Induction of cytolytic T lymphocytes against pediatric solid tumors in vitro using autologous dendritic cells pulsed with necrotic primary tumor.
OBJECTIVE
Effective and generally applicable methods for generating cancer vaccines in children have not been defined. Dendritic cells (DCs) are the most potent professional antigen-presenting cells capable of activating primary cytolytic T cells. We tested the ability of DCs generated from pediatric patients' peripheral blood monocytes and pulsed with a necrotic tumor to activate autologous tumor-specific cytolytic T cells.


METHODS
Tumor and peripheral blood cells were obtained from pediatric patients undergoing biopsy or resection for advanced solid tumors according to an institutional research board-approved protocol and after acquiring informed consent from them. To generate DCs, we treated peripheral blood monocytes with granulocyte-macrophage colony stimulating factor and interleukin (IL)-4. Maturation was induced with a cytokine cocktail (CC) containing tumor necrosis factor-alpha, IL-6, IL-1beta, and prostaglandin E2. The DC phenotype was assayed using flow cytometry. Tumor necrosis was induced by exposure to UV-B irradiation (1000 mJ). Dendritic cells pulsed with a UV-B-treated primary tumor and matured with CC were used to stimulate autologous peripheral blood lymphocytes weekly. Tumor-specific cytolytic activity was assayed using 4-hour 51Cr release.


RESULTS
Peripheral blood monocytes isolated from pediatric patients differentiated into immature DCs (CD14-, MHCII+ [major histocompatibility complex], CD80(low), CD86(low)) in the presence of granulocyte-macrophage colony stimulating factor and IL-4. Cytokine cocktail induced maturation of DCs, as characterized by increased expressions of MHCII, CD83, CD80, and CD86. Patients' peripheral blood lymphocytes stimulated in vitro with DCs loaded with a necrotic primary tumor and matured with CC specifically lysed autologous neuroblastoma in 7 of 9 patients.


CONCLUSIONS
Dendritic cells generated from the peripheral blood of children with advanced solid tumors and pulsed with a necrotic primary tumor undergo maturation and effectively stimulate autologous tumor-specific cytolytic T cells in vitro. We describe a simple method for generating a vaccine capable of activating cytotoxic T cells against pediatric solid tumors that does not require the genetic identification of tumor-associated antigens.
Shilyansky
Joel
J
Division of Pediatric Surgery, Department of Surgery, Medical College of Wisconsin, Milwaukee, WI 53226, USA.
Jacobs
Paulette
P
Doffek
Kara
K
Sugg
Sonia L
SL
D000293:Adolescent; D019496:Cancer Vaccines / Q000276:immunology*; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D002648:Child; D002675:Child, Preschool; D003713:Dendritic Cells / Q000276:immunology*; D006801:Humans; D007223:Infant; D008213:Lymphocyte Activation; D009336:Necrosis; D009369:Neoplasms / Q000276:immunology / Q000628:therapy*; D013602:T-Lymphocytes, Cytotoxic / Q000276:immunology*
D019496:Cancer Vaccines