Nobel Prize lecture: Drew Weissman, Nobel Prize in Physiology or Medicine 2023

I now have the great pleasure to introduce this year's Nobel laat in physiology and Medicine Dr Drew vicman he was born in Lexington Massachusetts he received his MD PhD degrees from Boston University in 1987 and did his clinical training at Best Israel the Aon Medical Center at Harvard Medical School his postdoctoral work was performed at the National Institutes of Health with Dr Anthony foui he then moved to perilman school of medicine at the University of Pennsylvania in 1997 where he met Dr Kiku he's the Roberts family professor in vacine development and director of the Penn Institute of RN Innovations when Dr wisan was working late in the lab which was often the case his wife was sometimes asked by her friends what her husband was doing she then often sarcastically answered oh Drew is saving the world as it turned out he eventually did Dr vicman we are very much looking forward to your Noble lectured entitled nucleoside modified Mr LMP Therapeutics [Music] please thank you very much so I need to thank so many people um I I'll do that throughout my entire talk but I need to thank the Nobel Foundation the Nobel selection committee uh and all of the scientists here for being present for these talks from Katie and I most of all I have to thank Katie because Katie is the person who introduced me to RNA I was a dendri celf scientist from the NIH who had lots of dreams uh Katie supplied the RNA and together we made those dreams come true so I wanted to start by talking about vaccines because that's where I started when I came to pen I had an interest in vacines mostly because dendritic cells which are antigen presenting cells are the principle cell to Target with the vaccine so I wanted to develop ways of loading dendritic cells with the view of making better vaccines I had every method I could think of except mRNA and that's where I luckily met Katie at the copy machine there are many different types of vaccine platforms and everybody in this room has had many of these maybe not all uh they started off hundreds of years ago with live viruses attenuated virus es the the first one was for by a physician Edward Jenner in England who noticed that milkmaids were not developing small pox and when he looked at the cows he noticed the cows had pox Legions that looked a lot like human small poox he cut open those pox lesions and injected them into people and they were protected that was likely the first vaccine ever developed after that a number of other platforms developed of inactivated viruses those are the influenza viruses that us old folks get we also have protein vaccines like tetanus toxoid viral vectors these are the adenoviral vectors that were developed for covid-19 and DNA and RNA so what I wanted to talk to you about first was the nucleus modus mRNA LNP vaccine platform and we started this about 10 years ago and we used a couple of model systems the ones I'll show you now are for influenza and we chose influenza because it's a yearly infection that sweeps the world and it changes every year which means you have to change the vaccine unfortunately it means we have to guess what next year's viruses are going to be and sometimes we get it right and often we don't so influenza in the United States leads to about 30 to 60,000 deaths per year and very large number of people get infected with the virus and are out for days to a week with symptoms when we first started doing this work this was the literature the these were what were reported for responses to vaccines and vaccine manufacturers always compare to what the old vaccines did when they develop a new vaccine so live virus inactivated virus and other delivery routes gave neutralization tighter around one to 100 and that was considered pretty good 1 to 40 is protective but we we took the approach well let's encode the hemog glutenin protein which is the major surface protein of influenza as an mRNA and we immunized animals with that and we compared that to two other types of standard vaccination you'll see in the middle that's an activated virus on the left are live virus in the nose but the MRNA blew us away I did this with a collaborator who ran an influenza lab it took him a month to get these results because he kept having to repeat them and dilute them more and more and he came storming into my office saying what did you do to these samples it's impossible to have a tighter this high and he was accusing me of giving him falsified samples to to make his life miserable I I explained to him know that these are the real results and and the titer were 50 times higher than an inactivated virus they were five times higher than a live virus vaccine typically when you come up with a new vaccine platform if you double the previous vaccine biotech and pharmaceutical companies are happy when they saw a 50-fold increase you can imagine their Joy but as an immunologist I wanted to understand why we were getting such enormous tighter so this is a cartoon of how B cells react to viruses antigens vaccines proteins anything that's foreign and follicular B cells recognize and pick up particles