Lacey G. Campbell

Recognition of HLA-A2-restricted mammaglobin-A-derived epitopes by CD8+ cytotoxic T lymphocytes from breast cancer patients.

A breast cancer-associated antigen, mammaglobin-A, is specifically expressed in 80% of primary breast tumors. The definition of immune responses against this highly expressed breast cancer-specific antigen should be of great value in the development of new therapeutic strategies for breast cancer. Thus, the purpose of this study was to identify HLA-A2-restricted mammaglobin-A-derived epitopes recognized by CD8+ cytotoxic T lymphocytes (CTL). We identified seven mammaglobin-A-derived candidate epitopes that bind the HLA-A2 molecule (Mam-A2.1-7) by means of a HLA class I-peptide binding computer algorithm from the Bioinformatics & Molecular Analysis Section of the National Institutes of Health. Subsequently, we determined that CD8+ CTLs from breast cancer patients reacted to the Mam-A2.1 (83–92, LIYDSSLCDL), Mam-A2.2 (2–10, KLLMVLMLA), Mam-A2.3 (4–12, LMVLMLAAL), Mam-A2.4 (66–74, FLNQTDETL), and Mam-A2.7 (32–40, TINPQVSKT) epitopes using an IFN-c ELISPOT assay. Interestingly, healthy individuals also showed high reactivity to the Mam-A2.2 epitope. Two CD8+ CTL lines generated in vitro against TAP-deficient T2 cells loaded with the candidate epitopes showed significant cytotoxic activity against the Mam-A2.1-4 epitopes. These CD8+CTL lines recognized a HLA-A2+breast cancer cell line expressing the Mam-A2.1 epitope. In addition, DNA vaccination of HLA-A2+/human CD8+ double-transgenic mice with a DNA construct encoding the Mam-A2.1 epitope and the HLA-A2 molecule induced a significant expansion of epitope-specific CD8+ CTLs that recognize the same HLA- A2+/Mam-A2.1+ breast cancer cell line. In conclusion, these results demonstrate the immunotherapeutic potential of mammaglobin-A for the treatment and prevention of breast cancer.

Inhibition of Obliterative Airway Disease Development in Murine Tracheal Allografts by Matrix Metalloproteinase-9 Deficiency

This study was designed to define the roles of matrix metalloproteinase (MMP)‐2 and MMP‐9 in obliterative airway disease (OAD) in heterotopic murine tracheal allografts, considered a suitable animal model for chronic lung allograft rejection. BALB/c tracheal allografts were transplanted into MMP‐2‐deficient (−/−) and MMP‐9−/− mice. Also, wild‐type recipients were treated with doxycycline, a nonspecific MMP inhibitor. After 10, 20 and 30 days, allografts were analyzed for OAD development, intragraft levels of MMP‐2 and MMP‐9 and the frequency and cytokine/chemokine production profile of alloreactive T cells. Allografts transplanted into wild‐type mice developed OAD lesions within 30 days. These allografts revealed significant upregulation of both MMP‐2 and MMP‐9. Allografts transplanted into MMP‐9−/− and doxycycline‐treated recipients did not develop OAD. In contrast, allografts transplanted into MMP‐2−/− mice developed OAD lesions with normal kinetics. Interestingly, MMP‐9−/− recipients showed an enhanced T cell alloreactivity associated with an abnormal profile of cytokine/chemokine production. The enhanced T cell alloreactivity in MMP‐9−/− mice was mediated by enhanced dendritic cell stimulatory capacity as well as enhanced T cell responsive capacity. These results suggest that MMP‐9 plays an important role in the pathogenesis of OAD and may represent a target for the therapeutic intervention of chronic lung allograft rejection.

Different roles for matrix metalloproteinase-2 and matrix metalloproteinase-9 in the pathogenesis of cardiac allograft rejection

Recent studies have shown an increased expression of several matrix metalloproteinases (MMP) during cardiac, renal and pulmonary allograft rejection. To further define the roles of MMP‐2 and MMP‐9 in the pathogenesis of cardiac allograft rejection, BALB/c cardiac allografts were transplanted into MMP‐2‐deficient (−/−) and MMP‐9−/− mice. Allografts rejected by wild‐type mice revealed a significant increase in MMP‐2 and MMP‐9 expression. MMP‐2‐deficiency significantly prolonged allograft survival time. Functioning allografts harvested from MMP‐2−/− mice showed lower cellular infiltration and fibrosis than rejected allografts harvested from MMP‐2+/+ mice at the same time. In contrast, MMP‐9‐deficiency significantly decreased allograft survival time. Functioning allografts harvested from MMP‐9+/+ mice showed lower cellular infiltration and fibrosis than rejected allografts harvested from MMP‐9−/− mice at the same time. MMP‐2−/− recipients showed decreased T‐cell alloreactivity mediated by a defect in dendritic cell stimulatory and T‐cell responsive capacities. In contrast, MMP‐9−/− recipients showed increased T‐cell alloreactivity mediated by a significant increased in dendritic cell stimulatory and T‐cell responsive capacities. These results indicate that MMP2 and MMP‐9 play significantly different roles in the process of cardiac allograft rejection.

Generation of CD8+ cytotoxic T lymphocytes against breast cancer cells by stimulation with mammaglobin-A-pulsed dendritic cells

Mammaglobin-A is exclusively expressed by breast cancer cells. Thus, mammaglobin-A-specific T cell immune responses may be useful for the design of new breast cancer-specific immunotherapies. We show herein that CD8+ T cells generated against recombinant mammaglobin-A-pulsed dendritic cells display a marked cytotoxic activity against mammaglobin-A-positive breast cancer cell lines. This study indicates the immunotherapeutic potential of this novel antigen for the treatment of breast cancer.