Phenotypic Signatures and Molecular Mechanisms of Cancer Cell Invasion

Cancer;invasion;Chemical & Biomolecular Engineering

San, Angela C.Wirtz, Denis ;

Johns Hopkins University

关键词: Cancer;    invasion;    Chemical & Biomolecular Engineering;   

Others  :  https://jscholarship.library.jhu.edu/bitstream/handle/1774.2/37109/SAN-THESIS-2014.pdf?sequence=1&isAllowed=y

瑞士|英语

来源: JOHNS HOPKINS DSpace Repository

【 摘 要 】

The leading cause of cancer-related deaths is when cancer progresses to the metastatic stage, spreading into secondary sites.In order to combat metastasis, emerging research in cancer biology examines signaling pathways and mechanisms that are involved in the malignant transformation of cancer cells. An early stage in metastasis is invasion, where cells detach from their original environment and invade into surrounding tissue.Despite the significance of cell invasion, a complete picture of how invasion takes place is lacking, partially due to the limitations of complicated, time consuming, and costly platforms currently used to study this process. In this thesis, we utilized a 2.5D assay that combines the ease of 2D cell culture techniques with the physiological relevance of the 3D environment.Cells were first plated and allowed to spread on collagen coated 2D substrates, followed by the casting of 1 μm-thick 3D collagen gels on top of the cells.Unlike other invasion assays, this platform features a clearly defined 2.5D interface between the substrate and gel, which allowed high-resolution imaging of the morphological transition of cells from 2D to 3D and accurate quantification of the fraction of cells that invaded into the gel.Moreover, data collection and analysis do not require highly technical machinery or software.The novel functions of our assay led us to discover that cells instantaneously form microspikes that outline the entire cell body (within two hours after collagen gel casting) that may play a role in invasion.We further investigated the factors that influence invasion and discovered that the invasion of fibrosarcoma HT1080 cells is due to active cellular processes rather than simple diffusion and do not depend on matrix concentration or substrate rigidity.However, at low concentrations of collagen, HT1080 cells can invade independent of actomyosin proteins, cell-ECM binding, and the degradation of the surrounding tissue.Furthermore, we investigated the potential similarities of microspikes to filopodia and their involvement in invasion.Our preliminary results revealed that the depletion of filopodia-related proteins had no effects on microspike formation or invasion.Through examining various cell lines, we were able to establish a positive correlation between the quantity of microspikes formed and invasion capabilities, which can be used as a tool to quickly predict whether a cell line can invade from 2D substrates to 3D matrices.Finally, the phenomenon observed in this system defies durotaxis; the movement of cells from a stiff substrate to a softer gel is a novel discovery.Our system allows analysis at high resolution to observe changes at the single cell level as well as accurately quantify invasion percentages at the larger scale.This system can be used to delve deeper into the connection between the formation of microspikes and invasion, potentially uncovering the elusive first steps of invasion.The advantages that this novel assay provides over current methodologies can act as a solution to the drawbacks of current invasion studies.Furthermore, it can serve as an example for the design of future invasion assays that will be high throughput, low cost, and accessible.