Quantitation of HLA-A*0201 bound tumor associated antigens on a peptide pulsed B cell line.
CTLs recognize 8- to 10-mer peptides on MHC class I molecules. Recent studies have shown that human CTLs kill autologous tumor cells in an HLA-restricted and peptide-specific manner, and that artificial pep- tides can stimulate tumor-specific CTLs both in vitro and in vivo. Accordingly, several human clinical trials using such peptides are ongoing worldwide. In such methods, the amount of peptide-MHC complexes that remain on the cell surface of APCs after peptide administration is crucial, because CTL activation depends on the number of ligated TCRs and co-stimulation. However, it remains uncertain how many peptide-MHC complexes are reconstituted and remain on live cells after peptide administration. We herein examined the binding affinities of five HLA-A*0201 restricted peptides-four TAAs and one HIV antigen-to HLA-A*0201 molecules and their decay rates on a live B cell line using tandem mass spectrometry. Our experiments showed that nearly 10(5) peptide-MHC complexes per cell could be reconstituted on a cell surface by pulsing a high dose of peptide even if the binding affinities were intermediate or low. However, the decay rates observed for these pep- tide-MHC complexes on a B cell line were faster than previously estimated.
Wataya
H
H
Department of Genetics, CREST (Core Research for Evolutional Science and Technology), Kyushu University, 812-8582, Fukuoka, Japan.
Kamikawaji
N
N
Nakanishi
Y
Y
Takayama
K
K
Hara
N
N
Sasazuki
T
T
D000818:Animals; D000951:Antigens, Neoplasm / Q000276:immunology / Q000378:metabolism*; D002144:Callithrix; D002460:Cell Line; D002462:Cell Membrane / Q000737:chemistry / Q000276:immunology / Q000378:metabolism; D004306:Dose-Response Relationship, Immunologic; D054303:HIV Reverse Transcriptase / Q000276:immunology / Q000378:metabolism; D015789:HLA-A2 Antigen / Q000276:immunology / Q000378:metabolism*; D006801:Humans; D006825:Hybridomas; D008562:Membrane Glycoproteins / Q000276:immunology / Q000378:metabolism; D051379:Mice; D009363:Neoplasm Proteins / Q000276:immunology / Q000378:metabolism; D010446:Peptide Fragments / Q000138:chemical synthesis / Q000276:immunology / Q000378:metabolism*; D011485:Protein Binding / Q000276:immunology; D018719:Erb-b2 Receptor Tyrosine Kinases / Q000276:immunology / Q000378:metabolism; D021241:Spectrometry, Mass, Electrospray Ionization; D016159:Tumor Suppressor Protein p53 / Q000276:immunology / Q000378:metabolism; D058951:gp100 Melanoma Antigen
D000951:Antigens, Neoplasm; D054303:HIV Reverse Transcriptase; D015789:HLA-A2 Antigen; D008562:Membrane Glycoproteins; D009363:Neoplasm Proteins; D010446:Peptide Fragments; D018719:Erb-b2 Receptor Tyrosine Kinases; D016159:Tumor Suppressor Protein p53; D058951:gp100 Melanoma Antigen; C091577:HER2-neu-derived peptide (654-662); C072013:MAGEA3 protein, human; C489519:Mageb3 protein, mouse; C548637:PMEL protein, human; C548642:Pmel protein, mouse