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1062. Functional Correction of T Cells from Wiskott-Aldrich Syndrome Patients by Retroviral and Lentiviral Vector-Mediated Gene Transfer
STEM CELL GENE THERAPY FOR GENETIC DISEASES 1060. Functional Analysis of an Oncoretrovirus Vector for Human Gamma-Globin in Mouse Models of Beta Thalassemia Tamon Nishino,1 Julie Tubb,1 Yumiko Nishino,1 George Stamatoyannopoulos,1 David W. Emery.1 1 Medicine / Medical Genetics, University of Washington, Seattle, WA. Many advances have been made over the past few years in the development of recombinant virus vectors for human globin genes designed to treat patients with beta-thalassemia and sickle cell disease. However, many of these advances have relied on the use of lentivirus vectors. We have recently described the development of an oncoretrovirus vector for human gamma-globin that is resistant to silencing position effects and expresses gamma-globin at 6 ±4% of total endogenous alpha-globin in normal mice transplanted longterm with transduced marrow [Blood 100:2012, 2002]. In order to determine whether this is a potentially therapeutic level of expression, we performed studies in a mouse model for beta-thalassemia intermedia. Transduction and transplantation of bone marrow cells from heterozygous Hbbth-3/+ donors into myeloablated syngeneic recipients resulted in a modest improvement in both the RBC count (3.8±0.1 x106/ul vs. 2.6±1.1 for mock and 6.0±1.8 for wild type) and total hemoglobin (6.7±0.2 g/dl vs. 4.7±1.9 for mock and 12.2±3.3 for wild type). However, this effect was only seen early after engraftment and in mice with greater than 25% gamma-expressing RBC. Likewise, studies with this vector in a mouse model for severe beta-thalassemia major also showed only a partial therapeutic effect. In this case, transduction and transplantation of fetal liver cells from Hbbth-3/Hbbth-3 homozygotes into congenic C57BL6 recipients increased survival to 34±20 days (n=13), compared to only 16±7 days for the mock-transduced controls (p=0.006). However, the level of gamma-globin provided by the optimized vector was not sufficient to support long-term survival in this model. In order to determine the amount of gamma-globin necessary for a full therapeutic correction in these mouse models of beta-thalassemia, we crossed Hbbth-3/+ heterozygotes with a transgenic line expressing gamma-globin at a level 2-3 fold higher (10-15% of total endogenous alpha-globin) than that of the optimized oncoretrovirus vector. Compound heterozygotes exhibited a significant improvement in both RBC counts (11.4±0.3 x106/ul vs. 10.5 ±0.5 for Hbbth-3/+ and 11.6±0.7 for w.t.) and total hemoglobin (15.0±0.4 g/dl vs. 13.1 ±0.6 for Hbbth-3/+ and 17.9±1.5 for w.t.), suggesting a nearly full correction. However, the level of gamma-globin produced by the transgene was insufficient to rescue the Hbb th-3/Hbb th-3 embryonic lethality phenotype. These studies are currently being repeated with a transgenic line expressing gamma-globin at twice the level of the original line. Taken together these studies demonstrate that an optimized oncoretrovirus vector for human gamma-globin is capable of functionally improving the phenotypes of two mouse models of beta-thalassemia, but that full correction of these phenotypes will require further improvement in the level of vector expression.
