Ribosomal DNA integrating rAAV-rDNA vectors allow for stable transgene expression

Leszek Lisowski, Ashley Lau, Zhongya Wang, Yue Zhang, Feijie Zhang, Markus Grompe, Mark A. Kay

Research output: Contribution to journalArticle

22 Citations (Scopus)

Abstract

Although recombinant adeno-associated virus (rAAV) vectors are proving to be efficacious in clinical trials, the episomal character of the delivered transgene restricts their effectiveness to use in quiescent tissues, and may not provide lifelong expression. In contrast, integrating vectors enhance the risk of insertional mutagenesis. In an attempt to overcome both of these limitations, we created new rAAV-rDNA vectors, with an expression cassette flanked by ribosomal DNA (rDNA) sequences capable of homologous recombination into genomic rDNA. We show that after in vivo delivery the rAAV-rDNA vectors integrated into the genomic rDNA locus 8-13 times more frequently than control vectors, providing an estimate that 23-39% of the integrations were specific to the rDNA locus. Moreover, a rAAV-rDNA vector containing a human factor IX (hFIX) expression cassette resulted in sustained therapeutic levels of serum hFIX even after repeated manipulations to induce liver regeneration. Because of the relative safety of integration in the rDNA locus, these vectors expand the usage of rAAV for therapeutics requiring long-term gene transfer into dividing cells.

Original languageEnglish (US)
Pages (from-to)1912-1923
Number of pages12
JournalMolecular Therapy
Volume20
Issue number10
DOIs
StatePublished - Oct 2012

Fingerprint

Dependovirus
Ribosomal DNA
Transgenes
Factor IX
Liver Regeneration
Insertional Mutagenesis
Homologous Recombination
Clinical Trials
Safety
Therapeutics
Serum

ASJC Scopus subject areas

  • Molecular Biology
  • Molecular Medicine
  • Genetics
  • Drug Discovery
  • Pharmacology

Cite this

Lisowski, L., Lau, A., Wang, Z., Zhang, Y., Zhang, F., Grompe, M., & Kay, M. A. (2012). Ribosomal DNA integrating rAAV-rDNA vectors allow for stable transgene expression. Molecular Therapy, 20(10), 1912-1923. https://doi.org/10.1038/mt.2012.164

Ribosomal DNA integrating rAAV-rDNA vectors allow for stable transgene expression. / Lisowski, Leszek; Lau, Ashley; Wang, Zhongya; Zhang, Yue; Zhang, Feijie; Grompe, Markus; Kay, Mark A.

In: Molecular Therapy, Vol. 20, No. 10, 10.2012, p. 1912-1923.

Research output: Contribution to journalArticle

Lisowski, L, Lau, A, Wang, Z, Zhang, Y, Zhang, F, Grompe, M & Kay, MA 2012, 'Ribosomal DNA integrating rAAV-rDNA vectors allow for stable transgene expression', Molecular Therapy, vol. 20, no. 10, pp. 1912-1923. https://doi.org/10.1038/mt.2012.164
Lisowski, Leszek ; Lau, Ashley ; Wang, Zhongya ; Zhang, Yue ; Zhang, Feijie ; Grompe, Markus ; Kay, Mark A. / Ribosomal DNA integrating rAAV-rDNA vectors allow for stable transgene expression. In: Molecular Therapy. 2012 ; Vol. 20, No. 10. pp. 1912-1923.
@article{674ce2f9567d4b8b83f947c682d41e8d,
title = "Ribosomal DNA integrating rAAV-rDNA vectors allow for stable transgene expression",
abstract = "Although recombinant adeno-associated virus (rAAV) vectors are proving to be efficacious in clinical trials, the episomal character of the delivered transgene restricts their effectiveness to use in quiescent tissues, and may not provide lifelong expression. In contrast, integrating vectors enhance the risk of insertional mutagenesis. In an attempt to overcome both of these limitations, we created new rAAV-rDNA vectors, with an expression cassette flanked by ribosomal DNA (rDNA) sequences capable of homologous recombination into genomic rDNA. We show that after in vivo delivery the rAAV-rDNA vectors integrated into the genomic rDNA locus 8-13 times more frequently than control vectors, providing an estimate that 23-39{\%} of the integrations were specific to the rDNA locus. Moreover, a rAAV-rDNA vector containing a human factor IX (hFIX) expression cassette resulted in sustained therapeutic levels of serum hFIX even after repeated manipulations to induce liver regeneration. Because of the relative safety of integration in the rDNA locus, these vectors expand the usage of rAAV for therapeutics requiring long-term gene transfer into dividing cells.",
author = "Leszek Lisowski and Ashley Lau and Zhongya Wang and Yue Zhang and Feijie Zhang and Markus Grompe and Kay, {Mark A.}",
year = "2012",
month = "10",
doi = "10.1038/mt.2012.164",
language = "English (US)",
volume = "20",
pages = "1912--1923",
journal = "Molecular Therapy",
issn = "1525-0016",
publisher = "Nature Publishing Group",
number = "10",

