276. Enhancement of Cytotoxic T-Lymphocyte Responses Against Carcinoembryonic antigen (CEA) Using Genetically Modified Dendritic Cells Expressing CEA and CD40 Ligand

Abstract

Carcinoembryonic antigen (CEA), one of the well-characterized tumor-associated antigens is commonly expressed by a wide variety of adenocarcinomas. However, as a self-antigen it is poorly immunogenic due to immune tolerance. Although several human-leukocyte-antigen (HLA)-restricted epitopes within the CEA protein that can be recognized by human T lymphocytes have been identified, CEA expressing tumor cells are generally weakly recognized by the immune system. Therefore, development of immunotherapeutic strategies to break its tolerance is necessary to generate effective anti-tumor immunity against CEA expressing malignancies. In this study we evaluated the efficiency of genetically modified human peripheral blood monocyte-derived dendritic cell (MoDC) expressing CEA and immunostimulatory molecule CD40 ligand (CD40L) to generate effective immune response against autologous as well as tumor cell lines expressing CEA. The recombinant human immunodeficiency virus (HIV)-1-based self-inactivating lentiviral vectors expressing the full length CEA (Lenti-CEA) or human CD40L (Lenti-CD40L) under the transcriptional control of human PGK promoter provided a highly efficient reproducible gene transfer into immature MoDC. After single exposure to Lenti-CEA and/or Lenti-CD40L vector, more than 80% of DCs showed cell surface expression of CEA alone and/or CD40L without any toxicity. Notably, the Lenti-CD40L transduced DCs showed increased cell surface expression of maturation marker CD83 and molecules involved in antigen presentation (CD80, CD86, CD54, HLA-ABC and HLA-DR), secreted high amounts of T-cell immunostimulatory cytokine interleukin (IL)-12, and showed enhanced T-cell stimulatory function. Importantly, the CEA/CD40L transduced DCs were able to generate CEA-specific T-cell responses ex vivo, which was higher than that of DC transduced with CEA alone and matured with recombinant soluble CD40L or prostaglandin E-2 either alone or in combination with tumor necrosis factor-a. The T-cell lines generated ex vivo with CEA/CD40L expressing DCs comprised both CD8+ and CD4+ T-cells possessing polyclonal T-cell receptor repertoire, showed increased antigen-specific proliferative response, and secreted high levels of interferon-|[gamma]| and IL-2 upon antigen recall. Importantly, the CEA-specific T-cell lines were able to specifically recognize CEA expressing autologous target cells as well as CEA positive tumor cell lines in MHC restricted manner. Although cytotoxicity toward target cells was mediated primarily by CD8+ T-cells, interestingly both CD8+ and CD4+ T-cells were able to lyse CEA expressing target cells. Taken together, these results suggest that lentiviral vector-mediated expression of CEA and CD40L in DC may be a promising strategy for the rational development of DC vaccines or antigen-specific T-cells for adoptive immunotherapy of CEA expressing malignancies.