Multifunctional nanobubbles carrying indocyanine green and paclitaxel for molecular imaging and the treatment of prostate cancer.

BACKGROUND: Combining ultrasound imaging with photoacoustic imaging provides tissue imaging with high contrast and resolution, thereby enabling rapid, direct measurements and the tracking of tumour growth and metastasis. Moreover, ultrasound-targeted nanobubble destruction (UTND) provides an effective way to deliver drugs, effectively increasing the content of the drug in the tumour area and reducing potential side effects, thereby successfully contributing to the treatment of tumours. RESULTS: In this study, we prepared multifunctional nanobubbles (NBs) carrying indocyanine green (ICG) and paclitaxel (PTX) (ICG-PTX NBs) and studied their applications in ultrasound imaging of prostate cancer as well as their therapeutic effects on prostate cancer when combined with UTND. ICG-PTX NBs were prepared by the mechanical oscillation method. The particle size and zeta potential of the ICG-PTX NBs were 469.5 ± 32.87 nm and - 21.70 ± 1.22 mV, respectively. The encapsulation efficiency and drug loading efficiency of ICG were 68% and 2.52%, respectively. In vitro imaging experiments showed that ICG-PTX NBs were highly amenable to multimodal imaging, including ultrasound, photoacoustic and fluorescence imaging, and the imaging effect was positively correlated with their concentration. The imaging effects of tumour xenografts also indicated that ICG-PTX NBs were of good use for multimodal imaging. In experiments testing the growth of PC-3 cells in vitro and tumour xenografts in vivo, the ICG-PTX NBs + US group showed more significant inhibition of cell proliferation and the promotion of cell apoptosis compared to the other groups (P < 0.05). Blood biochemical analysis of the six groups showed that the levels of aspartate aminotransferase (AST), phenylalanine aminotransferase (ALT), serum creatinine (CRE) and blood urea nitrogen (BUN) in the ICG-PTX NBs and the ICG-PTX NBs + US groups were significantly lower than those in the PTX group (P < 0.05). Moreover, H&E staining of tissue sections from vital organs showed no obvious abnormalities in the ICG-PTX NBs and the ICG-PTX NBs + US groups. CONCLUSIONS: ICG-PTX NBs can be used as a non-invasive, pro-apoptotic contrast agent that can achieve multimodal imaging, including ultrasound, fluorescence and photoacoustic imaging, and can succeed in the local treatment of prostate cancer providing a potential novel method for integrated research on prostate cancer diagnosis and treatment.

Construction of Nucleolin-Targeted Lipid Nanobubbles and Contrast-Enhanced Ultrasound Molecular Imaging in Triple-Negative Breast Cancer.

PURPOSE: To construct aptamer AS1411-functionalized targeted lipid nanobubbles that could simultaneously target abnormally highly expressed nucleolin (NCL) on tumor tissue and neovasculature. Additionally, the study of their contrast-enhanced ultrasound molecular imaging capabilities in vitro and in vivo to explore new methods and approaches for the early and accurate diagnosis of triple-negative breast cancer (TNBC). METHODS: First, the targeted lipid-nucleic acid molecules were constructed by an amide reaction. Then, the targeted lipid nanobubbles (AS1411-NBs) and nontargeted lipid nanobubbles (NBs) were prepared by membrane hydration, mechanical vibration and centrifugal floatation. The physicochemical characteristics and contrast-enhanced ultrasound imaging capabilities of AS1411-NBs and NBs were compared and analyzed in vitro and in vivo. RESULTS: There were no significant differences between the AS1411-NBs and NBs in their concentration, average particle size or ultrasound imaging capabilities in vitro (P > 0.05). However, AS1411-NBs could simultaneously target NCL in tumor tissue and neovasculature to effectively prolong the duration of contrast-enhanced ultrasound imaging compared to NBs in vivo. The area under the time-intensity curve was significantly different between AS1411-NBs and NBs (P < 0.001), and the drug loading capacity of the AS1411-NBs was also significantly higher than that of the NBs (P < 0.05). CONCLUSIONS: Aptamer AS1411-functionalized targeted lipid nanobubbles could significantly prolong the duration of contrast-enhanced ultrasound imaging to achieve dual-targeted ultrasound molecular imaging of tumor tissue and neovasculature. AS1411-NBs also have higher drug loading and targeted drug delivery capabilities compared with NBs, which can provide new methods and approaches for the early accurate diagnosis and effective treatment of TNBC.

Targeted Nanobubbles Carrying Indocyanine Green for Ultrasound, Photoacoustic and Fluorescence Imaging of Prostate Cancer.

