[Identification of T cell epitopes from ovarian cancer associated anti-idiotype antibody].
OBJECTIVE
To identify the T cell epitopes from ovarian cancer associated anti-idiotypic antibody 6B11 in order to explore the molecular basis of 6B11 induced cellular immune responses against ovarian cancer.


METHODS
Potential human leukocyte antigen (HLA) A0201 ligands were predicted by using SYFPEITHI algorithm and tested by the T2 binding assay for screening of HLA-A2 binding peptides from 6B11 complimentary determining region (CDR). Cytotoxic T lymphocytes (CTL) to 6B11 or peptides were generated by 3 rounds of in vitro stimulation with 6B11 or peptide-pulsed dendritic cells (DC), and then tested by (51)Cr-release assay to ascertain the CTL epitope of 6B11. Cell proliferation assay was performed by using 6B11 specific CTL as responder cells and peptide-pulsed DC as stimulators to ascertain the helper T lymphocyte (Th) epitope of 6B11. Cytokine assay and interferon-gamma ELISPOT assay were performed to verify the CTL and Th epitope of 6B11 further.


RESULTS
Light chain CDR3 peptide (VL CDR3) of 6B11 induced specific CTL to kill HLA-A2 and target antigen positive ovarian cancer cells, which could be blocked by anti human major histocompatibility complex (MHC) I antibody. Heavy chain CDR3 peptide (VH CDR3) of 6B11 stimulated the proliferation of 6B11-primed CTL, which could be blocked mainly by anti human MHC-II antibody, and further experiments showed that part of the VH CDR3 peptide-primed CTL killed K562 cells. Peptides in VL CDR3 and VH CDR3 were the CTL and Th epitope mimicking the original antigen of 6B11 respectively. Collaboration of 6B11 CTL and Th epitope, or 6B11 CTL epitope and keyhole limpet hemocyanin (KLH), the former was more powerful in inducing specific cellular immune responses against ovarian cancer. 6B11 or corresponding CTL and Th epitope specific CTL secreted high levels of interleukin-2 (1630, 1503 ng/L) and interferon-gamma (5620, 5421 ng/L), while basal level of interleukin-4 was detected (253, 274 ng/L). ELISPOT assay confirmed the existence of specific interferon-gamma secreting cells in 6B11 or epitopes specific CTL (196/1 x 10(6) T cell, 184/1 x 10(6) T cell).


CONCLUSIONS
VL CDR3 and VH CDR3 peptides of 6B11 are the CTL and Th epitopes mimicking original antigen which could duplicate the activity of intact 6B11 to induce the cellular immune responses against ovarian cancer. The results have significant theoretical and practical value in application of anti-idiotypic antibody as anti tumor vaccine. The acquired CTL and Th epitopes as constituents of future multiple peptides against ovarian cancer could be used in peptide vaccine based ovarian cancer therapy.
Li
Wei
W
Department of Gynecologic Oncology Center, Peking University People's Hospital, Beijing 100044, China.
Cui
Heng
H
Chang
Xiao-Hong
XH
Cheng
Hong-Yan
HY
Cheng
Ye-Xia
YX
Feng
Jie
J
Fu
Tian-Yun
TY
D000888:Antibodies, Anti-Idiotypic / Q000276:immunology* / Q000494:pharmacology; D000912:Antibodies, Neoplasm / Q000276:immunology; D000951:Antigens, Neoplasm / Q000276:immunology* / Q000378:metabolism; D015496:CD4-Positive T-Lymphocytes / Q000276:immunology; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D022801:Complementarity Determining Regions / Q000276:immunology; D003713:Dendritic Cells / Q000276:immunology*; D004797:Enzyme-Linked Immunosorbent Assay; D018984:Epitopes, T-Lymphocyte / Q000276:immunology*; D005260:Female; D006801:Humans; D010051:Ovarian Neoplasms / Q000276:immunology* / Q000628:therapy; D013602:T-Lymphocytes, Cytotoxic / Q000276:immunology*
D000888:Antibodies, Anti-Idiotypic; D000912:Antibodies, Neoplasm; D000951:Antigens, Neoplasm; D022801:Complementarity Determining Regions; D018984:Epitopes, T-Lymphocyte