The early expansion of a defective NKG2Apos/CD56dim/CD16neg NK cell subset represents a therapeutic target in haploidentical HSCT

Natural Killer (NK) cells are the first lymphocyte population to reconstitute early after non myelo-ablative and T cell-replete haploidentical hematopoietic stem cell transplantations (h-HSCTs) with post-transplant infusion of cyclophosphamide. The present study characterizes the transient and predominant expansion starting from the 2nd week after h-HSCT of a donor-derived unconventional subset of CD56dim/CD16neg (uCD56dim) NK cells expressing remarkable high levels of NKG2A and low levels of NKp46. Both transcription and phenotypic profiles indicated that uCD56dim NK cells are a distinct NK cell subpopulation with features of late differentiation, yet retaining proliferative capability and functional plasticity to generate conventional CD56bright/CD16pos NK cells in response to IL-15 plus IL-18. uCD56dim NK cells represent by far the largest NK cell subset detectable in the following 7 weeks after h-HSCT and they also express high levels of the activating receptors NKGD and NKp30 as well as of the lytic granules Granzyme-B and Perforin. Nonetheless, uCD56dim NK cells displayed a defective cytotoxicity that could be reversed by blocking the inhibitory receptor CD94/NKG2A. These data open new important perspectives to better understand the ontogenesis/homeostasis of human NK cells and to develop a novel immune-therapeutic approach by targeting the inhibitory NKG2A check point, thus enhancing NK cell alloreactivity early after h-HSCT. FACS-sorted NK cell subsets from the peripheral blood of healthy donors and their h-HSCT recipients were seeded at 0.1 x 106 cells/ml and cultured in presence of 30 U/ml rhIL-15 (Miltenyi) and 100 ng/ml rhIL-18 (MBL International Corporation) for 14 days. NK cell proliferation was determined by the analysis of 5-(and 6)-carboxyfluorescein diacetate succinimidyl ester (CFSE; Life Technologies) dilution. Before stimulation, cells were stained with 2 M CFSE for 7 min at 37C and then washed in FBS and medium.