? Gene therapy trials would benefit from the ability to determine the location, magnitude, and change in magnitude over time of the expression of delivered genes. Thus far, gene transfer efficiency has been evaluated by obtaining tissue biopsies at predetermined times post treatment. This method of determining gene transfer efficiency is undesirable due to its invasiveness and its inability to generate a global picture of gene transfer, since it is limited to the small piece of tissue(s) examined. Numerous groups have been developing methods for non-invasively imaging gene transfer. We have utilized the human somatostatin receptor subtype 2 (hSSTR2) as a reporter gene along with various radiolabeled somatostatin analogs for gamma ray imaging of gene transfer. However, this system is limited by 1) overexpression of hSSTR2 may have an impact on cell physiology and homeostasis; 2) hSSTR2 is endogenously expressed on certain human tissues; and 3) somatostatin analogs are eliminated through the kidneys which can interfere with signal detection. Therefore, we hypothesize that the hSSTR2 reporter system can be improved through modification of the receptor itself by uncoupling the receptor from G proteins and through the development of novel targeting ligands that recognize a novel epitope inserted into hSSTR2.
The first Aim will investigate the ability of somatostatin analogs radiolabeled with positron emitters to quantify different levels of hSSTR2 expression, in vitro and in vivo.
The second Aim will construct diabodies and minibodies targeted toward an alternative epitope inserted into hSSTR2 and evaluate these constructs in a manner similar to the somatostatin analogs in Aim 1. These first two Aims are intended to determine the most optimal radiopharmaceutical as a reporter probe for PET imaging.
The third Aim will focus on making and evaluating mutants of hSSTR2 that uncouple the receptor from G proteins.
The final Aim will use the best mutant hSSTR2 (developed in Aim 3) and the most optimal radiopharmaceutical to determine the activity and efficacy of a therapeutic gene. This is intended to demonstrate that the system can be used to non-invasively detect therapeutic gene transfer through PET imaging. This proposal should result in improvement of the existing hSSTR2 reporter system for imaging gene transfer. ? ? ?
|Lears, K A; Parry, J J; Andrews, R et al. (2015) Adenoviral-mediated imaging of gene transfer using a somatostatin receptor-cytosine deaminase fusion protein. Cancer Gene Ther 22:215-21|
|Parry, Jesse J; Chen, Ronald; Andrews, Rebecca et al. (2012) Identification of critical residues involved in ligand binding and G protein signaling in human somatostatin receptor subtype 2. Endocrinology 153:2747-55|
|Chen, Ron; Parry, Jesse J; Akers, Walter J et al. (2010) Multimodality imaging of gene transfer with a receptor-based reporter gene. J Nucl Med 51:1456-63|
|Eiblmaier, Martin; Andrews, Rebecca; Laforest, Richard et al. (2007) Nuclear uptake and dosimetry of 64Cu-labeled chelator somatostatin conjugates in an SSTr2-transfected human tumor cell line. J Nucl Med 48:1390-6|