(a) Six mutation clusters (marked a–f) constructed from MAF at diagnosis (D) and relapse (R). The splice mutation in PMS2 (A725_E12splice) is placed to cluster d because it is located on a haploid region due to loss of chromosome 7p at relapse. (b) Comparison of mutation spectra of relapse-specific clusters (c–f) and all diagnosis (a+b) mutations. Relapse-specific clusters (d,e) show significantly higher (P<0.05, Fisher’s exact test; indicated by an asterisk) transition mutation rates (and mutation counts) than clusters c and a+b. (c) Clonal lineages from diagnosis to relapse and the ambiguous lineage of cluster e,f are resolved with the input from mutation spectrum analysis. Specifically, clone 5, which contains mutation cluster e with a total of 98 SNVs at a MAF of 0.15, can be considered as a descendant of either clone 4 or clone 3. As the mutation spectrum for cluster e is highly enriched for transition changes, consistent with cluster d mutations in clone 4 but significantly different (P=0.01, Fisher’s exact test) from that of cluster c mutations in clone 3, it is placed as a descendant of clone 4 instead of clone 3. Clone 6 contains 21 lineage-specific mutations of cluster f with a MAF of ~0.05. It would have an ambiguous clonal linage that could have descended from clone 3, or clone 4 or clone 5 if compatibility of population frequency is the only criteria for determining lineage. It was placed under clone 3 as its mutation spectrum is significantly different from that of clones 4 and 5 but not from that of clone 3. (d) Probe intensity of SNP array in germline (G), diagnosis (D) and relapse (R) on chromosome 7. Isochromosome 7q (that is, loss of 7p and gain of 7q) demonstrated a low signal intensity due to low tumour purity at diagnosis and is present in all tumour cells at relapse.