DNA binding chelates for nonviral gene delivery imaging

Abstract

Noninvasive in vivo monitoring of gene delivery would provide a critically important information regarding the spatial distribution, local concentration, kinetics of removal and/or biodegradation of the expression vector. We developed a novel approach to noninvasive gene delivery imaging using heterobifunctional peptide-based chelates (PBC) bearing double-stranded DNA-binding groups and a technetium-binding amino acid motif. One of such chelates: Gly-Cys(Acm)-Gly-Cys(Acm)-Gly-Lys4- (oxotechnetate). The psoralen moiety (a DNA binding group of PBC) allowed linking to double-stranded DNA upon short-term irradiation with the near UV range light (>320 nm). Approximately 30-40% of added 99mTc-labeled PBC was nonextractable and coeluted with a model pCMV-GFP vector during the gel-permeation chromatography. Nuclear imaging of 'naked' DNA and DNA complexes with lipid-based transfection reagents ('lipoplexes') has been performed after systemic or local administration of 99mTc-PBC-labeled DNA in mice. Imaging results were corroborated with the biodistribution using 99mTc-PBC and 32P-labeled DNA and lipoplexes. A markedly different biodistribution of 99mTc PBC-labeled DNA and lipoplexes was observed with the latter being rapidly trapped in the liver, spleen and lung. 99mTc PBC-DNA was used as an imaging tracer during in vivo transfection of B16 melanoma by local injection of 'naked' 99mTc PBC-DNA and corresponding lipoplexes. As demonstrated by nuclear imaging, 99mTc PBC-DNA lipoplexes showed a slower elimination from the site of injection than 99mTc PBC-DNA alone. This result correlated with a higher expression of marker mRNA and green fluorescent protein as determined using RT-PCR and immunohistochemistry, respectively.