Retroviral Vector-mediated Gene Transfer Research Articles

The use of retroviral vectors for gene therapy-what are the risks? A review of retroviral pathogenesis and its relevance to retroviral vector-mediated gene delivery

Retroviral vector-mediated gene transfer has been central to the development of gene therapy. Retroviruses have several distinct advantages over other vectors, especially when permanent gene transfer is the preferred outcome. The most important advantage that retroviral vectors offer is their ability to transform their single stranded RNA genome into a double stranded DNA molecule that stably integrates into the target cell genome. This means that retroviral vectors can be used to permanently modify the host cell nuclear genome. Recently, retroviral vector-mediated gene transfer, as well as the broader gene therapy field, has been re-invigorated with the development of a new class of retroviral vectors which are derived from lentiviruses. These have the unique ability amongst retroviruses of being able to infect non-cycling cells. Vectors derived from lentiviruses have provided a quantum leap in technology and seemingly offer the means to achieve significant levels of gene transfer in vivo.The ability of retroviruses to integrate into the host cell chromosome also raises the possibility of insertional mutagenesis and oncogene activation. Both these phenomena are well known in the interactions of certain types of wild-type retroviruses with their hosts. However, until recently they had not been observed in replication defective retroviral vector-mediated gene transfer, either in animal models or in clinical trials. This has meant the potential disadvantages of retroviral mediated gene therapy have, until recently, been seen as largely, if not entirely, hypothetical. The recent clinical trial of γc mediated gene therapy for X-linked severe combined immunodeficiency (X-SCID) has proven the potential of retroviral mediated gene transfer for the treatment of inherited metabolic disease. However, it has also illustrated the potential dangers involved, with 2 out of 10 patients developing T cell leukemia as a consequence of the treatment. A considered review of retroviral induced pathogenesis suggests these events were qualitatively, if not quantitatively, predictable. In addition, it is clear that the probability of such events can be greatly reduced by relatively simple vector modifications, such as the use of self-inactivating vectors and vectors derived from non-oncogenic retroviruses. However, these approaches remain to be fully developed and validated. This review also suggests that, in all likelihood, there are no other major retroviral pathogenetic mechanisms that are of general relevance to replication defective retroviral vectors. These are important conclusions as they suggest that, by careful design and engineering of retroviral vectors, we can continue to use this gene transfer technology with confidence.

Development and application of gene therapy technologies

The success of hematopoietic stem cell gene therapy for X-linked severe combined immunodeficiency (X-SCID) was a major breakthrough in the field of gene therapy. However, two patients treated with this gene therapy developed leukemia at a later time, and retroviral vector-mediated gene transfer was considered to trigger leukemogenesis; i.e. insertional mutagenesis caused activation of LMO 2 gene, which was one step toward leukemia development. To cope with this serious problem, basic studies are required to improve the safety of retroviral vectors and to develop the method for site-specific integration of transgenes. In addition, we have to develop technologies such as selective amplifier genes (SAGs), the system for selective expansion of transduced cells, in order to obtain therapeutic efficacy of hematopoietic stem cell gene therapy in many other disorders. Moreover, clinical applications of AAV vector are promising from the standpoint of safety issue, because this vector is derived from non-pathogenic virus. AAV vector is appropriate for gene transfer into neurons, muscles, and hepatocytes. For example, gene therapy for Parkinson's disease is investigated using AAV vectors. Genetic manipulation is also one of the indispensable technologies in the field of regeneration medicine, and further promotion of basic research is important.

