Extended Data Figure 4: Predictors of plasma VAF.

a, Plasma VAFs of SNVs detected in plasma in 46 patients who were ctDNA-positive (two or more SNVs detected). Clonal (blue) and subclonal (red) VAFs are indicated, mean is shown as a horizontal line. Driver variants are shown as triangles. b, Mean clonal VAF correlated with maximum tumour size measured in a post-surgical specimen (pathologic size, n = 46). Grey vertical bars represent range of clonal VAF; shaded red background indicates the 95% confidence interval. c, Filtering steps taken to define a group of ctDNA-positive patients with volumetric data considered adequate to model tumour volume and plasma VAF. d, Scatter plot showing mean clonal VAF relative to tumour volume for TRACERx (blue dots and fitted blue line, n = 37) and VAF relative to volume for previously published data¹⁶ based on CAPP-seq analysis of ctDNA (orange dots and orange fitted line, n = 9). Orange shaded background indicates the 95% confidence interval based on CAPP-seq data. e, Mean clonal VAF correlated with tumour volume × tumour purity (cancer cell volume), n = 37. Shaded red background indicates the 95% confidence interval. f, Association between the number of cancer cells and VAF of clonal SNVs in plasma based on linear modelling of Extended Data Fig. 4e and the assumption that a cancer cell volume of 1 cm³ contains 9.4 × 10⁷ cells¹⁷. g, Detected subclonal SNVs were mapped back to M-seq-derived tumour phylogenetic trees (process illustrated in the key). Detected private subclones (subclones identified within only a single tumour region) are coloured red. Shared subclones (subclones detected in more than one tumour region) are light blue. Subclonal nodes were sized on the basis of the maximum recorded CCF. The top row of the phylogenetic trees represent subclonal nodes targeted by primers within that patient’s assay panel, the bottom row represent subclonal nodes detected in ctDNA, within this row grey subclonal nodes represent subclones that were not detected in ctDNA.