Analysis of clonal hierarchy points toward cohesin as a secondary mutation. (A-B) Exemplary serial samples illustrating clonal architecture of cohesin family genes. (C) VAFs of STAG2 mutations by deep sequencing in serial samples. For each STAG2 mutant patient with available serial samples, the VAF in each sampling is shown. Although only 2 patients were found to harbor a STAG2 mutation in the dominant clone at the first time point, 7 out of 8 patients (87.5%) who were initially found to have a subclonal STAG2 mutation underwent clonal expansion, becoming the dominant clone at the time of transformation (P = .05). (D) VAFs of SMC3 and RAD21 mutations by deep sequencing in serial samples. For 4 patients with SMC3 mutations and the 1 patient with a RAD21 mutation with available serial samples, the VAF in each sampling is shown. All of these 5 patients harbored subclonal cohesin mutations at the first time point, with 2 out of 4 (50%) of the SMC3 mutant patients and the 1 out of 1 RAD21 mutant patient undergoing clonal expansion, becoming the dominant clone at the time of transformation.