Shufang Chang

Ultrasound-Mediated Destruction of LHRHa-Targeted and Paclitaxel-Loaded Lipid Microbubbles for the Treatment of Intraperitoneal Ovarian Cancer Xenografts

Ultrasound-targeted microbubble destruction (UTMD) is a promising technique to facilitate the delivery of chemotherapy in cancer treatment. However, the process typically uses nonspecific microbubbles, leading to low tumor-to-normal tissue uptake ratio and adverse side effects. In this study, we synthesized the LHRH receptor-targeted and paclitaxel (PTX)-loaded lipid microbubbles (TPLMBs) for tumor-specific binding and enhanced therapeutic effect at the tumor site. An ovarian cancer xenograft model was established by injecting A2780/DDP cells intraperitoneally in BALB/c nude mice. Microscopic imaging of tumor sections after intraperitoneal injection of TPLMBs showed effective binding of the microbubbles with cancer cells. Ultrasound mediated destruction of the intraperitoneally injected TPLMBs yielded a superior therapeutic outcome in comparison with other treatment options. Immunohistochemical analyses of the dissected tumor tissue further confirmed the increased tumor apoptosis and reduced angiogenesis. Our experiment suggests that ultrasound-mediated intraperitoneal administration of the targeted drug-loaded microbubbles may be a useful method for the treatment of ovarian cancer.

Targeted microbubbles for ultrasound mediated gene transfection and apoptosis induction in ovarian cancer cells

Ultrasound-targeted microbubble destruction (UTMD) technique can be potentially used for non-viral delivery of gene therapy. Targeting wild-type p53 (wtp53) tumor suppressor gene may provide a clinically promising treatment for patients with ovarian cancer. However, UTMD mediated gene therapy typically uses non-targeted microbubbles with suboptimal gene transfection efficiency. We synthesized a targeted microbubble agent for UTMD mediated wtp53 gene therapy in ovarian cancer cells. Lipid microbubbles were conjugated with a Luteinizing Hormone-Releasing Hormone analog (LHRHa) via an avidin-biotin linkage to target the ovarian cancer A2780/DDP cells that express LHRH receptors. The microbubbles were mixed with the pEGFP-N1-wtp53 plasmid. Upon exposure to 1 MHz pulsed ultrasound beam (0.5 W/cm(2)) for 30s, the wtp53 gene was transfected to the ovarian cancer cells. The transfection efficiency was (43.90 ± 6.19)%. The expression of wtp53 mRNA after transfection was (97.08 ± 12.18)%. The cell apoptosis rate after gene therapy was (39.67 ± 5.95)%. In comparison with the other treatment groups, ultrasound mediation of targeted microbubbles yielded higher transfection efficiency and higher cell apoptosis rate (p<0.05). Our experiment verifies the hypothesis that ultrasound mediation of targeted microbubbles will enhance the gene transfection efficiency in ovarian cancer cells.

Ultrasound-mediated destruction of paclitaxel and oxygen loaded lipid microbubbles for combination therapy in ovarian cancer xenografts

We have synthesized multifunctional oxygen and paclitaxel loaded microbubbles (OPLMBs) for ultrasound mediated delivery of combination therapy in an ovarian cancer xenograft model. In comparison with other therapeutic options, intravenous injection of OPLMBs followed by ultrasound mediation yielded a superior therapeutic outcome. Immunohistochemical analyses of the dissected tumor tissue confirmed the increased tumor apoptosis and the reduced VEGF expression after treatment. Western Blot tests further confirmed the decreased expressions of HIF-1α and P-gp. Our experiment suggests that ultrasound mediation of OPLMBs may provide a promising drug delivery strategy for the combination treatment of ovarian cancer.

Ultrasound-mediated destruction of LHRHa-targeted and paclitaxel-loaded lipid microbubbles induces proliferation inhibition and apoptosis in ovarian cancer cells.

Although paclitaxel (PTX) is used with platinum as the first line chemotherapy regimen for ovarian cancer, its clinical efficacy is often limited by severe adverse effects. Ultrasound-targeted microbubble destruction (UTMD) technique holds a great promise in minimizing the side effects and maximizing the therapeutic efficacy. However, the technique typically uses nontargeted microbubbles with suboptimal efficiency. We synthesized targeted and PTX-loaded microbubbles (MBs) for UTMD mediated chemotherapy in ovarian cancer cells. PTX-loaded lipid MBs were coated with a luteinizing hormone-releasing hormone analogue (LHRHa) through a biotin-avidin linkage to target the ovarian cancer A2780/DDP cells that express the LHRH receptor. In the cell culture studies, PTX-loaded and LHRHa-targeted MBs (TPLMBs) in combination with ultrasound (300 kHz, 0.5 W/cm(2), 30 s) demonstrated antiproliferative activities of 41.30 ± 3.93%, 67.76 ± 2.45%, and 75.93 ± 2.81% at 24, 48, and 72 h after the treatment, respectively. The cell apoptosis ratio at 24 h after the treatment is 32.6 ± 0.79%, which is significantly higher than other treatment groups such as PTX only and no-targeted PTX-loaded MBs (NPLMBs) with or without ultrasound mediation. Our experiment verifies the hypothesis that ultrasound mediation of ovarian cancer-targeted and drug-loaded MBs will enhance the PTX therapeutic efficiency.

