T and NK cell gene therapy for haemophagocytic lymphohistiocytosis due to perforin deficiency

Supervisors: Professor Bobby Gaspar and Dr Marlene Carmo

Haemophagocytic lymphohistiocytosis (HLH) is a severe disorder of immune dysregulation characterized by failure of effector cell cytotoxicity in which affected individuals suffer multiorgan complications due to hypercytokinemia and histiocytic infiltration1. Mutations in the perforin gene give rise to the most common form of HLH2. On contact with virally infected target cells, effector T and NK cells form an immunological synapse with exocytosis of specialized granules that mediate cell cytotoxicity. Perforin is contained within these granules and upon release forms pores on the target cell membrane allowing the passage of granzymes which initiate apoptotic pathways, eventually leading to target cell death3. Mutations in the perforin gene (PRF) leads to a failure of pore formation and target cell cytotoxicity in effector cells resulting in a proliferation of highly activated cytotoxic T and NK cells, which in turn lead to hypercytokinaemia and subsequent clinical sequelae of HLH. Treatment of this disease is currently limited to the control of infectious triggers and chemotherapeutic regimes to control the initial dysregulation, but permanent correction can only be achieved by allogeneic hematopoietic stem cell (HSC) transplantation which has high mortality in mismatched donor transplants4. The development of alternative strategies based on gene transfer may have great benefit for such patients.

Experiments performed in our lab show that introduction of the correct copy of the perforin gene into HSCs can restore the T and NK cell defects seen in perforin deficient mice and that reconstituted mice are protected against development of HLH after viral challenge. These initial experiments provide a proof of principle for this gene therapy approach. However, it is known that hematopoietic stem cell transplant into patients with active disease have a survival rate of only 50%1 and this may also affect the results of hematopoietic stem cell gene therapy.

The main aim of this project is to develop a control therapy for HLH due to perforin deficiency using corrected autologous effector T and NK cells. This therapy can be used in patients refractory to conventional chemotherapy or in patients prior to hematopoietic stem cell transplant that have active disease. To achieve this aim we need to choose the best population of effector cells to give correction of the cytotoxic defects, develop a vector system that gives good transduction efficiency in effector cells leading to correction of functional defects both in vitro and in vivo and finally verify if introduction of perforin reconstituted cells can control or ameliorate virus induced disease in the mouse model of HLH. Amelioration of disease in such experiments will provide encouraging data to support correction of human effector cells to try and control disease in chemotherapy refractory patients.

1) Henter JI et al. Blood. 2002;100(7):2367-2373.
2) Gholam C et al. Clin Exp Immunol. 2011;163(3):271-283.
3) Katano H, Cohen JI. 2005;128(6):739-750.
4) Bode SF et al. Arthritis Res Ther. 2012;14(3):213.
5) Jordan MB et al. Blood. 2004;104(3):735-743