Enhancement of antitumor immunity against B16 melanoma tumor using genetically modified dendritic cells to produce cytokines.
Dendritic cells (DC) that have been genetically modified to express cytokine genes may be novel tools for inducing antitumor immune responses. In the present study, the pMX retroviral vector was modified to express the mouse IL-2 (mIL-2pMX) and mouse IL-12 (mIL-12pMX) genes. Supernatants from 293 cells transfected with pMX retroviral vectors were harvested and used to transduce mouse lin- bone marrow (BM) progenitor cells. After 48 h co-culture with pseudotype retrovirus, BM cells were cultured for 12 days in the presence of mGM-CSF, mSCF and mTNF-alpha to obtain a DC-enriched fraction. Flow cytometric analysis showed that GFP protein expression in these cultures was 20-40% and that 40-50% of the cultured BM cells were positive for the DC marker, DEC205. About 60% of cells sorted for DEC205 also expressed GFP. The supernatants of DC-mIL-2 and DC-mIL-12 cultured for 48 h contained 5.2 +/- 0.15 and 33.9 +/- 2.6 ng cytokine protein per milliliter, respectively. Intratumoral injection of DC-mIL-2 or DC-mIL-12 on days 8 and 15 after the intradermal injection of 1 x 105 B16F10 cells, resulted in a significant reduction in tumor size by day 21, as compared with mice treated with unmodified DC or DC-GFP. Longer term analysis as assessed at day 42 revealed that B16 tumor-bearing mice treated with cytokine gene-modified DC survived significantly longer than mice from other groups. Spleen cells obtained from DC-treated mice were specifically sensitized for the generation of CTL by subsequent restimulation with gene-modified DC. These results suggested that DC genetically modified to express IL-2 or IL-12 can induce potent antitumor responses against well-established, poorly immunogenic B16F10 tumors. Gene Therapy (2000) 7, 2113-2121.
Growth Factor Division, National Cancer Center Research Institute, 5-1-1 Tsukiji, Chuo-ku, Tokyo 104, Japan.
Y
Y
Akiyama
M
M
Watanabe
K
K
Maruyama
F W
FW
Ruscetti
R H
RH
Wiltrout
K
K
Yamaguchi
D000818:Animals; D003713:Dendritic Cells / Q000276:immunology*; D015870:Gene Expression; D015316:Genetic Therapy / Q000379:methods*; D005822:Genetic Vectors / Q000008:administration & dosage; D049452:Green Fluorescent Proteins; D006412:Hematopoietic Stem Cells / Q000276:immunology; D016219:Immunotherapy, Adoptive / Q000379:methods*; D015552:Injections, Intralesional; D018664:Interleukin-12 / Q000235:genetics* / Q000276:immunology; D007376:Interleukin-2 / Q000235:genetics* / Q000276:immunology; D008164:Luminescent Proteins / Q000235:genetics; D008546:Melanoma, Experimental / Q000628:therapy*; D051379:Mice; D008810:Mice, Inbred C57BL; D012190:Retroviridae / Q000235:genetics; D014161:Transduction, Genetic
D007376:Interleukin-2; D008164:Luminescent Proteins; D049452:Green Fluorescent Proteins; D018664:Interleukin-12