title: Novel lactoferrin-conjugated amphiphilic poly(aminoethyl ethylene phosphate)/poly(L-lactide) copolymer nanobubbles for tumor-targeting ultrasonic imaging.
abstract: In the study reported here, a novel amphiphilic poly(aminoethyl ethylene phosphate)/poly(L-lactide) (PAEEP-PLLA) copolymer was synthesized by ring-opening polymerization reaction. The perfluoropentane-filled PAEEP-PLLA nanobubbles (NBs) were prepared using the O1/O2/W double-emulsion and solvent-evaporation method, with the copolymer as the shell and liquid perfluoropentane as the core of NBs. The prepared NBs were further conjugated with lactoferrin (Lf) for tumor-cell targeting. The resulting Lf-conjugated amphiphilic poly(aminoethyl ethylene phosphate)/poly(L-lactide) nanobubbles (Lf-PAEEP-PLLA NBs) were characterized by photon correlation spectroscopy, polyacrylamide gel electrophoresis, Fourier transform infrared spectroscopy, and transmission electron microscopy. The average size of the Lf-PAEEP-PLLA NBs was 328.4±5.1 nm, with polydispersity index of 0.167±0.020, and zeta potential of -12.6±0.3 mV. Transmission electron microscopy imaging showed that the Lf-PAEEP-PLLA NBs had a near-spherical structure, were quite monodisperse, and there was a clear interface between the copolymer shell and the liquid core inside the NBs. The Lf-PAEEP-PLLA NBs also exhibited good biocompatibility in cytotoxicity and hemolysis studies and good stability during storage. The high cellular uptake of Lf-PAEEP-PLLA NBs in C6 cells (low-density lipoprotein receptor-related protein 1-positive cells) at concentrations of 0-20 µg/mL indicated that the Lf provided effective targeting for brain-tumor cells. The in vitro acoustic behavior of Lf-PAEEP-PLLA NBs was evaluated using a B-mode clinical ultrasound imaging system. In vivo ultrasound imaging was performed on tumor-bearing BALB/c nude mice, and compared with SonoVue(®) microbubbles, a commercial ultrasonic contrast agent. Both in vitro and in vivo ultrasound imaging indicated that the Lf-PAEEP-PLLA NBs possessed strong, long-lasting, and tumor-enhanced ultrasonic contrast ability. Taken together, these results indicate that Lf-PAEEP-PLLA NBs represent a promising nano-sized ultrasonic contrast agent for tumor-targeting ultrasonic imaging.
authors.affiliation: College of Life Science and Technology, Huazhong University of Science and Technology, Wuhan, People's Republic of China ; College of Pharmacy, Hubei University of Science and Technology, Xianning, People's Republic of China.
authors.forename: Binhua
authors.initials: B
authors.lastname: Luo
authors.affiliation: Department of Urology, Union Hospital of Tongji Medical College, Huazhong University of Science and Technology, Wuhan, People's Republic of China.
authors.forename: Huageng
authors.initials: H
authors.lastname: Liang
authors.affiliation: Department of Urology, Union Hospital of Tongji Medical College, Huazhong University of Science and Technology, Wuhan, People's Republic of China.
authors.forename: Shengwei
authors.initials: S
authors.lastname: Zhang
authors.affiliation: Department of Medical Ultrasound, Union Hospital of Tongji Medical College, Huazhong University of Science and Technology, Wuhan, People's Republic of China.
authors.forename: Xiaojuan
authors.initials: X
authors.lastname: Qin
authors.affiliation: College of Life Science and Technology, Huazhong University of Science and Technology, Wuhan, People's Republic of China.
authors.forename: Xuhan
authors.initials: X
authors.lastname: Liu
authors.affiliation: College of Life Science and Technology, Huazhong University of Science and Technology, Wuhan, People's Republic of China ; National Engineering Research Center for Nanomedicine, Huazhong University of Science and Technology, Wuhan, People's Republic of China.
authors.forename: Wei
authors.initials: W
authors.lastname: Liu
authors.affiliation: Department of Urology, Union Hospital of Tongji Medical College, Huazhong University of Science and Technology, Wuhan, People's Republic of China.
authors.forename: Fuqing
authors.initials: F
authors.lastname: Zeng
authors.affiliation: Department of Biomedical Engineering, University at Buffalo, State University of New York, Buffalo, NY, USA.
authors.forename: Yun
authors.initials: Y
authors.lastname: Wu
authors.affiliation: College of Life Science and Technology, Huazhong University of Science and Technology, Wuhan, People's Republic of China ; National Engineering Research Center for Nanomedicine, Huazhong University of Science and Technology, Wuhan, People's Republic of China.
authors.forename: Xiangliang
authors.initials: X
authors.lastname: Yang
mesh_terms: D000818:Animals; D049109:Cell Proliferation; D003952:Diagnostic Imaging / Q000379:methods*; D005030:Ethylenes; D005910:Glioma / Q000175:diagnosis*; D006461:Hemolysis / Q000187:drug effects; D061307:Human Umbilical Vein Endothelial Cells / Q000187:drug effects; D007781:Lactoferrin / Q000737:chemistry*; D051379:Mice; D008807:Mice, Inbred BALB C; D008819:Mice, Nude; D008823:Micelles; D045423:Microbubbles; D010710:Phosphates; D011091:Polyesters / Q000737:chemistry*; D011108:Polymers / Q000737:chemistry*; D017207:Rats, Sprague-Dawley; D014407:Tumor Cells, Cultured; D014465:Ultrasonics / Q000379:methods*
chemical_list: D005030:Ethylenes; C490098:LTF protein, human; D008823:Micelles; D010710:Phosphates; D011091:Polyesters; D011108:Polymers; C033616:poly(lactide); C036216:ethylene; D007781:Lactoferrin
keywords: PAEEP-PLLA copolymer; SonoVue® microbubbles; in vitro acoustic behavior; in vivo ultrasonic imaging