Acute myeloid leukemia impairs natural killer cells through the formation of a deficient cytotoxic immunological synapse

Abstract

Acute myeloid leukemia (AML) cells are killed by allogeneic NK cells. However, autologous NK cells from AML patients express decreased levels of activating receptors, and show reduced cytotoxicity. Here, we investigated how interactions between NK and AML cells might cause loss of NK-cell activity in patients. Our results show that AML cell lines and primary blasts alter the NK-cell phenotype, reducing their cytotoxic potential upon prolonged contact. Downregulation of NK-cell-activating receptors was contactdependent and correlated with conjugate formation. Time-lapse imaging of HL60 AML cell line and NK-cell interactions showed a high proportion of noncytolytic contacts. Studies of NK-cell immunological synapses revealed a defect in lytic synapse formation. Namely, despite correct F-actin and LFA-1 recruitment, polarization of lytic granules toward primary blasts or AML cell lines was reduced. The NK-AML cell line synapses showed impairment of CD3ζ recruitment. Attempts to correct these synapse defects by cytokine stimulation of NK cells improved conjugate formation, but not granule polarization. Pretreatment of AML cell lines with the immunomodulating molecule lenalidomide significantly enhanced granule polarization. We speculate that combining immunomodulatory drugs and cytokines could increase AML cell sensitivity to autologous NK cells and reinforce the activity of allogeneic NK cells in adoptive immunotherapy.Keywords: Acute myeloid leukemia r Cancer immunity r Cytotoxicity r Immunological synapse r NK-cell-activating receptors Additional supporting information may be found in the online version of this article at the publisher's web-site [6,7]. AML cells express ligands interacting with NK-cell-activating receptors, making them susceptible to killing by NK cells [8,9]. However, autologous NK cells do not effectively control AML growth in patients. Some mechanisms have been suggested for AML escape, including downregulation of the ligands for NK-cellactivating receptors, secretion of soluble forms of these ligands, or upregulation of ligands for NK-cell inhibitory receptors [10]. Reduced toxicity has also been linked to downregulation of activating receptors on NK cells, particularly NKG2D and the natural cytotoxicity receptors (NCRs) NKp46 and NKp30 [11][12][13]. Upon remission, the NK-cell phenotype and function normalize, suggesting that the presence of AML blasts is responsible for the defects observed [12]. Interestingly, once out of the AML environment, NK cells that had previously expressed low levels of activating receptors regain their activity, becoming cytotoxic toward autologous leukemic blasts in mice [13]. This further implicates contact with AML cells in NK-cell activity down-modulation.Interaction between NK cells and a cancerous cell involves a series of steps resulting in the formation of an immunological synapse (NKIS) [14]. Upon first contact, the NK-cell cytoskeleton is reorganized in the zone of contact with the target cell (TC) [15]. Lytic granules are then polarized and their cont...

Defective NK Cells in Acute Myeloid Leukemia Patients at Diagnosis Are Associated with Blast Transcriptional Signatures of Immune Evasion

“…First, there is no correlation between the expression levels of the activating NK cell receptors and their cognate ligands on the tumor (data not shown; Ref. 49), and second, NK cell receptor's downregulation cannot be directly related to defects in IFN-g secretion after cytokinemediated stimulation. In this work, we showed a reciprocal relationship between the AML transcription profile and NK cell status in patients at diagnosis.…”

“…Previous studies described NK cell defects in AML patients including a decreased expression of NKp30 and NKp46 (10,47) and DNAM-1 (48). Such downregulation may be related to the engagement of the NK cell activating receptors by their ligands on the leukemic blasts (48,49). However, in vivo NK cell's defects are only partially explained by such mechanisms.…”

Natural killer cell immunosenescence in acute myeloid leukaemia patients: new targets for immunotherapeutic strategies?

“…In addition, AML blasts obtained from patients with an NCR low phenotype are susceptible to NK cell-mediated lysis using NCR bright NK cells, suggesting once more that AML blasts can be lysed by NK cells. Altogether, based on these observations it has been proposed that at some stage during AML development, NK cells turn out to be overwhelmed by the increasing number of leukaemic blasts, and their functionality goes down [14,21,31,69,73].…”

Loss of DNAM-1 ligand expression by acute myeloid leukemia cells renders them resistant to NK cell killing

“…Initially, it has been demonstrated that NK cells form low-frequency conjugates with AML cells. 35 Downstream of this early event, NK-AML cell conjugation events may not necessarily result in formation of a cytotoxic synapse, in which the tumor cell is successfully lysed. 35 Finally, intrinsic resistance to apoptosis may play a role in resistance to NKcell-mediated killing, downstream of granule entry.…”

“…35 Downstream of this early event, NK-AML cell conjugation events may not necessarily result in formation of a cytotoxic synapse, in which the tumor cell is successfully lysed. 35 Finally, intrinsic resistance to apoptosis may play a role in resistance to NKcell-mediated killing, downstream of granule entry. 36 Here, we demonstrate that poor NK cell function against AML cells originates through an early conjugation defect, which is deregulated through aberrant expression of DNAM-1 ligands.…”