Imaging-Guided RNAi Therapy By Using High Density Lipoprotein-mimicking Nanoparticles

RNAi therapeutics have been hailed as the personalized medicine of the future and have shown early promise in clinical trials. However, challenges remain in systemic delivery of the siRNA to the correct tissues and transporting them into the cytoplasm of targeted cells, at safe, therapeutic levels. In addition, to conceive and optimize experimental treatment strategies, RNAi therapeutics development also requires noninvasive assessment of the siRNA delivery to tissues of interest. Herein, we developed a multifunctional HDL-like theranostic nanoparticle by integrating near-infrared (NIR) fluorescence probe and therapeutic siRNA on high-density lipoprotein-mimicking peptide-phospholipid scaffold (HPPS) nanoparticles for better RNAi therapeutics by efficient systemic delivery of siRNA and imaging-guided RNAi therapy strategy.1) We demonstrated its direct cytosolic delivery of siRNA in vitro, thereby bypassing endosomal trapping. Our study first elucidates this unique delivery mechanism for improving cytosolic drug delivery. Multifluorophore-labeled HPPS nanoparticles were developed. Fluorescence imaging was utilized to examine HPPS transporting payloads into cells step by step through sequential inhibition studies. HPPS specifically recognizes and binds to SR-BI, then interacts with SR-BI, which results in direct transport of payload molecules into the cell cytoplasm without entire particles internalization. The cytosolic transport of payloads occurred through a temperature-and energy-independent pathway, and was also different from actin-and clathrin-mediated endocytosis. Furthermore, this transport was significantly inhibited by disruption of lipid rafts using filipin or methyl-β-cyclodextrin. And demonstrated the cytosolic delivery of payloads by HPPS via scavenger receptor class B type I (SR-BI) targeting is predominately mediated through a lipid rafts/caveolae-like pathway by multifluorophore-labelling method and sequential inhibition studies.2) Next, We investigate the in vivo implementation of HPPS for siRNA delivery in a KB xenograft cancer model, and demonstrated that HPPS prolonged the blood circulation time of cholesterol-modified siRNA (chol-siRNA) by a factor of four, improved its biodistribution and facilitated its uptake in SR-BI overexpressed tumors. For therapeutic targeting to the bcl-2gene, the HPPS-chol-si-bcl-2nanoparticles down regulated Bcl-2protein, induced enhanced apoptosis (2.5-fold) in tumors when compared with controls (saline, HPPS, HPPS-chol-si-scramble and chol-si-bcl-2) and significantly inhibited tumor growth with no adverse effect. So HPPS is a safe, efficient nanocarrier for RNAi therapeutics in vivo.3) Finally, we investigate the imaging-guided RNAi therapy by our nanoplatform in a more clinical revalent-PC3orthotopic prostate cancer model, and demonstrated this nanoplatform can selectively target to and accumulate in tumor to achieve higher target drug concentrations with lower overall body exposure. The NIR fluorescence was only observed in malignant tumor cells, whereas healthy prostate tissue, SV and testis showed little to no fluorescence, allowing in vivo detection of orthotopic tumor by non-invasive FMT/CT (Fluorescence Molecular Tomography/Computed Tomography) dual imaging technique and further assessment of drug delivery in deep tissues. Simultaneously, the ferried bcl-2-siRNA significantly knocked down the bcl-2gene in tumor, induced tumor tissue apoptosis, and ultimately resulted in tumor growth hindrance. Such multimodal HDL-like nanotheranostic platform provides a useful tool in personalized medicine for detection of tumor, assessment of drug delivery, effective treatment planning, and finally evaluation of therapeutic response.