Abstract P4-01-11: Circulating tumor cells expressing the prostate specific membrane antigen PSMA indicate worse outcome in triple-negative breast cancer patients at primary diagnosis
Authors: Kasimir-Bauer Sabine , Keup Corinna , Hoffmann Oliver , Hauch Siegfried , Kimmig Rainer , Bittner Ann-Kathrin 
Published: March 2020

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. We here comprehensively analyzed mRNA profiles of CTCs from TNBC pts before and after neoadjuvant chemotherapy including the expression of prostate related genes [androgen receptor (AR), its splice variant ARV7, the prostate specific antigen (PSA) and the prostate specific membrane antigen (PSMA)] to further elucidate these markers as targets for personalized treatment. Methods: 2 × 5 ml blood of 34 TNBC pts before and 22 pts after therapy were analyzed for CTCs applying positive immunomagnetic selection using the AdnaTest EMT-2/Stem Cell Select (QIAGEN, Germany). Subsequently, cDNA was pre-amplified for specific genes using TaqMan PreAmp Master Mix according to in house designed assays. A 19 gene qPCR panel was performed including AKT2, ALK, AR, AURKA, BRCA1, EGFR, ERCC1, ERBB2, ERBB3, KIT, KRT5, MET, MTOR, NOTCH1, PARP1, PIK3CA, SRC, 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, Germany), cDNA was further analyzed for the expression profile of AR, ARV7, PSA and PSMA, respectively. Results: Before therapy, at least one of the prostate related genes was detected in 50% of the pts, resulting in the expression of AR in 35%, ARV7 in 24%, PSMA in 18%, and PSA in 15% of cases, respectively. Notably, in 75% of AR-positive cases, ARV7 was also expressed on CTCs. After therapy, prostate related genes were only detected in two pts, expressing AR, ARV7 and PSA (n=1) and AR (n=1), respectively. Before therapy, the presence of PSMA-positive CTCs alone (p=0.026) or in combination with ARwildtype/ARV7-positive CTCs (p=0.014) significantly correlated with early relapse. Interestingly, this expression pattern was highly related to the expression of genes belonging to the ERBB family (EGFR, ERBB2, ERBB3; p=0.034). Either ERRB family or PSMA expression on CTCs in TNBC was detected in about 70% of the cases analyzed with 90% relapses predicted. Conclusion: Although AR and its splice variant were detected on CTCs in one third of the pts before therapy, these markers do not seem to be relevant with regard to be considered as targets for additional treatment options. In contrast, although expressed in a minority of pts, PSMA-positive CTCs at primary diagnosis indicated worse outcome and should be included in CTC-multimarker panel analysis to identify pts at higher risk for relapse. This study is ongoing and the analysis of more pts will finally show whether these results can be confirmed.
Citation Format: Sabine Kasimir-Bauer, Corinna Keup, Oliver Hoffmann, Siegfried Hauch, Rainer Kimmig, Ann-Kathrin Bittner. Circulating tumor cells expressing the prostate specific membrane antigen PSMA indicate worse outcome in triple-negative breast cancer patients at primary diagnosis [abstract]. In: Proceedings of the 2019 San Antonio Breast Cancer Symposium; 2019 Dec 10-14; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2020;80(4 Suppl):Abstract nr P4-01-11.