Detection and Analysis of Circulating Tumor Cells (CTCs) in Patients With Breast Cancer Using a Novel Microfluidic and Raman Spectrum Device

Circulating tumor cells (CTCs) are a new type of breast cancer molecular marker. CTCs in peripheral blood originate from breast cancer (primary and metastatic lesions) shedding. Utilization of CTCs as novel and noninvasive tests for diagnosis confirmation, therapy selection, and cancer surveillance is a rapidly growing area of interest. At present, there is a great challenge to create an effective platform that can isolate these cells, as they are extremely rare: only 1-10 CTCs are present in a 7.5mL of a cancer patient's peripheral blood. The majority of the CTC capture methods are based on EpCAM expression as a surface marker of tumor-derived cells. However, EpCAM protein expression levels can be significantly down regulated during cancer progression as a consequence of the process of epithelial to mesenchymal transition. Although many technologies have been reported to achieve the capture and counting of CTCs, these methods study little information of cells and limited biological information can be obtained, leading to a lack in clinical diagnosis.

Microfluidics has demonstrated great potential as an effective technique for the medical and biological sciences. Inspired by the bed topography in river meanders, here, the investigators report a novel river meander-like cross-section in helical microchannels for size-based inertial focusing and enrichment, aiming to realize more functional geometries as well as reduce the extensive laborious requirement in traditional fabrication process. This device can facilitate particle focusing at a larger scale than traditional channels. Compared to the circular, rectangular and trapezoidal channels, the river meander-like microfluidic channel can successfully realize 26 μm particle focusing with a thinner focusing band in a shorter channel length. Also, in a single test, this structure can achieve 85.4% recovery and the enrichment ratio of 1.86 of spiked MDA-MB-231 cells in the whole blood, overcoming the dependence on traditional cell manipulation microfluidic devices. These results indicate that this river meander-like microfluidic chip has the great potential of size-based cell/particle sorting and enrichment for clinical application. Meanwhile, Surface-enhanced Raman scattering (SERS) has such advantages as high sensitivity, flexible excitation wavelength, high spectral resolution, non-invasiveness to biological samples, resistance self-fluorescence, photobleaching, etc., which is considered as a promising and powerful real-time detection technology for unlabeled cells. But SERS spectrum of cell contains information about different molecules, so it usually requires complex data interpretation. Therefore, the application of Raman spectroscopy combined with chemometrics in biological problems has attracted more and more attention. However, analysis result of chemometrics can be influenced by the complex background of Raman spectroscopy. Therefore, a pre-processing is needed to remove these influencing factors.

Based on these previous work, the investigators have developed a novel technology for the detection of CTCs of breast cancer. The detection platform constructed by the combination of microfluidic chip and surface-enhanced Raman spectroscopy is used to detect the blood sample of the subject, obtain the high quality of breast cancer circulating tumor cells from the aspects of cell specificity, surface structure and molecular activity of surface-enhanced Raman spectrum information, and extract the characteristics of different breast cancer circulating tumor cells by related signal feature extraction methods, establish a standard surface-enhanced Raman spectral feature database for the main types of breast cancer circulating tumor cells.

This technology aims to build a platform allowing for cell detection, synthesize particles for surface plasmon Raman enhancement, fabricate microfluidic devices, study the collection and analysis of the Raman spectra of CTCs, establish the database of CTC Raman spectra and develop the evaluation method of the biological detection. The primary objective is to demonstrate that the CTC assay counts technology can distinguish between healthy subjects and malignant breast cancer subjects. The secondary objective is to demonstrate that the CTCs detection technology can evaluate the efficacy of chemotherapy and neoadjuvant chemotherapy, as well as dynamic treatment monitoring and prognosis evaluation.