Doxorubicin-loaded hollow gold nanospheres for dual photothermal ablation and chemoembolization therapy.

BACKGROUND: Doxorubicin-loaded hollow gold nanospheres (Dox@HAuNS) are a promising technology for simultaneous trans-arterial tumor-targeted chemotherapy delivery and thermal ablation. We evaluated the efficacy of intra-arterial delivery of Dox@HAuNS followed by photothermal ablation (PTA) in a rabbit model of liver cancer. Adult New Zealand white rabbits (N=25) were inoculated with VX2 tumors into the left lobe of the liver. The animals were then randomized to sham surgery (N=5), PTA only (N=3), Dox@HAuNS only (N=5), HAuNS + PTA (N=5), and Dox@HAuNS + PTA (N=7). Nanoparticles were delivered as an emulsion with Lipiodol (Guerbet, France) via a trans-arterial approach. Following nanoparticle delivery, PTA was performed using an 808nm fibered laser at 1.5W for 3 minutes. Thermography during PTA demonstrated a sustained elevation in tumoral temperature in both HAuNS + laser and Dox@HAuNS + laser treatment groups relative to animals that underwent laser treatment without prior nanoparticle delivery. RESULTS: There was a significant decrease in tumor volumes in all three treatment arms relative to control arms (P = 0.004). Concentrations of intratumoral doxorubicin were significantly greater in animals treated with laser compared to those that were not treated with laser (P< 0.01). CONCLUSIONS: Doxorubicin-loaded HAuNS is a promising therapeutic agent for dual ablation/chemoembolization treatment of liver cancer.

Targeted Drug Delivery and Image-Guided Therapy of Heterogeneous Ovarian Cancer Using HER2-Targeted Theranostic Nanoparticles.

Cancer heterogeneity and drug resistance limit the efficacy of cancer therapy. To address this issue, we have developed an integrated treatment protocol for effective treatment of heterogeneous ovarian cancer. Methods: An amphiphilic polymer coated magnetic iron oxide nanoparticle was conjugated with near infrared dye labeled HER2 affibody and chemotherapy drug cisplatin. The effects of the theranostic nanoparticle on targeted drug delivery, therapeutic efficacy, non-invasive magnetic resonance image (MRI)-guided therapy, and optical imaging detection of therapy resistant tumors were examined in an orthotopic human ovarian cancer xenograft model with highly heterogeneous levels of HER2 expression. Results: We found that systemic delivery of HER2-targeted magnetic iron oxide nanoparticles carrying cisplatin significantly inhibited the growth of primary tumor and peritoneal and lung metastases in the ovarian cancer xenograft model in nude mice. Differential delivery of theranostic nanoparticles into individual tumors with heterogeneous levels of HER2 expression and various responses to therapy were detectable by MRI. We further found a stronger therapeutic response in metastatic tumors compared to primary tumors, likely due to a higher level of HER2 expression and a larger number of proliferating cells in metastatic tumor cells. Relatively long-time retention of iron oxide nanoparticles in tumor tissues allowed interrogating the relationship between nanoparticle drug delivery and the presence of resistant residual tumors by in vivo molecular imaging and histological analysis of the tumor tissues. Following therapy, most of the remaining tumors were small, primary tumors that had low levels of HER2 expression and nanoparticle drug accumulation, thereby explaining their lack of therapeutic response. However, a few residual tumors had HER2-expressing tumor cells and detectable nanoparticle drug delivery but failed to respond, suggesting additional intrinsic resistant mechanisms. Nanoparticle retention in the small residual tumors, nevertheless, produced optical signals for detection by spectroscopic imaging. Conclusion: The inability to completely excise peritoneal metastatic tumors by debulking surgery as well as resistance to chemotherapy are the major clinical challenges for ovarian cancer treatment. This targeted cancer therapy has the potential for the development of effective treatment for metastatic ovarian cancer.

Improving Sensitivity and Specificity of Amyloid-ß Peptides and Tau Protein Detection with Antibiofouling Magnetic Nanoparticles for Liquid Biopsy of Alzheimer's Disease.

