ALBERT

Description: Opportunistic viral infections and relapse are major complications in patients after T cell depleted allogeneic stem cell transplantation (TCD alloSCT). Since the application of unmodified donor lymphocyte infusion (DLI) early after alloSCT results in a high risk of graft versus host disease (GVHD), infusion of selected populations of virus-specific donor T cells can be an effective approach to safely restore anti-viral immunity early after alloSCT. As part of the EU FP7 consortium T Control, in this phase I/II study the feasibility and safety of the generation and administration of selected populations of donor-derived T cells targeting multiple antigens (Ag) is assessed. The multi Ag-specific T cell products contained T cells targeting cytomegalovirus (CMV), Epstein Bar virus (EBV) and adenovirus (AdV) as well as T cells targeting tumor associated Ag (TAA) and minor histocompatibility Ag (MiHA) to boost the graft versus leukemia (GVL) reactivity. To assess efficacy, in-vivo appearance or expansion of Ag-specific T cells, and the effect on viral reactivations and/or disease relapse was evaluated for 20 weeks after infusion until regular DLI was applied. HLA-A*02+ patients treated for a hematological malignancy with an HLA-matched TCD alloSCT from a CMV and/or EBV seropositive donor were included in this study. 6-8 weeks after alloSCT, T-cells directed against HLA-A*02-restricted peptides of CMV, EBV and AdV, and the TAA NY-eso, WT-1, RHAMM, PRAME and proteinase 3 were isolated using the reversible streptamer-nanobead technology (Juno) by cliniMACS selection out of the naïve and/or memory T cell compartment from 2*10^9 donor PBMC. Depending on the HLA-typing of the patient/donor additional streptamers targeting viral peptides in A*01, A*24, B*07 or B*08 were added to the selection procedure. In case of patient/donor MiHA disparity in the GvL direction, the HLA-A*0201/HA-1h streptamer was also added. This procedure allows purification of T cells under GMP conditions in 1 day. 20 multi Ag-specific T cell products were generated of which 19 met the release criteria. These products consisted of 0.5-12*10^6 cells containing purities of 46,0-94,3% target Ag-specific CD8+ T cells within the T cell compartment. In all products CMV and/or EBVas well as AdV virus-specific memory T cells were isolated comprising 99% of the target Ag-specific CD8+ T cells, while the other 1% included the TAA and MiHA specificities or CMV specific T cells from seronegative donors. 17 products were administered without infusion-related complications or GVHD; 2 patients experienced GVHD before infusion and did not receive their product. Of the 14 patients evaluated at this stage, 13 completed the follow-up period until DLI and 1 patient died during follow-up. No product-related adverse events were reported. In all 5 CMV- patients and in 3/9 CMV+ patients no CMV reactivation and no expansion of CMV-specific T cells was observed. The other 6 CMV+ patients experienced CMV reactivations. In 2/6 patients who received the product from a CMV+ donor CMV-specific T cells were detected with tetramer analysis and CMV was cleared. From the other 4 reactivating patients with CMV- donors 3 had circulating CMV-specific T cells already at the moment of infusion, whereas in 1 patient CMV-specific T cells clearly expanded after infusion, resulting in viral clearance in all 4 patients. All 14 donors were EBV+. In 7 patients EBV reactivations were observed, which coincided in 2/7 patients with the appearance of EBV-specific T cells and subsequent clearance of the virus. In 4/7 patients EBV reactivation was cleared without clear expansion of EBV-specific T cells. However, 1 patient required treatment for an EBV-PTLD, although ultimately EBV-specific T cells appeared and EBV was cleared. In none of the patients AdV DNA loads were detected in the follow-up period, while in 1 patient expansion of AdV-specific T cells was observed. Expansion of TAA and MiHA-specific T cells could not be demonstrated in-vivo using tetramer staining. More sensitive techniques will be required to visualize these cells. 2 patients showed relapse of their malignancy before DLI infusion. In this clinical study, we have shown that the reversible streptamer technology based generation and adoptive transfer of donor-derived multi Ag-specific T cell products is feasible and safe and can be used as a strategy to prevent viral infections in the interval between TCD alloSCT and DLI. Disclosures Germeroth: Juno Therapeutics: Employment.

