HSV-sr39TK Positron Emission Tomography and Suicide Gene Elimination of Human Hematopoietic Stem Cells and Their Progeny in Humanized Mice

Abstract

Engineering immunity against cancer by the adoptive transfer of hematopoietic stem cells (HSC) modified to express antigen-specific T-cell-receptors (TCR) or chimeric antigen receptors (CAR) generates a continual supply of effector T-cells, potentially providing superior anti-cancer efficacy compared with the infusion of terminally differentiated T-cells. Here we demonstrate the in vivo generation of functional effector T-cells from CD34-enriched human peripheral blood stem cells (PBSC) modified with a lentiviral vector designed for clinical use encoding a TCR recognizing the cancer/testes antigen NY-ESO-1, co-expressing the PET/suicide gene sr39TK. Ex vivo analysis of T-cells showed antigen- and HLA-restricted effector function against melanoma. Robust engraftment of gene-modified human cells was demonstrated with PET reporter imaging in hematopoietic niches such as femurs, humeri, vertebrae, and the thymus. Safety was demonstrated by the in vivo ablation of PET signal, NY-ESO-1-TCR bearing cells, and integrated lentiviral vector genomes upon treatment with ganciclovir (GCV), but not with vehicle control. Our study provides support for the efficacy and safety of gene-modified HSCs as a therapeutic modality for engineered cancer immunotherapy.

Immuno-PET of Murine T Cell Reconstitution Postadoptive Stem Cell Transplantation Using Anti-CD4 and Anti-CD8 Cys-Diabodies

“…Currently many stem or progenitor cell therapies involving T cell receptor (TCR) or chimeric antigen receptor (CAR) targeting epitopes expressed on malignant cells are under development for clinical translation (17–20). Previous work utilizing PET to detect hematopoietic stem cell transfer and immune cell engraftment employs reporter genes to image total cell engraftment as opposed to lineage specific repopulation (14, 21). Here we report the development of anti-CD4 and -CD8 cDbs radiolabeled with 89 Zr for direct immunoPET detection of CD4 + and CD8 + T cells with the goal of detecting helper and cytotoxic lymphocyte repopulation after HSC therapy.…”

Domain-swapped T cell receptors improve the safety of TCR gene therapy

“…Humanized mice were generated as previously described (Gschweng et al, 2014). Briefly, HLA-A*0201 + , CD34-enriched human peripheral blood stem cells were stimulated for 24 hr before transduction with spin concentrated virus encoding the indicated constructs at a vector concentration of 1 x 10 8 transduction units/mL.…”

Advances in PET Detection of the Antitumor T Cell Response

“…Animals that received a bone marrow transplant with cells engineered to express HSV-TK were challenged with a viral induced sarcoma and this allowed for the detection of a primary anti-tumor T cell response in vivo (Shu et al, 2005). The dual functionality and efficacy of sr39TK as a reporter and suicide gene was recently validated in a humanized mouse model (Gschweng et al, 2014; Vatakis et al, 2011). Human hematopoietic stem cells were transduced to express a TCR and sr39TK.…”

“…To ablate the engineered cells therapeutic doses of ganciclovir was given. Efficacy was determined by follow-up [ 18 F]-FHBG scans that displayed little to no signal in ablated animals (Gschweng et al, 2014). …”