Transient in vivo selection of transduced peripheral blood cells using antifolate drug selection in rhesus macaques that received transplants with hematopoietic stem cells expressing dihydrofolate reductase vectors

Abstract

One of the main obstacles for effective human gene therapy for hematopoietic disorders remains the achievement of an adequate number of genetically corrected blood cells. One approach to this goal is to incorporate drug resistance genes into vectors to enable in vivo selection of hematopoietic stem cells (HSCs). Although a number of drug resistance vectors enable HSC selection in murine systems, little is known about these systems in large animal models. To address this issue, we transplanted cells transduced with dihydrofolate resistance vectors into 6 rhesus macaques and studied whether selection of vector-expressing cells occurred following drug treatment with trimetrexate and nitrobenzylmercaptopurineriboside-phosphate. In some of the 10 administered drug treatment courses, substantial increases in the levels of transduced peripheral blood cells were noted; however, numbers returned to baseline levels within 17 days. Attempts to induce stem cell cycling with stem cell factor and granulocyte-colony stimulating factor prior to drug treatment did not lead to sustained enrichment for transduced cells. These data highlight an important species-specific difference between murine and nonhuman primate models for assessing in vivo HSC selection strategies and emphasize the importance of using drugs capable of inducing selective pressure at the level of HSCs.

Improved transduction of human sheep repopulating cells by retrovirus vectors pseudotyped with feline leukemia virus type C or RD114 envelopes

“…This can be accomplished by either improved HSC transduction or the modification of HSCs with drug-selectable genes or growth regulators. [59][60][61][62][63][64] Our observations about superior transduction with RD114-pseudotyped retrovirus particles are consistent with previous data in canine, nonhuman primate, and NOD-SCID mouse xenograft model systems. [40][41][42][43][44][45] Our data are the first demonstration of transduction of primary hematopoietic cells with recombinant FeLV-C retrovirus particles.…”

Methylguanine DNA Methyltransferase-Mediated Drug Resistance-Based Selective Enrichment and Engraftment of Transplanted Stem Cells in Skeletal Muscle

“…Strategies for selective enrichment of donor hematopoietic stem cells to enhance engraftment were initially developed as a treatment for myelosuppression. There are many studies which have investigated various drug resistance-mediated in vivo selection enrichment strategies [7][8][9][10][11][12][13] among which, the most effective strategy uses O 6 -methylguanine-DNA-methyltransferase (MGMT) (P140K) gene-mediated drug resistance [14][15][16][17][18][19]. MGMT (P140K) is a mutant form of the drug resistance gene MGMT.…”

Gene therapy with drug resistance genes

“…While successful in the murine model, the same approach 27 was not able to achieve stable in vivo selection with the DHFR in a non-human primate model. 40 The explanation given for this lack of efficacy is that large animals have fewer cycling HSCs than mice.…”