Unraveling Cancer Metastatic Cascade Using Microfluidics-based Technologies.

Cancer has long been a leading cause of death. The primary tumor, however, is not the main cause of death in more than 90% of cases. It is the complex process of metastasis that makes cancer deadly. The invasion metastasis cascade is the multi-step biological process of cancer cell dissemination to distant organ sites and adaptation to the new microenvironment site. Unraveling the metastasis process can provide great insight into cancer death prevention or even treatment. Microfluidics is a promising platform, that provides a wide range of applications in metastasis-related investigations. Cell culture microfluidic technologies for in vitro modeling of cancer tissues with fluid flow and the presence of mechanical factors have led to the organ-on-a-chip platforms. Moreover, microfluidic systems have also been exploited for capturing and characterization of circulating tumor cells (CTCs) that provide crucial information on the metastatic behavior of a tumor. We present a comprehensive review of the recent developments in the application of microfluidics-based systems for analysis and understanding of the metastasis cascade from a wider perspective.

Dynamic CTC phenotypes in metastatic prostate cancer models visualized using magnetic ranking cytometry.

Tumors can shed thousands of cells into the circulation daily. These circulating tumor cells (CTCs) are heterogeneous, and their phenotypes change dynamically. Real-time monitoring of CTC phenotypes is crucial to elucidate the role of CTCs in the metastatic cascade. Here, we monitor phenotypic changes in CTCs in mice xenografted with tumors with varying aggressiveness during cancer progression and a course of chemotherapy to study the metastatic potential of CTCs and changes in the properties of these cells in response to treatment. A new device that enables magnetic ranking cytometry (MagRC) is employed to profile the phenotypic properties of CTCs. Overall, CTCs from metastatic xenografts in mice display dynamic and heterogeneous profiles while non-metastatic models had static profiles. Decreased heterogeneity followed by a reduction in metastasis incidence was observed after a course of chemotherapy administered to highly metastatic xenografts. Phenotypic profiling of CTCs could be employed to monitor disease progression and predict therapeutic responses.

Isolation of Phenotypically Distinct Cancer Cells Using Nanoparticle-Mediated Sorting.

Isolating subpopulations of heterogeneous cancer cells is an important capability for the meaningful characterization of circulating tumor cells at different stages of tumor progression and during the epithelial-to-mesenchymal transition. Here, we present a microfluidic device that can separate phenotypically distinct subpopulations of cancer cells. Magnetic nanoparticles coated with antibodies against the epithelial cell adhesion molecule (EpCAM) are used to separate breast cancer cells in the microfluidic platform. Cells are sorted into different zones on the basis of the levels of EpCAM expression, which enables the detection of cells that are losing epithelial character and becoming more mesenchymal. The phenotypic properties of the isolated cells with low and high EpCAM are then assessed using matrix-coated surfaces for collagen uptake analysis, and an NAD(P)H assay that assesses metabolic activity. We show that low-EpCAM expressing cells have higher collagen uptake and higher folate-induced NAD(P)H responses compared to those of high-EpCAM expressing cells. In addition, we tested SKBR3 cancer cells undergoing chemically induced hypoxia. The induced cells have reduced expression of EpCAM, and we find that these cells have higher collagen uptake and NAD(P)H metabolism relative to noninduced cells. This work demonstrates that nanoparticle-mediated binning facilitates the isolation of functionally distinct cell subpopulations and allows surface marker expression to be associated with invasiveness, including collagen uptake and metabolic activity.

Profiling Functional and Biochemical Phenotypes of Circulating Tumor Cells Using a Two-Dimensional Sorting Device.

During cancer progression, tumors shed circulating tumor cells (CTCs) into the bloodstream. CTCs that originate from the same primary tumor can have heterogeneous phenotypes and, while some CTCs possess benign properties, others have high metastatic potential. Deconstructing the heterogeneity of CTCs is challenging and new methods are needed that can sort small numbers of cancer cells according to their phenotypic properties. Here we describe a new microfluidic approach that profiles, along two independent phenotypic axes, the behavior of heterogeneous cell subpopulations. Cancer cells are first profiled according to expression of a surface marker using a nanoparticle-enabled approach. Along the second dimension, these subsets are further separated into subpopulations corresponding to migration profiles generated in response to a chemotactic agent. We deploy this new technique and find a strong correlation between the surface expression and migration potential of CTCs present in blood from mice with xenografted tumors. This system provides an important new means to characterize functional diversity in circulating tumor cells.

Tracking the dynamics of circulating tumour cell phenotypes using nanoparticle-mediated magnetic ranking.

Profiling the heterogeneous phenotypes of rare circulating tumour cells (CTCs) in whole blood is critical to unravelling the complex and dynamic properties of these potential clinical markers. This task is challenging because these cells are present at parts per billion levels among normal blood cells. Here we report a new nanoparticle-enabled method for CTC characterization, called magnetic ranking cytometry, which profiles CTCs on the basis of their surface expression phenotype. We achieve this using a microfluidic chip that successfully processes whole blood samples. The approach classifies CTCs with single-cell resolution in accordance with their expression of phenotypic surface markers, which is read out using magnetic nanoparticles. We deploy this new technique to reveal the dynamic phenotypes of CTCs in unprocessed blood from mice as a function of tumour growth and aggressiveness. We also test magnetic ranking cytometry using blood samples collected from cancer patients.

Interrogating Circulating Microsomes and Exosomes Using Metal Nanoparticles.

A chip-based approach for electrochemical characterization and detection of microsomes and exosomes based on direct electro-oxidation of metal nanoparticles (MNPs) that specifically recognize surface markers of these vesicles is reported. It is found that exosomes and microsomes derived from prostate cancer cells can be identified by their surface proteins EpCAM and PSMA, suggesting the potential of exosomes and microsomes for use as diagnostic biomarkers.