Faculty Collaboration Database

Transfer of the human glucocerebrosidase gene into hematopoietic stem cells of nonablated recipients: successful engraftment and long-term expression of the transgene. Blood 1995 Aug 01;86(3):1218-27

Date

08/01/1995

Pubmed ID

7620175

DOI

10.1182/blood.v86.3.1218.1218

Scopus ID

2-s2.0-0029152181 (requires institutional sign-in at Scopus site)   43 Citations

Abstract

In trying to develop methods of gene therapy for Gaucher disease that will avoid the morbidity and mortality associated with bone marrow (BM) ablation, we transplanted BM stem cells transduced with a retroviral vector containing the human glucocerebrosidase cDNA into normal, nonablated, syngeneic mice. Donor BM from untreated male mice or treated with 5-fluorouracil (5-FU) was transduced ex vivo using a standard 4-day transduction protocol. Recipient female mice were injected one time only or once daily for 5 consecutive days or once a week for 5 consecutive weeks using 2 x 10(7) (untreated BM) or 2 x 10(6) (5-FU-treated BM) cells per injection. Initial transduction efficiency into colony-forming unit-spleen (CFU-S) was 80% to 100%. Recipient analysis was performed at least 6 months after the last transplantation. The best engraftment of donor stem cells, up to 5% by secondary CFU-S analysis, was obtained with multiple injections of transduced BM not previously treated with 5-FU. Polymerase chain reaction (PCR) amplification for both the transgene and the Y chromosome identified the progeny of transduced stem cells in various hematopoietic and non-hematopoietic organs. The copy number of the transgene in stem cells was 0.13 to 2.8. Transgene expression was shown by reverse transcriptase-PCR, in situ hybridization, and immunohistochemistry. No serious side effects of the procedure were noted. We conclude that multiple transplants of retrovirally transduced BM cells into nonablated recipients may be a safe and effective therapeutic modality for a number of genetic hematopoietic disorders.

Author List

Schiffmann R, Medin JA, Ward JM, Stahl S, Cottler-Fox M, Karlsson S

Author

Jeffrey A. Medin PhD Professor in the Pediatrics department at Medical College of Wisconsin

MESH terms used to index this publication - Major topics in bold

Animals
Base Sequence
Bone Marrow
DNA Primers
Female
Gene Transfer Techniques
Genes
Glucosylceramidase
Hematopoietic Stem Cell Transplantation
Hematopoietic Stem Cells
Humans
In Situ Hybridization
Male
Mice
Mice, Inbred BALB C
Molecular Sequence Data
Transduction, Genetic