Brain Malformations Caused by Retroviral Vector-Mediated Gene Transfer of Hippocampal Cholinergic Neurostimulating Peptide Precursor Protein into the CNS via Embryonic Mice Ventricles

Hippocampal cholinergic neurostimulating peptide precursor protein (HCNP-pp) is a unique multifunctional protein, being not only the precursor of HCNP, which promotes the phenotype development of septo-hippocampal cholinergic neurons, but also the binding protein of phosphatidylethanolamine, ATP, Raf-1 kinase (known as ‘Raf-1 kinase inhibitory factor’ in peripheral organs), and serine protease. We obtained a high-titer retroviral vector harboring HCNP-pp cDNA by the use of a modified packaging cell line and centrifugation, and by injecting it into embryonic mouse ventricles, we investigated the function of its gene product within the central nervous system (CNS). We found that efficient transduction into hippocampal pyramidal neurons can be achieved by injecting the vector into embryonic brain ventricles on embryonic day 14 (E14). Three days after receiving the intraventricular injection of the high-titer HCNP-pp retrovirus vector on E14, the tissues around the ventricles showed an overexpression of HCNP-pp. This was accompanied by a reduced amount of activated MEK and Erk (as analyzed by histochemical and Western blot methods), suggesting that HCNP-pp also regulates the MAP-kinase cascade within the CNS. Surprisingly, mouse brains that received the HCNP-pp retroviral vector showed massive malformation of the hippocampus and cerebellum when examined 30 days after birth. This shows that strictly regulated HCNP-pp gene expression is necessary for the normal development of the mouse brain, and that the moderate overexpression achieved by retroviral vector-mediated gene transfer is sufficient to cause severe abnormality of entire brain structures.

A simple and efficient method for constructing an adenoviral cDNA expression library

cDNA expression cloning is a powerful method for the identification of genes that are able to confer a selectable phenotype on specific cell types. An adenovirus vector is characterized by several advantages over plasmid DNA and retroviral vector-mediated gene transfer, such as broad host range and high infectivity. However, an expression cloning protocol using the adenovirus vector has not been reported. We describe here a simple and efficient method for constructing adenovirus cDNA expression libraries based on Cre-lox-mediated in vitro recombination between adenoviral shuttle plasmid cDNA libraries and adenoviral genomic DNA tagged with terminal protein. In a model experiment, EGFP clones present at the frequency of 0.003% in the shuttle plasmid library could be efficiently converted to adenoviral vector in a 6-cm dish under optimized conditions, indicating that high-complexity libraries harboring low-abundance cDNAs can be produced. The efficiency of this system was demonstrated by the isolation of cDNA for CD2 (frequency less than 1 in 0.3 × 104 transcripts in T cells) from the human T cells. This effective and versatile method can facilitate the functional identification of genes for a variety of purposes.

Performance- and safety-enhanced lentiviral vectors containing the human interferon-β scaffold attachment region and the chicken β-globin insulator

AbstractRetroviral vectors are the most efficient means of stable gene delivery to hematopoietic stem cells (HSCs). However, transgene expression from retroviral vectors is frequently subject to the negative influence of chromosomal sequences flanking the site of integration. Toward the development of autonomous transgene expression cassettes, we inserted the human interferon-β scaffold attachment region (IFN-SAR) and the chicken β-globin 5′ DNase I hypersensitive site 4 (5′HS4) insulator both separately and together into a series of self-inactivating (SIN) lentiviral vector backbones. Transduced cells of the human CD34+ hematopoietic progenitor line KG1a—pooled populations as well as individual clones harboring single integrants—were analyzed for reporter expression during culture periods of up to 4 months. Vectors carrying both the 5′HS4 insulator and the IFN-SAR consistently outperformed control vectors without inserts as well as vectors carrying either element alone. The performance of a set of vectors containing the murine stem cell virus long terminal repeat as an internal promoter was subsequently assessed during in vitro monocytic differentiation of transduced primary human CD34+ cord blood cells. Similar to what was observed in the KG1a hematopoietic progenitor cell model, optimal reporter expression in primary monocytes was obtained with the vector bearing both regulatory elements. These findings indicate that the 5′HS4/IFN-SAR combination is particularly effective at maintaining open chromatin domains permissive for high-level transgene expression at early and late stages of hematopoietic development, and thus could be of utility in HSC-directed retroviral vector–mediated gene transfer applications.

1062. Functional Correction of T Cells from Wiskott-Aldrich Syndrome Patients by Retroviral and Lentiviral Vector-Mediated Gene Transfer

1062. Functional Correction of T Cells from Wiskott-Aldrich Syndrome Patients by Retroviral and Lentiviral Vector-Mediated Gene Transfer

Retroviral vectors to study cell differentiation.

