High-dimensional mass cytometry analysis of NK cell alterations in Acute Myeloid Leukemia identifies a subgroup with adverse clinical outcome

Abstract

Natural killer (NK) cells are major anti-leukemic immune effectors. Leukemic blasts have a negative impact on NK cell function and promote the emergence of phenotypically and functionally impaired NK cells. In the present work, we highlight an accumulation of CD56-CD16+ unconventional NK cells in acute myeloid leukemia (AML), an aberrant subset initially described as being elevated in patients chronically infected with HIV-1. Deep phenotyping of NK cells was performed using peripheral blood from patients with newly-diagnosed AML (N=48, HEMATOBIO cohort, NCT02320656) and healthy subjects (N=18) by mass cytometry. We evidenced a moderate to drastic accumulation of CD56-CD16+ unconventional NK cells in 27% of patients. These NK cells displayed decreased expression of NKG2A as well as the triggering receptors NKp30, and NKp46, in line with previous observations in HIV-infected patients. High-dimensional characterization of these NK cells highlighted a decreased expression of three additional major triggering receptors required for NK cell activation, NKG2D, DNAM-1, and CD96. A high proportion of CD56-CD16+ NK cells at diagnosis was associated with an adverse clinical outcome, with decreased overall survival (HR=0.13; P=.0002) and event-free survival (HR=0.33; P=.018), and retained statistical significance in multivariate analysis. Pseudo-time analysis of the NK cell compartment highlighted a disruption of the maturation process, with a bifurcation from conventional NK cells toward CD56-CD16+ NK cells. Overall, our data suggest that the accumulation of CD56-CD16+ NK cells may be the consequence of immune escape from innate immunity during AML progression.

Dynamic evaluation of blood immune cells predictive of response to immune checkpoint inhibitors in NSCLC by multicolor spectrum flow cytometry

“…Upon target recognition, CD16+CD56dim NK cells release perforin and granzyme granules and mediate antibody-dependent cellular cytotoxicity through CD16 (FcɣRIII) to clear cancer cells. In contrast, increased CD16+CD56-NK cells were associated with immune escape from innate immunity during AML progression (42).…”

High dimensional analysis reveals distinct NK cell subsets but conserved response to stimulation in umbilical cord blood and adult peripheral blood

“…The regulation NK cell functions through immune checkpoints has recently generated much interest given its importance to clinical outcomes [28]. Our data indicate that TIM-3, TIGIT, and CD96 are the main immune checkpoints expressed by unstimulated NK cells from healthy donors, an observation that corroborates previous reports [25,28,34]. Interestingly, while immune (H-I) NK cells were expanded as per (A-F) and stimulated overnight with 10 ng/ml IL-12, 50ng/ml IL-15, 50 ng/ml IL-18.…”

“…We applied a 20-color flow cytometry panel (Table S2) to define immune checkpoint and NK cell receptor expression on UCB and APB NK cells. As per previous studies [28], NK cells were defined as CD3 − CD33 − CD19 − cells expressing either CD56 or CD16 (Figure S1). This gating strategy ensured the exclusion of CD16 + monocytes but also excluded a minor population of CD56 bright cells positive for NKG2D, NKp46, and NKp30 (Figure S2) that might correspond to previously described CD33 + NK cells [29].…”

Combined flow cytometry natural killer immunophenotyping and KIR/HLA-C genotyping reveal remarkable differences in acute myeloid leukemia patients, but suggest an overall impairment of the natural killer response

“…Chretien et al (78) were able to show that AML patients with a particular mutation (inversion 3) developed a CD16+ CD56unconventional NK subpopulation, representing more than 20% of the total NK cells. Combined with the lower percentages of conventional NKs, this might offer the basis for an important immune escape mechanism.…”