1061. Reconstitution of the IL-12 Signaling β1) KO Pathway in IL-12 Receptor beta-1 (IL-12Rβ Mice upon Transplantation of Retrovirally Transduced Hematopoietic Stem Cells Marita Bosticardo,1 Francesco Novelli,2 Jean-Laurent Casanova,3 Fabio Candotti.1 1 Genetics and Molecular Biology Branch, NHGRI/NIH, Bethesda, MD, United States; 2Centro Oncologico Ematologico Subalpino, Centro Ricerche Medicina Sperimentale, Ospedale San Giovanni Battista, Torino, Italy; 3Unité Clinique d’Immunologie et d’Hématologie Pédiatriques, Hôpital Necker-Enfants Malades, Paris, France. The functionality of the IL-12-mediated signaling pathway is critical for the elimination of intracellular pathogens. Patients carrying genetic defects in the IL-12 signaling pathway (IL-12 p40 or IL12Rβ1) indeed show increased susceptibility to weakly pathogenic strains of mycobacteria and salmonella. Infections can be treated by administration of IFN-γ and antibiotics; however, reversion of patients’ susceptibility by corrective gene transfer could be beneficial. We were able to restore the IL-12-mediated signaling pathway in PHA-activated T cells blasts of IL-12Rβ1 deficient patients through retroviral-mediated gene correction. In addition, to test the feasibility and safety of retroviral-mediated gene correction in vivo, we established a murine model of gene therapy in IL-12Rβ1deficient mice. Lineage-negative cells isolated from bone marrow of IL-12Rβ1-deficient mice were cultured for 5 days and transduced using a retroviral vector carrying the murine IL-12Rβ1 cDNA. This resulted in a high percentage of IL-12Rβ1+ cells (50-100%). Genecorrected bone marrow cells were transplanted into lethally irradiated recipient mice, which were then analyzed at 8-16 weeks after the transplant. Treated mice did not show adverse effects upon gene therapy. We detected IL-12Rβ1 expression in peripheral blood lymphocytes, spleen and thymus from the majority of treated mice. Moreover, splenocytes isolated from mice transplanted with genecorrected bone marrow cells acquired the ability to respond to IL12, as demonstrated by the production of IFN-γ, which is completely impaired in IL-12Rβ1 KO mice. However, membrane expression of the IL-12Rβ1 chain as well as the production of IFN-γ in response to IL-12 where significantly lower than those detected in wild type control mice. In vivo experiments of IFN-γ production and challenge with intracellular pathogens are planned to establish if the degree of gene correction reached in our experimental model is sufficient to overcome their immunodeficiency.
1062. Functional Correction of T Cells from Wiskott-Aldrich Syndrome Patients by Retroviral and Lentiviral Vector-Mediated Gene Transfer Loïc Dupré,1 Sara Trifari,1 Francesco Marangoni,1 Antonia Follenzi,2 Antonio Bernad,3 Silvana Martino,4 Shigeru Tsuchiya,5 Luigi Naldini,1,2 Claudio Bordignon,1 Alessandro Aiuti,1 MariaGrazia Roncarolo.1 1 San Raffaele Telethon Institute for Gene Therapy (HSR-TIGET), Milan, Italy; 2IRCC, University of Turin, Turin, Italy; 3National Center for Biotechnology, Autonomous University of Madrid, Madrid, Spain; 4Department of Pediatrics, University of Turin, Turin, Italy; 5Department of Pediatric Oncology, Tohoku University, Sendai, Japan. Wiskott-Aldrich syndrome (WAS) is an X-linked primary immunodeficiency characterized by infections, severe hemorrhage, and lymphomas resulting in a median survival below the age of 20. WAS is caused by mutations in the gene encoding the WAS protein (WASP) which is expressed in hematopoietic cells and is crucial for actin cytoskeleton organization. Since limited success has been reported with bone marrow transplantation, gene therapy could
Molecular Therapy Vol. 7, No. 5, May 2003, Part 2 of 2 Parts Copyright ® The American Society of Gene Therapy
S409
STEM CELL GENE THERAPY FOR GENETIC DISEASES represent an alternative treatment for this disease. Defective T lymphocytes play a central role in the pathogenesis of WAS and we demonstrated that WASP is required for the assembly of the immunological synapse and for optimal T-cell activation. In this study, we compared the efficiency of different MMLV-based oncoretroviral and HIV-based lentiviral vectors in transferring the WASP gene into T lymphocytes to restore normal immune functions. Stable packaging cell lines designed to produce MMLV-based oncoretroviral vectors were associated with low virus titers and infectivity, which correlated with high WASP expression. These low titers could be overcome by transient transfection and production, which resulted in higher titers and higher transduction efficiency in patients T cells. The use of transiently produced HIVbased lentiviral vectors led to higher transduction rates, compared to oncoretroviral vectors. All vectors tested, including oncoretroviral vectors containing the WASP cDNA either under the LTR or the internal SV-40 promoter and lentiviral vectors containing the WASP cDNA either under the PGK promoter or the autologous WASP promoter, led to normal levels of WASP expression in the transduced T cells. Correction of the functional defects, including proliferation and IL-2 production, was achieved in transduced patients T cells. Functional correction of transduced T cells was further proven by normal clustering of the lipid raft marker GM1 upon TCR activation, thus indicating normal immunological synapse assembly. These results show that WASP-encoding retroviral and lentiviral vectors can lead to functional restoration of WAS T cells, providing the basis for the development of further gene therapy approaches for WAS patients using either T cells or hematopoietic stem cells as target cells.
