| Autor: |
J H, Slingsby, D, Baban, J, Sutton, M, Esapa, T, Price, S M, Kingsman, A J, Kingsman, A, Slade |
| Rok vydání: |
2000 |
| Předmět: |
|
| Zdroj: |
Human gene therapy. 11(10) |
| ISSN: |
1043-0342 |
| Popis: |
A number of stable producer cell lines for high-titer Mo-MuLV vectors have been constructed. Development has previously centered on increasing end-point titers by producing maximal levels of Mo-MuLV Gag/Pol, envelope glycoproteins, and retroviral RNA genomes. We describe the production yields and transduction efficiency characteristics of two Mo-MuLV packaging cell lines, FLYA13 and TEFLYA. Although they both produce 4070A-pseudotyped retroviral vectors reproducibly at1 x 10(6) LFU ml(-1), the transduction efficiency of unconcentrated and concentrated virus from FLYA13 lines is poor compared with vector preparations from TEFLYA lines. A powerful inhibitor of retroviral transduction is secreted by FLYA13 packaging cells. We show that the inhibitory factor does not affect transduction of target cells by RD114-pseudotyped vectors. This suggests that the inhibitory factor functions at the level of envelope-receptor interactions. Phosphate starvation of target cells shows a two-fold increase in Pit2 receptor mRNA and causes some improvement in FLYA13 virus transduction efficiency. Western blots show that FLYA13 viral samples contain an eight-fold higher ratio of 4070A envelope to p30gag than that of virus produced by TEFLYA producer cell lines. This study correlates overexpression of 4070A envelope glycoprotein in retroviral preparations with a reduction of transduction efficiency at high multiplicities of infection. We suggest that TEFLYA packaging cells express preferable levels of 4070A compared with FLYA13, which not only enables high-titer stocks to be generated, but also facilitates a high efficiency of transduction of target cells. |
| Databáze: |
OpenAIRE |
| Externí odkaz: |
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