Cancer Treatments For Dogs

[applause] elaine ostrander:hi, everybody. so thank you so much for coming. so for the row of people sitting behind me,you do not get to fill this out until you've heard me speak. so put your pens and pencilsdown there, all right? so how many -- how many people in the audience have dogs? yeah,so good this is a great, dog-friendly audience, because we're going to be talking today aboutdogs, and a couple of different aspects: we're going to talk about health as well as morphology,you know, why does are different shapes, and how all that data sort of comes together toinform us about human health and human biology as well. so we'll leave some time at the end;we'll have lots of time for questions. so

this is the closest ancestor to the dog. whoknows what we're looking at here? audience response:wolf. elaine ostrander:what kind of wolf? gray wolf, right, of course. so dogs were believed to have domesticatedfrom gray wolves. the latest data suggests maybe around 13,000 years ago. so evolutionarily,that's really a -- that's a drop in the bucket. i mean, that's not very long to go at all,and so one of the things we're always sort of thinking about and pondering about whenwe look at our pets, wandering around our house is, you know, is whatever they're doingcaused by genes that are embedded in this wolf genome, or is this something new that'scome up during the process of domestication?

and that's one of the things that we go backto and we ask ourselves over and over and over as we look at all of our friends. so how many of you recognize your dog breedup there? all right, there's got to be some golden retriever owners, i'm betting? there'sprobably some schnauzer owners, right? maybe some hound owners? you know, a weimaranerowner or two? so these are a small number of the 175 breeds of dog that are recognizedby the american kennel club. the american kennel club is the largest registering bodyof dogs in the united states today, but they are not by any means the only registeringbody. there is the united kennel club, and several other kennel clubs as well, and thereare kennel clubs actually all over the world

that in aggregate recognize 493 differentdog breeds today -- 493 different dog breeds today. and those dog breeds show an extraordinaryamount of variation, right? so we would say that there's probably three things you needto keep in mind as we're going to talk about dogs for the next 45 minutes, or so. it'simportant to remember that, even though all of these guys look very, very different -- theyhave a different body size, different coat colors, different head shapes, different leglengths -- all dog breeds are members of the same species. so they all have the same karyotypeor the same chromosomal organization; they all have the same genome organization; andthey can be crossed to do produce fertile offspring. now, clubs like the american kennelclub don't really encourage you to cross dogs

from one breed to the next, and as a matterof fact, to be, for instance, a registered golden retriever, your parents had to be akc-registeredgolden retrievers, and your grandparents in turn had to be akc-registered golden retrievers. so one of the reasons people like me studydog breeds is because each one represents sort of a closed population. so if you thinkabout human populations that are isolated, you know, people who live in finland or iceland,or people who have lived on islands for many, many generations, or the bedouin pedigrees,geneticists like to study those kinds of populations because there isn't a lot of add mixture fromthe outside. they really have sort of a set number of different alleles at each geneticloci, and that makes the problem of studying

complex traits, like diabetes, and cancer,and epilepsy actually an awfully lot easier. the only problem in human genetics is thatthere's a limited number of such isolated populations, whereas in the dog world we have493 of them. and so we actually in my lab have been working hard to get dna samplesfrom each one of those 493 breeds. we have a great relationship with the american kennelclub. we don't breed any dogs. we don't keep any dogs in kennels, but if you go to a dogshow or an obedience trial, or a specialty or an agility trial, anywhere here in thetri-state area, you're probably going to find someone from my lab there taking cheek swabsor handing out kits or collecting blood samples. and we've now got a set of 50,000 dna samplesin my laboratory. so that's pretty impressive,

huh? so when we look at these dog breeds, we reallysee extremes of variation, and one of the things that makes a breed a breed is thatthat variation breeds true. so if you cross one golden retriever to another golden retriever,the puppies are all going to pretty much look the same, and there's some nice examples uphere for you. you know, if you cross one dalmatian to another, they're all pretty much goingto look the same. cross one boxer to another, they're all pretty much going to look thesame. and the american kennel club is really strict about the things that are important,and that define each breed. and it's different for different breeds. for some breeds it'stheir coat color; for some, it's how tall

they are; for some, it's how long their legsare; for some, it's the shape of their face; how far apart their eyes are; whether theirears are perked or whether they're down; and when a dog goes into a dog show, those areall the things that the judges are actually grading them on. so because breeders havebeen breeding for those traits for years and years and years and years and years, thoseare the things that in my lab we're trying to find the genes that control them, and indoing so, we're sort of getting a vocabulary of growth regulation that we really can'tget from studying worms, and flies, and mice, and rats, and all those other traditionalmodel organism systems. now, i really like this picture, and don'tworry if you can't read the writing here,

but what we've done is we took 1,000 dna samplesfrom dogs representing 85 different breeds. so we took about 12 different dogs from eachbreed, and we tested their genome at 100,000 different positions, and we were looking atthe variation in the a's and those t's and c's and g's that you're always hearing about.and then we fed it all into a computer program, and we said, tell us how all the dog breedsrelate one to another; and that's what this wheel -- and you can really follow the colorcoding in the round -- is telling you so well. so if you look at your upper right, you seethere are the red, and you see spaniels, the american cocker spaniel, the english cockerspaniel, the english springer spaniel, the cavalier king charles spaniel, the irish waterspaniel, the brittany spaniel, and so on.

