Abstract P1-01-19: Circulating tumor cells in triple-negative breast cancer patients express prostate related genes and show different genetic profiles in US and German patient cohorts
Authors: Kasimir-Bauer S , Gao H , Bittner A-K , Plappert L , Feniuk N , Ueno NT , Cohen L , Valero V , Woodward WA , Alvarez RH , Hoffmann O , Kimmig R , Reuben JM 
Published: March 2017

Abstract
Background: Triple negative breast cancer (TNBC) is known for its aggressive behavior and poor prognosis since treatment options are limited. Specific biomarkers are urgently needed to treat patients (pts) accordingly. In this regard, circulating tumor cells (CTCs) are discussed to be an ideal surrogate marker for individualized treatment options. Using a multi-marker gene panel for the characterization of CTCs, we recently demonstrated that CTCs in TNBC pts and non-TNBC pts showed different genetic profiles including the expression of the androgen receptor (AR). We here compared CTCs of TNBC pts in two different national Institutions in the US (mainly Afro-American women) and Germany (Caucasian women) to identify potentially ethnical differences and to further explore AR expression related similarities of prostate cancer to TNBC.
Methods: 2x5 ml blood of TNBC pts before therapy (n=18 from the Dep. of Gynecol. and Obstetrics, Essen, GER; n=31 from the MDA Cancer Center Houston, US) were analyzed for CTCs applying positive immunomagnetic selection targeting EpCAM, EGFR and HER2 using the AdnaTest EMT-2/Stem Cell Select (QIAGEN Hannover GmbH, Germany). Subsequently, cDNA was pre-amplified for specific genes using TaqMan PreAmp Master Mix according to in house designed assays. Establishment of a 19 gene qPCR panel was performed for the markers PI3K, AKT2, ERCC1, AURKA, HER2, HER3, EGFR, ALK, AR, BRCA1, c-KIT, c-MET, KRT5, mTOR, NOTCH1, PARP1, SRC1, CD45 (leucocyte control) and GAPDH (housekeeping gene) as well as an internal reference. The cutoff was calculated, taking the false positive rate in healthy donors into account, and defined as Ct (cutoff) - Ct (sample) - [Ct (CD45cutoff) – Ct (CD45sample)]. Using the AdnaPanel Prostate Cancer (QIAGEN Hannover GmbH, Germany), cDNA obtained from a subset of 14 pts was further analyzed for the expression profile of AR, prostate specific antigen (PSA) as well as prostate specific membrane antigen (PSMA).
Results: The overall CTC-positivity rate (at least one of the markers expressed) was comparable between both sites (GER: 78%; US: 65%). Similar overexpression frequencies were found for AKT2, AR, AURKA, c-KIT, NOTCH1 and SRC at both sites. However, ALK, BRCA1, HER3 and KRT5 were predominantly found in CTCs of German pts while these genes were not or only weakly expressed in CTCs of the US group. In contrast, c-MET, ERCC1, HER2 and PARP were mainly expressed in CTCs of the US group and rarely in the German group. Interestingly, EGFR was not detected in both groups. Analyzing the overexpression of prostate related genes in a subgroup of 14 TNBC pts resulted in the expression of AR in 21%, PSA in 14%, and PSMA in 36% of cases, respectively.
Conclusion: Although we observed similar gene expression profiles in both patient cohorts, clear differences in some gene expression frequencies were detected at both sites. Whether these findings can be related to ethnic differences will be subject of further investigations. Furthermore, we were able to demonstrate that prostate related genes were frequently detectable which so far have not been discussed in the context of breast cancer.
Citation Format: Kasimir-Bauer S, Gao H, Bittner A-K, Plappert L, Feniuk N, Ueno NT, Cohen L, Valero V, Woodward WA, Alvarez RH, Hoffmann O, Kimmig R, Reuben JM. Circulating tumor cells in triple-negative breast cancer patients express prostate related genes and show different genetic profiles in US and German patient cohorts [abstract]. In: Proceedings of the 2016 San Antonio Breast Cancer Symposium; 2016 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2017;77(4 Suppl):Abstract nr P1-01-19.