All things Embryo
Transcript from the interview with ASU School of Life Sciences Professor Tsafrir Mor.
Science Studio Podcast Vol 24
Transcript from the interview with ASU School of Life Sciences Professor Tsafrir Mor.
Science Studio Podcast Vol 24
[music]
Peggy Coulombe: Hi this is Peggy Couloumbe and welcome to Science Studio. Today we are going to talk about "pharming" but not in the usual sense of the word. This "pharming" is spelled with a "ph" like "phat" and links plant biology with medicine and biotechnology.
Our guest today is Tsafrir Mor, an assistant professor in the School of Life Sciences and a researcher at the Biodesign Institute at ASU. He studies the use of plants as platforms for the production of useful especially therapeutic polypeptides, proteins and enzymes and pursues solutions and antidotes to disease such as HIV/AIDS and bioterrorism.
Welcome Tsafrir.
Tsafrir Mor: Hey.
Peggy: So tell me, how did you start on this path to becoming a "pharmer"? Did you always know you would go into the sciences?
Tsafrir: I entered the sciences...probably yes. It was something that was always fascinating. As a child, I had a father who was pursuing a Ph.D. in plant physiology, so the plant direction was also probably sown at that age. But I did my Ph.D. actually I worked in the same lab also as an undergrad – on something very basic. I was working on something very basic, the dynamics of photo system two, working of something very basic in biology the inhibition of photosynthesis by high light was something that I, toward the end of my dissertation work, I became progressively disenchanted with and frustrated with how basic it is.
I didn't see the connection to humans and what the general world really needs. I was helping my advisor to write a grant proposal and we wrote, of course, when he justified that working on photosynthesis is something which is of course the main source of our food and I thought, "This is so far fetched from what we are actually doing in the lab. That's not the way I want to go."
It so happened that a few years before I joined the lab, my advisor took a sabbatical in Michigan in the lab of Charlie Arntzen.
Peggy: Charlie Arntzen is now at the School of Life Sciences and a Regent’s Professor and also a member of the Biodesign Institutes. Oh, but go on.
Tsafrir: They wrote several seminal papers on photoinhibition, so I knew of Charlie Arntzen as a great scientists working on photosynthesis, on photoinhibition, and we heard rumors that he's bailing out, that he's switching, that he's moving to something completely different vaccines in bananas. Everyone in the lab was joking, "He's gone bananas."
Peggy: [laughs]
Tsafrir: I was actually thinking, "Why not?" I followed the literature as it unfolded from his lab. When the time was right to look for a place to do a post doc, I contacted him and there I was, in Ithaca, New York, working on something completely different.
Peggy: So your interest in genetics and genetic engineering came at that time?
Tsafrir: Yes. Even during my work on photoinhibition, we were designing and creating mutants that would enable us to study the process, so I value molecular biology and genetic engineering as something also contributing to basic science, but most certainly the use of such techniques in order to create something very useful was something that I started to work on when I got to Ithaca.
Peggy: So it was mostly interest in science and the interest...
Tsafrir: ...To human welfare. As time goes by, I am starting to look back that that's not necessarily my distaste for basic science. It’s something that I grew out of, so to speak, so the pendulum shifted from me just wanting to be a scientist of discovery, working on something very basic...full stop, moving to something very applied, has now probably shifted back and centers on what I feel now, as someone a little older and a little bit wiser perhaps, something which is cross fertilizing. Working on basic science and working on applied projects seems to be the right way to go.
Peggy: How did you choose to focus on protein engineering in plants?
Tsafrir: The idea was that plants and at that time, during the early 1990s, the idea of using transgenic animals was something that was only starting to come about. But as plant biologists, we were ahead of that flock and we could produce a proteins in plants pretty much like our microbiologist friends have been doing for decades before, and we could produce a whole range of proteins that those microbiologists and bacteriologists in particularly could not even dream of, because plants are eukaryotic organisms. They have a lot of the machinery that is present mammalian cells. Actually, they have all of that. There have slight nuances and small differences, but pretty much it's very closely related in many ways.
So you could produce many different types of proteins in plants that you could not do otherwise. Of course, plants as we all know are relatively inexpensive to produce. All you need basically are sun, water, and fertilized soil, and they'll grow making a lot of sense as a production system, especially if you are intending some of its application to go to poor countries in the third world, where a lot of the need exists but very little of the resources to fulfill this need exist.
