Anti CMV and/or Anti Adenovirus IFN-g-Positive CD4+ CD8+ T Lymphocytes for Treatment of Viral Infections After Allogeneic HSC Transplantation: First Results
Authors: Dal Cortivo Liliane , Creidy RITA , Gabrion Aurélie , Leruez-Ville Marianne , Heritier Sebastien , Touzot Fabien , Cros Guilhem , Maury Sebastien , Xhaard Alienor , Ouachee Marie , Hamon Ingrid , Picard Capucine , Cambouris Christophe , Abina Salima Hacein Bey , Blanche Stephane , Cavazzana-Calvo Marina 
Published: October 2019

Abstract
Abstract 1906
Reactivation of latent viruses such as cytomegalovirus (CMV) and adenovirus (AdV) is responsible for infections which may be life-threatening in HSCT recipients. In the post-transplantation period, severity and frequency of these infections depend on (a) the degree of donor-recipient HLA incompatibility and (b) the intensity of immunosuppressive therapy used to prevent immunological complications. Antiviral drugs may be partially effective, often toxic and cannot always control those viral infections.T cell immunity plays a major role in the control of viral infections. It has been demonstrated that the transfer of donor T lymphocytes specifically directed against viral antigens is capable of preventing, controlling and clearing viral infection (Feuchtinger T et al., 2004 and 2010). The present project aimed the evaluation of specific, cell-based immunity against CMV and AdV by injection of IFN-g-positive CD4+and CD8+ donor T lymphocytes isolated ex vivo after stimulation with viral peptides.

Methods:
Our protocol was designed for pediatric or adult patients treated by allogeneic HSCT and matching the following inclusion criteria: (1) biological and/or clinical symptoms of CMV and/or AdV infection 2) no response or contraindication to conventional antiviral treatment and (3) no or low grade pre-existing aGvHD at inclusion (≤ grade II) controlled by corticoids (<1 mg/kg). Antiviral treatments are allowed during the inclusion period.
Donor IFN-g-positive T lymphocytes are isolated with the CliniMACS Cytokine Capture System (Miltenyi Biotech) after incubation with viral peptide pools.
Primary evaluation criterion is the efficacy of the treatment on CMV viral load 21 days after the first injection. In the event of a negative or partial response and the absence of aGvHD, a second injection may be scheduled.
Secondary evaluation criteria are (1) the occurrence of de novo aGvHD or aggravation of existing aGvHD, (2) the evolution of clinical symptoms potentially related to the infection, (3) the demonstration of biological in vivo expansion of injected T lymphocytes (as evidenced by the IFN-g secretion capacity and specific tetramer assays) and (4) for AdV infection, evaluation of efficacy (viral load, in vivo expansion of transfused lymphocytes, clinical symptoms) and the safety (occurrence of aGvHD) of this immunotherapy.

Results:
From September 2010 to July 2012, 9 patients were included: 3 male adults (46–54 years, 1 CLL, 1 CML and 1 AA, 2 geno- and 1 pheno-identical transplantation) and 6 children (age: 7–25 months, sex ratio F/M: 4/2, 4 FLH, 1 SCID and 1AA, 4 haplo, 1 geno- and 1 pheno-id transplantation).
4/9 patients were treated for CMV, 3/9 for AdV and 2/9 for CMV and AdV reactivation. 5/9 patients received 2 cytotoxic T lymphocytes (CTL) injections. Mean number of CD3 IFN-g positive cells injected was 4206/kg (1167–6000/kg) with 55% and 69% of CD4 and CD8 anti CMV-T cells and 56% and 61% of CD4 and CD8 anti AdV T cells respectively.
Mean delay of first immunotherapy was 109 days (28–270) after transplantation.
2/9 patients were not evaluable due to early death (<21 days post injection) and 1/9 patient died of graft failure 43 days after CTL injection without efficacy on infectious evolution. 6 patients are still alive: 4 with complete, 1 with partial remission of virus replication and 1 recently included, is still under evaluation. An in vivo expansion of transfused CTL was observed (mean expansion was 33 and 35 fold for CD8-IFN-g and CD4-IFN-g positive cells respectively 42 days after injection) in parallel with the decrease of viral load in all alive patients. No aGvHD was detected in the 5/6 evaluated patients. One of 6 presenting cGvH at inclusion need increase of corticotherapy 3 months after second injection of CTL One patient presenting with CMV retinitis received 2 CTL injections without worsening of retina lesions which healed.

Conclusion:
The CliniMACS Cytokine Capture System allows the isolation of virus-specific T cells in a brief delay (24 hours) with a satisfactory enrichment of both CD4 and CD8 T cells.
First results show efficacy of virus-specific T cells injection on viral load without signs of aGvHD in 5/6 evaluable patients. More patients need to be included in this trial in order to confirm these encouraging results.

Disclosures:
Cambouris: Miltenyi Biotec: Employment.