antigens adant and get activated they get help from a CD4 helper cell and they become something known as a germinal center B cell the germinal center B cells are critical because those are the cells that rapidly proliferate Affinity mature and produce Long Live plasma cells and memory B cells so we set up an assay where we measured these three different types of B cells but we did it in an antigen specific manner so what that means is we took ha hemog gluten and protein and fluorinated it and then added it to a flow cytometry panel where we could identify each of these subpopulations of cells so we could count how many B cells in this mouse spleen or lymph node recognize the antigen in the vaccine and we saw not surprisingly that for all three there was about 50 times more antigen specific B cells with the RNA LNP vaccine so that explained the incredibly High tighter it didn't explain where they came from we forgot about some mice actually Norby he's here somewhere forgot about some mice uh and we found them about 13 months later and decided Well let let's look and see what the what what's going on so we took bone marrow out of these mice and we measured Long Live plasma cells The Long Live plasma cells are what makes antibody that appears in your circulation and they can be there for your lifetime we measured antigen specific hemog gluten and specific plasma cells in the bone marrow and the numbers were enormous 0.005% of all nucleated cells were making hemog gluten in antibody that's higher than any progenitor cell in the bone marrow it's an enormous number but we had another question which is is the antibody response any good some vaccines are great at making antibodies but they're not good at making neutralizing antibodies that are protective so in the influenza field there's 18 different subtypes types of influenza A and typically two of influenza B and the main ones that infect people and that you get vaccinated with currently are H1N1 and H3 n2s but there's many different families and these families reside in different animal species so Aven influenza which can possibly cross over into humans and it occurs all the time versus pig influenzas which crossed over in 2009 the current H1N1 was a pig influenza these are new pandemic viruses that cross over from animals and change the world of influenza so they're always a fear every year we guess what the influenza is going to be and sometimes we're right and you get decent protection but none of these vaccines protect against a pandemic crossover virus that can appear at any time so what we did is we took these animals and we they were immunized with an H1 and we challenged them with a very distant H5 it shares about 60% homology with the vaccine and we challenged these animals that had been immunized with an H1 ha and they were completely protected so this told us not only were we making enormous antibody tighters but we were making enormous neutralization and protection tighter so these vaccines are now in phase one clinical trials in people being developed as universal influenza vaccines these are vaccines that will prevent infection from mutated virus from crossover over viruses from birds or pigs or any Source possible and it's the hope for the future instead of having to make a new vaccine every year we make a universal vaccine and it protects people for 10 or 20 or someone no number of years but as an immunologist I had a problem our vaccine did not make sense and the issue was Katie showed you that nucleus modified mRNA has no adant activity it doesn't induce any inflammatory cyto kindes vaccines require adant adant stimulate the immune system and say hey this is a foreign antigen you need to do something as far as we could see there was no adant in this vaccine the lipid nanop particles had not been described as binding any adant stimul ating receptors so we were confused typically most vaccines have an Adent they say oh they're th1 th2 biased doesn't matter they're both inflammatory but there are many different types of CD4 helper cells and we focused in on one particular type known as T folicular helper cells T folicular helper cells and this brings us back to the journ germinal Center B cells they're required to form a germinal center and in the germinal center they Supply help to the germinal center B cells to get them to Affinity mature to get them to mature and become isotype switched Long Live plasma cells Long Live memory cells so without tfhs T folicular helper cells you get a poor antibody response so we investigated was this vaccine somehow inducing tfhs in an un previously undescribed adant activity so we turned to the monkey model and we did this because we could measure antigen specific tfhs and it also allowed us to do something else we compared the RNA LNP vaccine to a standard protein vaccine with a very potent tfh inducing adant double stranded RNA we measured tfhs both Total and antigen specific and as you can see the MRNA lnps just blew the double stranded RNA out of the park we went back and calculated on average most adant alum mf59 uh others double stranded RNA about 5% of the CD4 helper cells are tfh phenotypes the rest are