A prototype transduction tag system (delta LNGFR/NGF) for noninvasive clinical gene therapy monitoring.
The dramatic expansion of clinical gene therapy trials requires the development of noninvasive clinical monitoring procedures, which provide information about expression levels, expression kinetics, and spatial distribution of transduced therapeutic genes. With the development of such procedures, invasive sampling of tissue probes from patients potentially could be reduced significantly. In this study, an experimental platform for the rational design and in vitro testing of suitable receptor-ligand couples as components of future transduction tag systems for noninvasive gene therapy monitoring applications was developed. Initially, the feasibility of the delta LNGFR/nerve growth factor (NGF) transduction tag system was investigated; this system employs a mutated version of the low-affinity nerve growth factor receptor (p75mut or delta LNGFR) lacking the entire cytoplasmic domain. Specific binding of 125I-radiolabeled NGF was demonstrated for two stable delta LNGFR-transduced cell lines, but not for delta LNGFR-negative parental control cell lines. An additional binding analysis performed in a MicroImager directly confirmed binding of radiolabeled ligands (125I-NGF, 125I-anti-p75 monoclonal antibody) to the p75mut expressed on intact target cells, but not on control cells. Subsequent binding studies employing NGF radiolabeled with the positron-emitting isotope 124I demonstrated a specific binding for LNGFR+ PC12 cells. Consequently, the first in vitro proof of a transduction tag approach based on the specificity of the 124I-NGF/LNGFR interaction was provided, which opens up the possibility for future noninvasive positron emission tomography monitoring in clinical gene therapy trials.
Lauer
U M
UM
Internal Medicine I, Medical University Clinic Tübingen, Germany. ulrich.lauer@uni-tuebingen.de
Staehler
P
P
Lambrecht
R M
RM
Oberdorfer
F
F
Spiegel
M
M
Wybranietz
W A
WA
Gross
C D
CD
Gregor
M
M
D016475:3T3 Cells; D000818:Animals; D000911:Antibodies, Monoclonal / Q000378:metabolism; D001666:Binding Sites, Antibody; D002460:Cell Line; D015972:Gene Expression Regulation, Neoplastic; D005819:Genetic Markers; D015316:Genetic Therapy / Q000379:methods*; D007457:Iodine Radioisotopes / Q000378:metabolism; D051379:Mice; D020932:Nerve Growth Factor / Q000096:biosynthesis / Q000235:genetics*; D016716:PC12 Cells; D051381:Rats; D020800:Receptor, Nerve Growth Factor / Q000096:biosynthesis / Q000235:genetics* / Q000276:immunology; D012190:Retroviridae / Q000235:genetics*; D017384:Sequence Deletion / Q000235:genetics; D014055:Tomography, Emission-Computed / Q000379:methods; D014161:Transduction, Genetic
D000911:Antibodies, Monoclonal; D005819:Genetic Markers; D007457:Iodine Radioisotopes; D020800:Receptor, Nerve Growth Factor; D020932:Nerve Growth Factor