Ultrasound mediated destruction of multifunctional microbubbles for image guided delivery of oxygen and drugs

We synthesized multifunctional activatible microbubbles (MAMs) for ultrasound mediated delivery of oxygen and drugs with both ultrasound and fluorescence imaging guidance. Oxygen enriched perfluorocarbon (PFC) compound was encapsulated in liposome microbubbles (MBs) by a modified emulsification process. DiI dye was loaded as a model drug. The ultrasound targeted microbubble destruction (UTMD) process was guided by both ultrasonography and fluorescence imaging modalities. The process was validated in both a dialysis membrane tube model and a porcine carotid artery model. Our experiment results show that the UTMD process effectively facilitates the controlled delivery of oxygen and drug at the disease site and that the MAM agent enables ultrasound and fluorescence imaging guidance of the UTMD process. The proposed MAM agent can be potentially used for UTMD-mediated combination therapy in hypoxic ovarian cancer.

Indocyanine-green-loaded microballoons for biliary imaging in cholecystectomy

Abstract. We encapsulate indocyanine green (ICG) in poly[(D,L-lactide-co-glycolide)-co-PEG] diblock (PLGA-PEG) microballoons for real-time fluorescence and hyperspectral imaging of biliary anatomy. ICG-loaded microballoons show superior fluorescence characteristics and slower degradation in comparison with pure ICG. The use of ICG-loaded microballoons in biliary imaging is demonstrated in both biliary-simulating phantoms and an ex vivo tissue model. The biliary-simulating phantoms are prepared by embedding ICG-loaded microballoons in agar gel and imaged by a fluorescence imaging module in a Da Vinci surgical robot. The ex vivo model consists of liver, gallbladder, common bile duct, and part of the duodenum freshly dissected from a domestic swine. After ICG-loaded microballoons are injected into the gallbladder, the biliary structure is imaged by both hyperspectral and fluorescence imaging modalities. Advanced spectral analysis and image processing algorithms are developed to classify the tissue types and identify the biliary anatomy. While fluorescence imaging provides dynamic information of movement and flow in the surgical region of interest, data from hyperspectral imaging allow for rapid identification of the bile duct and safe exclusion of any contaminant fluorescence from tissue not part of the biliary anatomy. Our experiments demonstrate the technical feasibility of using ICG-loaded microballoons for biliary imaging in cholecystectomy.

Ultrasound-mediated destruction of oxygen and paclitaxel loaded lipid microbubbles for combination therapy in hypoxic ovarian cancer cells

We synthesized oxygen and paclitaxel (PTX) loaded lipid microbubbles (OPLMBs) for ultrasound mediated combination therapy in hypoxic ovarian cancer cells. Our experiments successfully demonstrated that ultrasound induced OPLMBs destruction significantly enhanced the local oxygen release. We also demonstrated that OPLMBs in combination with ultrasound (300 kHz, 0.5 W/cm(2), 15s) yielded anti-proliferative activities of 52.8 ± 2.75% and cell apoptosis ratio of 35.25 ± 0.17% in hypoxic cells at 24h after the treatment, superior to other treatment groups such as PTX only and PTX-loaded MBs (PLMBs) with or without ultrasound mediation. RT-PCR and Western blot tests further confirmed the reduced expression of HIF-1α and MDR-1/P-gp after ultrasound mediation of OPLMBs. Our experiment suggests that ultrasound mediation of oxygen and drug-loaded MBs may be a useful method to overcome chemoresistance in the hypoxic ovarian cancer cells.