th1 th2 other types with mRNA lnps over 50% of the CD4 helpers are T folicular helper cells so we had enormous induction of these critically important CD4 helper cells and that's the reason why we get such potent antibody tighter with the covid-19 vaccines the tighter of antibodies with these vaccines is typically three to five times higher than what you get after infection with covid-19 and that's rare to have a vaccine that works much better than live virus or live pathogen infection so it this gave us the mechanism for why we got such potent immune responses the next question we investigated was that what about the lnps or the RNA was responsible so we did a simple experiment we took empty lnps and mixed them with protein antigen other people use Alum or mf59 or a variety of their adant we asked could lnps be the adant and that's exactly what we saw when we mix the l&p empty l&p with the protein we got high levels of T folicular helper cells and high levels of antibody production so it turns out that the LNP is an adant it's just not a typical aent we looked at what kind of cyto kindes the l&p induced and it didn't induce typical cyto kindes which are usually type one interferons it induced il6 il6 is a potent stimulus for T folicular helper cells so that the story started to come together that the lmps are a potent adant that induced T folicular helper cells in part through il6 induction and by a lack of type 1 interferon induction with that we wanted to work on better vaccines and we've taken a number of approaches we've developed probably close to 30 different vaccines I'm working with a couple researchers at kolinska right now to develop different vaccines but we tried something unusual that we figured nobody else in the world would try because we were crazy and people knew that um we made a vaccine where we took one ha from every subtype of influenza so that's a vaccine that has 20 different rnas mixed together and nobody believed it would work and I didn't believe it would work but I did it anyway but when we put this vaccine into an animals in both mice and ferrets this is what happens when you put in only H1 ha you get a very potent antibody response to H1 you get a little bit of cross reactivity to group one ha and you get nothing to group two when we put all 20 ha rnas together we got equal antibody responses to every single protein this surprised us first because we were getting 20 antigen responses in a vaccine and there was no antigen dominance and the critical fear of of multicomponent vaccines is one antigen will dominate and the others won't respond we've done this with a couple of different RNA vaccines we've never seen antigen dominance we get a good response to all 20 uh Haas so I only say this when pharmaceutical exacts aren't in the audience but in theory I could see a day when we bring our children to the pediatrician for their every thre Monon vaccination and instead of going at 1 month 3 6 12 16 18 24 uh up until they're 18 years old we could be giving kids one IM immunization of RNA at 1 month 6 months 2 years and be done and instead of getting 20 vaccines per year for a number of years they could get a few would greatly simplify parents' lives would drive pharmaceutical EXA insane because they would lose all the vaccines they were producing I don't know if it'll ever happen but we can dream about it so we developed a new vaccine platform that we could encode just about any antigen we could put as many antigens in as we wanted we've done 20 we've got a vaccine with 75 going right now I'll let you know if that works uh it gave incredibly potent antibody responses and the two mechanisms were first Katie showed Long Live protein production we see protein production for up to 10 days immune systems like that because you're constantly loading the immune system with antigen and a specific induction of T folicular helper cells that drives the antibody responses now I I I would often be asked questions at talks like this if you look at the personalized cancer vaccines that Mna and biotch are doing right now and they both reported great results in melanoma and good results in pancreatic cancer people ask well what's the difference between these two vaccines the principal difference is biotch uses unmodified RNA and Mna uses nucleoid modified mRNA but the other critical difference is modna uses lipid nanoparticles like the covid-19 vaccine and biotch uses a lipoplex which is a lipid shell with an aquous interior that lipoplex has no adant activity so they use unmodified RNA to supply adant to their vaccine Mna uses the l&p Aden activity when you compare these two together we asked who makes better t- cell responses and we compared using lnps 100% modified gave decent t- cell responses but unmodified gave much better much more potent and that's why the unmodified likely work better in cancer vaccines the problem is is when you make this number of te- cells with unmodified RNA you don't make good anybody responses and that's why the clinical trial failed we tried this in a monkey model for an HIV vaccine and we again mixed different concentrations of modified and unmodified mrnas and we saw the