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P Hoogerbrugge

Publications and source records attributed to P Hoogerbrugge.

5 recordsLinked to original sources
PubMed1999

In vivo methotrexate selection of murine hemopoietic cells transduced with a retroviral vector for Gaucher disease.

The studies described were performed to investigate whether in vivo selection of retrovirus-transduced hemopoietic cells is feasible starting from a low percentage of transduced hemopoietic stem cells (PHSCs). The vector used is an amphotropic bicistronic retroviral vector carrying a cDNA for human lysosomal glucocerebrosidase (hGC) for treatment of Gaucher disease and a methotrexate (MTX) resistant mutant cDNA encoding human dihydrofolate reductase (DHFR). We tested the effect of MTX selection in mice that were either myeloablated or not before infusion of transduced cells. In addition, we determined whether repeated administration of transduced bone marrow cells has an additional effect on the percentage of hGC expressing cells. The results obtained have shown that, in myeloablated mice transplanted once with transduced bone marrow and treated twice weekly with 10 mg/kg of MTX for a total of 6 months, a two- to three-fold increased numbers of hGC expressing cells could be detected in both peripheral blood and bone marrow as compared with non-MTX treated mice. In mice transplanted with transduced bone marrow once every 2 weeks for a total of four times, percentages of hGC expressing cells were not significantly increased as compared with mice transplanted once. In non-ablated mice neither MTX selection nor multiple infusions of transduced bone marrow resulted in detection of hGC expressing cells 6 months after transplantation, indicating that the success of in vivo selection using MTX is highly dependent on the ratio of transduced hemopoietic stem cells transplanted versus residing and untransduced stem cells.

Animals↗
PubMed1997

Retroviral stem cell gene therapy.

Long-term in vivo gene transfer studies in mice have shown that recombinant murine retroviruses are able to infect murine hemopoietic stem cells with high efficiency. Taken together the results indicated that the proviral structure was present at high frequency in circulating hemopoietic cells resulting in significant expression levels. Because of the success of these murine studies, it was believed that gene therapy would soon be applicable to treat a wide variety of congenital or acquired human diseases associated with the hemopoietic system. However, results from gene transfer studies in nonhuman primates and first human clinical trails have indicated that murine retrovirus infection of primate hemopoietic stem cells is inefficient. Although there are essential differences between the murine and primate gene therapy studies with respect to the recombinant viruses and transduction protocols used, these differences cannot solely account for the differences observed in infection efficiency. Therefore, in recent years effort has been spent on the identification of factors limiting retroviral transduction of primate hemopoietic stem cells. Increasing knowledge concerning hemopoiesis and retroviral infection has helped in identifying a number of limiting factors. Novel transduction strategies and tools have been generated which attempt to circumvent these limiting factors. These factors as well as the strategies that showed increased retroviral infection of primate hemopoietic stem cells will be discussed.

Animals↗
PubMed1997

Development of safe and efficient retroviral vectors for Gaucher disease.

We have generated amphotropic and Gibbon ape leukemia (GaLV) viruses carrying either a full-length (IG-GC2) or a shortened glucocerebrosidase cDNA (IG-GC4). For all recombinant retroviruses, a single infection was sufficient to augment glucocerebrosidase activity in unselected Gaucher type I and type II fibroblasts to levels which can be considered therapeutic. Transfer efficiency of the glucocerebrosidase cDNA into normal human and Gaucher type I CD34+ cells, using supernatant transduction, ranged from 4 to 50% as established on vector-positive CFU-GM. In these experiments, GaLV and amphotropic virus were equally efficient in transducing early human progenitors. Importantly, mixing amphotropic and GaLV pseudotyped retroviruses resulted in significantly higher transduction efficiencies as compared with single infections, up to 70% vector-positive CFU-GM. Glucocerebrosidase activity, measured in the progeny of human CD34+ cells, increased up to 460% compared with mock-infected CD34+ cells. Upon transduction of Gaucher CD34+ bone marrow cells the glucocerebrosidase deficiency was reversed.

Antigens, CD34↗