Gene expression analysis of SCC tumor cells in muscle tissue.

The purpose of this study was to evaluate microarray technology of HNSCC cells in muscle tissue. 200 SCCVII tumor cells were injected intramuscularly into the right flank of ten C3H/Km mice each. One week later the animals were killed and the tissue taken out. Histology (H&E staining) and microarray of the tissue were performed. Histology showed a few tumor cells between the muscle fibers. Microarray technology showed different gene expression pattern of the muscle tissue with SCCVII cells in comparison with normal muscle tissue. Only those genes showing a fold change difference of 5 or higher were considered. Gene expression analysis revealed changes in the expression levels of SCCVII cells in muscle tissue in 220 genes. Significant gene expression differences between SCCVII cells in muscle tissue and pure muscle tissue could be seen.

Effect of continuous high intensity focused ultrasound in a squamous cell carcinoma tumor model compared to muscle tissue evaluated by MRI, histology, and gene expression.

The purpose of this study was to investigate the effect of the continuous mode of high intensity focused ultrasound (HIFU) in a mouse head and neck cancer model (SCCVII) compared to muscle tissue. HIFU was applied to SCCVII tumors and to muscle tissue in C3H/Km mice using a dual ultrasound system (imaging 6 MHz/therapeutic 1 MHz). A continuous HIFU mode (total time 20 sec, intensity 6730.6 W/cm(2)) was applied. Three hours after HIFU treatment pre- and post-contrast T1-wt, T2-wt images, and a diffusion-wt STEAM sequence were obtained. After MR imaging, the animals were euthenized and the treated tumor and muscle tissue was taken out for histology and functional genomic analysis. T2 images showed increased signal intensity, post-contrast T1 showed a decreased contrast uptake in the central parts in the tumor tissue as well as in the muscle tissue. In addition a significant higher diffusion coefficient was found in both tissue types. Histological evaluation (H&E, Immunohistochemistry) of the tumors and the muscle tissue revealed areas of significant necrosis. In the tumor tissue 23 genes were up-regulated (> 2 fold change) and 4 genes were down-regulated (< -2 fold change). In the muscle tissue 29 genes were up-regulated and 17 genes down-regulated. Thirteen genes were up-regulated in both tissue types, 8 genes only in the SCCVII tissue, and 11 genes only in the muscle tissue. The use of HIFU treatment on tumor and muscle tissue results in dramatic changes in gene expression. The expression of some genes are tissue specific, the expression of other genes are independent of the tissue type.

Gene expression profiles, histologic analysis, and imaging of squamous cell carcinoma model treated with focused ultrasound beams.

OBJECTIVE: The purpose of our study was to evaluate the effect of short-pulse high-intensity focused ultrasound (HIFU) on inducing cell death in a head and neck cancer model (SCCVII [squamous cell carcinoma]) compared with continuous HIFU to get a better understanding of the biologic changes caused by HIFU therapy. MATERIALS AND METHODS: HIFU was applied to 12 SCCVII tumors in C3H/Km mice using a dual sonography system (imaging, 6 MHz; therapeutic, 1 MHz). A continuous HIFU mode (total time, 20 seconds; intensity, 6,730.6 W/cm2) and a short-pulse HIFU mode (frequency, 0.5 Hz; pulse duration, 50 milliseconds; total time, 16.5 minutes; intensity, 134.4 W/cm2) was applied. Three hours later, MR images were obtained on a 1.5-T scanner. After imaging, the treated and untreated control tumor tissue samples were taken out for histology and oligonucleotide microarray analysis. RESULTS: Prominent changes were observed in the MR images in the continuous HIFU mode, whereas the short-pulse HIFU mode showed no discernible changes. Histology (H and E, TUNEL [terminal deoxynucleotidyl transferase-mediated dUTP {deoxyuridine triphosphate} nick-end labeling], and immunohistochemistry) of the tumors treated with the continuous HIFU mode revealed areas of significant necrosis. In the short-pulse HIFU mode, the H and E staining showed multifocal areas of coagulation necrosis. TUNEL staining showed a high apoptotic index in both modes. Gene expression analysis revealed profound differences. In the continuous HIFU mode, 23 genes were up-regulated (> twofold change) and five genes were down-regulated (< twofold change), and in the short-pulse HIFU mode, 32 different genes were up-regulated and 16 genes were down-regulated. CONCLUSION: Genomic analysis might be included when investigating tissue changes after interventional therapy because it offers the potential to find molecular targets for imaging and therapeutic applications.

