Successful expansion but not complete restriction of tropism of adeno-associated virus by in vivo biopanning of random virus display peptide libraries

TY - JOUR

T1 - Successful expansion but not complete restriction of tropism of adeno-associated virus by in vivo biopanning of random virus display peptide libraries

AU - Michelfelder, Stefan

AU - Kohlschütter, Johannes

AU - Skorupa, Alexandra

AU - Pfennings, Sabrina

AU - Müller, Oliver

AU - Kleinschmidt, Jürgen A

AU - Trepel, Martin

PY - 2009

Y1 - 2009

N2 - Targeting viral vectors to certain tissues in vivo has been a major challenge in gene therapy. Cell type-directed vector capsids can be selected from random peptide libraries displayed on viral capsids in vitro but so far this system could not easily be translated to in vivo applications. Using a novel, PCR-based amplification protocol for peptide libraries displayed on adeno-associated virus (AAV), we selected vectors for optimized transduction of primary tumor cells in vitro. However, these vectors were not suitable for transduction of the same target cells under in vivo conditions. We therefore performed selections of AAV peptide libraries in vivo in living animals after intravenous administration using tumor and lung tissue as prototype targets. Analysis of peptide sequences of AAV clones after several rounds of selection yielded distinct sequence motifs for both tissues. The selected clones indeed conferred gene expression in the target tissue while gene expression was undetectable in animals injected with control vectors. However, all of the vectors selected for tumor transduction also transduced heart tissue and the vectors selected for lung transduction also transduced a number of other tissues, particularly and invariably the heart. This suggests that modification of the heparin binding motif by target-binding peptide insertion is necessary but not sufficient to achieve tissue-specific transgene expression. While the approach presented here does not yield vectors whose expression is confined to one target tissue, it is a useful tool for in vivo tissue transduction when expression in tissues other than the primary target is uncritical.

AB - Targeting viral vectors to certain tissues in vivo has been a major challenge in gene therapy. Cell type-directed vector capsids can be selected from random peptide libraries displayed on viral capsids in vitro but so far this system could not easily be translated to in vivo applications. Using a novel, PCR-based amplification protocol for peptide libraries displayed on adeno-associated virus (AAV), we selected vectors for optimized transduction of primary tumor cells in vitro. However, these vectors were not suitable for transduction of the same target cells under in vivo conditions. We therefore performed selections of AAV peptide libraries in vivo in living animals after intravenous administration using tumor and lung tissue as prototype targets. Analysis of peptide sequences of AAV clones after several rounds of selection yielded distinct sequence motifs for both tissues. The selected clones indeed conferred gene expression in the target tissue while gene expression was undetectable in animals injected with control vectors. However, all of the vectors selected for tumor transduction also transduced heart tissue and the vectors selected for lung transduction also transduced a number of other tissues, particularly and invariably the heart. This suggests that modification of the heparin binding motif by target-binding peptide insertion is necessary but not sufficient to achieve tissue-specific transgene expression. While the approach presented here does not yield vectors whose expression is confined to one target tissue, it is a useful tool for in vivo tissue transduction when expression in tissues other than the primary target is uncritical.

KW - Amino Acid Sequence

KW - Animals

KW - Breast Neoplasms

KW - Capsid

KW - Dependovirus

KW - Female

KW - Gene Transfer Techniques

KW - Genetic Therapy

KW - Genetic Vectors

KW - Humans

KW - Mice

KW - Mice, Transgenic

KW - Molecular Sequence Data

KW - Peptide Library

KW - Polymerase Chain Reaction

KW - Transduction, Genetic

KW - Tropism

KW - Tumor Cells, Cultured

KW - Evaluation Studies

KW - Journal Article

KW - Research Support, Non-U.S. Gov't

U2 - 10.1371/journal.pone.0005122

DO - 10.1371/journal.pone.0005122

M3 - SCORING: Journal article

C2 - 19357785

VL - 4

SP - e5122

JO - PLOS ONE

JF - PLOS ONE

SN - 1932-6203

IS - 4

ER -