Selective removal of alloreactive cells from haematopoietic stem cell grafts: graft engineering for GVHD prophylaxis

One of the main goals in allogeneic bone marrow transplantation is the abro gation of graft-versus-host disease with the preservation of antileukaemia and antiviral activity. We have established a novel system for the selectiv e removal of alloreactive lymphocytes from donor grafts while retaining an effective allogeneic response to third-party stimulator cells. Initial feas ibility studies were done with unrelated HLA-mismatched pairs and then exte nded into the matched setting. Mononuclear cells from HLA-matched donors we re cocultured with irradiated recipient cells prestimulated with cytokines (gamma-IFN and TNF-alpha) in a modified mixed lymphocyte culture (MLC), All oreactive donor lymphocytes were identified by expression of CD69, an early activation marker and selectively removed by paramagnetic bead sorting. Th e remaining 'non-alloreactive' lymphocytes were tested in proliferative ass ays against the original matched recipient and to a third-party donor. A me an depletion of proliferative capacity to 11.5 +/- 9.9% of the original mat ched recipient response was achieved while the residual third-party respons e was largely preserved at 77.8 +/- 20.9% which should translate into impro ved immune reconstitution and preservation of antiviral activity. The non-a lloreactive lymphocytes could also possess functional antileukaemia activit y. Moreover, the alloreactive cells are easily recoverable in this selectiv e T cell depletion strategy for cryopreservation and ready for immediate ac cess as therapeutic donor lymphocyte infusions in cases of frank relapse po st transplant.