Session 7: Autoimmunity and Allergy

Transcript of Part 3: Discovery and Development of Natalizumab for the Treatment of Multiple Sclerosis

00:00:07.02	Hi.
00:00:07.22	I'm Ted Yednock and I'm an immunologist trained at the University of California in San Francisco.
00:00:11.17	And for a number of years, I worked for a company called Elan pharmaceuticals.
00:00:15.09	At Elan, and we developed a drug called Tysabri for the treatment of multiple sclerosis.
00:00:20.09	So, I'm going to talk about the identification and development of Tysabri in two lectures.
00:00:26.04	Today, I'm going to talk about the scientific underpinnings of Tysabri and how we think
00:00:31.02	it works by inhibiting immune cell infiltration of the central nervous system,
00:00:35.15	preventing the damage associated with the autoimmunity in MS.
00:00:40.11	I'll also talk about the preclinical work and the clinical studies that we did in order
00:00:44.10	to demonstrate that the drug was safe and efficacious.
00:00:48.03	In the second lecture, I'll talk about how we believe the selective mechanism of this drug
00:00:52.20	allowed for the emergence of a very rare but serious brain infection called
00:00:57.27	progressive multifocal leukoencephalopathy, or PML.
00:01:01.19	And I'll also talk about the complexity that that caused for patients, and physicians,
00:01:06.25	and the FDA, as well as the companies, in trying to understand how PML was associated
00:01:12.07	with Tysabri.
00:01:13.22	So, at first, I...
00:01:15.05	I will say a few words about multiple sclerosis.
00:01:22.00	So, MS is an autoimmune disease, of the CNS, as I mentioned.
00:01:26.02	It's involved in the destruction of myelin.
00:01:27.25	So, there's a very strong T and B cell response against myelin.
00:01:31.27	And once myelin is lost, you have impaired nerve conduction and neuronal loss.
00:01:36.11	Now, MS is... can happen anywhere in the central nervous system, so it can affect any part
00:01:40.26	of your body.
00:01:41.27	A patient may feel tingling in an arm or hand, or they may have loss of vision, or issues
00:01:49.15	with control of bladder function, or... or not be able to walk.
00:01:54.06	It affects about 300,000 patients in the United States.
00:01:57.06	It's more common in women than men.
00:01:59.12	And, as I've alluded to, it's a devastating disease and it affects people in the
00:02:02.25	prime of life, between the ages of 20 and 40.
00:02:06.04	But it can be seen at any age.
00:02:08.23	Now, usually, MS is a relapsing... it begins as a relapsing-remitting disease.
00:02:14.05	So, the attacks can come along very quickly, you know, vision loss can happen very quickly,
00:02:19.17	and then, over a period of days to months, there will be a degree of recovery to varying levels.
00:02:25.08	But after 15 or 20 years, the disease often becomes progressive.
00:02:30.18	And this happens in about 65% of patients.
00:02:33.23	And now, it's more of a chronic neurodegenerative disease, without the acute inflammatory attacks.
00:02:40.12	So, MRI is actually a very useful diagnostic and monitoring tool for multiple sclerosis.
00:02:46.14	So, this is an MRI image.
00:02:48.14	And what... what is often done in MS is that patients are given a contrasting agent called
00:02:53.22	gadolinium.
00:02:54.22	It's injected intravenously, and normally it's excluded from the brain.
00:02:58.27	However, in MS, when there's an active lesion, the blood-brain barrier is leaky and so gadolinium
00:03:04.03	will enter the brain and show up very prominently in an MRI.
00:03:08.03	Now, if you were to look at this lesion through a microscope, you would see this.
00:03:13.17	This is a... a blood vessel in multiple sclerosis.
00:03:17.09	And what's happened here is there are immune cells traveling throughout the body in the bloodstream
00:03:22.27	and... now, normally, they just go right through the brain.
00:03:25.10	But in the case of MS, there's something happening to the blood vessel wall, and the immune cells
00:03:30.12	will adhere to the vessel wall, migrate in, and accumulate in these large clusters around
00:03:36.15	the blood vessel.
00:03:37.15	And this basically is ground zero for demyelination and neuronal damage.
00:03:41.18	So, in 1990, we had a very simple hypothesis, and that was that if we could inhibit immune
00:03:48.00	cell attachment to the blood vessel wall we could prevent their migration into the brain
00:03:51.19	and all the damage associated with that.
00:03:53.26	And so... when I say adhesion molecule, picture Velcro.
