Welcome with President Thomas F. Rosenbaum; Session I: The Gut-Brain Connection

good morning good morning Caltech alumni family and friends come and sit down and join us yeah we're back alumni day it's back we're here after the pandemic it's been 2019 since 2019. so welcome um first I want to say it's happening live in Pasadena right now um and I'm going to ask you as you come in to please silence your cell phones but do leave them

on and we'll tell you why in a moment and just note your emergency exits but I should say it's not just a 300 of us here in Beckman today there are over 400 people joining us from techers from all around the globe and they can also check their emergency exits if they wish um I am standard Singh low Caltech alumna BS physics 1983. and in 1983s

here okay all right and host of The Daily radio minute the lowdown on science syndicated by NPR and KPCC now called last a show actually created by Caltech almost 20 years ago by then yay by then PR head Bob O'Rourke under the tutelage of then president David Baltimore um who is brilliant and has a sense of humor too I'm also a proud member of the techer

community of more than 25 000 alumni living and working all over your planet so I'm going to be hosting this first ever hybrid alumni day where we'll be here and people are watching elsewhere as I said um speaking of which are are in-person guests today for this exciting informative interactive and fully Caltech event here to open the date officially now that you're seated I bring you

the Sonja and William davidau presidential chair professor of physics Caltech president Thomas F Rosenbaum a warm welcome to you Tom if I can call you Tom after that okay thank you welcome to all I I want to start with a shout out to Sandra so thank you so much for doing this uh we've been playing Hollywood Squares for the last few years uh but she's a

Mainstay of these events now and uh it's just great to be able to do it in person I also want to thank the speakers where else but Caltech on one day can you get more than three percent of the faculty engaging with our lumps and I want to thank all of you for showing up and those online as well nearly a thousand people or we'll get

there all together sharing in the intellectual or past that we have to present uh so I was coming out Parson Gates where my administrative offices the other day uh yesterday and I saw a number of Tours some of you may have been part of them and I was thinking that the campus has really changed uh of course for those of you who go far enough back

the campus ended at San Pasquale uh it's now of course extending up to Del Mar and in fact you'll find Caltech buildings north of that but a lot of new buildings if you haven't had a chance to walk around the hemitman center is a gathering place for all members of the community has a rehearsal Hall in it it has a much expanded Red Door Cafe where

you can just sit at a table and hear all kinds of great conversations going on all about science and engineering new undergraduate residents the Bechtel residence which allows us yet again now to have all our undergraduates on campus for all four years which I think is really important part of the experience here uh if you go to the northwest corner there's a new center for Neuroscience

a big Copper Building which has lots of interactive spaces which brings together members students faculty post docs staff from all six divisions and if you go inside uh now the Lind Hall of mathematics and physics which used to be Sloan and Gates Thomas which houses mechanical engineering you'll see a complete gut renovation where the facade has been preserved and there's more to come come back in

a few years and we'll have a new center for Quantum Precision measurement the panel I think you'll hear about that how this is an organizing principle quantum mechanics to bring together individuals from lots of disciplines chemistry biology computer science physics Applied Physics engineering who want to use entanglement to be able to build new devices to better interrogate nature and we also will launch next year the

Resnick sustainability Institute Building which is on the west side of Campus as a new entrance and importantly on its second floor will house all new freshman chemistry Labs which now be sustainability Laboratories so a lot of change but the essence of Caltech Remains the Same and you can look at Gordon Moore as a wonderful example of that right Pioneer in the semiconductor industry thinking of him

because of course he passed away uh in March um and and he said if everything you try works you aren't trying hard enough and and I think that's just perfect it's how we've been it's how we challenge ourselves right if if you get an easy solution you aren't pushing yourself hard enough you aren't asking important enough questions you aren't switching to a new field which may

be more productive than what you're working in now even though that's a hard thing to do we see that for our faculty we see that for our students we see that for our alumni and this characteristic of pushing ourselves of pushing ourselves together to discover something new about the universe to develop technologies that improve people's lives continues to be an animating Spirit here at Caltech so

look forward to seeing you at many seminar days and alumni reunions to come thank you thank you so much president Rosenbaum inspiring words uh and So speaking of pushing ourselves as a community today before we begin our first of first of six fascinating sessions I'd like to introduce you to our new friend slido Applause for slido you don't know what it is but you know we're

voyaging into the unknown so today your q a will be facilitated via yes your phone okay so let's take a deep breath if you go to um you see that QR code I want to say good phones are going up Checkers good okay so if you go there it'll take you to the app slido where you can ask questions while our first speaker is speaking and

for instance and if you don't feel like typing on your phone you can actually ask you can actually say for instance I would say quick demo I've been here at alumni day since 1965 as a child so I would say where are the donuts send okay now it'll take a moment if you're looking at the app which I hope that you are for them to okay