In vitro effect of focused ultrasound or thermal stress on HSP70 expression and cell viability in three tumor cell lines.

RATIONALE AND OBJECTIVES: In this study, we compared the effect of focused ultrasound with the effect of thermal stress on the induction of a heat inducible promoter in an in vitro model using three tumor cell lines (M21, SCCVII, and NIH3T3). MATERIALS AND METHODS: We used a reporter construct that was generated using the stress-inducible promoter from the gene encoding a murine 70-kilodalton heat shock protein (Hsp70A.1) and a luciferase (luc) reporter plasmid. High-intensity focused ultrasound (HIFU) was applied in two different modes. In the first mode, an increasing voltage at constant pulse duration and in the second mode a constant voltage at increasing pulse duration was applied. HIFU or thermal stress was delivered over a range of temperatures (36-52 degrees C) for 5 minutes, and resulting luciferase activity was measured in live cells using a cooled charge-coupled device camera as a measure of reporter gene transcription. Luciferase activity was measured at set time intervals for a total of 108 hours post-stress. RESULTS: Both methods induced the hsp70 promoter; however, the luciferase activity under the influence of HIFU, independent of the applied mode, and thermal stress differs despite the fact that the temperature was the same. In the M21 tumor cell line, the maximum luciferase activity after focused ultrasound application was 4818 +/- 1521% at a temperature of 48 degrees C and after thermal stress 4468.2 +/- 1890.2% at a temperature of 52 degrees C with a viability of 72.3 +/- 5.2% and 85 +/- 3.4%, respectively. In the SCC tumor cell line, the maximum luciferase activity after focused ultrasound application was 6743.0 +/- 3281.4% and after only thermal stress exposure was 3910.6 +/- 2189.0% at a temperature of 44 degrees C and 50 degrees C, respectively. At the highest luciferase activity, the portion of vital cells was 72.5 +/- 8.4% and 72.5 +/- 5.9% respectively. In the NIH3T3 tumor cell line the highest luciferase activity of 428510.6 +/- 26526.8% was seen at a temperature of 42 degrees C applying focused ultrasound. Under thermal stress it was 29221.3 +/- 7205.0% at a temperature of 50 degrees C. At the highest luciferase activity, the viability analysis showed 75.3 +/- 9.2% and 72.3 +/- 7.9% viable cells, respectively. CONCLUSIONS: Focused ultrasound induces hsp70 expression like thermal stress alone; however, HIFU is capable of inducing expression at lower temperatures than heat stress alone, indicating that nonthermal effects also play a role on the induction of hsp70.

Angiogenesis is required for successful bone induction during distraction osteogenesis.

UNLABELLED: The role of angiogenesis during mechanically induced bone formation is incompletely understood. The relationship between the mechanical environment, angiogenesis, and bone formation was determined in a rat distraction osteogenesis model. Disruption of either the mechanical environment or endothelial cell proliferation blocked angiogenesis and bone formation. This study further defines the role of the mechanical environment and angiogenesis during distraction osteogenesis. INTRODUCTION: Whereas successful fracture repair requires a coordinated and complex transcriptional program that integrates mechanotransductive signaling, angiogenesis, and osteogenesis, the interdependence of these processes is not fully understood. In this study, we use a system of bony regeneration known as mandibular distraction osteogenesis (DO) in which a controlled mechanical stimulus promotes bone induction after an osteotomy and gradual separation of the osteotomy edges to examine the relationship between the mechanical environment, angiogenesis, and osteogenesis. MATERIALS AND METHODS: Adult Sprague-Dawley rats were treated with gradual distraction, gradual distraction plus the angiogenic inhibitor TNP-470, or acute distraction (a model of failed bony regeneration). Animals were killed at the end of distraction (day 13) or at the end of consolidation (day 41) and examined with muCT, histology, and immunohistochemistry for angiogenesis and bone formation (n = 4 per time-point per group). An additional group of animals (n = 6 per time-point per group) was processed for microarray analysis at days 5, 9, 13, 21, and 41. RESULTS AND CONCLUSIONS: Either TNP-470 administration or disruption of the mechanical environment prevented normal osteogenesis and resulted in a fibrous nonunion. Subsequent analysis of the regenerate showed an absence of angiogenesis by gross histology and immunohistochemical localization of platelet endothelial cell adhesion molecule in the groups that failed to heal. Microarray analysis revealed distinct patterns of expression of genes associated with osteogenesis, angiogenesis, and hypoxia in each of the three groups. Our findings confirm the interdependence of the mechanical environment, angiogenesis, and osteogenesis during DO, and suggest that induction of proangiogenic genes and the proper mechanical environment are both necessary to support new vasculature for bone induction in DO.