Description: Viral infections and disease relapses are major complications between T cell depleted allogeneic stem cell transplantation (TCD alloSCT) and donor lymphocyte infusion (DLI). The prophylactic infusion of selected donor T cells may be an effective method to restore anti-viral and anti-tumor immunity early after TCD alloSCT. In this phase I/II study, we aimed to prevent these complications by the infusion of donor-derived T cell products containing CD8+ T cells directed against cytomegalovirus (CMV), Epstein-Barr virus (EBV) and adenovirus (AdV) antigens (Ag), tumor associated Ag (TAA) and minor histocompatibility Ag (MiHA) 6-8 weeks after TCD alloSCT. The feasibility of multi-Ag specific T cell product generation and safety of early administration were investigated. Furthermore, clinical outcome and immune reconstitution were assessed. HLA-A2+ patients treated for a hematological malignancy with a ≥9/10 HLA-matched TCD graft from a CMV+ and/or EBV+ (un)related donor were eligible for inclusion. Donor T cells directed against HLA-A2-restricted peptides of CMV, EBV, AdV, and TAA NY-ESO-1, WT-1, RHAMM, PRAME and proteinase 3 were simultaneously isolated under GMP conditions in 1 day, using MHC I-Streptamers and CliniMACS selection out of the naïve and memory T cell compartment from 2*109 PBMC. Depending on patient/donor HLA-typing, additional MHC I-Streptamers targeting viral peptides in HLA-A1, -A24, -B7, or -B8 were added to the procedure, as well as the HLA-A2/HA-1h MHC I-Streptamer in case of MiHA disparity in the GVL direction. The incidence of GVHD, mortality, disease relapses and viral reactivations was monitored until 6 months after alloSCT. In addition, follow-up samples were stained with tetramers to analyze in vivo reconstitution of target-Ag specific T cells. Products were generated for 27/28 included patients that showed stable engraftment after TCD alloSCT; 1 patient died early after alloSCT. All donors were EBV+, 14/27 donors were CMV+. 26/27 products were successfully generated and contained a median of 5.2*106 cells (range 0.4 - 26.5*106) with a median purity of target-Ag specific T cells within the CD3+ compartment of 80.4% (range 46.0-99.9). Target-Ag specific T cells consisted for 99% of virus-specific memory T cells, while the remaining 1% included TAA, MiHA and naïve virus-specific T cells. 24 patients received their product (median 58 days after alloSCT; range 51-107) without infusion related complications; in 2 patients the infusion was cancelled due to acute skin GVHD on the day of infusion. During follow-up, 1 patient experienced skin GVHD requiring immunosuppressive therapy. One patient died due to complications of a nephrotic syndrome, probably unrelated to product infusion. Four patients showed disease relapse and were treated accordingly; coinciding expansion of TAA- or MiHA-specific T cells was not observed. None of the patients experienced AdV reactivations; in 2 patients AdV-specific T cells expanded after infusion. Four patients had detectable EBV loads; in one of these patients the EBV reactivation progressed to EBV-PTLD and he was treated with rituximab followed by DLI. In 2 patients, the rise in EBV load led to expansion of EBV specific T cells in vivo. 4/5 CMV+ patients with a CMV- donor had ongoing CMV reactivations at the moment of product infusion, CMV specific T cells could already be detected just before product infusion and reactivations were cleared within 7-84 days. Of the 8 CMV+ patients with a CMV+ donor, 3 patients remained free of CMV reactivations, 4 had already a reactivation at the moment of product infusion and 1 patient developed a CMV reactivation during follow up. One patient had progressed to CMV pneumonia within 2 weeks after infusion, requiring ganciclovir. In all 5 CMV reactivating patients, T cells directed against 2-3 CMV-specific epitopes expanded significantly and viral loads declined. TCR sequencing of the CDR3 region illustrated in vivo persistence and expansion of target-Ag specific T cells present in the product. In conclusion, we have shown that MHC I-Streptamer isolation based generation and adoptive transfer of donor-derived multi Ag-specific T cell products is safe and feasible. Moreover, efficacy of the prophylactic infusion is illustrated by expansion of target-Ag specific T cells in patients coinciding viral reactivations and the prevention of viral complications in the majority of patients between TCD alloSCT and DLI. Disclosures Germeroth: Juno Therapeutics: Employment, Equity Ownership.