CRISPR Cas9: Will it Cure Aging? — Talk by Oliver Medvedik at D.N.A. Conference

hello everybody so my name is Oliver medve deck I'm uh right now currently I'm a professor at a small University in New York City Cooper Union so I work in the bioengineering department there and before that I was a co-founder of gen space which is a community biotech lab and along with Keith kameido here we then co-founded lifespan dot IO so my interest in aging goes back kind of a long way so I did my PhD with dr. David Sinclair at Harvard University and I've been sort of out of the aging field for a while I've been out on a periphery observing this but nevertheless this is something that drew me to science initially because it's such a fundamental thing that can be answered finally with the tools that we have at our disposal so I'm going to be talking about some of these tools that we have I'm going to try to ground this in tools that we have available now and also just going out of a little bit out of the context of the talk as Damian said I'll talk a little bit about gene therapy but mainly I'm going to talk about some of the tools that are used in gene therapy such as CRISPR cast 9 and basically what it is and how it works so I hope this doesn't devolve into a total lecture because I give those a lot University but I'm also going to be talking about some of the technologies that are outside of gene therapy that are currently being utilized to affect positively lifespan and where I think this is going and what are going to be some of the early tools that we'll see therapies coming about so this is sort of the coolest picture I could find of CRISPR and cast line in action on the internet and I guess everybody's asking CRISPR will it cure aging I mean the answer to that for me is maybe right so yes or no maybe it's a tool it's got its limitations and I think it has its purpose in in this but we've got to basically ground ourselves in its limitations so I'm going to just kind of quickly overview and discuss how we're going to use it so there's been a lot of you know talk about controversies whether it's going to destroy us to save us certainly the scientific magazine science nature are not above this type of whipping up a frenzy about CRISPR and caste 9 seek-and-destroy targeted destruction you can go online you can get these two fools these plasmids sonic companies such as ad gene for easy genome editing so these tools can be used at community biotech labs like gen space which I work at and I teach a CRISPR class there so basically people come in from the public and they learn how to use these technologies and they actually apply them to engineering genes in an organism in our case yeast so just to run through giving everybody a background because I'm going to assume here you don't some of you or most of you don't know exactly how this tool works those of you who do or experts please bear with me but I'm just going to give you a very fast overview over my view of biotechnology so biotech basically started in 1970s and we've been using these tools called restriction endonucleases which chop DNA up but they do it in in vivo that's one of the problems limitations we still use them but we can't really use them in organisms and we can only use it in discreet location so we have to use this in test-tube so we have to use it to chop up DNA splice it put it back into organism so very slow been discovered in the 70s but we still work with it in late 80s site specific recombination technologies came into being and these are technologies that use enzymes that a little bit more sophisticated than the restriction enzymes they don't just cut DNA but they can cut DNA and essentially loop pieces out so if you make organisms that have something integrated in one location that are flanked by specific sequences and you have another organism another Mouse that's making this enzyme if you cross these mice you can have this location basically modified so these are basically used in the field of transgenics in transgenic mice so you could use this technology to see what happens if you switch a certain gene on or off in different cells so it's a very powerful tool but still there's a limitation this cutting only happens in one location so with that in 1990s to now the development has been using programmable endonucleases so just like the restriction enzymes these things cut so an indoor nucleus is an enzyme that just cuts DNA so everything we've been talking about is just cutting DNA just cutting in its specific locations and these zinc finger nucleases and talents which stands for transcription activator like endonucleases are basically what are referred to in the field as fusion proteins so they're not naturally found unlike restriction enzymes but in this case you take a one enzyme and you hook it up to modules that recognize different parts of the genome so you can custom design these to cut wherever you want in a genome so this is great the only issue is that you know these are patented there's companies that use them but even if there weren't patented it's still really tricky to do protein design so you have to design one it takes weeks months and then apply it so that's where CRISPR comes in so CRISPR really simplified the cutting of DNA so really all biotechnology most of its been cutting DNA at a particular location and that's all CRISPR does it just cuts crew cuts at one location very well very specifically with almost zero percent off target cutting at this point so what's CRISPR well