same thing 100% modified gave relatively low t- cell responses as you increase the modification you increase the amount of t- cell response we compared that to a chimp Ado vaccine this is similar to what Oxford developed the responses for the chimp Ado were actually a little bit lower but statistically they're about the same so it tells you that depending on how you make the MRNA vaccine you can change the characteristics you can change the amount of antibody or the amount of te- cell responses to address this we tried a different approach we asked could we add cyto kind agents to change the vaccine and in this case we're looking at i12 il12 is a potent t- cell inducing cine and we did a simple experiment we took a microgram of ovalbumin and coding RNA and mixed it with a microgram of either empty or il12 containing uh rnas and put them into mice that had a th000 cd8 t- cells that had a t- cell receptor for ovalbumin so these weren't naive mice but they only had a thousand antigen specific cells so we could easily measure their expansion the addition of I 12 enormously about 10-fold increase the activation and expansion of these cd8 positive T cells they were present everywhere in the spleen and lymph nodes and interestingly they were affector te- cells il12 induces affector te- cells so you can see here the klrg1 positive 127 negative those are affectors memories are 127 positive klrg1 negative so we greatly expanded the number of affector t-c cells without affecting memory t-c cells we're using this now for new types of cancer vaccines that will be used in patients who have genetic deficiencies associated with cancer braa is the the most commonly w un known uh but the idea here is that you treat people before they develop cancer um we know that it's five or 10 years that cancer cells first start to appear before you've got full-fledged large tumors that impair function if we treat these people maybe every 5 years with a vaccine that only makes affector te- cells will will clean out clear away kill all of the transformed cells uh and maybe completely prevent cancer from ever appearing in these patients another critically important thing is that adding 12 unlike unmodified RNA doesn't lower and actually improves the antibody response so this is with I 12 addition it's about over half a log higher antibody tighters so now we've made a vaccine that gives better te- cell responses and better antibody responses at Penn we've got a bunch of different vaccines in clinical development these include many different pathogens including malaria TB HIV hcv we have vaccines for food allergies like peanut allergies have vaccines for en environmental allergens like dust mites and we've got a number of vaccines for autoimmune diseases that are all in development Katie showed that already so I can jump ahead what I wanted to next talk to you about is what I see as the future of mRNA Therapeutics so if you ask anybody Who develops nucleic acid or vir viral Therapeutics the biggest problem is targeting it's getting the nucleic acid or the virus to the cells of Interest lipid nanoparticles go to the liver they also go to dendu cells that makes them a good vaccine but what if you wanted to send them to the heart or the brain or the lung or the bone marrow they don't go there so we developed a way of targeting lipid nanoparticle by adding targeting molecules to the surface uh it's a little complicated chemistry that I don't need to go into but we essentially add any targeting molecule to a peg lipid that sits in the lipid nanoparticle it doesn't change the morphology of the lipid nanop particles it makes them a little bit bigger the surface charge Remains the Same so all of those are critical most importantly it doesn't impair function so these are in in red these are pcam expressing cells and the lnps are labeled with an anti pecam antibody and they bind very well they make luciferase when we add them to endothelial cells only when you add Pam the endothelia cells take up the lnps and make the RNA so not only can they target but they're functional we injected these into a mouse and then analyzed where the activity was so in the absence of targeting all the activity is in the liver when they're pcam targeted we've switched activity to the lungs and there's about equal amounts of activity in the lungs so we switched where these lmps are targeted to and here we can effic efficiently deliver to lungs as an immunologist I had an interest in modifying immune cells te- cells in particular were interesting they could be modified in a variety of ways you could deliver manage into them as a vaccine you can deliver cyto kindes to change their function you can deliver car molecules to kill tumor cells the the list goes on and on but there's a difficulty with te- cells they don't have endocytic activity so what that means is that they're unable to take up nucleic acid particles our idea was we could combine targeted with getting into the cell by targeting a molecule that endocytosis after it's bound and that's exactly what we saw when we added targeted lmps to purified CD4 positive te cells in the absence of targeting we had no uptake with