Brian Wadzinski, PhD, Associate Professor of Pharmacology at Vanderbilt University talks about how alpaca antibodies are leading to new research.
Interview conducted by Ivanhoe Broadcast News in October 2019.
Tell me, why study alpacas?
WADZINSKI: Starting with my research, I work in the area of cell signaling, which is basically the process whereby an external signal is converted into an appropriate cellular response. We work with a family of enzymes known as protein phosphatases; I have worked on them for almost 30 years now. And one of the things that’s very important for these studies is to develop specific antibodies recognizing the phosphatases – the antibodies been very useful to increase our understanding of the function and regulation of these enzymes. I’ve worked with a number of different companies in the past to try to develop antibodies for these proteins, and it has been hit or miss. Roughly three years ago, I was at a point in my career where I was having a little bit of difficulty getting funding for my research and I had to think outside the box. That kind of forced me to step outside a little bit and explore some of the things that have worked for me over the course my career. As I mentioned, antibodies have proven to be very useful to my research, so I started exploring the idea of starting my own company where I could have access to animals to generate antibodies on my own. I was fortunate enough to collaborate with Ben Spiller – a Vanderbilt colleague. Together we looked into alpacas as a host for generating antibodies. Turns out that alpacas and other members of the camel family – old world camels and llamas – have a very unique form of antibody that has a lot of different applications. One can use them for research, diagnostics, and more recently for therapeutic purposes.
Are you harvesting the antibodies within the alpaca? Or are you taking their blood samples and creating antibodies?
WADZINSKI: So essentially what we do is we’re studying a molecule of interest. Mine is protein serine threonine phosphatase 2A – that’s the protein I’ve studied my whole career. We’re taking a fragment of that protein or the protein itself and immunizing the animals, basically injecting a small amount of that protein into an animal – like a flu shot – and doing that every two weeks for five to seven times. Then we have the vet come out to collect a little bit of blood from the animal. We’re not hurting the animals. We test that blood for the presence of antibodies recognizing our protein of interest. If that’s positive, we can then go on using a molecular biology approach to isolate a specific fragment of an antibody found in alpacas. These antibody fragments, also called nanobodies, can be used for a wide variety of applications in the laboratory.
And when you inject them with this protein, is it hit or miss? Or is it the alpaca blood that you’re always finding the signaling molecule?
WADZINSKI: We have had a pretty good response. It turns out that for all animals, the immune system is remarkable. We have gotten a positive response on greater than 90 percent of the antigens, the molecules, that we put into these animals. We inject them subcutaneously – that is just underneath the skin. Approximately two months later we test for the presence of antibodies. And that response has been very high – greater than 90 percent.
Is there a particular reason why?
WADZINSKI: The production of antibodies is a way the body uses to fight off infections, so it’s kind of a natural process that occurs in the body. Any time there’s a foreign substance in the body, our immune system attacks it and generates specific antibodies recognizing the foreign substance. In this case, our molecule of interest is a foreign substance that the alpaca’s immune system is recognizing and attacking by generating specific antibodies that help facilitate the removal of the foreign substance.
Is there something special within the alpaca blood that helps you with this research?
WADZINSKI: The alpacas actually have two different types of antibodies – the so-called conventional antibodies that we all have. They also have what’s called a heavy chain only antibody. And the nature of this heavy chain only antibody is such that one can use a relatively straightforward molecular biology approach to isolate a fragment of that antibody that actually recognizes the molecule to which it’s targeted. That small fragment has a wide variety of applications. It is heat stable, acid stable, and there’s good evidence that it passes the blood brain barrier. So, we can use it for a large number of research purposes as well as diagnostic applications.
And can you go into your own research – when you inject it into the animals and then you get the response back, what do you do with that?
WADZINSKI: We’re studying one of the protein phosphatase 2A subunits – namely PPP2R5D. It turns out that that subunit is mutated in patients that have this syndrome known as Jordan’s syndrome. They are characterized with intellectual disability, a large head, low muscle tone, and they have autistic like features. We put fragments of that protein or the protein itself into the animal and then harvest the antibodies from the animal, the so-called polyclonal antibodies. But we’re also taking the peripheral blood mononuclear cells, which are produced in the animal. We can isolate the small antibody fragments, also known as nanobodies, from those cells.
