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A mouse mutant p53 product recognized by CD4+ and CD8+ T cells
1Ludwig Institute for Cancer Research, New York Unit, Memorial Sloan-Kettering Cancer Center, NY 10021.
Abstract:
The T-cell response to mutated and normal p53 products of BALB/c-derived Meth A sarcoma was analyzed. Meth A p53 is known to have three missense point mutations in codons 132, 168, and 234, and 24 peptides containing wild-type or mutated sequences at the three mutation sites were constructed. Spleen cells from BALB/c or (BALB/c x C57BL/6)F1 mice immunized with p53 peptides were sensitized in vitro with the corresponding peptides. Because Meth A is resistant to cytotoxic T cells, the sensitive P1-HTR cell line, which expresses a low level of p53 lacking the Meth A p53 mutations, was chosen as a target, either pulse-labeled with p53 peptides or transfected with plasmids containing coding sequences from Meth A p53. One peptide, a nonamer containing the codon 234 mutation (234CM), induced CD8+ cytotoxic T cells that lysed 234CM-pulsed P1-HTR cells in an H-2Kd-restricted fashion. P1-HTR cells pulsed with the corresponding wild-type peptide were only weakly lysed by 234CM-reactive cytotoxic T cells. P1-HTR cells pulsed with other wild-type or mutated p53 peptides were not lysed by 234CM-reactive cytotoxic T cells, nor could these peptides, including 234CW (the wild-type counterpart to 234CM), elicit cytotoxic cells. P1-HTR cells transfected with plasmids coding for the 234CM sequence and expressing high p53 levels were weakly lysed by 234CM-reactive cytotoxic T cells. However, lysis of one of the transfectants was significantly increased by pretreatment with interferon gamma. A proliferative response of CD4+ T cells was elicited by immunization with 234CM and 234CW, but not with other p53-related peptides. The specificity of 234CM-induced CD4+ T cells for 234-region peptides was broader than the reactivity of 234CM-reactive cytotoxic T cells. Mice immunized with 234CM in incomplete Freund's adjuvant showed heightened resistance to Meth A challenge.
Insights
A specific mutated p53 peptide (234CM) induced CD8+ cytotoxic T cells that target cancer cells. This peptide also stimulated CD4+ T cells and enhanced resistance to Meth A sarcoma challenge in mice.
Area of Science:
- Immunology
- Oncology
- Molecular Biology
Background:
- The p53 tumor suppressor protein is frequently mutated in cancer.
- Meth A sarcoma in BALB/c mice harbors specific missense mutations in p53.
- Understanding T-cell responses to mutated p53 is crucial for cancer immunotherapy.
Purpose of the Study:
- To analyze the T-cell response to mutated and wild-type p53 peptides from Meth A sarcoma.
- To identify specific p53 epitopes that elicit cytotoxic T-cell and helper T-cell responses.
- To evaluate the potential of p53-derived peptides in cancer immunotherapy.
Main Methods:
- Synthesis of 24 peptides containing wild-type or mutated sequences of Meth A p53.
- In vitro sensitization of mouse spleen cells with p53 peptides.
- Cytotoxicity assays using peptide-pulsed or transfected P1-HTR cells as targets.
- T-cell proliferation assays to assess CD4+ T-cell responses.
- In vivo tumor challenge experiments in mice.
Main Results:
- A nonamer peptide with the codon 234 mutation (234CM) induced CD8+ cytotoxic T cells recognizing H-2Kd-restricted targets.
- CD8+ T-cell lysis was specific for the 234CM peptide and enhanced by interferon-gamma treatment of target cells.
- Both 234CM and its wild-type counterpart (234CW) elicited CD4+ T-cell proliferation with broader peptide specificity.
- Immunization with 234CM peptide conferred heightened resistance to Meth A sarcoma challenge in mice.
Conclusions:
- The codon 234 mutation in Meth A p53 generates a specific epitope recognized by CD8+ cytotoxic T cells.
- p53-derived peptides can induce both cellular and humoral immune responses.
- Targeting mutated p53 with specific peptides holds promise for developing novel cancer immunotherapies.