Tracking of multimodal therapeutic nanocomplexes targeting breast cancer in vivo.

Nanoparticle-based therapeutics with local delivery and external electromagnetic field modulation holds extraordinary promise for soft-tissue cancers such as breast cancer; however, knowledge of the distribution and fate of nanoparticles in vivo is crucial for clinical translation. Here we demonstrate that multiple diagnostic capabilities can be introduced in photothermal therapeutic nanocomplexes by simultaneously enhancing both near-infrared fluorescence and magnetic resonance imaging (MRI). We track nanocomplexes in vivo, examining the influence of HER2 antibody targeting on nanocomplex distribution over 72 h. This approach provides valuable, detailed information regarding the distribution and fate of complex nanoparticles designed for specific diagnostic and therapeutic functions.

HIV-based vectors and angiogenesis following rabbit hindlimb ischemia.

BACKGROUND: Numerous medical and surgical options exist for the treatment of vessel ischemia, which some patients fail or cannot tolerate. These investigations were designed to determine the effects of lentiviral-delivered vascular endothelial-derived growth factor (VEGF) and angiopoietin-2 (Ang-2) on collateralization in a rabbit model of hindlimb ischemia. MATERIALS AND METHODS: Self-inactivating human immunodeficiency virus (HIV)-based vectors were constructed encoding VEGF or Ang-2, co-transfected with vesicular stomatitis virus glycoprotein (VSV G) into 293T cells, and vector supernatants (1 x 10(8) IU/ml after concentration) were harvested. New Zealand white rabbits had ligation of either the right or left external iliac artery and excision of the ipsilateral femoral artery. Ten days later, empty, VEGF, or VEGF+Ang-2 vector supernatant was injected intramuscularly (IM) into the ipsilateral thigh. Ankle systolic blood pressure (SBP) ratios were recorded and venous blood samples collected on postoperative days (POD) 10, 25, and 40. On POD 40, run-off angiography was performed to measure vessel collateralization. Capillary density was determined by thin sectioning of muscle. RESULTS: A significant increase was noted in SBP in the VEGF-treated animals over time. Capillary density was not elevated despite significantly increased large vessel collateralization in rabbits receiving VEGF, which was counteracted by Ang-2. Antibodies against vector components were detected in exposed serum. CONCLUSIONS: Arterial collateralization and SBP increased significantly following VEGF vector administration, which was reversed by the Ang-2 vector. Development of antibody against VSV G can limit repeated injections of vector. Future experiments will involve the addition of other pro-angiogenic factors, repeated vector administration, and alternative routes of vector delivery.

Identification of synthetic endothelial cell-specific promoters by use of a high-throughput screen.

Transcriptional targeting is a desirable property for many gene transfer applications. Because endothelial cells line most blood vessels, they are attractive candidates for the introduction of therapeutic gene products. As a proof-of-concept study, we attempted to identify a synthetic, endothelial cell-specific promoter by use of a high-throughput screen involving self-inactivating (SIN) human immunodeficiency virus type 1 (HIV-1)-based vectors. Select duplex oligodeoxynucleotides recognized by transcription factors and located 5' of endothelial cell-specific mRNA transcripts were randomly ligated and cloned upstream of a minimal ICAM-2 promoter driving enhanced green fluorescent protein (eGFP) in a SIN HIV-1-based vector. Vesicular stomatitis virus G protein-pseudotyped particles were prepared from a library of >10(6) vector recombinants and used to transduce an endothelial cell line. The highest eGFP expressers were repeatedly sorted, and the synthetic promoters were recovered and retested by a luciferase reporter. Several promoters were active and specific to endothelial cells of varied species, with high selectivity indexes and inducibility under hypoxia-mimetic conditions. One in particular was then introduced back into a SIN HIV-1-based vector to confirm its endothelial cell activity and specificity. This study suggests that SIN vectors may be used in a high-throughput manner to identify tissue-specific promoters of high activity, with potential applications for both transcriptional targeting and gene transfer.

Knockout mouse model for Fxr2: a model for mental retardation.

Fragile X syndrome is a common form of mental retardation caused by the absence of the FMR1 protein, FMRP. Fmr1 knockout mice exhibit a phenotype with some similarities to humans, such as macro-orchidism and behavioral abnormalities. Two homologs of FMRP have been identified, FXR1P and FXR2P. These proteins show high sequence similarity, including all functional domains identified in FMRP, such as RNA binding domains. They have an overlap in tissue distribution to that of FMRP. Interactions between the three FXR proteins have also been described. FXR2P shows high expression in brain and testis, like FMRP. To study the function of FXR2P, we generated an Fxr2 knockout mouse model. No pathological differences between knockout and wild-type mice were found in brain or testis. Given the behavioral phenotype in fragile X patients and the phenotype previously reported for the Fmr1 knockout mouse, we performed a thorough evaluation of the Fxr2 knockout phenotype using a behavioral test battery. Fxr2 knockout mice were hyperactive (i.e. traveled a greater distance, spent more time moving and moved faster) in the open-field test, impaired on the rotarod test, had reduced levels of prepulse inhibition, displayed less contextual conditioned fear, impaired at locating the hidden platform in the Morris water task and were less sensitive to a heat stimulus. Interestingly, there are some behavioral phenotypes in Fxr2 knockout mice which are similar to those observed in Fmr1 knockout mice, but there are also some different behavioral abnormalities that are only observed in the Fxr2 mutant mice. The findings implicate a role for Fxr2 in central nervous system function.