CRISPR stands for clustered regularly interspaced palindromic repeats that's a long name and it's basically a protein that interacts with an RNA and RNA is another little molecule that's like DNA and this complex then homes in on different locations and the cool thing about CRISPR is the part that does the homing is the RNA well you're probably asking so what why not just make it a protein and go there well the really cool thing is RNA and snoop laic acids are really easy to work with any lab can basically manufacture these nucleic acids very readily or outsource them to companies that will do that for you for a few bucks so this is that's really what let's why CRISPR is exploding is because everybody can custom design and a nucleus is now to cut wherever they want really simply and really cheaply so what CRISPR is and just like the other enzymes I talked about this all came from nature so every tool that we use is basically coming from nature and we repurpose it just like every other tool so if we look at the phylogenetic tree of life the three major domains bacteria archaea Eukarya these two used to be clumped together but now they've found to be genetically distinct the CRISPR cache lines system is found in about 40 percent of bacteria 90% of archaea not I don't think anybody's discovered any in Eukarya which is us like animals and plants so what is CRISPR do in the natural world well this is to me even cooler than using it as a tool this is basically comes through basic science it's a bacterial acquired immune system so until 2010 2009 if you were to ask somebody does any organism that's beyond an advanced organism like a lizard or a bird or a human have an acquired immune system the answer would be no the open up any biological textbook that came out you know like me the last edition of molecular cell biology or two editions ago this is not going to be in it right there you're just going to say flat out in the immuno immunology section it doesn't they won't even say that bacteria don't have an immune system because there's nothing to say everybody knows that it doesn't have an immune system but it does and the cool thing about this is that essentially the way this immune system works is it has a record of the DNA of viruses that infect the bacteria and they put it into the genome of the organism in this case the bacteria and this array basically is like a database a viral database of viruses that recently infected the bacteria and these little bits of DNA are converted to rnas these things called guide rnas which then when they bind to the kass protein which is made by the bacteria this guide RNA specifies where this cast is going to go and do the cutting in this case to the viral genome and like viral databases on your laptop this is also continuously updated so the old sequences come off the new sequences come in because the bacteria only has a limited space to push all this information so this stuff has been engineered by an a through billions of years ago right so this this is the the original antiviral software literally so that's not even analogy so I'm not going to go through all of this but this is just a timeline of how long this has been in development and from the 1980s to 90s I'm just going to say that these weird sequences were found using high-throughput genomic sequencing when they compared bacterial sequences they notice this CRISPR think popping up over and over and over and again and it took a while before somebody said AHA and this was this fellow Mojica and his lab and they suggested it might be a bacteriological immune system and then later laboratories around 2010 our pen TA and Doudna and jangula they basically started to use this technology as a tool you basically making a tool out of it and this is where really we've been seeing the explosion of where everybody's been talking about CRISPR here they've been talking about it as an application rather than the original you know really cool discovery and what it does in nature so just briefly you've got the protein you've got this RNA they come together to form a complex they find a home in on a piece of DNA and these blobs of the protein basically do the cutting so this little X here X here are two cuts on the DNA and it basically just makes it cut through DNA that's it very precisely and since then people have been making all sorts of variants of caste 9 different caste species I'm just using that term loosely different proteins so not only there are there different naturally-occurring caste lines but there's also modified ones that recognise things called proto space or adjacent motifs I just said it now forget it because I'm not going to refer to it now again but suffice it to say you have all these variants and how is caste 9 to be used in gene therapy well when you put a break in DNA that's a bad thing generally if you do it in cells randomly in your body that's associated with mutations some places you want them to happen like the immune system but most places you don't so the cell wants to repair it really quickly and the way it does so is two mechanisms one I'm not going to go over it's called nhej a non homology and joining that's like a quick and dirty way to repair but it makes errors another way is homology directed repair and this uses a template to copy off of and basically fix itself so it makes a precise repair and this is the system that you can subvert to try to sneak in something of your interest so how would this be used in in anti-aging or aging therapeutic