Creatine kinase, a magnetic resonance-detectable marker gene for quantification of liver-directed gene transfer.
We reported previously the in vivo detection of ectopic and transient expression of creatine kinase gene (ck) in the liver by phosphorus-31 magnetic resonance spectroscopy ((31)P MRS). Here we demonstrate the feasibility of using ck as a reporter gene to monitor the transfer of low-density lipoprotein receptor (LDLr) gene to LDLr(/) mice, a preclinical model for familial hypercholesterolemia. A recombinant adenovirus was generated that carries the creatine kinase gene (ck) and human LDL receptor gene (hLDLr) linked by an internal ribosomal entry site sequence. Intravenous injection of the adenovirus into LDLr(/)mice (1 x 10(11) viral particles/mouse) resulted in transduction of more than 90% of hepatocytes in the liver. Simultaneous expression of ck and LDLr was confirmed by Western analysis of the transduced livers. Through precise regulation of transgene expression in hepatocytes in vitro, an excellent correlation (R(2) = 0.96) between LDLr and ck expression was demonstrated over a wide range of viral dose. In vivo 31P MRS was employed to detect the metabolic product (i.e., phosphocreatine) of the creatine kinase protein (CK) reaction. CK activity, which is a true measure of ck gene expression, was quantified in vivo by magnetization transfer. Because ck is expressed abundantly in human muscle and brain but is absent from the liver, ck is useful to monitor any liver directed gene transfer. Use of the ck reporter would facilitate the clinical translation of gene therapy by providing a nondestructive readout of the level and duration of therapeutic gene expression.
Li
Zijun
Z
Department of Radiology, School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA.
Qiao
Hui
H
Lebherz
Corinna
C
Choi
Seok Rye
SR
Zhou
Xiangyang
X
Gao
Guanping
G
Kung
Hank F
HF
Rader
Daniel J
DJ
Wilson
James M
JM
Glickson
Jerry D
JD
Zhou
Rong
R
D000256:Adenoviridae / Q000235:genetics; D000818:Animals; D003402:Creatine Kinase / Q000235:genetics* / Q000378:metabolism; D005240:Feasibility Studies; D015870:Gene Expression; D017930:Genes, Reporter; D005819:Genetic Markers; D008099:Liver / Q000378:metabolism*; D009682:Magnetic Resonance Spectroscopy / Q000379:methods; D051379:Mice; D011973:Receptors, LDL / Q000235:genetics*; D014161:Transduction, Genetic
D005819:Genetic Markers; D011973:Receptors, LDL; D003402:Creatine Kinase