Peggy: So what kinds of plants do you use, and why those in particular?
Tsafrir: That again went through an interesting evolution. As I said earlier, the vaccine bananas of Charlie became really synonymous with this whole approach of molecular pharming. It's a very photogenic fruit and the pictures of Charlie smiling, holding a banana, were on daily newspapers.
Peggy: [laughs]
Tsafrir: However, there were several issues with banana per se. Banana is a plant which is very difficult to transform it takes about two years to create the transgenic plant, which then, of course, has to be screened and you have to look for a good expressive line and so on and so forth, and that definitely doesn't want to work on this type of organism as a prototype.
Another development that we experienced and we really experienced it in a very straightforward way – was that we were all basically plant biologists and we knew very little of immunology and we knew very little of the different forces that work together to bring a vaccine from an idea to an actual drug in the market and we faced severe criticism from a lot of scientists and regulators because we could not control the exact dosage of the intended vaccine in the plant.
So it also became clear that we could work on food plants, on crop plants, as much as we like but, in the end, the product would have to be delivered as a drug to the patient rather than as a commodity as a consumer, and we gradually switched back from working on very exotic plants like bananas to less exotic plants like tomatoes and potatoes and other crop plants to the good old vehicle host plant, which was tobacco.
Tobacco is a plant with ill repute; however, it’s extremely easy to transform, highly productive, and if you are looking at the plant as a production system, for which protein inside the plant you'll have to purify, then it doesn't really matter if it’s from a crop plant or not.
There are actually several advantages as well. Many of you are probably familiar with Protogen debacle a few years ago. Protogen was a small company in Texas, based in College Station that was also working on the heels of Charlie Arntzen, producing vaccines in plants.
They've chosen maize – corn as their vehicle, as their host plant, and there's no better place to grow corn in the U.S., probably in the world, than in Iowa. They had a small experimental field there, and they had actually taken a lot of precautions like the USDA requires. They had a buffer zone of soybean, and the nearest maize was so many miles away. I don't remember the specifics.
But the one thing that they did not come and do was to come in the next season and pull out the rogue plants that are coming up from seeds that were shed during the process of picking up the corn. The whole crop of soybeans of the next year that were grown in the same experimental field were contaminated with very, very little, but enough, of that Protogen vaccine product, which created of course a huge thing in the media which led to the fall of that small company.
So that whole story comes in order to teach us a lesson that, maybe using a non food plant for production of pharmaceuticals is actually a desirable thing and the other thing which comes out of it is that maybe Iowa in the center, the epicenter, of agriculture is not the best place to grow such plants and maybe a place like Arizona which has a certain natural types of containment.
Peggy: The extreme heat and no water...[laughs]
Tsafrir: Extreme heat and if you chose your places of growing the plants smartly, it is relatively away from any other type of farming. So both of these thinkings: avoiding food crop plants and choosing the place where you grow them - potentially even using green houses - although we are still hoping that we will be able to grow things in the open, because it is a lot less expensive.
Peggy: We are going to get back to some of the concerns about genetic engineering and transgenic plants a little later in the program, but I had a question. You said that some plants are more easily transformed. Can you tell me a little about that process and what makes a plant more acceptable or less acceptable?
Tsafrir: There are several ways to shove genes into the genomes of the plants. One of them is really that. It is the brute force method called the Gene Gun method or Biolistics, in which you are "shooting" small particles of gold or tungsten particles into plants cells and those particles will penetrate the different barriers of cell walls, the plasma membrane, the nuclear envelope and some of it will land in the nucleus where, if those particles were coated with appropriate DNA, they would be able to get incorporated.
This is the brute force, it usually results in many copies of the trans gene and that's associated with several problems that you might want to talk about later. In a problem called “gene silencing” plant perceive these trans genes and their products as foreign and they work very hard to eliminate their expression. So that's one method, the biolistic method.
By far the most common way for us, agrobacterium, it's a small soil bacteria universally found everywhere; it's responsible for creating a plant disease called Crown Gall disease. Those bacteria are very "smart." They have a way to introduce part of their DNA, part of their DNA which resides not in the genome of the bacteria, but in plasmids inside the bacteria. So they have a way to move part of that DNA into the plant cell and into the nucleus where it ‘s integrated.
