Differentiation of leukemic blasts is not completely blocked in acute myeloid leukemia

Abstract

Hematopoiesis, the formation of blood cells, involves the hierarchical differentiation of immature blast cells into mature, functional cell types and lineages of the immune system. Hematopoietic stem cells precisely regulate self-renewal versus differentiation to balance the production of blood cells and maintenance of the stem cell pool. The canonical view of acute myeloid leukemia (AML) is that it results from a combination of molecular events in a hematopoietic stem cell that block differentiation and drive proliferation. These events result in the accumulation of primitive hematopoietic blast cells in the blood and bone marrow. We used mathematical modeling to determine the impact of varying differentiation rates on myeloblastic accumulation. Our model shows that, instead of the commonly held belief that AML results from a complete block of differentiation of the hematopoietic stem cell, even a slight skewing of the fraction of cells that differentiate would produce an accumulation of blasts. We confirmed this model by interphase fluorescent in situ hybridization (FISH) and sequencing of purified cell populations from patients with AML, which showed that different leukemia-causing molecular abnormalities typically thought to block differentiation were consistently present in mature myeloid cells such as neutrophils and monocytes at similar levels to those in immature myeloid cells. These findings suggest reduced or skewed, rather than blocked, differentiation is responsible for the development of AML. Approaches that restore normal regulation of hematopoiesis could be effective treatment strategies.

Keywords: AML; differentiation block; mathematical model.

Fig. 1.

Conventionally hypothesized phylogeny of blood cell types in hematopoiesis. Adapted by permission of ref. , Springer Nature: Springer Science+Business Media, LLC, copyright 2009. Recent evidence supports a continuous differentiation process for hematopoiesis (21). Our model is independent of the particulars of the tree. Solid lines represent well-accepted differentiation paths. The dotted line represents a suspected path. LT-HSC is long-term hematopoietic stem cell. ST-HSC is short-term hematopoietic stem cell. MPP is multipotent progenitor. CMP is common myeloid progenitor. MEP is megakaryocyte-erythrocyte progenitor. GMP is granulocyte-macrophage progenitor. CLP is common lymphoid progenitor. DN is double negative thymocyte. DP is double positive thymocyte. NK is natural killer cell.

Fig. 2.

Model for reduced or skewed, rather than completely blocked, differentiation in AML. For each type of cell (σ), the model accounts for total cell number (N_σ), single-cell death rate (δ_σ), single-cell division rate (ρ_σ), and the probability (p_σ,τ) that each daughter cell differentiates into a particular type of cell (τ). The probability that the cell does not differentiate is p_σ,σ. (A) The ratio N_τ/N_σ can be effectively controlled by the (non)differentiation rate p_τ,τ. Since N_τ ≫ N_σ, we expect p_τ,τ $\underset{\approx }{<}$ 1/2. If p_τ,τ ≥ 1/2 and ρ_τ > 0, the system is no longer stable, and N_τ grows without bound. (B) In healthy hematopoiesis, differentiation rates in the hierarchy of cells are tightly regulated. This figure is a greatly simplified example with only 4 cell types and one feedback point. Here, A are stem cells, B and C are intermediate progenitors, and D is a mature cell type that does not divide or differentiate. (C–E) A mutation may perturb the differentiation rate of one of the cell types τ, so that p_τ,τ $\underset{\approx }{>}$ 1/2 rather than p_τ,τ $\underset{\approx }{<}$ 1/2, as shown in the panels.

Fig. 3.

Evolution over time with impaired differentiation in one compartment and regulation affecting either the growth or differentiation rate of compartment B representing intermediate progenitors. Here, A are stem cells, B and C are intermediate progenitors, and D is a mature cell type that does not divide or differentiate. (A) The growth in compartment A with impaired differentiation as p_A,A varies; all except p_A,A = 0.5 are exponential, but with different bases. (B) The healthy system. In Left, cell counts stay at their base levels, and in Right, the control of ρ_B (the growth rate of compartment B) is regulated and corrects for random up to ±25% changes in compartment D every cell cycle. (C) The ρ_B (the growth rate of B) is controlled and differentiation is impaired in (Left) compartment A, (Middle) B, or (Right) C. When differentiation in compartment B is impaired, the counts grow linearly, while all others are exponential. (D) The system with p_B,B (the differentiation rate of B) controlled and differentiation impaired in compartments A or C. The examples in D have a higher base growth rate than in C, which is why N_A grows differently in the 2 cases where its differentiation is impaired.

Fig. 4.

Leukemic genotypes are present at similar levels in both blast and mature, differentiated AML cells. Proportion of cells with (A−C) leukemic karyotype or (D) mutation, from 12 AML patients. Error bars represent a 95% binomial confidence interval, so smaller error bars in these measured proportions indicate that more cells of the given type were tested for the leukemic genotype. N for each point ranged from 2 to 112 cells; exact counts are provided in Dataset S2. T cell distributions were compared to each sorted leukemic cell population by 2-tailed Mann−Whitney U test: neutrophils (P = 1.3 × 10⁻⁴), monocytes (P = 3.9 × 10⁻⁴), and myeloid blasts (P = 1.3 × 10⁻⁵). Neutrophils and monocytes were not significantly different from myeloid blasts (P = 0.29 and 0.79, respectively), or from each other (P = 0.20).