Chromosomal instability is a major driver of intratumoral heterogeneity (ITH), promoting tumor progression. In the present study, we combined structural variant discovery and nucleosome occupancy profiling with transcriptomic and immunophenotypic changes in single cells to study ITH in complex karyotype acute myeloid leukemia (CK-AML). We observed complex structural variant landscapes within individual cells of patients with CK-AML characterized by linear and circular breakage–fusion–bridge cycles and chromothripsis. We identified three clonal evolution patterns in diagnosis or salvage CK-AML (monoclonal, linear and branched polyclonal), with 75% harboring multiple subclones that frequently displayed ongoing karyotype remodeling. Using patient-derived xenografts, we demonstrated varied clonal evolution of leukemic stem cells (LSCs) and further dissected subclone-specific drug–response profiles to identify LSC-targeting therapies, including BCL-xL inhibition. In paired longitudinal patient samples, we further revealed genetic evolution and cell-type plasticity as mechanisms of disease progression. By dissecting dynamic genomic, phenotypic and functional complexity of CK-AML, our findings offer clinically relevant avenues for characterizing and targeting disease-driving LSCs.

染色体不稳定性是瘤内异质性 （ITH） 的主要驱动因素，促进肿瘤进展。在本研究中，我们将结构变异发现和核小体占有率分析与单细胞中的转录组和免疫表型变化相结合，以研究复杂核型急性髓性白血病 （CK-AML） 中的 ITH。我们观察到 CK-AML 患者单个细胞内复杂的结构变异景观，其特征是线性和循环断裂-融合-桥接周期和染色体脱浆。我们在诊断或挽救 CK-AML 中确定了三种克隆进化模式 （单克隆、线性和分支多克隆），其中 75% 携带多个亚克隆，这些亚克隆经常表现出持续的核型重塑。使用患者来源的异种移植物，我们证明了白血病干细胞 （LSC） 的不同克隆进化，并进一步剖析了亚克隆特异性药物反应谱，以确定 LSC 靶向疗法，包括 BCL-xL 抑制。在配对的纵向患者样本中，我们进一步揭示了遗传进化和细胞型可塑性是疾病进展的机制。通过剖析 CK-AML 的动态基因组、表型和功能复杂性，我们的研究结果为表征和靶向疾病驱动的 LSC 提供了临床相关的途径。