00:03:57.22	So, we're trying to identify the little hooks on the... on the immune cell that allow it
00:04:01.22	to attach to the blood vessel wall, specifically at times of inflammation.
00:04:05.28	Now, since this is a brain disease, it's really important to be able to do these kind of studies
00:04:11.09	using an... an animal model.
00:04:12.23	So, the best animal model for multiple sclerosis is experimental autoimmune encephalomyelitis,
00:04:18.24	or EAE.
00:04:20.20	Now, in EAE... this is a... a... a model in the guinea pig, but we also use models in
00:04:25.19	mice and rats.
00:04:26.26	So, in this model, this is a brain... a slice through the brain of a guinea pig with EAE.
00:04:32.03	You can see a large blood vessel.
00:04:34.05	And in the blood vessel there are infiltrating immune cells, which are the blue cells, here.
00:04:39.23	And the section is stained with a red dye for myelin.
00:04:42.18	So, you can see that around the blood vessel there's a huge loss of myelin.
00:04:47.22	And then, also, the... the little green dots are axons, or nerve fibers, that are... are...
00:04:54.02	you're looking at them in cross-section.
00:04:56.07	And normally, over here, you can see that the nerve fibers are surrounded by red myelin,
00:05:01.12	so they're quite well insulated from each other.
00:05:03.27	But when the myelin is lost, the nerve cells or the nerve fibers are pushed around and clustered,
00:05:07.24	and the spacing between is occupied by immune cells and by edema.
00:05:11.15	And so you can see, with this kind of damage, how nerve conduction would be impaired.
00:05:18.18	This is the way the model progresses in animals.
00:05:21.03	So, in the guinea pig, we immunize the animals with spinal cord homogenate.
00:05:26.17	This causes a very strong response, again, of B cells and T cells to myelin.
00:05:31.19	And so that... by day 9 or 10, the immune cells will begin to infiltrate the
00:05:35.14	central nervous system, and you'll begin to see a degree of hind limb paralysis in the... in
00:05:40.01	the acute phase.
00:05:41.01	And then the animals begin to get a little bit better.
00:05:43.21	But then, by day 15 or so, the disease begins to, well, progress, and enters the chronic phase.
00:05:50.18	And the... the difference here is this is when demyelination begins.
00:05:53.20	So, once demyelination begins, the nerve fibers are pushed around, the immune cells are even
00:05:58.15	more activated, and the disease progresses with just more and more demyelination.
00:06:03.27	When we were first doing these experiments, we were a very small lab in a...
00:06:07.05	in a small company and we really didn't have a good model of EAE.
00:06:11.19	In fact, we weren't working in this area at all.
00:06:14.17	We... we were trying to develop therapeutics for Alzheimer's disease.
00:06:18.14	And we were trying to develop an animal model of Alzheimer's.
00:06:21.06	It would have been the world's first model, which... which we eventually did, but
00:06:25.15	our initial attempts weren't so good.
00:06:27.07	And in fact, we really were inducing a model in which there was a chronic inflammation.
00:06:33.21	And it's that kind of inflammation that initially got us interested in this area to begin with.
00:06:37.26	So, we took inflamed brains from these animals in this strange model of Alzheimer's disease,
00:06:43.24	and did a very simple experiment.
00:06:45.27	We took these sections and overlaid them or exposed them to a suspension of human lymphocytes.
00:06:51.20	And we found that the lymphocytes would adhere selectively to these inflamed vessels.
00:06:55.26	So, you can see these very dark blue, large circles are the human lymphocytes, and they're
00:07:02.06	adhering right in the center of these blood vessels, right where you would expect to see
00:07:06.06	blood cells in the circulation.
00:07:08.13	They don't bind elsewhere in the section and they didn't bind to non-inflamed vessels.
00:07:12.16	So, the fact that we were getting physical adhesion of these cells to inflamed vessels
00:07:18.21	suggests that this is probably a physiologically relevant... relevant interaction.
00:07:22.26	Because these ligands -- whatever these adhesive ligands... whatever they are -- were being
00:07:26.07	induced in the context of a whole brain.
00:07:29.04	We also isolated endothelial cells from rat brains and put them into culture,
00:07:34.25	stimulated them with TNF, and we found, there,
00:07:37.17	that lymphocytes would adhere here as well.
00:07:39.17	So, that was great.
00:07:41.00	We now had two different assays by which we could look at lymphocyte interaction with
00:07:44.21	inflamed endothelium, so we could screen for inhibitors of that interaction.
