Circulating tumor cells and novel biomarkers for prognostic and biological of breast cancer

The detection of microscopic disease in breast cancer has been evaluated in lymph nodes, bone marrow (primary breast cancer), and peripheral blood (metastatic disease) [1, 2]. Most of these studies demonstrated that the detection of microscopic disease in breast cancer patients contributes prognostic information and, in selected cases, can predict the efficacy of treatments [1, 2]. In primary breast cancer, the detection of microscopic disease in lymph nodes and bone marrow has led to a better understanding of the role of minimal residual disease (MRD). In metastatic breast cancer (MBC) reliable detection of circulating tumor cells (CTCs) had been obtained by using immuno-magnetic separation and subsequent analysis by the CellSpotter™ analyzer (Veridex LLC, a Johnson & Johnson company, Warren, NJ, USA). This technology is becoming a standard tool for the 'real-time' assessment of prognosis and response to treatment. This is particularly important in the context of advanced disease management, considering the incurable status of the disease and the increasing therapeutic options available that could at least contribute to improve palliation and impact on overall survival.

In fact, despite years of clinical research, the odds of achieving complete response, and hence major survival benefit, for patients with MBC remain extremely low. Only a few patients who achieve a complete response after chemotherapy remain in this state for prolonged periods of time, with some remaining in remission beyond 20 years. There are presently no reliable biological markers that can predict prognosis and monitor therapy effects in MBC.

The detection of CTCs in patients with MBC about to start a new line of treatment has been shown to predict progression-free survival (PFS) and overall survival (OS). This prognostic value was independent of the line of therapy (e.g. first-line versus second-line or more) [2, 3]. Moreover, in multivariate analysis CTCs demonstrated superior value compared with site of metastasis (e.g. visceral versus soft tissue/bone), type of therapy, and length of time to recurrence after definitive primary surgery. In recent analysis, detection of CTCs has also been found to be prognostic in patients with bone-only disease (not measurable disease). CTCs have been shown to be superior to standard tumor markers (e.g. Ca27-29) in predicting prognosis. Furthermore, the efficacy or benefit to systemic therapy could be predicted by the level of CTCs as early as 3–4 weeks after initiation of therapy. Patients with persistent of ≥ 5 CTCs demonstrated lack of response or progressive disease at the time of restaging by standard imaging modalities. Conversely, patients with < 5CTCs showed objective remission. These data clearly suggest that CTCs can be used as an early predictor of treatment efficacy and be extremely useful in sparing patients from futile therapy early in the course of their treatment.

Prospective clinical trials are presently being conducted in MBC to validate further the prognostic value of CTCs, possibly to use this diagnostic tool to better stratify patients with metastatic disease, eventually modifying the current staging system (International Stage IV Stratification Study [ISSS]). Patients with metastatic disease could be divided into the subcategories IV_A and IV_B, depending on the presence or absence of CTCs. Additional studies are presently assessing the survival benefit of early change in treatment based on the persistence of CTCs and the possibility of collecting the cells, after sorting for evaluation of biomarkers (RT-PCR, gene profiling). Exploratory studies in PBC are also being conducted.

This technology could be integrated with other new investigation tools to develop blood-based integrated platforms that will facilitate screening, diagnosis, prognosis and target discovery. A recent acquisition is represented by the use of glycan arrays [4].

Malignant transformation and tumor progression are associated with the specific changes in the complex surface carbohydrates known as tumor-associated carbohydrate antigens (TACAs). Production of autoantibodies against these abnormal carbohydrates during cancer progression is expected. A robust printed glycan array was recently fabricated that employs a library of over 200 well defined structures comprising carbohydrate sequences of N-glycans, O-glycans, glycolipids, and glycoproteins. This printed glycan array was used to simultaneously detect multiple specific antiglycan autoantibodies in sera from breast cancer patients.