Retroviral vector-mediated gene transfer has been contributing to studies in developmental biology, genetics and clinical science. Retroviruses are ideal tools for gene transfer into dividing cells both in vitro and in vivo, which has led to their expanding use in developing systems. In this review we will primarily review their use for ectopic gene expression and lineage analysis to study cell differentiation in murine and chick embryos.

Multidrug Resistance 1 Gene Transfer Can Confer Chemoprotection to Human Peripheral Blood Progenitor Cells Engrafted in Immunodeficient Mice

Myelosuppression is the main side effect of cancer chemotherapy. An improved rate of retroviral vector-mediated gene transfer to hematopoietic stem cells, shown in more recent clinical trials, has created the basis to test the concept of myeloprotective gene therapy. We transplanted clinical-scale human peripheral blood progenitor cell grafts (n = 2) transduced with retroviral vector SF91m3, which contains the human multidrug resistance 1 gene (MDR1), into nonobese diabetic/severe combined immunodeficient (NOD/SCID) mice. Engrafted mice of one cohort were protected from paclitaxel toxicity (p < 0.05) and we noted a similar trend in the second cohort. In paclitaxel-treated mice that had received gene-transduced cells we found a significant increase in gene marking (p < 0.05 - p < 0.01) or P-glycoprotein expression (p < 0.01) compared with their chemotherapy-naive counterparts. This is the first report showing that cytostatic drug resistance gene therapy can mediate chemoprotection of human clinically relevant stem cell populations with marrow engraftment potential.

In utero transplantation of fetal liver cells in the mucopolysaccharidosis type VII mouse results in low-level chimerism, but overexpression of β-glucuronidase can delay onset of clinical signs

Mice with the lysosomal storage disease mucopolysaccharidosis (MPS) VII, caused by a deficiency of beta-glucuronidase (GUSB), have signs of disease present at birth. Bone marrow transplantation (BMT) or retroviral vector-mediated gene transfer into hematopoietic stem cells can partially correct the disease in adult mice, and BMT performed at birth results in a better clinical outcome. Thus, treatment in utero may result in further improvement. However, this must be done without cyto-ablation, and the donor cells do not have a competitive repopulating advantage over host cells. Transplantation in utero of either syngeneic fetal liver hematopoietic stem cells marked with a retroviral vector, or allogeneic donor cells that constitutively express high levels of human GUSB from a transgene, resulted in only about 0.1% engraftment in the adult. Immuno-affinity enrichment of stem and progenitor cells of 5- to 10-fold resulted in significantly higher GUSB activities at 2 months of age, but by 6 months engraftment was about 0.1%. Attempts to further increase the number of stem and progenitor cells were deleterious to the recipients. Nevertheless, GUSB expressed during the first 2 months of life in MPS VII fetuses could delay the onset of overt signs of disease. This suggests that the expression of some normal enzyme activity beginning in fetal life may offer the possibility of slowing the progression of the disease until more definitive postnatal transplantation or gene transfer to stem cells could be accomplished.

Current Status of Retroviral Vector Mediated Gene Transfer into Human Hematopoietic Stem Cells

Genetic modification of hematopoietic stem cells (HSCs) has been proposed as a treatment strategy for a variety of hematologic diseases, tracking marked cells or conferring resistance to chemotherapeutic agents. Despite early enthusiasm, the results of clinical studies involving gene transfer into HSCs have not resulted in therapeutic benefits for the vast majority of treated patients. This review describes the limitations and advances that have been made in the areas of gene transfer vectors, identification of the appropriate HSCs to target for genetic modifications and the methods used to perform gene transfer.

Current Status of Retroviral Vector Mediated Gene Transfer into Human Hematopoietic Stem Cells

Genetic modification of hematopoietic stem cells (HSCs) has been proposed as a treatment strategy for a variety of hematologic diseases, tracking marked cells or conferring resistance to chemotherapeutic agents. Despite early enthusiasm, the results of clinical studies involving gene transfer into HSCs has not resulted in therapeutic benefits for the vast majority of treated patients. This review describes the limitations and advances that have been made in the areas of gene transfer vectors, identification of the appropriate HSCs to target for genetic modifications and the methods used to perform gene transfer.

