Moving Gene Therapy Forward

Mobile DNA elements that can integrate into 'foreign' DNA Monday, 04 May 2009 Gene therapy Gene therapy is the introduction of genetic material into a patient's cells resulting in a cure or a therapeutic effect. In recent years, it has been shown that gene therapy is a promising technology to treat or even cure several fatal diseases for which there is no attractive alternative therapy. Gene therapy can be used for hereditary diseases, but also for other diseases that affect heart, brain and even for cancer. Indeed, recent results suggest that gene therapy can be beneficial for patients suffering from aggressive brain cancer that would otherwise be lethal. A safe delivery of the genes? Despite the overall progress, there is still a need to develop improved and safer approaches to deliver genes into cells. The success of gene therapy ultimately depends on these gene delivery vehicles or vectors. Most vectors have been derived from viruses that can be tailor-made to deliver therapeutic genes into the patients' cells. However, some of these viral vectors can induce side-effects, including cancer and inflammation. Marinee Chuah, Thierry VandenDriessche, Eyayu Belay and their fellow VIB researchers at K.U. Leuven in collaboration with Zsuzsanna Iszvak and Zoltan Ivics and colleagues at the Max Delbrück Center in Berlin (Germany) have now developed a new non-viral approach that overcomes some of the limitations associated with viral vectors. Lessons from evolution Using the principles of evolution and natural selection, that were initially conceived by Charles Darwin, they have now developed an efficient and safe gene delivery approach based on non-viral genetic elements, called transposons. Transposons are mobile DNA elements that can integrate into 'foreign' DNA via a 'cut-and-paste' mechanism. In a way they are natural gene delivery vehicles. The researchers constructed the transposons in such a way that they can carry the therapeutic gene into the target cell DNA. Doing so, they obviate the need to rely on viral vectors. “We show for the first time that it is now possible to efficiently deliver genes into stem cells, particularly those of the immune system, using non-viral gene delivery,” says Marinee Chuah. “Many groups have tried this for many years but without success. We are glad that we could now overcome this hurdle,” claims Thierry VandenDriessche. Zsuzsanna Izsvak and Zoltan Ivics concur: “This transposon technology may greatly simplify the way gene therapy is conducted, improve its overall safety and reduce the costs.” The VIB researchers are further testing this technology to treat specific diseases including cancer and genetic disorders, in anticipation of moving forward and treat patients suffering from these diseases. ......... ZenMaster

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What Regulates microRNAs?

When Cells Reach Out and Touch Monday, 04 May 2009 MicroRNAs are single-stranded snippets that, not long ago, were given short shrift as genetic junk. Now that studies have shown they regulate genes involved in normal functioning as well as diseases such as cancer, everyone wants to know: What regulates microRNAs? Scientists at Johns Hopkins were surprised to find an elegantly simple answer: touch. In a new study, published online April 9 in the Proceedings of the National Academy of Sciences, the researchers discovered that cell-to-cell contact revs up the manufacture of these small but mighty molecules. “This study documents one of the very few clear examples of a stimulus that directly influences the global efficiency of microRNA production,” says Josh Mendell, M.D., Ph.D., an assistant professor in the McKusick-Nathans Institute of Genetic Medicine, Johns Hopkins University School of Medicine. “No one anticipated, including us, that the production of microRNAs is linked to how densely cells are packed together.” In what Mendell describes as an “accidental discovery,” the team was studying contact inhibition: a phenomenon in which non-cancerous cells growing in a dish stop multiplying when they touch each other. Cancer cells, on the other hand, lose contact inhibition and continue to proliferate even when they are touching. The researchers suspected that microRNAs might play a role in contact inhibition because whenever they studied these enigmatic bits – only about 20 or so genetic building blocks comprise a microRNA – they always saw more in the tissues of animals, where cells are packed together, relative to the amount they found in isolated cells growing in culture. To investigate, the team grew cancer cells and non-cancer cells to increasing densities in culture and, using a tool developed in the Mendell laboratory, measured the abundance of hundreds of microRNAs simultaneously. This analysis revealed that the more densely the cells were packed together, the more microRNA was produced in each cell. The scientists then examined microRNA production in five additional commonly studied human and mouse cell lines, including human breast cancer cells, human colorectal cancer cells and human pancreatic cancer cells. They also tested fruit fly cells to determine whether the phenomenon is restricted to mammals. In all tested cell lines, including the fruit fly cells, scientists observed a dramatic increase in microRNA abundance with increasing cell density. “All evidence points to the fact that physical contact – when cells actually touch each other – is the critical factor that revs up the production of microRNAs,” Mendell says. “Through additional experiments, we were able to identify the specific molecular steps at which microRNA production is affected. We expect that this phenomenon will profoundly influence how cells behave in normal development and disease.” The team’s finding has practical importance for researchers who are investigating a range of biological processes that are most conveniently studied in cells growing in culture, Mendell says. “Little did we know the manufacture of microRNAs was so potently influenced simply by growing cells to different densities. We now recognize that this is a critical parameter that must be closely monitored when performing experiments with microRNAs in tissue culture.” A better understanding of how microRNA production is regulated is important because a reduction in the abundance of these molecules has been linked to the development of certain cancers. To date, one barrier to understanding how microRNAs are regulated in normal development and in disease states has been the lack of a simple system by which scientists could turn on and off a molecular pathway that controls microRNA production. Now, it seems, they may be able to toggle that pathway using cell-to-cell contact. “If we can identify the mechanisms through which microRNA production is regulated in normal settings, such as under conditions of extensive cell-cell contact, we can then ask whether the same mechanisms block microRNA manufacture in diseases such as cancer,” Mendell says. “This might allow the development of small molecules or other methods to turn microRNA production back on for therapeutic benefit.” Reference: Cell–cell contact globally activates microRNA biogenesis Hun-Way Hwang, Erik A. Wentzel, and Joshua T. Mendell PNAS 2009 106:7016-7021, doi:10.1073/pnas.0811523106 ......... ZenMaster

