Current Gene Therapy - Volume 14, Issue 4, 2014

Conventional therapies for malignant cancer such as chemotherapy and radiotherapy are associated with poor survival rates owing to the development of cellular resistance to cancer drugs and the lack of targetability, resulting in unwanted adverse effects on healthy cells and necessitating the lowering of therapeutic dose with consequential lower efficacy of the treatment. Gene therapy employing different types of viral and non-viral carriers to transport gene(s) of interest and facilitating production of the desirable therapeutic protein(s) has tremendous prospects in cancer treatments due to the high-level of specificity in therapeutic action of the expressed protein(s) with diminished off-target effects, although cancer cell-specific delivery of transgene(s) still poses some challenges to be addressed. Depending on the potential therapeutic target genes, cancer gene therapy could be categorized into tumor suppressor gene replacement therapy, immune gene therapy and enzyme- or prodrug-based therapy. This review would shed light on the current progress of delivery of potentially therapeutic genes into various cancer cells in vitro and animal models utilizing a variety of viral and non-viral vectors.

Poly(ethylenimine) (PEI) is a cationic polymer extensively exploited for non-viral gene delivery; however, its wide application has been impeded by its cytotoxicity. PEI can assume either a branched or linear configuration. Whereas branched PEI (bPEI) is more chemically reactive and can form smaller complexes with DNA under salt-containing conditions, lPEI is generally less toxic and exhibits higher transfection efficiency. In this study, we cross-linked low-molecularweight lPEI with methyl β-cyclodextrin (MβCD) to form MβCD-lPEI (MLP). The structure of MLP was successfully characterized by NMR, FT-IR, MALDI-TOF and elemental analysis. In the standard serum-free transfection environment, MLP could effectively transfect glioblastoma, melanoma and hepatocarcinoma cells. A high transfection efficiency was maintained in the presence of serum. Apart from its high transfection efficiency, MLP was found to have negligible cytotoxicity over a wide range of concentrations and to exhibit a low membrane disruptive capacity ex vivo. MLP warrants further development as a promising gene delivery system for future research.

Vitamin D is a steroid hormone that regulates mineral homeostasis, bone metabolism and many other physiological processes. The active metabolite of vitamin D, 1α, 25-dihydroxyvitamin D (1,25D3), has broad spectrum antitumor activities and potentiates the effects of a number of chemotherapeutic agents. 1,25D3 exerts its anti-tumor effects mainly through genomic mechanisms involving the regulation of gene transcription through vitamin D response elements (VDREs). More recently, miRNAs have been shown to be regulated by 1,25D3. miRNAs are short non-coding RNAs that post-transcriptionally modulate the expression of a wide range of genes. Therefore, they have important regulatory roles in the development and progression of many diseases including cancer. This review focuses on the regulation of miRNA expression by 1,25D3 in cancer model systems and the contribution of the regulated miRNAs to the anti-tumor effect of 1,25D3. In addition, the impact of miRNAs on 1,25D3 signaling is discussed.

Lentiviral vectors (LVs) represent suitable candidates to mediate gene therapy for muscular dystrophies as they infect dividing and non-dividing cells and integrate their genetic material into the host genome, thereby theoretically mediating longterm expression. We evaluated the ability of LVs where a GFP reporter gene was under the control of five different promoters, to transduce and mediate expression in myogenic and non-myogenic cells in vitro and in skeletal muscle fibres and stem (satellite) cells in vivo. We further analysed lentivirally-transduced satellite cell-derived myoblasts following their transplantation into dystrophic, immunodeficient mouse muscles. The spleen focus-forming virus promoter mediated the highest gene expression in all cell types; the CBX3-HNRPA2B1 ubiquitously-acting chromatin opening element (UCOE) promoter was also active in all cells, whereas the human desmin promoter in isolation or fused with UCOE had lower activity in non-muscle cells. Surprisingly, the human skeletal muscle actin promoter was also active in immune cells. The human desmin promoter mediated robust, persistent reporter gene expression in myogenic cells in vitro, and satellite cells and muscle fibres in vivo. The human desmin promoter combined with UCOE did not significantly increase transgene expression. Therefore, our data indicate that the desmin promoter is suitable for the development of therapeutic purposes.

Tumor angiogenesis involves multiple signaling pathways that provide potential therapeutic targets to inhibit tumor growth and metastasis. Regarding the significant role of vascular endothelial growth factor (VEGF) in angiogenesis and tumor progression, VEGF sequence-specific small interfering RNA (siRNA) for anti-angiogenic tumor therapy are under development. In the present study, dual-modified liposomes (At-Lp) was designed by attaching two receptorspecific peptides, Angiopep and tLyP-1, which specifically targeting low-density lipoprotein receptor (LRP) for brain tumor targeting and neuropilin-1 receptor (NRP-1) for tumor penetration, respectively. Gene transfection and silencing, and antitumor effect of the At-Lp loaded with VEGF siRNA were evaluated in vitro and in orthotopic xenograft models of U87 MG tumor. The At-Lp significantly enhanced cellular uptake (2-fold) and down-regulated expression of VEGF in U87 MG glioblastoma cells compared with non-modified and single-modified liposomes. The internalization of the At-Lp into tumor cells was taken via the enhanced permeability and retention effect and receptor-mediated endocytosis, followed by an effective endosomal escape of loaded siRNA into the cytoplasm. The At-Lp showed great superiority in inhibition of tumor growth, anti-angiogenesis, expression of VEGF and apoptosis effect after in vivo application against nude mice bearing U87 MG glioblastoma without activation of system-associated toxicity and the innate immune response. These results demonstrated that the combination of two receptor-specific peptides-mediated liposomes presented a promising platform for effective targeting delivery of siRNA for cancer anti-angiogenic therapy.

Peripheral artery disease (PAD) is a highly prevalent disease, which still has unmet medical needs. Therapeutic angiogenesis for PAD, achieved by gene therapy, has achieved promising results in preclinical studies and early-phase clinical trials, yet few late-phase clinical trials have been conducted or have not shown efficacy. This article provides an overview of the progression of angiogenesis research in gene therapy field as it applies to PAD. The focus of angiogenic growth factors and clinical trials is introduced as a frontier of therapeutic angiogenesis. The article also includes insights into future directions from bench to bedside.

Gene therapy continues to grow as an emerging treatment strategy toward numerous diseases. However, such prospects are hindered by the use of viral vectors prompting significant safety concerns along with limitations concerning repeat administrations, size of delivered gene construct, scale-up, high production costs, contamination during production, and lack of desired tissue selectivity. Non-viral gene delivery demonstrates the potential to address the abovementioned limitations, but itself generally suffers from low efficacy. Continuing efforts have been made to develop innovative delivery systems, synthetic gene carriers, and DNA vectors in a concerted attempt to enhance gene delivery suitable for clinical applications. In this review, we focus on the advances in the design of novel DNA vectors catered to enhance transfection and transgene expression and their influences on the efficacy and safety of existing and emerging delivery systems and synthetic vectors for non viral gene delivery.