Alzheimer's disease (AD) is a growing global healthcare burden affecting the aging population and society. Given the lack of effective treatment to AD, early detection at the prodromal stage and timely monitoring of changes during progression are considered the best approach to control and intervene in disease progression. "Liquid biopsy" of AD biomarkers amyloid-ß peptides (Aßs) and tau proteins in the cerebrospinal fluid (CSF) or blood samples holds great promises for cost-effective, widely accessible, and easy-administrated noninvasive detection and follow-up of AD. However, current in vitro detection methods have not yet demonstrated sufficient sensitivity and specificity using neither Aßs nor tau proteins biomarkers. One major challenge of accurate detection and measurement of biomarker levels in biofluidic samples is the biofouling effect with nonspecific adsorption of unwanted biomolecules, such as various serum proteins, on the surface of targeted detecting agents or devices, causing false-positive and false-negative findings. In this study, antibiofouling polymer polyethylene glycol-block-allyl glycidyl ether (PEG-b-AGE) coated magnetic iron oxide nanoparticles (IONPs) capable of suppressing the nonspecific interactions with biomolecules, especially proteins, were investigated for the immunomagnetic capturing of Aß40 and Aß42 peptides and tau protein spiked in CSF- and serum-mimicking samples using corresponding antibodies conjugated as targeting ligands. Antibody-conjugated antibiofouling IONPs demonstrated improved specificity (>90%) and sensitivity (>95%) over those of antibody-conjugated magnetic micron beads (Dynabeads, ∼50% specificity and 30-40% sensitivity) widely used as magnetic separating agents under the same experimental conditions with the presence of nontargeted interfering proteins. The antibody-conjugated IONPs also exhibited significantly higher sensitivities (80-90%) and better performance of capturing Aßs and tau protein from the human whole blood samples than antibody-conjugated Dynabeads (∼20%).

Dual-targeting Wnt and uPA receptors using peptide conjugated ultra-small nanoparticle drug carriers inhibited cancer stem-cell phenotype in chemo-resistant breast cancer.

Heterogeneous tumor cells, high incidence of tumor recurrence, and decrease in overall survival are the major challenges for the treatment of chemo-resistant breast cancer. Results of our study showed differential chemotherapeutic responses among breast cancer patient derived xenograft (PDX) tumors established from the same patients. All doxorubicin (Dox)-resistant tumors expressed higher levels of cancer stem-like cell biomarkers, including CD44, Wnt and its receptor LRP5/6, relative to Dox-sensitive tumors. To effectively treat resistant tumors, we developed an ultra-small magnetic iron oxide nanoparticle (IONP) drug carrier conjugated with peptides that are dually targeted to Wnt/LRP5/6 and urokinase plasminogen activator receptor (uPAR). Our results showed that simultaneous binding to LRP5/6 and uPAR by the dual receptor targeted IONPs was required to inhibit breast cancer cell invasion. Molecular analysis revealed that the dual receptor targeted IONPs significantly inhibited Wnt/ß-catenin signaling and cancer stem-like phenotype of tumor cells, with marked reduction of Wnt ligand, CD44 and uPAR. Systemic administration of the dual targeted IONPs led to nanoparticle-drug delivery into PDX tumors, resulting in stronger tumor growth inhibition compared to non-targeted or single-targeted IONP-Dox in a human breast cancer PDX model. Therefore, co-targeting Wnt/LRP and uPAR using IONP drug carriers is a promising therapeutic approach for effective drug delivery to chemo-resistant breast cancer.

Tumor Penetrating Theranostic Nanoparticles for Enhancement of Targeted and Image-guided Drug Delivery into Peritoneal Tumors following Intraperitoneal Delivery.

