Download Purpose: Radionuclide gene therapy and immunotherapy have both

Purpose: Radionuclide gene therapy and immunotherapy have both been investigated
as potential cancer treatments, but have been found to be ineffective as single
therapies against aggressive tumors. In this study, I evaluated the tumoricidal
effects of combined hMUC1 vaccination and hNIS radioiodine gene therapy in an
animal tumor model.
Experimental Design: A stable cell line (CT26/hMUC1-hNIS-Fluc: CMNF) expressing
the hMUC1, hNIS, and Fluc genes was established using a retrovirus/lenti virus
system. In vitro survival rates of CMNF cells were determined using I-131 (1 mCi)
in vitro clonogenic assays. In addition, 1¡¿105 CMNF cells were transplanted
into 28 mice, and 4 and 11 days after tumor challenge, tumor bearing mice were
immunized intramuscularly with pcDNA (50§¶/50§¡) or pcDNA-hMUC1 (50§¶/50§¡), and
subsequently injected with PBS (100§¡) or I-131 (2mCi/100§¡ in saline)
intraperitoneally [4 groups (7 mice/group), which are referred to as the
pcDNA+PBS, phMUC1+PBS, pcDNA+I-131, and phMUC1+I-131 groups]. Tumor progression
was monitored using bioluminescence images and by taking caliper measurements.
Tumor free mice (%) were defined as mice with no bioluminescence at tumor sites.
Thirty-two days after the initial tumor challenge, mice in the pcDNA+I-131 and
phMUC1+I-131 groups were rechallenged with CMNF cells (5¡¿104 cells/mouse). To
investigate the hMUC1-associated immune response generated by combination
therapy, the levels of hMUC1-associated CD8+IFN¥ã+T cells and hMUC1-associated
CD8+IFN¥ã+T cells generated by hMUC1 antigen loaded CD11+ cells were examined.
In addition, the cytotoxic effect of T cells on CMNF cells was measured using
LDH assays.
Results: FACS, I-125 uptake, and luciferase assays showed that CMNF cells highly
expressed hMUC1, hNIS, and Fluc. In vitro clonogenic assays showed that the
survival rates of CMNF cells were markedly reduced to 13.6 ¡¾ 1.0% compared with
parental CT26 cells after treating them with 1 mCi of I-131. Marked tumor growth
inhibition was only observed in the phMUC1+I-131 group by bioluminescence at 32
days after initial tumor challenge. In terms of long-term tumor protection,
hMUC1-expressing cancer growth was completely inhibited in the phMUC1+I-131
group but not in the pcDNA+I-131 group after tumor re-challenge. Numbers of
tumor free mice (%) were higher in the phMUC1+I-131 group than in the other
three groups (pcDNA+PBS, phMUC1+PBS, pcDNA+I-131, and phMUC1+I-131, 0%, 0%, 14%,
and 85%, respectively). Furthermore, levels of hMUC1-associated CD8+IFN¥ã+T
cells were higher in the phMUC1+I-131 group than in the other three groups
(pcDNA+PBS, phMUC1+PBS, pcDNA+I-131, and phMUC1+I-131 groups, 53.5¡¾1.2,
91.7¡¾1.7, 676.5¡¾3.6, and 1278.7¡¾3.4, respectively, P<0.02,). In addition,
hMUC1-loaded CD11+cells in the phMUC1+I-131 group were found to be most
effective at generating hMUC1-associated CD8+IFN¥ã+ T cells (pcDNA+PBS,
phMUC1+PBS, pcDNA+I-131, and phMUC1+I-131, 7¡¾3, 31¡¾9, 86¡¾19, and 139¡¾15
cells, respectively, P<0.02,). Moreover, hMUC1-associated cytotoxic T cells
(CTLs) activities against CMNF cell were higher in the phMUC1+I-131 group than
in the other three groups (pcDNA+PBS, phMUC1+PBS, pcDNA+I-131, and phMUC1+I-131,
16¡¾2%, 20¡¾1%, 30¡¾2%, and 60¡¾2%, respectively, P<0.02). In the tumor rechallenge experiment, hMUC1-associated cytotoxic T cells (CTLs) activities
against CMNF cells were higher in the phMUC1+I-131 group than in the pcDNA+I-131
group (96¡¾5% and 8¡¾2%, respectively, P<0.001).
Conclusion: phMUC1 plus I-131 combination therapy generates more hMUC1associated immune response and synergistically induces a tumoricidal effect
against hMUC1 expressing cancers. Furthermore, the results of the present study
suggest that this combination therapy prevents tumor recurrence after complete
remission has been achieved
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