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Gold‐Nanoshelled Microcapsules: A Theranostic Agent for Ultrasound Contrast Imaging and Photothermal Therapy

The term theranostics, which is derived from “diagnostics” and “therapy”, refers to a treatment strategy that combines a diagnostic test and a specific therapy based on the test results. This integration of diagnostic imaging capability with therapy is critical in addressing the challenges of cancer heterogeneity and adaptation. Therefore, theranostic agents have received a great deal of recent research interest in cancer diagnosis and treatment. Among all the diagnostic imaging techniques, ultrasound imaging has a unique advantage because of its features of real-time, low-cost, high safety, and ease of incorporation into portable devices. With the use of ultrasound contrast agents (UCAs), the resolution and sensitivity of clinical ultrasound imaging have been greatly improved. Microcapsules composed of poly(lactic acid) (PLA), which has outstanding biocompatibility and biodegradability, show good ultrasound contrast-enhancing capabilities and other advantages: they have good mechanical strength and are thus stable, they can load either hydrophilic or hydrophobic species or both, and they are surface-charged and have functional groups on the surface so that they could be easily modified to introduce further practical features. Gold nanostructures exhibit good biocompatibility as well as excellent optical and electronic properties, thus allowing use in biological and medical applications. Gold nanoshells have a spherical dielectric core particle surrounded by a thin nanoscale gold shell. By controlling the thickness of the gold shell and the diameter of the core, the plasmon resonance and the resulting optical absorption of gold nanoshells can be tuned to the near-infrared (NIR) region, where the absorption of human tissues is minimal and penetration is optimal. On the other hand, the strong optical absorption of nanoshells can rapidly increase the local temperature under NIR irradiation. Therefore, the gold nanoshells can be used as photoabsorbers for remote NIR photothermal ablation therapy. Lasers and photoabsorbers such as gold nanostructures are used to carry out cancer treatment in photothermal therapy. However, the location and size of cancers must be identified before therapy, the treatment procedure needs to be monitored in real time during therapy, and the effectiveness has to be assessed after therapy. Contrast-enhanced ultrasound imaging could be the technique of choice to address these tasks. Therefore, the development of goldnanoshell-based UCAs could operate as a multifunctional theranostic agent for imaging-guided photothermal therapy. We have developed a novel multifunctional theranostic agent based on gold-nanoshelled microcapsules (GNS-MCs) by electrostatic adsorption of gold nanoparticles as seeds onto the polymeric microcapsule surfaces, followed by the formation of gold nanoshells by using a surface seeding method (Figure 1). The polymeric microcapsules were generated from PLA and polyvinyl alcohol (PVA) materials by employing the water-in-oil-in-water (W/O/W) double-emulsion method, and were negatively charged with a zeta potential of about 25 mV. Upon exposure to positively charged poly(allyl-

Gold nanoshelled liquid perfluorocarbon magnetic nanocapsules: a nanotheranostic platform for bimodal ultrasound/magnetic resonance imaging guided photothermal tumor ablation.

Imaging guided ablation therapy has been applied in both biomedical research and clinical trials and turned out to be one of the most promising approaches for cancer treatment. Herein, the multifunctional nanocapsules were fabricated through loading perfluorooctylbromide (PFOB) and superparamagnetic iron oxide nanoparticles (SPIOs) into poly(lactic acid) (PLA) nanocapsules (NCs), followed by the formation of PEGylated gold nanoshell on the surface. The resulting multi-component NCs were proved to be able to act as nanotheranostic agent to achieve successful bimodal ultrasound (US)/magnetic resonance imaging (MRI) guided photothermal ablation in human tumor xenograft models non-invasively. Such a single theranostic agent with the combination of real-time US and high-resolution MR imaging would be of great value to offer more comprehensive diagnostic information and dynamics of disease progression for the accurate location of therapeutic focusing spot in the targeted tumor tissue, showing great potential as an effective nanoplatform for contrast imaging guided photothermal therapy.

