Introduction of 65 kDa Antigen of Mycobacterium tuberculosis to Cancer Cells Enhances Anti‐Tumor Effect of BCG Therapy

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Abstract
Bacillus Calmette Guerin (BCG) immunotherapy has anti‐tumorigenic effects against bladder cancer. To improve the efficacy of BCG therapy, we introduced the gene encoding the 65 kDa heat shock protein (hsp) of Mycobacterium tuberculosis into a mouse malignant melanoma cell line (B16). An expression vector harboring the 65 kDa antigen gene was transfected into B16 using Lipofectamine, then expression of the antigen was confirmed by RT‐PCR and Western blotting. Several cell lines expressing 65 kDa antigen were established (B16/65kDa). We also established a control cell line transfected with the vector alone (B16/con). All cell lines (B16, B16/con, B16/65kDa) were injected intraperitoneally into syngeneic mice with or without BCG prior immunization and the development of tumor ascites was examined. To analyze the mechanism of the anti‐tumor effect, CD4 T cells or CD8 T cells were depleted in vivo by administering the corresponding monoclonal antibody. B16/65kDa expressed the 65 kDa hsp of M. tuberculosis. The tumor growth of B16/65kDa was slightly retarded in naive mice, but significantly inhibited by BCG. The anti‐tumor effect was totally abrogated in mice deficient in CD4 T cells, suggesting that CD4 T cells are involved in this process. The 65 kDa hsp of M. tuberculosis was expressed after gene transduction in a malignant melanoma cell line and significantly enhanced the anti‐tumor effect of BCG immunotherapy. CD4 T cells play an important role in this anti‐tumor effect.