Safety and Clinical Efficacy Of Rapidly-Generated Virus-Specific T Cells With Activity Against Adv, EBV, CMV, HHV6 and BK Virus Administered After Allogeneic Hematopoietic Stem Cell Transplant
Authors: Papadopoulou Anastasia , Katari Usha L , Gerdemann Ulrike , Martinez Caridad , Leung Kathryn , Carrum George , Gee Adrian P. , Vera Juan F , Krance Robert A. , Brenner Malcolm K. , Rooney Cliona M , Heslop Helen E. , Leen Ann 
Published: October 2019

Abstract

Viral infections remain a major cause of morbidity and mortality after allogeneic HSCT. We and others have demonstrated that the adoptive transfer of virus-specific T cells (VSTs) specific for EBV, CMV and Adv antigens can treat infections that are impervious to conventional therapies, but broader implementation and extension to additional problematic viruses has been limited by competition between viral antigens and time-consuming/laborious manufacturing. We therefore developed a simplified 10-day system for generating a single preparation of VSTs with activity against 12 antigens from 5 viruses (EBV, CMV, Adv, BK, HHV6) that commonly cause post-transplant morbidity and mortality. We report our initial clinical results using these pentavalent pVSTs.
With NHLBI-PACT support, we prepared 35 clinical-grade pVSTs from PBMCs (3x107 cells/sample) that we exposed to overlapping peptide libraries spanning immunogenic Adv (Hexon, Penton), CMV (pp65, IE1), EBV (LMP2, EBNA1, BZLF1), BK (Large T, VP1) and HHV-6 (U11, U14, U90) antigens. Exposure was followed by a 9-11 day expansion phase in a G-Rex device in the presence of IL4+7, producing a mean of 374x106 T cells (range 99-713x106). These lines were polyclonal, comprising both CD4+ (57±5%) and CD8+ (35±5%) cells and retained expression of the memory markers CD45RO+CD62L+ (58±8%). Their specificity was dependent on the prior viral exposure of the cell donor; 32/35 lines had activity against Adv (Hexon: 446±153; Penton:317±108 SFC/2x105), 20/35 against CMV (IE1: 337±141; pp65 1059±479), 26/35 against EBV (LMP2: 175±87; EBNA1: 116±44; BZLF1: 129±88), 18/35 against BK (Large T: 130±67; VP1: 231±104) and 21/35 against HHV-6 (U90: 66±50; U11: 36±18; U14: 82±21). None of the lines reacted against recipient PHA blasts (mean Cr51release of 1% at a 20:1 E:T ratio).
We have administered pVSTs to 10 allogeneic HSCT recipients in a dose escalation study; 4 on DL1 (5x106/m2), 4 on DL2 (1x107/m2) and 2 on DL3 (2x107/m2). There were no immediate infusional toxicities, and no de novo acute GvHD, demonstrating the in vivo safety of these pVSTs even after a single exposure to viral antigens in vitro. Three patients received the cells as viral prophylaxis (days 38-43 post-HSCT) and all remain well and virus-infection free at up to 3 months post-treatment. The other 7 patients received the cells as treatment for one or more active infections between days 59-139 post-HSCT. Based on viral load measurements by day 42 post-infusion, the pVSTs were successful in controlling active CMV (1 complete (CR) and 1 partial response (PR)), EBV (2 CRs, including a case of frank PTLD); Adv (1 CR); HHV6 (1 CR); and BK (3 CR, 1 PR, 1NR) infections. Of note, 3 of our BK virus responders had tissue disease with severe hemorrhagic cystitis and all had marked improvement or disappearance of hematuria following infusion. One subsequently had an episode of transient but severe bladder pain in association with inflammation seen on cytoscopy coincident with a 6 log fall in urine BK viral load. Our only non-responder was a patient with BK infection whose line lacked activity for this virus, likely reflecting the serostatus of the donor. In addition, 3 patients subsequently reactivated other viruses than those for which they were initially treated, but all cleared these infections by week 12, without requiring additional cell infusions (CMV: 1CR; EBV: 1CR; BK: 1CR; HHV6: 1CR). Finally, 1 patient received pVSTs under a single patient protocol as an emergency treatment for widespread and bulky rituximab-resistant EBV-PTLD. Post pVST there was an immediate decline in her EBV viral load with complete and sustained resolution of PTLD, coincident with an increase in circulating EBV-specific T cells. However, the profound anti-tumor activity mediated by the rapidly-expanding EBV-directed T cells also produced a transient systemic inflammatory response syndrome, which was controlled with steroids and anti-TNFR antibody, with no long term adverse effects.
Thus, infusion of pVSTs as prophylaxis or treatment has been safe and is associated with the appearance of virus-reactive T cells in peripheral blood that have been able to control infection with all 5 targeted viruses. We are currently exploring the extension of this platform to include additional clinically relevant viruses and are planning to assess the activity of these cells in the 3rd party setting for broader implementation.

Disclosures:
Off Label Use: Virus-specific CTLs manufactured under an investigator-initiated IND. Vera:Wilson Wolf Corporation: Consultancy, Research Funding.