And how does that translate into potentially helping to save these kids and basically save lives?
WADZINSKI: At this point it’s strictly research focused as far as the antibodies go. Jordan’s Syndrome is a rare disease, and there’s not much known about how the mutated protein alters the protein’s function. One of the ways that investigators have approached this problem is to develop antibodies that help facilitate studies aimed at better understanding the structure, function, and regulation of these enzymes. We are tackling these questions by generating alpaca-derived antibodies and nanobodies because not only can we use them to visualize the protein itself, but we can also exploit the small specialized antibodies to potentially modulate the protein in either a positive or negative fashion. Currently we don’t know if these mutations are inhibitory or activating in terms of how they work. So, to have a plethora of different tools to be able to study this will increase our understanding and hopefully pave the way for future development of treatments or potentially even therapeutics.
Recently there was a drug approved using llama antibodies. Can you go a little bit into that?
WADZINSKI: I think the first camelid-derived antibody that’s been used as a drug came out this year and it is for a blood clotting disorder. I’m not exactly sure of the physiology there, but this is camelid-derived antibody that has been developed into a therapeutic. There are a number of different conventional antibodies that are currently being used as drugs. You can tell if a drug is an antibody if the generic name ends in “ab”. I think the number of antibody drugs has increased exponentially over the last several years. As far as the antibodies go, or the antibodies derived from camelids – I think they have a wide variety of research applications but also the potential to develop into diagnostics or possibly even therapeutics. There are a number of different investigators and companies across the world that are working towards that goal.
And are they contacting you?
WADZINSKI: No, many of these companies have their own antibody platforms already. The initial discovery of these unique antibodies in camelids goes back probably twenty five years – in Belgium at the University of Leuven. Investigators have seen the potential in these unique antibody fragments. Several companies have also been studying these antibodies and their potential applications.
What are some of the conditions that these antibodies can potentially help with?
WADZINSKI: When you think of antibodies, as I said, they can be used for research, diagnostic, and therapeutic purposes. From a diagnostic standpoint, the small nature of the nanobodies allows better visualization of various cancers because the nanobodies are more tissue permeable. Surgeons could potentially use a fluorescently-tagged nanobody recognizing a cancer protein to more clearly see the margins of the tumor and surgically remove it. As far as therapeutic development, the small nature nanobodies could potentially allow disruption of protein-protein interactions, which has been a challenge in drug discovery. Small molecules aren’t as good for disrupting protein-protein interactions, but nanobodies have a larger binding surface – they might be better for disrupting certain protein-protein interactions. And if you can identify a protein-protein interaction that you want to disrupt to lead to a therapeutic benefit, those are the ones that you might want to target with nanobodies.
And as a whole, what impact do you believe these antibodies and your research in general can have on the medical field?
WADZINSKI: A lot of different investigators and companies are working on different targets. My own research is largely focused on Jordan’s syndrome, and this research is being supported by Jordan’s Guardian Angels. If there is anything I can do with these antibodies to help us figure out what is wrong with the mutated PPP2R5D protein and help us find a route forward in terms of potentially treating this disease – that would be awesome. Working with these families has been a tremendous source of motivation and inspiration for me as a scientist. Getting to meet them and seeing that some of the research you’re doing, although a long ways from a cure, could lead us in the right direction that could ultimately help these children is truly motivational.
Anything that I didn’t ask you that you believe people should know?
WADZINSKI: I think the one thing that I think gets left out, from my perspective, is there are other companies and investigators developing antibodies for research, diagnostic, therapeutic purposes. I think one of the beautiful things about what we’ve been doing is it’s allowed us to engage three different groups of people – a group of scientists here at Vanderbilt, a group of farmers out in rural Tennessee, and the families who have children that are afflicted with this syndrome. Being able to bring together these three diverse groups of people has been beautiful, in my opinion. You’ve got scientists learning from the farmers. You’ve got the farmers learning about antibodies and their potential applications. Then of course you have the families and patients who have simply been awesome, and are bridge that brings us all together. From a personal standpoint, knowing that some of this research could impact them in the long run, I think that’s been the most rewarding for me.
END OF INTERVIEW
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