Previous study with CytoSorter® in BC showed that CTC could reflect tumor burden and could be used as a diagnostic aid in BC screening and early diagnosis [8]. First we compared CTCs with BC patients’ clinicopathological features and found that CTCs were associated with tumor stage and tumor size, but not with lymph node metastasis, patients’ age and tumor molecular subtype and histology grade. Latest study in the lab shows that mesenchymal type of CTCs in BC is more invasive and more related with lymph node and distant metastasis [9]. We used epithelial cell adhesion molecule (EpCAM) antibody to capture the epithelial types of CTCs, which may explain that CTCs are associated with tumor size, but not with lymph node involvement. Only confirmed BC patients were enrolled in this study. If CTC positive was defined as any patient with CTC ≧ 2, CTC positive rates were not correlated with tumor stage, tumor size, lymph node involvement, and tumor molecular subtypes as shown in Table 6. CTC negative rates were 0 in stage III patients and in patients at N2 and N3 stages. But CTC negative rates were comparable in other group of patients. CTC detection rates (positive rates) in stage I and II BC were 92.9% (71.6%) and 87.2% (76%), respectively, which is in line with the previous finding that CTC can be used as an screening aid for early stage BC [8].
Next, we compared CTCs with other BC screening imaging methods, such as US, MG and MR. Our analysis showed CTCs were more associated with US than MR, and showed least correlation with MG. MG is an X-ray imaging to detect for presence of tumor or lump in breast, and is currently the most efficient screening method recommended by American Cancer Society (ACS) and AJCC to detect early BC. The sensitivity of MG depends on the patient’s age and breast composition. Sensitivity is higher in women over 50 and with fatty breasts [10]. High breast density leads to the FNR of MG. In fact, MG had the highest FNR of 30.6% in this study, followed by US (15.8%) and MR (12.8%). Breast is a mixed structure, containing both dense tissue (i.e., glandular tissue and connective tissue, together known as fibroglandular tissue) and fatty tissue. On a MG film, fatty tissue appears dark, whereas fibroglandular tissue appears as white areas. Tumors usually have similar density as fibroglandular tissue and therefore, it is harder to detect tumor in women with denser breasts with MG. FNR occur more frequently among younger women as well because younger women are more likely to have dense breasts [10]. As Oriental women usually have dense breasts, MG is less sensitive in Chinese females [11]. MG was not associated with any clinicopathological features of BC in this study, which may explain why CTCs were not correlated with the MG. MG FNR are surprisingly more observed in stage II patients (38.2%) and in patients at T2 stage (44%). MG FNR in other groups of BC patients were ranging from 15–30%.
US uses sound waves to make a computer picture of the inside of the breast. US can differentiate fluid-filled cysts from solid masses, therefore, it is most useful for looking at some breast changes, such as lumps. BC detection rate with US is comparable with MG, with a greater proportion of invasive and node-negative cancers among US detections [12, 13]. US usually cannot detect the small calcification points and is less sensitive to detect tumors less than 5 mm or deep in the breast. Since most BC are superficial tumors and Chinese women usually have dense breasts, US was more sensitive for BC detection than MG in our study. FNR of US was negatively associated with patients’ TNM stage. US showed higher FNR in stage I patients, and in patients at T1 and N0 stage. US usually generates more false-positive results (FPR) [13], which cannot be confirmed in our study since no healthy volunteer or patients with benign breast diseases were enrolled in this study.
Although MG and US are currently the mostly used imaging techniques for BC detection in practice, the low sensitivity and specificity of these two imaging tools resulted in a demand for new imaging modalities and breast MR has become increasingly important in the detection and delineation of BC [13, 14]. A breast MR uses magnets and radio waves to take pictures of the breast. Breast MR is the most sensitive method for detection of BC and has more diagnostic value on pretherapeutic BC staging, monitoring of primary systemic therapies and solving problematic diagnostic situations where direct biopsy is not possible [14]. MR has the lowest FNR of 12.8% in our study. Similar to US, most MR FNR were observed in stage I BC patients, and in patients at T1 and N0 stage.
Medical imaging modalities showed different FNR in patients with different molecular subtypes of tumor. MR and US showed lowest FNR in human epidermal growth factor receptor 2 (HER2) over-expression BC patients, while MG had a similar FNR in all 4 molecular subtypes. The general limitation of imaging modalities for BC screening is that most methods cannot detect tumors smaller than certain size. That is why US and MR showed higher FNR in early stage BC (stage I) and in patients with smaller tumor (T1). Previous study in the lab showed that MR has the highest sensitivity of BC screening in Chinese women, followed by US and then MG [4]. But in Stage I BC patients, CTC, US, MG, and MR showed similar FNR ranging from 26–28%.
BSGI is a molecular breast imaging procedure that shows the metabolic activity of breast lesions [15]. MR-TIC can reflects the hemodynamic features of a specific lesion, especially about the vascular structure around tumor. CTCs were not associated with BSGI nor MR-TIC. Bansal et al used flow cytometry to detect CTCs in 114 BC patients and found that CTC positivity did not show any correlation with the tumor immunohistochemical profile [16]. Taken together, CTC enumeration cannot be used as a biomarker to reflect the molecular signature of BC. CTCs are heterogeneous. Some are epithelial type, and some are mesenchymal type. Some may express stem-cell markers, such as CD44, or other proteins, such as PD-L1 or HER2. Therefore, profiling of CTCs is considered as a promising tool to reveal the molecular signature of BC, allowing for a better personal treatment rearrangement.
Our results show CTCs are more associated with US than MR or MG. Comparing the consistence between CTCs and medical imaging examinations in BC detection, CTCs show the highest consistence of 70.3% with US, followed by 68.2% with MR and 58.3% with MG, which may explain why CTCs are more correlated with US than MR, and show least correlation with MG. Interpretation of medical imaging examinations require a radiologist’s expertise. An inexperienced expert might misinterpret the imagings to generate false results. Compared to CTCs, medical imaging examinations might be considered as a diagnostic tool with more human error bias.
Lastly, we liked to know whether conjugation of CTC would decrease FNR of imaging examinations for BC detection as well. Table 10 showed that combination of CTC with medical imaging examination decreased their FNR for BC detection, especially for MG. FNR of MG decreased from 30.6–8.3%, even more than in conjugation with US. Medical imaging examinations combine with CTCs counts would improve the diagnostic potency for BC detection.