and if you look over down at about 4:00, you'llsee the sort of bull mastiff-type dogs -- the miniature bull terrier, the french bulldog,the bulldog, the boxer, and so on. now, we have this hanging in my lab and weuse this every single day as we're developing a hypothesis. so if i'm going to study somethinglike epilepsy, and i've got a whole bunch of samples from english springer spaniels-- those are the red up there at about 2:00 -- then i can actually probably go out andget dna from affected american cocker spaniels, and irish water spaniels, and cavalier kingcharles spaniels that are also affected, and i can probably be correct when i hypothesizethat they all got the disease because they carry a mutation in the same gene. and that'sbecause they all share a common set of founders;

that's why they're all colored in red. andi study epilepsy in the mastiff-like breeds -- well, they probably have a mutation ina different gene, but when i look at the mastiff, and the bull mastiff, or the bulldog, or theboxer, and the french bulldog, again, they're all colored in kind of that teal color. theyprobably got the disease because, again, they share a mutation in the same gene, and theylikely got it from a founder. so how long ago did this founder live? well,how long ago do you think most dog breeds were developed? you know, domestication occurredabout 13,000 years ago. how long do you think most dog breeds have been in existence? anyguesses? female speaker:two hundred.

elaine ostrander:you're pretty close. about 300 years. so most dog breeds were developed in europe, mostwere developed by fanciers during the victorian times. so most of them have only been aroundreally for 200 or 300 hundred years. so again, evolutionarily, we're not even talking abouta drop in the bucket; we're talking about a, you know, a perspiration drop in the bucket.i mean, as tiny as you can really get. so we can make these kinds of hypothesis whenit comes to a morphology or when it comes to disease susceptibility, and we're usuallyright. and the other thing is that the genes that we end up finding by studying dogs usuallyturn out to be important for the same things in humans. it's just that, gosh, it's a loteasier to go to my freezer of 50,000 dog dna

samples, and pick out a few related breeds,and look up their health records, and find out what they've got and what they don't have,and correspond with their owners, and start studying a pool of affected and a pool ofunaffected dogs in order to find a disease gene of interest. okay, so this is actually one of my favorite,favorite dog pictures. this is a harlequin great dane. it's skeletally among the largestof the dog breeds, and down here is a little chihuahua, which is skeletally among the smallestof the dog breeds. now, we've been studying dog morphology for several years in my lab,and we've published papers describing genes that control body size, and leg length, andskull shape, and fur, and we rely on samples

from these extremes in the population in orderto do studies on things like body size. again, remember, both of these guys are members ofthe same species, and they could be crossed -- these two guys here -- to produce fertileif singularly unattractive, but nevertheless fertile offspring, and you might want to givea little thought as to who would be the male and who would be the female when you, youknow. all right, so what do we have over here? [laughter] elaine ostrander:okay, so this is, again, a chihuahua; this is zeus. i had to put this picture in. zeusactually holds the 2013 guinness book of world record as the tallest dog. so he is 44 inchesat the withers, or the shoulders. this is

not photo shopped. this is actually how tallzeus really is. and so we've been collecting samples from dogs at these extremes, as wellas everything in the middle, in order to fine genes that are responsible for this dramatic,dramatic difference in body size that exists within this one species. and i can tell youfrom a couple of papers that we've published that there are about a half a dozen genesthat account for most of that size variation. and so i didn't put their names up, but ifyou're interested i can tell you they're the insulin-like growth factor i gene; there area couple genes on the x chromosome; there's the insulin growth factor receptor; [unintelligible],hmga 2, stt 2, growth hormone receptor. and so some of these may sound familiar to you,because they've been shown in studies of mice

to be important in controlling mouse bodysize. the only thing is you don't learn a lot from studying mice in this case, becauseyou don't find mice that differ in size by 40 fold, right? i mean, wouldn't that be trulyfrightening, i mean, if you did, right? but of course, we do find that in dogs. so we've spent a lot of time identifying thegenes and the mutations that account for variation in body size. and we just published this,and i know it's kind of complicated, but i wanted to show it to you, because i'm actuallyreally excited about the implications of this study. so if you look on the left, you seea bunch of bricks that are either yellow or red, and i know it's probably hard to readon the bottom, but on the bottom is the name

of each one of these genes; one at a time:growth hormone, igf 1 -- so on until we get to igf 1. and you don't have to worry about,you know, really which gene is which. going along the vertical are the weight in poundsof different dog breeds that we've assayed. and then what the red and the yellow is tellingyou is that at each one of these genes, there's two possible alleles, or there's two possiblemutations or variants that can occur. one is an ancient variant that we find in wolves,and one is a new variant that's only been on the planet for a few hundred years, andwe only ever see it in domestic dogs. now, the yellow is the ancient variant that wefind in wolves, and red is the new variant that we've only ever seen in dogs. now, what'sreally striking is if we look at really little

dogs that weigh like zero to five pounds,all the way across you see red, telling us that those dogs have the new variant, theone that we only see in dogs at every single one of these genes, but as dogs get biggerand bigger and bigger, that drops off until it's basically mustard yellow. and so allthose big dogs are big because they carry the dna variant that came from the ancientwolf. so that's kind of cool. so we took that data and we said, how predictiveis it? you know, if i actually take what the data's predicting versus what i really seein a panel of 500 dogs, how good is the predictive value of my half a dozen genes? and that'swhat this line over here on you right is telling you. and the fact that you get a pretty goodline from all of those data points is telling

you just those six genes, if we could assaythem in every single puppy that was born, would have actually have great predicativevalue for what the final size of the dog is going to be. we'd be right to about 82 percent.so think about this. this means that you could go to the pound, you could get a puppy, youcould get a cheek swab, you could get it assayed at these six genes, and you would know whatthe final size of that puppy's going to be pretty close to accurate, to about 82 percentaccurate. so that's really amazing. six genes, just six genes control that much variation. now, one of the things about this study isit actually only holds true for dogs that weigh up to 90 pounds, and if you think aboutthe giant breeds -- the great danes, the newfoundlands,

