Fig. 4: SPRINTER reveals a link between clone proliferation and metastatic seeding, and clone-specific ART present in distinct metastatic clades.

a, Tumor phylogeny was reconstructed for SPRINTER’s single-cell clones (tree leaves) from patient CRUKP9145 (colored by sample, with clones uniquely shaded). Seeding clones (dark gray) and ancestral clones (white with border colored according to inferred anatomical site) were inferred, with some clones harboring ctDNA-tracked SNVs (Roman numerals). b, Phylogeny from a with clones colored by SPRINTER’s S fractions. c, Across samples (anatomical location indicated as circles on body map), metastatic migrations (arrows) were inferred, and metastatic clades (blue, green and pink with corresponding clones indicated in tree) were defined based on primary tumor seeding clones. The figure is created with BioRender.com. d, In the two main phylogenetic branches containing different metastatic clades (top row), SPRINTER inferred ART (colored rectangles) for each clone (second row) for genes (left) known to impact proliferation or metastatic potential, with reference replication timing derived from normal cells shown (left column). ART is supported by related gene expression changes measured using bulk RNA sequencing (right heatmap), with late-to-early and early-to-late ART associated with increased and decreased gene expression, respectively (P values derived using a two-sided Wald test with a Benjamini–Hochberg correction with family-wise error rate = 0.05). *P < 0.1, **P < 0.05 and ***P < 0.01. e, For each SPRINTER clone (dot) in the primary tumor (dark blue) or metastases (orange), the seeding genetic distance (x axis) computed with respect to the closest seeding clone based on either SNVs (left) or CNAs (right) was compared to SPRINTER’s S fraction (y axis) using two-sided Pearson correlation tests (correlation coefficients and P values reported), and the 95% CI was calculated for linear regressions (shaded areas). f, For each ctDNA-tracked clone (dot), a ctDNA shedding index (x axis) was calculated using the frequency of SNVs for either (left) SPRINTER single-cell clones or (right) previous bulk clones and compared to the maximum S fraction inferred from descendant SPRINTER clones (y axis). In each case, a two-sided Spearman correlation test was performed (with correlation coefficients and P values reported), and the 95% CI was calculated for linear regressions (shaded areas).