Colorectal cancer (CRC) is one of the leading causes of death worldwide1. The high mortality of colorectal cancer patients is tightly linked to late diagnosis, undetected recurrence, and distant metastases2. Approximately half of patients will develop distant metastasis after CRC resection, usually leading to poor prognosis3. In clinical practices, adjuvant chemotherapy is used in high-risk patients to eradicate the occult micrometastatic tumor cells before metastatic disease becomes clinically evident. Currently, patients classified with TNM above stage IIB, elevated preoperative carcinoembryonic antigen (CEA), angio-invasive growth, tumor perforation or obstruction and insufficient lymph node sampling (< 10 nodes) are eligible for adjuvant therapy4. Some patients receive adjuvant therapy without having presumed micro-metastasis because of a lack of high sensitivity and specificity of the known risk factors for disease recurrence. Other patients are classified as having a low risk for disease recurrence and do not receive adjuvant therapy, but progress with recurrence. The early detection of micrometastases could identify the patients who are most (and least) likely to benefit from adjuvant therapy. There is an unmet clinical need in disease management for better tools, reliable biomarkers, and simpler tests for early detection of disease recurrence or micro-metastasis.
Metastasis is a multistep process caused by dissemination of malignant cells from the primary tumor5. Cancer cells which undergo intravasation result in circulating tumor cells (CTCs) in the bloodstream or lymphatic system and carry the potential for metastatic tumor formation at distant sites6. Several studies have shown that the number of CTCs correlates with progression-free survival and overall survival in metastatic CRC patients7–10. These studies were primarily conducted with stage IV patients where distant metastasis had already occurred, providing limited evidence in understanding if CTC count predisposes the development of metastasis. In non-metastatic CRC, different non-standardized detection methods of CTC have made inter-study comparisons difficult, and only limited data exists about the prognostic role of CTC11–15. Because of extremely low frequency of CTC in non-metastatic CRC, larger volume of peripheral blood was used to analyzed the correlation between CTC number and outcome 14,16. Mesenteric vein blood obtained during surgery showed higher CTC amount than peripheral blood17. In this study, mesenteric vein blood and peripheral blood were both used in a CTC enumeration assay with a demonstrated enhancement of the sensitivity of CTC detection.
CTC isolation remains a challenge due to the scarcity and heterogeneity of these cells in blood. Label-free approaches to cell isolation, such as by size, typically suffer from low recovery, clogging of filters, complicated integration of external force fields, low cell purity, and loss of smaller rare cells limiting their broad utility18. Antibody-based methods to isolate rare cells based on expression of surface marker proteins typically use immunomagnetic isolation by magnetic fields using antibodies immobilized to magnetic beads19; microfluidics approaches with antibodies immobilized on a microfluidic chip20; or fluorescence-activated methods where rare cells are detected and sorted based on laser-induced fluorescence of fluorophore-labeled antibodies21. A new developed microfluidic platform, for CTC isolation, antibody labeling, and fluorescence imaging, allows for consistent measurement over the whole study period. This study validated the accuracy, linearity, limits of blank and detection, and the reproducibility of this microfluidic platform and hardware system. It also explored the prognostic value of CTC presence before and after surgical resection in patients with potentially curable disease, focusing on non-metastatic CRC.