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  • Title: Retrovirus vectors designed for efficient transduction of cytotoxic or cytostatic genes.
    Author: Ui M, Takada M, Arai T, Matsumoto K, Yamada K, Nakahata T, Nishiwaki T, Furukawa Y, Tokino T, Nakamura Y, Iba H.
    Journal: Gene Ther; 1999 Oct; 6(10):1670-8. PubMed ID: 10516715.
    Abstract:
    It is difficult to establish stable packaging cell lines producing retrovirus vectors for the expression of anti-oncogenes with cytotoxic or cytostatic potential, because these genes would also affect the growth of the packaging cell lines. To overcome this problem, we designed a transcriptional unit pBabeLPL for vector RNA production, in which the transcription of the exogenous genes is completely suppressed by the presence of a preceding insertion containing the puromycin resistance gene (puro) and a poly(A) addition signal. This insertion is flanked by a tandem pair of loxP, and is designed to be excised after the introduction of Cre recombinase, when transcription of the exogenous gene will be started from the 5'-LTR. The transcriptional unit car- rying LacZ or p53 as the exogenous gene was introduced into a previously constructed prepackaging cell lines PtG-S2, in which the expression of VSV-G is also designed to be initiated by the introduction of Cre recombinase, while the gag-pol gene is expressed continuously. After the introduction of Cre recombinase by an adenovirus vector, LacZ- or p53-expressing VSV-G-pseudotyped retrovirus vectors with the designed structure were produced at high virus titers. The p53 virus was shown to be able to transduce p53 into the entire population of several human cancer cell lines and to induce their growth arrest at the G1 phase, indicating that this vector-producing system will be advantageous for human gene therapy.
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