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Optimized adeno-associated virus (AAV)-protein phosphatase-5 helper viruses for efficient liver transduction by single-stranded AAV vectors: therapeutic expression of factor IX at reduced vector doses.

Abstract：

:Abstract Our studies have shown that coinjection of conventional single-stranded adeno-associated virus 2 (ssAAV2) vectors carrying the enhanced green fluorescent protein (EGFP) gene with self-complementary (sc) AAV2-T cell protein tyrosine phosphatase (TC-PTP) and scAAV2-protein phosphatase-5 (PP5) vectors resulted in an approximately 16-fold increase in EGFP expression in primary murine hepatocytes in vivo [Jayandharan, G.R., Zhong, L., Li, B., Kachniarz, B., and Srivastava, A. (2008). Gene Ther. 15, 1287-1293]. In the present studies, this strategy was further optimized to achieve transgene expression at reduced vector/helper virus doses. These included the use of scAAV helper viruses containing (1) hepatocyte-specific promoters, (2) tyrosine-mutant AAV2 capsids, and (3) additional AAV serotype vectors known to efficiently transduce hepatocytes. The hepatocyte-specific transthyretin (TTR) promoter was approximately 6- to 7-fold more efficient than the Rous sarcoma virus (RSV) promoter; tyrosine-mutant AAV2 capsids were approximately 6- to 11-fold more efficient than the wild-type AAV2 capsids; and the AAV8 serotype helper virus was approximately 16-fold more efficient than AAV2 serotype helper virus. With these modifications, the vector dose of the helper virus could be further reduced by approximately 50-fold. Last, coadministration of scAAV8-PP5 helper virus increased coagulation factor IX expression from an ssAAV2 vector by approximately 7- to 10-fold, thereby achieving therapeutic levels at lower vector doses. No adverse effect on hepatocytes was observed under any of these experimental conditions. The strategy presented here should be adaptable to any ssAAV transgene cassette and, specifically, liver-directed applications of ssAAV2 vectors containing larger genes that cannot be encapsidated in scAAV vectors.

journal_name

Hum Gene Ther

journal_title

Human gene therapy

authors

Jayandharan GR,Zhong L,Sack BK,Rivers AE,Li M,Li B,Herzog RW,Srivastava A

doi

10.1089/hum.2009.100

subject

Has Abstract

pub_date

2010-03-01 00:00:00

pages

271-83

issue

eissn

1043-0342

issn

1557-7422

journal_volume

pub_type

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Optimized adeno-associated virus (AAV)-protein phosphatase-5 helper viruses for efficient liver transduction by single-stranded AAV vectors: therapeutic expression of factor IX at reduced vector doses.