Clonal differences underlie variable responses to sequential and prolonged treatment

Cancer cells exhibit dramatic differences in gene expression at the single-cell level, which can predict whether they become resistant to treatment. Treatment perpetuates this heterogeneity, resulting in a diversity of cell states among resistant clones. However, it remains unclear whether these differences lead to distinct responses when another treatment is applied or the same treatment is continued. In this study, we combined single-cell RNA sequencing with barcoding to track resistant clones through prolonged and sequential treatments. We found that cells within the same clone have similar gene expression states after multiple rounds of treatment. Moreover, we demonstrated that individual clones have distinct and differing fates, including growth, survival, or death, when subjected to a second treatment or when the first treatment is continued. By identifying gene expression states that predict clone survival, this work provides a foundation for selecting optimal therapies that target the most aggressive resistant clones within a tumor. A record of this paper’s transparent peer review process is included in the supplemental information. •Melanoma clones were traced through multiple rounds of treatment•Cells within clones maintain gene expression similarities over 2 months of treatment•Different clones have distinct fates during continued treatment•Initial treatment can cause changes in sensitivity to secondary treatment Schaff et al. show that resistant cancer cell clones are different from one another, yet the cells within a clone remain similar during treatment. They also show that transcriptional differences between clones result in different cell fates when treated sequentially with different agents or continuously with the same agent.