The major obstacles in intraperitoneal (i.p.) chemotherapy of peritoneal tumors are fast absorption of drugs into the blood circulation, local and systemic toxicities, inadequate drug penetration into large tumors, and drug resistance. Targeted theranostic nanoparticles offer an opportunity to enhance the efficacy of i.p. therapy by increasing intratumoral drug delivery to overcome resistance, mediating image-guided drug delivery, and reducing systemic toxicity. Herein we report that i.p. delivery of urokinase plasminogen activator receptor (uPAR) targeted magnetic iron oxide nanoparticles (IONPs) led to intratumoral accumulation of 17% of total injected nanoparticles in an orthotopic mouse pancreatic cancer model, which was three-fold higher compared with intravenous delivery. Targeted delivery of near infrared dye labeled IONPs into orthotopic tumors could be detected by non-invasive optical and magnetic resonance imaging. Histological analysis revealed that a high level of uPAR targeted, PEGylated IONPs efficiently penetrated into both the peripheral and central tumor areas in the primary tumor as well as peritoneal metastatic tumor. Improved theranostic IONP delivery into the tumor center was not mediated by nonspecific macrophage uptake and was independent from tumor blood vessel locations. Importantly, i.p. delivery of uPAR targeted theranostic IONPs carrying chemotherapeutics, cisplatin or doxorubicin, significantly inhibited the growth of pancreatic tumors without apparent systemic toxicity. The levels of proliferating tumor cells and tumor vessels in tumors treated with the above theranostic IONPs were also markedly decreased. The detection of strong optical signals in residual tumors following i.p. therapy suggested the feasibility of image-guided surgery to remove drug-resistant tumors. Therefore, our results support the translational development of i.p. delivery of uPAR-targeted theranostic IONPs for image-guided treatment of peritoneal tumors.

Hepatocellular Carcinoma: Intra-arterial Delivery of Doxorubicin-loaded Hollow Gold Nanospheres for Photothermal Ablation-Chemoembolization Therapy in Rats.

Purpose To determine if combretastatin A-4 phosphate disodium (CA4P) can enhance the tumor uptake of doxorubicin (Dox)-loaded, polyethylene glycol (PEG)-coated hollow gold nanospheres (HAuNS) mixed with ethiodized oil for improved photothermal ablation (PTA)-chemoembolization therapy (CET) of hepatocellular carcinoma (HCC) in rats. Materials and Methods Animal experiments were approved by the institutional animal care and use committee and performed from February 2014 to April 2015. Male Sprague-Dawley rats (n = 45; age, 12 weeks) were inoculated with N1S1 HCC cells in the liver, and 8 days later, were randomly divided into two groups of 10 rats. Group 1 rats received intrahepatic arterial injection of PEG-HAuNS and ethiodized oil alone; group 2 received pretreatment with CA4P and injection of PEG-HAuNS and ethiodized oil 5 minutes later. The gold content of tumor and liver tissue at 1 hour or 24 hours after injection was quantified by using neutron activation analysis (n = 5 per time point). Five rats received pretreatment CA4P, PEG-copper 64-HAuNS, and ethiodized oil and underwent micro-positron emission tomography (PET)/computed tomography (CT). In a separate study, three groups of six rats with HCC were injected with saline solution (control group); CA4P, Dox-loaded PEG-coated HAuNS (Dox@PEG-HAuNS), and ethiodized oil (CET group); or CA4P, Dox@PEG-HAuNS, ethiodized oil, and near-infrared irradiation (PTA-CET group). Temperature was recorded during laser irradiation. Findings were verified at postmortem histopathologic and/or autoradiographic examination. Wilcoxon rank-sum test and Pearson correlation analyses were performed. Results PEG-HAuNS uptake in CA4P-pretreated HCC tumors was significantly higher than that in non-CA4P-pretreated tumors at both 1 hour (P < .03) and 24 hours (P < .01). Mean ± standard deviation of tumor-to-liver PEG-HAuNS uptake ratios at 1 hour and 24 hours, respectively, were 5.63 ± 3.09 and 1.68 ± 0.77 in the CA4P-treated group and 1.29 ± 2.40 and 0.14 ± 0.11 in the non-CA4P-treated group. Micro-PET/CT allowed clear delineation of tumors, enabling quantitative imaging analysis. Laser irradiation increased temperature to 60°C and 43°C in the tumor and adjacent liver, respectively. Mean HCC tumor volumes 10 days after therapy were 1.68 cm3 ± 1.01, 3.96 cm3 ± 1.75, and 6.13 cm3 ± 2.27 in the PTA-CET, CET, and control groups, respectively, with significant differences between the PTA-CET group and other groups (P < .05). Conclusion CA4P pretreatment caused a higher concentration of Dox@PEG-HAuNS to be trapped inside the tumor, thereby enhancing the efficacy of anti-HCC treatment with PTA-CET in rats. © RSNA, 2016 Online supplemental material is available for this article.

