The genetic information of a cancerous tumour is typically obtained by sequencing the tumour cells together, rather than analysing them at the single-cell level. However, we are now beginning to realise the value in understanding cancer at the single-cell level, as the sub-clones could provide important insights into how the cancer progresses, its spread and why it becomes drug-resistant.
Researchers at the USC and 10x Genomics have developed a new technique that uses high-throughput single-cell DNA sequencing that creates a higher resolution into cancer studies than ever before. The researchers used a microfluidic droplet-based single-cell sequencing method, where they simultaneously sequenced the genomes of close to 1,5000 cells, revealing genetic diversity which was previously hidden in a well-studied melanoma cell line.
David Craig, the co-director of the Institute of Translational Genomics at Keck School of Medicine of USC said they used this approach to examine a standard cancer cell-line, examined thousands of times by many different labs but what was “really surprising” was that this technology uncovered a complexity that they did not expect. The cell line consistently became a mixture of different cell types but re-examining previous work with this new information has provided new insights into tumour evolution.
The study demonstrated the ability of single-cell sequencing for studying possible evolutionary trajectories of cancer cells.
The researchers used a process called “single-cell copy number profiling”, an emerging technique developed by 10x Genomics. To conduct the study, they performed shallow single-cell sequencing of genomic DNA across 1475 cells from the COLO829 cell line. This tumour-line has been characterised using multiple technologies and is a benchmark for evaluating somatic alterations. In some of the past studies, COLO829 has shown conflicting or indeterminate copy number, and therefore single-cell sequencing provides a tool for gaining insight.
The analysis revealed at least four major sub-populations of cells, known as clones, that are expected to have mutated from the original cancer cell. The authors wrote that “based on clustering, break-point and loss of heterozygosity analysis of aggregated data from sub-clones, we identified distinct hallmark events that were validated within bulk sequencing and spectral karyotyping”.
Lead author and assistant professor Velazquez-Villarreal said that studying cancer at this higher resolution can help discover information that lower resolution bulk sequencing misses. This was further supported by John Carpten, study author and co-director of the USC Institute for Translational Genomics, who said that there may be a small population of cells in a tumour that has acquired a change that makes them resistant to therapy, but if you were to take a tumour and grind it up and sequence it, you may not notice that change. However, when you go to the single-cell level, you not only see it but you can see the specific populations of cells that have acquired the change.
The researchers concluded that they hope that more cancer researchers will focus on single-cell sequencing. They are now using their technique to study genetic diversity in clinical cancer specimens to help understand the early molecular changes that can lead to aggressive and tough-to-treat cancers.
If you are interested in single-cell analysis, be sure to check out our virtual series in July – Single Cell ONLINE, a free to attend 4-part webinar series available live and on-demand.
Image credit: Medical photo created by kjpargeter – www.freepik.com