}

TY - JOUR

T1 - Ribosomal DNA integrating rAAV-rDNA vectors allow for stable transgene expression

AU - Lisowski, Leszek

AU - Lau, Ashley

AU - Wang, Zhongya

AU - Zhang, Yue

AU - Zhang, Feijie

AU - Grompe, Markus

AU - Kay, Mark A.

PY - 2012/10

Y1 - 2012/10

N2 - Although recombinant adeno-associated virus (rAAV) vectors are proving to be efficacious in clinical trials, the episomal character of the delivered transgene restricts their effectiveness to use in quiescent tissues, and may not provide lifelong expression. In contrast, integrating vectors enhance the risk of insertional mutagenesis. In an attempt to overcome both of these limitations, we created new rAAV-rDNA vectors, with an expression cassette flanked by ribosomal DNA (rDNA) sequences capable of homologous recombination into genomic rDNA. We show that after in vivo delivery the rAAV-rDNA vectors integrated into the genomic rDNA locus 8-13 times more frequently than control vectors, providing an estimate that 23-39% of the integrations were specific to the rDNA locus. Moreover, a rAAV-rDNA vector containing a human factor IX (hFIX) expression cassette resulted in sustained therapeutic levels of serum hFIX even after repeated manipulations to induce liver regeneration. Because of the relative safety of integration in the rDNA locus, these vectors expand the usage of rAAV for therapeutics requiring long-term gene transfer into dividing cells.

AB - Although recombinant adeno-associated virus (rAAV) vectors are proving to be efficacious in clinical trials, the episomal character of the delivered transgene restricts their effectiveness to use in quiescent tissues, and may not provide lifelong expression. In contrast, integrating vectors enhance the risk of insertional mutagenesis. In an attempt to overcome both of these limitations, we created new rAAV-rDNA vectors, with an expression cassette flanked by ribosomal DNA (rDNA) sequences capable of homologous recombination into genomic rDNA. We show that after in vivo delivery the rAAV-rDNA vectors integrated into the genomic rDNA locus 8-13 times more frequently than control vectors, providing an estimate that 23-39% of the integrations were specific to the rDNA locus. Moreover, a rAAV-rDNA vector containing a human factor IX (hFIX) expression cassette resulted in sustained therapeutic levels of serum hFIX even after repeated manipulations to induce liver regeneration. Because of the relative safety of integration in the rDNA locus, these vectors expand the usage of rAAV for therapeutics requiring long-term gene transfer into dividing cells.

UR - http://www.scopus.com/inward/record.url?scp=84866973619&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=84866973619&partnerID=8YFLogxK

U2 - 10.1038/mt.2012.164

DO - 10.1038/mt.2012.164

M3 - Article

C2 - 22990671

AN - SCOPUS:84866973619

VL - 20

SP - 1912

EP - 1923

JO - Molecular Therapy

JF - Molecular Therapy

SN - 1525-0016

IS - 10

ER -