IGF1 Receptor Targeted Theranostic Nanoparticles for Targeted and Image-Guided Therapy of Pancreatic Cancer.

Overcoming resistance to chemotherapy is a major and unmet medical challenge in the treatment of pancreatic cancer. Poor drug delivery due to stromal barriers in the tumor microenvironment and aggressive tumor biology are additional impediments toward a more successful treatment of pancreatic cancer. In attempts to address these challenges, we developed IGF1 receptor (IGF1R)-directed, multifunctional theranostic nanoparticles for targeted delivery of therapeutic agents into IGF1R-expressing drug-resistant tumor cells and tumor-associated stromal cells. These nanoparticles were prepared by conjugating recombinant human IGF1 to magnetic iron oxide nanoparticles (IONPs) carrying the anthracycline doxorubicin (Dox) as the chemotherapeutic payload. Intravenously administered IGF1-IONPs exhibited excellent tumor targeting and penetration in an orthotopic patient-derived xenograft (PDX) model of pancreatic cancer featuring enriched tumor stroma and heterogeneous cancer cells. IGF1R-targeted therapy using the theranostic IGF1-IONP-Dox significantly inhibited the growth of pancreatic PDX tumors. The effects of the intratumoral nanoparticle delivery and therapeutic responses in the orthotopic pancreatic PDX tumors could be detected by magnetic resonance imaging (MRI) with IONP-induced contrasts. Histological analysis showed that IGF1R-targeted delivery of Dox significantly inhibited cell proliferation and induced apoptotic cell death of pancreatic cancer cells. Therefore, further development of IGF1R-targeted theranostic IONPs and MRI-guided cancer therapy as a precision nanomedicine may provide the basis for more effective treatment of pancreatic cancer.

Theranostic nanoparticles carrying doxorubicin attenuate targeting ligand specific antibody responses following systemic delivery.

Understanding the effects of immune responses on targeted delivery of nanoparticles is important for clinical translations of new cancer imaging and therapeutic nanoparticles. In this study, we found that repeated administrations of magnetic iron oxide nanoparticles (IONPs) conjugated with mouse or human derived targeting ligands induced high levels of ligand specific antibody responses in normal and tumor bearing mice while injections of unconjugated mouse ligands were weakly immunogenic and induced a very low level of antibody response in mice. Mice that received intravenous injections of targeted and polyethylene glycol (PEG)-coated IONPs further increased the ligand specific antibody production due to differential uptake of PEG-coated nanoparticles by macrophages and dendritic cells. However, the production of ligand specific antibodies was markedly inhibited following systemic delivery of theranostic nanoparticles carrying a chemotherapy drug, doxorubicin. Targeted imaging and histological analysis revealed that lack of the ligand specific antibodies led to an increase in intratumoral delivery of targeted nanoparticles. Results of this study support the potential of further development of targeted theranostic nanoparticles for the treatment of human cancers.

MR imaging enables measurement of therapeutic nanoparticle uptake in rat N1-S1 liver tumors after nanoablation.