In that piece of DNA that they are moving in, you have several things that are beneficial for the bacteria. You have genes that are producing plant hormones that promote growth of the plant cells and thereby creating the gall formation, this almost tumor-like structure on the plant. But more importantly, they are able to synthesize, using some of the genes that are in that transfer DNA, the DNA which moves into the nucleus. They have genes for the synthesis of specialized amino acids that are used as metabolites by the bacteria, but the plants don't know how and what to do with it. So basically it's hijacking the carbon metabolism of the plant for its own sake.
We are eliminating these two elements, the hormone producing genes as well as the genes responsible for the synthesis of these opines. We are placing them with our gene of interest. The bacteria still would mediate the transfer of this piece of DNA and the integration into the plant genome, but of course with the gene of interest, our favorite gene.
Then you can get rid of the bacteria by applying antibiotics and you select for the presence of your gene and it's pretty straight forward. Also we rely on the fact that plants are amazing in their ability to regenerate by new limbs, so to speak, with a little bit of helping with plant hormones one can expedite that.
So it's relatively straightforward to do. However, not all plants are as susceptible to agrobacterium as tobacco, which was the first plant to be transformed. And other members of the solinacious family: tomatoes, potatoes, and petunia. These are all very easy to transform plants. Some other plants are a little bit more difficult in the sense that you have to devise special ways to A. co-cultivate the bacteria together with explants. As well as to regenerate whole plants, which is another kind of art.
So at this point we don't know necessarily why some plants are more amenable for these types of treatment than others.
Another limitation, which is also is encountered in many ways also in the realm of transgenic animals, is the fact that we cannot control, so far, the place where we are inserting our gene, which means that the gene can be inserted in areas of the genome which are not very productive as far as expression of genes.
It could fall within a very important gene for the plant, thereby knocking it out, creating a mutant which, of course, which would not be very useful. So you have to select among the different plants not only for high level expresser, a plant that is expressing a lot of your foreign gene but also that is robust as far as its agronomic properties are. We have all these issues that we have to tackle.
Peggy: So let me go back to another issue that we touched on. In a lot of European countries, they have struggled with the concept of genetically modified crops, particularly for human and animal consumption, and humans have shifted qualities of plants through selective breeding for generations. What are the specific concerns around transgenic systems? You mentioned a little bit about the field in Iowa and the lack of containment and the lack of, I guess, essentially cleansing the field after the first generation of crops.
Tsafrir: Right. The concern there is self-imposed rule, essentially. The USDA has a regulation that dictates how those transgenic plants are being grown in the field. And what are the containment issues and although we know of no particular health concern that would come from these types of genetically modified plants, we still are taking these precautions of not mixing them with food stuff.
So, the concern there was that this particular company did not follow the rules. They did not do what they were supposed to be doing and the problem was not that people were at risk but that following the rule was at risk and was actually breached.
What are the concerns? The concerns are multiple and in many cases they are not really science based, but that doesn't mean they are not real, or in the minds of the people that raise them. One concern is that the selectable marker that we are using for generation of these plants - so we have to make sure that we have transgenic plants so we are using an extra gene that we are introducing which confers herbicide resistance or antibiotics resistance- so some people have concerns that this might get into the environment; therefore, get into either close relatives or weed relatives of plants or get into bacteria. I think that chances there are not extremely high, but there are already people that are developing ways to create transgenic plants and select for them without the need for selectable marker or with a selectable marker that then can be excised. That's something which is being addressed as part of the science.
Other issues have to do with the supposed allergenicity of some of these plant derived antigens. Again that is something which is very hard to predict and very hard to control and the risk is again very minimal. People have food allergies even without the help of protein engineering and transgenic organisms.
From a use of plants for a platform for the production of pharmaceuticals, one has to again remember that these are not going to be available in your local supermarkets as a vaccine de jour kind of crop. So you are not going into the supermarket and say, do I want to take the Hepatitis B bananas or the Norwalk virus tomatoes?
It's going to be a pharmaceutical and it is going to be delivered by or handed to people by medically trained personnel. So in that respect there is again little issue. I think Europeans are coming around. The initial fears are kind of more subdued and we are becoming more careful, as well.