1063. HIV Mediated Expression of Bruton´s Tyrosine Kinase in Hematopoietic Stem Cells Promotes B Cells Development but Not Restore Immunoglobulin Production in X-Linked Immunodeficient Mice Hiroko Tanabe,1,2 Koichi Miyake,1 Takashi Shimada.1 Deaprtment of Biochemistry and Molecular Biology, Nippon Medical School, Tokyo, Japan; 2Department of Internal Medicine 1, Nippon Medical School, Tokyo, Japan. 1
X-linked agammaglobulinemia (XLA) is characterized by profound hypogammaglobulinemia affecting all isotypes, an absence of antigenspecific antibodies, and less than 1% of the normal number of B cells. Mutations of the Bruton´s tyrosine kinase (Btk) gene have been implicated in the pathogenesis of human XLA. A specific mutation was also found in the mouse Btk (mBtk) gene in a spontaneously occurring mouse model of immunodeficiency, murine X-linked immunodeficiency (Xid). Although the clinical picture of the mouse model is mild compared to that of XLA patients, Xid mice should act as a useful model to evaluate new strategies for treatment of patients with XLA. As a step toward gene therapy for patients with XLA, we constructed a VSV-pseudotyped HIV vector containig the human Btk (hBtk) gene under the control of the internal murine stem cell virus (MSCV) promoter. Freshly isolated and magnetically selected hematopoietic stem cells (HSCs) from Xid mice (5-10 weeks) were transduced with the HIV vector without cytokine stimulation. The vector sequence was detected in more than 90 % of colony-forming units in vitro by PCR. Transduced HSCs (1 - 3 x 105) were injected into irradiated adult Xid mice (4 weeks old) through the tail vein. All mice were sacrificed 30 weeks post-transplantation for examination of immunological function. The HIV vector sequence and hBtk mRNA were detected in bone marrow, spleen and peripheral blood. The number of differentiated B cells (IgMlowIgDhigh) was increased in treated animals. However, there was little or no restoration of serum immunoglobulin concentrations and antibody response to NP-Ficoll challenge (T-cell-independent S410
type II antigen). We also Similarly, the immunological reconstitution was not achieved even if the Xid mice was treated by stem cell gene therapy within 48 hours after birth. To elucidate the reason why gene transfer of the Btk gene into HSCs was inadequate, we generated chimeric mice that received a mixture of 10% normal HSCs from CBA/J mice and 90% mutant-HSCs from Xid mice after lethal irradiation. Normalization of IgM and IgG3 concentrations was confirmed 7 weeks after transplantation, indicating that partial reconstitution with normal bone marrow cells is sufficient for phenotypic correction of Xid mice. The ratio of human Btk+ B cells in total B cells in Xid mice treated by HIV mediated gene therapy was 60 % in bone marrow and peripheral blood, but markedly decreased to 5 % in spleen, suggesting that bone marrow cells expressing Btk may have selective growth disadvantage during B cell development. In contrast, the ratio in mice treated by partial bone marrow transplantation was 25 % in bone marrow, but increased to 60 % in spleen. These results indicate that HIV mediated expression of Btk in bone marrow stem cells promotes B cell development, but not sufficient to restore immunoglobulin production in Xid mice. Other factors in addition to Btk may be required for normal B lymphopoiesis.