00:07:49.12	What we did is made thousands of antibodies against the lymphocyte surface, at random,
00:07:54.22	and screened for antibodies that would inhibit this.
00:07:56.25	So, in this assay, we could screen thousands of antibodies.
00:08:00.15	And in this one, we could screen umm... perhaps in dozens.
00:08:03.07	It was a much more cumbersome assay.
00:08:06.00	But, nonetheless, both assays gave us exactly the same answer.
00:08:09.22	And that is that alpha 4 beta 1 integrin is very important for lymphocytes and monocytes
00:08:15.28	to adhere to the... to the inflamed brain vessel.
00:08:18.16	So, in the presence of anti-alpha 4 or anti-beta 1, you can see that there was no longer any
00:08:24.00	human lymphocyte attachment to the vessels in this section.
00:08:29.09	So, this is what we thought was going on.
00:08:33.10	Lymphocytes and monocytes expressed alpha 4 beta 1 integrin and they were binding to
00:08:37.01	some ligand expressed by inflamed brain endothelial cells.
00:08:41.19	So, we also knew in 1990... and, in fact, I was...
00:08:45.01	I was kind of disappointed with this observation.
00:08:48.19	Because we knew that alpha 4 integrin was expressed by many cells in the circulation
00:08:53.13	-- lymphocytes, monocytes, eosinophils, basophils.
00:08:56.22	Unfortunately, it was low or negative on neutrophils, red blood cells, and platelets.
00:09:02.06	However, we had been hoping to find sort of a brain-specific adhesion molecule that was
00:09:06.15	expressed by a small subset of... of pathology-causing immune cells.
00:09:10.27	But, in fact, it was expressed very broadly.
00:09:13.20	Nonetheless, as you'll see, I believe that alpha 4 beta 1 integrin turned out to be
00:09:17.23	a pretty selective adhesion molecule for the CNS.
00:09:22.00	The molecule, alpha 4, had been cloned the year before by Yoshi Takada and Martin Hemler's group.
00:09:27.06	And a number of groups were also identifying the ligands for alpha 4.
00:09:32.13	For example, Elizabeth Wayner had found that an alternatively spliced form of fibronectin
00:09:36.20	was a ligand for alpha 4, and also Roy Lobb at Biogen had identified VCAM-1,
00:09:43.14	expressed by human umbilical vein endothelial cells in culture.
00:09:47.18	So... but no one really knew how alpha 4 beta 1 worked in vivo and how it contributed to
00:09:52.16	disease pathology.
00:09:54.00	And that's when we go back to umm... the need for a good animal model of EAE.
00:09:58.15	So, the model that we had with our Alzheimer's-inflamed brain wasn't really going to be good enough
00:10:03.21	for that.
00:10:04.21	So, we went down the street and visited Larry Steinman at Stanford.
00:10:09.19	And Larry Steinman is a world expert in multiple sclerosis and in EAE.
00:10:14.10	And we took... we went to him and showed him our results with alpha 4 beta 1 integrin,
00:10:18.17	and we had antibodies against alpha 4, and we asked him if he could help us test it in
00:10:23.05	a... in a really good model of EAE.
00:10:25.09	And that's exactly what he did.
00:10:27.03	Ni...
00:10:28.03	Nathan Karin, who was a postdoc in his lab at the time, was working with a rat model
00:10:32.21	of EAE.
00:10:33.21	So, the first thing we did was confirm that using brain... sections of inflamed brain
00:10:38.11	from animals with... rats with EAE, that we had the same sort of lymphocyte adhesion,
00:10:43.04	and that it was completely dependent upon alpha 4 integrin.
00:10:46.14	Nathan then tested one of our antibodies against alpha 4 and found that it was very protective
00:10:52.00	in preventing disease in these... in this animal model.
00:10:54.21	So, that was great.
00:10:56.09	Based on that, we... we started bringing in a number of different EAE models into our...
00:11:01.07	our little lab.
00:11:02.26	This was one, again, in the guinea pig.
00:11:04.25	And here we wanted to characterize more thoroughly what... how alpha 4 integrin works, how quickly
00:11:10.12	it works, and the impact that it can have on the disease.
00:11:13.15	So, in this study, we labeled lymphocytes with indium-111, which is a very high gamma emitter.
00:11:20.19	And we found that if you inject the cells back into the bloodstream they will migrate
00:11:25.18	to the brain.
