A combination of circulating tumor cell (CTC) and cancer stem cell (CSC) markers to predict the prognosis of breast cancer.
Authors: Sun Jingxin , Marx Angela , Alexander Angela , Wiley Jie , Kai Megumi , Valero Vicente , Lim Bora 
Published: June 2023

e13007 Background: CTCs are tumor cells that shed from either primary tumor or metastatic foci and can provide more information about the tumor's biological entity than circulating tumor DNA (ctDNA), or exosomes do. The current FDA-approved CTC assay misses the CSC phenotypes, which are pivotal in driving tumorigenesis, metastasis, resistance to treatment, and cancer recurrence. Therefore, we combine CTC and CSC markers to determine the prognosis. We hypothesized that the CSC markers on CTC correlate with poorer outcomes. Methods: Peripheral blood samples of qualified breast cancer patients were collected at diagnosis and every 6 months during follow-up. EpCAM, MUC-1, and HER2 are used as CTC markers, while TWIST, AKT, PI3K, and ALDH-1 are used to detect CSC. According to the expression level of the above markers, patients were categorized into four groups: NN negative with both CTC and CSC markers, NP expressing CSC markers, PN expressing CTC markers, and PP positive with both CTC and CSC markers. The progression-free survival (PFS) and overall survival (OS) of all four groups were calculated by Kaplan-Meier model from the date of surgery until the disease progression and death/the last follow-up of the patients, respectively. Results: We screened 100 patients aged 18 years or older with confirmed stage II to stage IV breast cancer and selected 81 patients for the study. Fifteen patients dropped out of the study due to loss of follow-up or incomplete clinical information leaving 66 evaluable patients. There are 55 Caucasians (83%), 5 African Americans (7.5%), 5 Hispanics (7.5%), and 1 Asian (1.5%). The median age of the patients is 52.5 years old (29-81), and the median follow-up is 22.6 months (2.5-65). Most patients have stage III disease (60 and 91%) followed by 4 stage IV (6%) and 2 Stage II (3%). The median time between the first sample date and surgery is 172 days (-148 to 270). The NN group has the most patients (41 and 62%) compared to NP group (16 and 24%) and PN group (6 and 9%). The PP group has the least number of patients (2 and 3%) and is not observed in patients with positive hormonal receptors (HR). The PN group has a higher ratio of HR-HER2+ patients than NP group does. The HR and HER2 statuses were checked in the four groups. Interestingly, the NN and NP groups are mostly HR+HER2- patients, while the PN group is mainly HR- patients regardless of the HER2 status. The PP group only has HR- patients. The PFS curves start to flatten at 12 months and separate among the four groups, with the highest PFS percentage in the NP (81.2%) and NN (77.8%) followed by the PN (62.5%) and PP (50%) group. The OS percentage is 70% in NN, 66% in PN, 60% in NP, and 50% in PP, similar to the PFS pattern. Conclusions: Checking both CTC and CSC markers help triage the prognosis of breast cancer patients. CTC and CSC markers observed in breast cancer patients with different HR or HER2 status can be further investigated for potential correlations.