B. Fisk et al., MASS-SPECTROMETRIC ANALYSIS OF NATURALLY PROCESSED PEPTIDES RECOGNIZED BY OVARIAN TUMOR-ASSOCIATED CD8(+) CTL, International journal of oncology, 10(1), 1997, pp. 159-169
Antigens recognized by cytotoxic T cells (CTL) are expressed as peptid
es presented by MHC class I molecules. To isolate peptides from the MH
C molecule HLA-A2.1 and identify epitopes that define the activity pro
file of ovarian CD8(+) CTL, peptides were separated by reverse-phase h
igh-pressure liquid chromatography (HPLC), and analyzed by electrospra
y ionization-tandem mass spectrometry (ES-MS). HLA-A2.1-bound peptides
were extracted from the ovarian tumor line SKOV3 transfected with the
HLA-A2.1 (clone 1E4) and C1R.A2 cells transfected with HCA-A2.1 and H
ER-2 (clone HER-2.J) by immunoaffinity chromatography. At least five p
eaks of distinct retention times (termed 1, 2A, 2B, 2C, and 3) were re
cognized by an ovarian HER-2(high) (HER-2(hi)) tumor-associated HLA-A2
(+), CD8(+) CTL line. ES-MS analysis was performed for peak 2B peptide
s from both types of cells. In the four consecutive fractions of peak
2B, at least 27 and 16 ion species of mass-to-charge (m/z) ratio betwe
en 760-1300 were detected in 1E4 and HER-2.J cells, respectively. The
abundance of four 1E4 and six HER-2.J ions believed to be peptides in
four consecutive HPLC fractions in this peak matched the CTL activity
profile. Of these, two ions with actual m/z ratios 497.3-498.4 and 792
.8-793.2, were found in the peak 2B from both types of cells. Since li
ttle is known about the tumor Ag recognized in human cancers, characte
rization of these ions may lead to identification of novel tumor Ag in
breast and ovarian cancers. This may also be useful in developing qua
ntitative approaches to the identification of tumor Ag and the determi
nation of epitope density on tumor and normal cells. This may help cha
racterize the relationship between tumor immunity and epitope toleranc
e in human epithelial cancers.