Immunotherapy of EBV Post Solid Organ Transplantation Lymphoproliferative Disease with Genetically Engineered Haploidentical T Cells.

Abstract Epstein Barr Virus (EBV) associated post-transplantation lymphoproliferative disease (PTLD) is commonly treated with reduction of immune suppression, Rituximab and chemotherapy, but few options exist for individuals with resistant disease. Donor lymphocyte infusions (DLI) from the patient's original donor have provided responses in PTLD following allogeneic marrow transplantation, but HLA matched allogeneic donors are generally not available for solid organ transplantation recipients. A renal-pancreas transplant recipient with refractory PTLD was treated with an infusion of haploidentical T cells genetically engineered to allow eradication of these cells by the administration of ganciclovir should severe GVHD or organ rejection be observed. A retrovirus capable of expressing the herpes simplex virus thymidine kinase (HSV-tk) gene to provide negative selection and the human nerve growth factor receptor (NGFR) for positive selection was used to transduce donor T cells (1.2 x 109) 48 hours post activation with anti-CD3/CD28 beads. The transduction frequency was 37.2% based on NGFR expression; following selection with a biotinylated anti-NGFR antibody and anti-biotin magnetic beads using the Miltenyi CliniMACS a 98.1% NGFR+ product was obtained. Engineered T cells (106/kg) were infused following a lympho-depletion regimen consisting of fludarabine 125 mg/m2 and cyclophosphamide 60 mg/kg. Platelet (>50,000) and neutrophil (>500) recovery occurred 10 and 12 days post cell infusion, respectively. A bone marrow evaluation on day +21 revealed 8.2% of nucleated cells in the marrow to be of donor origin by VNTR analysis. On day +30 peripheral blood VNTR testing documented 28.6% of mononucleated cells to be donor in origin. CT imaging 4 weeks after infusion of the engineered T cells documented decreased size of the primary tumor in the breast. After the initial neutrophil and platelet recovery, neutropenia and thrombocytopenia recurred. A bone marrow evaluation on day 37 revealed decreasing cellularity, possibly consistent with GVHD associated aplasia; VNTR analysis of this marrow sample revealed 20.9% of cells to be of donor origin. The decision was made to treat the patient with ganciclovir (GCV) to eliminate donor T cells contributing to aplasia. Unfortunately, the patient expired within 48 hours of beginning GCV due to an infectious complication. On autopsy, no evidence of previously biopsy proven EBV PTLD was found in the lung and the buccal surface of the mouth. In addition, much of the primary breast mass was free of PTLD. Lymphocytic infiltration of hepatic portal tracts was observed, consistent with GVHD. No evidence of rejection was noted in the transplanted kidney or pancreas. In summary, an excellent response of a refractory EBV PTLD was obtained using therapy including haploidentical T cells engineered for negative selection, and extended persistence of the cells was documented was for greater than 1 month. In the future, a secondary drop in blood counts will be used to start CGV for negative selection of these cells in vivo. This novel strategy may provide effective therapy to transplant recipients without donors who invariable die of their underlying secondary malignancy.