Much Like a Crime Scene, Cancer’s Evolution Can be Reconstructed

By Deborah Borfitz 

November 14, 2024 | Investigators at Weill Cornell Medicine have combined several powerful technologies to reconstruct how cancer spreads from the prostate to metastatic sites elsewhere in the body. Bioluminescence imaging, CRISPR/Cas9-based barcoding, and innovative computational methods for tracing the movement of cancer from tissue to tissue were used in creating a roadmap revealing the small number of aggressive cells that seed cancer’s often deadly migration.  

A shared interest in evolutionary biology brought the research team together with colleagues at Cold Spring Harbor Laboratory to trace the trajectory of clonal populations of cancer cells from the primary tumor to metastatic sites such as the liver or bone. Their mission was to learn what special evolutionary adaptations permitted them to make the journey, says Dawid Nowak, Ph.D., assistant professor of pharmacology in medicine at Weill Cornell Medicine. 

The technology being used causes cancer cells to leave “breadcrumbs” as they move through the body, enabling barcode sequencing to produce precise, high-resolution information about their travels, according to Adam Siepel, Ph.D., professor and chair of the Simons Center for Quantitative Biology and co-leader of the Cancer Genetics and Genomics Program at Cold Spring Harbor Laboratory. The hope now is to learn the biology underlying changes in metastatic disease using technologies that allow the gene expression patterns of cancer cells to be traced along with their evolutionary trajectories. 

The defining patterns of metastatic seeding in prostate cancer were described in a study that was published recently in Cancer Discovery (DOI: 10.1158/2159-8290.CD-23-1332). Researchers used a mouse model of aggressive metastatic cancer (Evolution in Cancer of the Prostate, or EvoCaP) together with computational software (Evolutionary Lineage Tracing in R, or EvoTraceR) to track tumor clones containing readable barcodes. 

“It’s a way of bringing some of the tools we normally use to study viruses or evolution over millions and millions of years... to bear on cancer over these much shorter timeframes where tumors develop inside of an individual human body,” says Siepel. The long-term objective is to explore ways to counteract the evolutionary strategies of cancer cells—genetic as well as epigenetic and metabolic—with adaptive, organ-specific therapies that block the metastatic process, Nowak adds. 

Integrating Technologies

Cancer-tracking maps have been built for other cancer types, but this is believed to be the first one specific to prostate cancer, says Nowak. It could also be easily adapted to other cancer types since genetic alterations in the mouse model are induced in the prostate using lentiviral injections.  

Bioluminescence imaging, used to track cancer progression in live mice, is a well-established technology for visualization of cancer progression in preclinical models, he notes. In addition, cancers are also positive for enhanced green fluorescent protein, which can be used as a marker to distinguish diseased from healthy cells and thereby allow the downstream analysis of their movement—on top of the additional monitoring layer using a Cas9-based recordable barcode strategy. 

"Having a system where we can follow tumors in live mice and then visually detect tumor cells in dissected organs allows us to pinpoint exactly where we can find each of the barcodes as the cancer cells move throughout the body," states Ryan Serio Ph.D., lead author in the study and a postdoc in the Nowak lab.  

One of the elements in the lentivirus injected into the prostate of the mice is a barcode comprised of a sequence of 260 base pairs, which in cells allows Cas9 enzymes that make edits to the barcode, Nowak explains. The barcode thereby gets integrated into the genome so that the marks made by Cas9 get passed to the next generation of the cell. 

Thanks to the EvoTraceR software, developed jointly by Nowak’s and Siepel’s labs, researchers could use short sequences of DNA as genetic barcodes to trace the movement of individual cancer cells in a high-throughput manner. The analysis generates FASTQ files that store sequence data and quality scores for that data, which get collected from organs based on the DNA extracted from different tissues with those cell-embedded barcodes. 

As a postdoc at Cold Spring Harbor Laboratory, Nowak worked with cancer biologist Lloyd Trotman, Ph.D., whose focus was on metastatic prostate cancer. Although Nowak and Siepel first met a decade ago, it was a few years before the “stars came into alignment” for their collaboration. 

By then, Nowak had his new permanent position at Cornell and the tools in place to do this type of biology reconstruction. He and Siepel decided to team up on a grant proposal for the clonal lineage tracing study and the rest is history.  

View Forward

Researchers elsewhere have similarly produced combined data on barcodes and gene expression, says Siepel, adding that the information will inform new computational methods his lab is working on that can be applied to these datasets as well as those being generating by Nowak. 

Bladder cancer and metastatic bladder cancer are among his newer areas of focus, Nowak says. His system for understanding the kinetics of cancer metastasis is applicable even to “non-cancer diseases where some kind of common clonal expansion takes place.” 

What’s intriguing about prostate cancer is that it typically exhibits “organ tropism,” whereby primary cancer cells preferentially spread to the bone in lieu of other organs, he continues. Untangling the reason for that affinity, and the adaptations employed, could one day help personalize treatments to different metastatic sites. The unknowns at present are whether cells’ preference for one site over another is “a random process, a little bit more of a deterministic process, or a combination of both.”   

Among the forces driving evolutionary solutions in cancer is the actual therapy used to treat it, says Nowak. His lab is therefore looking at the effect of androgen deprivation therapy—the primary systemic treatment for prostate cancer—on the evolutionary trajectory of the disease using his clonal lineage tracing system, to better block resistant cancer cells from adapting and thriving under therapeutic selection. 

“Cancer itself is an evolutionary process,” Siepel points out. “Tumors evolve in a similar way as populations of organisms evolve, in this case with cells competing with one another and the cells that tend to grow faster becoming dominant. It is evolution gone wrong, in the sense that they are not the cells we would like to proliferate.”