Preclinical demonstration of lentiviral vector-mediated correction of immunological and metabolic abnormalities in models of adenosine deaminase deficiency

Denise A. Carbonaro, Lin Zhang, Xiangyang Jin, Claudia Montiel-Equihua, Sabine Geiger, Marlene Carmo, Aaron Cooper, Lynette Fairbanks, Michael L. Kaufman, Neil J. Sebire, Roger P. Hollis, Michael P. Blundell, Shantha Senadheera, Pei Yu Fu, Arineh Sahaghian, Rebecca Y. Chan, Xiaoyan Wang, Kenneth Cornetta, Adrian J. Thrasher, Donald B. KohnH. Bobby Gaspar

Research output: Contribution to journalArticle

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Abstract

Gene transfer into autologous hematopoietic stem cells by γ-retroviral vectors (gRV) is an effective treatment for adenosine deaminase (ADA)-deficient severe combined immunodeficiency (SCID). However, current gRV have significant potential for insertional mutagenesis as reported in clinical trials for other primary immunodeficiencies. To improve the efficacy and safety of ADA-SCID gene therapy (GT), we generated a self-inactivating lentiviral vector (LV) with a codon-optimized human cADA gene under the control of the short form elongation factor-1α promoter (LV EFS ADA). In ADA -/- mice, LV EFS ADA displayed high-efficiency gene transfer and sufficient ADA expression to rescue ADA-/- mice from their lethal phenotype with good thymic and peripheral T- and B-cell reconstitution. Human ADA-deficient CD34+ cells transduced with 1-5 × 107 TU/ml had 1-3 vector copies/cell and expressed 1-2x of normal endogenous levels of ADA, as assayed in vitro and by transplantation into immune-deficient mice. Importantly, in vitro immortalization assays demonstrated that LV EFS ADA had significantly less transformation potential compared to gRV vectors, and vector integration-site analysis by nrLAM-PCR of transduced human cells grown in immune-deficient mice showed no evidence of clonal skewing. These data demonstrated that the LV EFS ADA vector can effectively transfer the human ADA cDNA and promote immune and metabolic recovery, while reducing the potential for vector-mediated insertional mutagenesis.

Original languageEnglish
Pages (from-to)607-622
Number of pages16
JournalMolecular Therapy
Volume22
Issue number3
DOIs
StatePublished - 2014

Fingerprint

Adenosine Deaminase
Insertional Mutagenesis
Genes
Peptide Elongation Factor 1
Severe Combined Immunodeficiency
Hematopoietic Stem Cells
Severe combined immunodeficiency due to adenosine deaminase deficiency
Codon
Genetic Therapy
B-Lymphocytes
Complementary DNA
Transplantation
Clinical Trials
Phenotype
Safety
Polymerase Chain Reaction

ASJC Scopus subject areas

  • Molecular Biology
  • Molecular Medicine
  • Genetics
  • Drug Discovery
  • Pharmacology

Cite this

Preclinical demonstration of lentiviral vector-mediated correction of immunological and metabolic abnormalities in models of adenosine deaminase deficiency. / Carbonaro, Denise A.; Zhang, Lin; Jin, Xiangyang; Montiel-Equihua, Claudia; Geiger, Sabine; Carmo, Marlene; Cooper, Aaron; Fairbanks, Lynette; Kaufman, Michael L.; Sebire, Neil J.; Hollis, Roger P.; Blundell, Michael P.; Senadheera, Shantha; Fu, Pei Yu; Sahaghian, Arineh; Chan, Rebecca Y.; Wang, Xiaoyan; Cornetta, Kenneth; Thrasher, Adrian J.; Kohn, Donald B.; Gaspar, H. Bobby.

In: Molecular Therapy, Vol. 22, No. 3, 2014, p. 607-622.