PURPOSE: To test the hypothesis that magnetic resonance (MR) imaging can quantify intratumoral superparamagnetic iron oxide (SPIO) nanoparticle uptake after nanoablation. MATERIALS AND METHODS: SPIO nanoparticles functionalized with doxorubicin were synthesized. N1-S1 hepatomas were successfully induced in 17 Sprague-Dawley rats distributed into three dosage groups. Baseline tumor R2* values (the reciprocal of T2*) were determined using 7-tesla (T) MR imaging. After intravenous injection of SPIO nanoparticles, reversible electroporation (1,300 V/cm, 8 pulses, 100-µs pulse duration) was applied. Imaging of rats was performed to determine tumor R2* values after the procedure, and change in R2* (ΔR2*) was calculated. Inductively coupled plasma mass spectrometry was used to determine intratumoral iron (Fe) concentration after the procedure, which served as a proxy for SPIO nanoparticle uptake. Mean tumor Fe concentration [Fe] and ΔR2* for each subject were assessed for correlation with linear regression, and mean [Fe] for each dosage group was compared with analysis of variance. RESULTS: ΔR2* significantly correlated with tumor SPIO nanoparticle uptake after nanoablation (r = 0.50, P = .039). On average, each 0.1-ms(-1) increase in R2* corresponded to a 0.1394-mM increase in [Fe]. There was no significant difference in mean SPIO nanoparticle uptake among dosage groups (P = .57). CONCLUSIONS: Intratumoral SPIO nanoparticle uptake after nanoablation can be successfully quantified noninvasively with 7-T MR imaging. Imaging can be used as a method to estimate localized drug delivery after nanoablation.

The role of functionalized magnetic iron oxide nanoparticles in the central nervous system injury and repair: new potentials for neuroprotection with Cerebrolysin therapy.

Functionalized Magnetic Iron Oxide Nanoparticles (FMIONPs) are being explored for the development of various biomedical applications, e.g., cancer chemotherapy and/or several other radiological or diagnostic purposes. However, the effects of these NPs per se on the central nervous system (CNS) injury or repair are not well known. This review deals with different aspects of FMIONPs in relation to brain function based on the current literature as well as our own investigation in animal models of CNS injuries. It appears that FMIONPs are innocuous when administered intravenously within the CNS under normal conditions. However, abnormal reactions to FMIONPs in the brain or spinal cord could be seen if they are combined with CNS injuries e.g., hyperthermia or traumatic insults to the brain or spinal cord. Thus, administration of FMIONPs in vivo following whole body hyperthermia (WBH) or a focal spinal cord injury (SCI) exacerbates cellular damage. Since FMIONPs could help in diagnostic purposes or enhance the biological effects of radiotherapy/chemotherapy it is likely that these NPs may have some adverse reaction as well under disease condition. Thus, under such situation, adjuvant therapy e.g., Cerebrolysin (Ever NeuroPharma, Austria), a suitable combination of several neurotrophic factors and active peptide fragments are the need of the hour to contain such cellular damages caused by the FMIONPs in vivo. Our observations show that co-administration of Cerebrolysin prevents the FMIONPs induced pathologies associated with CNS injuries. These observations support the idea that FMIONPs are safe for the CNS in disease conditions when co-administered with cerebrolysin. This indicates that cerebrolysin could be used as an adjunct therapy to prevent cellular damages in disease conditions where the use of FMIONPs is required for better efficacy e.g., cancer treatment.

Active targeting using HER-2-affibody-conjugated nanoparticles enabled sensitive and specific imaging of orthotopic HER-2 positive ovarian tumors.

Despite advances in cancer diagnosis and treatment, ovarian cancer remains one of the most fatal cancer types. The development of targeted nanoparticle imaging probes and therapeutics offers promising approaches for early detection and effective treatment of ovarian cancer. In this study, HER-2 targeted magnetic iron oxide nanoparticles (IONPs) are developed by conjugating a high affinity and small size HER-2 affibody that is labeled with a unique near infrared dye (NIR-830) to the nanoparticles. Using a clinically relevant orthotopic human ovarian tumor xenograft model, it is shown that HER-2 targeted IONPs are selectively delivered into both primary and disseminated ovarian tumors, enabling non-invasive optical and MR imaging of the tumors as small as 1 mm in the peritoneal cavity. It is determined that HER-2 targeted delivery of the IONPs is essential for specific and sensitive imaging of the HER-2 positive tumor since we are unable to detect the imaging signal in the tumors following systemic delivery of non-targeted IONPs into the mice bearing HER-2 positive SKOV3 tumors. Furthermore, imaging signals and the IONPs are not detected in HER-2 low expressing OVCAR3 tumors after systemic delivery of HER-2 targeted-IONPs. Since HER-2 is expressed in a high percentage of ovarian cancers, the HER-2 targeted dual imaging modality IONPs have potential for the development of novel targeted imaging and therapeutic nanoparticles for ovarian cancer detection, targeted drug delivery, and image-guided therapy and surgery.

