Effect Of C-terminal Polyarginine Domain On Indocyanine Green Encapsulation With HBc VLPs

Indocyanine green(ICG)is a safe and effective in vivo imaging agent.However,the instability in aqueous solution and rapid clearance rate in blood limit its clinical application.In recent years,the continuous development of bionic nanotechnology provides a new idea for the integration of cancer diagnosis and treatment.Hepatitis B virus core protein virus-like nanoparticles(HBc VLPs),as natural protein biomolecules with high safety and biocompatibility,have been proved by our experimental team that it can be used to load ICG.However,the relationship between the structure of HBc VLPs and the encapsulated amount of ICG is still not clear,and there are few related studies.It has been shown that the C-terminal polyarginine domain(CTD)of HBc VLPs affect the charge distribution without affecting particle self-assembly,which regulated the ability of particles to bind nucleic acids.Therefore,it is expected to regulate the drug loading efficiency of HBc VLPs by regulating the amino acid sequence of CTD.In this study,three RGD-HBc VLPs(RH VLPs)with different C-terminal lengths were prepared by truncating the CTD of HBc VLPs and inserting the tumor-targeting peptide RGD.ICG was loaded into three RH VLPs to study the differences in encapsu-lation amounts and drug activity among the nanocomplexes.The result showed that RH183 VLPs,RH160 VLPs and RH144 VLPs had good self-assembly characteristics and uniform particle size with 30.90±1.33 nm,32.95±0.77 nm and 35.88±1.75 nm,respectively.The ability of RH VLPs to bind nucleic acids was decreased with the truncation of CTD,which may be related to the fact that the truncation of CTD leads to fewer arginine blocks provided by RH VLPs.The encapsulation result showed that the encapsulation efficiency of RH160/ICG was 32.77±1.23%,and was 1.15 times and 1.68 times that of RH183/ICG and RH144/ICG,respectively.Excessive nucleic acids may occupy the internal space of particles and arginine fragments,resulting in reduced encapsulation efficiency of RH183/ICG.In vitro release studies demonstrated that RH183/ICG and RH160/ICG could stably and efficiently release ICG in acidic environments,while RH144/ICG was easy to release drugs in physiological environme-nts,which reduced the effective amount of drug reaching tumor tissues.In vitro cell experiments showed that 4T1 mouse breast cancer cells high-efficiently taken up RH160/ICG,which significantly increased cell surface temperature and produced ROS after 10 min of 1.5 W/cm~2laser irradiation.The lowest cell survival rate with 200μg/m L RH160/ICG treatment reached 24.73±9.08%,which effectively killed tumor cells.In vivo imaging and photothermal transformation experiment showed that RH160/ICG significantly enhanced the accumulation ability and prolonged the in vivo fluorescence retention time of ICG at the tumor site.The temperature of mice tumor site in the RH160/ICG group effectively increased with more than 43℃ after 10 min of 1.5 W/cm~2 laser irradiation,which indicated that RH160/ICG had superior biological tumor imaging and in vivo photothermal transformation ability.In conclusion,truncating CTD regulate the efficiency of drug encapsulation with RGD-HBc VLPs,which affects the drug activity of nanocomplexes.The highest encapsulation efficiency of RH/ICG VLPs reach 32.77±1.23%when truncating HBc VLPs160 aa.Moreover,RH160/ICG has significant advantages in tumor imaging and photothermal transformation.This study provides a new strategy for the research of the nanomedicine carrier platform based on HBc VLPs and further expanding the diagnostic and therapeutic applications of ICG.