the saint bernards -- these six genes onlyhave about 5 percent predictive value. so there's probably lots of genes responsiblefor giantism [phonetic sp] in dogs that i actually haven't found yet, and that's oneof the things that my lab is in the process of doing so that we can extend this line,and make it bigger and bigger and bigger until we can actually understand all the genes controllingthat full range of body size, going all the way up to, you know, 180, 200 pounds. now, as we've begun to publish more and moreof this data, people have gotten really excited, and they've kind of picked out different thingsthat they want to study, or that breeders would like to try and breed for. so i havepeople in my lab studying leg length, and

studying skull shape; others are continuingto study body size, some are even studying performance. and we built a big dataset basedon 1,000 different dogs from 85 breeds, and we also included 500 wild canids -- so coyotes,wolves -- from all over the world. and we tested their dna at 1,000 different pointsin the genome, and we made that dna publicly available without any restrictions, withoutany patents to anyone who wants it. so you know, we really encourage other people totry and think about these problems as well. now, i like this particular story. this isone that was led by heidi parker [phonetic sp], and she was a graduate student in mylab, and then she went on to be post-doc, and then she went on to be a staff scientist,and she's been with me for 15 years, and i

actually don't think she's ever going to leave.so heidi has been really interested in short-legged dogs. and there's about 20 such breeds. they'recalled chondrodysplastic. they have a ratio of height to body length less than zero. sothey sort of have a normal head, and a normally proportioned body, but then they have thesevery short and thick legs. in the dog world, we would say their structure is well-bonedor heavy, and their forelimbs, like you can see on that basset hound, are often sort ofbowed, or a little bit curved out. so heidi came to me one day, and she said"i want to try and find the genes responsible for this trait across these 20 breeds." andi said, well, you know, this could be a really hard problem. it could be that they all sharea common mutation, but these breeds were developed

for different purposes. some were developedto be -- to go down rabbit holes; some are fox hunters; some are companions; some areratters. so i said i'm not so sure they're all going to have the same mutation in thesame gene. and she said, "well, i think this is a really interesting problem; i'm goingto try it anyway." and she's not being totally unbiased here, because you see that doxenand that basset hound? those are actually her dogs. elaine ostrander:yeah. and so, you know, when you come to my lab and you work, people are often motivatedby things that they have with their own dogs, and you know, some of my graduate studentsare top -- have top show dogs, or top dogs

in the agility ring. so it's not -- i've actuallyhad mushers come and be graduate students in my lab. so, again, motivated by tryingto understand the underlying genetics. so heidi went after the gene for chondrodysplasia.now, i know this is -- looks like just a bunch of lines to you, but what you're looking athere in the alternating gray and black, each one represents a different dog chromosome;and dogs have 38 chromosomes, and we also included the x, but we didn't put in the y,because we figured there was nothing important on the y anyway. and then what we're doingis we're comparing cases, who are chondrodysplastic, to what we call control, which are all theother breeds that are not chondrodysplastic; and we're looking at 100,000 different pointsin the genome, and we're saying, is there

a chromosome that has a data point that'ssignificantly different between cases and controls? and you can see this is very significantlydifferent here on canine chromosome 11. and for those of you who've calculate p-values,the p-value here 10 to the negative 102. so .0000000 -- put 102 o's, and then a one, andthat's how statistically significantly this is. so this is hugely statistically significant. so we decided to follow up on this, and theway we did it is we looked for something that evolutionary biologists call a selective sweep.so what's a selective sweep? well, when you have a selective sweep, you assume, as i'vetold you before, that there's an ancestral mutation that occurred many, many generationsago. in this case, before dogs were divided

up into lots of different breeds. and thendog breeders breed, and they breed, and they select for different things, and there's alot of scrambling of chromosomes, but the mutation stays, because they're always stillselecting for that one trait; in our case, chondrodysplasia. and so now, when we lookat modern day dogs, their chromosome may look nothing like the ancient chromosome, exceptfor where the mutation is, and in the space right around the mutation. and so we lookfor that region of commonality, and when we find a region of commonality across a groupof breeds who have a trait, then we know the mutation has to be somewhere in there, andthat turns out to be exactly the case here. so this is an old fashioned gel, and i putup because i think probably some of you in

college had a -- have had a chance in scienceclasses to run a gel. a gel simply separates dna based on its size. the control dogs arethings like greyhounds, and boxers, and cocker spaniels, and if you look at the top, youdon't really see anything of the gel. but when you look at the case dogs, and theseof course basset hounds, and the doxens, and the pekingese, you see a bright yellow -- abright white band. and that's telling you that all of these cases have some extra dnathat's responsible for this trait that's not present in the group of controls. so rightaway we know that what our mutation is -- it's not a single base pair change, and it's nota loss of dna. all these chondrodysplastic breeds have acquired extra dna. and in fact,they've acquired an extra copy of a gene called

fibroblast growth factor iv. now, they didn't acquire any of the regulatorymachinery that tells it when to turn on or when to turn off, but they acquired the fullsequence of the gene. and so actually the genes around it, they sort of parasitize theregulatory machinery from genes around this -- what we call retro gene, and that's tellingit to be expressed in fetal chondrocytes, and you know what that's doing? that's closingthe growth plates prematurely. so the legs never elongate as long as they should. thisgene is expressed, the growth plate closes, and er, er, er, er, er [phonetic sp] -- theleg can't elongate to its full and natural length. and every single one of those 20 breedsi showed you has exactly that gene, including

this, which is the corgi breed. so this was really exciting, and we were ableto publish this 'science,' not because the editors of 'science' care about corgis, althoughi think they should, but because this was sort of a new way to screw up the genome thathad never really been described before in mammals. and the other thing is, we of courseknow that there are humans who suffer from forms of what we've historically called dwarfism,but it's really chondrodysplasia, and we don't always know what's causing that. so now thisgene goes into that lexicon, into that vocabulary as something that we need to think about whenwe look at those. so this is a really neat example, just by studying a phenotype thatdog breeders have been breeding for for a