00:11:26.18	So, you can just take out the brain and do a gamma count and see how many cells there.
00:11:30.07	But, in the presence of anti-alpha 4, inhibition was 80 or 90%.
00:11:35.11	So, in this short-term migration assay, lasting about 18 hours, alpha 4 integrin was clearly
00:11:41.09	affecting the ability of cells to migrate into the brain.
00:11:45.17	And then we treated guinea pigs with the antibody, just at the onset of disease.
00:11:51.06	This was like day 10 or 11, just when paralysis was beginning but before demyelination had happened.
00:11:57.28	And what we found is that by day 20 we had a very strong impact in preventing demyelination.
00:12:02.28	So, in the control-treated animal, you can see that they're... we have stained these
00:12:07.19	sections of spinal cord with a blue dye for myelin, and there's an extensive loss of myelin
00:12:14.01	by day 20 in the control animals.
00:12:16.01	However, in the presence of anti-alpha 4, the myelin is almost completely intact.
00:12:20.05	So, there was very strong protection against the loss of myelin.
00:12:26.22	We next... we next treated animals at day 20.
00:12:30.15	So, this is after demyelination has already occurred.
00:12:34.02	And we treated them for a long time -- 56 days -- because you wanted to see if it would
00:12:38.06	impact the disease and if that impact would last.
00:12:42.04	And what we saw was this: a very strong reversal of the paralysis and... that lasted for
00:12:47.17	the entire time of the treatment.
00:12:50.19	Okay, so we did this experiment a number of times, looking at different time points to
00:12:54.19	see what impact that we would have on myelin.
00:12:58.07	So, in this study, we treated the animals after demyelination had already occurred,
00:13:05.03	and then we inhibited alpha 4 integrin for various periods of time.
00:13:09.17	We found that after 20 or 40 days of treatment that we actually would allow myelin to repair.
00:13:15.26	So, the way we knew this was that under control conditions myelin is either present or it's not.
00:13:22.21	So, over here, you can see that... here is myelin, this is an area where the myelin
00:13:27.08	has been eliminated, and the... the line between the two is very, very stark.
00:13:31.25	So, it's either there or it's not.
00:13:34.17	However, in animals that had been treated with an inhibitor of alpha 4 integrin,
00:13:37.23	we saw this very pale blue pallor, which we confirmed by electron microscopy to be regeneration
00:13:44.16	of myelin.
00:13:45.16	So, we believe that what was happening is that by inhibiting immune cell infiltration
00:13:49.14	into the CNS we were dampening down this inflammatory response and basically creating an environment
00:13:55.22	that was permissive for remyelination.
00:13:57.22	So, these are the studies that led to the development of Tysabri, which is a... we took
00:14:05.20	our best antibody against alpha 4 integrin and humanized it by CDR grafting.
00:14:10.16	And this was done in collaboration with the MRCC in London.
00:14:14.19	And fortunately, the antibody retained the full potency of its murine parent.
00:14:22.00	Just to give a perspective on scale, this is a model of an antibody next to a model
00:14:26.26	of an integrin.
00:14:28.10	Umm... the... the antibody actually binds to the head group, up here, which is the
00:14:33.02	business end of the integrin.
00:14:34.02	This is where it binds to ligands.
00:14:35.22	Now, recently, the crystal structure of natalizumab bound to the head groups of alpha 4 integrin
00:14:42.04	has actually been solved, by Yamel Yu in Timothy Spring... in Tim Springer's group.
00:14:48.11	So, this is looking at the crystal structure, just at the head groups of the integrin umm...
00:14:54.23	and the blue is beta 1 and the green is alpha 4.
00:14:58.18	And the ligand binding site -- this is for VCAM-1 -- is right in between the two.
00:15:03.03	So, there are critical contact residues on both sides of the line.
00:15:06.12	Now, natalizumab, he found, bounds right next to this.
00:15:10.11	So, the binding actually doesn't overlap.
00:15:12.26	And, in fact, if you take the business end of VCAM, the first domain, it will bind just
00:15:18.16	fine in the presence of... of Tysabri.
00:15:21.20	However, the second domain of VCAM causes steric hindrance, so that, when natalizumab...
00:15:29.10	when natalizumab is bound in the presence of the full-length ligand, its binding energy
00:15:34.10	is greatly decreased because of the steric hindrance.
00:15:37.06	What they also noticed in the lab was three critical contact residues within the natalizumab
00:15:43.14	binding site that help to explain observations that we had made a number of years earlier.