targeting we had very high levels so not only did we bind the cell but we allowed the cell to take up the lmps and transl the MRNA we injected these into animals we saw an increase in spleen activity with targeting the spleen has about 20% CD4 positive t- cells in them but we we did something interesting we re-imaged the animals after we removed all of the organs on the previous slide and when we did that all of a sudden lymph nodes lit up these are parotic and inguinal lymph nodes so what this means is that we gave lipid nanop particles targeted to cd4t cells intravenously into the circulation they escaped from the circulation when into the tissues into the draining lymphatics back to the draining lymph nodes where they targeted CD4 positive T cells were able to be taken up and the RNA was translated so we could complete that entire process here we purifi the cd4s and show that the activity is there so we were able to deliver nucleic acid Therapeutics to a particular cell type in Vivo we used a model system known as the creck system what that involves is you put a stop code on in front of a fluorescent protein and you put what are known as locks P sites specific DNA sequences and then you deliver a cre recombinase mRNA the cre recombinat cuts out the stop codon and it makes the cells Express the fluorescent protein when we did that we saw enormous levels of fluorescent protein in both splenocytes and lmph nodes this is the fluorescent protein expression there's almost nothing without targeting very high levels with targeting but from an HIV point of view there was something more important this axis is an activation marker HIV forms latency in unactivated resting CD4 positive t- cells so if you're delivering a therapeutic for HIV it has to Target be taken up and be translated in resting cells and that's exactly what we saw here the resting cells had equal levels of Gene Rec combination so this is now in a MAAC experiment looking to cut HIV out of the Genome of latently infected cells we saw very high levels of Gene Rec combination both in spleen and lymph node we've looked at a variety of other tissues and see the same thing so our conclusion ions were that we could figure out how to Target specific cell types in Vivo we then went on to test another model system so if you come from pen Penn clinically developed car T cells the first clinical trials were done there the first two drugs were FDA approved at pen so P pen has a close relationship to car T cells what a car T cell is it it's an altered cd8 killer cell so cd8 killer cells recognize a peptide derived from a virally infected or oncogenic cell in association with the host MHC they're very effective at killing the problem is is just about everybody in this room has a different peptide and many different mhc's so they're not translatable from one person to the next what a C cell does is it puts a piece of an antibody that recognizes a tumor Associated antigen on the surface of the te- cell that way you can make killer te- cells that kill any tumor or any cell with that antigen cd19 was the first one to to be developed the problem with car T cells this is what it takes to make a carart T Cell you start with a patient you lucer the patient that's a machine that has a cuge in it that spins their blood takes out the white cells gives everything back it does that over and over and over over a few hours and it takes out a couple billion te- cells you then have to infect those te- cells with lent viruses in culture you then have to stimulate them for 10 days to expand how many there are and then you can give those back to the patient and then about 70% of the time they kill the tumor and cure the patient the problem is it cost half a million dollars a dose because it's 10 days in a very fancy lab um under GMP conditions um these sites are available in in the US and in Europe there's one in China there's I think one in South America there is none in subsaharan Africa so they're very limited in where they are because of the cost and the E expertise needed to do it we had the idea since we can now target cells in Vivo could we make car T cells in Vivo and the thought was well if we targeted the LNP to bind to all t- cells and then we put a car molecule in the MRNA the T the lnps would be taken up by the te- cells the RNA would be translated and they' put the car on the surface of the t- cell we chose a car that recognized activated fiberblast and a model of cardiac fibrosis so I apologize for the echocardiography readings but what what you need to realize is the gray are normal animals and these are just different measures of heart function as a a an internist this is what I look at this is how much blood is pumped out by the heart so in a normal heart it's about 70% of the blood is pumped out when a heart becomes fibrotic due to hypertension that reduces and the lower it it goes the worse off the patient is we treated these mice with a single dose of lmps that targeted te- cells and we return their heart function to normal their ejection fractions returned to normal the size of their ventricles everything returned to normal we stained the heart muscle for fibrosis the