couple of hundred years. in a bunch of healthydogs using dna from my freezer, we've been able to figure out a whole new mechanism forscrewing up the genome, and we've been able to add a gene to the medical genetics vocabularythat turns out to then become very important. so this to us is really a huge success. so we've gone on to do that in several otherways, and i'm going to give you one more example in morphology, and then i'll give you oneexample in disease, and then we'll have some time for questions in the end. what are these? audience response:[inaudible] elaine ostrander:absolutely. these are all skulls from dogs,

and these are pictures we took down here atthe smithsonian. turns out they have a lot of skulls in the back room. elaine ostrander:right. and these are all different dog breeds, and they differ in both shape, and what else? audience response:size. elaine ostrander:and size, exactly. so when jeff shaunenbeck [phonetic sp] joined my lab, he said "i wantto find the genes that control this. i want to understand the genetics of the skull shapeand size in dogs." and i said, well, i don't know. this sounds like a really hard problem,but jeff had one of these giant leonberger

dogs with a big round, kind of fluffy face,and so, you know, this is what he wanted to do, and so that's what we did. now, the firstproblem we had was we don't know how -- wow do you quantitate a skull, right? i mean chondrodysplasticis easy. the dogs either have it or they don't have it; and body size is easy -- you measurethem or you weigh them. how do you actually quantitate a skull? it turned out to be ahard problem, but we solved it, and what we do is we have something called a microscribedigitizer, and we touch each skull at 51 different landmarks, and that sends data to a computer,and then in the end, the computer takes the 51 data pieces and it draws a three dimensionalpicture of what that particular skull looks like.

so here i'm showing you top and bottom andside views of a particular dog's skull, and everywhere there's a red or a blue number,that's one of those data points that we've gotten. so the palate can be long or short-- that's the roof of your mouth -- for a dog that has a long or a short snout; thatangle in picture c between the rostrum and the nose, that can be, you know, pretty mucha ski slope, or it can be at a right angle, like it is in a newfoundland. we would saythey have a roman nose, or a very high forehead; and there's variation actually in every oneof these traits. now, we've actually been fortunate to travel around the world. we'vemeasured about 1,000 skulls from 161 different breeds, and people always ask me, where doyou go to do this? and we go to lots of museums,

and certainly the smithsonian is the firstplace we went, but we've been to museums and universities all over the country, and actuallyall over the world. the university in switzerland actually has something like 2,000 canine skullsthat we're about halfway through measuring now. but i have to admit, there are a group ofpeople in the united states that have a lot of skulls in their basement, and i don't why,but they do. and they all call me, and they say, you know, i got a bunch of dog skulls.why don't you come down and measure them? this is alex, one of the people in my lab,and she's measuring the dog skulls. this is a gentleman in california, who called us tofly out and go down to his basement, and measure

all the skulls, and you can see he has allkinds of animal skulls. we really don't know why. we don't ask those questions. we justmeasure the dog skulls, and get out of there, but yeah, there's a lot of these people inamerica. they all want to friend me on facebook. you know, it's a whole culture thing. so anyway,but these people have been very generous with their collections, and we've actually gottena lot of data from skulls where we could verify the breed and we could verify the age. okay, so this is -- this is in some way sortof a tragic slide. so this is really what motivated jeff to begin this project. so inthe left-hand column are a set of human conditions that are really different kinds of craniofacialabnormalities, and i know not -- you're not

familiar with most of those words, but theydescribe different abnormalities that we see unfortunately in humans, often associatedwith particular syndromes. and next to them are breeds where the breeders are actuallytrying to breed that in as part of the breed standard, and really one of the examples thati use -- and this is not something that the american kennel club is doing, this is notan akc breed, but this dog shown here is a pachã³n navarro, and the breed standard includeshaving this deep cleft that goes all the way from the outside of the snout, all the waydown in to the roof of the mouth. so you know, this is not something that american breedersare advocating, but you know, it is something that, you know, we see and so we want to tryand find those genes, because we think that'll

help us understand something about cleft lipand cleft palate, and we're actually interested in all of these craniofacial features. so the one that we've been doing the mostwork with is called brachycephaly, and that means having a very pushed-in face. so ifyou think about your saturday morning cartoons, those of you that are old enough, and some-- you know, something happens and, you know, the face just kind of accordions in like gnaaa[phonetic sp] -- kind of like that, right? and so that's the kind of appearance you seein the pug or the cane corso, and that's very different than what you see in the afghanand the bull terrier, which have very elongated noses, and that's referred to as a dolichocephalicphenotype. so a long nose versus that very

pushed-in face. so we've been going afterthese kinds of genes, and we do the exact same experiment over and over, we comb thegenome, and we look for evidence of a selective sweep, and that's what the data actually lookslike. so these are base pair positions along canine chromosome 30, and you see a certainlevel of chatter, and then when you get right here, you can see this big dip. and that'stelling us that there's a selective sweep there, that that's a place where there's alot of homogeneity, that -- something breeders have been selecting on for years and yearsand years and years. so they don't know that there's a gene under there. they don't knowwhat the gene is they've been selecting on, but we're going to find it, and we're goingto tell them what it is.

okay, so here are a set of 12 dog breeds thatmy lab has now sequenced. we've sequenced these breeds pretty deeply so we've got apretty good genomic sequence, and we picked these breeds for lots and lots of reasons.we have lots of different studies going on, and there's another actually 40 dog breedsthat we and others have sequenced as well, but these were the first 12. and partly, wepicked them because if you think about going very brachycephalic to very dolichocephalic,you have a really nice continuum. so we will be able to use this data to try and figureout what's underneath that little v, that statistical blip that the breeders have beenselecting on over and over by comparing the brachy- and dolichocephalic dogs.

