Fibroblasts genetically engineered to secrete interleukin 12 can suppress tumor growth and induce antitumor immunity to a murine melanoma in vivo.
Interleukin 12 (IL-12), a disulfide-linked heterodimeric cytokine produced primarily by macrophages, is composed of light (p35) and heavy (p40) chains. It binds to a receptor on T-cells and natural killer cells, promoting the induction of primarily a TH1 response in vitro and in vivo. To determine whether paracrine IL-12 secretion can alter tumor cell growth or promote antitumor immunity, we have developed a delivery system using genetically engineered fibroblasts in murine tumor models. NIH3T3 cells were stably transfected to express 100-240 units/10(6) cells/48 h of IL-12 using expression plasmids carrying both the murine p35 and p40 genes of murine IL-12. The effects of paracrine secretion of IL-12 on tumor establishment and vaccination models were examined using the poorly immunogenic murine melanoma cell line (BL-6) in C57BL/6 mice. To determine the effects of IL-12 on tumor formation, nonirradiated BL-6 cells were inoculated s.c. into C57BL/6 mice admixed with NIH3T3 cells transfected with both subunits of mIL-12 (3T3-IL-12) or with cells transfected with only the neomycin phosphotransferase gene (3T3-Neo). Compared to mice given injections of BL-6 alone, the day of emergence of detectable tumors was significantly delayed in mice given injections of BL-6 admixed with 3T3-IL-12, but not in mice with BL-6 admixed with 3T3-Neo. Effectiveness in this system was related to the amount of IL-12 expressed by the 3T3-IL-12. To determine the ability of locally secreted IL-12 at the tumor site to induce antitumor immunity, 10(6) irradiated tumor cells mixed with 3T3-IL-12 or 3T3-Neo were injected as a vaccine, and the response to a tumor challenge was subsequently examined. With a tumor challenge of less than 1 x 10(5) nonirradiated BL-6 cells, significant delay of establishment of tumor was noted with a relatively small amount of IL-12 secretion (1.2 units/5 x 10(5) cells/48 h). Larger amounts of secreted IL-12 provided no additional therapeutic benefit. Histological examination of tumor inoculum with 3T3-IL-12 secreting a high level of IL-12 showed peritumoral accumulation of macrophages, a characteristic capsule around the tumor composed of palisades of fibroblasts, and decreased numbers of CD4+ cells in the tumor. These results suggest that local delivery of IL-12 inhibits tumor growth in a dose dependent manner but leads to the development of an antitumor immune response when IL-12 is expressed at the tumor site at the relatively small amount indicated above.(ABSTRACT TRUNCATED AT 400 WORDS)
Tahara
H
H
Department of Surgery, University of Pittsburgh School of Medicine, Pennsylvania.
Zeh
H J
HJ
Storkus
W J
WJ
Pappo
I
I
Watkins
S C
SC
Gubler
U
U
Wolf
S F
SF
Robbins
P D
PD
Lotze
M T
MT
D016475:3T3 Cells; D000818:Animals; D001483:Base Sequence; D002455:Cell Division / Q000276:immunology; D002460:Cell Line; D005347:Fibroblasts / Q000378:metabolism*; D007167:Immunotherapy; D018664:Interleukin-12; D007378:Interleukins / Q000235:genetics / Q000378:metabolism*; D019868:Kanamycin Kinase; D008545:Melanoma / Q000276:immunology / Q000473:pathology / Q000517:prevention & control*; D051379:Mice; D008969:Molecular Sequence Data; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000235:genetics*; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000378:metabolism*; D014162:Transfection
D007378:Interleukins; D012333:RNA, Messenger; D018664:Interleukin-12; D017853:Phosphotransferases (Alcohol Group Acceptor); D019868:Kanamycin Kinase