Selection for invasive tumour cells reveals a role for MAPK signalling in cell elasticity regulation

[摘要] The metastatic spread of cancer cells is a major contributor to cancer patient deaths. In order to disseminate from one part of the body to another, invasive tumour cells must perform a complex cascade of steps. Common to several stages of the metastatic process is the ability of tumour cells to squeeze through narrow spaces. One adaptation that allows cancer cells to adjust to confined environments is a change in the cell mechanical properties, which results in diminished cell stiffness. Such an altered cellular physical property is thought to contribute to the invasive and metastatic properties of cancer cells. There is little currently known about the factors or signalling pathways that modulate cell stiffness; therefore, the identification of factors that modify tumour cell plasticity could identify potential drug targets for anti-metastasis chemotherapy.I hypothesized that within the distributions of parameters in tumour cell line populations, it would be possible to select for cells with an augmented ability to squeeze through narrow gaps, thus highlighting factors that contributed to cell deformability. To identify factors that help cancer cells migrate through confined spaces, MDA MB 231 human breast cancer cells and MDA MB 435 melanoma cells underwent 3 rounds of selection, using tissue culture inserts with 3 μm pores. This selection approached allowed me to isolate cell populations with enhanced abilities to pass through psychical constrictions, as well as augmented invasive abilities in vitro and in vivo. Additionally, populations of small volume cells were selected from parental MD MB 231 cells by flow cytometry. By obtaining the matching cell size control, I found that increased pore invasion was not solely dependent on small cell or nuclei size. In fact, properties unique to the pore-selected invasive cells were: decreased actin cytoskeleton anisotropy and cell rigidity (Young’s modulus) as determined by Atomic Force Microscopy (AFM).To identify signalling pathways that were associated with observed cytoskeleton and elasticity changes, RNA sequencing was performed on parental, pore-selected and flow-selected MDA MB 231 cells, and parental and pore-selected MDA MB 435 cells and a stringent comparison was performed. Such an approach allowed identification of common genes, which were used for further analysis by applying GSEA. The most common gene signature was found to associate with 3 increased signalling through the Ras-Raf-MEK-ERK mitogen-activated protein kinase (MAPK) pathway. Blocking the activity of the MAPK/ERK pathway with two pharmacologically distinct MEK inhibitors resulted in actin stress fibre restoration, increased cell stiffness and restrained cell invasion through collagen matrices. I was able to identify, for the first time that changes in cytoskeletal organisation mediated by increased signalling through the MAPK/ERK pathway resulted in cell plasticity alterations. Therefore, drugs that block Ras-MAPK signalling would likely provide clinical benefit by reversing the effect of this signalling pathway on tumour cell plasticity and subsequently restraining migratory and invasive capabilities.

[发布机构] University:University of Glasgow;Department:Institute of Cancer Sciences

[关键词] Q Science (General)