now, you've heard a lot about the genome project,and usually people talk about the human genome project. when i hear about the genome project,i think about the canine genome project, because that's what's really important to me. so iknow this, again, looks like a checkerboard, but each one of the rows is giving us informationabout the sequence variation we saw in those 12 dog breeds. so we started out with 190,000base pairs, and 2,000 possible variants, and then we started filtering and filtering. wegot it down to 85,000 variants and 48 variants, and in the end, when we applied the sequenceof all those other breeds, we were able to find the single base pair in the single genethat turns out to be important, and it's how canine chromosome 30 contributes to that facialphenotype. so the gene is called bone morphogenesis

protein 3 -- kind of makes sense it's a bonemorphogenesis protein -- and it is a single base pair change that changes one amino acid,phenylalanine to lucy [phonetic sp]. one amino acid. now, when jeff came to me with that data,i said, gosh, you know, i believe you, but in order to publish this, we're, you know,we're going to have to have more proof. and so jeff did lots of statistical studies, andthey all looked really, really good, and we wrote it up, and i said, you know, it's prettygood, but i think we're just going to need just a little -- a little bit more proof toget it into one of the fancy journals. and jeff said, "well, okay. i can knock that geneout in zebra fish, and i can make a pug-nose

fish." and i said, "you're going to make apug-nose zebra fish?" and he said, "i'm going to make you a pug-nose zebra fish." and sothere's a technique he used, and he knocked that gene out, and that is exactly what jeffmade. so let's forget the top row for a second -- i'llcome back to that -- but if you look where it says e, f, and g, that's a zebra fish,you know, several -- about 48 hours after the embryo was fertilized, and it didn't getinjected with the stuff that knocked out the gene. and then the blue is the cartilage fromthe top of the jaw, the top jaw, and the g is the bottom jaw. and now, the other twoare examples of fish that did get injected with what we call morpholinos, and they knockthat gene out, and you can see that the jaw

is gone, especially that bottom jaw. look,there's almost no blue staining in j and m, and even the top jaw is pretty screwed upas well. so in essence, jeff made me a pug-nose zebra fish, and indeed, in doing so demonstratedthat by knocking out just that one gene, we're able to dramatically affect the jaw structure.and so this gene, in fact, does turn out to be important in one particular type of humancraniofacial abnormality. so this is another example of how, you know,we started with healthy dogs, long-lived dogs, but they had a particular phenotype, thissort of pushed in face, and we've been finding the genes responsible for that. and there'snot just one; it turns out there's actually several. and in aggregate, they are responsiblefor the dramatic difference between being

brachycephalic with a pushed in nose, or dolichocephalicwith that elongated nose. and we can prove we're right by going to some of these modelorganisms like zebra fish, which are these sort of little tiny fish that people use inthe lab sometimes. okay, so those are examples of how my labhas been studying morphology. and right about this point in time, someone usually says tome, well, you know, that's great, but -- gosh, dogs have an awful lot of diseases. do youactually, directly study any of those diseases as well, and if you do, are they telling ussomething about human disease? and the answer to both those questions is a resounding yes.so here are the top 10 genetic diseases in dogs, and what do you notice is number one?

audience response:cancer. elaine ostrander:right. about one in how many humans will get cancer in their lifetime? anybody? about onein four people will get cancer, some kind of cancer at some point in their life. theymay not die of it, but they'll get it, and about one in three dogs will get cancer intheir lifetime as well. how many of you have had a dog or a cat who got cancer? yeah, right.so in my lab we do in fact study several different types of cancer. now, this is one of my favoritedog pictures ever. it was actually sent to me by a very well-known, and very generousdog breeder, and she breeds -- what are these? audience response:poodles.

elaine ostrander:standard poodles. right, these are standard poodles. so about four years ago, we startedgetting phone calls from people who owned standard poodles, telling us that the dogswere getting a particular kind of cancer, and it's called squamous cell carcinoma. andoddly, it was occurring in the toes, and sadly the way the veterinarians have to treat itis they have to actually remove the toes. and so that's horrible for the dogs, it'shorrible for the owners. if the dog's a show dog, it won't be after that, and people obviouslydon't want to breed to those dogs after that, and so this is really a big deal for thiscommunity, but what was so interesting about this is they said, you know, we only eversee the disease in black standard poodles,

and we never see it in white standard poodles.and so we've now looked at hundreds of standard poodles, and we see it in black and brown,very, very, very, very, dark gray, but we don't see it in the white, or the cream, orthe apricot dogs. so we thought, well, you know, there's a lot of selection for coatcolor in standard poodles. maybe they've inadvertently selecting for a cancer gene as well, and thatin fact turns out to be the case. now, let me just tell you that we study lotsof kinds of cancer in my lab, and in fact, across the world dog geneticists study lotsof kind of cancer. so those of you who have long-limbed breeds like scottish deer houndsor irish wolf hounds probably worry about osteosarcoma. we see tons of bladder cancerin scotties, and westies, and shelties, and

we actually have a paper we're writing aboutthat now that comes again from sequencing tumors, the dna from tumors that we find inthose dogs. if you've got a bernese mountain dog, a super wonderful dog breed that's reallyincreasing in popularity, or a flat-coated retriever, one in four, one in six of thosedogs will get malignant histiocytosis or histiocytic sarcoma. stomach cancer, we see in the belgiansheepdog, the belgian tervuren, as well as in the chow chow. universally lethal; dogsdon't survive it. and the idea here is we study these in dogs because the breed structure,as i told you at the beginning, simplifies the overall problem. we'll see shared geneticsamong affected dogs, and it'll be distinct from what we see of healthy dogs of the sameor, remembering the wheel, very related dog

breeds. so this is a picture of squamous cell carcinoma.you can see that the toe is sort of blown out. it's the most common nail bed cancerin dogs. if you are a giant schnauzer, your chances of getting this are 22-fold higherthan the average mixed breed dog walking down the street. if you are a briard, your chancesare 10-fold higher than the average mixed breed dog walking down the street, and ifyou're a standard poodle, they're about 6-fold higher, although standard poodles are wherewe see most of this, because they're the most popular of those three breeds. and again,in the cases of the standard poodles, we only see it in the black dogs, not in the white.for really complicated reasons in the briard,

