Current Gene Therapy - Volume 16, Issue 6, 2016

Monoclonal antibodies (mAbs) based-therapies are currently one of the most successful strategies to treat immune disorders, cancer and infectious diseases. Vectors derived from adenoassociated virus (AAV) are very attractive to deliver the genes coding the mAbs because they allow long-term expression thus, reducing the number of administrations. They can also penetrate biological barriers such as the blood-brain-barrier to transduce cells localized in immunoprivileged organs. Recent animal studies with AAV have demonstrated the capacity of AAV to deliver sufficient quantity of antibodies to confer an efficient immunoprotection against chronic and infectious diseases for several months to years. The treatment was successfully applied either for prophylaxis or therapeutic use, depending on the disease and its progression. In this review, we discuss the advantages and the limitations of AAV for mAb and immunoadhesin delivery. Recent advances in vector design and antibody engineering are also presented. Optimization of the vector design can improve the kinetic and the level of mAbs expression whereas protein engineering can enhance transgene product properties. Furthermore, an exhaustive review of pre-clinical studies for chronic diseases including Alzheimer disease, amyotrophic lateral sclerosis and cancer is presented as well as for infectious diseases.

In the past decade, miRNAs have been extensively attracted the scientist’s attentions as tumor suppressors or oncogenes that have been implicated in tumor progression, metastasis and intrinsic resistance to various cancer therapies. microRNA-21 (miR-21) demonstrates a potential oncogenic function and targets tumor inhibitor proteins in almost all types of cancer. miR-21 overexpression has been studied in terms of cell proliferation, migration, invasion, metastasis, and apoptosis regulation. Inhibition of miRNA expression using antisense technology by various nanovectors of different sizes, shapes and compositions has been evolved progressively to overcome the barriers confronted by miRNA delivery. Application of miR-21 antisense oligonucleotides for treating cancerous cells has become a promising achievement for cancer therapy. Moreover, miR-21 can mediate resistance to radiation and chemotherapy. The expanding role of miR-21 functions in human cancers with an emphasis on its regulatory targets and mechanisms, miR-21 related achievements against cancer promotion have been discussed.

Gene transfer and oncolytic viruses provide new therapeutic approaches for the treatment of hematologic malignancies. However, it is still too early to introduce gene delivery or oncolytic viruses into standard clinical protocol. It is very important to discuss the obstacles that gene transfer and oncolytic virotherapy face for the further clinical application for the treatment of hematologic malignancies, and updating the advances made to overcome them. The major concerns in this review include the approaches of the development of immuno-stimulatory gene transfer mediated-vaccination for leukemia therapy, RNAi-based therapy for leukemia and enhancement of sensitivity of target malignant cells to virotherapy and alteration of host immune response to favor oncolytic viruses. We conclude with a perspective on the future of the gene therapy and virotherapy for the treatment of hematologic malignancies, emphasizing the problems we should solve and the technological requirements for further clinical applications.

Objective: This study was aimed to explore the potential of a non-invasive monocytes-based delivery system to transport therapeutic genes into the diseased brain. The study was conducted by first establishing the optimized conditions for lentiviral vector (LV)-mediated gene transfer into freshly isolated monocytes, followed by investigating the inflamed-brain homing efficiency and in vivo cell-mediated transgene expression by carrier monocytes in a mouse model with acute sub-regional neuroinflammation. Materials & Methods: Using a newly optimized spin-infection method, up to 35% of freshly isolated monocytes were successfully transduced with the LV system DHIV-101 at M.O.I. of 10. Meanwhile, cell trafficking and accumulation were detected in the inflamed brain regions in high density following intravenous (IV) administration of freshly isolated monocytes, confirming the suitability of using monocytes as cellular vehicles targeting the brain. However, LV transduced monocytes (TD-MO) displayed significantly reduced homing efficiency into the brain, possibly due to the initial unhealthy cellular states following LV transduction. Although the presence of transgene in the brain was confirmed by PCR, transgene expression was not detected within the inflammatory central nervous system (CNS) sites by RT-PCR or ELISA. Instead, high density of functional TD-MO and their transgene products were detected in the spleen. Conclusion: In conclusion, this study demonstrated that IV-infused monocytes were able to migrate into the brain, and remain viable and functional in vivo following LV transduction, warranting more indepth research to fully establish the conditions in order to reduce cellular toxicity induced by vector transduction, and thus enhance the brain homing efficiency of carrier monocytes.

Background: Gene therapy in mammalian cells requires vectors exhibiting long-term stability and high expression. Episomal gene expression vectors offer a safe and attractive alternative to those that integrate into the host cell genome. Materials & Methods: In the present study, we developed a new episomal vector based on the insulator, chicken hypersensitive site 4 (cHS4). The cHS4 element was artificially synthesized, cloned into the pEGFP-C1 vector, and used to transfect Chinese hamster ovary (CHO) and human Chang liver cells. The stably transfected cell colonies were further cultured in either the presence or absence of G418 selection. Fluorescence in situ hybridization (FISH) analysis and vector rescue experiments demonstrated that the vector replicated episomally in both CHO and human Chang liver cells. Compared with episomal vectors mediated by matrix attachment region sequences, the cHS4 element-containing vector yielded increased transgene expression levels, transfection efficiency, and stability during long-term culture. The vector was present at a very low copy number in the cells and was stably maintained over more than 100 generations without selection pressure. Conclusion: In conclusion, apart from a few free vector forms, the cHS4-containing vector mainly replicates episomally in mammalian cells and out- performs comparable systems in terms of yielding both higher expression levels and stability levels.

Background: Oncolytic viruses such as live-attenuated, vaccine strains of measles virus (MV) have recently emerged as promising cancer treatments, having shown significant antitumor activity against a large variety of human tumors. Objective: Our study aims at determining which parameters define the sensitivity of human melanoma cells to oncolytic MV infection. Methods: We analyzed both in vitro and in vivo the oncolytic activity of MV against a panel of human melanoma cell established in our laboratory. We tested whether either type I interferons or the interferon pathway inhibitor Ruxolitinib could modulate the sensitivity of these cells to oncolytic MV infection. Results: Human melanoma cells exhibit varying levels of sensitivity to MV infection in culture and as tumor xenografts. As these differences are not explained by their expression level of the CD46 receptor, we hypothesized that antiviral immune responses may be suppressed in certain cell resulting in their inability to control infection efficiently. By analyzing the type I IFN response, we found that resistant cells had a fully functional pathway that was activated upon MV infection. On the contrary, sensitive cell showed defects in this pathway. When pre-treated with IFN-α and IFN-β, all but one of the sensitive cell became resistant to MV. Cells resistant to MV were rendered sensitive to MV with Ruxolitinib. Conclusion: Type I interferon response is the main determinant for the sensitivity or resistance of melanoma to oncolytic MV infection. This will have to be taken into account for future clinical trials on oncolytic MV.