[Identification of a naturally presented MAGE-A3 epitope on the surface of HLE cell line by mass spectrometry].
OBJECTIVE
To identify a naturally presented HLA-A2-restricted epitope of MAGE-A3 antigen, FLWGPRALV (MAGE-A3(271 - 279)), on the surface of a human hepatocellular carcinoma (HCC) cell line HLE.


METHODS
Synthetic peptide FLWGPRALV, served as positive control target, was analyzed by HPLC and HPLC-ESI-TOF-MSMS, in order to determine its HPLC elution time, mass-spectrometric characteristics and the lowest detection limitation by the two approaches. 3 x 10(9) HLE cells were collected, peptides naturally presented by major histocompatibility complex (MHC) molecules on the cell surface were isolated by mild acid elution, and concentrated by lyophilization, then the mixtures of peptides were fractioned by HPLC. The ingredient ranged from 2 min before the elution time determined by the synthetic peptide to 2 min after that was collected, concentrated by lyophilization, and analyzed by HPLC-ESI-TOF-MSMS, to identify the existence of the MAGE-A3(271 - 279) peptide.


RESULTS
The HPLC-ESI-TOF-MSMS detection provided an evidence for the existence of a doubly charged ion of (m/z)(2) 529.9, which was further analyzed by collision induced dissociation. The doubly charged ion was ultimately identified as the MAGE-A3(271 - 279) peptide, its amino sequence was FLWGPRALV and its molecular weight was 1058.4 Da.


CONCLUSIONS
MAGE-A3(271 - 279) epitope could be naturally presented by HLA-A2 molecules to the surface of HCC cell line and MAGE-A3(271 - 279) peptide may have potential immunotherapeutic value in HCC patients.
Peng
Ji-run
JR
Center of Hepatobiliary Surgery, Peking University People's Hospital, Beijing 100044, China.
Dong
Nan
N
Liu
Hu-wei
HW
Leng
Xi-sheng
XS
D000595:Amino Acid Sequence; D017951:Antigen Presentation; D000951:Antigens, Neoplasm / Q000032:analysis* / Q000302:isolation & purification; D006528:Carcinoma, Hepatocellular / Q000276:immunology / Q000473:pathology; D045744:Cell Line, Tumor; D002851:Chromatography, High Pressure Liquid; D018984:Epitopes, T-Lymphocyte / Q000032:analysis* / Q000302:isolation & purification; D015789:HLA-A2 Antigen / Q000276:immunology; D006801:Humans; D008113:Liver Neoplasms / Q000276:immunology / Q000473:pathology; D013058:Mass Spectrometry; D009363:Neoplasm Proteins / Q000032:analysis* / Q000302:isolation & purification
D000951:Antigens, Neoplasm; D018984:Epitopes, T-Lymphocyte; D015789:HLA-A2 Antigen; C072013:MAGEA3 protein, human; D009363:Neoplasm Proteins