Hi. Welcome to Dog Genes Tell Surprising Tales, Part II. I’m Elaine Ostrander, and I’m from the National Human Genome Research Institute, which is one of the National Institutes of Health. So today, we’re going to be talking about disease. And we’re gonna try and relate what we’ve learned about disease in dogs to what we need to learn about disease in humans. So, let’s start by reminding ourselves how much dogs and humans share both genetic and genomic features, and as a result several advances as well. So, dogs and humans share a lot of the same medical concerns. There’s very similar disease risk for most of the common disorders. The disease presentation, again, for most disorders is very similar, and that includes the underlying pathology. And how dogs and humans respond to various treatments is also quite similar. Dogs and humans also share a lot at the genetic level. The organization of the chromosomes and the genes is pretty much the same. So, the same genes are in the same order on chromosomes. The dog genome is just divided into more chromosomes. Over the years, my lab, together with those of collaborators, has been able to build maps that have allowed us to navigate the dog genome very, very efficiently. There are a large number of genomic resources that are available for doing dog genetics today. Over 800 dogs have had their genome fully sequenced. And this data, and portions of this data, have become very useful at finding disease genes or genes that control virtually anything that we care about in dogs. Studies of disease in dogs have illuminated similar conditions in humans, and those include cancer, autoimmune disease, neurologic disease. Really, anything you care about in human health we care about in veterinary health. And today, we’re gonna focus our discussion on cancer. So, I’ve listed for you, here, the top ten diseases that we see in dogs. And you can see at the top of the list is cancer. And that’s because about one in three dogs will get cancer in their lifetime, which is pretty similar to what we see in humans. So, this is really a major concern in veterinary medicine. So, we’re gonna go through some examples that illuminate some of the points I think are important. And we’re gonna start with a disease called canine hereditary multifocal renal cystadenocarcinoma and nodular dermatofibromatosis. I know that’s pretty much a mouthful, but we’ll just call it RNCD. So, this is a disease that we see in German Shepherds. And it’s an autosomal dominant form of kidney cancer. Diagnosis can be made early on by observation of these microscopic renal cysts. Now, we also see skin-fibrofolliculomas. We can see lung cysts and pneumothoraxes, but the really important take-home is that if you have a mutation in the relevant gene the chances of getting renal cancer go up 7-fold. And that’s a bad disease. That’s a bad disease for dogs, and it’s a bad disease for humans. So, we really want to understand as much as we can about this. Now, several years ago, a young woman from Norway came into my lab and she said, you know, I’d really like to study this disease. I’d like to be a graduate student in your lab. Her name was Thora Jonasdottir. And I said, well, I don’t know, maybe, I’m not sure. And then she showed me this pedigree. And my eyes really bugged out, because this is an extraordinary pedigree by genetic standards, no matter what organism it is that you’re studying. So, in this pedigree, as always, the circles are females, the males are squares. And those individuals that are affected, like this one here, are colored in black. And in fact, it’s this individual with the red circle who’s responsible for all the problems. So, he’s affected with the disease. And he spent time with an awful lot of females. And in every case, every litter was big, and it produced a combination of white and black, affected… unaffected and affected individuals. So, when I looked at this pedigree, I realized, oh my gosh, we should be able to find the disease gene just using one single pedigree, because this pedigree is so remarkable. And indeed, that turned out to be the case. So, I know the numbers and letters are distracting. Those are markers, like the SNP we talked about in Part 1, that we use to navigate our way up and down the relevant chromosome. And in fact, we were able to use that data to get to a really tiny, tiny portion of the chromosome, where the disease gene was. So, it turns out that the disease in both humans — and in humans, the disease is called Birt-Hogg-Dube syndrome — and dogs is caused by mutations in the gene folliculin. Folliculin is a tumor suppressor, so its job is to keep cancer from happening. So when it’s mutated, it can’t do its job, and cancer is more likely to happen. There’s a signaling link between folliculin and the mTOR pathway and cancer susceptibility. So it fits in a space that’s not really unexpected. Now, this is an interesting case, because the gene region or the locus was first found in dogs, but the actual mutations were found in humans. So, it’s a great example where putting together data from humans and dogs really allowed us to make a very rapid advance in a disease that was both interesting and really important. Now, I should tell you that canine pedigrees like the one I showed you in the previous slide, they’re not common but they certainly do occur. And when we see them, we jump on them, and we show them to everyone because they’re so remarkable. In humans, you just don’t see these kind of pedigrees. So again, it’s the bringing together of human resources and the canine data that really helps us advance these diseases