Unbiased quantitative assessment of Her-2 expression of circulating tumor cells in patients with metastatic and non-metastatic breast cancer.

Background Circulating tumor cells (CTCs) can provide the basis for a liquid biopsy and may guide the use of targeted therapies. We report on unbiased quantification of Her-2 protein expression of CTCs. Patients and methods Her-2 assessment of CTCs was carried out using the CellSearch(®) system in 103 metastatic (M1) and 88 non-metastatic (M0) breast-cancer patients. Expression of Her-2 on CTCs was determined by a manual review and an automated algorithm using Her-2- fluorescein isothiocyanate (FITC) fluorescence of leukocytes to determine the Her-2-expression threshold in each sample. Results Her-2 expression of CTCs varied greatly within and among patients compared with Her-2 expression of leukocytes. In M1 patients, a threshold of 75% of Her-2 positive CTCs in patients with ≥5 CTCs was set. Applying this threshold, 9% of M1 patients with Her-2-negative primary tumors had Her-2-positive CTC status and 29% of M1 patients with Her-2-positive primary tumors had Her-2-negative CTC status. No Her-2 discrepancy was observed between CTCs and primary tumors in M0 patients. Conclusions Our findings demonstrate that Her-2 expression is heterogeneous among CTCs within each patient. We show the feasibility of unbiased quantitative and reproducible assessment of treatment targets on CTCs, opening a path towards personalized treatment.

Challenges in the stratification of breast tumors for tailored therapies.

Studying the molecular stratification of breast carcinoma is a real challenge considering the extreme heterogeneity of these tumors. Many patients are now treated following recommendation established at several NIH and St Gallen consensus conferences. However a significant fraction of these breast cancer patients do not need adjuvant chemotherapies while other patients receive inefficacious therapies. High density gene expression arrays have been designed to attempt to establish expression profiles that could be used as prognostic indicators or as predictive markers for response to treatment. This review is intended to discuss the potential value of these new indicators, but also the current weaknesses of these new genomic and bioinformatic approaches. The combined analysis of transcriptomic and genomic alteration data from relatively large numbers of well annotated tumor specimens may offer an opportunity to overcome the current difficulties in validating recently published non overlapping gene lists as prognostic or therapeutic indicators. There is also hope for identifying and deciphering signal transduction pathways driving tumor progression with newly developed algorithms and semi quantitative parameters obtained in simplified in vitro or in vivo models for specific transduction pathways.

Zyxin is up-regulated during megakaryocytic differentiation of human UT-7/c-mpl cells.

To characterize genes involved in megakaryocytic commitment, we compared expression profiles of bipotent cells (UT-7/c-mpl) with those of the same cells induced to differentiate towards megakaryopoiesis in the presence of TPO. Using cDNA arrays, we showed that 12 out of 2260 genes changed their expression level after 6h of TPO stimulation. One of these genes encodes for zyxin, a cytoskeleton protein component. Zyxin is up-regulated at the mRNA and protein levels in UT-7/c-mpl cells in response to TPO confirming the reliability of the cDNA array technology. Similarly, when CD34 positive cells were induced to differentiate into megakaryocytes, zyxin mRNA was accumulated. Furthermore, when megakaryocytes were allowed to spread on fibrinogen, formation of stress fibers and lamellipodia was induced and zyxin was localized at the picks of actin stress fibers. These results suggest an important role for zyxin during megakaryocytic differentiation and more precisely in the regulation of the integrin mediated adhesion process in megakaryocytes.