Description:
<jats:title>Abstract</jats:title>
<jats:p>Aims: Bone morphogenetic proteins (BMP) play an important role in early embryonic development as well as in organ homeostasis and in tumorigenesis. We could show previously that BMP-2 enhance migratory and invasive properties of tumor cells. Furthermore, BMP-2 affects the expression of genes which are involved in the regulation of apoptosis. A crucial step in metastasis formation is the liberation of tumor cells from the primary site, the entrance into circulation and the long-lasting persistence in the peripheral blood. Therefore we asked whether BMPs might contribute to the vitality of circulating tumor cells.</jats:p>
<jats:p>Methods: Leukocytes and circulating epithelial cells from the peripheral blood of breast cancer patients were labelled with fluorescent antibodies and magnetically enriched with the MACS® system. After separation of single vital cells with the aureka® device, a single cell multiplex RT PCR was performed using the AmpliGrid system. The PCR products were analysed by urea-polyacrylamide gel electrophoresis. For real-time single cell PCR demonstrating the expression level of selected genes the Fastlane system (Qiagen, Hilden) was used. Localization of phosphorylated receptor Smads was analysed by laser scanning microscopy after one hour incubation with 100 ng/ml BMP-2.</jats:p>
<jats:p>Results: Circulating epithelial cells were identified by using the cell surface antigen EpCam. Vital EpCam-positive cells were picked and subjected to multiplex RT-PCR covering 5 components of the BMP signaling cascade as well as RPL13A for control. 121 of 265 selected cells expressed RPL13A. These cells were used for further analysis. Most of these cells expressed BMP-2 (54%), BMPR-IA (46%), BMPR-IB (82%). Transcripts of the type II receptor BMPR-II (35%) and the putative BMP antagonist BMP-3 (25%) were presen to a lower extend. The expression level of BMP-2 was comparable in individual EpCam-positive cells, thus at least an autocrine stimulation of the BMP signaling pathway can be assumed. For studying the responsiveness of circulating epithelial cells towards a BMP stimulus these cells were enriched from peripheral blood and incubated with BMP-2. Vital cells showed a predominant nuclear staining for pSmad1 demonstrating a functional BMP signalling network.</jats:p>
<jats:p>Conclusion: We show that key components of the BMP signaling pathway are expressed in leukocytes and epithelial cells in the peripheral blood of breast cancer patients. This indicates that the BMP signaling network potentially contributes to the activity and survival of circulating epithelial cells.</jats:p>
<jats:p>This work was supported by the Dr. Rainald-Stromeyer-Stiftung</jats:p>
<jats:p>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 LB-36. doi:10.1158/1538-7445.AM2011-LB-36</jats:p>