context well this is just one paper many meta-analysis of genetic variants associated the human exceptional longevity so people are because of high-throughput sequencing everybody's looking for what's the difference between a super centenarian and the rest of us so all of these different changes basically so some people have more efficient enzymes and so on and so forth so recently Christopher cast line was used as a genome editing tool for correcting this these dystrophin mutations and they used a technique of CRISPR called multiplexing where you can express many of these guide RNAs and do several different chain plots in this case about twenty changes all at once to fix a genetic locus so it's easy for me to envision that you can use this technology to modify stem cells and improve them and before you put them back into the patient it would be probably a lot of multiplexing involved depending on how many locations were looking at so that's that's kind of like the the I guess the fundamental aspect of gene therapy and genetic engineering but there's a lighter side to it and that you've all heard of the genome and some of you probably heard of the epigenome so genetic languages has many hierarchies and DNA is not just found in a long string but it's wound up in these three-dimensional hierarchical organizations and in order for things to be read they have to be unwound right and rewound so you have this three dimensionality that has to be traversed by all the enzymes and how genes are accessed depends on how it's packed so there are certain chemical modifications that are put onto the genome either on these little look like meatballs here but these meatballs are histones so if you put certain chemical groups on these these nucleosomes they can an unwind the DNA or cause it to compact likewise you can put modifications on DNA this is chemical language that we are only just beginning to understand right now we know like four or five different modifications but I spoke to Chris Mason and works at Cornell and he studies this and he said we found up to 50 different chemical groups and we don't know what most of them do so that just gives you a taste of how new this stuff is so I call this a wee bit of Lamarr because you've seen a few slides later that these epigenetic modifications are influenced environmentally yet they are inherited so one type of modification that you'll hear a lot of when you read the literature are these what are called CPG Islands these are just seas and G's which are just the two letters of the four-letter alphabet of the DNA and you can have these little chemical groups called methyls attached to particular nucleotides and where you have a lot of methyl groups the DNA is enclosed up and you can't be read or if it's unmethylated it's opened up so the methylation status determines how a cell reads its DNA it's like the file cabinet is open or file cabinet is closed and that's how all the cells your body that's how they're differentiated for the most part you all have for the most part the same genome but because different regions have different methylation patterns and lots of other epigenetic patterns the initial cell the zygote as it develops through through a development will develop into the 200-plus different cell types or somatic adult stem cells throughout your body to make the different tissues right so this is all through epigenetic modifications and for the most part these changes you want to want them to stay the same throughout your life you don't want them to kind of change around right you don't want your neuron to turn into a - a muscle cell or you know you won't you want it to kind of stay wrap the stay around in the way it is but unfortunately these epigenetic modifications could also change or mutate so just like mutations in DNA which is sort of in its in the in the hard code you can get changes in the epigenetic status and genes are turned on that shouldn't be turned on right so and that's that's been seen in aging so this is just a slide from paper which shows that these epigenetic changes can also be hereditarily passed on so certain environmental factors like diet can affect the passing on of methylation patterns to offspring so that's why I called it the Lamarck light so those of you who read about Lamarckian ism yes the environment can potentially you know pass on changes in a certain context in this case so these epigenetic changes these when you look at different regions of the epigenome they vary they start to you know you start to get the mesh relation patterns changes somebody gets older and this can correlate with healthier cells or younger cells and you can actually estimate you know how healthy or young a cell is by looking at some of these epigenetic modifications the way it connects the CRISPR is that CRISPR isn't just used for cutting it's just really such a flexible tool that you can do something you can basically all CRISPR does I just mentioned it cuts but cuts it in specific locations and at RNA takes it to that location but what if the CRISPR didn't cut at that location well the RNA would still bring the CRISPR I mean the Cassadine protein over it just wouldn't be functional this one is just called D cast nine d stands for dead cast nine that just mutated the endonuclease port part so it goes there and it does nothing but if you design it to attach to a different protein in this case they used a methyl transferase you can home in this CRISPR two different locations and have it alter the epigenome