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TALEN-mediated genetic inactivation of the glucocorticoid receptor in cytomegalovirus-specific T cells

Laurie Menger, Agnes Gouble, Maria A.V. Marzolini, Annette Pachnio, Katharina Bergerhoff, Jake Y. Henry, Julianne Smith, Martin Pule, Paul Moss, Stanley R. Riddell, Sergio A. Quezada and Karl S. Peggs

Key points

  • The GR gene can be inactivated in Streptamer®-selected CMV-specific CD8+ T cells using TALEN.

  • The GR gene inactivation endows T cells with resistance to the immunosuppressive effects of corticosteroids in vitro and in vivo.

Abstract

Cytomegalovirus (CMV) infection is responsible for substantial morbidity and mortality following allogeneic hematopoietic stem cell transplantation (HSCT). T cell immunity is critical for control of CMV infection and correction of the immune deficiency induced by transplantation is now clinically achievable by the adoptive transfer of donor-derived CMV-specific T cells. It is notable, however, that most clinical studies of adoptive T cell therapy exclude patients with graft-versus-host disease (GvHD) receiving systemic corticosteroid therapy, which impairs cellular immunity. This group of patients remains the highest clinical risk group for recurrent and problematic infections. Here, we address this unmet clinical need by genetic disruption of the glucocorticoid receptor (GR) gene using electroporation of transcription activator-like effector nucleases (TALEN) mRNA. We demonstrate efficient inactivation of the GR gene without off-target activity in Streptamer®-selected CMV-specific CD8+ T cells (A2-NLV), conferring resistance to glucocorticoids. TALEN-modified CMV-specific T cells retained specific killing of target cells pulsed with the CMV peptide NLV in the presence of dexamethasone (DEX). Inactivation of the GR gene also conferred resistance to DEX in a xenogeneic-GvHD model in sub-lethally irradiated NOD-scid IL2rγnull (NSG) mice. This proof of concept provides the rationale for the development of clinical protocols for producing and administering high-purity genetically-engineered virus-specific T cells that are resistant to the suppressive effects of corticosteroids.

  • Submitted August 19, 2015.
  • Accepted October 16, 2015.