The two defining themes of our laboratory is gene therapy and cancer immunotherapy. We intend to dwell into the basics and focus on the molecular mechanisms of viral gene delivery, with special reference on innate immune responses against viral vectors. In parallel, we work on both basic and preclinical aspects of genetically modified Natural Killer (NK) cells and NK cell-derived exosomes as a nanomedicine approach in cancer immunotherapy. More specifically, the aims of the research program in our laboratory are:
- To develop novel and more efficient approaches for the ex vivo genetic modification of human cells in order to facilitate practical and low-cost clinical trials of gene therapy.
- To investigate the feasibility and the potential efficiency genetically modified NK cells and NK cell-derived exosomes in the treatment of cancer.
GENE DELIVERY USING LENTIVIRAL VECTORS
The major hurdle in gene therapy is the delivery of the gene-of-interest into the cell. When it comes to hijacking cellular defense mechanisms and carrying genetic information into the cell, nature presents us with an evolutionarily perfected carrier: viruses. However, using viruses for gene delivery has problems of its own. In order to get rid of the pathogenicity of the virus and ensure that the virus will not be replicating and spreading, all viral genes and sequences except those necessary for packaging of the viral genome are removed during viral vector production.
As with wild-type viruses, intracellular recognition of viral components by pattern recognition receptors is a possible mechanism of cellular response against viral vectors. It is well known that toll-like receptors (TLRs) and RIG-I-like receptors (RLRs) play a major role in the detection of internalized viral particles and induction of an anti-viral state. Many wild-type viruses have developed elaborate schemes to avoid detection by these receptors and increase their virulence. In the case of viral vectors, the removal of various viral genes that counteract host responses but are dispensable for vector production is often preferred due to safety and practicality considerations. Inevitably, this would render viral vectors more prone to inducing strong innate responses upon target cell infection. It is possible that TLR or RLR mediated detection of viral vector components might activate an anti-viral response, negatively affecting the efficiency of viral gene delivery.
In order to test this hypothesis, we have previously attempted to use small molecule inhibitors of TLR and RLR signaling preceding lentiviral transduction, and observed a dramatic increase in gene delivery efficiency. Therefore, it might be possible to state that the lentiviral RNA is recognized by one or more of these receptors and an anti-viral response is triggered.
These results present a proof-of-principle for the feasibility of such approaches for enhancement of gene therapy applications. Overcoming the intracellular defense mechanisms against gene delivery vectors will lead to a dramatically increased delivery rate of the transgene. This may have broad technical applications in order to improve the efficiency of genetic modification of a wide variety of cell types. Because of the innate resistance of NK cells against viral infections, compared to other cell types, they are a good model to get detailed insights about cellular response mechanisms against viral vectors.
NK CELL-DERIVED EXOSOMES IN CANCER IMMUNOTHERAPY
In the light of recent findings demonstrating that human NK cells release exosomes, which can rapidly eliminate various tumor cell lines, we now know that exosomes released by activated NK cells express typical NK cell markers, such as CD56, as well as cytotoxic proteins, like perforin. Furthermore, the anti-tumor effect of NK cell-derived exosomes (NK-exosomes) opens up the possibility of adoptive transfer of ex vivo manufactured NK-exosomes, which still requires extensive investigation in vitro and in vivo. Exosomes from B, T, epithelial, and especially those derived from dendritic cells have been widely characterized. However, among the exosomes released by normal cells of the hematopoietic system, those secreted by NK cells have largely been ignored.
In view of the important role of NK cells as a major component in tumor immune surveillance, we investigate NK-exosomes as surrogates of this cell population. It appears conceivable that NK-exosomes may have a role in both paracrine and systemic control of the immune response and tumor surveillance. Thus, NK cell derived exosomes might have a future complementary role in cancer immunotherapy.