TY - JOUR
T1 - High-level erythroid-specific gene expression in primary human and murine hematopoietic cells with self-inactivating lentiviral vectors
AU - Moreau-Gaudry, Francois
AU - Xia, Ping
AU - Jiang, Gang
AU - Perelman, Natalya P.
AU - Bauer, Gerhard
AU - Ellis, James
AU - Surinya, Katherine H.
AU - Mavilio, Fulvio
AU - Shen, Che Kun
AU - Malik, Punam
PY - 2001/11/1
Y1 - 2001/11/1
N2 - 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 [I8] enhancers) and WPRE expressed at levels higher than the HS2/βpromoter 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 I8/HS-40-containing vector encoding a hybrid human β/γglobin gene led to 43% to 113% human γ-globin expression/copy of the mouse α-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.
AB - 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 [I8] enhancers) and WPRE expressed at levels higher than the HS2/βpromoter 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 I8/HS-40-containing vector encoding a hybrid human β/γglobin gene led to 43% to 113% human γ-globin expression/copy of the mouse α-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.
UR - http://www.scopus.com/inward/record.url?scp=0035525735&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=0035525735&partnerID=8YFLogxK
U2 - 10.1182/blood.V98.9.2664
DO - 10.1182/blood.V98.9.2664
M3 - Article
C2 - 11675336
AN - SCOPUS:0035525735
VL - 98
SP - 2664
EP - 2672
JO - Blood
JF - Blood
SN - 0006-4971
IS - 9
ER -