Activity of broad-spectrum T cells as treatment for AdV, EBV, CMV, BKV, and HHV6 infections after HSCT

Abstract

It remains difficult to treat the multiplicity of distinct viral infections that afflict immunocompromised patients. Adoptive transfer of virus-specific T cells (VSTs) can be safe and effective, but such cells have been complex to prepare and limited in antiviral range. We now demonstrate the feasibility and clinical utility of rapidly generated single-culture VSTs that recognize 12 immunogenic antigens from five viruses (Epstein-Barr virus, adenovirus, cytomegalovirus, BK virus, and human herpesvirus 6) that frequently cause disease in immunocompromised patients. When administered to 11 recipients of allogeneic transplants, 8 of whom had up to four active infections with the targeted viruses, these VSTs proved safe in all subjects and produced an overall 94% virological and clinical response rate that was sustained long-term.

Trial registration: ClinicalTrials.gov NCT01570283.

Conflict of interest statement

Competing interests: The authors declare that they have no competing interests relevant to the subject matter of this article. JFV is a consultant for Wilson Wolf Manufacturing. CMR, MKB and HEH have a licensing agreement with Cell Medica for EBV-specific T cells to treat EBV-associated lymphoma or nasopharyngeal carcinoma. MKB is on the scientific advisory board of CAnVAC and has share options in both CanVAc and bluebird bio. The Center for Cell and Gene Therapy has a research collaboration with Celgene. None of these relationships is relevant to the subject matter presented in this manuscript. The trial is registered at www.clinicaltrials.gov as NCT01570283.

Copyright © 2014, American Association for the Advancement of Science.

Figures

Figure 1. Cell expansion and immunophenotype of mVSTs generated for clinical use

Panel A shows the T cell expansion of mVSTs achieved over a 9–11 day period based on cell counting using trypan blue exclusion. Panel B shows the phenotype of the mVST cell lines on the day of cryopreservation. Each symbol represents an individual T cell line (n=48), and the black bar (–) represents the mean.

Figure 2. Virus-specific activity of mVST lines

Panel A shows the spectrum of specificity of each mVST cell line vs the age of the donor from whom the T cell line was generated. Each circle represents a T cell line (n=48). The mean is represented as a black line. Panel B shows the estimated probability of activity against one or more viruses compared to the donor’s age, analyzed by the binomial regression model. Each circle represents one donor. Panel C shows the frequency of AdV (Hexon and Penton), CMV (IE1 and pp65), EBV (LMP2, EBNA1, BZLF1), BKV (Large T and VP1), and HHV6 (U11, U14 and U90) reactive T cells in mVST cell lines as measured by IFNγ ELIspot. Only T cell lines with a total of ≥30 SFC/2×105 for a given virus were considered to be positive. Each symbol represents the mean ± SEM SFC/2×105 input cells of an individual T cell line. Control was IFNγ release in response to stimulation with no pepmix.

Figure 3. In vivo expansion and clinical benefits of mVSTs in subjects infected with one virus

Panel A shows the AdV viral load in blood (copies/ml) and frequency of AdV-specific T cells before and after infusion in subject P4052 who was treated for an AdV infection. Panel B shows the BKV viral load in blood (copies/ml) and frequency of BKV-directed T cells before and after infusion in subject P3850 who was treated for a BKV reactivation. In both cases dotted lines represent the viral loads and solid lines the frequency of VSTs as measured by IFNγ ELIspot and results are presented as average SFC/5×105 PBMCs. The arrow indicates the time of mVST infusion.

Figure 4. In vivo expansion and clinical benefits of mVSTs in subjects with two viral infections/reactivations

Panels A and B show the CMV and BKV viral load in blood (copies/ml) and frequency of CMV− and BKV-reactive T cells before and after infusion in subject P3908 who was treated for a CMV infection and had a subsequent BKV reactivation. Panels C and D show the BKV load (copies/ml) in blood and urine respectively, and frequency of BKV-specific T cells before and after infusion in subject P3925 who was treated for BKV infection. Panel E shows the frequency of BKV-reactive T cells expanded from a bladder biopsy sample taken from patient P3925, versus unstimulated T cells, which served as a negative control. Panel F shows the EBV viral load in blood (copies/μg DNA), and frequency of EBV-specific T cells before and after infusion in subject P3925 who reactivated EBV. Panels G and H show the BKV and EBV loads (copies/ml and copies/μg DNA, respectively) in blood and frequency of BKV-and EBV− directed T cells before and after infusion in subject P4165 who had both BKV and EBV infections. In all cases dotted lines represent the viral loads and solid lines the frequency of VSTs measured by IFNγ ELIspot and results are presented as average SFC/5×105 PBMCs. The arrow indicates when the mVSTs were administered.

Figure 5. In vivo expansion and clinical benefits of mVSTs in subjects with three viral infections/reactivations

Panels A, B and C show the CMV, EBV and BKV viral loads detected in the peripheral blood (copies/ml, copies/μg DNA and copies/ml, respectively) and frequency of CMV−, EBV− and BKV− reactive T cells before and after infusion in subject P3987 who was treated for CMV, EBV and BKV. Panels D, E and F show HHV6, EBV and BKV viral loads detected in the peripheral blood (copies/ml, copies/μg DNA and copies/ml, respectively) and frequency of HHV6−, EBV− and BKV-reactive T cells before and after infusion in subject P3940 who was treated for active HHV6 and BKV and subsequently reactivated EBV. In all cases dotted lines represent the viral loads and solid lines the frequency of VSTs measured by IFNγ ELIspot and results are presented as average SFC/5×105 PBMCs. The arrow indicates the infusion of mVSTs.

Figure 6. In vivo expansion and clinical benefits of mVSTs in one subject with four viral infections/reactivations

Panel A shows the EBV viral load (copies/μg DNA) in blood and the frequency of EBV-specific T cells in subject P3022 who was treated for EBV-PTLD. Panel B shows the PET scan of the subject P3022 before and after mVSTs. Panels C and D show the levels of BKV in blood and urine while Panels E and F show the HHV6 and CMV viral loads in the peripheral blood (copies/ml and copies/cell, respectively). The frequency of BKV−, HHV6− and CMV− reactive T cells in blood before and after infusion of mVSTs is also shown. In all cases dotted lines represent the viral loads and solid lines the frequency of VSTs measured by IFNγ ELIspot and results are presented as average SFC/5×105 PBMCs. The arrow indicates the infusion of mVSTs.