Delivery of a Retroviral Vector Expressing Human β-Glucuronidase to the Liver and Spleen Decreases Lysosomal Storage in Mucopolysaccharidosis VII Mice

Mucopolysaccharidosis VII (MPS VII) is caused by beta-glucuronidase (beta-gluc) deficiency and results in lysosomal storage due to the inability to degrade glycosaminoglycans. Transfer of a beta-gluc gene into the liver reduces hepatic pathology as well as storage in other organs via uptake of secreted protein. A Moloney murine leukemia-based retroviral vector expressing the human beta-gluc cDNA was injected intravascularly into MPS VII mice during hepatocyte replication, which was induced with im injection of an adenoviral vector that transiently expressed hepatocyte growth factor (Ad.CMV. HGF). This procedure resulted in transduction of approximately 1% of hepatocytes, 1% of normal liver enzyme activity, and a reduction in lysosomal storage in the liver at 3.5 months. Surprisingly, controls that received retroviral vector without HGF had transduction of nonparenchymal cells in the liver, significant levels of enzyme and RNA in the liver at 2 but not 3.5 months, and reduced lysosomal storage at 3.5 months. Transduction was also achieved in the replicating cells of the spleen, where lysosomal storage was reduced. An approach using a retroviral vector without a growth factor might temporarily reduce lysosomal storage in the liver and spleen in humans. Addition of HGF might be used to augment and prolong gene transfer.

Retroviral vector-mediated gene transfer into umbilical cord blood-derived megakaryocyte and platelet progenitors.

Optimizing platelet engraftment following hematopoietic stem cell transplantation is essential for minimizing transplant-related morbidity, particularly following umbilical cord blood transplantation (UCBT), where platelet engraftment frequently takes >60 days. One strategy for optimizing platelet engraftment following UCBT is to study or alter the genetic program of megakaryocyte/platelet (Mk/plt) progenitors. Retroviral vector gene transfer has previously proven useful for studying the biology of hematopoietic stem cells; however, procedures for transducing UCB cells of the Mk/plt lineage with retroviral vectors have not been described. We report here that Mk/plt progenitors generated from UCB progenitors can be efficiently transduced with retroviral vectors. Transduced Mk/plt cells were identified and quantitated by expression of a vector transgene encoding a truncated version of the human nerve growth factor receptor (NGFR). Vector-mediated NGFR expression could be readily detected in Mk/plt progenitors defined by immunophenotypic, morphologic, and functional criteria. In addition, NGFR expression persisted in mature anucleate platelets generated from the transduced Mk/plt progenitors. These methods may be useful for introducing genetic elements into Mk/plt progenitors to study various aspects of platelet development and biology and for marking ex vivo expanded Mk/plt progenitors to determine their contribution to engraftment.

Interleukin-2 gene transfer into human transitional cell carcinoma of the urinary bladder.

Transitional cell carcinoma of the bladder is one of the human cancers most responsive to immunotherapy, and local interleukin-2 (IL-2) production appears to be an important requirement for immunotherapy to be effective. In this study, we engineered two human bladder cancer cell lines (RT112 and EJ) to constitutively release human IL-2 by retroviral vector-mediated gene transfer. Following infection and selection, stable and consistent production of biologically active IL-2 was demonstrated at both the mRNA and the protein level. Morphology, in vitro growth rate and proliferation, as well as other cytokine gene mRNA or membrane adhesion receptor expression, were not altered in IL-2 transduced cells as compared to their parental or control vector-infected counterparts. Moreover, IL-2 engineered cells lost their tumorigenicity into nu/nu mice and the mechanism of rejection appeared to involve multiple host effector cell populations, among which a prominent role was played by neutrophils and radiosensitive cells. These findings may offer support to the development of an IL-2-based gene therapy approach to human bladder cancer. 1999 Cancer Research Campaign

The long internal loop of the alpha 3 subunit targets nAChRs to subdomains within individual synapses on neurons in vivo.

