T Cell Immunotherapy for Adenoviral Infections of Stem Cell Transplant Recipients.
Authors: Leen Ann M. , Myers Gary D. , Weiss Heidi , Gee Adrian P. , Leung Kathryn S. , Krance Robert A. , Brenner Malcolm K. , Rooney Cliona M. , Heslop Helen E. , Bollard Catherine M. 
Published: April 2020

Abstract
Adenoviruses remain a major cause of mortality and morbidity in hematopoietic stem cell transplant (HSCT) recipients. To date, no effective anti-adenoviral treatment is available, presenting a potential role for adoptive T cell therapy, as an increased risk of infection may be correlated with lack of endogenous T cell immunity. Therefore the goal of this project was to evaluate the use of adenovirus-specific cytotoxic T lymphocyte (CTL) lines for the prophylaxis and treatment of adenoviral infections post-HSCT. CTLs are initiated by stimulation with donor-derived mononuclear cells transduced with a recombinant adenovirus type 5 vector pseudotyped with a type 35 fiber (MOI 200 virus particles (vp) per cell). Responder cells are expanded by weekly restimulation with autologous EBV-transformed B lymphoblastoid cell lines (LCL) transduced with the same vector using an MOI of 500 vp. Nine of the 11 CTL lines manufactured for clinical use were predominantly CD4+ (range 12–95%), but all had a CD8+ component (range 3.5–79%). In chromium release assays, all CTL lines showed specific killing of both adenovirus and EBV antigen-bearing target cells. Recognition of adenovirus and EBV targets was not significantly different; at 20:1 effector:target ratio there was 18.5 ± 14% specific lysis of adenovirus targets and 23 ± 15% lysis of EBV targets while uninfected recipient target cells were not lysed (1.5 ± 2%). We identified the hexon as the immunodominant target antigen in the CTL lines. To test the hypothesis that these polyclonal CD4+ and CD8+ hexon-specific T cells can provide protection against adenovirus infections in HSCT recipients, we administered donor-derived bi-virus-specific CTL lines to pediatric recipients of allogeneic stem cell transplant. So far, we have treated 7 patients in this phase I dose escalation study (using the continual reassessment model); 2 patients on dose level 1 (5x106/m2), 2 on dose level 2 (1.5x107/m2) and 2 on dose level 3 (4.5x107/m2). Of 6 patients who received CTL as prophylaxis, all remain negative for adenovirus and EBV and there were no adverse events attributable to CTL infusion. Post-infusion, samples for immune monitoring were collected for at least 3 months and analyzed by pentamer and IFN-g ELISPOT assays in batches to minimize inter-assay variablility. An increase in the frequency of EBV-specific T cells could be detected in the blood by 2 weeks post infusion in all evaluated patients, likely due to the presence of this latent virus in all patients. However, there was no significant rise in the adenovirus-specific T cell frequency post-infusion. The seventh patient received the bi-virus-specific CTL at a dose of 5x106/m2 as treatment for adenovirus disease. This patient had adenoviral pneumonia requiring ventilatory support. Following the CTL infusion the patient made a dramatic clinical recovery, was weaned off mechanical ventilation, and discharged a month after the infusion. In contrast to the prophylaxis group, this patient had a substantial rise in adenovirus-specific T cells detected in the peripheral blood by 4 weeks post-CTL infusion, which coincided with his clinical recovery and clearance of the virus from the tracheal aspirate, suggesting that the infused cells were effective in vivo. Adoptive immunotherapy for adenovirus appears to be safe but expansion of adenovirus-specific CTL in vivo may require the presence of antigen to stimulate the infused cells.