00:15:48.12	So, when you're developing a drug, it's important to have animal species for efficacy testing,
00:15:53.05	but you also need to have two different animal species for toxicology.
00:15:56.27	You need to be able to show that it's safe in at least two species.
00:16:00.28	So, we were disappointed because our... our best model for EAE at the time was in the rat.
00:16:06.22	However, the rat has a mutation in one of these critical residues, and we found that
00:16:10.16	the antibody didn't react.
00:16:12.28	There also was an EAE model in the marmoset monkey, which was a model that was set up
00:16:18.05	at UCSF.
00:16:19.05	And, again, we found that our antibody did not react with the monkey, which... we were
00:16:22.11	really surprised, because it's very closely related to the human.
00:16:26.12	But now it makes sense with... with the finding that there's a mutation in one of these critical
00:16:31.02	amino acids.
00:16:32.12	So, we did find, however, that the... the antibody reacted with cynomolgus monkey and
00:16:38.20	with guinea pig.
00:16:39.20	And, as you can see, that the sequence of these... of the three critical contact residues
00:16:44.03	in these species are identical to those in human.
00:16:46.27	And these are the species that we had chosen for our toxicology.
00:16:52.21	So, in order to develop a drug, you need to do a lot of toxicology studies.
00:16:58.21	This is just a subset of the things that we did.
00:17:00.21	And I'm not going to go through all these, other than to mention a couple findings.
00:17:05.18	First of all, in the first study up here, it was a six-month study in cynomolgus monkey
00:17:10.15	involving very high doses of... of natalizumab.
00:17:13.08	Umm... natalizumab, by the way, is the generic name for Tysabri.
00:17:17.12	So, we did 60 milligrams per kilogram dosing every week.
00:17:22.12	And, in patients, we find that 3 milligrams per kilogram dosed every month
00:17:27.06	is what it takes for efficacy.
00:17:29.15	So, in fact, in the monkeys, we were achieving blood levels more than a thousand-fold that
00:17:33.28	you would find in humans.
00:17:35.10	So, at the end of 6 months, we looked to see what impact it would have, particularly on
00:17:40.02	the immune system.
00:17:41.03	Remember, alpha 4 integrin is expressed by almost all circulating mononuclear cells.
00:17:46.00	And so we were quite interested to see what would happen in the immune system.
00:17:50.17	And much to our relief, we found that hematopoiesis... bone marrow was... was very much intact, so
00:17:57.10	lymphocytes and red blood cells were being made normally.
00:18:00.05	And this was important because alpha 4 integrin had been shown to be involved in hematopoiesis.
00:18:04.11	We also did immunotoxics... immunotox... immunotoxicity profiling and found that if you vaccinated
00:18:13.20	animals in the presence of Tysabri the... they responded just... just fine to the vaccine.
00:18:18.24	There was a slight delay in the initial antibody response, but the animals quickly caught up.
00:18:25.01	And other than that, in these studies we also looked at the impact of natalizumab on implantation,
00:18:31.03	in development, and... as well as on development of the fetus during the full pregnancy,
00:18:37.16	and found very little impact there.
00:18:39.25	So, to summarize our... our preclinical and our toxicology studies, functional screens
00:18:47.09	are... are really important when trying to understand a disease process.
00:18:50.10	And this may be an obvious thing to say, but I'm just so impressed with the simplicity
00:18:54.26	of the tissue assay that we had performed, looking at the endogenous induction of ligands
00:19:01.13	involved in this inflammatory process.
00:19:03.19	In fact, this tissue assay is something that was established by Stamper and Woodruff in the 70s,
00:19:08.22	and had... has been used successfully for identifying a number of immune adhesion receptors.
00:19:14.20	Our other findings were that anti-alpha 4 was effective in multiple models of CNS inflammation,
00:19:19.19	whether it was a very funny Alzheimer's model or EAE.
00:19:22.18	And, in fact, in different labs around the world, it's been shown to be effective in
00:19:27.10	rat, mouse, guinea pig, and primate EAE, as well as
00:19:31.03	two different models of CNS viral inflammation.
00:19:34.14	So, these are models in which there's a CNS infection by a virus, the immune system goes
00:19:39.26	in to fight the virus and causes damage, and we found that anti-alpha 4 was actually protective there.
00:19:46.04	It protected against the damage and the... the body was still able to effectively fight
00:19:51.00	the virus.