Imaging guided photothermal therapy using iron oxide loaded poly(lactic acid) microcapsules coated with graphene oxide

A novel multifunctional theranostic agent has been successfully fabricated by loading iron oxide nanoparticles into poly(lactic acid) (PLA) microcapsules followed by surface functionalization with graphene oxide. Both in vitro and in vivo experiments proved that the resulting microcapsules could serve as contrast agents to simultaneously enhance ultrasound, magnetic resonance and photoacoustic imaging. The composite microcapsules show good biocompatibility and rapid response to magnetic fields. Due to the strong absorption of the near-infrared light, the composite microcapsules could efficiently kill cancer cells upon NIR laser irradiation. In addition, it was found that such a photothermal effect could be obviously enhanced by applying an external magnetic field. In a nutshell, this multifunctional microcapsule can be developed as a promising platform that integrates multimodality imaging and therapy capabilities for effective cancer theranostics.

Polypyrrole Hollow Microspheres as Echogenic Photothermal Agent for Ultrasound Imaging Guided Tumor Ablation

Ultrasound (US) imaging provides a valuable opportunity to administer photothermal therapy (PTT) of cancer with real-time guidance to ensure proper targeting, but only a few theranostic agents were developed by physically grafting near infrared (NIR)-absorbing inorganic nanomaterials to ready-made ultrasound contrast agents (UCAs) for US imaging guided PTT. In this paper, NIR absorbing hollow microspheres were generated from polypyrrole merely using a facile one-step microemulsion method. It was found that the obtained polypyrrole hollow microspheres (PPyHMs) can act as an efficient theranostic agent not only to enhance US imaging greatly, but also exhibit excellent photohyperthermic effects. The contrast consistently sustained the echo signals for no less than 5 min and the NIR laser light ablated the tumor completely within two weeks in the presence of PPyHMs. More importantly, no use of additional NIR absorber substantially minimizes an onetime dose of the theranostic agent.

Engineering of perfluorooctylbromide polypyrrole nano-/microcapsules for simultaneous contrast enhanced ultrasound imaging and photothermal treatment of cancer.

A versatile oil-in-water emulsion method has been explored for constructing water-dispersible polypyrrole (PPy) nano-/microcapsules with a soluble PPy complex as multifunctional photothermal agents for tumor ablation. In this work, both PPy nanocapsules (280.4 ± 79.0 nm) and microcapsules (1.31 ± 0.45 μm) with liquid perfluorooctylbromide (PFOB) core could be obtained by simply tuning the process energy for emulsion formation from ultrasonication to homogenization. Owing to the encapsulated liquid PFOB and strong near-infrared (NIR) absorption of PPy shell, the resulted PPy capsules showed great promise in ultrasound imaging guided photothermal ablation of tumor cells without inducing any significant side effect. Thus, it is anticipated that fine-tuning of the other encapsulated drugs or functional materials in PPy capsules would foster avenues for the development of multifunctional platforms for cancer treatments.

Bifunctional gold nanorod-loaded polymeric microcapsules for both contrast-enhanced ultrasound imaging and photothermal therapy

A novel bifunctional theranostic agent has been fabricated through the combination of acoustic responsive poly(lactic acid) microcapsules and near infrared absorbed gold nanorods that holds the potential of simultaneous ultrasound contrast diagnostic imaging and photothermal therapy.

Contrast ultrasound-guided photothermal therapy using gold nanoshelled microcapsules in breast cancer

OBJECTIVES The purpose of this study was to test whether dual functional gold nano-shelled microcapsules (GNS-MCs) can be used as an ultrasound imaging enhancer and as an optical absorber for photothermal therapy (PTT) in a rodent model of breast cancer. METHODS GNS-MCs were fabricated with an inner air and outer gold nanoshell spherical structure. Photothermal cytotoxicity of GNS-MCs was tested with BT474 cancer cells in vitro and non-obese diabetes-SCID (NOD/SCID) mice with breast cancer. GNS-MCs were injected into the tumor under ultrasound guidance and treated with near-infrared (NIR) laser irradiation. The photothermal ablative effectiveness of GNS-MCs was evaluated by measuring the surface and internal temperature of the tumor as well as the size of the tumor using histological confirmation. RESULTS NIR laser irradiation resulted in significant tumor cell death in GNS-MCs-treated BT474 cells in vitro. GNS-MCs were able to serve as an ultrasound enhancer to guide the intratumoral injection of GNS-MCs and ensure their uniform distribution. In vivo studies revealed that NIR laser irradiation increased the intratumoral temperature to nearly 70°C for 8 min in GNS-MCs-treated mice. Tumor volumes decreased gradually and tumors were completely ablated in 6 out of 7 mice treated with GNS-MCs and laser irradiation by 17 days after treatment. CONCLUSION This study demonstrates that ultrasound-guided PTT with theranostic GNS-MCs is a promising technique for in situ treatment of breast cancer.