at different sites so you can basically stick on whatever enzyme you want you can have it a satellite a site or deacetylated site or whatever so now you have this tool kit with CRISPR cast nine can't go in go to sites and cut them and with the right template you can change either the hard genetic code or the slightly I'll call it softer genetic code of the epigenome and modify it so we have this now tool at our disposal to do those changes but there's probably gonna be a lot of changes and and one critical problem is accepts accessibility getting it to where it needs to go in all these locations so that's where we're at with basically with CRISPR and it's the story still evolving it's still basically more variants are being built more applications are being designed and with this I want to take you now just to move on to a different approach that's being applied a little earlier to treatment of aging like effects you know cells that don't don't perform metabolically the way they should perform and I'm just going to throw out a term here that I sort of invented I'll call it augmentive a softer touch we all take supplements or some of us take supplements vitamins when you say supplement to me it sounds like a dietary supplement like you're taking something you're lacking in your in your diet I just said well let me call these augmentis because this isn't really you don't need them in your diet but your body is depleted of them as it grows older cells that need to make certain things don't make them anymore right some small molecules can enter into the cells and basically boost the activity of enzymes that are now faltering so you have small molecules like resveratrol pterostilbene nicotinamide right beside rapamycin a lot of you have heard of these molecules that affect core and sirtuins there's also biologics which are larger things like proteins DDF 11 para bios's experiments this is a factor that influences muscle growth aisle 33 it's been some really impressive studies done with mice showing the reversal of Alzheimer's in an Alzheimer's mouse model so beta amyloid plaques were being cleared using this this il 33 which also drops down and decreases with patients that have Alzheimer's so company out there Elysium which was founded by Lenny gurantee who's a former P I of David Sinclair you can get nicotinamide ribose ID and pterostilbene now at the market and what they do is basically they affect 31 which is an enzyme that basically accelerates the deacetylation of those meatballs the histones and that affects the epigenetic status of certain genes so here we have now a chemical modulation of the epigenetic status and that's already at the market still you know being worked on as for that as far as the biologics are concerned gdf 11 this result as I mentioned came from a para by OSA study by Amy wagers at Harvard and this is where they this is to me Frankenstein in science at its finest two mice were basically had their vascular systems connected and old mouse and a young mouse so you had the blood rejuvenating another Mouse right so sort of a countess Bathory type of thing taking place here the question is well what's going on here right I mean what's what is rejuvenating what and one such molecule was discovered gdf 11 which is a protein that basically declines with age and is required for muscle stem cells to differentiate another class of molecules that are therapeutics and this is a class of its own and this is really something really new that just came out is primarily work done at the Campisi lab arsenal it expand these are really interesting molecules a lot of them have been repurposed from cancer drugs so drugs that basically cause cells to die if you're apoptosis technically if they're if they're expressing the wrong proteins and these things are found to kill off senescence cells and when cells basically proliferate in the body right so they'll divide divide divide they'll eventually some of them will wear out they'll senesce they'll stop dividing and they'll not immediately apoptosis but they'll sort of linger and release these inflammatory chemicals and basically wreck the neighborhood of the other cells so these thus analytics target the cysts the pathways that are on in these senescence cells and they basically oblate them right so it basically gets rid of these negative stimuli and this is a really cool technology because it's probably going to be very important you know people talk about stem cell replacements getting stem cells out processing them basically such as using maybe CRISPR cast nine to modify them then reinvent is bad these stem cells are going to malfunction so having Senna lytx on hand will potentially go hand in hand with this type of treatment so that's where I see you know this is sort of all the most recent technologies and the ones that are are to me the most well-developed there's going to be there a lot more that I haven't talked about that some of the speakers will but I want to sort of project now a little bit as to how I see these working so this is a statistical history of human lifespan so this is our normal lifespan is as a Gompertz curve this is a hundred percent of things or humans are alive here this is our maximal age 125 give or take and this is our average lifespan in the wild this is when we're taking care of my mother nature we die off at 35 so thanks mom so when we have civilization we have improved safety and diet so no predation taking place we ship this the average lifespan shifts so people still