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Identification of cyclin B1 as a shared human epithelial tumor-associated antigen recognized by T cells.

Kao H, Marto JA, Hoffmann TK, Shabanowitz J, Finkelstein SD, Whiteside TL, Hunt DF, Finn OJ - J. Exp. Med. (2001)

Bottom Line: We also detected memory T cells specific for one or more of these peptides in patients with breast cancer and squamous cell carcinomas of the head and neck (SCCHN).Immunohistochemical analysis of tumor lines and tissue sections showed cyclin B1 overexpression and aberrant localization in the cytoplasm instead of the nucleus.Sequencing genomic DNA and cDNA corresponding to P1-P6 region showed that differences in COOH-terminal residues were not due to either DNA mutations or errors in transcription, suggesting a high error rate in translation of cyclin B1 protein in tumors.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Genetics and Biochemistry, University of Pittsburgh, Pittsburgh, PA 15261, USA.

ABSTRACT
We eluted peptides from class I molecules of HLA-A2.1(+) breast adenocarcinoma and loaded reverse phase high-performance liquid chromatography (HPLC) fractions onto dendritic cells to prime naive CD8(+) T cells. Fractions that supported growth of tumor-specific cytotoxic T lymphocytes were analyzed by nano-HPLC micro-ESI tandem mass spectrometry. Six HLA-A2.1-binding peptides, four 9-mers (P1-P4) differing in the COOH-terminal residue, and two 10-mers (P5 and P6) with an additional COOH-terminal alanine, were identified in one fraction. Peptide sequences were homologous to cyclin B1. We primed CD8(+) T cells from another HLA-A2.1(+) healthy donor with synthetic peptides and generated P4-specific responses. We also detected memory T cells specific for one or more of these peptides in patients with breast cancer and squamous cell carcinomas of the head and neck (SCCHN). T cells from one patient, restimulated once in vitro, could kill the tumor cell line from which the peptides were derived. Immunohistochemical analysis of tumor lines and tissue sections showed cyclin B1 overexpression and aberrant localization in the cytoplasm instead of the nucleus. Sequencing genomic DNA and cDNA corresponding to P1-P6 region showed that differences in COOH-terminal residues were not due to either DNA mutations or errors in transcription, suggesting a high error rate in translation of cyclin B1 protein in tumors.

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Cyclin B1 protein is overexpressed in SCCHN. A and B, SCCHN cell Line PCI-13. C–J, SCCHN tumor sections. A, C, E, G, and I, original magnification, ×10. B, D, F, H, and J, original magnification, ×20.
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fig6: Cyclin B1 protein is overexpressed in SCCHN. A and B, SCCHN cell Line PCI-13. C–J, SCCHN tumor sections. A, C, E, G, and I, original magnification, ×10. B, D, F, H, and J, original magnification, ×20.

Mentions: To rule out that the intense staining of cyclin B1 observed in the tumor cell lines might be a result of a prolonged in vitro culture, we examined a tumor cell line as well as tumor tissue sections obtained from the SCCHN patients whom we had analyzed for cyclin B1–specific T cell responses. Fig. 6, A and B , shows intense cytoplasmic staining of cyclin B1 in the tumor cell line PCI-13, derived from the tumor of patient A in Fig. 3. Very high expression of cyclin B1 in the cell line correlates with strong cyclin B1–specific T cell responses observed in this patient. Intense cytoplasmic cyclin B1 staining was also observed in the tumor tissue samples (Fig. 6, C and D; E and F) of two other patients who exhibited cyclin B1-specific T cell responses (patients C and D, respectively; Fig. 3). No cyclin B1 staining was detected in the normal mucosa surrounding the tumor. The tumor seen in Fig. 6, G and H, showed weak and diffuse cyclin B1 staining that was not convincingly positive. Patient B from whom the tumor was obtained did have cyclin B1–specific T cell responses (Fig. 3). The same weak staining was seen in the tumor shown in Fig. 6, I and J, derived from a patient who did not exhibit any HLA class I–restricted T cell responses against the cyclin B1 peptides (data not shown).