Quantitative Assessment of Hepatic Fibrosis by Contrast-enhanced Ultrasonography

OBJECTIVE To explore the contrast-enhanced ultrasonographic features for quantitative assessment of hepatic fibrosis. METHODS 86 patients with chronic viral hepatitis B were enrolled in this study from March 2007 to August 2009. The patients were classified into 5 groups (S(0)-S(4)) according to fibrosis stage evaluated with ultrasound guided liver biopsy. New contrast-enhanced ultrasonography (CEUS) features including area under the time-intensity curve (TIC) of portal venous phase/hepatic arterial phase (Qp/Qa) and intensity of portal venous phase/hepatic arterial phase (Ip/Ia) were used to detect the blood supply ratio (portal vein/hepatic artery) in each group. Arrival time of portal vein trunk (Tp) and decreasing rate of TIC (β) were also analyzed. RESULTS Qp/Qa and Ip/Ia decreased from S(0) to S(4), while Tp and β increased. These 4 features were significantly correlated with the degree of fibrosis (P<0.001) and were significantly different among the five groups (P<0.001). Sensitivity and specificity of Ip/Ia were 80% and 86% for groups ≥S(1), 75% and 86% for groups ≥ S(2), 71% and 84% for groups ≥ S(3), and 76% and 80% for group S(4), respectively. Sensitivity and specificity of Qp/Qa were 70% and 88% for groups ≥ S(1), 80% and 76% for groups ≥ S(2), 74% and 70% for groups ≥ S(3), and 81% and 95% for group S(4), respectively. CONCLUSION Ip/Ia and Qp/Qa could be adopted as reliable, non-invasive features for quantitative assessment of hepatic fibrosis.

Porphyrin-grafted Lipid Microbubbles for the Enhanced Efficacy of Photodynamic Therapy in Prostate Cancer through Ultrasound-controlled In Situ Accumulation

Photodynamic therapy (PDT) holds promise for focal therapy of prostate cancer (PCa). However, the therapeutic efficacy needs improvement, and further development of PDT for PCa has challenges, including uncertainty of photosensitizers (PSs) accumulation at the tumor site and difficulty in visualizing lesions using conventional ultrasound (US) imaging. We have developed novel porphyrin-grafted lipid (PGL) microbubbles (MBs; PGL-MBs) and propose a strategy to integrate PGL-MBs with US imaging to address these limitations and enhance PDT efficacy. Methods: PGL-MBs have two functions: imaging guidance by contrast-enhanced ultrasound (CEUS) and targeted delivery of PSs by ultrasound targeted microbubble destruction (UTMD). PGL-MBs were prepared and characterized before and after low-frequency US (LFUS) exposure. Then, in vitro studies validated the efficacy of PDT with PGL-MBs in human prostate cancer PC3 cells. PC3-xenografted nude mice were used to validate CEUS imaging, accumulation at the tumor site, and in vivo PDT efficacy. Results: PGL-MBs showed good contrast enhancement for US imaging and were converted into nanoparticles upon LFUS exposure. The resulting uniquely structured nanoparticles avoided porphyrin fluorescence quenching and efficiently accumulated at the tumor site through the sonoporation effect created with the assistance of US to achieve excellent PDT efficacy. Conclusions: This is the first preclinical investigation of MBs applied in PDT for PCa. PGL-MBs possess favorable CEUS imaging effects to enhance the localization of tumors. PGL-MBs with LFUS control PS accumulation at the tumor site to achieve highly effective PDT of PCa. This strategy carries enormous clinical potential for PCa management.