that we care about for both. Alright. So, that was our first example. It’s also true, and as many dog owners will tell you, that there are many breeds that have elevated risks for specific cancers. So for instance, osteosarcoma is particularly common in Scottish Deerhounds, Irish Wolfhounds, very long-lived breeds, Great Danes and Rottweilers. Bladder cancer we see in the Scottish Terrier, the West Highland White Terriers, and the Shetland Sheepdogs. Histiocytic sarcoma in the Bernese Mountain Dog and the Flat-coated Retriever. As a matter of fact, 25% of each of those breeds are going to go on to die of the disease. It’s a terrible disease for those two breeds. Gastric cancer we see in the Belgian breeds, and we see it in the Chow Chow, shown here. This is a very lethal disease in humans. It’s a very lethal disease in dogs. Veterinarians tell me that diagnosis to death in dogs can really just be a few weeks. So, that’s one we’d really like to understand as well. So, we’re gonna talk a little bit about invasive bladder cancer. We want to identify the loci, the genes, and the variants that predispose dogs to invasive bladder cancer. And we’re gonna do it using some population structure-based approaches. So, we want to specifically find the loci that increase risk of getting the disease and the drivers in the tumors that make those tumors so successful. And ultimately, of course, we’d like to be involved in developing diagnostics for invasive cell… invasive transitional cell carcinoma of the bladder, which we’re going to be calling InvTCC. And my lab has also been involved in a transcriptome analysis of invasive transitional cell carcinoma, the bladder tumors, to help identify predictive features as well as drivers of that tumor progression. These are the three breeds that are at pretty much the highest risk of getting the disease. And over here I have the Scottie. If you’re a Scottish Terrier, your odds of getting the disease are 22-fold higher than the average mixed-breed dog walking down the street… 22-fold higher. There’s some amazing genetics buried in that breed. Alright. So, from a human genetics viewpoint, why do we care? Well, in humans, the same disease, the cause is unknown in about 50% of people who get it. Front line therapy, removal of the bladder, is a complex and it’s an expensive operation. At least half of the patients who undergo cystectomy will subsequently die of metastatic cancer. Now, there are chemotherapeutic approaches. They can initially be successful, but resistance then usually ensues. And there’s really no second line treatment. Metastasis is common. And the median survival times really haven’t changed in the last 20 years. So, we like dog models. It’s really the best model for human invasive bladder cancer for the reasons that are listed for you here. So, it is spontaneous. This isn’t something that has to be induced, like a lot of mouse cancers. The age of onset, symptoms, drug response, site of metastases, and pathology are all very, very, very similar between humans and dogs. So, one of the first things that we did was to look at a series of tumors. These were bladder tumors provided by a very valued collaborator, Debbie Knapp, from a series of different dog breeds. And when we looked at them, we found that 85% of the tumors carried a mutation in a gene called Braf. And the mutation — every tumor — was at the exact same position. It was amino acid 600, and it was this valine to glutamic acid change. So, that set off all kinds of bells, because mutations in the exact same gene in the exact same position… position are really important in human cancers. As a matter of fact, we see them in about 40% of melanoma tumors; we see them in thyroid cancers, as well as in colorectal cancers. In humans, if you have this mutation in your tumor, it’s associated with poor outcomes, and the same thing is true in dogs. So, we were really interested to find this because it represents a target, a druggable target. So now, for canine bladder cancer, we have some ideas about how to treat it, because there are drugs developed for human melanoma and others that directly target this particular mutation. So, we now have a route to advancing our studies. This result also gave us a route into some diagnostics. So, we’ve been doing sequencing of urine samples from dogs who have this disorder. And we can tell pretty early on if a dog, in fact, has bladder cancer. And we have a sensitivity where if there’s even one mutation, one mutant sequence read, in the background of 10,000 normal reads we can pick it up. So, this is a place for genomics has really, really helped us advance something about a disease that we care about a lot in canine health — it’s pretty common — and in human health as well. Now, the last tumor I want to tell you about is called transmissible tumor. As a matter of fact, I call this part of the talk “The Long and Lonely Life of a Canine Venereal Transmissible Tumor” because this is a venereal tumor. So, this is what we call a clonally transmissible parasite. Now, not a parasite like a virus, or a bacteria, or a little bug crawling around. This is a transmissible cancer. So, there was a single founder, who we’ll talk about, thousands of years ago who originally got the tumor. And the tumor has now spread from dog to dog to dog to dog via sexual transfer. So, that is, one dog has the tumor and it spreads to the other dog by the transfer of malignant cells. So, that’s kind of scary, isn’t it? It’s endemic everywhere in the world except Antarctica. It is the world’s oldest-known continuously propagating somatic cell lineage. We know it’s