[rAAV/BA46-transfected dendritic cells can induce specific cellular immunity].
OBJECTIVE
To study the feasibility of transfecting breast cancer BA46 gene into dendritic cells (DCs) using adeno-associated virus (AAV) to induce specific cellular immunity.


METHODS
Mononuclear cells (DC precursor) were isolated from the peripheral blood of healthy donors by density gradient centrifugation and infected with rAAV/BA46/Neo virus stock (transfection group) or pulsed with 293 cell lysate (control group). In both groups, maturation of the DC precursor was induced by granulocyte-macrophage colony-stimulating factor (GM-CSF), interleukin-4 (IL-4) and tumor necrosis factor-alpha(TNF-alpha). On day 7, the DCs were collected and mixed with T cells at the ratio of 1 to 20 to induce cytotoxic T lymphocytes (CTL). The capacity of the DCs in stimulating T lymphocyte proliferation was assessed using (3)H-thymidine incorporation assay. The expressions of interferon-gamma (IFN-gamma), IL-4, CD4, CD8, CD25 and CD69 in the CTLs were analyzed with cytometry, and the cytotoxicity of the CTLs was evaluated with (51)Cr-release assay using BA46-positive breast cancer cell line Hs578T as the target.


RESULTS
The DCs transfected with BA46 gene exhibited potent capacity to stimulate T lymphocyte proliferation. The CTL population induced by the transfected DCs expressed high levels of CD8, CD69 and IFN-gamma, and showed strong cytotoxicity against BA46-positive breast cancer cell line Hs578T, which was BA46 antigen-specific and MHC-limited.


CONCLUSIONS
The success in BA46 gene transfer in the DCs that induce specific cellular immunity provides the experimental basis for breast cancer immunotherapy using genetically modified cells.
You
Chang-xuan
CX
Department of Oncology, Nanfang Hospital, Southern Medical University, Guangzhou 510515, China. ycx6026@yahoo.com
Su
Jin
J
Liao
Wang-jun
WJ
Zhang
Jun-yi
JY
Liu
Yong
Y
Paul
L Hermonat
LH
Luo
Rong-cheng
RC
D000954:Antigens, Surface / Q000235:genetics / Q000378:metabolism*; D001943:Breast Neoplasms / Q000235:genetics / Q000276:immunology; D002478:Cells, Cultured; D003713:Dendritic Cells / Q000276:immunology* / Q000378:metabolism; D000229:Dependovirus / Q000235:genetics* / Q000378:metabolism; D005260:Female; D016178:Granulocyte-Macrophage Colony-Stimulating Factor / Q000494:pharmacology; D006801:Humans; D007111:Immunity, Cellular; D007167:Immunotherapy; D015847:Interleukin-4 / Q000494:pharmacology; D008894:Milk Proteins / Q000235:genetics / Q000378:metabolism*; D013602:T-Lymphocytes, Cytotoxic / Q000276:immunology*; D014162:Transfection
D000954:Antigens, Surface; C098942:MFGE8 protein, human; D008894:Milk Proteins; D015847:Interleukin-4; D016178:Granulocyte-Macrophage Colony-Stimulating Factor