Harnessing Endogenous Systems for Cancer Therapy

Abstract

In the recent decade, two strategies in particular have attracted attention due to the prospect of significantly improving cancer treatment: Gene silencing therapy and immunotherapy. Both strategies work by manipulating endogenous mechanisms and theoretically promise very strong effect on the diseased cells and minimal effect on the healthy ones. This thesis regards the investigation of important mechanistic aspects of gene silencing mediated by delivery of small interfering RNA (siRNA) using synthetic vectors (Project I) as well as the development of a delivery platform for targeted immunotherapy (Project II). Transfer into the clinic of therapies based on gene silencing by siRNA delivered by synthetic vectors has yet to happen. A major reason is the lack of efficiency in the delivery process, partly due to insufficient understanding of cellular uptake and processing of the siRNA-containing particles. Project I aims to provide new mechanistic understanding of intracellular processing and vector interaction with target cells by investigating siRNA delivery using branched polyethyleneimine (bPEI), which is a well-known synthetic vector for DNA delivery, and comparing the properties of bPEI with a lipid derivative thereof (DOPE-PEI). We demonstrate mechanistic differences between the bPEI conjugate and conventional bPEI with respect to siRNA condensation and intracellular processing and also show that lipid conjugation of bPEI results in markedly different formulation requirements compared to the conventional PEIs. However, lipid conjugation did not sufficiently reduce the inherent toxicity associated with high molecular weight PEI, and lipid conjugation of bPEI did also not change the ability of bPEI to affect lysosomal pH as a function of time. In contrast to gene silencing therapy, cancer immunotherapy is starting to produce positive results in the clinic. A major target in cancer immunotherapy is the immunosuppressive tumor microenvironment generated directly or indirectly by the tumor. Tumor tissues have been shown to be heavily infiltrated by macrophages and DCs but due to the immunosuppressive environment they frequently adopt an in-active or tumor-promoting phenotype. Project II describes the development of a platform which enables the highly specific targeting of monocytes and DCs in the bloodstream. Using this platform to deliver a TLR7 agonist, we were able to demonstrate activation of the targeted cells and increased potency of the agonist. While the described platform targets selected immune cells in the blood and not in itself targets the tumor tissue, we believe that because tumor associated inflammation has been shown to recruit monocytes and DCs to the tumor tissues, our strategy could be an elegant and efficient way of providing activated monocytes, monocyte-derived macrophages, and DCs to the tumor site. If the duration of cytokine secretory activity extends post-extravasation, this will not only provide activated innate immune cells to the tumor site, but may also contribute to the re-polarization of the tumor microenvironment thereby promoting antitumor immunity.