Efficient clinical scale gene modification via zinc finger nuclease-targeted disruption of the HIV co-receptor CCR5.
Since HIV requires CD4 and a co-receptor, most commonly C-C chemokine receptor 5 (CCR5), for cellular entry, targeting CCR5 expression is an attractive approach for therapy of HIV infection. Treatment of CD4(+) T cells with zinc-finger protein nucleases (ZFNs) specifically disrupting chemokine receptor CCR5 coding sequences induces resistance to HIV infection in vitro and in vivo. A chimeric Ad5/F35 adenoviral vector encoding CCR5-ZFNs permitted efficient delivery and transient expression following anti-CD3/anti-CD28 costimulation of T lymphocytes. We present data showing CD3/CD28 costimulation substantially improved transduction efficiency over reported methods for Ad5/F35 transduction of T lymphocytes. Modifications to the laboratory scale process, incorporating clinically compatible reagents and methods, resulted in a robust ex vivo manufacturing process capable of generating >10(10) CCR5 gene-edited CD4+ T cells from healthy and HIV+ donors. CD4+ T-cell phenotype, cytokine production, and repertoire were comparable between ZFN-modified and control cells. Following consultation with regulatory authorities, we conducted in vivo toxicity studies that showed no detectable ZFN-specific toxicity or T-cell transformation. Based on these findings, we initiated a clinical trial testing the safety and feasibility of CCR5 gene-edited CD4+ T-cell transfer in study subjects with HIV-1 infection.
Department of Pathology and Laboratory Medicine, University of Pennsylvania Perelman School of Medicine, Philadelphia, PA 19104, USA.
Dawn A
DA
Maier
Andrea L
AL
Brennan
Shuguang
S
Jiang
Gwendolyn K
GK
Binder-Scholl
Gary
G
Lee
Gabriela
G
Plesa
Zhaohui
Z
Zheng
Julio
J
Cotte
Carmine
C
Carpenito
Travis
T
Wood
S Kaye
SK
Spratt
Dale
D
Ando
Philip
P
Gregory
Michael C
MC
Holmes
Elena E
EE
Perez
James L
JL
Riley
Richard G
RG
Carroll
Carl H
CH
June
Bruce L
BL
Levine
D000260:Adenoviruses, Human / Q000235:genetics; D019264:Adoptive Transfer; D000818:Animals; D018106:CD28 Antigens / Q000276:immunology; D017252:CD3 Complex / Q000276:immunology; D004262:DNA Restriction Enzymes / Q000235:genetics* / Q000378:metabolism; D005260:Female; D005822:Genetic Vectors / Q000008:administration & dosage / Q000009:adverse effects / Q000235:genetics / Q000592:standards*; D015658:HIV Infections / Q000235:genetics* / Q000276:immunology* / Q000628:therapy; D006801:Humans; D008213:Lymphocyte Activation / Q000276:immunology; D008297:Male; D051379:Mice; D010641:Phenotype; D019713:Receptors, CCR5 / Q000235:genetics* / Q000276:immunology; D013601:T-Lymphocytes / Q000276:immunology / Q000378:metabolism; D014161:Transduction, Genetic / Q000379:methods / Q000592:standards; D014183:Transplantation, Heterologous; D016335:Zinc Fingers / Q000235:genetics*
D018106:CD28 Antigens; D017252:CD3 Complex; D019713:Receptors, CCR5; D004262:DNA Restriction Enzymes