00:19:52.00	So, again, it was suggest... suggesting that... that natalizu... natalizumab would be safe.
00:19:57.20	There was a lot of criticism of EAE models, because it doesn't exactly mimic the processes
00:20:04.07	of multiple sclerosis.
00:20:05.26	But in a case like this, where we're focusing on the immune infiltration aspect, I think
00:20:10.25	it's actually a very good model.
00:20:12.20	Because immune cells need to enter the brain to effectively induce EAE, and they also need
00:20:17.05	to go into the brain to cause MS.
00:20:19.24	So, by studying mechanisms involved in this infiltration process, I think that EAE reflects
00:20:25.22	what's happening in humans.
00:20:28.08	And then, finally, natalizumab appears to be safe and well tolerated in preclinical safety studies.
00:20:33.17	Again, this was important because... because of alpha 4 integrin's broad distribution.
00:20:38.00	It just speaks to the fact that toxicology is an empirical science.
00:20:42.12	You really don't know until you test it.
00:20:45.15	And then, even then, it's important to remember that you always need to be vigilant.
00:20:50.19	So, these are the studies that led to the clinical program for natalizumab.
00:20:55.19	In the first case, we looked at a Phase I study in healthy volunteers and found that
00:21:00.14	the antibody was well tolerated with low immunogenicity.
00:21:03.07	And this is just with a single dose.
00:21:05.11	And we also were able to determine that it has a 12 day half-life, which is sufficient
00:21:10.02	for once a month dosing at 3 milligrams per kilogram.
00:21:13.26	So, reasonable dosing.
00:21:16.15	As we saw in the animal models, there was a two-fold increase in the number of circulating lymphocytes.
00:21:22.14	This is reflec... this is consistent with the drug's mechanism of action.
00:21:26.11	It's making it a bit more difficult for cells to get out of the blood stream into tissues.
00:21:30.02	And so you quickly establish a new equilibrium, which involves more lymphocytes in the blood.
00:21:36.13	But this two-fold increase really represents just the higher end of normal range.
00:21:41.24	Based on those studies, it went into a Phase II study.
00:21:44.02	And... and initially we... our idea was to treat acute flares in MS.
00:21:49.26	So, a patient would have a flare, they would come in to the doctor, get a dose of natalizumab,
00:21:54.21	and hopefully we would quickly dampen down that flare and prevent the damage associated
00:21:59.02	with it.
00:22:00.02	In fact, what we found... and this is with two doses covering two months... we found
00:22:05.18	that there was no impact at all on the time of resolution or the outcome of that relapse.
00:22:12.02	So apparently, by the time a patient is feeling the onset of a relapse, the immune cells are
00:22:17.05	already in the brain, and by inhibiting more it really does not have an impact.
00:22:22.00	However, what we did notice in the study was that by the end of the second month the new
00:22:28.12	lesion development in MRI... by MRI had basically stopped.
00:22:31.28	So, this is looking at the cumulative number of new gadolinium-enhancing lesions by MRI.
00:22:38.11	And you can see that in the placebo group, in red, that there's a pretty consistent accumulation
00:22:43.02	of new lesions.
00:22:44.05	But, by the fourth week on natalizumab, these lines are beginning to diverge.
00:22:49.02	And so that... by the eighth week, the second month, when the drug is still on board,
00:22:53.11	you can see that new lesion activity had... had almost stopped.
00:22:57.05	At the next time point, when the drug was gone, MRI activity began to return.
00:23:01.14	So, this result led us to the idea that, since the drug appeared to be safe, perhaps we could
00:23:06.08	take it into MS for chronic treatment -- prevent the development of new lesions and hopefully
00:23:11.07	have an impact on the progression of disease.
00:23:14.12	So, this led to the our Phase IIb study, and this was a six-month study involving
00:23:21.18	monthly dosing as well as MRIs, which are indicated by the Ms, and then we follow the patients
00:23:27.27	for six months after that.
00:23:29.24	210 patients, looking at placebo and two different doses of drug.
00:23:34.11	And the primary endpoint would be looking... was looking at lesion burden, as well as relapse rate.
00:23:40.23	This is what we found.
00:23:41.27	So, in the months before treatment began, and at the time of treatment, all three groups
00:23:46.17	had sim... very similar levels of MRI activity.
00:23:51.00	And then, looking at the placebo group, they maintain this level of activity pretty consistently
00:23:55.11	over the six-month period.
