Supplementary material
infection, NLV
VIRUS, DISEASE, RECRUITMENT, INFECTION, PROLIFERATIVE
The Supplementary Material for this article can be found online at: Study design overview. Patients were randomized at visit 1 before allogeneic stem cell transplantation between day -28 and -7. Transplant procedure was carried out according to standards of the participating centers. A small fraction of the stem cell graft (3-5% of all cells) was shipped along with donor serum to the manufacturing site at the University Hospital of Erlangen. Manufacturing was started at the day of transplantation. If the T cell product could not be released or manufacturing failed, patients were able to cross over from the treatment group into the control group. Twenty-four to 48 h before each T cell product application patients were screened for acute GvHD including steroid treatment and signs of infection. If patients were not eligible for T cell product application this visit was repeated as additional visit 7 days later. The time frame for this assessment was 42-days opening a window for the first treatment from day 30 to day 72. Patients in the control group were monitored accordingly. End of study visit was 204 days after allogeneic stem cell transplantation.Click here for additional data file.Study recruitment overview. Schematic overview of the patient flow of all patients screened. In the treatment arm 3 patients crossed over to the control arm, as the product could not be released, or patient refused study medication. Four patients we not considered eligible for the study medication at visit 2 and were excluded from the study. In the control arm 2 patients were excluded due to relapse and the development of post transplant proliferative disease (PTLD).Click here for additional data file.Pat 11, Epitope specific T cell expansion during manufacturing. PBMC derived from the stem cell graft were analyzed before peptide stimulation (Day 0) and after 9 days of expansion (Day 9) by flow cytometry. Patient 11 shared 5 out the 6 HLA types for which peptides from CMV and EBV were available. Cells were gated on CD8+ T cells. Upper panel (red frame) shows specificities of CD8 for CMV epitopes, lower panel for EBV epitopes respectively (yellow frame).Click here for additional data file.Pat 11, T cell subsets and activation during manufacturing. PBMC derived from the stem cell graft were analyzed before peptide stimulation Day 0 and after 9 days of expansion Day 9 by flow cytometry. Cells were gated on CD3+ T cells. Click here for additional data file.Flow cytometry of epitope specific T cells after adoptive transfer. Example of flow cytometric assessment of virus specific T cells in the peripheral blood of patient 5, 14 days after the second infusion of the T cell product. The patient was positive for HLA*A02:01 and B*08:01. As shown in four epitopes (CLG, GLC, YVL, FLY) presented on HLA A*02:01 and two epitopes (QAK, RAK) presented on HLA*B08:01 could be used to stimulate donor-derived EBV-specific T cells. Two CMV derived epitopes (NLV, VLE) with restriction to HLA A*02:01 and two epitopes with restriction to HLA B*08:01 were available in the peptide pools to stimulate CMV specific T cells. The same peptides were used to load HLA pentamers for flow cytometric analysis. Click here for additional data file.Study Inclusion/Exclusion criteria. Inclusion criteria are shown for patients and donors separately. Only patients with serologic positive (CMV, EBV) 10/10 HLA-matched donors were included in the study. In addition, the study protocol required the presence of at least one of the following HLA class I loci: A*01:01, A*02:01, B*07:02, B*08:01, B*35:01, C*07:02. Before each T cell product transfusion “additional inclusion criteria” were applied as indicated to prevent infusion when acute GvHD or treatment with steroids, or signs of infection are present.Click here for additional data file.