We are not trying to shove our technology into the faces, or literally into the throats of people who don't want them, but we are doing a lot of fieldwork as far as education. Eventually if there will be a vaccine that will be able to help mitigate the HIV epidemic and be available inexpensively to people in Africa, I think people would have little objection to this.
Another development or the major player at this point is a company in Europe is to avoid charging for these plants all together and use what they’ve coined “deconstructed plant viruses” as the agent of engineering. So, we use wild-type plants. We engineer plant viruses to over express a certain protein. All of this is done under containment in green houses or in growth chambers. There are no transgenic plants involved and the viruses are very smartly engineered, so that they are not able to propagate by themselves. So it is not something which would have environmental consequences.
Peggy: One line of research at your laboratory pursues, this is a quote from your website: "The evaluation of transgenic plants as a source for recombinant human acetylcholinesterase to provide protection from non conventional warfare agents, is chemical, organophosphate intoxication and biological, as in immunomodulator." First, tell me something about how nerve agents typically affect the nervous system.
Tsafrir: So we all know that the nerve impulses travel down protrusions in nerve cells, the axons and the dendrites, as an electric pulse, creating depolarization on the membrane. However, in the place where two different nerve cells meet, or a nerve cell and a target cell, for example a muscle cell meet, there is a small gap called synapse and in the synapse the neurotransmitters have to be secreted out of the cell, travel the gap, bind to receptors and initiate a whole cascade of responses. If it's another nerve cell usually initiation of nerve impulse in the next cell. In many neurons the neurotransmitter is acetylcholine, which obviously binds to the acetylcholine receptor. In order for the nerve cells in the nerve system to insure and all or none kind of response, the signal has to be caught as soon as it has being formed. That is, acetylcholine is released into the synapse, binds to the receptors and soon after it should be eliminated and the whole system should be ready for another impulse.
The elimination of the neurotransmitter, acetylcholine, is the aim of a very unique enzyme called acetylcholinesterase. It hydrolyzes acetylcholine to acetate and choline and it is one of the fastest enzymes that we know of. Its activity is in the realm of diffusion. So the speed that this enzyme is catalyzing the reaction is limited only by the ability of the substrate to diffuse into the active site.
The fact that such an important enzyme meant that many organisms are using this as a way to control these reactions. If they are plants that would like to eliminate herbivores for example, many insects eat them, they would produce anti cholinesterase in their plant tissues that the insects would digest and therefore die. Similarly, other animals might use it as part of their predation strategy. So certain snakes have got anti cholinesterase in their venom.
All of these naturally occurring anti cholinesterases are very potent, but they are not the most feared at this point. How many of us actually encounter mambas frequently? Probably not...
Peggy: [laughs]...not very often.
Tsafrir: The fact that nature can do it also means that people can do it. During the early ‘30s people were investigating a new group of newly synthesized chemicals called organophosphates. As anti cholinesterase occurs in pesticides. The thrust of this effort was in Germany, which during the ‘30s was controlled of course by the Nazis, which had a very broad definition of "pests" at that time.
So although they were originally developing it, organophosphates, as pesticides, their extreme toxicity toward mammals meant that they could be very useful "as ways to eliminate human beings." By the end of the war the stockpiles of Nazi Germany had tons of the three, what we call, classical nerve agents or more popularly called, nerve gases. None of them are actually gases; they all are liquids at room temperature which are aerosolized and used as vapors, essentially.
Peggy: My grandfather got gassed in France during World War I.
Tsafrir: World War I, so World War I the gas that was mostly used was different types of mustard gas. However, yes, that's exactly what intended use was. We have no evidence that these stockpiles were ever used in combat but the fact was that the German military was stockpiling meant that they full intentions of doing it.
And soon after the technology spread to the Allied forces that adopted it very readily, both the American side, the NATO side, as well as the Russians and the Warsaw Pact side, were very active in producing new and more lethal types of organophosphates. We had the VX, the Russians had their own isomer of that called Russian VX, and cycle-sarin and other types of organophosphates were added to the mix.
It's a relatively simple chemistry that’s involved in making those chemicals. Basically anyone with a chemical set can produce those and we actually did find that they were used in some of the not as developed countries as we think. Iraq had impressive stockpile that they have used against the Kurds during the late ‘80s, when Saddam Hussein was still a major ally of the U.S. The end result was thousands and thousands of dead Kurds.