Identification of cyclin B1 as a shared human epithelial tumor-associated antigen recognized by T cells.

Kao H, Marto JA, Hoffmann TK, Shabanowitz J, Finkelstein SD, Whiteside TL, Hunt DF, Finn OJ - J. Exp. Med. (2001)

Cyclin B1 protein is overexpressed in SCCHN. A and B, SCCHN cell Line PCI-13. C–J, SCCHN tumor sections. A, C, E, G, and I, original magnification, ×10. B, D, F, H, and J, original magnification, ×20.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2195974&req=5

fig6: Cyclin B1 protein is overexpressed in SCCHN. A and B, SCCHN cell Line PCI-13. C–J, SCCHN tumor sections. A, C, E, G, and I, original magnification, ×10. B, D, F, H, and J, original magnification, ×20.
Mentions: To rule out that the intense staining of cyclin B1 observed in the tumor cell lines might be a result of a prolonged in vitro culture, we examined a tumor cell line as well as tumor tissue sections obtained from the SCCHN patients whom we had analyzed for cyclin B1–specific T cell responses. Fig. 6, A and B , shows intense cytoplasmic staining of cyclin B1 in the tumor cell line PCI-13, derived from the tumor of patient A in Fig. 3. Very high expression of cyclin B1 in the cell line correlates with strong cyclin B1–specific T cell responses observed in this patient. Intense cytoplasmic cyclin B1 staining was also observed in the tumor tissue samples (Fig. 6, C and D; E and F) of two other patients who exhibited cyclin B1-specific T cell responses (patients C and D, respectively; Fig. 3). No cyclin B1 staining was detected in the normal mucosa surrounding the tumor. The tumor seen in Fig. 6, G and H, showed weak and diffuse cyclin B1 staining that was not convincingly positive. Patient B from whom the tumor was obtained did have cyclin B1–specific T cell responses (Fig. 3). The same weak staining was seen in the tumor shown in Fig. 6, I and J, derived from a patient who did not exhibit any HLA class I–restricted T cell responses against the cyclin B1 peptides (data not shown).

Bottom Line: We also detected memory T cells specific for one or more of these peptides in patients with breast cancer and squamous cell carcinomas of the head and neck (SCCHN).Immunohistochemical analysis of tumor lines and tissue sections showed cyclin B1 overexpression and aberrant localization in the cytoplasm instead of the nucleus.Sequencing genomic DNA and cDNA corresponding to P1-P6 region showed that differences in COOH-terminal residues were not due to either DNA mutations or errors in transcription, suggesting a high error rate in translation of cyclin B1 protein in tumors.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Genetics and Biochemistry, University of Pittsburgh, Pittsburgh, PA 15261, USA.

ABSTRACT
We eluted peptides from class I molecules of HLA-A2.1(+) breast adenocarcinoma and loaded reverse phase high-performance liquid chromatography (HPLC) fractions onto dendritic cells to prime naive CD8(+) T cells. Fractions that supported growth of tumor-specific cytotoxic T lymphocytes were analyzed by nano-HPLC micro-ESI tandem mass spectrometry. Six HLA-A2.1-binding peptides, four 9-mers (P1-P4) differing in the COOH-terminal residue, and two 10-mers (P5 and P6) with an additional COOH-terminal alanine, were identified in one fraction. Peptide sequences were homologous to cyclin B1. We primed CD8(+) T cells from another HLA-A2.1(+) healthy donor with synthetic peptides and generated P4-specific responses. We also detected memory T cells specific for one or more of these peptides in patients with breast cancer and squamous cell carcinomas of the head and neck (SCCHN). T cells from one patient, restimulated once in vitro, could kill the tumor cell line from which the peptides were derived. Immunohistochemical analysis of tumor lines and tissue sections showed cyclin B1 overexpression and aberrant localization in the cytoplasm instead of the nucleus. Sequencing genomic DNA and cDNA corresponding to P1-P6 region showed that differences in COOH-terminal residues were not due to either DNA mutations or errors in transcription, suggesting a high error rate in translation of cyclin B1 protein in tumors.

Show MeSH
Related in: MedlinePlus