the question and then it will it will come up and then there it is as they wear the donuts and then you can like on on the right you can actually hit a like if you like someone's question and it'll move up in the queue of course while our first speaker is speaking do not ask where the donuts are they're on Saturn I think you saw

out there um so these These are actually questions for our speaker and of course we want you to be focusing on the lecture but also if you have a question that goes along the way you can ask and we'll be able to address it likely in the Q a so um and and if you can't log on you can go to slido.com and enter and the

code is Caltech one case doesn't matter for the first seminar lecture and then Caltech 2 will be the second one Etc and we'll be here to support you along the way okay enough with the phones you should have all the programs with those agendas are virtual friends you can use slido also and you can find it on the WebEx platform now here's a brief outline of

our day now that you've had that great breakfast we get a little insight into whether we might actually think with our gut we'll take a step into a fascinating future as we learn how Caltech moves us from quantum theory into Quantum science since we'll already be in the future one of caltech's newest faculty will take us into the world of artificial intelligence and how machines learn

what we might really be thinking about them and if you've been following the chat GPT um but I think it'll be especially fascinating just before lunch we'll ask you to hang on for just a few minutes you'll be the first to hear who's actually been selected for the 2023 distinguished alumni Awards and you'll see an actual film Premiere don't worry it's just a few minutes long

but it's very exciting we'll break for lunch those of you on campus can get the grab and go picnic lunches and explore lots of facilities are hosting open houses which you can find in your programs those of you at home I direct you down the hall to your fridge to your microwave whatever just be sure to check your emergency exits in your own homes but please

be back at Beckman or at your screen by 2PM after lunch where we go where no researcher has gone before exploring extraterrestrial environments on our solar system and beyond we'll explore our hidden this is really a favorite of mine I have to say an unhidden fears with expert from the fear Lab at Caltech I don't know if you went to Cal if you've gone to Caltech

the whole thing was a fear lab but anyway Fizz 106. anyone AMA 95 anyone I'd be interested in hearing what courses were especially fearful for you for me it was all of them even computer science one was and in case you feel like running out you're running out of steam at the end of the day will end the day discovering how Caltech is driving new technologies

with renewable energy and welcome each talk will be followed by a 15-minute break so you get a snack some water a donut and please remember to use your new friend slido so you can pose questions to our speakers today and the first is Caltech one and uh and you can try it out okay so our first session indeed is the gut brain connection with Dr Sarkis

masmanian the Lewis B and Nelly Sue professor of microbiology and Heritage principle investigator Dr masmanian was born in Lebanon to two Armenian refugees his family moved to the U.S in 1973. he enrolled at UCLA in 1990 planning to major in English before Switching gears to graduate in microbiology earning his PhD in 2002. if a thereafter with the Harvard Medical School and University of Chicago masmanian won

a MacArthur genius grant for his work on the human microbiome the founder of two biotech companies and so much more masmanian has explored the fascinating relationships between humans bacteria and Myriad vaccines antibiotics and sanitary practices we put in place to control our environment or at least attempt to the question how does gut bacteria influence the development and function of the immune and nervous systems today Dr

mezmanian leads us on an examination of the gut brain connection welcome Dr sarkas masamanian uh thank you for that amazing introduction uh and the opportunity to speak here today at seminar day about our work I will tell you about a story that has taken well over a decade to evolve and you'll appreciate it went from an initial idea to what is now an active clinical trial

and so I'm going to just walk you through the basic science and how that's informed our hopes to improve human health so the title is the gut microbiome and autism spectrum disorder from Discovery to drug and so I'm a microbiologist by training I work specifically on bacteria and even more specifically on gut bacteria the many trillions of cells that live in and on our bodies most

of which are in our lower gastrointestinal tract so if we actually look at the numbers of bacteria inside our colon when again most of them reside about 100 trillion bacterial cells in us and what that means is that we are mostly bacteria on a cellular level there are more bacteria in in our bodies and there are human selves by some estimates about three to five times

as many bacteria and even more astounding are the number of genes right so the microbiome refers to the the actual genes that are that are harbored in these bacteria uh there are about 150 times as many Gene families or unique types of genes so we view our microbiome in our genome as a collaboration that come together to to affect our health and and uh in some

cases impact disease and I'll tell you about one of those examples and the microbiome has been referred to an alien or foreign organ because it has a metabolic capacity of the liver and the actual weight of the of the adult brain and so in a number of different clinical and pre-clinical settings a microgram has been linked to neurological metabolic and immunological conditions I'll just list them

here and this is really just a small sample of the types of different conditions and some cases diseases and disorders where the microbiome has been implicated most of this work is still early meaning that a lot of the work is still in pre-clinical models mostly Mouse models but other model systems as well and we are now as a community beginning to translate some of this work

and apply it to to clinical settings but I have to say there's a lot of excitement because the the data at least in the animal model shows that the microbiome is a critical contributor in some cases causal to the development of disease and perhaps we can get similar data in humans and give us some actionable items to improve human health and so the story I'll tell

