IL-12 directs further maturation of ex vivo differentiated NK cells with improved therapeutic potential.

Dorit Lehmann,Dirk Groenewegen,Jan Spanholtz,Caterina Sturtzel,Bernhard Schlechta,Marleen Tordoir,Erhard Hofer,Dan Kaufman

doi:10.1371/journal.pone.0087131

Abstract

The possibility to modulate ex vivo human NK cell differentiation towards specific phenotypes will contribute to a better understanding of NK cell differentiation and facilitate tailored production of NK cells for immunotherapy. In this study, we show that addition of a specific low dose of IL-12 to an ex vivo NK cell differentiation system from cord blood CD34+ stem cells will result in significantly increased proportions of cells with expression of CD62L as well as KIRs and CD16 which are preferentially expressed on mature CD56dim peripheral blood NK cells. In addition, the cells displayed decreased expression of receptors such as CCR6 and CXCR3, which are typically expressed to a lower extent by CD56dim than CD56bright peripheral blood NK cells. The increased number of CD62L and KIR positive cells prevailed in a population of CD33+NKG2A+ NK cells, supporting that maturation occurs via this subtype. Among a series of transcription factors tested we found Gata3 and TOX to be significantly downregulated, whereas ID3 was upregulated in the IL-12-modulated ex vivo NK cells, implicating these factors in the observed changes. Importantly, the cells differentiated in the presence of IL-12 showed enhanced cytokine production and cytolytic activity against MHC class I negative and positive targets. Moreover, in line with the enhanced CD16 expression, these cells exhibited improved antibody-dependent cellular cytotoxicity for B-cell leukemia target cells in the presence of the clinically applied antibody rituximab. Altogether, these data provide evidence that IL-12 directs human ex vivo NK cell differentiation towards more mature NK cells with improved properties for potential cancer therapies.

Highlights

Natural killer (NK) cells are innate lymphocytes that exhibit cytotoxic and immunoregulatory functions upon activation
That preferentially low doses of interleukin 12 (IL-12) induce the generation of increased proportions of cells with expression of CD62L, CD16 and killer cell immunoglobulin-like receptors (KIRs) and a specific chemokine receptor repertoire without significantly affecting the amplification of the cells during differentiation
The recently established ex vivo differentiation system developed by Glycostem Therapeutics for large scale generation of human NK cells is of interest for studies of human ex vivo NK cell development and holds great potential for adoptive immunotherapies of cancer [15,16,17]. This system is amenable to modifications, which opens the possibility to study specific effects of individual cytokines on human NK cell differentiation and to generate tailored NK cell products with specified phenotypes and functions to target malignancies

Summary

Introduction

Natural killer (NK) cells are innate lymphocytes that exhibit cytotoxic and immunoregulatory functions upon activation In humans these functions are correlated with two distinct NK cell phenotypes, namely the preferentially cytokine producing CD56bright NK cells that are most prominently found in secondary lymphoid tissues and the blood resident CD56dim NK cells preferentially exerting killing of virus-infected and transformed cells [1,2,3]. Both NK cell subtypes express a typical range of activating and inhibiting receptors balancing their activity.

Objectives

Methods

Results

Conclusion