we actually see it in black as well as whitedogs, and we figured out why that is. it's actually a very, very complicated geneticstory, but let me tell you a little bit about standard poodles. so we, again, combed the genome with our 100,000points of variation, and we found a signal on canine chromosome 15. it wasn't as strongas what we saw when we were looking at those morphologic traits, because breeders haven'tbeen trying to breed cancer into dogs the way their trying to breed, you know, shortlegs or large body size; but nevertheless, it's there and it's in a lot of dogs. now,i think this is the last data slides that i'll show you. and so what we did is we founda region on canine chromosome 15. it was about

a million base pairs long. we sequenced itin lots of affected and non-affected dogs, and everywhere that there was a possible mutation,there is a triangle. and then, we defined a region that, you know, maybe if we wereliberal in our thinking, was about 500,000 bases in size; and if we're conservative inour thinking, it's about 800,000 bases in size. and what's cool is there's only onegene in that region, and that gene is called kit ligand, and we knew instantly that wehad found the right gene, because it's a gene that's important in coat color, but it's alsoa gene that has been shown to be important in cancer. so we did a lot of work. it tookabout three years, and we in fact, found the mutation, and in this case, we again foundsort of a new and interesting way that the

genome gets screwed up. so it turns out that the mutation is, again,extra dna; it's not a deletion. it's an insertion of about 5,000 bases, and it can be presentone, two, three, four, five, or six times. and the more times it is present, the morethese green proteins bind; and the more these green proteins bind the more they ramp upproduction of this gene kit ligand; and the higher you ramp up production of this, thegreater the chances are that you're going to get the cancer. so if you're a dog whohas this insert present on both of your chromosomes four or more times, boy, your chances of gettingcancer are really, really high. if you have it maybe four times on one chromosome, threeon the other, it's sort of moderate. but four

is really the threshold. if both of your chromosomeshave it repeated three times or two times or one time, or any combination of thereof,no chance you're going to get cancer. no chance. and we've looked at hundreds and hundredsof dogs, and we've never even found one. so this is one of these sort of threshold dealswhere you have to get kit ligand ramped up to a certain point in order for it to go aheadand cause the cancer. so it makes it hard to develop a genetic predictive test, butpeople are in the process of doing that. we found out why white standard poodles didn'tget this. it isn't because they didn't carry the mutation, the four-four genotype. theydidn't -- they do indeed have chromosomes where this insert is present multiple times,but they have a compensatory mutation on another

gene called mc1r, and it completely knocksthis out. so it's a case where they have a bad mutation, but they have a good mutationand good triumphs over evil in this case, and so they never, ever get this, even thoughthey carry the bad genotype. so it's an important lesson, because just looking for the presenceor absence of the bad genotype isn't really totally predictive. you also have to lookfor the presence or absence of the compensatory mutation, and all the white standard poodleshave it, none of the black standard poodles have it. and so that explains the differencein what we observed in coat color. so i'm going to stop there. we have about15 minutes to ask questions, or 12 minutes or so. i hope i've showed you that dogs area really fun system for looking at both simple

and complex traits, including susceptibilityto cancer, which is of course a very complex trait. when we study morphology in dogs, welearn things that are important about development of all mammals, and that includes humans aswell. for both canine and human health, these studies of cancer have become very, very important,and there are labs really at vet schools, as well as at non-vet schools that are studyingevery conceivable kind of cancer, as well as all the other common human diseases: epilepsy,diabetes, heart disease -- you know, whatever you've got including morphologic traits youdon't like, like baldness or obesity or things like that. those -- there are labs all overthe world that are studying those things as well.

many of these studies are long term; someare short term. there are a number of disease genes that we've been able to find the mutationfor. we've been able to develop a genetic test, and breeders are now using those genetictests to make really good decisions to produce healthier, more long-lived dogs. so you don'tsee kidney cancer in the german shepherd anymore, and we've been able to wipe out collie eyeanomaly, which is a degenerative disease of collies and border collies, and a number ofherding breeds, as well as several other diseases. and this has all been done in collaborationwith breeders and owners and veterinarians actually all over the world. samples are always needed so if any of youhave a really interesting dog breed -- i gave

this talk yesterday, and a man came up tome afterwards and said "i have a shiba inu if you want dna from that." yes, we do wantdna if you've got an interesting dog breed. we love your labs, we love your golden retrievers,we love your german shepherds, but we probably have enough dna on those, but the more esotericbreeds, we're still always collecting more dna from. and you know, great progress canbe made, but you know, it is necessary to get the dna samples, and allow us the timeto do our work. so for the breeders in the audience, i know many of you have contributedsamples, and you wonder how long it's going to take. wel, sometimes it's a year, sometimesit's going to be five or six years, because some of these problems end up being simple;some of them end up being very complex, but

our goal is always to make available to yousome sort of a diagnostic test. so i'm going to go ahead and stop there, andallow you to -- turn up -- maybe turn up the lights? turn up the lights, and i'll go aheadand take questions. okay? elaine ostrander:fire away. female speaker:[inaudible] elaine ostrander:this slide? how do we find out what? female speaker:how do you find out the disease risk? elaine ostrander:how do we find out the disease risk? so what we did is we got dna from a lot of poodleswho had the disease and a lot of poodles who