00:23:57.03	However, in the groups that were treated with natalizumab,
00:24:00.06	and this is both for the 3 and the 6 milligram group,
00:24:03.12	you can see that MRI activity almost completely stopped.
00:24:06.19	So, again, by the second month, activity was very, very low.
00:24:10.08	Now, I have to say I...
00:24:11.15	I was sitting in the audience of the company when we first saw these results and it...
00:24:19.07	it really took my breath away.
00:24:20.15	This is... this was a day that I will remember in my career.
00:24:23.07	And I think that it... it really changed the way I look at things from then on out,
00:24:28.07	because as a scientist working in the bench it's really hard to believe that you can have an impact
00:24:32.20	on human disease.
00:24:33.22	And so this...this really made me believe that and, as I said, has had a long-lasting impact.
00:24:41.08	The drug also affected the number of relapses.
00:24:43.28	So, there was about a 50% reduction in the number of relapses, as well as a number of
00:24:48.02	patients who had a relapse.
00:24:50.00	And also decreased the need for steroid rescue.
00:24:52.25	So, these patients, if they had a flare, could... had the option of having steroid and on natalizumab
00:24:59.11	they did not feel the need to do that.
00:25:02.25	This is looking at the same data, the MRI data.
00:25:07.06	So, the blue line is 6 milligrams per kilogram, the red line is 3 milligrams per kilogram.
00:25:13.23	And so... you can see that... you can see that the drug is... for 3 milligrams per kilogram,
00:25:20.26	when it begins to go away, MRI activity returns.
00:25:24.25	And with 6 milligrams per kilogram, it takes a little bit longer for... for activity to resume.
00:25:29.07	You can overlay lymphocyte counts on top of this.
00:25:33.06	So, this is looking at the number of lymphocytes in the bloodstream.
00:25:36.22	And at a very beginning, you can see that there's this... this increase to lymphocyte
00:25:39.22	counts that's maintained very stably throughout the treatment period.
00:25:43.01	But then, in the 3 milligram per kilogram due... dose group, you can see that as the
00:25:47.12	drug goes away the lymphocyte counts drop, and the MRI activity goes up.
00:25:52.07	And with the 6 milligram per kilogram group, you can see this is... this is basically shifted
00:25:56.11	by a month.
00:25:57.11	So, the cell counts drop and the MRI activity comes up a little bit later.
00:26:01.07	So, this is very consistent.
00:26:02.16	This finding is very consistent with the mechanism of action that we're thinking.
00:26:06.07	As blood cells leave the... as immune cells leave the bloodstream, the MRI activity would
00:26:12.14	resume.
00:26:13.14	Alright, so our... for our Phase III studies in relapsing-remitting multiple sclerosis,
00:26:19.08	in order to get approval for a drug you actually have to have two separate Phase III studies.
00:26:24.03	And so both of these were about a thousand patients.
00:26:26.15	Umm... the first one involved monotherapy, so it was just Tysabri versus placebo.
00:26:31.11	The second one was on... with patients who were already on interferon beta, which was
00:26:36.05	the standard of care for MS patients.
00:26:38.28	And in this case, these patients were having disease breakthrough even on interferon.
00:26:42.27	So, the trial was natalizumab... natalizumab plus interferon versus interferon by itself.
00:26:49.05	So, this was a randomized double-blind study, as were the Phase II, and involved patients
00:26:55.08	who had to have at least one relapse in the prior year to the study.
00:26:58.14	It was a two-year treatment with an early assessment at one year.
00:27:04.24	And the findings were very consistent with what we saw in Phase II.
00:27:07.13	The number of gadolinium-enhancing lesions, both in first year... in the first year,
00:27:12.28	as well as the second year, were inhibited by 92%.
00:27:15.17	So, very strong inhibition of... of this apparently acute inflammatory reaction.
00:27:21.15	And this translated to a 68% reduction in relapse rate, again, both in the first year
00:27:26.26	and the second year, as well as in the third year.
00:27:30.02	This is an open label extension study at this point, so there isn't a placebo control because
00:27:34.02	everyone's on drug.
00:27:35.13	But, in all three years, the disease activity or the relapse activity was reduced to the
00:27:40.22	same basal level.
00:27:45.21	And this is an interesting finding.
00:27:47.05	This is a subset analysis looking at how different levels of activity would impact the efficacy
00:27:54.03	of the drug.
00:27:55.03	So, this is comparing patients who had had one relapse at the onset of when they were
00:27:59.14	coming into the trial in the previous year versus three relapses in the previous year.