A novel antiangiogenesis therapy using an integrin antagonist or anti-Flk-1 antibody coated 90Y-labeled nanoparticles.

PURPOSE: Integrin alpha(v)beta(3) and vascular endothelial growth factor receptor 2 (Flk-1) have been shown to be involved in tumor-induced angiogenesis. Selective targeting of upregulated alpha(v)beta(3) and Flk-1 on the neovasculature of tumors is a novel antiangiogenesis strategy for treating a wide variety of solid tumors. In the studies described here, we investigated the potential therapeutic efficacy of two three-component treatment regimens using two murine tumor models. METHODS AND MATERIALS: The treatment regimens used nanoparticle (NP) based targeting agents radiolabeled with (90)Y. The small molecule integrin antagonist (IA) 4-[2-(3,4,5,6-tetrahydropyrimidin-2-ylamino)ethoxy]-benzoyl-2-(5)-aminoethylsulfonylamino-beta-alanine, which binds to the integrin alpha(v)beta(3), and a monoclonal antibody against murine Flk-1 (anti-Flk-1 MAb) were used to target the NPs. Murine tumor models K1735-M2 (melanoma) and CT-26 (colon adenocarcinoma) were used to evaluate the treatment efficacy. RESULTS: In K1735-M2 and CT-26 tumors, a single treatment with IA-NP-(90)Y (14.2 microg/g IA, 5 or 6 microCi/g (90)Y) caused a significant tumor growth delay compared to untreated control tumors, as well as tumors treated with IA, IA-NP, and NP-(90)Y, respectively (p < 0.025, Wilcoxon test). In K1735-M2 tumors, a single treatment with anti-Flk-1 MAb-NP-(90)Y (0.36 microg/g anti-Flk-1 MAb, 5 microCi/g (90)Y) also caused a significant tumor growth delay (p < 0.05, Wilcoxon test) compared to untreated tumors, as well as tumors treated with anti-Flk-1 MAb, anti-Flk-1 MAb-NP, and conventional radioimmunotherapy with (90)Y-labeled anti-Flk mAb. Anti-CD31 staining showed a marked decrease in vessel density in tumors treated with anti-Flk-1 MAb-NP-(90)Y, which was associated with a high level of apoptotic death in these tumors, as shown by terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling staining. CONCLUSIONS: The present studies provide proof of principle that targeted radiotherapy works using different targeting agents on a nanoparticle, to target both the integrin alpha(v)beta(3) and the vascular endothelial growth factor receptor. These encouraging results demonstrate the potential therapeutic efficacy of the IA-NP-(90)Y and anti-Flk-1 MAb-NP-(90)Y complexes as novel therapeutic agents for the treatment of a variety of tumor types.

Comparing genomic and histologic correlations to radiographic changes in tumors: a murine SCC VII model study.