Image-guided local delivery strategies enhance therapeutic nanoparticle uptake in solid tumors.

Nanoparticles (NP) have emerged as a novel class of therapeutic agents that overcome many of the limitations of current cancer chemotherapeutics. However, a major challenge to many current NP platforms is unfavorable biodistribution, and limited tumor uptake, upon systemic delivery. Delivery, therefore, remains a critical barrier to widespread clinical adoption of NP therapeutics. To overcome these limitations, we have adapted the techniques of image-guided local drug delivery to develop nanoablation and nanoembolization. Nanoablation is a tumor ablative strategy that employs image-guided placement of electrodes into tumor tissue to electroporate tumor cells, resulting in a rapid influx of NPs that is not dependent on cellular uptake machinery or stage of the cell cycle. Nanoembolization involves the image-guided delivery of NPs and embolic agents directly into the blood supply of tumors. We describe the design and testing of our innovative local delivery strategies using doxorubicin-functionalized superparamagnetic iron oxide nanoparticles (DOX-SPIOs) in cell culture, and the N1S1 hepatoma and VX2 tumor models, imaged by high resolution 7T MRI. We demonstrate that local delivery techniques result in significantly increased intratumoral DOX-SPIO uptake, with limited off-target delivery in tumor-bearing animal models. The techniques described are versatile enough to be extended to any NP platform, targeting any solid organ malignancy that can be accessed via imaging guidance.

Designed fabrication of unique eccentric mesoporous silica nanocluster-based core-shell nanostructures for pH-responsive drug delivery.

A novel and facile strategy using poly(acrylic acid) (PAA) as a nanoreactor and template has been proposed and applied for the first time to fabricate a novel and unique class of multifunctional eccentric Fe3O4@PAA/SiO2 core-shell nanoclusters (NCs) consisting of a single Fe3O4 nanoparticle (NP), PAA, and eccentric SiO2 NCs that are composed of a large number of small fluorescent SiO2 NPs. Interestingly, the resulting eccentric PAA shell around Fe3O4 NPs as a high water-absorbent polymer is like a "reservoir" to absorb and retain water molecules inside its net structure to confine the growth of small SiO2 NPs inside the PAA networks, resulting in the formation of an eccentric SiO2 NC with aggregated pores. The thicknesses of uniform and well-dispersed SiO2 NCs can also be precisely controlled by varying the amount of tetraethyl orthosilicate (TEOS). Importantly, the synthetic method has been confirmed to be universal and extended to other functional NPs with different compositions and shapes as eccentric cores. Furthermore, the as-prepared multifunctional eccentric Fe3O4@PAA/SiO2 core-shell NCs combined fluorescence imaging, ultrahigh drug loading capacity (1.13 mg doxorubicin/mg eccentric NCs), and pH-responsive drug release into one were taken as an example to study the applications in simultaneous fluorescence imaging and pH responsive drug delivery into prostate cancer PC3M cells.

DOT corrected fluorescence molecular tomography using targeted contrast agents for small animal tumor imaging.

PURPOSE: To demonstrate diffuse optical tomography (DOT) corrected fluorescence molecular tomography (FMT) for quantitatively imaging tumor-targeted contrast agents in a 4T1 mouse mammary tumor model. PROCEDURES: In the first set of experiments, we validated our DOT corrected FMT method using subcutaneously injected 4T1 cells pre-labeled with a near-infrared (NIR) Cy 5.5 dye labeled recombinant amino-terminal fragment (ATF) of the receptor binding domain of urokinase plasminogen activator (uPA), which binds to uPA receptor (uPAR) that is highly expressed in breast cancer tissues. Next, we apply the DOT corrected FMT method to quantitatively evaluate the ability of sensitive tumor imaging after systemic delivery of new uPAR-targeted optical imaging probes in the mice bearing 4T1 mammary tumors. These uPAR-targeted optical imaging probes are ATF peptides labeled with a newly developed NIR-830 dye being conjugated to magnetic iron oxide nanoparticles (IONPs). RESULTS: Our results have shown that DOT corrected FMT can accurately quantify and localize the injected imaging probe labeled 4T1 cells. Following systemic delivery of the targeted imaging nanoprobes into the mice bearing orthotopic mammary tumors, specific accumulation of the imaging probes in the orthotopic mammary tumors was detected in the mice that received uPAR targeted NIR-830-ATF-IONP probes but not in the mice injected with non-targeted NIR-830-mouse serum albumin (MSA)-IONPs. Additionally, DOT corrected FMT also enables the detection of both locally recurrent tumor and lung metastasis in the mammary tumor model 72 hrs after systemic administration of the uPAR-targeted NIR-830-labeled ATF peptide imaging probes. CONCLUSIONS: DOT corrected FMT and uPAR-targeted optical imaging probes have great potential for detection of breast cancer, recurrent tumor and metastasis in small animals.