been around for thousands of years. Now, it’s transmitted during a month-long period of evasion from the host immune defenses, and then it can later be eliminated. Sometimes it’s lethal; sometimes it’s not. We’re gonna talk about those host immune defenses in just a moment. What’s important is that all of the tumors, all the dogs running around in the world with the tumor today, if you look at their tumor sequence you can see that they have shared origins. There’s a lot of places where there are haplotypes or sequence that’s identical. So, they have a very strong genetic identity, we would say, with one another. And it’s markedly distinct from what we call their transient host. The transient host is the dog who walked into the clinic who had the tumor. So, we can look at DNA sequence and we can distinguish those things. So, the question that we asked in my lab is, can the CTVT genome, the genome sequence, reveal the biological mechanisms that underlie the tumor’s dogged appearance in canids around the world today? And that’s my only pun, so you have to laugh at it. Alright. So, let’s think about how this happened. So there’s a dog, thousands of years ago. He’s what we’re gonna call the founder, and he unfortunately gets this tumor. Alright? He or she eventually… apparently gets this tumor. It is a myeloid tumor, just so you know that. And then lots of different things happen over thousands of years. There are some places where there are lineages that just simply end as indicated by the red X’s. There are other places where lineages go on and on and on ’til we have dogs that we can sample today, like this dog in Brazil or this dog here in Australia. And there are other places where lineages started and they just ended. So, when we began this study, we only had two things. We had the DNA sequence of a dog… the tumor, excuse me… the tumor sequence of a dog from Brazil, and we had the tumor sequence of a dog from Australia. We did not have the sequence of the actual dogs. We only had the sequence of their tumors. And what we wanted to do was figure out what it was that was allowing this tumor to be so successful. So in order to do that, we had to separate the ancient canine genome from the somatic genome or the tumor genome. So, if you… if you take a single dog and he has this tumor, and you just do sequencing of the tumor, there’s gonna be a lot of things in that sequence. There’s going to be DNA sequence from the host, the dog that walked into your office. There’s going to be DNA sequences from the ancient founder. There’s gonna be DNA sequence from the tumor. There’s gonna be lineage-specific mutations. There’s gonna be some systematic errors. It’s gonna be a mess. Now, if you add a second dog, as we were able to do, you simplify the problem. Because you get rid of the DNA sequence from the host that walks into your office. They cancels… they cancel each other out, and you’re left with two things that you really care about. You’re left with the DNA sequence of the original founder, and you’re left with the DNA sequence of the tumor. And that’s what we want to separate. We want to know what has made this tumor so successful. These are the things that we want to separate. Alright. So, how are we going to separate those? Remember, our goal is to understand the genomic underpinnings of CTVT’s remarkable behavior. So, we really want to figure out what we can learn about the origins of CTVT. And we also want to learn something about that original dog in whom the tumor first occurred, and of course that dog’s been dead for thousands and thousands of years. Now, this wasn’t available at the time we did this study, but studies like this are really enhanced, now, by the fact that there’s a worldwide consortium that’s formed called Dog10K. So, the goal of Dog10K is to sequence 10,000 dog genomes in the next 5 years. And there are people from 18 countries… 18 members from 9 different countries who are part of this consortium. And they’re listed for you here. So, this is a really exciting collaboration in canine genomics, and it’s gonna make the kinds of studies I’m going to tell you about in just a moment an awful lot easier. Alright. So… how are we doing this? Well, remember, we’re gonna be starting out over here with the founder, who… you know, we don’t have anything from him. But we have the sequence of these two tumors. And because we have two tumors, the potential number of mutations — insertion-deletion, structural variants, single-nucleotide variants — is initially potentially very large, but it reduces when we compare the sequence of these two tumors. And so then we can go ahead and we can take those variants that we see down here and we can say, do we find them in our catalog of whole-genome sequences of hundreds of dogs that our lab is assembled? Now, these are dogs have been sequenced all over the world, and we’ve assembled the data. If the answer is yes, well, that’s probably part of the sequence of that original founder. And if the answer is no, that’s what we care about. Those are probably some of the mutations that occurred in that original tumor that made it so spectacularly successful. And what do those mutations turn out to be. Well, they turn out to be mutations in all aspects of somatic participation in immune surveillance. So, every part of the immune system that’s important in recognizing self versus non-self has been destroyed over the years by this tumor. And so it’s exquisitely adapted to its niche as a transmissible allograft. We don’t… our immune system… or, the dog’s immune system doesn’t recognize it as foreign. So, it just marches along