The HER2/neu-derived peptide p654-662 is a tumor-associated antigen in human pancreatic cancer recognized by cytotoxic T lymphocytes

The protooncogene HER2/neu encodes a 185-kDa transmembrane protein with extensive homology to the epidermal growth factor receptor. It is overexpressed in several human cancers of epithelial origin, such as pancreatic cancer. Previously, we demonstrated that cytotoxic T lymphocytes (CTL) derived from breast, ovarian, and non-small cell lung cancer recognized a peptide derived from HER2/neu. To evaluate whether this HLA-A2-binding peptide is a tumor-associated antigen (TAA) in pancreatic cancer, the ability of HER2/neu-reactive CTL to lyse human pancreatic carcinoma cells was tested. CTL were generated from tumor-associated T lymphocytes from HLA-A2+ HER2/neu+ breast and ovarian cancer patients. All CTL recognized autologous and allogeneic HER2/ neu+ tumor cells in an HLA-A2-restricted fashion. Furthermore, all CTL recognized p654-662 (GP2) derived from HER2/neu. These CTL also recognized HER2/neu+ pancreatic cancer cells in an HLA-A2-restricted fashion. HER2/neu+ HLA-A2- pancreatic cancer were not or only poorly lysed. Repeated stimulation of HLA-A2+ PBL from pancreatic cancer patients using the HER2/neu-derived peptide resulted in specific recognition of this peptide and, more importantly, HER2/neu+ pancreatic tumors in an HLA-A2-restricted fashion. Autologous HLA-A2+ fibroblasts or HLA-A2+ malignant melanoma cells were not recognized. HLA-A2- peptide-stimulated T lymphocytes showed no significant cytotoxicity. These results demonstrate that this HER2/neu-derived peptide is a shared TAA among several adenocarcinomas including pancreatic carcinoma, suggesting a common mechanism of recognition of these human tumors by T lymphocytes. The identification of the HER2/neu-derived peptide GP2 as a TAA in pancreatic cancer provides an opportunity for the design of novel immunotherapy and vaccine strategies.     

Heat shock induces transient p53-dependent cell cycle arrest at G1/S

The HER-2/neu proto-oncogene is frequently amplified or overexpressed in human breast and ovarian cancers, and is significantly correlated with shorter survival. We have previously reported that the adenovirus type 5 early region 1A (E1A) gene product can repress HER-2/neu overexpression by repressing HER-2/neu promoter activity, and suppress the tumorigenic potential of HER-2/neu-overexpressing ovarian cancer cells. To examine E1A tumor suppressor function in breast cancer, we transduced E1A in vitro by adenovirus into both HER-2/neu-overexpressing and low expressing human breast cancer cell lines. In HER-2/neu-overexpressing cells, E1A greatly inhibited tumor cell growth in vitro. However, in HER-2/neu low expressing cancer cell lines, E1A had no significant effect on cell growth in culture medium. To test the therapeutic efficacy of E1A, we used both adenovirus-mediated and cationic liposome-mediated E1A gene delivery systems in an orthotopic breast cancer animal model. An advanced breast cancer model was established by inoculation of HER-2/neu-overexpressing human breast cancer cells in mammary fat pad and treated by local injections of either replication-deficient adenovirus expressing E1A, Ad.E1A(+) or a liposome-E1A DNA complex. As controls, mice bearing tumors were also treated with Ad.E1A(-) which is virtually the same adenovirus as Ad.E1A(+) except that E1A is deleted, a liposome-E1A frame-shift mutant DNA complex, or just PBS. In mice bearing a HER-2/neu-overexpressing breast cancer cell line, E1A delivered either by adenovirus or liposome significantly inhibited tumor growth and prolonged mouse survival compared with the controls. In fact, 60-80% of E1A-treated mice lived longer than 2 years versus only 0-20% of control mice (P<0.05). Western blot analysis showed that E1A protein was expressed in tumor tissue and immunohistochemical analysis showed that HER-2/neu p185 protein expression was suppressed. Taken together, our results indicated that both adenovirus and cationic liposome delivery systems were effective in transfering E1A gene for tumor suppression in a HER-2/neu-overexpressing breast cancer model.     

Breast cancer and family history: a multivariate analysis of levels of tumor HER2 protein and family history of cancer in women who have breast cancer

BACKGROUND: The HER2 gene, located on the long arm of chromosome 17, codes for a protein with the characteristics of a growth factor receptor. In a preliminary study, we reported that high levels of tumor HER2 (erbB-2/neu) protein are associated with a family history of breast cancer (that is, one or more female blood relatives with breast cancer). METHODS: We have now collected a larger number of subjects (94) and performed a multivariate analysis of the independent variables family history of breast cancer, tumor estrogen receptor, age, and tumor DNA index. Family history of breast cancer was assessed by questioning the patient, in many cases by telephone. RESULTS: HER2 levels were significantly higher in women with a family history of breast cancer (p = 0.015, two-tailed t-test). The 27 women with family history were predominantly postmenopausal, mean age 61 +/- 2.3 (mean +/- SEM), versus a mean age of 56 +/- 1.7 for the 67 women with no family history. Of the 27 women with a family history of breast cancer, 13 had a first-degree relative (mother or sister) with the disease. The remaining 14 women had other relatives (grandmothers, aunts, cousins, or a niece) with breast cancer. The results of multiple linear regression analysis, with HER2 as the dependent variable, showed that family history of breast cancer was significantly associated with elevated HER2 levels in the tumors (p = 0.0038), after controlling for the effects of age, tumor estrogen receptor, and DNA index. CONCLUSIONS: The association of family history of breast cancer and elevated tumor HER2 protein suggests that postmenopausal familial breast cancer may be associated with altered HER2 expression.     

DNA methylation in the promoter region of the p16 (CDKN2/MTS-1/INK4A) gene in human breast tumours

The p16 (CDKN2/MTS-1/INK4A) gene is one of several tumour-suppressor genes that have been shown to be inactivated by DNA methylation in various human cancers including breast tumours. We have used bisulphite genomic sequencing to examine the detailed sequence specificity of DNA methylation in the CpG island promoter/exon 1 region in the p16 gene in DNA from a series of human breast cancer specimens and normal human breast tissue (from reductive mammaplasty). The p16 region examined was unmethylated in the four normal human breast specimens and in four out of nine breast tumours. In the other five independent breast tumour specimens, a uniform pattern of DNA methylation was observed. Of the nine major sites of DNA methylation in the amplified region from these tumour DNAs, four were in non-CG sequences. This unusual concentration of non-CG methylation sites was not a general phenomenon present throughout the genome of these tumour cells because the methylated CpG island regions of interspersed L1 repeats had a pattern of (almost exclusively) CG methylation similar to that found in normal breast tissue DNA and in DNA from tumours with unmethylated p16 genes. These data suggest that DNA methylation of the p16 gene in some breast tumours could be the result of an active process that generates a discrete methylation pattern and, hence, could ultimately be amenable to therapeutic manipulation.