A squirmy system for studying cancer’s spread

The spread of cancer cells from their points of origin to other parts of the body makes cancer more deadly and difficult to treat. While researchers strive to understand the mechanisms that drive this process and find new ways to stop the spread, cancer cell metastasis is challenging to study. In vitro assays don’t fully recapitulate the complex environmental and mechanical cues that coordinate a cell’s invasion through extracellular material. And in most in vivo systems, metastasis occurs deep within the tissue, beyond the reach of light microscopes. In these models, “even if you could visualize it, it would be hard to catch a cell in the act of invading. It's kind of like trying to watch a snow leopard in the Himalayan mountains,” said David Sherwood, a cell and molecular biologist at Duke University.

To more easily investigate cancer cell metastasis, Sherwood’s team takes advantage of a normal, controlled cellular invasion event that occurs in the roundworm species Caenorhabditis elegans. During the worm’s development, a specialized uterine cell called the anchor cell breaks through the tough basement membranes that enclose the uterine and vulval tissues, connecting the two regions to enable reproduction. The process relies on much of the same cellular machinery as metastasis, including enzymes that degrade the extracellular matrix and feet-like invadopodia protrusions that drill into the surrounding structures, but is much more predictable and amenable to visual analysis. By capturing images of anchor cells as they invade in C. elegans, Sherwood’s team can directly observe the effect of manipulating specific genes on cell invasion, paving the way for new strategies for combating metastasis.

We can watch this invasive cell interact with the basement membrane and actually see what happens to the basement membrane, watch specialized invasive structures that the cell deploys to interact with the basement membrane, and understand at a deep, fundamental level how the cell invades.
- David Sherwood, Duke University

The researchers developed transgenic strains of C. elegans featuring genetically encoded fluorophores that emit bright colors in the anchor cell and basement membrane. Leveraging this visualization tool and the consistent timing and location of anchor cell invasion, the team can image the process inside the 50 micrometer-thick, translucent worm body using a light microscope. “We can watch this invasive cell interact with the basement membrane and actually see what happens to the basement membrane, watch specialized invasive structures that the cell deploys to interact with the basement membrane, and understand at a deep, fundamental level how the cell invades,” Sherwood said.

To use this model to gain insights into metastasis, the researchers isolated anchor cells from thousands of C. elegans at the time of invasion and analyzed their transcriptomes (1). They pinpointed more than 100 new genes that are upregulated in invasive anchor cells compared to other cell types in the worm and that have mammalian counterparts. They engineered Escherichia coli expressing double-stranded RNA that knocks down each gene from this group and fed the bacteria to the transgenic worms. The team observed that knockdown of certain genes impaired the anchor cell’s ability to breach the basement membrane, indicating that these genes are important for cellular invasion and therefore promising drug targets for metastasis.

When the researchers knocked down the gene encoding TCT-1, the C. elegans equivalent of the multifunctional, cancer-associated translationally controlled tumor protein (TCTP), they saw a strong delay in invasion, suggesting that TCTP could play a role in halting metastasis. They also found that a subset of the upregulated genes is involved in the biosynthesis of ribosomes and that knocking these genes down interfered with invasion. The cell prepares to invade by building more ribosome protein factories, ramping up production of the equipment it needs to battle the basement membrane. “That could be something that's harnessed to basically stop invasive cells before they can even become invasive,” Sherwood said.

Sherwood hopes that this work sheds light on combinations of genes that promote invasion through different mechanisms and may show a synergistic effect when knocked out together. This approach is also important for selectively targeting cancer cell metastasis without affecting normal processes in healthy cells. “How do we specifically knock out invasion? ...That's absolutely going to be through combinatorial targeting of proteins, so, knocking out multiple different pathways at the same time to just hit the Achilles heel of the invasion event,” Sherwood said.