Ruthenium Complexes And Multifunctional Nanoparticles Loaded With Drugs And SiRNA For Anti-tumor Activity And Its Targeting Imaging

Malignant tumor is one of the most common types of threat for human health. However, current chemotherapy drugs administered in this fashion are toxic to many healthy cell types, often resulting in severe side effects for the patient. The resistance of neoplastic cells to chemotherapeutic agents is a significant problem in the clinical management of malignant tumors. The traditional single-target antitumor drugs is very difficult to achieve a ideal therapeutic effect. Therefore, drug development of antitumor drugs with high efficiency, low toxicity and strong target is the focus of the future research.Among the potential metal-based candidates, ruthenium, possesses several favorable properties suited to rational anti-cancer drug design, and ruthenium complexes have emerged as leading players by showing promising results. Ruthenium complexes could induced telomere DNA forming G-quadrome structure to inhibit the activity if telomerase, and a number of ruthenium complexes have shown potential utility in chemotherapy and photodynamic therapy as evidenced by in vitro and in vivo studies, another advantage of Ruthenium complexes is that can selectively inhibit tumor invasion and metastasis.RNA interference(RNAi) is a powerful tool for sequence-specific suppression of genes and has potential applications for targeted gene therapy. Nanoparticles delivery system load siRNA and antitumor drugs, and combining with use recognition targets molecular, effectively improve the therapeutic effect of traditional antitumor drugs. These theories lay a substantial theory basis for exploitation and application of composite nanoparticles for cancer diagnosis and treatment.Based on the above research background, this paper we studied interaction and mechanism of Ruthenium(II) complexes with human telomere sequence HTG21 DNA; We developed an siRNA-delivery system for VEGF, that consists of two selenium nanoparticles SeNPs and G2/PAH-Cit/SeNPs. G2/PAH-Cit/SeNPs is a pH-sensitive delivery system that is capable of enhancing siRNA loading, thus increasing siRNA release efficiency and subsequent target gene silencing both in vitro and in vivo; Magnetic nanoparticle delivery sysetems loaded with double chemotherapy drugs DOX and siRNA for tumor magnetic targeted therapy and multimodal imaging.This thesis consists of five chapters:Chapter 1. We briefly summarized the G-quadruplex structure characteristics and biological function, introduced the vascular growth factor VEGF and bFGF effects on tumor blood vessels and the significance roles of β-catenin genetic in the tumorigenesis, development; mainly introduced the research progress of gene therapy for tumor and detailed necessary factors of the nanoparticles for genes and drug delivery systems.Chapter 2. This study aims to clarifythe anticancer effect of metal complexes as noveland potent telomerase inhibitorsand cellular nucleus target drug.First, the chiral selectivity of the compounds and their ability to stabilize quadruplex DNA were studied via absorption and emission analyses, circular dichroism spectroscopy, fluorescence-resonance energy transfer melting assay, electrophoretic mobility shift assay, and polymerase chain reaction stopassay. The two chiral compounds selectively induced and stabilized the G-quadruplex of telomeric DNA with or without metal cations. These results provide newinsights into the development of chiral anticanceragents for G-quadruplex DNA targeting. Telomerase repeat amplification protocol reveals the higher inhibitory activity of Λ-[Ru(phen)2(p-HPIP)]2+ against telomerase, suggesting that Λ-[Ru(phen)2(p-HPIP)]2+may be a potential telomerase inhibitor for cancer chemotherapy. MTT assay results show that these chiral complexes have significant antitumor activities in HepG2 cells. More interestingly, cellular uptakeand laser-scanning confocal microscopic studies reveal the efficient uptake of ?