1064. Investigation into the Possible Use of HoxB4 in Order To Enhance Reconstitution of Gene Modified Haemopoietic Stem Cells Following Bone Marrow Transplantation Michael D. Milsom,1 Lorna B. Woolford,1 Dorothy Gagen,1 Rachel Duxbury,1 Claire M. Heyworth,2 Geoff Margison,3 Leslie J. Fairbairn.1 1 Gene Therapy Group, Paterson Institute for Cancer Research, Manchester, United Kingdom; 2Experimental Haematology Group, Paterson Institute for Cancer Research, Manchester, United Kingdom; 3Carcinogenesis Group, Paterson Institute for Cancer Research, Manchester, United Kingdom. We have previously described the construction of a novel tricistronic retroviral vector, which facilitates the high level coexpression of HoxB4 and the O6-alkylguanine-DNA alkyltransferase P140K mutant (ATaseP140K), along with the selective marker gene enhanced green fluorescent protein (eGFP). In order to assess the relative selective advantage conferred upon haemopoietic stem cells (HSCs) transduced with this vector, we have devised a novel in vivo competitive repopulation assay that enables a direct comparison of the performance of two retroviral vectors within the same mouse. Using this assay, we find that the tricistronic vector described above is able to confer a profound post-engraftment selective advantage over bone marrow cells transduced with a vector containing only ATaseP140K and eGFP. Furthermore, following treatment with O6benzylguanine followed by temozolomide, HSCs containing this tricistronic vector expanded to provide virtually all of the peripheral blood cells in the recipient mice. Clearly this not only demonstrates the potential use of HoxB4 to facilitate the in vivo selection of HSCs containing a second therapeutic gene, but also reveals the possibility of using HoxB4/ATaseP140K co-expression to enable a further enhanced expansion of gene modified bone marrow cells. In an effort to evaluate the safety implications of using HoxB4 in any future gene therapy protocol, we utilised the FDCP-mix haemopoietic progenitor cell line as a model system in an attempt to elucidate the mechanism of action of HoxB4. We found that retroviral mediated constitutive expression of HoxB4 lead to a block/delay in myeloid differentiation as measured by analysis of both cellular morphology and colony forming ability following a differentiation stimulus. We are currently investigating the nature of this perturbation of differentiation in order to determine if this biological phenomenon significantly contributes to the observed improvement in haemopoietic reconstitution described above. Molecular Therapy Vol. 7, No. 5, May 2003, Part 2 of 2 Parts
Copyright ® The American Society of Gene Therapy
1061. Reconstitution of the IL-12 Signaling β1) KO Pathway in IL-12 Receptor beta-1 (IL-12Rβ Mice upon Transplantation of Retrovirally Transduced Hematopoietic Stem Cells Marita Bosticardo,1 Francesco Novelli,2 Jean-Laurent Casanova,3 Fabio Candotti.1 1 Genetics and Molecular Biology Branch, NHGRI/NIH, Bethesda, MD, United States; 2Centro Oncologico Ematologico Subalpino, Centro Ricerche Medicina Sperimentale, Ospedale San Giovanni Battista, Torino, Italy; 3Unité Clinique d’Immunologie et d’Hématologie Pédiatriques, Hôpital Necker-Enfants Malades, Paris, France. The functionality of the IL-12-mediated signaling pathway is critical for the elimination of intracellular pathogens. Patients carrying genetic defects in the IL-12 signaling pathway (IL-12 p40 or IL12Rβ1) indeed show increased susceptibility to weakly pathogenic strains of mycobacteria and salmonella. Infections can be treated by administration of IFN-γ and antibiotics; however, reversion of patients’ susceptibility by corrective gene transfer could be beneficial. We were able to restore the IL-12-mediated signaling pathway in PHA-activated T cells blasts of IL-12Rβ1 deficient patients through retroviral-mediated gene correction. In addition, to test the feasibility and safety of retroviral-mediated gene correction in vivo, we established a murine model of gene therapy in IL-12Rβ1deficient mice. Lineage-negative cells isolated from bone marrow of IL-12Rβ1-deficient mice were cultured for 5 days and transduced using a retroviral vector carrying the murine IL-12Rβ1 cDNA. This resulted in a high percentage of IL-12Rβ1+ cells (50-100%). Genecorrected bone marrow cells were transplanted into lethally irradiated recipient mice, which were then analyzed at 8-16 weeks after the transplant. Treated mice did not show adverse effects upon gene therapy. We detected IL-12Rβ1 expression in peripheral blood lymphocytes, spleen and thymus from the majority of treated mice. Moreover, splenocytes isolated from mice transplanted with genecorrected bone marrow cells acquired the ability to respond to IL12, as demonstrated by the production of IFN-γ, which is completely impaired in IL-12Rβ1 KO mice. However, membrane expression of the IL-12Rβ1 chain as well as the production of IFN-γ in response to IL-12 where significantly lower than those detected in wild type control mice. In vivo experiments of IFN-γ production and challenge with intracellular pathogens are planned to establish if the degree of gene correction reached in our experimental model is sufficient to overcome their immunodeficiency.
