Reduced Graft-Versus-Host Disease-Inducing Capacity of T Cells After Activation, Culturing, and Magnetic Cell Sorting Selection in an Allogeneic Bone Marrow Transplantation Model in Rats

Abstract

Graft-versus-host disease (GvHD), a major complication of allogeneic bone marrow transplantation, has been ascribed to mature T cells in the graft. Because T cells play an important role in engraftment of the bone marrow and decrease the probability of relapse of leukemia, a treatment strategy was developed to preserve the benefits of T cells in the graft and to control the severe complications of GvHD. This can be accomplished by the genetic modification of donor T cells with a suicide gene that allows their selective in vivo elimination and subsequently the abrogation of GvHD. For clinical benefit the alloreactivity of herpes simplex virus thymidine kinase (HSV-TK) gene-transduced T cells should be retained. Therefore, we investigated the influence of gene transduction and the selection procedure on T cells. We demonstrated that activation and culturing of T cells reduce their capacity to induce lethal GvHD in an allogeneic rat bone marrow transplantation model. Furthermore, positive immunomagnetic selection of gene-transduced T cells resulted in loss of the GvHD-inducing capacity of HSV-TK(+) T cells directly after MACS (magnetic cell sorting) selection; this loss could be recovered by a 1-day expansion of the selected T cells. No effect on alloreactivity was observed to be caused by the gene transduction procedure. Our study resulted in the development of an optimized culture and gene transduction protocol with preservation of T cell alloreactivity. Treatment of transplanted rats with ganciclovir resulted in a rapid reduction in the number of HSV-TK(+) T cells in the peripheral blood and in increased survival of the animals.

Factors affecting human T cell engraftment, trafficking, and associated xenogeneic graft-vs-host disease in NOD/SCID β2mnull mice

“…Data from both mouse and human allogeneic studies have shown that donor T cells exhibit decreased allo-reactivity and GVHD potential following ex vivo activation and expansion [2][3][4][5][6]. This loss of allo-reactivity is extremely undesirable because both the conversion to full donor chimerism and the anti-leukemia effect after allogeneic HSCT are mediated, at least in part, by alloreactive donor T cells.…”

“…Many of the therapeutic strategies that are being developed to control GVHD while maintaining the beneficial graft-versus-leukemia and/or graft-versus-infection effects provided by donor lymphocyte infusion after allogeneic HSCT require ex vivo T cell stimulation and expansion. Unfortunately, multiple preclinical and clinical studies have demonstrated that these ex vivo expanded T cells exhibit decreased survival and function in vivo, including reduced alloreactivity and GVHD potential [2][3][4][5][6]. Recently, others and we described an ex vivo method for expansion of human T (huT) cells, using anti-CD3 and anti-CD28 monoclonal antibodies that are covalently attached to superparamagnetic microbeads (CD3/CD28 beads) [7,8].…”

Bone-Marrow-Derived Cells Contribute to Glomerular Endothelial Repair in Experimental Glomerulonephritis

“…18 In short, donor BM cells were obtained by flushing the cavity of WR rat tibias and femurs with RPMI 1640 (Invitrogen, Carlsbad, CA). A single-cell suspension was obtained by filtering the BM through a cell strainer (100-m filter, Falcon; Becton Dickinson Labware, Franklin Lakes, NJ).…”

Functional and phenotypic variations in human T cells subjected to retroviral-mediated gene transfer

“…In addition, we observed that the IL-2 proliferation response was significantly higher in samples that had been subjected to transduction under standard stimulatory conditions (1 mg/ml anti-CD3i/ anti-CD28i) compared to those activated with 1 mg/ml anti-CD3i (Figure 1a). Since previous studies showed that cell selection could alter the functionality -including the GVHD reactivity -of transduced cells, 17,19,28 we have investigated whether or not the immunomagnetic cell sorting of cells expressing the selectable tNGFR transgene modifies the proliferative response of T cells to IL-2. The experiments in Figure 1b show that no differences in the proliferative response of T cells to IL-2 were observed between transduced unselected cells with respect to the NGFR þ (96% NGFR þ ) and NGFR À (o10% NGFR þ ) fractions collected after immunoselection.…”