RATIONALE AND OBJECTIVES: To investigate the correlation between the temporal changes in T1- and T2-weighted contrast-enhanced magnetic resonance imaging (MRI), histologic evaluation, and genomic analysis using oligonucleotide microarrays in a murine squamous cell carcinoma tumor models. MATERIALS AND METHODS: The squamous cell carcinoma (SCC VII) cell line was used to initiate subcutaneous tumors in mice. This mouse model has been used as a model for human head and neck carcinomas. Animals were imaged using contrast enhanced MRI (CE-MRI). Different stages of tumor growth were defined based on changes in the T1- and T2-weighted MRI patterns. The contrast enhancing (CE) and nonenhancing (NE) regions of the tumors were marked and biopsied for oligonucleotide microarray and histologic analysis. Tumors with no differential contrast enhancement were used as controls. RESULTS: Distinct temporal stages of tumor progression can be defined using both T1- and T2-weighted CE-MRI and microarray analysis. The early stage tumors show a homogeneous contrast enhancement pattern in the T1- and T2-weighted images with no significant differential gene expression from the center and periphery of the tumor. The more advanced tumors that show discrete regions of contrast enhancement in the post-contrast T1-weighted MRIs and tissues from the CE and NE regions show distinctly differential gene expression profiles. Histologic analysis (hematoxylin-eosin stain) showed that the samples obtained from the periphery and center of the early stage tumors and the CE and NE regions from these more advanced tumors were similar. The gene expression profiles of late-stage tumors that showed changes in T2-weighted MRI signal intensity were consistent with tissue degradation in the NE region, which also showed characteristic signs of tissue necrosis in histologic analysis. CONCLUSION: These results show that temporal changes in T1- and T2-weighted CE-MRI are related to distinct gene expression profiles, and histologic analysis may not be sufficient to detect these detailed changes. As tumors progress, discrete regions of post-contrast T1 enhancement are identified; these regions have distinct gene expression patterns despite similar histologic features. In late-stage tumors, regions of T2 signal changes are observed which correspond with tissue necrosis.

Molecular imaging and therapy directed at the neovasculature in pathologies. How imaging can be incorporated into vascular-targeted delivery systems to generate active therapeutic agents.

We have discussed the impact of molecular imaging on clinical and preclinical medicine. We have presented the potential problems of delivering the effective therapeutic dose and the properties that can help contribute to the drug efficacy. The rationale for the design of new antiangiogenic agents that can be used for imaging and therapy was presented. Finally, results from imaging and targeted nanoparticle based therapies were presented. In vivo imaging of angiogenic tumors using anti-alpha(v)beta3 -targeted polymerized vesicles composed of the murine antibody LM609 attached to NPs labeled with the MR contrast agent gadolinium in the V2 carcinoma model in rabbits. MRI studies using this targeted contrast agent revealed large areas of alpha(v)beta3 integrin expression in tumor-associated vasculature that conventional MRIs failed to show. Other investigators have used microemulsions conjugated to an antibody targeted against alpha(v)beta as imaging agents. These materials also show contrast enhancement of tumor vasculature undergoing angiogenesis. Other markers, such as the PECAM-1 (CD-31), VCAM-1 (CD54) and VEGF receptor (flk-1), have been shown to be upregulated on tumor endothelium and associated with angiogenesis but have not been used in imaging studies. Furthermore, by modification of the NPs, we were able to use this imaging agent as an antiangiogenic gene delivery system. The results from these studies are very promising and are being further pursued.

Mechanic effect of pulsed focused ultrasound in tumor and muscle tissue evaluated by MRI, histology, and microarray analysis.

The purpose of this study was to investigate the effect of pulsed high-intensity focused ultrasound (HIFU) to tumor and muscle tissue. Pulsed HIFU was applied to tumor and muscle tissue in C3H/Km mice. Three hours after HIFU treatment pre- and post-contrast T1-wt, T2-wt images and a diffusion-wt STEAM-sequence were obtained. After MR imaging, the animals were euthenized and the treated tumor and muscle was taken out for histology and functional genomic analysis. In the tumor tissue a slight increase of the diffusion coefficient could be found. In the muscle tissue T2 images showed increased signal intensity and post-contrast T1 showed a decreased contrast uptake in the center and a severe contrast uptake in the surrounding muscle tissue. A significant increase of the diffusion coefficient was found. Gene expression analysis revealed profound changes in the expression levels of 29 genes being up-regulated and 3 genes being down-regulated in the muscle tissue and 31 genes being up-regulated and 15 genes being down-regulated in the SCCVII tumor tissue. Seven genes were up-regulated in both tissue types. The highest up-regulated gene in the tumor and muscle tissue encoded for Mouse histone H2A.1 gene (FC=13.2±20.6) and Apolipoprotein E (FC=12.8±27.4) respectively MHC class III (FC=83.7±67.4) and hsp70 (FC=75.3±85.0). Immunoblot confirmed the presence of HSP70 protein in the muscle tissue. Pulsed HIFU treatment on tumor and muscle tissue results in dramatic changes in gene expression, indicating that the effect of pulsed HIFU is in some regard dependent and also independent of the tissue type.

