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Effect of deletion and the site of insertion in double copy anti-tat retroviral vectors: viral titres and production of anti-tat mRNA.

Abstract：

:In attempts to further develop murine leukemia virus (MLV) based retroviral vectors for gene therapy, we investigated vector production and antisense expression from retroviral constructs with U3 deletions or insertions. Promoter elements in the U3 region of the 3' LTR of the vector pLXSN were deleted and replaced with DNA encoding the HIV anti-tat gene under control of the tRNAmet promoter to produce a double copy self inactivating vector (DC-SIN). DC-SIN constructs were compared to vectors containing the anti-tat cassette inserted at 5 different sites of the U3 region (DC-insertions). Titres of DC-SIN and DC-insertion vectors were similar but approximately 10 fold lower than parental pLXSN. Cells transduced with DC-SIN and DC-insertion vectors all expressed anti-tat mRNA. Transcripts from the MLV-LTR were detected in cells transduced with DC-insertion but not DC-SIN vectors or a vector with the anti-tat cassette between CAAT and TATA boxes of the promoter, indicating inactivation of the viral promoter in the latter vectors. Cells transduced with constructs of either design showed comparable efficacy of protection against HIV challenge. Thus, no U3 insertion site was preferred for virus production. Insertion of a tRNA promoter between CAAT and TATA boxes and the DC-SIN design which would not introduce an active RNA pol II promoter into the genome are attractive for further development of safe gene therapy agents.

journal_name

Arch Virol

journal_title

Archives of virology

authors

Carr JM,Calvert JK,Kumar R,Burrel CJ,Li P

doi

10.1007/s007050170029

keywords:

subject

Has Abstract

pub_date

2001-01-01 00:00:00

pages

2191-200

issue

11

eissn

0304-8608

issn

1432-8798

journal_volume

146

pub_type

杂志文章

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