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Induction and characterization of cytotoxic T-lymphocytes recognizing a mutated p21ras peptide presented by
A Van Elsas1, H W Nijman, C E Van der Minne
1Department of Clinical Oncology, University Hospital, Leiden, The Netherlands.
Abstract:
The ras oncogene is frequently found to be activated in human cancer through point mutations at codons 12, 13 or 61. We explored whether these altered p21ras protein sequences contain peptide sequences that can activate naive CD8+ cytotoxic T lymphocytes (CTL). Several wild-type and mutated p21ras peptides were identified that carry a binding motif for human leukocyte antigen (HLA)-A*0201. Two peptides were found to bind strongly to this allele. CD8+ CTL bulk cultures specifically reacting with one of these peptides could be induced, using processing-defective T2 cells loaded with peptide CLLDILDTAGL as stimulators. The peptide is derived from p21ras, position 51-61, and carries a 61 Gln-->Leu mutation. In contrast, a 9-mer peptide CLLDILDTA corresponding to amino acid sequence 51-59 of wild-type p21ras did not yield reactive CTL cultures. T-cell clones with low affinity for the 11-mer peptide were isolated from CLLDILDTAGL-reactive bulk cultures. These T cells did not lyse melanoma cells transfected with 61-Leu N-ras, although lysis was found when these transfectants were pulsed with the 11-mer peptide. Possibly, T cells of higher affinity may be required to demonstrate processed peptide on the cell surface. The combined experiments suggest that a peptide derived from mutated p21ras can be recognized by HLA class I-restricted CTL, whereas an analogous wild-type p21ras peptide may not be immunogenic.
Insights
Mutated ras oncogenes in cancer can produce peptides that activate CD8+ cytotoxic T lymphocytes (CTL). These cancer-specific peptides may be recognized by the immune system, unlike normal p21ras peptides.
Area of Science:
- Oncology
- Immunology
- Molecular Biology
Background:
- Ras oncogenes are frequently activated in human cancers via point mutations.
- These mutations lead to altered p21ras protein sequences.
Purpose of the Study:
- To investigate if mutated p21ras protein sequences contain peptides that can activate naive CD8+ cytotoxic T lymphocytes (CTL).
- To identify specific mutated peptides recognized by human leukocyte antigen (HLA)-A*0201.
Main Methods:
- Identification of wild-type and mutated p21ras peptides with HLA-A*0201 binding motifs.
- Induction of CD8+ CTL bulk cultures using peptide-loaded, processing-defective T2 cells.
- Isolation and characterization of T-cell clones.
Main Results:
- Two p21ras peptides strongly bound to HLA-A*0201.
- A mutated p21ras peptide (position 51-61, 61 Gln-->Leu) induced reactive CTL cultures, while the wild-type equivalent did not.
- Isolated T-cell clones showed low affinity and did not lyse target cells unless peptide-pulsed.
Conclusions:
- Peptides derived from mutated p21ras can be recognized by HLA class I-restricted CTL.
- Mutated p21ras peptides may be immunogenic in cancer, while wild-type peptides may not be.
- Higher affinity T cells might be necessary for effective recognition of processed mutated peptides on cancer cells.