Delineating the genetic pathway of Natural Killer cell production

We have identified the master gene (E4bp4) that causes blood stem cells to turn into disease-fighting ?Natural Killer? (NK) immune cells. This discovery could one day help scientists boost the body?s production of these frontline tumour-killing cells, creating new ways to treat cancer and other diseases including those caused by viral or parasite infections. NK cells ? a type of white blood cell - are a major component of the human body s innate, quick-response immune system. They provide a fast frontline defence against tumours, viruses and bacterial infections, by scanning the human body for cells that are cancerous or infected with a virus or a bacterial pathogen and killing them.
NK cells - along with all other types of blood cell, both white and red - are continuously generated from blood stem cells in the bone marrow over the course of a person?s lifetime. We would like to progress to a drug treatment for cancer patients which reacts with the protein expressed by their E4bp4 gene, causing their bodies to produce a higher number of NK cells than normal, to increase the chances of successfully destroying tumours. If increased numbers of the patient?s own blood stem cells could be coerced into differentiating into NK cells, via drug treatment, we would be able to bolster the body?s cancer-fighting force. This research proposal seeks to map out the genetic pathway controlled by the E4bp4 ?master gene?. If we know the components required to produce NK cells we may be able to use drugs to act selectively on one of those components to increase NK cell numbers and hence their disease fighting capability in sufferers.

Natural Killer (NK) cells are a distinct lineage of lymphocytes with both cytotoxic and cytokine (e.g. IFN-? and TNF) producing effector functions. NK cells play a central role in regulation of both the innate and adaptive immune response. They are associated with multiple functions and multiple pathologies such as cancer, microbial infections and autoimmune disease. In order to understand the basis of NK cell maturation and to manipulate NK cell numbers for therapeutic application, it is necessary to define the genetic pathway underlying NK cell development. We have recently identified the first gene, E4bp4, found to determine NK cell specific lineage determination (Gascoyne et al., 2009, Nature Immunology, published online Sept 13). By generating E4bp4-/- mice we showed that this gene is critical for NK cell development but not for NKT cells or CD8+ memory T cell production. There are no NK cells in the periphery of E4bp4-/- mice and the mice lack any detectable NK cell mediated cytotoxicity. E4bp4 has the remarkable ability to increase NK cell production when expressed in normal haematopoietic progenitors (HPCs), this highlights the possibility of manipulating NK cell numbers for therapeutic purposes. This research seeks to identify the genetic pathway used by E4bp4 to regulate NK cell production. DNA microarrays will be used to compare E4bp4-/- and +/+ HPCs to determine have their expression deregulated in the absence of E4bp4. Verification of targets will be carried out by real time PCR on -/- versus +/+ HPCs and wild type HPCs transduced by E4bp4. Chromatin immunoprecipitation will be used to define E4bp4 direct target genes. The functional of candidate target genes will be assessed by their retroviral-mediated overexpression and also shRNA-mediated knockdown in HPCs for readout by in vitro NK cell production assay. The function requirement of E4bp4 for the function and survival of mature NK cells (both mouse and human) will also be tested.