Radiolabeled AMO-phospholipid enveloped nanoparticle for cellular delivery and in vivo imaging in cervical cancer xenografts

Due to low ability to pass through the cell membrane and lack of proof in target delivery in vivo, anti-microRNA oligonucleotide (AMO) has poor target ability to microRNA in the cytoplasm, which results in low AMO targeting efficiency and limits AMO success in vivo application. In this study, AMO-phospholipids conjugates were developed to provide the characteristics of the two phospholipids to self -assemble via hydrophilic AMO and hydrophobic phospholipid tails to overcome the AMO backbone structures and enhance the AMO delivery efficiency. This AMO-phospholipids enveloped nanoparticles was radiolabeled with technetium -99 m (99mTc) for cellular delivery and in vivo imaging. Methods: 99mTc-radiolabeled miR-155 targeted AMO with partial 2 '-O-methyl (2 '-OMe) and phosphorothioate (PS) chemical modification was prepared via the conjugation to N-hydroxysuccinimide derivative of S-acetylmercaptoacetyltriglycine (NHS-MAG3). This radiolabeled AMO is chemically conjugated with phospholipids to form AMO-phospholipid. The AMO-phospholipids served as a component of nanoparticles to embed the hydrophobic part. AMO-phospholipids fused with polycarboxybetaine (PCB) based distearoyl phosphoethanolamine-polycarboxybetaine (DSPE-PCB) modified cationic lipids to form AMO-phospholipidcoated nanoparticles (AMO-NP), which delivered AMO into cytoplasm in vitro and into tumor in vivo. Further, this radiolabeled AMO-NP was confirmed by gel electrophoresis and evaluated for its serum stability, cytotoxicity, inhibitory ability and cellular uptake in HeLa cells, as well as the fluorescent distribution in vitro. Moreover, its distribution and in vivo imaging were performed HeLa tumor -bearing mice after the injection of radiolabeled AMO-NP. Results: The gel electrophoresis of 99mTc labeled AMO-NP showed good conjugation between AMO and 99mTc. It also kept highly stable in human serum. Western blotting results showed that AMO-NP could up -regulate the expression of C/EBP beta protein, one of target proteins of miR-155 in HeLa cells, proving the ability in the specific binding and inhibition. The cellular uptake of 99mTc labeled AMO-NP was significantly higher than that of bare AMO. Furthermore, FAM labeled AMO-NP displayed its ability of specific and effective distribution in tumor cells and a good stability in cellular level. After the system administration, the distribution and in vivo imaging results revealed the significant difference between 99mTc labeled AMO-NP and negative control probe in HeLa tumor xenograft models, which suggested the specific binding and effective delivery ability of 99Tc labeled AMO-NP in vivo. Conclusion: This study supports the effectiveness and efficiency of cellular and in vivo delivery of 99mTc radiolabeled AMO-NP in cervical tumor and suggests a prospective candidate for future tumor imaging.