Development of Targeted siRNA Therapeutics for Triple Negative Breast Cancer and Liver Fibrosis

Abstract

The objective of this dissertation is to develop various systems to deliver small interfering RNA (siRNA) for the treatment of triple negative breast cancer (TNBC) and liver fibrosis. siRNAs targeting vascular endothelial growth factor (VEGF) and IκB kinase ε (IKBKE) were used for treating TNBC, while siRNA targeting poly(rC) binding protein 2 (PCBP2) was used for treating liver fibrosis. In Chapter 1, we briefly introduced the background about TNBC and liver fibrosis. We also presented the Statement of the Problems and Objectives. In Chapter 2, we reviewed the molecular mechanisms and potential treatments for TNBC and liver fibrosis. In Chapter 3, we developed a poly(ethyleneimine) (PEI) conjugated biodegradable multiblock polymer to form nanocomplexes with VEGF siRNA for TNBC treatment. The nanocomplex is able to deliver the VEGF siRNA into TNBC cells with a high transfection efficiency and low cytotoxicity. In vitro activity studies showed that the siRNA nanocomplexes significantly inhibit migration and invasion of TNBC cells. More importantly, the VEGF siRNA nanocomplex efficiently inhibit tumor growth in vivo and successfully downregulate VEGF expression in the tumor. These results suggested that VEGF siRNA is a promising anti-tumor agent for TNBC therapy, and the PEI 1800 conjugated bio-degradable multiblock polymer is a promising system to deliver siRNAs to TNBC cells. In Chapter 4, we developed a CD44-targeting, cholesterol modified cationic peptide nanocomplex to co-deliver IKBKE siRNA and cabazitaxel (CTX) for TNBC therapy. IKBKE siRNA significantly inhibits the proliferation, migration, and invasion of TNBC cells but has no apoptosis-inducing effect. In vivo studies also indicated that IKBKE siRNA can inhibit TNBC tumor growth. CD44-targeting CHA/CP/siRNA/CTX nanocomplex showed the synergistic effect of IKBKE siRNA and cabazitaxel on the inhibition of invasiveness and growth of TNBC tumors with an enhanced CD44 specific targeting effect. CHA/CP/siRNA/CTX nanocomplexes also exhibited a significant antitumor effect through IKBKE siRNA and cabazitaxel in vivo. Thus, IKBKE siRNA may be a promising anti-tumor agent for TNBC therapy, and co-delivery of IKBKE siRNA and cabazitaxel through a CD44-targeting nanocomplex is a potential strategy for TNBC treatment. Moreover, this multifunctional delivery system can also provide a good option for combined gene therapy and chemotherapy. In Chapter 5, we prepared three neutravidin-based siRNA nanocomplexes with different targeting ligands to deliver PCBP2 siRNA to hepatic stellate cells (HSCs). Insulin-like growth factor 2 receptor (IGF2R) is overexpressed in activated HSCs and therefore can be utilized for HSC-specific drug delivery. Compared to vitamin A and cholesterol, the IGF2R-specific peptide exhibited the highest targeting effect to human LX-2, rat HSC-T6 cell line, and activated primary rat HSCs. Accordingly, the IGF2Rspecific peptide coupled nanocomplex demonstrated higher silencing activity of PCBP2 and better inhibition on the migration of activated HSCs. The IGF2R-specific peptide coupled nanocomplex also showed the highest uptake in the liver and lowest uptake in the lung and kidney of the rats with CCl4-induced liver fibrosis.