died at a you know people still lived long this is just an average right so people in ancient Greece still could go up to a hundred hundred twenty-five but most of them didn't because most of them got you know shot by an arrow and got an infection there's no antibiotic so the average drops so we have safer environment better diet and this has been shifting continuously medications like antibiotics but as you can see here this maximal isn't moving you're basically pushing this and there's a limit to how far you can push this until you get to this point and for most people this is enough so I can imagine lifespans if you put use these augmentive Xand sena lytx you'll maximize all the cells that you have basically make them as healthy as efficient they are until the stem cells are mostly depleted and you basically drop dead because of organ failure maybe that's the best way to go but the question is you know can we do this can we basically just take this curve and just keep going and there's no physical reason why we can't so I can imagine certainly if we have stem cell replacements and certainly stem cell replacements that have been modified using let's say it may be a CRISPR system where you have stem cells that are have been improved to be the best stem cells representative of perhaps somebody who has stem cells of a super centenarian and you start taking some sort of augmentive x' and you infuse them back you can potentially just keep doing that and just have that basically go on and on and on so that's that's my speculation there so there are challenges obviously to overcome what should we be doing to kind of keep the ball rolling in this direction so what I think is that we definitely need to identify and better characterize adult stem cells so we have over 200 plus different cell types what is it about a stem cell that makes a stem cell right so right now it's just a characterization you just have a parts list if it does all these things then it's a stem cell right we don't have anything there's no mathematical proof of a stem cell right you just have this list so that's the best we can do so if we can characterize which are the stem cells those are the ones we could target for therapy improved vectors for more effective gene therapy delivery so I've been just talking about the therapy itself but how do you get the therapy there people use modified viruses and they have limitations such as how much information you can pack into them so we have to improve vectors for how we get this stuff delivered identification of epigenetic states correlated with aging that's ongoing we have to compare one population to another population so what are all those epigenetic differences and what are they doing and of course identification of alleles or different types of point mutations or changes in the DNA that correlate with increased longevity so we can basically then apply that to a gene therapeutic approach and of course more and better screens to identify you were in better augmenta Vince analytic candidates and one such screen were basically funding or have funded lifespan Dario successfully and more and better clinical trials and I'm going to put this in parenthesis open source to characterize additional augmentive x' because I think for we're you know most of us here are impatient and you know at least in the United States you've got the FDA and if we do think the typical way every single one of these compounds is going to cost about a half a billion dollars and take ten years to develop so we'll be long dead before we can you know rejuvenate ourselves and that's clearly not workable so things are moving in that direction I think some of you have heard about the Apple research kit which basically allows people to sort of outsource a clinical type trial and basically become both the experimenter and the data point itself right so and then that data can then be accessed by everybody where else working in the field that can potentially accelerate discovery of such compounds so to sum up more basic research is definitely needed in my opinion we still don't have a solid theoretical understanding of aging and longevity we do know enough to start implementing a lot of these a lot of the results that we have so with that thank you and I'll take any questions yeah okay so the question is that I know that now there is also one thing CPF one that is quite popular and some people think that it may be even better than CRISPR maybe you can explain in what cases CPP CPF one is more promising and when in what case is not and well basically your opinion about this yeah so I believe so I so we're going a little bit in the rabbit hole with with with with Kris Pearn Cassadine CPF one is from it so that's not Cassadine is from a s pyogenes is this one type of bacteria that they then it's been isolated but lots of like I mentioned lots of other bacteria have different types of CRISPR systems and CPF one is one of them and I believe one advantages that it's much smaller and it's more so if it's smaller you can pack it in better into a in a vector so like let's say if you use a virus like an adenovirus there's a limit to how much stuff you can pack in so to deliver to a cell and one thing that people are trying to overcome is trying to get as much of the caste line information loaded into the genome as possible so CPF one is I believe even smaller so it's more efficient in that in that way so yeah there might be other advantages but that's that's the one I know of do you have any