Natural tumor-reactive T-cells can be rapidly expanded from human unfractionated peripheral blood and licensed for adoptive immunotherapy (AIT)
Pathangey LB*1, McCurry DB*1, Gendler SJ1,2,3,4, Dominguez AL1, Gorman J1, Pathangey G1, Mihalik LA4, Dang Y5, Disis ML5 and Cohen PA1,3,4 Departments of 1Research, 2Biochemistry and Molecular Biology, 3Immunology, and 4Hematology and Oncology, Mayo Clinic in Scottsdale, 5Tumor Vaccine Group, University of Washington. Seattle, WA
Effective adoptive immunotherapy has proved elusive for many types of human cancer, often due to difficulties achieving robust expansion of natural tumor-specific T-cells from peripheral blood. We hypothesized that antigen(Ag)-driven T-cell expansion might best be triggered in vitro by acute activation of innate immunity to mimic a life-threatening infection. Unfractionated PBMC were subjected to a two-step culture, first synchronizing their exposure to exogenous Ag with TLR agonist (TLRa) activation of type-1 polarized myeloid DC to enhance processing and presentation of Ag, followed by IL-7 driven T-cell expansion. Step 1 exposure to GM-CSF plus paired TLRa (resiquimod and LPS) stimulated abundant IL-12 and upregulated co-stimulatory molecules and CD11c expression within the myeloid (CD33+) subpopulation. Added synthetic long peptides (>20aa) derived from widely expressed oncoproteins (MUC1, HER2/neu and CMVpp65), were presented to CD4+ T-cells and cross-presented to CD8+ T-cells. Step 2 exposure to exogenous IL-7 produced selective and sustained expansion of both CD4+ and CD8+ peptide-specific T-cells with a predominant IFN-γ-producing T1-type by 16 days, as well as the Ag-specific ability to lyse tumor targets. Regulatory T-cells were minimally propagated. Such cultured T-cells can be expanded in scaled-up culture vessels and successfully cryopreserved. This mechanistically rational culture sequence, effective for unvaccinated donors or cancer patients, enables rapid preparation of T-cells recognizing tumor-associated Ag expressed by most human cancers, including pancreas, breast and glioblastomas.
NIH P50 CA102701, RO1 136632, Foundations: Chao, Lefkofsky and Mayo