Metal Organic Frameworks (MOFs) as non-viral gene delivery vehicles for gene therapy in prostate cancer

<p>Prostate cancer (PC) is a leading cause of cancer-related mortality and morbidity in men. Despite recent progress, there is an urgent need to enhance therapeutic efficacy for PC patients. Gene therapy involves the alteration of genetic expression to treat diseases and has shown promising potential in preliminary clinical trials. It is widely considered to be a particularly suitable therapy for PC. Two powerful approaches to alter gene expression are the traditional RNA interference (RNAi) and the revolutionary clustered regularly interspaced short palindromic repeats (CRISPR/Cas9) mediated knockdown and gene editing. However, cellular delivery of therapeutic genes is still at an early stage and calls for additional technological breakthroughs to realise its full potential. Selection of ideal delivery vehicles remains one of the primary challenges for successful gene therapy that can be taken to clinics.</p> <p>Metal organic frameworks (MOFs) are an emerging group of porous materials with well-defined coordination chemistry. They represent a unique platform by virtue of their high porosity, encapsulation capacity and controllable surface functionalities. In nanobiotechnology, a major attraction of MOFs is their ability to incorporate biomolecules and drugs for cellular delivery. Biomolecules such as proteins are well studied, indeed almost routinely researched, for MOF- based delivery. However, each biomolecule is vastly and necessarily different. With respect to MOFs, nucleic acids are a niche where there is limited information. Despite the published studies available on DNA loaded MOFs, prior to this thesis, even the fundamentally vital aspect of DNA - its functional activity of genetic expression - remained completely unexplored within these complexes.</p> <p>The principal aim of this thesis is to establish MOF-based delivery of intact functional genes; efficient and targeted gene delivery with minimal/no cellular toxicity is investigated for the purposes of gene therapy in PC.</p> <p>Chapter 1 provides a thorough literary background that forms the basis of this research. The need for investigating gene therapy for a disease such as PC and the reasons for looking into novel materials like MOFs are explained.</p> <p>Chapter 2 evaluates the potential of the most common biocompatible MOFs - zinc- based Zeolitic Imidazolate Frameworks (ZIFs), zirconium-based Universitetet I Oslo (UiOs) and iron-based Materials of Institute Lavoisier (MILs) for gene delivery. Synthesis conditions and gene loading efficiency were the critical factors from the material selection perspective. Rapid in-situ synthesis under physiological conditions, successful gene loading and low concentration requirements favoured ZIF-8-based MOFs.</p> <p>Chapter 3 delves into the details of ZIF-based gene delivery. Encapsulation of a complete gene-set and cellular expression of the gene delivered by the MOF composites is elucidated. Using a non-specific plasmid as a proof-of-concept genetic macromolecule, successful transfection in PC cells is demonstrated. Gene expression occurs over relatively prolonged periods where the cargo nucleic acid is released gradually in order to maintain sustained expression. Most importantly, the encapsulated gene retains its functional integrity. </p> <p>After establishing gene delivery, Chapter 4 proceeds to investigate gene therapy and its essentially specific nature - the ability to cause chromosomal gene editing by targeting a specific faulty gene in the host cell nucleus. An endogenously dysregulated gene in PC called ribosomal protein SA (RPSA) is targeted using RNAi as well as CRISPR/Cas9 delivered via ZIF-based MOFs. A phytochemical from green tea is further used to enhance uptake of the delivery system. Most importantly, not just cytoplasmic, but host genome editing by the MOF-delivered genes are clearly established.</p> <p>The thesis is concluded in Chapter 5 with a thorough discussion on the aforementioned novel results discovered from Chapters 2-4. Limitations, further scope and future directions of MOF-based gene therapy are considered. This research thus establishes ZIF-8 polymorphs as a promising gene delivery system targeted towards PC therapy; a beneficial step towards advancement in knowledge in the fields of MOF technology, cancer research and gene therapy.</p>