Different types of neurotransmitter receptors coexist within single neurons and must be targeted to discrete synaptic regions for proper function. In chick ciliary ganglion neurons, nicotinic acetylcholine receptors (nAChRs) containing alpha 3 and alpha 5 subunits are concentrated in the postsynaptic membrane, whereas alpha-bungarotoxin receptors composed of alpha 7 subunits are localized perisynaptically and excluded from the synapse. Using retroviral vector-mediated gene transfer in vivo, we show that the long cytoplasmic loop of alpha 3 targets chimeric alpha 7 subunits to the synapse and reduces endogenous nAChR surface levels, whereas the alpha 5 loop does neither. These results show that a particular domain of one subunit targets specific receptor subtypes to the interneuronal synapse in vivo. Moreover, our findings suggest a difference in the mechanisms that govern assembly of interneuronal synapses as compared to the neuromuscular junction in vertebrates.

Retroviral vector-mediated gene transfer into keratocytes in vitro and in vivo

PURPOSE: To determine the potential of somatic gene transfer as a technique for modulating corneal wound healing after superficial keratectomy. METHODS: The transduction of human and rabbit keratocytes with β-galactosidase and herpes simplex virus thymidine kinase genes was performed. In vitro, human and rabbit keratocytes were transduced with retroviral vectors bearing β-galactosidase or HStk (herpes simplex virus thymidine kinase) genes. In vivo, rabbit keratocytes were transduced by topical application of vector supernatant after a superficial keratectomy. In vitro and in vivo, expression of the β-galactosidase gene was examined with histochemical staining. In vitro, ganciclovir cytotoxicity in HStk gene-transduced keratocytes and bystander effect in co-cultures of HStk(+) and HStk(−) keratocytes were measured by determining the degree of confluency of cells in 6-well plates after 10 days of incubation. Corneal haze in rabbits was measured after transduction with Hstk and subsequent treatment with topical ganciclovir. RESULTS: In vitro, both human and rabbit keratocytes were transduced successfully with both β-galactosidase and HStk genes. Transduction efficiency was greater with human (22%) than with rabbit (16%) cells, and both HStk-transduced cell lines showed dose-dependent ganciclovir cytotoxicity and a significant bystander effect. In vivo, expression of β-galactosidase within vimentin-positive corneal stromal cells confirmed transduction of keratocytes in the rabbit after superficial stromal keratectomy with an efficiency of 25% to 40%. Postoperative application of topical ganciclovir reduced corneal stromal haze in rabbits. CONCLUSIONS: The ability to genetically transduce stromal keratocytes provides a new strategy for understanding the important cellular and molecular events that influence corneal wound healing, thus offering a potential approach to decrease or prevent corneal haze and scarring after superficial keratectomy.

Expression of human bone morphogenic protein 7 in primary rabbit periosteal cells: potential utility in gene therapy for osteochondral repair.