you about today it revolves around autism spectrum disorder and some of the the behavioral symptoms that are associated with autism so our laboratory works on a number of different uh conditions again in animal models that span Parkinson's Alzheimer's disease and anxiety we started working on autism uh very early on actually just a few years after I joined Caltech and I'll tell you that story in a

minute and so autism spectrum disorder is defined by core behavioral traits that include deficits or reductions in social communication that includes verbal and non-verbal forms of communication as well as repetitive behavior and that's what's shown here in these core symptoms but there are many comorbidities or Associated behaviors and phenotypes associated with autism and those are listed here on the periphery and what caught our eye many

years ago were these gastrointestinal symptoms and again about 13 14 years ago there were just some initial reports anecdotal uh suggesting that there may be an increase in GI symptoms such as constipation diarrhea bloating in the autism population compared to that of the of a healthy control population as well as immune dysfunction and metabolic abnormalities which are associated as I just mentioned uh with the microbiome

so we thought perhaps there was a link between gut bacterium and even the gut and the development or at least the the progression of autism and of course there's a number of different Associated traits with autism and those are listed here and I'll just draw our attention to anxiety and to aggression because those will become important in a few minutes for the prevalence of autism is

rising there's a latest estimates from the CDC indicate that one in 36 births in the United States result in an Autism diagnosis and you can see the the really steep trajectory of the increase in in diagnosis of autism there's controversy in terms of what is driving this increase at least what we know is that increases in awareness and changes in diagnostic criteria have led to a

large portion of this increase but there are still aspects of of the rate of increased autism that do not remain explained and I will suggest that the environment plays some role in that and because our environments have been changing over the last several decades and perhaps that's contributing to autism but I also want to be cautious in mentioning that this is still a hypothesis despite a

lot of data from many labs around the world we still really don't know what are the environmental components or the environmental risks associated with autism but clearly this is a a growing social media medical and economic burden the current estimates are there's up to 75 million people worldwide with an Autism diagnosis and this is an estimate of of the actual prevalence of autism largely in the

developed world because it's quite difficult to diagnose autism there are no blood tests that are reliable there are no brain scans that are reliable for diagnosing autism it's this requires a trained behavioral individuals or people with behavioral training to to assess autism based on standard questionnaires and other diagnostic tools and so these are very hard to implement in developing countries and so one may wonder that

the actual incidence of autism may be greater than what we actually know as I've already mentioned the the rate of autism is increasing and this makes uh the you know the rapid rate of increase makes autism the fastest growing developmental disorder I've mentioned that economic burden already in a person's lifespan about the cost to to both society and individuals about two and a half million dollars

that's including both therapy and loss wages and opportunities and uh again there are these gastrointestinal issues associated with autism and we now know that the microbiome the collection of bacteria viruses fungi and other microorganisms in the intestines of individuals so autism look different than that of people who don't have an Autism diagnosis and so of course this is an association that there's a change what causes

that change what's driving that change we don't know and as I mentioned the implications of the change in autism on the the symptoms itself is something that we are we and many other people are studying and so uh Sandra mentioned you know one of the interesting aspects of Caltech and Tom mentioned how we get out of our comfort zones and and reach for the stars in

the research that we did and I became interested in this in the conversation with Paul Patterson some of you may know Paul was a neuroscientist here at Caltech he sadly passed away in 2014 but Paul and I began a collaboration as I mentioned very early on in my career here at Caltech and this is a neuroscientist Paul was interested in in how environment shapes behavior and

a microbiologists coming together and trying to afford new new ideas and and new uh discoveries so what Paul had had done when I first started working with him is develop a mouse model of autism that's now widely validated use all around the world based on epidemiologic data that an infection during pregnancy a very severe infection oftentimes associated with hospitalization leads to an increased rate of diagnosis

in the child when the individual is born and so Paul modeled this epidemiologic data by essentially giving flu to to pregnant mice and so that's depicted here and so when the mice had flu again during pregnancy they resolved the flu the fever goes away and then when The Offspring the pups are born to this pregnant Mouse those those animals have features of autism and those features

are changes in social behaviors changes in communication increased anxiety and changes in sensory issues as well as structural changes in the brain in terms of how what types of cells move to different regions of the brain as the brain is developing so this was very exciting to us to be able to have a model that we can test this hypothesis that perhaps the gut was involved

and again you know this was quite the hunch when we when we started working on this where a new model of autism was still being developed was the testing ground to understand is there an implication for the gut is there a a role for the microbiome in any aspect of autism in this particular Mouse model and I'll just cut to the chase is that when we

first started working on this you know one of my students went to Paul's laboratory they already had an ongoing experiment and uh Paul being a neuroscientist told me of course we throw away the intestines of these animals we're interested in the brain and so my student went harvested the intestines and on the first experiment we found some really fascinating results the microbiome was different between the

