AM2011-5242