Intracellular Delivery of Anti-pPKCθ (Thr538) via Protein Transduction Domain Mimics for Immunomodulation.

TitleIntracellular Delivery of Anti-pPKCθ (Thr538) via Protein Transduction Domain Mimics for Immunomodulation.
Publication TypeJournal Article
Year of Publication2016
AuthorsE Ozay, I, Gonzalez-Perez, G, Torres, JA, Vijayaraghavan, J, Lawlor, R, Sherman, HL, Garrigan, DT, Burnside, AS, Osborne, BA, Tew, GN, Minter, LM
JournalMol Ther
Date Published2016 Dec
KeywordsAnimals, Antibodies, Monoclonal, Humanized, Cell Differentiation, Cell Proliferation, Cell-Penetrating Peptides, Graft vs Host Disease, Humans, Immunomodulation, Isoenzymes, Leukocytes, Mononuclear, Lymphocyte Activation, Mice, Phosphorylation, Protein Kinase C, Protein Kinase C-theta, Signal Transduction, Th1 Cells

Targeting cellular proteins with antibodies, to better understand cellular signaling pathways in the context of disease modulation, is a fast-growing area of investigation. Humanized antibodies are increasingly gaining attention for their therapeutic potential, but the collection of cellular targets is limited to those secreted from cells or expressed on the cell surface. This approach leaves a wealth of intracellular proteins unexplored as putative targets for antibody binding. Protein kinase Cθ (PKCθ) is essential to T cell activation, proliferation, and differentiation, and its phosphorylation at specific residues is required for its activity. Here we report on the design, synthesis, and characterization of a protein transduction domain mimic capable of efficiently delivering an antibody against phosphorylated PKCθ (Thr538) into human peripheral mononuclear blood cells and altering expression of downstream indicators of T cell activation and differentiation. We used a humanized, lymphocyte transfer model of graft-versus-host disease, to evaluate the durability of protein transduction domain mimic:Anti-pPKCθ modulation, when delivered into human peripheral mononuclear blood cells ex vivo. We demonstrate that protein transduction domain mimic:Antibody complexes can be readily introduced with high efficacy into hard-to-transfect human peripheral mononuclear blood cells, eliciting a biological response sufficient to alter disease progression. Thus, protein transduction domain mimic:Antibody delivery may represent an efficient ex vivo approach to manipulating cellular responses by targeting intracellular proteins.

Alternate JournalMol Ther
PubMed ID27633441
PubMed Central IDPMC5167783
Grant ListP01 CA166009 / CA / NCI NIH HHS / United States
T32 GM108556 / GM / NIGMS NIH HHS / United States
T32 GM135096 / GM / NIGMS NIH HHS / United States