Protein transduction of dendritic cells for NY-ESO-1-based immunotherapy of myeloma.
Myeloma vaccines, based on dendritic cells pulsed with idiotype or tumor lysate, have been met with limited success, probably in part due to insufficient cross-priming of myeloma antigens. A powerful method to introduce myeloma-associated antigens into the cytosol of dendritic cells is protein transduction, a process by which proteins fused with a protein transduction domain (PTD) freely traverse membrane barriers. NY-ESO-1, an immunogenic antigen by itself highly expressed in 60% of high-risk myeloma patients, was purified to near homogeneity both alone and as a recombinant fusion protein with a PTD, derived from HIV-Tat. Efficient entry of PTD-NY-ESO-1 into dendritic cells, confirmed by microscopy, Western blotting, and intracellular flow cytometry, was achieved without affecting dendritic cell phenotype. Experiments with amiloride, which inhibits endocytosis, and N-acetyl-l-leucinyl-l-norleucinal, a proteasome inhibitor, confirmed that PTD-NY-ESO-1 entered dendritic cells by protein transduction and was degraded by the proteasome. Tetramer analysis indicated superior generation of HLA-A2.1, CD8+ T lymphocytes specific for NY-ESO-1(157-165) with PTD-NY-ESO-1 compared with NY-ESO-1 control protein (44% versus 2%, respectively). NY-ESO-1-specific T lymphocytes generated with PTD-NY-ESO-1 secreted IFN-gamma indicative of a Tc1-type cytokine response. Thus, PTD-NY-ESO-1 accesses the cytoplasm by protein transduction, is processed by the proteasome, and NY-ESO-1 peptides presented by HLA class I elicit NY-ESO-1-specific T lymphocytes.
Batchu
Ramesh B
RB
Myeloma Institute for Research and Therapy, Section for Gene and Immunotherapy, University of Arkansas for Medical Sciences, Little Rock, Arkansas 72205, USA.
Moreno
Amberly M
AM
Szmania
Susann M
SM
Bennett
Grant
G
Spagnoli
Guilio C
GC
Ponnazhagan
Selvarangan
S
Barlogie
Bart
B
Tricot
Guido
G
van Rhee
Frits
F
D000584:Amiloride / Q000494:pharmacology; D000951:Antigens, Neoplasm / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology*; D003713:Dendritic Cells / Q000276:immunology* / Q000378:metabolism; D005347:Fibroblasts / Q000502:physiology; D015234:HLA-A Antigens / Q000276:immunology; D015789:HLA-A2 Antigen; D006801:Humans; D016219:Immunotherapy, Adoptive / Q000379:methods*; D008565:Membrane Proteins / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology*; D009000:Monocytes / Q000276:immunology; D009101:Multiple Myeloma / Q000276:immunology* / Q000628:therapy*; D046988:Proteasome Endopeptidase Complex / Q000276:immunology / Q000378:metabolism; D013601:T-Lymphocytes / Q000276:immunology; D014161:Transduction, Genetic
D000951:Antigens, Neoplasm; C104948:CTAG1B protein, human; D015234:HLA-A Antigens; C435939:HLA-A*02:01 antigen; D015789:HLA-A2 Antigen; D008565:Membrane Proteins; D000584:Amiloride; D046988:Proteasome Endopeptidase Complex