Herpes Simplex Virus Thymidine Kinase Gene Transfer for Controlled Graft-versus-Host Disease and Graft-versus-Leukemia: Clinical Follow-up and Improved New Vectors

Abstract

We previously demonstrated that severe graft-versus-host disease (GvHD), associated with the therapeutic infusion of donor lymphocytes after allogeneic marrow transplantation (BMT), can be efficiently controlled by the SFCMM-2-mediated expression of the herpes simplex virus thymidine kinase (HSV-tk) suicide gene into the allogeneic lymphocytes. This was achieved by selective elimination of transduced lymphocytes by ganciclovir (GCV) infusion. Despite the positive results of the pilot clinical trial, two vector-related limitations were observed. The induction of a strong immune response against genetically modified cells was observed in two patients. In addition, the only patient who developed chronic GvHD showed only partial response to ganciclovir treatment. In an attempt to overcome these limitations, we developed a new generation of vectors. The neomycin resistance (neoR) gene was removed from the SFCMM-3 and SFCMM-4 retroviral vectors. These vectors are less immunogenic and able to confer higher ganciclovir sensitivity to transduced human lymphocytes. All the vectors carry a modified form of the low-affinity nerve growth factor receptor cDNA, as cell surface selectable marker (deltaLNGFR). The vectors were compared in terms of gene transfer efficiency, and ability to confer high and specific sensitivity to ganciclovir. The SFCMM-3 vector, carrying the entire HSV-tk gene driven by the LTR promoter, allows efficient transduction of human T lymphocytes and confers the highest GCV sensitivity to transduced lymphocytes with a high and a low proliferation index. The expression of the deltaLNGFR marker allows an easier in vitro manipulation and a faster immune selection of transduced cells compared with neoR selection. Finally, the elimination of the neoR gene removes a potent immunogen from transduced lymphocytes.

Functional impairment of human T-lymphocytes following PHA-induced expansion and retroviral transduction: implications for gene therapy

“…In our hands, 0.5 g/ml PHA or 0.5 g/ml iCD3 plus 1 g/ml iCD28 led to levels of transduction comparable to what many other published protocols achieve. 3,7,14,15,27,30,31 Most likely, the incorporation in the protocol of elements such as retronectin, three rounds of infection, and a retroviral envelope with enhanced host range and the capacity to be concentrated, were critical to the results obtained. However, further analysis of these aspects is not possible from the design and results of this study.…”

Development of gene therapy in association with clinically used cytotoxic deoxynucleoside analogues

“…[27][28][29] Two T-lymphoblastoid cell lines were transduced with Dm-dNK and showed an increase in sensitivity toward pyrimidine nucleoside analogues such as BVDU, but not to the antiherpetic nucleoside analogues GCV, acyclovir (ACV) and penciclovir (PCV). 30 Immuno-suppressed patients often suffer from herpes virus infections, and if GCV or ACV is used to treat these infections, the compounds will also be activated in the suicide genecarrying T cells if HSV-TK is used as the suicide gene.…”

T cell transduction and suicide with an enhanced mutant thymidine kinase

“…One case of chronic GVHD was only partially responsive to ganciclovir, and vector-related limitations included evidence of an immune response against the transgenes in two patients. 10 The group has now treated over 40 patients using this strategy, but results have yet to be fully evaluated. 11 Tiberghien et al 8 used HSV-TK transduced T cell infusions to prevent leukemic relapse in 14 patients undergoing BMT for haematological malignancy.…”