Cytotoxicity of Cord Blood Derived Her2/neu-specific Cytotoxic T Lymphocytes against Human Breast Cancer in vitro and in vivo

Abstract

The Her2/neu oncogene encodes a transmembrane protein with homology to the epidermal growth factor receptor. Overexpression of this gene contributes to the aggressiveness of breast cancer and poor prognosis. Therefore, Her2/neu is an ideal target molecule for generating effective cytotoxic T lymphocytes (CTLs) against breast cancers. This study reports on the generation of Her2/neu-specific CTL from umbilical cord blood mononuclear cells (UCBC) using dendritic cells primed with Her2/neu-derived peptide (KIFGSLAFL, E75) for immunostimulation. The CTLs showed specific cytotoxicity to Her2/neu high expressing MDA-453 but not toward Her2/neu low expressing MDA-231 human breast cancer cells. Similarly generated CTLs stimulated with irrelevant peptide pulsed dendritic cells did not show significant cytotoxicity towards breast cancer targets. The phenotypes of cells in culture showed high percentage of CD3+, CD4+ and CD8+T cells as determined by flow cytometry. However, the antibody mediated blocking assay demonstrated that only HLA-Class I restricted CD8+ cells are involved in the cytotoxicity. Furthermore, in vivo studies showed that treatment of SCID mice bearing MDA-453 tumor with Her2/neu-specific CTLs resulted in significant inhibition of tumor growth compared to untreated tumor bearing control mice. These results demonstrate that human umbilical cord blood mononuclear cells are a good source for generating Her2/neu-specific CTLs against human breast cancer both in vitro and in vivo.

Molecular basis of aggressive disease in chronic lymphocytic leukemia patients with 11q deletion and trisomy 12 chromosomal abnormalities

“…Mononuclear cells (MNCs) were isolated from heparinized whole blood from CLL patients using Accu-Prep™ lymphocyte separation medium by Ficoll-Hypaque density centrifugation method (Accurate Chemical and Scientific Corp., Westbury, NY, USA) as described previously (11,14). The immunophenotypes of the isolated MNCs were determined by flow cytometry with flurochrome (phycoerythrin/PE or fluoroscein isothiocyanate/FITC)-labeled antibodies specific for cell surface markers (11).…”

Ultrastructural basis of enhanced antitumor cytotoxicity of cord blood-derived CTLs: A comparative analysis with peripheral blood and bone marrow

“…PB is currently the source of choice for immunotherapy in clinical trials, although the response rates in these trials are not as high as was initially hoped. It has been shown that UCB effector cells can be stimulated in vitro to acquire nonspecific effector cytotoxicity equal to or higher than that of PB (43), as well as specific antitumor effects that surpass PBderived cytotoxic cells (28)(29)(30). We have shown that CTLs generated from UCBC have superior antitumor cytotoxic effects than PB-derived CTLs against two different target tumor cell lines.…”

“…Tumor-specific cytotoxic effects of UCBC have also been shown, both in vitro and in vivo (25)(26)(27)(28)(29)(30). Head-to-head comparisons of UCB-and PB-derived effector cells suggest that, after stimulation, UCB acquires cytotoxic abilities that equal or surpass that of PB cells.…”

“…Head-to-head comparisons of UCB-and PB-derived effector cells suggest that, after stimulation, UCB acquires cytotoxic abilities that equal or surpass that of PB cells. Wang et al (29) and Joshi et al (30) showed that, after priming both UCB-DC and PB-DC with the Her2/neu peptide, cytotoxic T-lymphocytes (CTL) were able to selectively kill Her2/neu expressing cell lines. Their results also showed increased cytotoxic effects of UCB-CTLs over PB-CTLs.…”

Immunity to breast cancer in mice immunized with fibroblasts transfected with a cDNA expression library derived from small numbers of breast cancer cells

“…1,2 The antitumor immune responses following immunization with effective vaccines can be of sufficient magnitude to prolong the lives of experimental animals, and patients, providing a compelling rationale for the use of tumor vaccines in cancer therapy. 3,4 In breast cancer, TAA such as MUC-1, 5 HER-2/neu, 6 MAGE-1, 7 BAGE, 8 mammaglobin 9 and MENA 10 have been identified as potential targets of CTLs. It is likely that these are only several representations of an undefined, and possibly large number of TAA expressed by breast cancer cells.…”