mice that had the behavioral issues and the mice that behaved quote unquote normal as well as changes in gut physiology the way the intestines were performing were different in these two groups of mice and so we had an idea that perhaps one of the ways to interrogate this this initial finding was to intervene at the microbial level and my laboratory at the time was working on

an organism called bactroyce fragilis which is found about half of the the human population so half of us here and online have bactrace fragilis in our colon and we had been studying this organism because it's of its uh quite a strong problem or anti-inflammatory effects meaning that it was able to suppress inflammation and the context in which we were interested in bacterase fragilis was to correct

issues in models of Crohn's Disease ulcerative colitis and other inflammatory conditions I'm not going to go through the details but we had worked out many many aspects of of the mechanism of this organism was the molecule it's making that mediates these anti-inflammatory effects what receptors does it bind to what the cell types that respond and how does It ultimately suppress inflammation and so we applied this

organism back Trace vigils to the mouse model of autism to this Mia maternal immune activation model and then we behaviorally tested these mice and to our surprise adding the probiotic adding this human commensal organism this beneficial bacteria to mice corrected many of their behavioral ngi symptoms so my time is short so I'm going to just give you a couple of examples of this prior working get

and then segue into some of our new discoveries so we tested anxiety in these animals so and anxiety is tested by an open field test where we place mice in this Arena and mice are timid shy and anxious animals they stay close to the walls you may have already noticed this in your in your backyard or other places you've seen a mouse because they just feel

safe and closer to the walls but they venture out into the center to look for food or mates or or just curious and then we measure the time that might spend in the center the more time they spend the more bold they are the less time they spend the more anxious they are so these are the animals in the third bars that were The Offspring of

the mice that were given the flu offspring of the mice that were given essentially an infection during pregnancy and you can see they enter the center less than the control animals in the first bar and then we measure the time they spend the center and they spend less time in the center compared to the control animals and when we gave them the probiotic back trace for

drugs you can see in the third and fourth parts that these animals now enter the center more often and spend more time in the center again compared to their wild type counterparts suggesting that we've overcome their anxiety phenotype by just giving this one organism it's the same with vocalization so mice vocalize through ultrasonic Communications and we can measure this using an electronic microphone and essentially even

see patterns of language in these animals and when we look at the Mia mice The Offspring of the Meister we're given flu you can see that these mice communicate less they the number of calls or decrease and the duration of their calls were decreased compared to these controlled animals again a trait that's corrected by bacteriosphagellosis the small organism so there are many other aspects of the

physiology of the Mi animals that we interrogated and understood in this initial study but we became interested in what is linking the gut in the brain how is this signal sent by bacteria from the intestines presumably to the brain to mediate these beneficial effects and so we looked at the immune system we found some interesting Pathways that were activated by the organism that suppressed inflammation but

then we became interested in the metabolic conditions that are associated with this disorder because as I mentioned there's metabolic changes in individuals of autism which we also observed in the animal model and so we did an experiment called untargeted metabolomics where we're just surveyed all the small molecules in the circulation of the different groups of mice and try to understand what is the chemical fingerprint what's

the signature of of the different molecules in the blood of these animals that may differentiate how they're behaving and how they're responding to the probiotic and so these are the control animals so these are mice just given saline or salt water not given the flu and then these are the animals that were given uh whose parents were given the or the mother was given the flu

and you can see there's an increase in this one molecule called four ethylphenyl sulfate or four EPs and so here's the molecule here and this was quite astounding to us so we looked at a number of different molecules that had many thousands of which found several dozen that would change and this was the molecule that was the most highly increased between the control animals and the

animals with the autism-like features and this metabolite was also decreased in those organ in those animals that were given bacteries for jealous it was also really interesting about this molecule here's the chemical structure for EPS is that it's chemically very similar to another molecule called paracrystal sulfate and why this was a really exciting Discovery for us was that Pericles sulfate had been shown to be increased

in the urine of children with autism so it was a urinary biomarker if you will for for autism and again we had discovered this molecule there's very very little known about it about the fourth femal sulfate there were just a couple of papers describing its synthesis since chemistry had not been associated with any biological phenotypes not found in any animals or humans at the time and

so again we thought maybe this is just more than a coincidence that the molecule that is most highly dysregulated in the animals that have or don't have behavioral issues is very structurally related to animals or radio structure related to a molecule that's increased in human autism so this is a mouse Discovery and so the next thing or one of the next things that we did was

to try and validate whether or not this this molecule has actually increased in individuals with autism in a human population again the metabolite 40ps had not been highly studied at all and so we were very fortunate to work with Paul Ashwood at UC Davis and Lester Fasano at Massachusetts General Hospital to obtain both fecal samples and what I'm showing you here plasma samples from a large

cohort of individuals with autism or typically developing controls and found that about a third of the autism population in this cohort expressed very very high levels of four ethylphenyl sulfate the molecule is detectable in every single sample we've done many studies in humans now subsequent to this and this has been validated in a number of cohorts now we all have this molecule but it's just elevated