Targeted delivery of siRNA-generating DNA nanocassettes using multifunctional nanoparticles.

Molecular therapy using a small interfering RNA (siRNA) has shown promise in the development of novel therapeutics. Various formulations have been used for in vivo delivery of siRNAs. However, the stability of short double-stranded RNA molecules in the blood and efficiency of siRNA delivery into target organs or tissues following systemic administration have been the major issues that limit applications of siRNA in human patients. In this study, multifunctional siRNA delivery nanoparticles are developed that combine imaging capability of nanoparticles with urokinase plasminogen activator receptor-targeted delivery of siRNA expressing DNA nanocassettes. This theranostic nanoparticle platform consists of a nanoparticle conjugated with targeting ligands and double-stranded DNA nanocassettes containing a U6 promoter and a shRNA gene for in vivo siRNA expression. Targeted delivery and gene silencing efficiency of firefly luciferase siRNA nanogenerators are demonstrated in tumor cells and in animal tumor models. Delivery of survivin siRNA expressing nanocassettes into tumor cells induces apoptotic cell death and sensitizes cells to chemotherapy drugs. The ability of expression of siRNAs from multiple nanocassettes conjugated to a single nanoparticle following receptor-mediated internalization should enhance the therapeutic effect of the siRNA-mediated cancer therapy.

Cadmium-free quantum dots as time-gated bioimaging probes in highly-autofluorescent human breast cancer cells.

We report cadmium-free, biocompatible (Zn)CuInS(2) quantum dots with long fluorescence lifetimes as superior bioimaging probes using time-gated detection to suppress cell autofluorescence and improve the signal : background ratio by an order of magnitude. These results will be important for developing non-toxic fluorescence imaging probes for ultrasensitive biomedical diagnostics.

uPAR-targeted optical imaging contrasts as theranostic agents for tumor margin detection.

Complete removal of tumors by surgery is the most important prognostic factor for cancer patients with the early stage cancers. The ability to identify invasive tumor edges of the primary tumor, locally invaded small tumor lesions, and drug resistant residual tumors following neoadjuvant therapy during surgery should significantly reduce the incidence of local tumor recurrence and improve survival of cancer patients. In this study, we report that urokinase plasminogen activator (uPA) and its receptor (uPAR) are the ligand/cell surface target pair for the development of targeted optical imaging probes for enhancing imaging contrasts in the tumor border. Recombinant peptides of the amino terminal fragment (ATF) of the receptor binding domain of uPA were labeled with near infrared fluorescence (NIR) dye molecules either as peptide-imaging or peptide-conjugated nanoparticle imaging probes. Systemic delivery of the uPAR-targeted imaging probes in mice bearing orthotopic human breast or pancreatic tumor xenografts or mouse mammary tumors led to the accumulation of the probes in the tumor and stromal cells, resulting in strong signals for optical imaging of tumors and identification of tumor margins. Histological analysis showed that a high level of uPAR-targeted nanoparticles was present in the tumor edge or active tumor stroma immediately adjacent to the tumor cells. Furthermore, following targeted therapy using uPAR-targeted theranostic nanoparticles, residual tumors were detectable by optical imaging through the imaging contrasts produced by NIR-dye-labeled theranostic nanoparticles in drug resistant tumor cells. Therefore, results of our study support the potential of the development of uPAR-targeted imaging and theranostic agents for image-guided surgery.