So that was one recent incident more close in time and also may be more close in scope of the kinds of problems that we might be facing today, is the use is sarin gas in the Toyko subway and a few other incidences in Japan during the 1990s by religious terrorist groups.
We know that Al-Qaeda for example is experimenting with sarin gas. There are pictures of using dogs as experimental animals to show they are able to do it. Has to be said that during the first Gulf War, and immediately afterwards, most of the stockpiles of Iraqi nerves agents were discarded.
Some people say that the plumes, from these cases of discards when those plants were bombed, actually released small amounts of sarin that by itself is not toxic, however, or at least not lethal, but is responsible for what became known as the Gulf War Syndrome, together with use of prophylactic against nerve agents called pyridostigmine.
Peggy: How does the military for example, protect itself from such agents?
Tsafrir: Obviously there are the physical barriers, the gas masks, wearing special types of uniform of clothes. However, there are also some medical treatments. So following the exposure, one can use a blocker of the acetylcholine receptor to prevent the over excitation.
Peggy: Let's back up a bit. What happens when the nerve agent blocks the enzyme?
Tsafrir: What happens when the nerve agent blocks the enzyme - of course acetylcholine cannot be removed from the synapse and every new impulse coming to the synapse releases more and more of it. You end up having what's known as a cholinergic crisis. You have flooding of the synapse with acetylcholine, which obviously binds to the receptor and constantly activates the post synaptic cells, if it's a muscle cell, creates the tonic convulsion, or if it's a nerve cell in the brain it leads to seizures. All of these things are seen in acutely poisoned individuals and animals.
Peggy: What are some of the ways that a protective agent might stop the whole process?
Tsafrir: One way of preventing it is to block the receptors. If you are blocking the receptor, even if you have a lot of acetylcholine in the synapse it will still not be able to activate the downstream processes. So that solves some of the issues. It doesn't solve the issues of permanent brain damage, for example, that might occur. It doesn't also prevent the incapacitance. These people that experience nerve agent, and therefore are being treated with this blocker called atropine, are very sick. They have to be evacuated to a hospital, and so on and so forth.
Another option that was explored very heavily during the first Gulf War was to give a chemical called pyridostigmine That's another type of an acetylcholinesterase inhibitor and that was always puzzling to me as a student when I studying this. How do you treat as a prophylactic agent, how do you treat nerve agent toxicity with another nerve agent? The trick that people that developed this were saying that that's a reversible blocker of the enzyme and it binds to the active sites and then is removed, unlike the organophosphates which binds in the active site and prevents it forever from being active again.
But there are other secrets here. The major secret here is the fact that pyridostigmine, like other anti cholinesterases, starts a whole feed back response within the cell to produce more acetylcholinesterase. That's understandable. You have a problem with an enzyme; it cannot perform its work and make more of the enzyme trying to overcome the block.
That's beneficial short term, however, this shooting up of the expression is something which is long term and would have long term consequences to the individual. Again, pyridostigmine together with an inadvertent expose to the sarin gas during some of the cases of bombing the chemical plants in Iraq during the first Gulf War that are blamed for the Gulf War Syndrome.
Peggy: How does your system uniquely promote solutions to counteract the effects of chemical and biological bio warfare agents?
Tsafrir: So what we are trying to do is, is to augment the natural defenses that we have against the nerve agent. That is, enzymes that are very similar to acetylcholinesterase or in fact another iso form of the enzyme which are circulating in our blood, the nerve agents in order to get to the synapses of the neuro muscular junctions or the brain, they have to go through the blood, and in the circulation then encounter those circulating cholinesterases that work as decoys. They inactivate, they bind and sequester these nerve agents. We can boost up, give an animal or human being, extra amounts of these enzymes and they would be in the blood and whenever there is exposure, they would take care of that excess inhibitor. The problem is that because the enzyme binds to the inhibitor and gets out of the game, it's a one to one kind of ratio. In order for us to be effective we need to employ large amounts of protein, which means we need to find a place to produce high amounts of this protein and that's where the plant production comes in.
