title: Surrogate in vitro activation of innate immunity synergizes with interleukin-7 to unleash rapid antigen-driven outgrowth of CD4+ and CD8+ human peripheral blood T-cells naturally recognizing MUC1, HER2/neu and other tumor-associated antigens.
abstract: 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-driven T-cell expansion might best be triggered in vitro by acute activation of innate immunity to mimic a life-threatening infection. Unfractionated peripheral blood mononuclear cells (PBMC) were subjected to a two-step culture, first synchronizing their exposure to exogenous antigens with aggressive surrogate activation of innate immunity, followed by γ-chain cytokine-modulated T-cell hyperexpansion. Step 1 exposure to GM-CSF plus paired Toll-like receptor agonists (resiquimod and LPS), stimulated abundant IL-12 and IL-23 secretion, as well as 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 reliably presented to CD4+ T-cells and cross-presented to CD8+ T-cells. Both presentation and cross-presentation demonstrated proteasomal and Sec61 dependence that could bypass the endoplasmic reticulum. Step 2 exposure to exogenous IL-7 or IL-7+IL-2 produced selective and sustained expansion of both CD4+ and CD8+ peptide-specific T-cells with a predominant interferon-γ-producing T1-type, as well as the antigen-specific ability to lyse tumor targets. Other γ-chain cytokines and/or combinations were initially proliferogenic, but followed by a contractile phase not observed with IL-7 or IL-7+IL-2. Regulatory T-cells were minimally propagated under these culture conditions. This mechanistically rational culture sequence, effective even for unvaccinated donors, enables rapid preparation of T-cells recognizing tumor-associated antigens expressed by the majority of human cancers, including pancreatic cancers, breast cancers and glioblastomas.
authors.affiliation: Department of Biochemistry and Molecular Biology, Mayo Clinic, Scottsdale, AZ, USA.
authors.forename: Latha B
authors.initials: LB
authors.lastname: Pathangey
authors.affiliation: Department of Biochemistry and Molecular Biology, Mayo Clinic, Scottsdale, AZ, USA.
authors.forename: Dustin B
authors.initials: DB
authors.lastname: McCurry
authors.affiliation: Department of Biochemistry and Molecular Biology, Mayo Clinic, Scottsdale, AZ, USA.|Department of Immunology, Mayo Clinic, Scottsdale, AZ, USA.|Department of Hematology and Oncology, Mayo Clinic, Scottsdale, AZ, USA.
authors.forename: Sandra J
authors.initials: SJ
authors.lastname: Gendler
authors.affiliation: Department of Biochemistry and Molecular Biology, Mayo Clinic, Scottsdale, AZ, USA.
authors.forename: Ana L
authors.initials: AL
authors.lastname: Dominguez
authors.affiliation: Department of Biochemistry and Molecular Biology, Mayo Clinic, Scottsdale, AZ, USA.
authors.forename: Jessica E
authors.initials: JE
authors.lastname: Gorman
authors.affiliation: Department of Biochemistry and Molecular Biology, Mayo Clinic, Scottsdale, AZ, USA.
authors.forename: Girish
authors.initials: G
authors.lastname: Pathangey
authors.affiliation: Department of Hematology and Oncology, Mayo Clinic, Scottsdale, AZ, USA.
authors.forename: Laurie A
authors.initials: LA
authors.lastname: Mihalik
authors.affiliation: Tumor Vaccine Group, Center for Translational Medicine in Women's Health, University of Washington, Seattle, WA, USA.
authors.forename: Yushe
authors.initials: Y
authors.lastname: Dang
authors.affiliation: Tumor Vaccine Group, Center for Translational Medicine in Women's Health, University of Washington, Seattle, WA, USA.
authors.forename: Mary L
authors.initials: ML
authors.lastname: Disis
authors.affiliation: Department of Immunology, Mayo Clinic, Scottsdale, AZ, USA.|Department of Hematology and Oncology, Mayo Clinic, Scottsdale, AZ, USA.
authors.forename: Peter A
authors.initials: PA
authors.lastname: Cohen
mesh_terms: D000951:Antigens, Neoplasm / Q000276:immunology* / Q000378:metabolism; D015496:CD4-Positive T-Lymphocytes / Q000276:immunology* / Q000378:metabolism; D018414:CD8-Positive T-Lymphocytes / Q000276:immunology* / Q000378:metabolism; D049109:Cell Proliferation / Q000187:drug effects; D002478:Cells, Cultured; D045142:Cross-Priming / Q000276:immunology; D016178:Granulocyte-Macrophage Colony-Stimulating Factor / Q000276:immunology / Q000494:pharmacology; D006801:Humans; D007093:Imidazoles / Q000494:pharmacology; D007113:Immunity, Innate / Q000187:drug effects / Q000276:immunology; D016219:Immunotherapy, Adoptive / Q000379:methods; D007371:Interferon-gamma / Q000276:immunology / Q000378:metabolism; D007376:Interleukin-2 / Q000276:immunology / Q000494:pharmacology; D015851:Interleukin-7 / Q000276:immunology / Q000494:pharmacology*; D007963:Leukocytes, Mononuclear / Q000276:immunology / Q000378:metabolism; D008070:Lipopolysaccharides / Q000276:immunology / Q000494:pharmacology; D008213:Lymphocyte Activation / Q000187:drug effects / Q000276:immunology; D018396:Mucin-1 / Q000276:immunology* / Q000378:metabolism; D009369:Neoplasms / Q000276:immunology / Q000628:therapy; D010455:Peptides / Q000276:immunology / Q000494:pharmacology; D018719:Erb-b2 Receptor Tyrosine Kinases / Q000276:immunology* / Q000378:metabolism; D050378:T-Lymphocytes, Regulatory / Q000276:immunology / Q000378:metabolism
chemical_list: D000951:Antigens, Neoplasm; D016178:Granulocyte-Macrophage Colony-Stimulating Factor; D007093:Imidazoles; D007371:Interferon-gamma; D007376:Interleukin-2; D015851:Interleukin-7; D008070:Lipopolysaccharides; D018396:Mucin-1; D010455:Peptides; D018719:Erb-b2 Receptor Tyrosine Kinases; C060192:MUC1 protein, human; C508053:ERBB2 protein, human; C402365:resiquimod
keywords: GM-CSF; IL-7; MUC1; PBMC; TLR agonists; adoptive therapy