Clonality and NGS-MRD. Schematic depiction of the polyclonal evolution and trackable somatic mutations from CH, through CHIP and CCUS to MDS/AML. Each colored dot within the cell represents a distinct mutation, with 2 different transformed clones (dark circles) developing over time and one outcompeting the other (so called clone sweeping³⁹). NGS of the bulk population can only detect genetic alterations with a frequency above the LOD, which depends on the error-corrected sequencing methodology used. The VAF represents the variant frequency within the bulk population without information on the co-occurrence of variants within a single subclone of that population that is under constant intrinsic competition and extrinsic pressure (treatment). Depending on the bulk composition, the same VAF can represent different mutational states on a single-cell level (demonstrated by chromosomes in the right lower corner of the figure), such as biallelic vs monoallelic mutation, homozygous vs hemizygous or heterozygous mutations. Importantly, such allelic imbalances, eg, biallelic TP53 mutation, are not limited to MDS/AML but can also be found in CH, CHIP, and CCUS. CTx, chemotherapy. The figure was adapted and modified from the study by Stauber et al.⁴⁰