General route to multifunctional uniform yolk/mesoporous silica shell nanocapsules: a platform for simultaneous cancer-targeted imaging and magnetically guided drug delivery.

Hollow mesoporous SiO(2) (mSiO(2)) nanostructures with movable nanoparticles (NPs) as cores, so-called yolk-shell nanocapsules (NCs), have attracted great research interest. However, a highly efficient, simple and general way to produce yolk-mSiO(2) shell NCs with tunable functional cores and shell compositions is still a great challenge. A facile, general and reproducible strategy has been developed for fabricating discrete, monodisperse and highly uniform yolk-shell NCs under mild conditions, composed of mSiO(2) shells and diverse functional NP cores with different compositions and shapes. These NPs can be Fe(3)O(4) NPs, gold nanorods (GNRs), and rare-earth upconversion NRs, endowing the yolk-mSiO(2) shell NCs with magnetic, plasmonic, and upconversion fluorescent properties. In addition, multifunctional yolk-shell NCs with tunable interior hollow spaces and mSiO(2) shell thickness can be precisely controlled. More importantly, fluorescent-magnetic-biotargeting multifunctional polyethyleneimine (PEI)-modified fluorescent Fe(3)O(4)@mSiO(2) yolk-shell nanobioprobes as an example for simultaneous targeted fluorescence imaging and magnetically guided drug delivery to liver cancer cells is also demonstrated. This synthetic approach can be easily extended to the fabrication of multifunctional yolk@mSiO(2) shell nanostructures that encapsulate various functional movable NP cores, which construct a potential platform for the simultaneous targeted delivery of drug/gene/DNA/siRNA and bio-imaging.

Selected-control fabrication of multifunctional fluorescent-magnetic core-shell and yolk-shell hybrid nanostructures.

The selected-control preparation of uniform core-shell and yolk-shell architectures, which combine the multiple functions of a superparamagnetic iron oxide (SPIO) core and europium-doped yttrium oxide (Y(2)O(3):Eu) shell in a single material with tunable fluorescence and magnetic properties, has been successfully achieved by controlling the heat-treatment conditions. Furthermore, the shell thickness and interior cavity of SPIO@Y(2)O(3):Eu core-shell and yolk-shell nanostructures can be precisely tuned. Importantly, as-prepared SPIO@Y(2)O(3):Eu yolk-shell nanocapsules (NCs) modified with amino groups as cancer-cell fluorescence imaging agents are also demonstrated. To the best of our knowledge, this is the first report on the selected-control fabrication of uniform SPIO@Y(2)O(3):Eu core-shell nanoparticles and yolk-shell NCs. The combined magnetic manipulation and optical monitoring of magnetic-fluorescent SPIO@Y(2)O(3):Eu yolk-shell NCs will open up many exciting opportunities in dual imaging for targeted delivery and thermal therapy.

Preparation of quantum dots encoded microspheres by electrospray for the detection of biomolecules.

In this paper, a novel method based on the electrospray technique has been developed for preparation of quantum dot (QD)-encoded microspheres for the fist time. By electrospraying the mixture of polymer solution and quantum dots solution (single-color QDs or multi-color QDs), it is accessible to obtain a series of composite microspheres containing the functional nanoparticle. Poly(styrene-acrylate) was utilized as the electrospray polymer materials in order to obtain the microsphere modified with carboxyl group on the surface. Moreover, to test the performance of the QD-encoded microsphere in bioapplication, it is carried out that immunofluorescence analysis between antigens of mouse IgG immobilized on the functional microsphere and FITC labeled antibodies of goat-anti-mouse IgG in experiment. To the best of our knowledge, this is the first report of QD-encoded microspheres prepared by electrospray technology. This technology can carry out the one-pot preparation of different color QD-encoded microspheres with multiple intensities. This technology could be also suitable for encapsulating other optical nanocrystals and magnetic nanoparticles for obtaining multifunctional microspheres. All of the results in this paper show that the fluorescence beads made by electrospray technique can be well applied in multiplex analysis. These works provide a good foundation to accelerate application of preparing microspheres by electrospray technique in practice.