Consistent, Persistent Expression from Modified Retroviral Vectors in Murine Hematopoietic Stem Cells

Retroviral vectors based on the Moloney murine leukemia virus (MoMuLV) have shown inconsistent levels and duration of expression as well as a propensity for the acquisition of de novo methylation in vivo. MoMuLV-based vectors are known to contain sequences that are capable of suppressing or preventi...

Full description

Saved in:
Bibliographic Details
Published in:Proceedings of the National Academy of Sciences - PNAS 1998-08, Vol.95 (17), p.10182-10187
Main Authors: Robbins, Paul B., Skelton, Dianne C., Yu, Xiao-Jin, Halene, Stephanie, Leonard, Earl H., Kohn, Donald B.
Format: Article
Language:English
Subjects:
Animals
Base Sequence
Biological Sciences
Bone marrow
Bone Marrow Transplantation
Colony-Forming Units Assay
DNA
DNA Methylation
DNA Primers - genetics
DNA, Recombinant - genetics
DNA, Recombinant - metabolism
Female
Founder Effect
Gene Expression
Gene Transfer Techniques
Genetic Vectors
Hematopoietic stem cells
Hematopoietic Stem Cells - cytology
Hematopoietic Stem Cells - metabolism
Leukemia
Male
Methylation
Mice
Mice, Inbred C57BL
Microbiology
Moloney murine leukemia virus - genetics
Repetitive Sequences, Nucleic Acid
Retroviral vectors
Retroviridae - genetics
Spleen
Stem cells
Transduction, Genetic
Transplantation
Transplantation, Isogeneic
Virus Integration - genetics
Citations: Items that this one cites
Items that cite this one
Online Access:Get full text
Tags: Add Tag
No Tags, Be the first to tag this record!
Description
Summary:Retroviral vectors based on the Moloney murine leukemia virus (MoMuLV) have shown inconsistent levels and duration of expression as well as a propensity for the acquisition of de novo methylation in vivo. MoMuLV-based vectors are known to contain sequences that are capable of suppressing or preventing expression from the long terminal repeat. Previously, we constructed a series of modified retroviral vectors and showed that they function significantly better than MoMuLV-based vectors in vitro. To test the efficacy of the modified vectors in hematopoietic stem cells in vivo, we examined gene expression and proviral methylation in differentiated hematopoietic colonies formed in the spleens of mice after serial transplantation with transduced bone marrow (2 degrees CFU-S). We found a significant increase in the frequency of expression with our modified vectors (>90% expression in vector DNA containing 2 degrees CFU-S) over the frequency observed with the standard MoMuLV-based vector (28% expression in vector containing 2 degrees CFU-S). Expression from the modified vectors was highly consistent, with expression in >50% of the vector-containing 2 degrees CFU-S from all 20 transplant recipients analyzed, whereas expression from the standard MoMuLV-based vector was inconsistent, with expression in 0-10% of the vector containing 2 degrees CFU-S from 8 recipients and expression in >50% of the vector-containing 2 degrees CFU-S from 4 other recipients. In addition, we established that the modified vectors had a lower level of DNA methylation than the control vector. These findings represent significant advances in the development and evaluation of effective retroviral vectors for application in vivo.
ISSN:0027-8424
1091-6490
DOI:10.1073/pnas.95.17.10182