Natural Killer cells and Innate Immunity

The immune system is classically divided into innate and adaptive. Adaptive immunity can be defined by the presence of cells (i.e. T and B lymphocytes in higher vertebrates) that clonally express a colossal repertoire of receptors (i.e. the T cell and the B cell antigen receptors), the diversity of which results from somatic DNA rearrangements.
Besides T and B cells, NK cells are lymphocytes of the innate immune system that can kill an array of target cells and secrete cytokines that participate to the shaping of the adaptive immune response and to tissue repair.
A feature of NK cells resides in their capacity to distinguish stressed cells (such as tumor cells, microbe-infected cells, cells which have undergone physical or chemical injuries) from normal cells.

Our laboratory is focused on understanding NK cell detection systems and NK cell tolerance.

 

Recently the team has revealed a curious paradox: by removing one of the main systems of NK cell activation in mice, they were able to increase anti-tumor properties of these cells and the resistance of the animals to viruses.

Overview

The NK cell detection system includes a variety of cell surface activating and inhibitory receptors, the engagement of which regulates NK cell activities. Thus, the integration of antagonistic pathways upon interaction with neighboring cells governs the dynamic equilibrium regulating NK cell activation and dictates whether or not NK cells are activated to kill target cells.
NK cells use inhibitory receptors to gauge the absence of constitutively expressed self-molecules on susceptible target cells. In particular, NK cells express MHC class I-specific receptors and ‘lose’ inhibitory signals when encountering MHC class I–deficient hematopoietic cells in several in vitro and in vivo models. As a consequence, NK cells can recognize ‘missing self’ on hematopoietic cells.
By interacting with MHC class I molecules that are constitutively expressed by most healthy cells in steady-state conditions but that may be lost upon stress, inhibitory MHC class I receptors provide a way for NK cells to ensure tolerance to self while allowing toxicity toward stressed cells.

Missing-self and NK cell education

MHC class I-specific inhibitory receptors and their ligands (Ly49 dimers and H-2 in mice; killer cell immunoglobulin-like receptors (KIRs) and HLA in humans) are highly polymorphic molecules encoded by multigenic, multiallelic families of genes that are inherited independently.
NK cells have thus to discriminate self in a context where self-molecules differ from individuals to individuals. Like T lymphocytes, NK cells are educated to self versus altered-self discrimination, but the molecular strategies involved in this education are different. T cell education involves the stimulatory T cell receptor whereas NK cell education is mediated through the engagement of the MHC class I-specific inhibitory receptors.

This education, also termed ‘‘tuning, licensing or arming’’ leads to the maturation of a NK cell functional repertoire (i.e., the ensemble of stimulation toward which NK cells are reactive), which is adapted to self-MHC class I environment. Consequently, NK cells in MHC class I-deficient hosts are hyporesponsive to stimulatory receptor stimulation and thereby tolerant to self. Other studies have reported that the hyporesponsiveness of NK cells grown in a MHC class I-deficient environment can be overcome by inflammatory conditions in NK cell environment. It remains that 2 types of self-tolerant NK cells coexist in vivo at steady-state: functionally competent NK cells, whose effector responses are inhibited by the recognition of self MHC class I molecules, and hyporesponsive NK cells that cannot detect self-MHC class I.

The molecular mechanisms underlying the MHC-dependent NK cell education are still unknown, but have been shown in mice to require a functional ITIM in the intracytoplasmic tail of Ly49 inhibitory receptors.

Several studies have suggested that the manipulation of NK cell missing-self recognition may have important clinical benefit in leukemic patient.
The manipulation of NK cell alloreactivity in these settings implies haploidentical hematopoietic transplantations, which are associated with considerable adverse effects, including graft versus host disease mediated by allogenic T cells. A safer strategy would be to block NK cell inhibitory receptors in an autologous setting. Such a strategy is currently tested in phase II clinical trials with a fully human anti-KIR monoclonal antibody (1-7F9 developed by Innate-Pharma). This monoclonal antibody recognizes KIR2D inhibitor receptors and blocks their interaction with the human MHC class I molecules HLA-C, leading to NK cell-mediated lysis of leukemic cells. However, one of the main concerns for using this therapeutic approach in humans is the risk of generating a strong reactivity against normal self-tissues and/or to interfere with NK cell education.

Therefore the precise understanding of NK cell education mechanisms is not only critical to describe this process as a model of education to self reactivity for cells of the innate immune system, but it is also pivotal for the development of innovative therapeutic strategies based on the manipulation of NK cell immunity.

Stress-induced self recognition

In addition to the recognition of microbial molecules by a variety of innate immune receptors, the so-called “infectious non-self recognition”, it has been shown that some receptors of innate immune cells can detect internal changes, leading to the concept of “stress-induced self recognition”.
This mode of detection is based on the recognition of molecules whose expression is barely detectable in steady-state conditions, but induced upon various forms of stress. A prototypical example of this mode of detection is illustrated by the activation of NK cells via engagement of the NKG2D receptor, which interacts with self-molecules selectively up-regulated on stressed cells.

Besides NKG2D, NK cell express an array of cell surface molecules, such as the Natural Cytotoxicity Receptors (NCR), which have been shown since more than a decade to be involved in the activation of NK cells by tumor cells. The NCR family includes NKp46, NKp44 and NKp30. However, the NCR ligands that are supposedly expressed on tumor cells and activate NK cells are still unknown at the noticeable exception of B7-H6 that we recently identified. One important aspect of our program is to characterize the nature and the regulation of these ligands.

NK cell memory

Learning, a hallmark of life, produces adaptation to new information. The immune system, like the nervous system, has this ability to learn from previous experience, such as a single encounter with the many pathogens that exist. The result is immunological memory that confers long-lasting protection.

Until now, immunological memory was thought to be a feature of the adaptive immune system. Unexpectedly, recent studies revealed that NK cells could be players in the persistence of immunity, although they have traditionally been considered to be part of the innate immune system. NK cells can retain in vivo an intrinsic memory of a prior in vitro activation, which is maintained across cell divisions.

These results prompt to research into the mechanisms that allow the boosted effector function of NK cells to be maintained across cell divisions, in particular the epigenetic marks associated with various stages of these cells’ activation.

Specific aims

A central issue in our project is to understand the biological function and mode of action of the NK cell, as a cellular system model of a player in innate immunity. Specifically our multidisciplinary approaches will be combined to address three main issues as follows:

  • How NK cells are educated?
  • How NK cells distinguish their targets from normal cells?
  • How NK cells participate to immunological memory?


Several new mouse models have been and will be generated in the lab, including a series of knock-in mice based on the use of a NK-specific promoter region, mice in which NK cell molecules will be tagged in vivo for imaging studies as well as mice obtained by random germline mutagenesis via ENU. Pan-genomic epigenetic studies will be also conducted.

Finally, advanced techniques of nanoscopy will be implemented. We believe that the completion of the proposed tasks from the nanoscopic to the macroscopic scales will contribute to draw an integrated view on the innate immune responses at molecular, cellular and organismal levels. This is expected to reassess the actual concepts on the links between NK activation, tissue homeostasis and stress response, and to help the design of NK cell-based therapies.