Clonal Evolution of Pre-Leukemic Hematopoietic Stem Cells in Human Acute Myeloid Leukemia
Authors: Majeti Ravi 
Published: October 2019

Acute myeloid leukemia (AML) is an aggressive malignancy of hematopoietic progenitors with poor clinical outcomes. Recent genome-scale sequencing efforts have determined that on average, an individual AML case is associated with 5 somatic mutations in recurrently mutated genes. This finding raises the important question of how AML develops from normal hematopoietic stem and progenitor cells. Given that AML is characterized by the sequential acquisition of genetic lesions in a single lineage of cells, and that all cells in the myeloid lineage, apart from HSC, are short-lived, we proposed a model in which serial acquisition of mutations occurs in self-renewing HSC. We investigated this model and the nature of founder mutations through the genomic analysis of de novo AML and patient-matched residual HSC. Using exome sequencing, we defined mutations present in individual AML genomes from 19 cases, and screened for these mutations in the residual HSC. We identified multiple mutations present in residual HSC retaining normal multilineage differentiation in vivo, including mutations in IDH1/2, TET2, DNMT3A, and genes encoding the subunits of the cohesin complex. Through single cell analysis, we determined that as we hypothesized, a clonal progression of multiple mutations occurs in HSC. From these studies, we identified patterns of mutation acquisition in human AML. Our findings support a model in which mutations in "landscaping" genes, involved in global chromatin changes such as DNA methylation, histone modification, and chromatin looping, occur early in the evolution of AML, while mutations in "proliferative" genes such as FLT3 and KRAS occur late. Using this approach, we identified pre-leukemic HSC in a larger cohort of AML patients, and determined that their frequency within the stem cell compartment at the time of diagnosis varied widely from undetectable to nearly 100% of the cells. Stratifying these patients into two groups with either high or low frequencies of pre-leukemic HSC demonstrated that patients in the high group had much worse overall and relapse-free survival than those in the low group, indicating that the presence of pre-leukemic HSC may be critical for eventual clinical outcomes. To further investigate the response of pre-leukemic HSC to treatment, we analyzed the persistence of pre-leukemic mutations in patients in remission and found CD34+ progenitor cells and various mature cells that harbor pre-leukemic mutations. These findings indicate that pre-leukemic HSC can survive induction chemotherapy, identifying these cells as a potential reservoir for the re-evolution of relapsed disease. Finally, through the study of several cases of relapsed AML, we demonstrate various evolutionary patterns for the generation of relapsed disease, and show that some of these patterns are consistent with involvement of pre-leukemic HSC. Thus, our studies of pre-leukemic HSC reveal the clonal evolution of AML genomes from founder mutations, suggest a potential mechanism contributing to relapse, and constitute a cellular reservoir that may need to be targeted for more durable remissions.

Disclosures

Majeti: Forty Seven, Inc.: Consultancy, Equity Ownership, Membership on an entity's Board of Directors or advisory committees.