Modulation of luciferase activity using high intensity focused ultrasound in combination with bioluminescence imaging, magnetic resonance imaging and histological analysis in muscle tissue.

This study investigates the effect of high intensity focused ultrasound (HIFU) to muscle tissue transfected with a luciferase reporter gene under the control of a CMV-promoter. HIFU was applied to the transfected muscle tissue using a dual HIFU system. In a first group four different intensities (802 W/cm2, 1401 W/cm2, 2117 W/cm2, 3067 W/cm2) of continuous HIFU were applied 20 s every other week for four times. In a second group two different intensities (802 W/cm2, 1401 W/cm2) were applied 20 s every fourth day for 20 times. The luciferase activity was determined by bioluminescence imaging. The effect of HIFU to the muscle tissue was assessed by T1-weighted +/- Gd-DTPA, T2-weighted and a diffusion-weighted STEAM sequence obtained on a 1.5-T GE-MRI scanner. Histology of the treated tissue was done at the end. In the first group the photon emission was at 3067.6 W/cm2 1.28 x 10(7) +/- 3.1 x 10(6) photon/s (5.5 +/- 1.2-fold), of 2157.9 W/cm2 8.1 +/- 2.7 x 10(6) photon/s (3.2 +/- 1.1-fold), of 1401.9 W/cm2 9.3 +/- 1.3 x 10(6) photon/s (4.9 +/- 0.4-fold) and of 802.0 W/cm2 8.6x +/- 1.2 x 10(6) photon/s (4.5 +/- 0.6-fold) compared to baseline. In the second group the photon emission was at 1401.9 W/cm2 and 802.0 W/cm2 14.1 +/- 3.6 x 10(6) photon/s (6.1 +/- 1.5-fold), respectively, 5.1 +/- 4.7 x 10(6) photon/s (6.5 +/- 2.0-fold). HIFU can enhance the luciferase activity controlled by a CMV-promoter.

Tumor tissue characterization evaluating the luciferase activity under the control of a hsp70 promoter and MR imaging in three tumor cell lines.

We investigated the luciferase activity under the control of a hsp70 promoter and MR imaging for three tumor cell lines. Three tumor cell lines, SCCVII, NIH3T3 and M21 were transfected with a plasmid containing the hsp70 promoter fragment and the luciferase reporter gene and grown in mice. Bioluminescence imaging of the tumors was performed every other day. MR imaging, pre- and post-contrast T1-wt SE, T2-wt FSE, Diffusion-wt STEAM-sequence, T2-time determination were obtained on a 1.5-T GE MRI scanner at a tumor size of 600-800 mm(3) and 1400-1600 mm(3). Comparing the different tumor sizes the luciferase activity of the M21 tumors increased about 149.3%, for the NIH3T3 tumors about 47.4% and for the SCCVII tumors about 155.8%. Luciferase activity of the M21 tumors (r=0.82, p<0.01) and the SCCVII tumors (r=0.62, p=0.03) correlated significant with the diffusion coefficient. In the NIH3T3 tumors the best correlation between the luciferase activity and the MRI parameter was seen for the SNR (T2) values (r=0.78, p<0.01). The luciferase activity per mm(3) tumor tissue correlated moderate with the contrast medium uptake (r=0.55, p=0.01) in the M21 tumors. In the NIH3T3 and SCCVII tumors a negative correlation (r=-0.78, p<0.01, respectively, r=-0.49, p=0.02) was found with the T2 time. Different tissue types have different luciferase activity under the control of the same hsp70 promoter. The combination of MR imaging with bioluminescence imaging improves the characterization of tumor tissue giving better information of this tissue on the molecular level.