in individuals with autism and if we look at those individuals with gastrointestinal symptoms there's about a nine-fold increase over people over the typically developing population and if we look at the autism population that don't have gastrointestinal symptoms it's a little less than five fold increase and so there's a correlation between GI symptoms gastrointestinal symptoms and this molecule so everything I've shown you today is or so

far on the talk is an association so we identified the molecule in mice it goes up and down based on a probiotic treatment it's elevated in populations of people with an Autism diagnosis so maybe it may may Mark individuals with autism might help us identify who they are but we will also want to understand is there a functional role for this molecule is it actually doing

something relevant to autism again whether it's behavioral gastrointestinal symptoms or otherwise and so the molecule it was it was not known how this molecule is actually produced but based on some additional experiments that we did we were convinced that the only way this metabolite for feral sulfate was synthesized was through bacteria we don't have the genes mice don't have the genes flies worms don't have the

genes to make four-ethylphenyl sulfate and so we work with Michael fischbach at Stanford to identify the bacterial genes that can turn tyrosine an amino acid that we that's found in our diet so all of us as we eat food ingest tyrosine and to three chemical conversions tyrosine is metabolized to Fort ethylphenol and so this is the bacterial reaction again three bacterial enzymes uh converter Center for

epiphenyl and then when fourth phenyl is absorbed in the body it is very rapidly sulfated in the liver to four ethylphenol sulfates so 4 EPS is the bioactive molecule and then we perform this chemical reaction because many of these phenolic molecules these molecules with a ring-like structure can be quite toxic and so we sulfate them so that we can excrete them in our urine and so

again Michael and his team were able to identify the enzymes bacterial enzymes that can produce for ethylphenol and then we engineered groups of bacteria that either produce for ethylphenol and that's a four EP positive group and another set of bacteria that will bioengineered not to produce for ethylphenol the same exact organisms identical in every way except the four EP positive bacteria contain these three genes and

then the 40p negative group bacteria do not contain the genes and then we colonize mice with these two groups of bacteria so we made now two different lines of mice with that in theory the only difference in the entire body entire entire chemical signature of these of these animals was this one molecule so of course it's an animal study it's reductionist and it's simplified in many

ways but it's also a very clean system it lets us get to the biology of this one molecule and so we showed that for ethophenol is producing the feces of the mice that were colonized with the four EP positive bacteria for EPS again once it's sulfate in the liver is found the urine and the blood of only the group that is producing the the metabolite in

the intestines and then very interestingly we showed that a small population of this molecule 40ps actually winds up in the brains of the animals and we think something similar is happening in people we haven't proven that yet but at least in these animal size this metabolite is getting into the brains of mice and then we uh of course we were just looking for for phenotype we're

trying to understand what this molecule does we had really no uh reason to believe that it affected any particular brain region or in different cell type because again it's a novel molecule so we performed an experiment where we essentially looked at which regions of the brain were more or less active between these two groups of mice relating those activity results to this one molecule and so

in collaboration with Daniel holschneider at USC we identified that there are several different brain regions that are more active in the four EP positive group compared to the four EP negative group and the regions that are more active are listed here and those are in red and there's several regions of the brain that are less active in the four EP positive group and when we saw

patterns like this we wondered whether or not there were changing connectivity changes in how regions of the brain were actually communicating with each other in fact we work on Mikhail Shapiro here at Caltech to do functional ultrasound experiment to actually show that indeed there were changes in in connectivity between different regions of the brain and why this was so important is that these changes in connectivity

have been associated with the autistic brain and so again we're modeling certain features of brain activity in the mouse in the mouse that's associated with autism and other neurodevelopmental disorders and then in a series of experiments again which I don't have time to tell you we narrowed down the activity of four APS to a particular cell type called oligodendrocytes so oligodendrocytes are cells that we and

mice have in our brains and these are cells that myelinate neurons and so neurons or nerve cells have long axons that's how they send signals over long distances and what myelin is is think of of the coating around the copper wire myelin coats the long axons of neurons and allows those electrical signals to be sent over long distances and what we showed is that the maturation

state of oligodendrocytes was dependent on this microbial molecule so has four EPS entered the brains the mice it Arrested or delayed the activation of oligodendrocytes and you can see here the four EP positive group have less mature oligodendrocytes in the four AP negative group and we do this by staining and several other methods as well this is quite profound that the actual function of this brain

cell was dependent on exposure to this microbial molecule and then we also showed that the reduction myelination reduction in algodendrocyte activation led to changes in myelination patterns in the brains of this and of these animals and again myelination changes are also associated with human autism so it felt like we were on the right track here but of course our main objective was to understand is this

one molecule capable of inducing behavioral symptoms through alterations and myelination and so we performed a battery of Behavioral tests and showing a test just for anxiety so this is the open field test that I've already introduced you to and you can see the mice that works Associated or exposed to four EP in their intestines that have four EPS in their brains are less bold they're much