didn't have the disease, and then we sequencedit to see how many times this was re-iterated, how many times this particular 5.7 base pairunit was repeated. and we actually knew from looking at the human genome, we knew exactlywhat these 5,000 base pairs do. we knew that this protein binds to them, and when it doesit ramps up production of kit ligand. so we can do statistics looking at dogs who havetwo copies, three copies, four copies, five copies, and we can look at them at 10 years,11 years, 12 years, 13 years, 14 years, and see who does and doesn't have cancer. andfrom that, we can figure out what the risk is for a dog who has three copies or fourcopies or five copies, and four is really the cutoff. if both of your chromosomes havethree copies or two copies, or one copy, we've

looked at hundreds of dogs, and not a oneof those dogs has this kind of cancer, but as soon as one chromosome gets four cancer-- has the repeat -- repeated four times, then we start to see the incidents of cancercreeping up, and the more you have, the higher it gets. okay? and there are you know formalstatistical tests that we can apply to that, and if you're interested, if sort of got astatistical mind, come see me afterwards and i'll give you the paper and the tests thatwe use. okay, other questions? yep? female speaker:i notice that you were [inaudible] dogs [inaudible]? elaine ostrander:sure. so the question was, is epilepsy indeed prominent in dogs, and yes, we see it in lotsand lots of dog breeds; and have we found

genes? we've been part of one study that identifiedone gene, published it with a group in belgium several years ago in science, but lots ofother people are looking at this problem, because you know, not only is it an importantproblem in human, it's a big deal in dogs. i mean, if your dog has epilepsy, i mean,that is a lifelong problem that you as a pet owner have to deal with, and certainly thosedogs don't show and people don't, you know, put them in the breeding pool if they know.and so some of those genes have been found; not all of them have been found. and thereare some breeds where it is a much worse problem than it is in other breeds. so that's oneof the areas where we see some of the most active and intense work. yeah, you had a question.

male speaker:this is sort of a general [unintelligible] not so much a question. elaine ostrander:okay, i probably won't know the answer, but go ahead. male speaker:well, i was just wondering what sort of breeds made up the dog reference gene? elaine ostrander:oh yeah, sure i do, because i picked the breed for the reference genome. so he asked me,what breed or breeds make up the dog reference genome? so it's one breed, and it's one singledog. so in 2001 i had the chance to pick the

dog that was going to be sequenced, and youknow, truthfully what i did is i probably looked at 100 different dogs of all breeds,and i picked the most in-breed dog i could find. and the reason for that is the way genomesare sequenced is not from the top of chromosome 1 to the bottom of chromosome 38. what theydo is they cut the dna up into a zillion little pieces, and then they sequence it all randomly,and then they have a computer put it back together. so if what you got from mom is reallydifferent than what you got from dad, it's a harder computational problem. if what yougot from mom and dad was pretty similar, it a much simpler computational problem. so i tested 100 dogs at 100 places in theirgenome, and there was one dog that was easily

the most in-breed. it was a boxer, and itwas from new york state, and as luck would have it, it was actually owned by a veterinarian.so when i went to him and i explained that his dog was the lucky winner -- i mean, heactually understood that this was really important. he provided us with an awful lot of dna, andhe understood that, you know, when the dog -- it was a pet, you know, like a lot of ourdogs are -- that when the dog died that we, you know, we would like to get some samplesfrom that dog, because it was, in fact, going to be the reference genome. and his only requestthat i not divulge his name, which i've never done, or his location, which i've never done.so he was great, he was fantastic, but it was a boxer, and her -- it was a she. almostall the reference genomes, except maybe maybe

-- are a she, because then we get good dataon the x chromosome. female speaker:have you guys found a gene related to like a yorkie, just randomly like [inuadible]? elaine ostrander:so is -- have we found a gene linked to fur loss in breeds like the yorkie? so i haven'tlooked at that. there are breeds, like the chinese crested or the mexican hairless, thereare breeds that have very little fur, except for some tufts at the top of their head anddown by their toes, and those genes have in fact been found, but those dogs, that's partof the breed standard; that's how they're supposed to look. when a dog blows all ofits fur, i mean -- and that's an anomalous

sort of thing -- i'm sure there are peoplelooking at it. i'm not one of them, and i'm not aware of a paper, but i could certainlytell where in the literature to look for it. yeah? male speaker:are you able to define what would be the top, like healthiest breeds? elaine ostrander:sure. yeah, so the question is what are the healthiest breeds, and it's a little bit ofa trick question, and the reason is because within every breed there are lines that arereally healthy, the breeders are very savvy, they've gotten dogs from multiple places inthe world that are members of that breed,

or multiple places in the united states thatare members of that breed, and they've worked really hard to maintain the hybrid rigor ofthat breed; and there are other breeders, you know, who have bred a lot of closely-relatedindividuals one to another, and their lines may look good, but they have a lot of healthproblems. so there's no one right answer. i mean, you can look on the internet and,you know, they'll certainly give you those -- that kind of information, and they'll saythings like, well, you see a lot of cancer in boxers or in golden retrievers; or yousee a lot of copper toxicosis in the bedlington terrier. you see -- you know, there's differentdiseases that tend to predominate in different breeds. there are some situations where diseaseis so predominate, like copper toxicosis was

in the bedlington, before the gene was foundthat i think that was a fair comment. you know, there are other cases where i thinkyou just have to really search. i mean, i wouldn't be hesitant to own a golden retrieveror you know, a german shepherd, or any of these other really popular breeds. i wouldjust work really hard to talk to people the breeder had sold to, and find dogs that liveda really long -- a really long life. in general, small dogs live longer for sure-- the terriers, the toy dogs -- in general, they do live longer. and dogs that have aworking job like border collies, boy, some of those live 17, 18 years. my border collielived to be 13 years. so breeds where there's a working lineage -- i mean, they have tohealthy to work -- those tend to be pretty

long-lived. the really big breeds tend tohave more heart difficulties and problems -- saint bernards, and you know, some of thesegiant breeds where they've just almost been bred to be too big for their heart. you know,they rarely live beyond the age of 10 or 11, because they almost always go for -- fromheart problems. so, you know, i always tell people to, you know, pick the breed you want,pick the breed that matches your family. there's all kinds of tests on the internet to helpyou do that. but then work with -- you know, really look around and take your time to finda healthy and reliable breeder. yeah? female speaker:would you say that breeds that are closer to the wolf, like in ancestral [inaudible]lineage, would you say that they are [inaudible]?