normal animals you see very little the fibrotic animals you see all this red fibrosis one treatment with lmps essentially cured these animals so instead of a half a million dollars 10 days specialized facilities now it can be an off the off-the-shelf drug somebody comes in with cardiac fibrosis heart disease is a number one killer in the world they get an injection they go home their heart is better this can also be expanded to many other diseases and many other applications the other thing I wanted to tell you about is bone marrow stem cell targeting and to me this is one of the most critical applications for RNA lnps currently there are thousands of genetic diseases of the bone marrow these include all of the immune deficiencies skid uh they include the phemia and most importantly it includes CLE cell anemia CLE cell anemia has 300,000 people a year are born with the disease they're mainly in subsaharan Africa but they're present throughout the world in the US they're about to approve uh FDA approve a gene therapy for sickle cell the problem is is it cost about a million to4 million per person you multiply a million time 300,000 that bankrupts the world so we need better ways of treating genetic diseases of the bone marrow the problem with bone marrow stem cells are their Rarity so this is the number of cells in the bone marrow of a mouse this is the number of repopulating bone marrow stem cells it it's a tiny fraction so hitting those rare cells is very difficult but we took the same approach that we did for CD4 we in this case we used a marker known as cd17 or C kit with a single treatment of mice and we're delivering a FL ENT protein we were able to hit about 90% of bone marrow stem cells incredibly efficient we went back to that cre lock mice and we followed them over time so the animals got a single treatment here and then we followed their blood over time looking for a fluorescent protein from the cre enzyme after a few weeks 100% of of the red cells white cells and granular sites were fluorescent so we had 100% efficiency of Gene editing the bone marrow of these mice now for stem cell transplant people they only believe Stem Cell results if you do a secondary transplant what that means is you take the bone marrow from these mice and you take it out you radiate a new group of mice to kill their bone marrows and then you put the new bone marrow cells in when we did that within a couple weeks 100% of red cells white cells and granulites were all Gene edited that meant we had 100% efficiency at Gene editing bone marrow stem cells in Vivo now that that's critical for certain diseases for for diseases like skid uh and other immune deficiency gencies if you fix half a percent of the stem cells you cure the disease but for Cle cell anemia you have to fix 25% of all of the repopulating bone marrow stem cells so with an efficiency of 100% targeting then however well we can Gene edit will likely give us potentials for cure this is now a cure where we can line people up on the Treet give them a single injection of RNA lnps and cure their disease no million dollars no fancy lab facilities just off-the-shelf injections it can also be used for a variety of other things we do bone marrow transplants for a number of different types of cancer you give people high doses of chemotherapy and sometimes radiation to prepare them for this there's a mortality rate of around 5% from that treatment now we can deliver toxic genes to bone marrow stem cells and selectively kill them mortality will be much much lower we can deliver therapeutic proteins to stimulate granulocytes or platelets or other needed cell types we can deliver really any kind of protein we're also able to Target a variety of other cells and tissues we can Target brain Katie talked about treating Strokes we're Now setting up to do studies on large animals delivering therapeutic proteins targeted to the brain we can deliver to the heart we can deliver to lungs we can deliver to te- cells we can deliver to any immune cell we continue to expand what we can Target So in theory some day gene therapy might be as simple as walking into a doctor's office having them take a vial out of the fridge injecting it into a person and curing the disease RNA Therapeutics has enormous potential for vaccines for genetic diseases for therapeutic protein delivery for the treatment of a variety of diseases I I joke with my lab people we haven't thought of everything that we can do with RNA and that's their job I'm I've done my job um but the future of RNA I think is really going to be enormous so I need to thank all of the people involved of course Katie who I started all of this work Norbert Pari and hameda pares and my lab were the leaders for vaccines and targeting technology people from a ous Therapeutics made the lipid nanoparticles vladakov's lab developed the targeting technology that we use for all of our invivo targeting I have to mention John Epstein because he's the CSO of my University um and um the many other labs that we've worked for thank you very much

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Length: 47min 35sec (2855 seconds)

Published: Fri Jan 05 2024