more anxious because they don't enter the center as often as the control for EP negative group and there's no changes in distance move so this is an important control experiment because if the animals were sick if this molecule was actually making them ill then they would be less likely to enter the center but there's no changes in actual Locomotion we interpret this as the mice are

choosing not to enter the center of the Arena if they're exposed to 4ep and then we supported this with additional behavioral tests and I'm showing the elevated plus mates where we place mice on a platform that's elevated over the ground by several feet and so mice feel much safer in the closed arms of the elevated plus maze but they do go exploring into the open arms

and so we can measure the amount of time that the mic spend in the open arms and the Terminus the open arms and you can see that the exploratory Behavior or the boldness of the animals is decreased in the 4ep positive group compared to the four EP negative group so we support this with a number of different experiments and experimental paradigms to really convince ourselves that

indeed for APS was inducing changes in behavior in these mice and so just to summarize what I've told you is that there are changes in the gut and the gut microbiome of individuals with autism that we can recapitulate in an animal model and there's several other animal models that show the same phenotype and when the microbiome changes the molecules that they're producing change as well so

they're metabolism that chemical repertoire of bacterial molecules is different when a mouse has features of autism and a person is diagnosed with autism and that leads to changes in the flux of molecules across the intestinal epithelium into the circulation in the case of four ethylphenol it's sulfated in the liver concentrations of four ethylphenol foreign buildup in the circulation and actually enter the brain and what this

molecule does is it arrests oligodendrocyte maturation maturation of those cells that I told you about leads to reduced myelination in several regions of the brain and altered behaviors and we think for APS is just the tip of the iceberg there are many many molecules that have been identified in humans that are different between a person with a diagnosis and without a diagnosis of autism and many

of those molecules share chemical and physical structure and structural similarities to uh for ethanol sulfate and so again we've intensively studied this one molecule but there are many many other molecules that we feel either will work with fluorothanol sulfate or may have different and various different phenotypes that they may induce on their own and so I'd like to summarize just this basic research that we've done

first using a bacteria and then learning from that bacteria to identify a particular molecule that's mediating these behavioral effects and so there were changes in the gut I didn't show you all this data but in that Mia model that I had mentioned there's a syndrome called leaky gut so these are the epithelial cells that line our intestines that keep everything out that it may be toxic

but let things in such as nutrients that we can that we ingest in our diet and when there's leaky gut meaning when the the seal between these epithelial cells is broken then more of these molecules can get into the circulation build up and get into the brain again this is what we saw in the Mia model and then when we gave bacteraseophagus it resealed the epithelium

and then reduced the number of these toxic or other amount of these toxic molecules that get into the circulation so we're developing bactrace fragilis as a probiotic and that work is is well underway but in the meantime there was we thought there was a faster path to perhaps intervening in this chemical communication between the gut and the brain and so we identified a particular small molecule

drug that binds these uh uh these metabolites that binds for ethylphenyl sulfate and I'll introduce you to the drug in a second but our hypothesis was again that we've learned about this the chemical transfer between the gut and the Brain through these basic research studies through using a probiotic but now that gave us the opportunity to intervene with a drug that's easier to develop pharmaceutically and

so the drug is called AST 120 it's not approved in in the United States but it's used in uh several Asian countries in chronic kidney disease what's really interesting is for ethanol sulfate and similar phenolic molecules are elevated in people with chronic kidney disease because their kidneys are failing again these are urinary metabolites that we excrete and when the kidneys fail they build up and asd-120

is given to these people to help with the build up of these toxic molecules and often the consequences of chronic kidney disease could be neurological conditions such as anxiety and brain fog and so what AST 120 is is it's a form of activated charcoal but it's a proprietary form that is specially engineered to bind this class of molecules and what's also interesting is that the drug

itself is not absorbed it does not enter into the bodies of mice or of humans and so as it's passing through the gastrointestinal tract it can take an orally it's binding these these small molecule toxins and then as it finds them it's uh and as the drug is excreted essentially lowers the concentration in the intestines and lowers the concentration in the circulation and we showed this

in mice and we actually show that the drug can overcome the behavioral effects of fourth fetal sulfate so this is the open field test in the elevated plus maze that I've already introduced to you and these two first bars are the data that I've already shown you these are the animals exposed to four ethylphenol and these are the mice that were not exposed to this microbe

metabolite and you can see the mice are more anxious in the for in a open field test as well as the elevated plus maze but what's quite interesting is that when we give these animals a drug even though the bacteria and their intestines are making for ethylphenol the mice overcome their anxiety in both of these tests and overcome many of the other behavioral conditions that these

mice have so again we're binding the molecule of these toxic molecules with a specific drug and this study enabled us to go into a clinical cohort to test some of these basic research findings in people and so uh I started a company in 2016 around this technology called axotherapeutics and axial's role is to essentially take the basic discoveries from our laboratory and translate them into people

