The goal of this study is to develop a novel and effective strategy for cancer gene therapy utilizing the Pseudomonas putida methioninase gene (METase) in combination with either: 1) the recombinant methioninase protein (rMETase), which itself has selective antitumor efficacy; 2) selenomethionine (SeMeth) as a prodrug analog cleaved by the METase gene product to toxic methylselenol. To achieve this goal, an adenoviral vector of the METase gene has been constructed (rAd-MBTase). Synergy between the rAd-METase and the recombinant methioninase protein (rMETase) was observed on human cells in vitro. The degree of synergy determined with the combination index (CI)isobologram method for all combinations of rMETase and rAd-METase tested resulted in CIs of less than 0.7, indicating strong synergy. rAd-METase was also combined with SeMeth as a prodrug. The rAd-METase infected cancer cells were up to 1000-fold more sensitive to SeMeth compared to uninfected cells due to the production of methylselenol by the METase gene transduced cells. The r-Ad-MET transduced cells also had a very strong bystander effect in the presence of SeMeth whereby only 3% of the cell population killed 80% or more of the remaining co-cultured cancer cells. The rAd-METase-SeMeth combination killed the cancer cells by an apoptotic mechanism. Based on the above results, we propose to develop METase gene therapy with the following specific aims: 1) Determine efficacy of adenoviral METase in a series of multiple types of human cancer compared to normal cells in vitro; 2) Determine optimal synergy of adenoviral MBTase in combination with rMETase in a series of human cancer compared to normal cells in vitro; 3) Determine optimal synergy of adenoviral METase in combination with SeMeth on a series of human cancer compared to normal cells in vitro; 4) Determine maximal bystander effect with adenoviral METase and SeMeth on human cancer cells in vitro; 5) Insert the tissue specific carcino-embryonic antigen (CEA) promoter in the rAd-METase vector. Selective efficacy will be determined against CEA-expressing tumor cells vs. other tumor and normal cells. The synergistic combination and prodrug approach should allow lower, and thereby safer, amounts of adenovirus to be used in vivo to avoid toxic effects recently observed in adenoviral-based gene therapy. Achievements of the specific aims of Phase I will enable realistic in vivo strategies of targeted METase gene therapy to be evaluated in Phase II.
Tumor targetable METase-gene vectors and prodrugs cleavable by the METase gene product will be developed in Phase II. A broad cancer market for these gene medicines is expected.
| Gupta, Anshu; Miki, Kenji; Xu, Mingxu et al. (2003) Combination efficacy of doxorubicin and adenoviral methioninase gene therapy with prodrug selenomethionine. Anticancer Res 23:1181-8 |
| Miki, K; Xu, M; Gupta, A et al. (2001) Methioninase cancer gene therapy with selenomethionine as suicide prodrug substrate. Cancer Res 61:6805-10 |