A commonly encountered problem in orthopedics is bone and cartilage tissue injury which heals incompletely or without full structural integrity. This necessitates development of improved methods for treatment of injuries which are not amenable to treatment using current therapies. An already large and growing number of growth factors which play significant roles in bone remodeling and repair have been identified in the past few years. It is well established that bone morphogenic proteins induce the production of new bone and cartilage. An efficient method of delivery of these growth factors by conventional pharmacological means has yet to be elucidated. We wished to evaluate the use of retroviral vector-mediated gene transfer to deliver genes of therapeutic relevance for bone and cartilage repair. To determine the feasibility of using amphotropically packaged retroviral vectors to transduce primary rabbit mesenchymal stem cells of periosteal origin, primary periosteal cells were isolated from New Zealand white rabbits, transduced in vitro with a retroviral vector bearing both the nuclear localized lacZ marker gene and the neo(r) gene, and selected in G418. We used a convenient model for analysis of in vivo stability of these cells which were seeded on to polymer scaffold grafts and implanted into rabbit femoral osteochondral defects. The nuclear localized beta-galactosidase protein was expressed in essentially 100% of selected cells in vitro and was observed in the experimental explants from animals after both 4 and 8 weeks in vivo, while cells transduced with a retroviral vector bearing only the neo(r) gene in negative control explants showed no blue staining. We extended our study by delivering a gene of therapeutic relevance, human bone morphogenic protein 7 (hBMP-7), to primary periosteal cells via retroviral vector. The hBMP-7 gene was cloned from human kidney 293 cell total RNA by RT-PCR into a retroviral vector under control of the CMV enhancer/promoter. Hydroxyapatite secretion, presumably caused by overexpression of hBMP-7, was observed on the surface of the transduced and selected periosteal cells, however, this level of expression was toxic to both PA317 producer and primary periosteal cells. Subsequently, the strong CMV enhancer/promoter driving the hBMP-7 gene was replaced in the retroviral vector by a weaker enhancer/promoter from the rat beta-actin gene. Nontoxic levels of expression of hBMP-7 were confirmed at both the RNA and protein levels in PA317 producer and primary periosteal cell lines and cell supernatants. This work demonstrates the feasibility of using a gene therapy approach in attempts to promote bone and cartilage tissue repair using gene-modified periosteal cells on grafts.

Retroviral vector-mediated gene transfer into keratocytes: in vitro effects of polybrene and protamine sulfate.

To determine the potential of somatic gene transfer as a novel technique for modulating corneal wound healing on a cellular level, the successful transduction of human keratocytes should be ascertained in vitro. In addition, the ability of different polycations to increase the transduction efficiency and their antiproliferative and cytotoxic effects should be assessed. To test transduction efficiency (X-Gal staining), cultured human keratocytes were incubated for 2 h with a retroviral vector bearing the beta-galactosidase gene, with and without the addition of polybrene or protamine sulfate. To test the antiproliferative and cytotoxic effects, cultured human keratocytes were incubated with various concentrations of polybrene and protamine sulfate (0.08 to 800 micrograms/ml) for 2, 24 and 72 h, and evaluations were performed by means of an XTT-based colorimetric assay and phase-contrast microscopy. Human keratocytes in vitro were transduced successfully with the beta-galactosidase gene (3.5 +/- 1.0%). Transduction efficiency was significantly (P < or = 0.01) improved by addition of a polycation (from 12.3 +/- 1.7% to 18.6 +/- 2.3%), but there was no significant difference between the effects of polybrene and those of protamine sulfate. Both drugs induced a highly significant dose-dependent inhibition of proliferation (P < 0.001). ID50 ranged from 11 to 22 micrograms/ml with polybrene and from 15 to 244 micrograms/ml with protamine sulfate. Only with doses of 80 and 800 micrograms/ml did protamine sulfate produce less antiproliferative effects than polybrene (P < or = 0.04). The lowest concentrations induced no morphological signs of cytotoxicity, whereas these signs were mild at 8 micrograms/ml and moderate to severe at the highest concentrations. Both polybrene and protamine sulfate can significantly improve the in vitro efficiency of successful retroviral vector-mediated gene transfer into keratocytes. Mild cytotoxic and moderate antiproliferative effects are to be expected in cultured keratocytes with a standard transduction procedure (8 micrograms/ml for 2 h).

High-efficiency retroviral vector-mediated gene transfer into human T lymphocytes.

Allogeneic bone marrow transplantation (aBMT) is an important treatment option for many haematological malignancies. Co-transplanted donor lymphocytes are the main cause of a common, in severe cases life-threatening, complication in aBMT- the graft-versus-host disease (GvHD). Depletion of T cells from the graft prevents severe GvHD, but at the same time leads to an impaired overall outcome of the transplantation due to increased incidence of graft rejection, opportunistic infections, and relapses. Infusion of genetically modified donor T cells, containing a Herpes simplex virus thymidine kinase (HSV-tk) suicide gene, should restore the “allogeneic immune advance”, especially the graft-versus-leukaemia (GvL) effect. In case of GvHD development, infused donor lymphocytes can be specifically eliminated by applying ganciclovir, an anti-Herpes drug commonly used in the clinics [1].