The effect of high intensity focused ultrasound on luciferase activity on two tumor cell lines in vitro, under the control of a CMV promoter.

In this study, we compared the effect of high intensity focused ultrasound (HIFU) and thermal stress on the luciferase activity, controlled by a cytomegaly virus (CMV) promoter in an in vitro model using two tumor cell lines (M21, SCCVII). HIFU was applied in a pulsed-wave mode with increasing voltage at constant pulse duration, or thermal stress was delivered over a range of temperatures (36-52 degrees C) for 5 min. The resulting luciferase activity was measured in live cells using a cooled CCD camera. Luciferase activity was measured at set time intervals over a total of 48 h post-stress. Compared to baseline, the luciferase activity of the M21 tumor cell line when exposed to HIFU was approximately 54.2+/-67.5% (p<0.01) higher at a temperature of 42 degrees C, and approximately 52.9+/-128.5% (p<0.01) higher at 44 degrees C. In the SCCVII tumor cell line, the luciferase activity after HIFU application was 55.4+/-66.6% (p<0.01) higher compared to baseline at a temperature of 42 degrees C. The M21 and SCCVII tumor cell line when exposed to thermal stress alone did not increase the luciferase activity. M21 and SCCVII tumor cells exposed to HIFU showed a maximum decrease in cell viability to 45.3+/-7.5% and 10.3+/-7.5%, respectively, and when exposed to thermal stress to 85.3+/-3.5% and 20.4+/-6.5%, respectively, compared to the untreated control. In M21 and SCCVII cells exposed to HIFU, free radicals could be detected using the dichlorofluorescein dye. Our findings demonstrate that HIFU can enhance the luciferase activity controlled by a CMV promoter. However it also has a higher damaging effect on the cells.

Comparison of continuous vs. pulsed focused ultrasound in treated muscle tissue as evaluated by magnetic resonance imaging, histological analysis, and microarray analysis.

The purpose of this study was to investigate the effect of different application modes of high intensity focused ultrasound (HIFU) to muscle tissue. HIFU was applied to muscle tissue of the flank in C3H/Km mice. Two dose regimes were investigated, a continuous HIFU and a short-pulsed HIFU mode. Three hours after HIFU treatment pre- and post-contrast T1-weighted, T2-weighted images and a diffusion-weighted STEAM sequence were obtained. After MR imaging, the animals were euthanized and the treated, and the non-treated tissue was taken out for histology and functional genomic analysis. T2 images showed increased signal intensity and post-contrast T1 showed a decreased contrast uptake in the central parts throughout the tissue of both HIFU modes. A significantly higher diffusion coefficient was found in the muscle tissue treated with continuous wave focused ultrasound. Gene expression analysis revealed profound changes of 54 genes. For most of the analyzed genes higher expression was found after treatment with the short-pulse mode. The highest up-regulated genes encoded for the MHC class III (FC approximately 84), HSP 70 (FC approximately 75) and FBJ osteosarcoma related oncogene (FC approximately 21). Immunohistology and the immunoblot analysis confirmed the presence of HSP70 protein in both applied HIFU modes. The use of HIFU treatment on muscle tissue results in dramatic changes in gene expression; however, the same genes are up-regulated after the application of continuous or pulsed HIFU, indicating that the tissue reaction is independent of the type of tissue damage.

Delivery of systemic chemotherapeutic agent to tumors by using focused ultrasound: study in a murine model.

