Applications and limitations of controlled GVL by HSV-TK donor lymphocytes infusion after T-depleted allogeneic bone marrow transplantation

ïn the context of T-depleted allogeneic bone marrow transplantation (BMT), delayed donor lymphocytes infusion plays a central therapeutic role in both graftversus-leukemia and immune reconstitution. However, the use of donor lymphocytes is limited by the risk of severe graft-vs-host disease (GvHD). Patients who relapsed or developed an Epstein-Barr virus (EBV)-induced lymphoma after BMT were treated with donor lymphocytes transduced with the thymidine kinase suicide gene of the Herpes Simplex virus (HSV-tk). HSV-tk confers to donor lymphocytes selective in vivo sensitivity to the drug ganciclovir allowing a specific treatment in case of GvHD thus avoiding immunosoppressive therapies (Bonini C. et al., Science 276:1719, 1997). However, three patients treated with HSV-tk genetically engineered donor T cells developed a specific immune response against the tk gene, which resulted in immediate elimination of genetically engineered cells. This phenomenon was observed in patients who received the first infusion of genetically modified donor cells late after BMT, documenting the role of the recipient immune reconstitution. One patient, who developed chronic GvHD, showed partial resistance to ganciclovir-mediated killing of transduced cells Circulating PBLs obtained from the patient before and after ganciclovir treatment showed no difference in ganciclovir sensitivity in vitro and high levels of expression of the cell surface marker gene LANGFR, suggesting that the ganciclovir resistance of chronic GvHD is mainly related to the cell-cycle dependence of the HSVTK/ganciclovir treatment and that no rearrangement occurred. To circumvent the intrinsic limitations of the HSV-tk based stategy, which are mainly immunogenicity and cell cycle dependence, a different suicide gene strategy has been developed. The strategy is based on the transfer of a fusion gene encoding for two copies of a modified FK506-binding-protein (FKBP) fused to the intracytoplasmic domain of FAS. The exposure of manipulated cells to a non toxic cell-permeable synthetic analog of FK506 results in binding of the FKBP components producing oligomerization of FKBPs and triggering of FAS-mediated cell death. In vitro data show that the ligand successfully triggers apoptosis in retroviral transduced human lymphocytes. This vector represents a new and promising suicide gene strategy potentially allowing to circumvent the intrinsic limitations of the timidine kinase based strategy.