doing its tumor thing, because it has so wisely knocked out anything that would make the host immune system… system recognize it as foreign. Now, this work was done largely by Brennan Decker and Brian Davis in my lab, and Brennan was a graduate student. And at this point, he wrote up a lovely paper on this and told me he was going to graduate. And I said, well, you know, I really want to know a little bit about that first dog thousands of years ago, that original canine. And he said, who cares. And I said, I care. You have to do it before I’m gonna let you graduate. And so, what Brennan did is he took the DNA sequence from those two tumors that we had, and he put it on a phylogenetic tree, as you see here. But in this case, what he did was he used DNA sequence to build that whole phylogenetic tree. And it turned out that that original dog, the original dog who had that very first mutation, is a Siberian Husky. Because the two tumor sequences grouped on the phylogenetic tree with the Siberian Husky. So, that’s really interesting, because these… the disease is something that occurred after domestication. It didn’t come up in the wolves. It didn’t come up in the coyotes. It’s something that came up post-domestication. So, somewhere in that 13-30,000-year range, that’s when this disease actually developed. We still see it today. We still see it all around the world today. And it’s a topic of intense interest. Now, transmissible tumors are something we worry about in humans. We don’t have them. We don’t have them yet. There are a couple of other examples where they occur. You may have heard about the story in Tasmania, with the Tasmanian Devils. And they also experience a transmissible tumor. And in fact, their tumor is quite aggressive. It’s a facial tumor, and it can affect breathing, it can affect eating. And it’s largely responsible for the rapid movement of the Tasmanian Devil towards extinction. So, that’s a place where we really care about it. And there’s also been reports in some marine life of having transmissible tumors. So, it’ll be interesting to see how all these stories evolve in the coming years. Okay, so I’ve told you an awful lot in the… in the last two videos. And I’m sure some of you are asking, how is it that you can help us? Well, you can help us in a lot of ways. These are the diseases that we study most intently: histiocytic sarcoma in Bernese Mountain Dogs and Flat-Coated Retrievers; bladder cancer in Scotties and Westies and Shelties; and gastric cancer in the Belgian Teurvern, the Belgian Sheepdog, and the Chow Chow. So, if you have dogs who have any of these disorders, please contact us — and I’ll tell you how to do that in just a moment — because we would love to get a sample from your dog and include him in the dog cancer genome project. If you have an older dog who’s very healthy and doesn’t have any of these cancers, their DNA could be used in our control dataset. So, we’d like you to contact us as well. Now, as you know, we study lots of different morphologic traits: body size, skull shape, bone structure. If you have an unusual dog breed, please contact us. We would love to get DNA and include it in the dog genome project from your dog. And if it’s particularly interesting, we may even include that 10,000 dog genome project. So, whether it’s an AKC-registered breed, a non-AKC-registered breed, an internationally recognized breed, you can go to our website, and I’ll show you that in a moment, and you can look at the breeds we’re most interested in. And of course we’re always interested in breed history and phylogeny. So, look at some of our data. Again, it’s available on our website. And if you, again, have an unusual breed that you think that’s a niche that we haven’t addressed, please contact us. We’d love to get a sample from your dog. So, today, these were some of my favorite dog breeds. And these are some of the ones that we’ve done quite a bit of study on. But there’s an awful lot that we don’t know yet about domestic dogs. And this is, as I said at the beginning, the largest experimental collaboration that’s probably ever been undertaken between scientists and the general public. So, we’d really love your contributions. Alright. So, in the… in the last several minutes, I’ve tried to show you that dogs are a great system for the study of both simple and complex traits. In the previous 30 minutes, we talked about morphology, and I showed you how canine morphologic studies illuminate multiple genes and pathways that are of interest to people studying, really, any aspect of mammalian biology. An important and reoccurring theme in dogs is that most phenotypes are controlled by a small number of genes of large effect. And that’s different than humans, where most phenotypes or traits are controlled by a large number of genes of very small effect. And finally, there are people in our community studying every disease you can possibly imagine. And my lab studies cancer, and that’s because canine cancers can help us understand the genetic underpinnings of human cancers. So, here’s our contact information. The person in our lab who collects all of our samples and will send you a kit that you can give to your veterinarian, who will then collect a sample, is Andrew Hogan, up there. His contact information is listed for you. And our website is listed for you as well. So, thank you for joining me today. I hope you’ve learned something new about your dog or about any dog that you care about. And I hope you can think about some interesting ways in which you could be part of our… of our big collaboration and our dog genome studies. Thank you.