| 1 | The Royal Society of Victoria | Using Cellular Reprogramming & CRISPR Technologies to Regenerate the R... | 1057 | 38 | 3 | 67.7 | positive | 10:04 | Hello. Thank you for the opportunity to present my research. As you mentioned, I'm a PhD student from the University of Melbourne from the Center for Higher Research Australia. I'm also going to talk about the retina. The aim of a PhD is using cellular reprogramming to regenerate the retina and to treat brain loss. Let's begin. I'm also going to ask you a question. I want you to think about how would you feel if you had to face your everyday life with your vision looking like this or in complete blindness. Seems impossible, right? Unfortunately, this is what people living with photoreceptor generation have to face every day. Photoreceptors are cells that are key for our vision. They are located at the back of the retina and they are responsible for detecting the signals that are in the light and that our brain needs to form the images that we see as vision. That's why when we have diseases that damage the photoreceptors, it has a profound impact in the quality of our vision and it can often lead into blindness. We have two types of photoreceptors on rods that regulate the vision during night or in dark conditions and cones that regulate vision during daylight and the detection of color. I am particularly interested in cones because that is the type of vision that we use most of the time. In fact, that is the type of vision that we're using at the moment to look at all these amusing presentations, not amusing but entertaining presentations. As I said, when we have a disease that affects the photoreceptors, it can often well, it results in visual impairment and it can develop into blindness. Blindness is a huge or global problem because it creates a profound impact in the quality of life of a patient and it also has a huge burden on our societies because it creates problems like isolation, psychological problems, not to mention the burden, the financial burden that it places on our health system. The data of photoreceptors is a process that is irreversible and in most cases, there are no treatments to restore vision once the photoreceptors are dead. The current approaches to treat these diseases is just to help the patients manage the disease or just prepare them to live alive with visual impairment or blindness, which is not ideal. Popular examples of diseases caused by photoreceptor dead are H-related molecular degeneration or AMD, which affects almost 196 million of people in the world and retinaity speckmentosa that affects 1.5 million worldwide. Luckily, there is a light at the end of the tunnel and novel technologies like gene therapies are promising avenue to treat vision loss. In fact, there's a prominent example called Loxeterna, which is a gene therapy that uses viruses and that it has been approved in many parts of the world, including Australia. Loxeterna treats a very specific type of retinaity speckmentosa, so unfortunately, it is only applicable to a very small percentage of patients living with visual impairment. So that's why in my PhD, I want to develop a treatment that would be applicable for all patients living with visual impairment caused by photoreceptor dead. It is based in a technology called cellular reprogramming that was awarded the Nobel Prize in the 2012 and when I first heard about this technology, it was amazed because before we used to think that the identity of material cells was at a definitive state and it couldn't be changed. But now we know that we can take cells of a patient like, let's say, skin cells and reprogram them to become other tissues like neurons, heart muscle and even photoreceptors. But how do you reprogram a cell? The answer is in our DNA. So basically, all the information that we have that we need to make all the tissues of our body is already in our DNA and the difference is how the cells use this information. For instance, I am particularly interested in a cell type called Mule Glyia that is in the retina and I want to reprogram it into photoreceptors because they have more acute supportive role in the retina. They can also regenerate, so it doesn't matter if we lose some of them by transforming them into photoreceptors. And they have also been shown to have stem cell activity in other organisms. So going back to Mule Glyia, at DNA, they have unlocked in their DNA the genes that they require to make Mule Glyia. But for instance, the genes that they need that encode for the recipe to make photoreceptors are locked. But with another technology that we use in our laboratory called CRISPR activation, we are able to activate specific genes whenever we want. So with this, we can unlock the genes that encode the recipe for making photoreceptors and then we can promote these cells to become confoltoreceptors. So with this approach, I want to develop a treatment that would stimulate Mule Glyia into becoming new cones in a degenerated retina with a single injection with the aim of replenishing the lost photoreceptors and potentially treating vision loss. The main challenge of cellular reprogramming is knowing the genes that you need to make the cell that you want in this case photoreceptors. So I developed a tool in the laboratory that allows me to detect their reprogram Mule Glyia into cones because they turn red, as you can see here. I coupled this tool with a potent technology called CRISPR activation screen that basically allows me to search all the genes within the human DNA to predict possible candidates that could make photoreceptors. In my experiment, I detected 196 genes that could potentially make reprogram Mule Glyia into photoreceptors and I selected a few of them because of their importance in cone photoreceptor development. Next, I tested different combinations of these genes to try to find some that would be effective in reprogramming Mule Glyia into photoreceptors and overall I tested more than 100 combinations and I was indeed able to detect some combinations that could reprogram Mule Glyia into photoreceptors because they turn red as I explained before with the tool that I developed. We were also very happy to see that our new cone photoreceptors were functional because they could respond to light which is a key characteristic of the photoreceptors. Now that I knew which genes could potentially or I could use to make con photoreceptors, we proceeded to do some preclinical studies on a red model that has photoreceptor degeneration and visual impairment. So what we did was to put this cone reprogramming genes into viruses and we injected these viruses into the eye of the rats and four weeks after we analyzed the effect it had in our inhibition. So we were very happy to see that our treatment was able to induce or promote a functional improvement in these rats for parameters that measure photoreceptor activity or the activity of all those cells within the retina. Well this is very exciting because it means that we're getting closer to at some day developing a treatment that could improve visual function after you lose the photoreceptors. I would like to finish summarizing my talk because I know that there was a lot of information. So the aim of a PhD was to develop a cellular reprogramming protocol that could allow you to generate new human cone photoreceptors. And I also show you how the viral delivery of these cone reprogramming genes improved the visual function in a rodent model that has photoreceptor degeneration. So overall our results provide an important preclinical evidence for using cell reprogramming as a gene therapy to treat photoreceptor loss. I would like to thank the members of my laboratory, the cellular reprogramming unit and of course all our collaborators because without them this wouldn't be possible. Thank you. | ↗ |