Use of oncoretroviral vectors in gene therapy for hemoglobinopathies has been impeded by low titer vectors, genetic instability, and poor expression. Fifteen self-inactivating (SIN) lentiviral vectors using 4 erythroid promoters in combination with 4 erythroid enhancers with or without the woodchuck hepatitis virus postregulatory element (WPRE) were generated using the enhanced green fluorescent protein as a reporter gene. Vectors with high erythroid-specific expression in cell lines were tested in primary human CD34(+) cells and in vivo in the murine bone marrow (BM) transplantation model. Vectors containing the ankyrin-1 promoter showed high-level expression and stable proviral transmission. Two vectors containing the ankyrin-1 promoter and 2 erythroid enhancers (HS-40 plus GATA-1 or HS-40 plus 5-aminolevulinate synthase intron 8 [18] enhancers) and WPRE expressed at levels higher than the HS2/beta -promoter vector in bulk unilineage erythroid cultures and individual erythroid blast-forming units derived from human BM CD34+ cells. Sca1(+)/lineage(-) Ly5.1 mouse hematopoietic cells, transduced with these 2 ankyrin-1 promoter vectors, were injected into lethally irradiated Ly5.2 recipients. Eleven weeks after transplantation, high-level expression was seen from both vectors in blood (63%-89% of red blood cells) and erythroid cells in BM (70%-86% engraftment), compared with negligible expression in myeloid and lymphoid lineages in blood, BM, spleen, and thymus (0%-4%). The 18/HS-40-containing vector encoding a hybrid human beta/gamma -globin gene led to 43% to 113% human gamma -globin expression/copy of the mouse alpha -globin gene. Thus, modular use of erythroid-specific enhancers/promoters and WPRE in SIN-lentiviral vectors led to Identification of high-titer, stably transmitted vectors with high-level erythroid-specific expression for gene therapy of red cell diseases.

High-level erythroid-specific gene expression in primary human and murine hematopoietic cells with self-inactivating lentiviral vectors / F., Moreau Gaudry; P., Xia; G., Jiang; Np, Perelman; G., Bauer; J., Ellis; Kh, Surinya; Mavilio, Fulvio; Ck, Shen; P., Malik. - In: BLOOD. - ISSN 0006-4971. - STAMPA. - 98:9(2001), pp. 2664-2672. [10.1182/blood.V98.9.2664]

High-level erythroid-specific gene expression in primary human and murine hematopoietic cells with self-inactivating lentiviral vectors

MAVILIO, Fulvio;
2001

Abstract

Use of oncoretroviral vectors in gene therapy for hemoglobinopathies has been impeded by low titer vectors, genetic instability, and poor expression. Fifteen self-inactivating (SIN) lentiviral vectors using 4 erythroid promoters in combination with 4 erythroid enhancers with or without the woodchuck hepatitis virus postregulatory element (WPRE) were generated using the enhanced green fluorescent protein as a reporter gene. Vectors with high erythroid-specific expression in cell lines were tested in primary human CD34(+) cells and in vivo in the murine bone marrow (BM) transplantation model. Vectors containing the ankyrin-1 promoter showed high-level expression and stable proviral transmission. Two vectors containing the ankyrin-1 promoter and 2 erythroid enhancers (HS-40 plus GATA-1 or HS-40 plus 5-aminolevulinate synthase intron 8 [18] enhancers) and WPRE expressed at levels higher than the HS2/beta -promoter vector in bulk unilineage erythroid cultures and individual erythroid blast-forming units derived from human BM CD34+ cells. Sca1(+)/lineage(-) Ly5.1 mouse hematopoietic cells, transduced with these 2 ankyrin-1 promoter vectors, were injected into lethally irradiated Ly5.2 recipients. Eleven weeks after transplantation, high-level expression was seen from both vectors in blood (63%-89% of red blood cells) and erythroid cells in BM (70%-86% engraftment), compared with negligible expression in myeloid and lymphoid lineages in blood, BM, spleen, and thymus (0%-4%). The 18/HS-40-containing vector encoding a hybrid human beta/gamma -globin gene led to 43% to 113% human gamma -globin expression/copy of the mouse alpha -globin gene. Thus, modular use of erythroid-specific enhancers/promoters and WPRE in SIN-lentiviral vectors led to Identification of high-titer, stably transmitted vectors with high-level erythroid-specific expression for gene therapy of red cell diseases.
2001
98
9
2664
2672
High-level erythroid-specific gene expression in primary human and murine hematopoietic cells with self-inactivating lentiviral vectors / F., Moreau Gaudry; P., Xia; G., Jiang; Np, Perelman; G., Bauer; J., Ellis; Kh, Surinya; Mavilio, Fulvio; Ck, Shen; P., Malik. - In: BLOOD. - ISSN 0006-4971. - STAMPA. - 98:9(2001), pp. 2664-2672. [10.1182/blood.V98.9.2664]
F., Moreau Gaudry; P., Xia; G., Jiang; Np, Perelman; G., Bauer; J., Ellis; Kh, Surinya; Mavilio, Fulvio; Ck, Shen; P., Malik
File in questo prodotto:
Non ci sono file associati a questo prodotto.
Pubblicazioni consigliate

Licenza Creative Commons
I metadati presenti in IRIS UNIMORE sono rilasciati con licenza Creative Commons CC0 1.0 Universal, mentre i file delle pubblicazioni sono rilasciati con licenza Attribuzione 4.0 Internazionale (CC BY 4.0), salvo diversa indicazione.
In caso di violazione di copyright, contattare Supporto Iris

Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/11380/305504
Citazioni
  • ???jsp.display-item.citation.pmc??? 25
  • Scopus 101
  • ???jsp.display-item.citation.isi??? 95
social impact