-[Ru(phen)2(p-HPIP)]2+ by HepG2 cells. This complex then enters the cytoplasm and tends to accumulate in the nucleus. This nuclear penetration of the ruthenium complexes and their subsequent accumulation are associated with the chirality of the isomers as well as with the subtle environment of the ruthenium complexes. Therefore, the nucleus can be the cellulartarget of chiral ruthenium complexes for anticancer therapy.Chapter 3. Studies have shown that ruthenium complexes have relatively strong anticancer activity, cell uptake of drugs have a crucial impact on the pharmacological activity, using autofluorescent of ruthenium complexes could effectively track cancer cells and drug distribution, transport accurately in real time. In this work, we present the synthesis and detailed characterization of two novel Ru(II) complexes with hydrophobic ancillary ligands, namely [Ru(bpy)2(5-idip)]2+(RBD) and [Ru(phen)2(5-idip)]2+(RPD)(5-idip = 2-indole-[4,5-f][1,10]phenanthroline).We have showed that RPD can enter the HeLa cells efficiently through non-endocytotic, but energy-dependent mechanism and first accumulated in lysosomes, and then escaped from the lysosomes and localize within the nuclei, efficiently lead to the inhibition of DNA transcription and translation and induced cell apoptosis. Further studies on the mechanism of apoptosis in HeLa cells demonstrate that RPD is able to induce mitochondria-mediated apoptosis in He La cells through activation of initiator caspase-9 and down-stream effector caspase-3 and-7 and cleavage of PARP. We have also demonstrated that RPD should bind to telomeric G-quadruplex DNA effectively and selectively, together with increased p21 and p16 expression. Our findings suggest that RPD induces HeLa cell apoptosis through mitochondria-mediated pathway and inhibition of telomerase activity. RPD may be a candidate for further evaluation as a chemotherapeutic agent for human cancers.Chapter 4. The utility of small interfering RNAs(siRNAs) has shown great promise in treating a variety of diseases including many types of cancer. While their ability to silence a wide range of target genes underlies their effectiveness, therapies application remain hindered by a lack of an effective delivery system. In this study, we sought to develop an siRNA-delivery system for VEGF, a known signaling molecule involved in cancer, that consists of two selenium nanoparticles SeNPs and G2/PAH-Cit/SeNPs. G2/PAH-Cit/SeNPs is a pH-sensitive delivery system that is capable of enhancing siRNA loading, thus increasing siRNA release efficiency and subsequent target gene silencing both in vitro and in vivo. In vivo experiments using G2/PAHCit/SeNPs@siRNA led to significantly higher accumulation of siRNA within the tumor itself, VEGF gene silencing, and reduced angiogenesis in the tumor. Furthermore, the G2/PAHCit/SeNPs delivery system not only enhanced anti-tumor effects on tumor-bearing nude mice as compared to SeNPs@si RNA, but also resulted in weak occurrence of lesions in major target organs. In sum, this study provides a new class of siRNA delivery system, thereby providing an alternative therapeutic route for cancer treatment. Chapter 5.In the field of biomedical applications, we designed and synthesized a new magnetic targeting multifunctional nanoparticles Fe3O4@TiO2@mTiO2, using for loading both chemotherapy drug DOX and siRNA as delivery system, realizing synergistic therapeutic effect of DOX and siRNA. The follow-up experiments proved that delivery system Fe3O4@TiO2@mTiO2 effectively loaded and control released of drug and gene. Fe3O4@TiO2@mTiO2-siRNA/DOX enter the cell rapidly under the effect of additional magnetic field. Further, additional magnetic field speed up cell uptake of cancer cells, and had no effect on the absorption patterns of endocytosis.Fe3O4@TiO2@mTiO2-siRNA/DOX effectivly identified tumor tissues through both MRI imaging and fluorescence imaging. In vivo studies, magnetic delivery systems Fe3O4@TiO2@mTiO2-siRNA/DOX effective targeted tumor tissues under the effect of additional magnetic field, and no obvious side effects of normal organs. Results show that magnetic targeting delivery system used for the joint treatment of adriamycin and siRNA has significant therapeutic effect on tumor.