evidence or you know anything about the acceleration of the declination of senescent cells in people with either an autoimmune disease in a very unhealthy lifestyle and things like that well yeah certainly if if there's a lot of stress to an organism senescent cells will accumulate accumulate I believe if you take certain toxins like chemotherapeutics for example Keith and I were recently or a while ago at a conference which basically tried to really kind of brought into horizons of what it's mean means but to age right so if you look at a lot of diseases in some way you can consider things that you don't normally consider to be aging to maybe fall under that umbrella by having cells sort of hyper senesce until you deplete them like being infected with the HIV basically your your immune system has to keep replicating more helper T cells over and over over again and there's a limitation to how many times these somatic stem cells can replicate and that's something that I don't think anybody really knows what what are the exact barriers to overcome in that continual replication of the somatic stem cells but suffice it to say they they run out and then that organ that part of the organ is gone depleted right so different parts of your different organs could actually senesce or get old at different rates as a result depending on where the damage accumulates in at what rate does that answer the question partly right thank you my question would be um is it possible to look at a person's genome a young person and create basically a backup of the data and then as a person ages and genome degrades that you just like put the old stuff in there as a backup with CRISPR or a similar technology right now that sounds like an excellent idea actually I don't know if anybody's doing that that second Stitz it's like the data equivalent of having your umbilical stem cells frozen right so then you can't rent in this case you're just you're just saving the epigenetic data rather than the stem cells themselves that's true but perhaps we can develop them in vitro and then see how what changes happen if you have a if you have basically like a the human on the chip model where you can do induced pluripotent stem cells and artificially make them into neurons then see hopefully the epigenetic genome of that will be correct hopefully that hasn't done yet cause I don't really need a good thing all right first do you bridge major semester not a good business model was somebody who had his sons skin to his port blood stored frozen some 12 years ago could you take that cord blood and make the kind of the kind of Avila Damien was just saying in other are they people out there right now the advantage of that store portlet I would say sure hey Keith not really a question but I want to kind of piggyback on that because actually Oliver I have talked about you can actually potentially go above and beyond this at some point if we get much more sophisticated information with companies like a human longevity they're trying to find out the DNA that maps to super nintendo centenarians and stuff you might be able to at some point take someone who's already old take their cell you know sequence it and with mathematics sort of come up with the better version of that person sequence make those cells then give them a stem cell infusion with that so that's down the road but like I would love to see like a stem cell bank like that down the road and we have sort of talked to sloan-kettering a little bit about that I did down the road any other questions so there are some of these augmentive molecules that are that are available right now that are known to protect stem cell populations I think mostly food and the oxidant mechanisms and then anti-inflammatory mechanisms and then what you see is that the organism gets more differentiating stem cells out of it that would that mean you just said that these stem cells can only divide so often would that mean that taking these things and getting more of this new cell influx that you're wearing yourself out faster oh yeah I haven't looked at that data but I'm question answer is unfortunately for me I don't know it could be that the cells as so as when the stem cells divide they a symmetrically divide one becomes a stem cell state remains a stem cell the other ones come differentiate and there are other mechanisms at play that you know cause certain damaged materials to stay in one cell versus another cell and I don't know how those molecules would that system so whether that would lead to eventual depletion I don't know certainly the the mouse models out there suggest done because you give them these compounds and they live longer and better so but I don't know if anybody's actually counted them you know so those are other experiments that need to be done alright thank you very much

Channel: Lifespan Extension Advocacy Foundation

Views: 60,206

Rating: 4.9292035 out of 5

Keywords: Oliver Medvedik, Life Extension Advocacy Foundation, Lifespan.io, CRISPR, Gene Therapy, CRIPSR CAS9, CRISPR/Cas9, Genspace, Citizen Science, Grass Roots, Longevity Research, Science Crowdfunding, transgenics, transgenic mice, ZFN, TALENs, histones, epigenetic, Lamark, Transhumanism, Life Extension Research, augmentives, rapamycin, resveratrol, Sirt1, parabiosis, senolytics, Aging, Ageing, SRF, Telomerase, IET, Institute of Exponential Sciences, Designing New Advances

Id: 5zHyUI13NDg

Length: 34min 10sec (2050 seconds)

Published: Thu Aug 11 2016

Reddit Comments

[u/dpwiz]

So, will it?