Targeted Microbubbles for Ultrasound Mediated Short Hairpin RNA Plasmid Transfection to Inhibit Survivin Gene Expression and Induce Apoptosis of Ovarian Cancer A2780/DDP Cells

Nonviral gene transfer by ultrasound-targeted microbubble destruction (UTMD) is an promising technique for RNA interference (RNAi) therapy. Targeting silence survivin gene may provide an important therapeutic option for patients with ovarian cancer. However, UTMD mediated RNAi therapy typically uses nontargeted microbubbles with suboptimal gene transfection efficiency. In this work, a LHRHa targeted microbubble agent and recombinant expression plasmid of shRNA targeting survivin gene (pshRNA survivin) were constructed for UTMD mediated pshRNA survivin therapy in ovarian cancer A2780/DDP cells that express LHRH receptors. The targeted microbubbles (TMBs) mixed with the pshRNA survivin were added to cultured ovarian cancer cells followed by ultrasound exposure (1 MHz, 0.5 W/cm(2)) for 30 s. After transfection for 48 h, the expression of survivin mRNA and protein were (0.36 ± 0.036) and (0.05 ± 0.02), respectively. The cell proliferation inhibitory rates at 24, 48, and 72 h after treatment are (42.08 ± 3.20)%, (54.60 ± 1.02)%, and (74.25 ± 2.14)%, respectively, and the apoptosis rate was (28.99 ± 2.70)%. The expression of apoptosis related protein caspase-9 and caspase-3 were (0.95 ± 0.09) and (2.6 ± 0.21). In comparison with the other treatment groups, ultrasound mediation of targeted microbubbles yielded higher RNAi efficiency and higher cell apoptosis rate and cell proliferation inhibitory rate (p < 0.05). Our experiment verifies the hypothesis that ultrasound mediation of targeted microbubbles will enhance RNAi efficiency in ovarian cancer cells. This novel method for RNA interference represents a powerful, promising no viral technology that can be used in the tumor gene therapy and research.

Oxygen and Indocyanine Green loaded microparticles for dual-mode imaging and sonodynamic treatment of cancer cells

Oxygen and Indocyanine Green (ICG) loaded microparticles (OI-MPs) were fabricated by a gas-driven coaxial flow focusing (CFF) process for dual-mode imaging and sonodynamic therapy (SDT). The produced OI-MPs agent showed stable optical properties, superior imaging depth in near infrared (NIR) fluorescence imaging, and enhanced acoustic contrast after ultrasound mediation. We hypothesized that encapsulating ICG and oxygen in microparticles would enhance reactive oxygen species (ROS) production in SDT. This hypothesis was validated in a cell-free environment. We further hypothesized that ultrasound mediated fragmentation of the OI-MPs would induce cytotoxicity and apoptosis of cancer cells. This hypothesis was validated in SKOV3 ovarian cancer cells. Our research demonstrated that OI-MPs can be potentially used as a dual-mode theranostic agent for image guided SDT with enhanced efficacy. Further study is needed to delineate the mechanism of ROS-induced cell apoptosis and optimize the OI-MPs formulation for the maximal anti-cancer potency.

Oxygen and indocyanine green loaded phase-transition nanoparticle-mediated photo-sonodynamic cytotoxic effects on rheumatoid arthritis fibroblast-like synoviocytes

Background Photodynamic therapy and sonodynamic therapy are developing, minimally invasive, and site-specific modalities for cancer therapy. A combined strategy PSDT (photodynamic therapy followed by sonodynamic therapy) has been proposed in this study. Here, we aimed to develop novel biodegradable poly(DL-lactide-co-glycolic acid) phase-transition nanoparticles simultaneously loaded with oxygen and indocyanine green (OI-NPs) and to investigate the cytotoxic effects and the potential mechanisms of OI-NP–mediated PSDT on MH7A synoviocytes. Methods The OI-NPs were prepared using a modified double emulsion method and the physicochemical properties were determined. The cellular uptake of OI-NPs was detected by confocal microscopy and flow cytometry. 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazoliumbromide assay, flow cytometry, and Hoechst 33342/propidium iodide double staining were used to determine the cytotoxic effect of OI-NP–mediated PSDT on MH7A cells. Fluorescence microscope and fluorescence microplate reader were used to detect reactive oxygen species (ROS) generation. Results The OI-NPs were a stable and efficient carrier to deliver oxygen and indocyanine green, and enhanced cellular uptake was observed in MH7A cells with the nanoparticles. OI-NP–mediated PSDT caused more serious cell damage and more evident cell apoptosis, compared with other groups. Furthermore, increased generation of intracellular ROS was detected in MH7A cells treated with PSDT. Interestingly, the OI-NP–mediated PSDT-induced cell viability loss was effectively rescued by pretreatment with the ROS scavenger N-acetylcysteine. Conclusion Multifunctional OI-NPs were successfully developed and characterized for the combined delivery of oxygen and indocyanine green, and OI-NP–mediated PSDT would be a potential cytotoxic treatment for MH7A cells. This study may provide a novel strategy for the treatment of RA and develop a model of theranostic application through phase-transition nanoparticle-mediated PSDT in the future.