So far, the only source for a similar enzyme called buterol cholinesterase was from outdated plasma, outdated blood samples. Which, if you combine all of the donated blood in the U.S., would give about 10 kilos of this enzyme which are not nearly enough for what one would need for the military as well as first responder needs. And of course, that's an exorbitant amount of money that is involved to purchasing all the blood bank samples in the U.S.
Peggy: What kind of success have you had producing this protein in plants?
Tsafrir: We produce several iso forms of this enzyme in plants. We have been very successful in showing that it is effective, and we are currently in collaboration with the USAMRICD [US Army Medical Research Institute of Chemical Defense], again by funding by NIH, we are working with the second generation of these agents, trying to create an enzyme that would cleave the nerve agents rather than sequester. So we'll need smaller amounts of this enzyme so people will not have to be injected with tons, well, figuratively speaking, with a lot of these enzymes.
Peggy: Can you tell me what kinds of discoveries have come about unexpectedly with your system?
Tsafrir: We talked quite a bit on the chemical side, but through my collaborations, and I have been lucky to have really good collaborators on the different projects that I am working on, we have been also exploring the possibilities of using the acetylcholinesterase to things that were not originally envisioned.
So this very interesting enzyme, apart from its distinct and well studied role in termination of signals in cholinergic synapse, also sits in a very interesting juxtaposition to control the nerve system, as well as other systems. And in fact one particular form of this enzyme which is produced under stress, able to control the immune system, and we are looking at ways of using the enzyme as well as peptides that are derived from it, as ways to control the innate immune system, which is the generalized first responders within the human body. First response against pathogens.
Incidentally, this enzyme is also very important in formation of plaques in the Alzheimer's patient brain. Just today, I was told by my collaborator that our paper was published in "Brain" to show that enzyme can disrupt the plaque formation. That's an in vitro study, but we are already initiating in vivo studies in mouse models of Alzheimer's disease.
So you see how an idea that started from and was backed by the good financing of the US military, actually has unexpected benefits, something which everyone face or will face in unfortunately their own future or the future of loved ones. So many people are affected by neurodegenerative disease like Alzheimer's disease and we may have the ability to contribute in this aspect as well.
Peggy: There's another line of research that you are also taking in your laboratory with support with the National Institutes of Health and it's the development of plant derived vaccines against sexually transmitted diseases with special focus on HIV/AIDS. Everyone knows how desperately we need an effective and affordable vaccine globally for this pandemic. So tell me something about your work and the utility of using the plant derived vaccine.
Tsafrir: Again, a bit of history about our involvement in this. So when Charlie Arntzen decided that he would like to create plant derived vaccines, he chose several disease models to work on. One of the ideas was to work on disease for which vaccine already is marketed, a subunit vaccine, a vaccine which is based from a protein produced in yeast in this case. So if we can produce the same type of protein in plants and show that it works just as well as the yeast vaccine, then that would be very easily accepted and the path for rich and glories is ”cake.”
Peggy: [laughs]
But that doesn't work that way because if you have a very good vaccine against Hepatitis B produced in yeast, who would want to try and back studies for developing something that would compete with this? So although people are still working on it, even here at ASU, notably Hugh Mason.
Peggy: I'd like to interject that Hugh Mason is an associate professor in the School of Life Sciences and also a member of the Center for Infectious Diseases and Vaccinology at the Biodesign Institute. Please continue.
Tsafrir: Together with Charlie, Hugh Mason is the pioneer in plant-derived vaccines. It's been relatively slow to progress for a lot of reasons, but one of them is what I have just said.
Another approach is to take a vaccine against a enteric disease, so it's a disease of the human intestine and so, therefore, the proteins and pathogens that are involved in it were evolved to survive the passage through the gut and so on and so forth. So that pathway to finding Norwalk-like vaccine which again Hugh Mason is proceeding.
In my early project was to try to tackle rotavirus which is a very important viral gastroenteritis-causing agent. But in all of this work we have very openly refused to work on HIV because it is such a complex type of disease that it was very clear that we’ll face not only challenges in bringing in a new technology, but also challenges, of course, to actually prove that we can make a vaccine, let alone plant derived vaccine. So Charlie wanted to tackle only a certain front and not try and beat everyone on their own turf.
But that's not what I had in mind. Because it was pretty boring to work, just to do another thing. Soon, I was actually in the first year I was here at ASU, we were contacted by a lab in France, headed by Morgane Bomsel and she sent us a draft for a grant proposal that she wrote that failed. In which it describes her approach and I was completely awe struck because it looked to me so straightforward and so obvious, that someone should have already thought about it and done it.
