Cancer gene therapy using a survivin mutant adenovirus
Mehdi Mesri, … , Richard W. Kim, Dario C. Altieri
Mehdi Mesri, … , Richard W. Kim, Dario C. Altieri
Published October 1, 2001
Citation Information: J Clin Invest. 2001;108(7):981-990. https://doi.org/10.1172/JCI12983.
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Cancer gene therapy using a survivin mutant adenovirus

Abstract

We have constructed a replication-deficient adenovirus encoding a nonphosphorylatable Thr34→Ala mutant of the apoptosis inhibitor survivin (pAd-T34A) to target tumor cell viability in vitro and in vivo. Infection with pAd-T34A caused spontaneous apoptosis in cell lines of breast, cervical, prostate, lung, and colorectal cancer. In contrast, pAd-T34A did not affect cell viability of proliferating normal human cells, including fibroblasts, endothelium, or smooth muscle cells. Infection of tumor cells with pAd-T34A resulted in cytochrome c release from mitochondria, cleavage of approximately 46-kDa upstream caspase-9, processing of caspase-3 to the active subunits of approximately 17 and 19 kDa, and increased caspase-3 catalytic activity. When compared with chemotherapeutic regimens, pAd-T34A was as effective as taxol and considerably more effective than adriamycin in induction of tumor cell apoptosis and enhanced taxol-induced cell death. In three xenograft breast cancer models in immunodeficient mice, pAd-T34A suppressed de novo tumor formation, inhibited by approximately 40% the growth of established tumors, and reduced intraperitoneal tumor dissemination. Tumors injected with pAd-T34A exhibited loss of proliferating cells and massive apoptosis by in situ internucleosomal DNA fragmentation. These data suggest that adenoviral targeting of the survivin pathway may provide a novel approach for selective cancer gene therapy.

Authors

Mehdi Mesri, Nathan R. Wall, Jia Li, Richard W. Kim, Dario C. Altieri

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Figure 1

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Construction and expression of pAd survivin vectors. (a) Map of pAdTrack...
Construction and expression of pAd survivin vectors. (a) Map of pAdTrack CMV and pAdEasy vectors. (b) GFP expression in transduced cultures. HeLa cells were infected with the indicated pAd vectors at moi of 50 for 8 hours, harvested after 48 hours, and analyzed by fluorescence microscopy. (c) Absence of replication-competent adenoviral particles. HeLa cells (8 × 104) were infected with pAd-T34A or pAd-GFP at moi of 1,250 and grown for 3 days at 37°C. Cell extracts were used to successively infect a second HeLa cell culture, and cells were analyzed by phase-contrast microscopy (Phase) or GFP expression (GFP) after an additional 2-day period. (d) Western blot analysis. Aliquots of HeLa or MCF-7 cells were infected with the indicated pAd vectors at moi of 50, harvested after 48 hours at 37°C, and protein-normalized extracts were analyzed by Western blotting with an Ab to survivin, XIAP, or control β-actin followed by chemiluminescence. Molecular-weight markers in kilodaltons are shown on the left. LITR, left-hand inverted terminal repeat; MW, molecular weight.