Research output: Contribution to journalArticle

Carbonaro, DA, Zhang, L, Jin, X, Montiel-Equihua, C, Geiger, S, Carmo, M, Cooper, A, Fairbanks, L, Kaufman, ML, Sebire, NJ, Hollis, RP, Blundell, MP, Senadheera, S, Fu, PY, Sahaghian, A, Chan, RY, Wang, X, Cornetta, K, Thrasher, AJ, Kohn, DB & Gaspar, HB 2014, 'Preclinical demonstration of lentiviral vector-mediated correction of immunological and metabolic abnormalities in models of adenosine deaminase deficiency', Molecular Therapy, vol. 22, no. 3, pp. 607-622. https://doi.org/10.1038/mt.2013.265
Carbonaro, Denise A. ; Zhang, Lin ; Jin, Xiangyang ; Montiel-Equihua, Claudia ; Geiger, Sabine ; Carmo, Marlene ; Cooper, Aaron ; Fairbanks, Lynette ; Kaufman, Michael L. ; Sebire, Neil J. ; Hollis, Roger P. ; Blundell, Michael P. ; Senadheera, Shantha ; Fu, Pei Yu ; Sahaghian, Arineh ; Chan, Rebecca Y. ; Wang, Xiaoyan ; Cornetta, Kenneth ; Thrasher, Adrian J. ; Kohn, Donald B. ; Gaspar, H. Bobby. / Preclinical demonstration of lentiviral vector-mediated correction of immunological and metabolic abnormalities in models of adenosine deaminase deficiency. In: Molecular Therapy. 2014 ; Vol. 22, No. 3. pp. 607-622.
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abstract = "Gene transfer into autologous hematopoietic stem cells by γ-retroviral vectors (gRV) is an effective treatment for adenosine deaminase (ADA)-deficient severe combined immunodeficiency (SCID). However, current gRV have significant potential for insertional mutagenesis as reported in clinical trials for other primary immunodeficiencies. To improve the efficacy and safety of ADA-SCID gene therapy (GT), we generated a self-inactivating lentiviral vector (LV) with a codon-optimized human cADA gene under the control of the short form elongation factor-1α promoter (LV EFS ADA). In ADA -/- mice, LV EFS ADA displayed high-efficiency gene transfer and sufficient ADA expression to rescue ADA-/- mice from their lethal phenotype with good thymic and peripheral T- and B-cell reconstitution. Human ADA-deficient CD34+ cells transduced with 1-5 × 107 TU/ml had 1-3 vector copies/cell and expressed 1-2x of normal endogenous levels of ADA, as assayed in vitro and by transplantation into immune-deficient mice. Importantly, in vitro immortalization assays demonstrated that LV EFS ADA had significantly less transformation potential compared to gRV vectors, and vector integration-site analysis by nrLAM-PCR of transduced human cells grown in immune-deficient mice showed no evidence of clonal skewing. These data demonstrated that the LV EFS ADA vector can effectively transfer the human ADA cDNA and promote immune and metabolic recovery, while reducing the potential for vector-mediated insertional mutagenesis.",
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AU - Carbonaro, Denise A.

AU - Zhang, Lin

AU - Jin, Xiangyang

AU - Montiel-Equihua, Claudia

AU - Geiger, Sabine

AU - Carmo, Marlene

AU - Cooper, Aaron

AU - Fairbanks, Lynette

AU - Kaufman, Michael L.

AU - Sebire, Neil J.

AU - Hollis, Roger P.

AU - Blundell, Michael P.

AU - Senadheera, Shantha

AU - Fu, Pei Yu

AU - Sahaghian, Arineh

AU - Chan, Rebecca Y.

AU - Wang, Xiaoyan

AU - Cornetta, Kenneth

AU - Thrasher, Adrian J.

AU - Kohn, Donald B.

AU - Gaspar, H. Bobby

PY - 2014

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N2 - Gene transfer into autologous hematopoietic stem cells by γ-retroviral vectors (gRV) is an effective treatment for adenosine deaminase (ADA)-deficient severe combined immunodeficiency (SCID). However, current gRV have significant potential for insertional mutagenesis as reported in clinical trials for other primary immunodeficiencies. To improve the efficacy and safety of ADA-SCID gene therapy (GT), we generated a self-inactivating lentiviral vector (LV) with a codon-optimized human cADA gene under the control of the short form elongation factor-1α promoter (LV EFS ADA). In ADA -/- mice, LV EFS ADA displayed high-efficiency gene transfer and sufficient ADA expression to rescue ADA-/- mice from their lethal phenotype with good thymic and peripheral T- and B-cell reconstitution. Human ADA-deficient CD34+ cells transduced with 1-5 × 107 TU/ml had 1-3 vector copies/cell and expressed 1-2x of normal endogenous levels of ADA, as assayed in vitro and by transplantation into immune-deficient mice. Importantly, in vitro immortalization assays demonstrated that LV EFS ADA had significantly less transformation potential compared to gRV vectors, and vector integration-site analysis by nrLAM-PCR of transduced human cells grown in immune-deficient mice showed no evidence of clonal skewing. These data demonstrated that the LV EFS ADA vector can effectively transfer the human ADA cDNA and promote immune and metabolic recovery, while reducing the potential for vector-mediated insertional mutagenesis.

AB - Gene transfer into autologous hematopoietic stem cells by γ-retroviral vectors (gRV) is an effective treatment for adenosine deaminase (ADA)-deficient severe combined immunodeficiency (SCID). However, current gRV have significant potential for insertional mutagenesis as reported in clinical trials for other primary immunodeficiencies. To improve the efficacy and safety of ADA-SCID gene therapy (GT), we generated a self-inactivating lentiviral vector (LV) with a codon-optimized human cADA gene under the control of the short form elongation factor-1α promoter (LV EFS ADA). In ADA -/- mice, LV EFS ADA displayed high-efficiency gene transfer and sufficient ADA expression to rescue ADA-/- mice from their lethal phenotype with good thymic and peripheral T- and B-cell reconstitution. Human ADA-deficient CD34+ cells transduced with 1-5 × 107 TU/ml had 1-3 vector copies/cell and expressed 1-2x of normal endogenous levels of ADA, as assayed in vitro and by transplantation into immune-deficient mice. Importantly, in vitro immortalization assays demonstrated that LV EFS ADA had significantly less transformation potential compared to gRV vectors, and vector integration-site analysis by nrLAM-PCR of transduced human cells grown in immune-deficient mice showed no evidence of clonal skewing. These data demonstrated that the LV EFS ADA vector can effectively transfer the human ADA cDNA and promote immune and metabolic recovery, while reducing the potential for vector-mediated insertional mutagenesis.

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