elaine ostrander:no. so she -- the question she asked is, are breeds that are more related to wolf healthier,and i guess i would say no, you know, in part because things that maybe look more wolfish,like the malamute or the husky, i mean, you know, there were clearly multiple domesticationevents that occurred in multiple places in the world, and you know, by and large, trulyit's the small dogs that are the healthiest and really do live the longest, and so i wouldn'tnecessarily say those are, you know, among the healthiest, although, you know, i hada malamute that lived to be like 15 or 16 so, you know, again dogs that have a workingfunction, but this was bought from a line of dogs that, you know, were involved in mushing,and so that's -- you know, those are going

to be healthy dogs so i wouldn't necessarilysay that. other questions? yeah? female speaker:do mixed breeds do better than pure breeds? elaine ostrander:do mixed breeds do better than pure breeds? you know, everybody thinks that they're goingto, and i have so many people come up to me and say "i bought a labradoodle or a" -- youknow, they have all these weird, cool names, and they said, you know -- and so -- hybrid,bigger, you know, i'm -- my dog's going to live to be 22, and then they're shocked whenthey don't. and the reason is because, you know, if the parent breeds or the parent lineswere themselves not healthy, particularly if they were not healthy because they hadthe same disease or the same mutation, you

haven't -- it doesn't do any good, right?and so you know golden retrievers, you know, they get a bad reputation for having a lotof inherited disease. i think there are some lines of golden retrievers that are wonderfulout there, but there are also some lines that really do have a lot of disease, and becausethey're so good with families, they're involved in an awful lot of crosses, and you know,you can say that of lots and lots of other breeds. so, you know, in general, what i likeabout pure bred dogs is you sort of know what you're getting. you know, most poodles i knowlive long, long, long, long periods of time. there an awesome breed in all three sizes.they have a reputation in the breed club for being very vigilant, being very careful, andso i don't necessarily think that's really

the best choice. yeah? female speaker:[unintelligible] you talking about being able to remove the mutation? so is that happeningwidely within that breed -- elaine ostrander:so -- female speaker:-- or is there some special [unintelligible]? elain ostrander:-- the question is, you know, removing mutations, does that happen widely? so it really isn'tremoving mutations in the sense of how that language is used, but it's really about breedingit out. so breeding a carrier not to another carrier, but to a healthy dog. and when italk to breed clubs, i tell them first thing,

don't throw all the carriers out of the breedingpool, because they're contributing so many good things that you're going to just endup with something else you didn't have a problem with before. but -- but take your carrierand breed him to a non-carrier. the carrier -- get the progeny tested, breed carriersto non-carriers, and gradually breed it out. and we've seen examples where a small breedhas thrown out all the carriers, and they have a disease that's really prevalent, andthen suddenly they have three other diseases that crop up as recessive. the american kennel club has just been wonderful.i mean, they ask us to come to all their meetings and their specialty events, and their trials,and we are inundated with invitations to come

and give talks about exactly these kinds ofissues, and labs like mine often make the data available without patenting it. sometimesit is patented, but often without patenting it, just so the tests can get out there andpeople can start using it. and breeders are some of the smartest geneticists in the world.they absolutely use it, because it is their livelihood. they want to breed healthier,more long-lived dogs, and if they're the first off the block with a reputation for usinggenomics to breed healthier more long-lived dogs, people love that -- people love that.and so i have found that this -- i mean, i have 50,000 dna samples in my freezer, andi can think of one incidence where i asked for a dna sample and i was turned down. goahead.

male speaker:my question is back to the mixing question. so in agriculture, the way you find new traits[unintelligible] is by crossing breeds so you will get the non-additive effects andthings like that. is there a push in the dog world to try these things out and see whathappens by mixing different breeds? elaine ostrander:no. no. there isn't. male speaker:[unintelligible]? elaine ostrander:right. so the question that was asked was, is there a push in the dog world to, you know,mix dogs of different breeds to try and figure out what's going on with the genetics of someof these traits? i just put this up because

it's my favorite picture in the slide show.and the answer to that is no. first of all, we don't breed or keep any dogs. so we'renot going to do that. and in the pure bred dog world, you know, people -- you know theconvention is you breed dogs of one breed only to members of the same breed, and that'sjust not the convention in that community, you know, to take a dog that may be a popularsire and breed him to a bunch of other breeds to figure out what's going on. that's justnot what is done. so it really isn't. and you know, i'm not in the management of theamerican kennel club so it's not really for me to say, but -- okay, one more? can takeone more question? all right. one more question. sure.

male speaker:you mentioned about 50,000 data points. are they all pure breed or do you also have mixedbreeds and mutts? elaine ostrander:so i take 100,000 data points, and i have 50,000 dna samples from dogs in my freezer.i have -- my lab has very few mixed breed dogs. almost all of those are from pure breddogs. they're not all american kennel club recognized breeds. you know, some of themare odd, you know, european or asian breeds. they're not all akc breeds, but they are purebreeds; they're not mixed breeds. but there are other labs that have focused their studieson mixed breeds. so there's one at cornell who specializes in village dogs, and he'straveled all over mexico, south america, africa,

to the outskirts of town on the garbage dumps,and he and his team sample hundreds and hundreds and hundreds of mixed breed dogs, and that's,you know, his thing. so, you know, you kind of can't do everything, and so that's been,you know, where my focus has been, but there are certainly people doing that. all right,i'm going to let you go. there's lots to see out in the museum, and i'll be around forquestions, and thank you all for your attention.