and so the really talented team at axial uh devised the clinical trial in Australia and New Zealand the demographics are here 26 individuals all adolescents that's what the their regulatory board allowed us to do they wouldn't allow us to go into the younger ages initially and you can see that most of the individuals who were enrolled in this trial had severe autism and then compliance was

was terrific over 95 percent of of the individuals took their Medicaid medication meaning that over 95 of the doses that were given were actually taken by people this is the the trial design and so there was a four-week evaluation where the enrollment criteria was a prior diagnosis of autism as well as gastrointestinal symptoms but we retested these individuals with again standard assessment tools and then took

samples to look at their microbiome and their metabolism if they met the criteria about autism diagnosis with gastrointestinal manifestations uh based on our assessment they were enrolled into the clinical trial where they were on drugs for two weeks at a low dose two weeks at a medium dose and then four weeks at a high dose so it's an eight week trial and then we took samples

from the beginning of the trial so just remember visit two is baseline or the beginning of the trial and then visit five is the end of the trial and then we also asked them to come back after four weeks and then retested the individuals and so again I'm going to show you data from people as they enter the trial and at the end of the trial

and so the the safety profile was Exquisite there were really no safety uh issues and this is what we expected because again the drug had been given to people but never to this patient population had never been tried in individuals with autism um and so met the safety criteria so that was one of the primary endpoints of the trial and when the other primary endpoints was

Target engagement and what that means is did we see a reduction in some molecular or cellular phenotype associated with the pathway that we were studying and so we met that criteria as well so Target engagement in our study meant lowering these microbial metabolites based on the research that I've shared with you this is again fourth phenosulfate as I mentioned there are a number of different uh

molecules associated with for the federal sulfate in terms of structure and all of these are produced by bacteria and you can see in all the individuals in the trial between after eight weeks on drug that the levels of these metabolites were greatly decreased in their circulation this is plasma data this is just a control molecule that does not bind AST 120 does not bind the drug

and you can see it's not decreased so quite gratifying that we met both of the primary endpoints of the trial the trial again it's a very small trial of 26 individuals and I should also mention that it was an open label trial meaning that everyone knew that they were on drugs so we're not controlling for the placebo effect which is very very critical in being able

to make uh claims about efficacy and so even with this small population we did see improvements in two different behaviors the first is anxiety as measured by the Pediatric anxiety rating scale so this is the entire cohort of 26 individuals at Baseline and at the end of trial and you can see there's a decrease in their anxiety scores but this didn't reach significance and the reason

why is because everyone who entered the trial doesn't have anxiety about 50 percent of the autism population can be is diagnosed with anxiety based on the pars so when we take those individuals which who had clinically meaningful anxiety in the pause a score of 10 and above means that there's anxiety you can see that enriched on this population that most of those individuals in eight weeks

reduced their levels of anxiety and this is the same with irritability so we use the aberrant behavioral checklist using the irritability subscale to look at the entire population and here across all individuals we did see significant decrease in irritability or aggression in the pars a score of 15 is considered oh sorry the ABC of score 15 is considered meaning clinically meaningful in terms of irritability and

you can see the majority of the individuals that had irritability going into the trial improved after eight weeks on drug so again we're quite pleased and excited about these results but still very early days we still need to control for for that placebo effect that I mentioned which is a very big confounder especially Neuropsychiatric conditions and and that's why we've now started a phase two trial

which will have both a placebo control and has a multiple Doses and what's really gratifying is that just a couple of weeks ago we were able to get FDA approval to go into younger populations so now this trial is enrolling individuals at five uh five years of age and above we have 27 sites total 24 in the US three in Australia and New Zealand and uh

you can actually find the trial at the autism study.com surprising that that domain name was not taken it's called a tapestry study and again it's enrolling now we have actually two sites in Los Angeles for those that are interested but again sites all across the country the study will read out in the early phases probably the first quarter of 2024 and so we'll have then a

pretty good idea of whether or not this drug is working in individuals with autism because we have that very very important Placebo control and I'll just leave you with this final concept that what we've learned over the years with this and other research in the laboratory again all based on uh very collaborative efforts both here in Caltech and and with institutions across the world is that

maybe this research is telling us that one can think about drugging the gut instead of the brain for various different conditions including Neuropsychiatric conditions such as autism and so the current strategy to address Neuropsychiatric neurodegenerative neurodegenerative and other neurological conditions is to get drugs into the brain which means that those drugs have to cross the blood-brain barrier this is very challenging to get enough of the

drug into the right parts of the brain but what it seems that has happened is that bacteria have figured this out they figured out exactly how to get molecules into the brain from our intestines but what the research has told us is that maybe we can intervene at the level of the gut and in those cases it's going to be you know it's not going to

be in all conditions but in those cases where the intestine and molecules from the intestine have a role in behavioral other neurological outcomes perhaps we can just deliver drugs to the gut and break this cycle break this path between the gut and the brain and obviously getting drugs into the gut is far easier than getting drugs into the brain and much less likely to cause side