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Bones Made From Human Skin Connective Tissue

Cartilage, bones and blood vessels created from connective tissue Friday, 01 May 2009 Cartilage, bones and the internal walls of blood vessels can be created by using common connective tissue cells from human skin. Researchers in reconstructive plastic surgery at Linköping University, Sweden, have successfully manipulated these tissue cells to take on different shapes depending on the medium they have been cultivated in. “This means that it will be much easier to produce autologous tissue, which is tissue created from the patient’s own body”, says Gunnar Kratz, Professor of Experimental Plastic Surgery and team leader for the research group. The results of the group’s research are now published in three simultaneous scientific articles. Bone, cartilage and blood vessels are important components in reconstructive surgery, where damaged tissue needs to be recreated. Minor fractures can heal spontaneously but for major bone damage and cartilage injuries, there is the need to transplant tissue from other parts of the patient’s body. Different strategies have been attempted to instead grow autologous tissue from stem cells, for example those present in bone marrow. These cells, however, can be difficult to harvest, cultivate and store. Compared to these cells connective tissue cells from human skin has great advantages. A small biopsy is often enough to collect a sufficient amount of cells. “They are the ‘weed’ cells of the body, very easy to collect and cultivate into the cell type required. They are also very suitable to use to create a personal cell bank”, Gunnar Kratz says. Together with his colleagues, he has developed a technique to grow bone-, cartilage- and endothelial-like cells from connective tissue cells. Endothelial cells are the building blocks for the inner walls of blood vessels. The technique has been used to create whole tissue in gelatine scaffolds and currently preparations are underway to transplant these complete tissue pieces into laboratory animals. In the studies, the researchers collected connective tissue cells from healthy skin left over from breast and stomach plastic surgery and used fat stem cells to provide a comparison. To ensure that the transformation was not a result of the fusion of different cells, connective tissue cells from one cloned cell were also used. The cell cultures were cultivated in four different environments optimised for bone, cartilage, fat and endothelium. After two to four weeks, the connective tissue cells had produced cartilage and bone mass to a greater extent than the fat stem cells had. The cells showed several functions normally only present in the genuine cell type. They could also be used to build three-dimensional tissues, to create capillary networks etc. The studies are the first in the world where the results have been able to show connective tissue cells from human skin transformed into other so called phenotypes and creating other types of tissue. “The dream is to be able to manipulate connective tissue cells in the human body to develop into specific cell types, for example to create bone cells for broken bones”, says Gunnar Kratz. Reference: Engineering three-dimensional cartilage- and bone-like tissues using human dermal fibroblasts and macroporous gelatine microcarriers Pehr Sommar, Sofia Pettersson, Charlotte Ness, Hans Johnson, Gunnar Kratz and Johan P.E. Junker Journal of Plastic Reconstructive & Aesthetic Surgery, 2009; DOI: 10.1016/j.bjps.2009.02.072 ......... ZenMaster

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