Retrovirus mediated gene transduction of human T-cell subsets

Allogeneic bone marrow transplantation (AlloBMT) can be curative for patients with leukaemia. Graft versus host disease (GVHD) is a potentially life threatening complication of AlloBMT mediated by the T cells contained within the graft. In order to be able to control GVHD, the allogeneic T cells may be transduced with a suicide gene such as herpes simplex virus thymidine kinase (HSV-tk). For this strategy to be successful, all subsets of T cells should be transduced to a similar extent. Also, the transduction protocol should not induce expression of unwanted homing receptors, nor should it lead to unwanted skewing of the T-cell receptor repertoire. We have studied the transduction efficiency of naïve and memory subsets of CD4+ and CD8+ T cells, and examined the transduced T-cell subsets for possible changes in T-cell receptor (TCR) repertoire and homing receptor expression.

The cells were transduced using a Moloney murine retroviral vector carrying a conjugate of the genes encoding the truncated form of the cell surface marker, low affinity nerve growth factor receptor (DeltaLNGFR) and HSV-tk. Transduction efficiency and homing receptor expression were quantified by flow cytometry. TCR repertoire was determined by spectratyping.

We obtained a transduction efficiency of 30-50% of the cells, with no difference between the T-cell subsets. Cell surface receptors responsible for homing to skin, gastrointestinal tract or lymph nodes were practically absent at the end of 2 weeks in culture. The activation procedure seemed to favour the expansion of certain T-cell clones over polyclonal populations. However, there was no difference in the TCR repertoire between transduced and non-transduced cells.

Changes in the composition of the T-cell subsets at the end of the cell culture were the results of the activation, and not the suicide gene transduction. The transduced T cells did not express unwanted homing receptors.