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Hybridoma 1995-Apr

p53 trans-dominantly suppresses tumor formation of human breast cancer cells mediated by retroviral bulk infection without marker gene selection: an expeditious in vitro protocol with implications towards gene therapy.

Només els usuaris registrats poden traduir articles
Inicieu sessió / registreu-vos
L'enllaç es desa al porta-retalls
I B Runnebaum
R Kreienberg

Paraules clau

Resum

Studies on the molecular basis of human breast cancer have demonstrated that mutational inactivation of the p53 tumor suppressor gene may be an essential step in the development of this cancer. We and others have previously shown that transfer of the wild-type p53 gene into cultured breast cancer cells reduced their malignant potential. We report here on a p53 gene transfer protocol based on a replication-incompetent retrovirus to efficiently inhibit tumor formation of cancer cells with endogenous mutant p53. The susceptibility of the cells to retroviral infection was determined with LZRNL transducing the lacZ reporter gene. A multiplicity of infection (moi) of 2 resulted in 90% of the exposed cell population in cytochemically detectable beta-galactosidase activity. Using the p53 vector Lhp53RNL with a moi of 2 was sufficient to completely supress tumor formation by the highly tumorigenic MDAMB231 breast cancer cells carrying a point missense mutation in codon 280. Even after 12 weeks, no vital tumors were histologically detectable. For comparison, established protocols were used to infect MDAMB231 cells with low moi with the p53 virus. Clones were expanded in G418-selective media for few weeks, pooled and injected into nude mice. Tumor formation occurred already after 1 week from G418-selected cells. Long-term expression of the p53 transgene was more stable in retrovirally bulk-infected and nonselected cells resulting in an efficient suppression of tumor formation. This approach may facilitate future studies on other growth suppressive genes that potentially qualify for in vivo gene therapy.

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