1062. Functional Correction of T Cells from Wiskott-Aldrich Syndrome Patients by Retroviral and Lentiviral Vector-Mediated Gene Transfer Loïc Dupré,1 Sara Trifari,1 Francesco Marangoni,1 Antonia Follenzi,2 Antonio Bernad,3 Silvana Martino,4 Shigeru Tsuchiya,5 Luigi Naldini,1,2 Claudio Bordignon,1 Alessandro Aiuti,1 MariaGrazia Roncarolo.1 1 San Raffaele Telethon Institute for Gene Therapy (HSR-TIGET), Milan, Italy; 2IRCC, University of Turin, Turin, Italy; 3National Center for Biotechnology, Autonomous University of Madrid, Madrid, Spain; 4Department of Pediatrics, University of Turin, Turin, Italy; 5Department of Pediatric Oncology, Tohoku University, Sendai, Japan. Wiskott-Aldrich syndrome (WAS) is an X-linked primary immunodeficiency characterized by infections, severe hemorrhage, and lymphomas resulting in a median survival below the age of 20. WAS is caused by mutations in the gene encoding the WAS protein (WASP) which is expressed in hematopoietic cells and is crucial for actin cytoskeleton organization. Since limited success has been reported with bone marrow transplantation, gene therapy could
Molecular Therapy Vol. 7, No. 5, May 2003, Part 2 of 2 Parts Copyright ® The American Society of Gene Therapy
S409
STEM CELL GENE THERAPY FOR GENETIC DISEASES represent an alternative treatment for this disease. Defective T lymphocytes play a central role in the pathogenesis of WAS and we demonstrated that WASP is required for the assembly of the immunological synapse and for optimal T-cell activation. In this study, we compared the efficiency of different MMLV-based oncoretroviral and HIV-based lentiviral vectors in transferring the WASP gene into T lymphocytes to restore normal immune functions. Stable packaging cell lines designed to produce MMLV-based oncoretroviral vectors were associated with low virus titers and infectivity, which correlated with high WASP expression. These low titers could be overcome by transient transfection and production, which resulted in higher titers and higher transduction efficiency in patients T cells. The use of transiently produced HIVbased lentiviral vectors led to higher transduction rates, compared to oncoretroviral vectors. All vectors tested, including oncoretroviral vectors containing the WASP cDNA either under the LTR or the internal SV-40 promoter and lentiviral vectors containing the WASP cDNA either under the PGK promoter or the autologous WASP promoter, led to normal levels of WASP expression in the transduced T cells. Correction of the functional defects, including proliferation and IL-2 production, was achieved in transduced patients T cells. Functional correction of transduced T cells was further proven by normal clustering of the lipid raft marker GM1 upon TCR activation, thus indicating normal immunological synapse assembly. These results show that WASP-encoding retroviral and lentiviral vectors can lead to functional restoration of WAS T cells, providing the basis for the development of further gene therapy approaches for WAS patients using either T cells or hematopoietic stem cells as target cells.
1063. HIV Mediated Expression of Bruton´s Tyrosine Kinase in Hematopoietic Stem Cells Promotes B Cells Development but Not Restore Immunoglobulin Production in X-Linked Immunodeficient Mice Hiroko Tanabe,1,2 Koichi Miyake,1 Takashi Shimada.1 Deaprtment of Biochemistry and Molecular Biology, Nippon Medical School, Tokyo, Japan; 2Department of Internal Medicine 1, Nippon Medical School, Tokyo, Japan. 1
X-linked agammaglobulinemia (XLA) is characterized by profound hypogammaglobulinemia affecting all isotypes, an absence of antigenspecific antibodies, and less than 1% of the normal number of B cells. Mutations of the Bruton´s tyrosine kinase (Btk) gene have been implicated in the pathogenesis of human XLA. A specific mutation was also found in the mouse Btk (mBtk) gene in a spontaneously occurring mouse model of immunodeficiency, murine X-linked immunodeficiency (Xid). Although the clinical picture of the mouse model is mild compared to that of XLA patients, Xid mice should act as a useful model to evaluate new strategies for treatment of patients with XLA. As a step toward gene therapy for patients with XLA, we constructed a VSV-pseudotyped HIV vector containig the human Btk (hBtk) gene under the control of the internal murine stem cell virus (MSCV) promoter. Freshly isolated and magnetically selected hematopoietic stem cells (HSCs) from Xid mice (5-10 weeks) were transduced with the HIV vector without cytokine stimulation. The vector sequence was detected in more than 90 % of colony-forming units in vitro by PCR. Transduced HSCs (1 - 3 x 105) were injected into irradiated adult Xid mice (4 weeks old) through the tail vein. All mice were sacrificed 30 weeks post-transplantation for examination of immunological function. The HIV vector sequence and hBtk mRNA were detected in bone marrow, spleen and peripheral blood. The number of differentiated B cells (IgMlowIgDhigh) was increased in treated animals. However, there was little or no restoration of serum immunoglobulin concentrations and antibody response to NP-Ficoll challenge (T-cell-independent S410