OBJECTIVE: To construct prostate-specific membrane antigen (PSMA)-targeting, indocyanine green (ICG)-loaded nanobubbles (NBs) for multimodal (ultrasound, photoacoustic and fluorescence) imaging of prostate cancer. METHODS: The mechanical oscillation method was used to prepare ICG-loaded photoacoustic NBs (ICG NBs). Then, PSMA-binding peptides were connected to the surface of ICG NBs using the biotin-avidin method to make targeted photoacoustic NBs, namely, PSMAP/ICG NBs. Their particle sizes, zeta potentials, and in vitro ultrasound, photoacoustic and fluorescence imaging were examined. Confocal laser scanning microscopy and flow cytometry were used to detect the binding ability of the PSMAP/ICG NBs to PSMA-positive LNCaP cells, C4-2 cells, and PSMA-negative PC-3 cells. The multimodal imaging effects of PSMAP/ICG NBs and ICG NBs were compared in nude mouse tumor xenografts. RESULTS: The particle size of the PSMAP/ICG NBs was approximately 457.7 nm, and the zeta potential was approximately -23.5 mV. Both confocal laser scanning microscopy and flow cytometry confirmed that the PSMAP/ICG NBs could specifically bind to both LNCaP and C4-2 cells, but they rarely bound to PC-3 cells. The ultrasound, photoacoustic and fluorescence imaging intensities of the PSMAP/ICG NBs in vitro positively correlated with their concentrations. The ultrasound and photoacoustic imaging effects of the PSMAP/ICG NBs in LNCaP and C4-2 tumor xenografts were significantly enhanced compared with those in PC-3 tumor xenografts, which were characterized by a significantly increased duration of ultrasound enhancement and heightened photoacoustic signal intensity (P < 0.05). Fluorescence imaging showed that PSMAP/ICG NBs could accumulate in LNCaP and C4-2 tumor xenografts for a relatively long period. CONCLUSION: The targeted photoacoustic nanobubbles prepared in this study can specifically bind to PSMA-positive prostate cancer cells and have the ability to enhance ultrasound, photoacoustic and fluorescence imaging of PSMA-positive tumor xenografts. Photoacoustic imaging could visually display the intensity of the red photoacoustic signal in the tumor region, providing a more intuitive imaging modality for targeted molecular imaging. This study presents a potential multimodal contrast agent for the accurate diagnosis and assessment of prostate cancer.

Efficacy evaluation and mechanism study on inhibition of breast cancer cell growth by multimodal targeted nanobubbles carrying AMD070 and ICG.

To construct targeted nanobubbles carrying both small-molecule CXCR4 antagonist AMD070 and light-absorbing material indocyanine green (ICG), and to study their in vitro multimodal imaging, as well as their mechanism and efficacy of inhibition of breast cancer cell growth. Nanobubbles carrying AMD070 and ICG (ICG-TNBs) were constructed by carbodiimide reaction and mechanical oscillation. The physical characteristics and in vitro multimodal imaging were determined. The binding potential of ICG-TNBs to human breast cancer cells were observed by laser confocal microscopy. CCK-8 and flow cytometry were used to analyze the role of ICG-TNBs + US in inhibiting proliferation and inducing apoptosis of tumor cells. Flow cytometry and Western blotting are used to analyse the ROS generation and molecular mechanisms. ICG-TNBs had a particle size of 497.0 ± 29.2 nm and a Zeta potential of -8.05 ± 0.73 mV. In vitro multimodal imaging showed that the image signal intensity of ICG-TNBs increased with concentration. Targeted binding assay confirmed that ICG-TNBs could specifically bind to MCF-7 cells (CXCR4 positive), but not to MDA-MB-468 cells (CXCR4 negative). CCK-8 assay and flow cytometry analysis showed that ICG-TNBs + US could significantly inhibit the growth of MCF-7 breast cancer cells and promote their apoptosis. Flow cytometry and Western blotting showed that ICG-TNBs + US could significantly raise generation of ROS, reduce the expression of CXCR4, inhibit phosphorylation of Akt, and increase the expression of Caspase3 and Cleaved-caspase3. This indicated that ICG-TNBs could effectively inhibit and block the SDF-1/CXCR4 pathway, thus leading to the apoptosis of MCF-7 cells. ICG-TNBs can specifically bind to CXCR4 positive breast cancer cells, furthermore inhibit growth and promote apoptosis of breast cancer cells combined with ultrasonic irradiation by blocking the SDF-1/CXCR4 pathway. This study introduces a novel concept, method and mechanism for integration of targeted diagnosis and treatment of breast cancer.

Preparation Of Nanobubbles Modified With A Small-Molecule CXCR4 Antagonist For Targeted Drug Delivery To Tumors And Enhanced Ultrasound Molecular Imaging.

PURPOSE: To construct nanobubbles (PTX-AMD070 NBs) for targeted delivery of paclitaxel (PTX) and AMD070, examine their performance in ultrasound molecular imaging of breast cancer and cervical cancer and their therapeutic effect combined with ultrasound targeted nanobubble destruction (UTND). MATERIALS AND METHODS: PTX-AMD070 NBs were prepared via an amide reaction, and the particle size, zeta potential, encapsulation rate and drug loading efficiency were examined. Laser confocal microscopy and flow cytometry were used to analyze the targeted binding ability of PTX-AMD070 NBs to CXCR4+ MCF-7 cells and C33a cells. The effect of PTX-AMD070 NBs combined with UTND on cell proliferation inhibition and apoptosis induction was detected by CCK-8 assays and flow cytometry. The contrast-enhanced imaging features of PTX-AMD070 NBs and paclitaxel-loaded nanobubbles were compared in xenograft tumors. The penetration ability of PTX-AMD070 NBs in xenograft tissues was evaluated by immunofluorescence. The therapeutic effect of PTX-AMD070 NBs combined with UTND on xenograft tumors was assessed. RESULTS: PTX-AMD070 NBs showed a particle size of 494.3±61.2 nm, a zeta potential of -22.4±1.75 mV, an encapsulation rate with PTX of 53.73±7.87%, and a drug loading efficiency with PTX of 4.48±0.66%. PTX-AMD070 NBs displayed significantly higher targeted binding to MCF-7 cells and C33a cells than that of PTX NBs (P<0.05), and combined with UTND manifested a more pronounced effect in inhibiting cell proliferation and promoting apoptosis than other treatments. PTX-AMD070 NBs aggregated specifically in xenograft tumors in vivo, and significantly improved the image quality. Compared with other treatment groups, PTX-AMD070 NBs combined with UTND exhibited the smallest tumor volume and weight, and the highest degree of apoptosis and necrosis. CONCLUSION: PTX-AMD070 NBs improved the ultrasound imaging effect in CXCR4+ xenograft tumors and facilitated targeted therapy combined with UTND. Therefore, this study provides an effective method for the integration of ultrasound molecular imaging and targeted therapy of malignant tumors.