No abstract

No abstract

The identification of antigenic peptides presented on the tumor cell surface by HLA class I molecules and recognized by tumor-specific cytotoxic T lymphocytes may lead to a peptide vaccine capable of inducing protective cellular immunity. We demonstrate that both HLA-A2-restricted breast and ovarian tumor-specific cytotoxic T lymphocytes recognize shared (3) and MART (melanoma antigen recognized by T cells) (4) in melanoma. The discovery of these genes and peptides substantiates the existence of such antigen systems, but unfortunately, melanoma accounts for only 3% of all malignancies and 1% of all cancer deaths (5).HLA-A2 is expressed in approximately 50% of Caucasians (6) and has been demonstrated to play a critical role in antigen presentation of both viral antigens (7) and tumor antigens from a variety of cancers (8-10). HER2/neu is a 185-kDa transmembrane glycoprotein with tyrosine kinase activity and extensive homology to the epidermal growth factor receptor (11). HER2/neu is ubiquitously expressed in many tumors and known to be overexpressed in approximately 30-40% of all ovarian and breast cancers (12).We have shown that tumor-specific CTLs can be demonstrated in the tumor-infiltrating lymphocytes (TILs) isolated from ovarian cancers (13). We have also found that HLA-A2 presents TAA in this disease (14). The level of expression of the HER2/neu oncogene in ovarian cancer positively correlates with recognition by HLA-A2+ ovarian tumor-specific CTLs, and HLA-A2+ melanoma cells transfected with the HER2/neu gene become sensitive to ovarian cancer-specific CTLs (15). Therefore, HLA-A2-presented, HER2/neuderived antigenic peptides may be recognized by ovarian cancer-specific CTLs and, potentially, by breast cancerspecific CTLs as well.In this study, we have searched the HER2/neu sequence (16) for HLA-A2-binding peptides (17), and we have found that both ovarian and breast cancer-specific CTLs recognize a nine-amino acid peptide from the transmembrane portion of the HER2/neu protein (GP2; amino acids 654-662). This peptide is widely expressed in HER2/neu+ tumors and is capable of inducing HLA-A2-restricted, tumor-specific CTL populations in vitro. METHODSGeneration of Tumor-Specific CTLs. Tumor-specific CTLs were generated from fresh tumor specimens obtained through the Departments of Surgery, Gynecologic Oncology, and Pathology at Brigham and Women's Hospital (BWH) and Beth Israel Hospital in Boston, under approval of the Institutional Review Boards. Solid tumor specimens were processed as previously described (14). Briefly, specimens were minced manually and enzymatically digested, and the lymphocytes and tumor cells were separated by centrifugation over discontinuous Ficoll (Organon Teknika-Cappel) gradients. TILs were suspended in RPMI-1640 medium (BioWhittaker) supplemented with 10% fetal calf serum (BioWhittaker) and antibiotics. Cultures were suspended at 5 x 105 cells per ml on solid-phase, anti-CD3 polystyrene plates (Orthoclone OKT3, Ortho Pharmaceuticals) for 48 h. T cells were then mainta...

To evaluate the safety, immune responses, and antitumor responses after the administration of α-galactosylceramide (αGalCer) KRN7000-pulsed PBMC cultured with IL-2 and GM-CSF (IL-2/GM-CSF-cultured PBMCs), a phase I-II study in patients with non-small cell lung cancer was conducted. Patients with advanced non-small cell lung cancer or recurrent lung cancer refractory to the standard therapy were eligible. αGalCer-pulsed IL-2/GM-CSF-cultured PBMCs (1 × 109/m2) were i.v. administered four times. Immune responses were monitored weekly. Twenty-three patients were enrolled in this study and 17 cases (73.9%) completed. No severe adverse event related to the treatment was observed. After the injection of αGalCer-pulsed IL-2/GM-CSF-cultured PBMCs, an increased number of IFN-γ-producing cells in the peripheral blood were detected in 10 patients (58.8%). Five cases remained as stable disease, and the remaining 12 cases were evaluated as progressive disease. The estimated median survival time (MST) of the 17 cases was 18.6 mo (range, 3.8 to 36.3 mo). Ten patients who displayed increased IFN-γ-producing cells (≥2-fold) showed prolonged MST (31.9 mo; range, 14.5 to 36.3 mo) as compared with poor-responder patients (n = 7) MST (9.7 mo; range, 3.8 to 25.0 mo) (log-rank test, p = 0.0015). The administration of αGalCer-pulsed IL-2/GM-CSF-cultured PBMCs was well tolerated and was accompanied by the successful induction of NKT cell-dependent immune responses. The increased IFN-γ-producing cells that result from αGalCer stimulation in PBMCs were significantly associated with prolonged MST. These results are encouraging and warrant further evaluation for survival benefit of this immunotherapy.