So I have done it a little bit of research around this and why would that work and why people avoid it. And what became clear was that people avoid it simply because they have their minds on completely different areas. In that it is something that is worth a shot.
What she's trying to tackle is the very early stages of virus penetration into the human body, which is in most cases, except for a very few cases of intra venal blood exposure, which happens with street drug use, and so on and so forth. But this is contributed globally to only a minute percentage of the incidents of the disease. In most cases it is sexually transmitted and the virus has to cross the mucosal barrier either in the vagina or in the lower intestinal, rectal and colon mucosa.
They are looking at one of the ways the virus is crossing the epithelium is via a process called transcytosis. That is kind of a shuttle mechanism: the virus binds to the outside of the cell, then receptor mediated endocytosis results in vesicle formation, the virus is engulfed in this vesicle. It does not infect the cell, the epithelial cells that it is crossing, but shuttled through those vesicles to the other side of the cell where it is being released by exocytosis into the blood stream and eventually find it's target CD4+ cells, the T cells which are the natural hosts for this virus and that's where infection really starts and chronic infection starts.
So we are looking at this very early step in the viral dissemination or entry into the body. And our strategy focus on trying to block this process of transcytosis and we have used the part of the envelope protein which Morgane Bomsel identified as responsible for this process. And we have effectively vaccinated animals; both mice and rabbits at this point, against it and the antibodies that these animals create are very effective in blocking the process.
So now we are continuing in several fronts on this project. We have collaboration with a group in Japan that have created a mouse model for HIV and they are vaccinating their animal models, their mice, and they will - after vaccination is complete - they will also challenge and see how effective we were in preventing the process of transmission.
We are also pursuing second generation vaccine candidates against, again, this area in the viral envelope, but by using other gadgets, other protein engineering gadgets and we are working together with Petra Fromme in the chemistry department. Petra Fromme is the preeminent structural biologist working on membrane proteins. In the world, one of the very few people that are very successful in this. She is trying to help us elucidate the structure of this part of the viral envelope which is membrane associated, membrane anchored. Hopefully by learning about the structure, we will be able to design better vaccine candidates.
Peggy: That is very exciting. So the next step would, after your animal models, would then be clinical trials?
Yes. The problem with HIV is that there isn't a very good animal model, so you cannot really test it in animals before you try it in people. People have created, like our Japanese friends, various models. One of the more accepted models is non human primate macaque model which is using a different virus. It is kind of a cross between the simian immunodeficiency virus and the human immunodeficiency virus, called SHIV expanded as Simian Human Immunodeficiency Virus.
But as far as the previous experience shows, it's not a very good predictor, and some people advocate going to clinical trials. Of course, clinical trials are a big step in money. We were hoping to be able to get our vaccine candidate into clinical trials as part of our NIH program. Right now, but at this point it looks like we're still a couple of years away from that.
Peggy: So where do you hope your research takes you in the future?
Tsafrir: Well, we have several projects that on ongoing right now and again from someone that started in plant biology, and now conducting animal experiments in the lab, starting the effects of various protein drugs and the physiology of the animals, I see that we are developing into these areas where we are not just is going to be the producer of proteins in plants, but we are actually going to study the effects of these drugs together with our collaborators.
Another aspect of our acetylcholinesterase research that I am very excited about is our realization that plants harbor analogous type of enzyme or enzymes and it is extremely curious first of all, what are these enzymes? Are they homologous to the mammalian, to the animal, system? And granted that plants don't have nerve systems, what do these enzymes do in plants? So again, that’s a basic type of research that is going on in my lab which I am happy about because it keeps things interesting and exciting, even if there is no immediate application in sight.
Peggy: Well, Tsafrir, I want to thank you for taking time to sit with me today and wish you the best of luck in your research. It is very exciting.
Tsafrir: Thank you.
Peggy: This is Peggy Coulombe and you have been listening to School of Life Sciences podcast, “Science Studio.” Our theme music comes from the website Magnatunes and was composed by Yongen from the collection, "Moonrise." The School of Life Sciences is in the College of Liberal Arts and Sciences on the Tempe campus of the Arizona State University.
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