1. ¹Center for Cell and Gene Therapy, Baylor College of Medicine, The Methodist Hospital, Texas Children's Hospital, Houston, Texas, USA
2. ²Dan L. Duncan Cancer Center, Baylor College of Medicine, Houston, Texas, USA
3. ³Clinic for Pediatric Oncology, Heinrich Heine University of Duesseldorf, Duesseldorf, Germany

Correspondence: Ann M Leen, Center for Cell and Gene Therapy, Baylor College of Medicine, 6621 Fannin Street MC 3-3320, Houston, Texas 77030, USA. E-mail: aleen@bcm.tmc.edu

Received 18 March 2009; Accepted 2 June 2009; Published online 7 July 2009.

Abstract

Viral infections cause morbidity and mortality in allogeneic hematopoietic stem cell transplant (HSCT) recipients. To prevent and treat these, we have produced and infused cytotoxic T lymphocytes (CTLs) with specificity for Epstein–Barr virus (EBV), cytomegalovirus (CMV), and adenovirus (Adv), and shown that small numbers of infused cells proliferate in vivo and protect against all three viruses. Despite these encouraging results, broader implementation of this approach is limited by the need for infectious virus material (EBV), expensive production of clinical grade adenoviral vectors, and a prolonged (8–12 weeks) period of manufacture. There is also competition between virus-derived antigens within antigen-presenting cells (APCs), limiting extension to additional agents. We now describe an approach that uses DNA nucleofection of dendritic cells (DCs) with DNA plasmids that encode a range of immunodominant and subdominant viral antigens from CMV, EBV, BK, and Adv. Within 10 days, this methodology provides multivirus-reactive CTLs that lack alloreactivity. We further demonstrate that nucleofected DC stimulation can be combined with interferon- (IFN-) capture technology to produce even more rapid multivirus-CTL products for treatment of acute infection. These CTL generation procedures should increase the feasibility and applicability of T-cell therapy.