Efficient human hematopoietic cell transduction using RD114- and GALV-pseudotyped retroviral vectors produced in suspension and serum-free media.
Retroviral vectors derived from the Moloney murine leukemia virus have been used in successful and promising gene therapy clinical trials. However, platforms for their large-scale production must be further developed. As a proof of principle, we reported the generation of a packaging cell line that produces amphotropic retroviral vectors in suspension and serum-free medium (SFM). In the present study, we have constructed and characterized two retroviral packaging cell lines designed for gene transfer in hematopoietic cells. These cell lines grow in suspension and SFM, and produce high-titer RD114- and gibbon ape leukemia virus (GALV)-pseudotyped vectors for a 3-month culture period. Viral particles released are as robust during repeated freeze-thaw cycles and on thermal inactivation at 37 degrees C as their counterparts produced in cells cultured adherently with serum. We also show that RD114- and GALV-pseudotyped vectors produced in suspension and SFM efficiently transduce human lymphocytes and hematopoietic stem cells. As these retroviral packaging cell lines distinctively maintain high vector titers while growing in suspension and SFM, we conclude that these cell lines are uniquely suitable for large-scale clinical-grade vector production for late-phase clinical trials involving gene transfer into hematopoietic cells.
Ghani
Karim
K
Centre de Recherche en Cancérologie de l'Université Laval, L'Hôtel Dieu de Québec, Centre Hospitalier Universitaire de Québec, Québec, Canada.
Wang
Xiuyan
X
de Campos-Lima
Pedro Otavio
PO
Olszewska
Malgorzata
M
Kamen
Amine
A
Rivière
Isabelle
I
Caruso
Manuel
M
D000818:Animals; D018952:Antigens, CD34 / Q000235:genetics / Q000378:metabolism; D002460:Cell Line; D003470:Culture Media; D016895:Culture Media, Serum-Free; D015316:Genetic Therapy / Q000379:methods; D005822:Genetic Vectors / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D006412:Hematopoietic Stem Cells / Q000378:metabolism / Q000821:virology*; D006801:Humans; D017989:Leukemia Virus, Gibbon Ape / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D012190:Retroviridae / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D013601:T-Lymphocytes / Q000821:virology; D014161:Transduction, Genetic; D014759:Viral Envelope Proteins / Q000235:genetics / Q000378:metabolism; D019065:Virus Assembly
D018952:Antigens, CD34; D003470:Culture Media; D016895:Culture Media, Serum-Free; D014759:Viral Envelope Proteins; D049452:Green Fluorescent Proteins