Retrovirus-mediated transfer of the cDNA for human glucocerebrosidase into peripheral blood repopulating cells of patients with Gaucher's disease.

Human Gene TherapyVol. 8, No. 17 News and CommentsRetrovirus-Mediated Transfer of the cDNA for Human Glucocerebrosidase into Peripheral Blood Repopulating Cells of Patients with Gaucher's Disease. University of Wisconsin, Madison, WisconsinPrincipal Investigator Friedrich SchueningPrincipal Investigator Friedrich SchueningSearch for more papers by this authorPublished Online:20 Mar 2008https://doi.org/10.1089/hum.1997.8.17-2143AboutSectionsPDF/EPUB ToolsPermissionsDownload CitationsTrack CitationsAdd to favorites Back To Publication ShareShare onFacebookTwitterLinked InRedditEmail FiguresReferencesRelatedDetailsCited byApplications of Gene TherapyGene therapy for β-thalassaemia: the continuing challenge1 October 2010 | Expert Reviews in Molecular Medicine, Vol. 12Therapy for Gaucher disease: Don’t stop thinking about tomorrowMolecular Genetics and Metabolism, Vol. 90, No. 2The evolving role of gene-based treatment in surgery4 November 2005 | British Journal of Surgery, Vol. 92, No. 12A review of gene therapy for haematological disordersBritish Journal of Haematology, Vol. 128, No. 1The Current Status of Gene Therapy in Autologous Transplantation2 November 2005 | Acta Haematologica, Vol. 114, No. 4Cell-based therapies for birth defects: A role for adult stem cell plasticity?25 August 2003 | Birth Defects Research Part C: Embryo Today: Reviews, Vol. 69, No. 3Macrophage-Specific Gene Targeting In VivoHematopoietic stem cell gene therapy for inherited bone marrow disorders: Past accomplishments and continued challenges16 May 2002 | Journal of Cellular Biochemistry, Vol. 85, No. S38Gene therapy for genetic haematological disorders and immunodeficienciesJournal of Internal Medicine, Vol. 249, No. 4Current Status of Retroviral Vector Mediated Gene Transfer into Human Hematopoietic Stem Cells1 July 2009 | Leukemia & Lymphoma, Vol. 41, No. 1-2Current Status of Retroviral Vector Mediated Gene Transfer into Human Hematopoietic Stem Cells1 July 2009 | Leukemia & Lymphoma, Vol. 41, No. 5-6Mild preconditioning and low-level engraftment confer methotrexate resistance in mice transplanted with marrow expressing drug-resistant dihydrofolate reductase activityBlood, Vol. 96, No. 4Mild preconditioning and low-level engraftment confer methotrexate resistance in mice transplanted with marrow expressing drug-resistant dihydrofolate reductase activityBlood, Vol. 96, No. 4Viral Vectors for Gene TransferDrugs, Vol. 60, No. 2Towards metabolic sink therapy for mut methylmalonic acidaemia: Correction of methylmalonyl-CoA mutase deficiency in T lymphocytes from a mut methylmalonic acidaemia child by retroviral-mediated gene transfer1 October 1999 | Journal of Inherited Metabolic Disease, Vol. 22, No. 7Transfert génique à visée thérapeutique dans les cellules souches hématopoïétiques et les cellules du système immunitaireAnnales de l'Institut Pasteur / Actualités, Vol. 10, No. 3-4Gene therapy for lysosomal storage diseases23 February 2005 | Expert Opinion on Investigational Drugs, Vol. 7, No. 10 Volume 8Issue 17Nov 1997 InformationCopyright 1997, Mary Ann Liebert, Inc.To cite this article:Principal Investigator Friedrich Schuening.Retrovirus-Mediated Transfer of the cDNA for Human Glucocerebrosidase into Peripheral Blood Repopulating Cells of Patients with Gaucher's Disease. University of Wisconsin, Madison, Wisconsin.Human Gene Therapy.Nov 1997.2143-2160.http://doi.org/10.1089/hum.1997.8.17-2143Published in Volume: 8 Issue 17: March 20, 2008PDF download