19F-NMR detection of lacZ gene expression via the enzymic hydrolysis of 2-fluoro-4-nitrophenyl beta-D-galactopyranoside in vivo in PC3 prostate tumor xenografts in the mouse.
Gene therapy shows promise for treating prostate cancer and has been evaluated in several clinical trials. A major challenge that remains is to establish a method for verifying transgene activity in situ. The lacZ gene encoding beta-galactosidase historically has been the most popular reporter gene for molecular biology. We have designed a 19F NMR approach to reveal lacZ gene expression by assessing beta-galactosidase (beta-gal) activity in vivo. The substrate 2-fluoro-4-nitrophenyl beta-D-galactopyranoside (OFPNPG) is readily hydrolyzed by beta-gal with a corresponding decrease in the 19F-NMR signal from OFPNPG and the appearance of a new signal shifted 4-6 ppm upfield from the aglycone 2-fluoro-4-nitrophenol (OFPNP). We report proof of principle in cultures of PC3 prostate cancer cells using 19F NMR spectroscopy and 19F chemical shift imaging. More importantly, we demonstrate for the first time the ability to differentiate wild-type and lacZ-expressing prostate tumor xenografts in mice using this approach.
Liu
Li
L
Department of Radiology, The University of Texas Southwestern Medical Center at Dallas, 5323 Harry Hines Blvd, Dallas, Texas 75390-9058, USA.
Kodibagkar
Vikram D
VD
Yu
Jian-Xin
JX
Mason
Ralph P
RP
D000230:Adenocarcinoma / Q000473:pathology; D000818:Animals; D045744:Cell Line, Tumor; D004790:Enzyme Induction; D005461:Fluorine; D017930:Genes, Reporter; D015316:Genetic Therapy / Q000379:methods; D006801:Humans; D006868:Hydrolysis; D007763:Lac Operon; D009682:Magnetic Resonance Spectroscopy / Q000379:methods*; D008297:Male; D051379:Mice; D009596:Nitrophenols / Q000032:analysis; D009598:Nitrophenylgalactosides / Q000378:metabolism*; D011471:Prostatic Neoplasms / Q000473:pathology; D011993:Recombinant Fusion Proteins / Q000032:analysis / Q000235:genetics / Q000378:metabolism; D013379:Substrate Specificity; D014162:Transfection; D014183:Transplantation, Heterologous; D001616:beta-Galactosidase / Q000032:analysis* / Q000235:genetics / Q000378:metabolism
C521953:2-fluoro-4-nitrophenyl galactopyranoside; D009596:Nitrophenols; D011993:Recombinant Fusion Proteins; D009598:Nitrophenylgalactosides; D005461:Fluorine; C088999:2-fluoro-4-nitrophenol; D001616:beta-Galactosidase