type II antigen). We also Similarly, the immunological reconstitution was not achieved even if the Xid mice was treated by stem cell gene therapy within 48 hours after birth. To elucidate the reason why gene transfer of the Btk gene into HSCs was inadequate, we generated chimeric mice that received a mixture of 10% normal HSCs from CBA/J mice and 90% mutant-HSCs from Xid mice after lethal irradiation. Normalization of IgM and IgG3 concentrations was confirmed 7 weeks after transplantation, indicating that partial reconstitution with normal bone marrow cells is sufficient for phenotypic correction of Xid mice. The ratio of human Btk+ B cells in total B cells in Xid mice treated by HIV mediated gene therapy was 60 % in bone marrow and peripheral blood, but markedly decreased to 5 % in spleen, suggesting that bone marrow cells expressing Btk may have selective growth disadvantage during B cell development. In contrast, the ratio in mice treated by partial bone marrow transplantation was 25 % in bone marrow, but increased to 60 % in spleen. These results indicate that HIV mediated expression of Btk in bone marrow stem cells promotes B cell development, but not sufficient to restore immunoglobulin production in Xid mice. Other factors in addition to Btk may be required for normal B lymphopoiesis.
1064. Investigation into the Possible Use of HoxB4 in Order To Enhance Reconstitution of Gene Modified Haemopoietic Stem Cells Following Bone Marrow Transplantation Michael D. Milsom,1 Lorna B. Woolford,1 Dorothy Gagen,1 Rachel Duxbury,1 Claire M. Heyworth,2 Geoff Margison,3 Leslie J. Fairbairn.1 1 Gene Therapy Group, Paterson Institute for Cancer Research, Manchester, United Kingdom; 2Experimental Haematology Group, Paterson Institute for Cancer Research, Manchester, United Kingdom; 3Carcinogenesis Group, Paterson Institute for Cancer Research, Manchester, United Kingdom. We have previously described the construction of a novel tricistronic retroviral vector, which facilitates the high level coexpression of HoxB4 and the O6-alkylguanine-DNA alkyltransferase P140K mutant (ATaseP140K), along with the selective marker gene enhanced green fluorescent protein (eGFP). In order to assess the relative selective advantage conferred upon haemopoietic stem cells (HSCs) transduced with this vector, we have devised a novel in vivo competitive repopulation assay that enables a direct comparison of the performance of two retroviral vectors within the same mouse. Using this assay, we find that the tricistronic vector described above is able to confer a profound post-engraftment selective advantage over bone marrow cells transduced with a vector containing only ATaseP140K and eGFP. Furthermore, following treatment with O6benzylguanine followed by temozolomide, HSCs containing this tricistronic vector expanded to provide virtually all of the peripheral blood cells in the recipient mice. Clearly this not only demonstrates the potential use of HoxB4 to facilitate the in vivo selection of HSCs containing a second therapeutic gene, but also reveals the possibility of using HoxB4/ATaseP140K co-expression to enable a further enhanced expansion of gene modified bone marrow cells. In an effort to evaluate the safety implications of using HoxB4 in any future gene therapy protocol, we utilised the FDCP-mix haemopoietic progenitor cell line as a model system in an attempt to elucidate the mechanism of action of HoxB4. We found that retroviral mediated constitutive expression of HoxB4 lead to a block/delay in myeloid differentiation as measured by analysis of both cellular morphology and colony forming ability following a differentiation stimulus. We are currently investigating the nature of this perturbation of differentiation in order to determine if this biological phenomenon significantly contributes to the observed improvement in haemopoietic reconstitution described above. Molecular Therapy Vol. 7, No. 5, May 2003, Part 2 of 2 Parts
Copyright ® The American Society of Gene Therapy