PURPOSE: To quantitatively determine the delivery of systemic liposomal doxorubicin to tumors treated with pulsed high-intensity focused ultrasound and to study the mechanism underlying this delivery in a murine model. MATERIALS AND METHODS: All animal work was performed in compliance with guidelines and approval of institutional animal care committee. C3H mice received subcutaneous injections in the flank of a cell suspension of SCC7, a murine squamous cell carcinoma cell line; mice (n = 32) in drug delivery study received unilateral injections, whereas mice (n = 10) in mechanistic study received bilateral injections. Tumors were treated when they reached 1 cm(3) in volume. In the drug delivery study, doxorubicin hydrochloride liposomes were injected into the tail vein: Mice received therapy with doxorubicin injections and high-intensity focused ultrasound, doxorubicin injections alone, or neither form of therapy (controls). Tumors were removed, and the doxorubicin content was assayed with fluorescent spectrophotometry. In the mechanistic study, all mice received an injection of 500-kDa dextran-fluorescein isothyocyanate into the tail vein, and half of them were exposed to high-intensity focused ultrasound prior to injection. Contralateral tumors served as controls for each group. Extravasation of dextran-fluorescein isothyocyanate was observed by using in vivo confocal microscopy. RESULTS: Mean doxorubicin concentration in tumors treated with pulsed high-intensity focused ultrasound was 9.4 microg . g(-1) +/- 2.1 (standard deviation), and it was significantly higher (124% [9.4 microg . g(-1)/4.2 microg . g(-1)]) than in those that were not treated with high-intensity focused ultrasound (4.2 microg . g(-1) +/- 0.95) (P < .001, unpaired two-tailed Student t test). Extravasation of dextran-fluorescein isothyocyanate was observed in the vasculature of tumors treated with high-intensity focused ultrasound but not in that of untreated tumors. CONCLUSION: Pulsed high-intensity focused ultrasound is an effective method of targeting systemic drug delivery to tumor tissue. Potential mechanisms for producing the observed enhancement are discussed.

Combined vascular targeted imaging and therapy: a paradigm for personalized treatment.

In order to be successful in personalizing treatment, methods for selecting patients as well as good surrogate biomarkers for monitoring the effects of treatment are required in addition to development of an efficacious targeted therapy. We have developed a polymerized nanoparticle platform technology that will allow us to put different targeting moieties on the surface of the particles in addition to loading the particles with different contrast and therapeutic agents. We have proven that these nanoparticles can be targeted to endothelial receptors and different payloads of contrast and therapeutic agents have been delivered to target cells with high target to background ratios. Using this combined vascular targeted imaging and therapy approach, we are optimistic that personalized treatment regimens can be developed for different disease processes such as cancer, inflammation, and ischemia.

Vascular-targeted molecular imaging using functionalized polymerized vesicles.

In this review we will discuss the use of multivalent polymerized vesicles (PVs) combined with magnetic resonance imaging (MRI) and gamma scintigraphy to image expression of vascular molecular receptors in vivo. Specifically, we will present our data on the use of this technology to design imaging agents toward specific vascular receptors in both a mouse and rabbit tumor model and in the EAE mouse, a model for human multiple sclerosis (MS). Examples will be shown where the in vivo specificity of the targeted molecular imaging agents was validated in the animal models. Since the PVs are designed to carry either contrast or therapeutic agents or both, we can potentially use vascular-targeted imaging for selecting patients and guiding vascular-targeted therapies in these patients. Using this combined vascular-targeted imaging and therapy approach, personalized treatment can potentially be delivered, maximizing efficacy and minimizing side effects.

Synthesis of polymerized thin films for immobilized ligand display in proteomic analysis.

We describe a new material for the display of biomolecular ligands for use in proteomic analysis. We report here on the construction of the first functionalized polymerized diacetylene thin films (PDTFs) for use in displaying immobilized ligands and their application in mass spectral proteomic analysis. Functionalized polymerized thin film surfaces were constructed with diacetylene-containing biotin lipid monomers designed for the capture of proteins (streptavidin) from a complex cellular lysate and detection with mass spectrometry (MS). These materials serve as a prototype for ligand-based spotted arrays amenable to high throughput screening. Functionalized PDTFs can be easily manufactured for customized microarrays and demonstrate high protein specificity and low nonspecific protein adsorption, and the resulting microarrays constructed from these materials are compatible with several different protein analysis platforms. Our results suggest that these materials have broad potential applications for use in mass spectral-based proteomic analysis.

Molecular imaging applications in nanomedicine.

The purpose of this article is to explore how molecular imaging techniques can be used as useful adjunts in the development of "nanomedicine" and in personalizing treatment of patients. The discussion focuses on in vivo applications at the whole organism level even though imaging can also play an important role in research at the cellular and subcellular level.