effects or other deleterious outcomes so again we're very very excited about the concepts not just the data and I'll just conclude by thanking the people who did the work the majority of the work that I just mentioned in the newer studies were performed by Brittany Needham who was a postdoc in our lab and now has our own laboratory at Indiana University these are some of the

other members of the lab that directly contributed to This research but our lab is very collaborative everyone works together in in trying to make some of these breakthroughs these are the the funding sources that helped support this work and these are the many many collaborators again both here in Caltech and elsewhere that really enable us to do these Cutting Edge research and of course a lot

of thanks to the families and individuals with autism both in the first phase trial as well as those that are entering the trial now thank you so much and I'm happy to take questions [Applause] thank you so much um because we went a little long with intro we're going to do five minutes of questions this will will not end right at 8 40 9 45 so

if you need to use the restroom you can we'll start the next session at 10 but we're going to do some questions and thank you for these great questions um Edward and kutulenk ask do you feel and there's a couple of questions that are ganged around the environmental issues do you feel that the rise of microplastics being introduced into our food chain this is our digestive

system could be a contributing factor to the rise of autism and there's a lot of questions about if you recommend people taking probiotic products whether it's microplastics or any other environmental exposure we simply don't know right and there's just simply not enough data to understand what it is about the environments that may be changing uh the incidence and or the diagnosis rates of autism and again

I want to be clear that the the role the environment in autism Still Remains unproven and controversial it's research like this and I want to say there's many other researchers around the world who are identifying both microbial and non-microbial meaning environmental contributors to autism largely in animal models but some translation now in people so again whether it's pollutants or other toxins food preservatives you know antibiotics

you know and certainly not vaccinations but other chemicals that were exposed to the research just simply isn't there yet to understand what are the environmental contributors to autism PK and many others were interested in what is the mean rate age and age range of autism diagnosis in the U.S and do you have a sense of how this has evolved over the past couple of decades the

diagnosis a the mean diagnosis age is about two and a half now in in the US and that is decreased over the years it was you know four or five years of age about a decade ago but again I'm not that doesn't it's not attributed to any changes in the diagnostic rates but just the fact that Physicians are is more aware of autism and they were

a decade ago and and prior to that and Physicians are just looking out for it right because it's not it's in many cases it could be quite subtle in some cases it could be very obvious and so the the decrease in the in the mean rate of diagnosis is just because Society is more aware of this condition and Jim Turman and others are interested should we

expect a spike in autism cases from children who gestated during covet 19. no it's a it's a great question and those studies are underway obviously the you know Society provided a great experimental uh setting a clinical uh trial if you will in society that um many many people are now following and so they're following those those pregnancies during covid those individuals who may have gotten coveted

during pregnancy their you know how severe their their inflammation and fevers were and then now tracking The Offspring to see what are the rates of diagnosis it's going to take several years certainly to get that data but many many groups around the world are tracking this or very clever Caltech Community is interested in of course the matter of I'm going to gang these questions soon with

an increase in monitoring genetic content of human waste streams so it's possible to look for beef fragilis in the human waste stream across time and biography and geography and um Can for EPS for be detected in Wastewater um I don't know certainly we can detect bacteries for drugs we can detect any organism in Wastewater but again but the molecule I don't know but again this the

organism nor the molecule is new to humans it's likely been with us for many millions of years why high levels have evolutionary resulted in these changes in Behavior I think maybe that's a different question and that's not certain I mean is this an evolutionary accident or did somehow bacteria intend to cause these these effects and what that you know the evolutionary forces that drove these interactions

were again I'm afraid to say are just not known what you're at Caltech you should know all right uh Gary Turner and some others are interested if only about one-third of people with Autism have access for EPS why were the clinical results so good um because we're we're the molecule itself sequestions the drug itself sequences many many different molecules and so there are likely other molecules

that are binding AST 120 binding the drug that are similar to for EPS but may have different uh properties and those just haven't been studied yet and so there's the best of our knowledge about 13 to 15 of these molecules that are related to four APS that the drug binds and so it's likely that some people you know you know again assuming that the model and

the hypothesis correct that some people's symptoms may be driven by four EPS on people's symptoms may be driven by other molecules but the drug binds them all and last question I think ganging together a couple of other medical conditions any potential related to Parkinson's treatments or any connections of gut towels to alzheimer's we've started now looking for these molecules and Parkinson's patients and uh and uh

ended Alzheimer's and they do seem to appear the effect isn't as high as they are in Neuropsychiatric conditions so I can tell you these metabolites appear to be elevated in schizophrenia and PTSD and so there may be differences in what types of neurological conditions that these molecules are associated with but if I were to venture a guess I would say that the pathways that by which

a microbiome impacts neurodegenerative conditions are likely going to be different than the pathways by which it's influencing your psychiatric conditions very much Dr Sarkis masmanian the questions fantastic we'll take a quick break start the next uh session right at 10 are quantum future see you in a moment thank you