00:28:04.22	And so you can see that patients who had high levels of disease activity, relapse activity
00:28:09.11	was inhibited down to the same level as everyone else.
00:28:12.07	So, it appears as though natalizumab has a very strong effect on patients with
00:28:16.23	highly active disease, inhibiting the... the relapse rate down to the same level.
00:28:24.14	And then, finally, this is the critical primary endpoint, at the second year of analysis.
00:28:29.04	And this was, how does it affect the overall progression of the disease in patients,
00:28:33.04	as measured by the standard EDSS scale?
00:28:36.24	And you can see that there's about a 54% reduction when looking at a sustained response over
00:28:41.28	six months.
00:28:42.28	Again, another subset analysis, looking at patients with highly active disease.
00:28:48.05	Again, there's an even stronger reduction, a 64% reduction, in the progression of EDS scores
00:28:54.22	in patients with... with more than two relapses and gadolinium lesions before they
00:28:59.15	came into the study.
00:29:02.28	And then, finally, this was not a point... this was not a formal endpoint in the study,
00:29:07.07	but nonetheless was a measure that was done.
00:29:09.23	And this is the MSFC scale.
00:29:12.22	And here you can see that the placebo patients... well, actually, what's interesting about this
00:29:16.08	scale is that it involves ability to walk, ability to have dexterity in the hands,
00:29:22.09	as well as cognitive function, a measure of cognitive function.
00:29:26.21	You can see that the placebo group maintained fairly stably over the two-year period, whereas
00:29:31.21	patients on natalizumab actually showed a level of improvement.
00:29:36.18	Importantly, these findings have been consistent across a number of studies.
00:29:42.04	These are actually looking at registrations and observational studies done by MS investigators
00:29:47.09	across the world, and very similar levels of efficacy with respect to relapse rate.
00:29:52.15	So, in summary, two years of treatment with natalizumab decreases disease activity, reduces
00:29:58.24	the risk of relapse by 68%, and reduces the risk of sustained disability progression by
00:30:05.04	42-54%.
00:30:06.04	To summarize the drug's safety in these clinical trials, these two-year clinical trials,
00:30:13.06	common adverse events were headache, fatigue, and arthralgias,
00:30:15.25	which are commonly seen in clinical trials.
00:30:18.22	There also were a few more infusion site reactions in natalizumab versus placebo.
00:30:24.21	Natalizumab actually causes a persistent anti... anti-natalizumab response in about 6% of patients.
00:30:32.04	And so, in these patients, actually, the drug loses this efficacy because the antibody response
00:30:36.26	causes it to be cleared very, very quickly, which is consistent with the infusion site reactions.
00:30:43.19	Importantly, there was a similar incidence of infections and malignancies between the
00:30:48.13	two groups.
00:30:49.13	In fact, if you look at this more closely, this is looking at the risk of all infections,
00:30:54.17	and this is for both studies combined.
00:30:57.05	So, both the monotherapy as well as the interferon therapy trial.
00:31:03.10	And you can see that there is no measurable difference in the rate of overall infection
00:31:08.16	between natali... natalizumab and placebo.
00:31:14.05	So, this is just the timeline of development, looking back in the beginning at 1990,
00:31:20.27	when the target was... of alpha 4 integrin was first identified for multiple sclerosis.
00:31:25.13	The antibody was humanized.
00:31:27.28	About five years later we started the clinical trials.
00:31:31.14	Biogen came in with Elan in about the year 2000.
00:31:36.01	And when developing a drug like this for multiple sclerosis, it's very expensive, because the
00:31:41.01	trials are large, there's a lot of MRI measurements, and so it's really important to have a partner.
00:31:45.18	And, in fact, Biogen was a great partner, because they already had a highly effective
00:31:51.03	drug for multiple sclerosis in the clinic and they knew a lot about... about the disease.
00:31:55.25	Shortly after the partnership was when we had the readout of the Phase II data,
00:32:00.16	you know, the day that... that changed my career.
00:32:03.01	So, I think that Biogen was very happy about that as well.
00:32:06.24	The phase III study was then shortly initiated.
00:32:09.20	The drug was approved in... in November of 2004.
00:32:13.14	The Phase... the second year of the Phase III study completed and read out shortly thereafter.
00:32:19.01	So, all said, 15 years from conception, 10 years of clinical experience involving 4,000 patients.
00:32:28.02	Now, in the second part of my talk, I'm going to discuss what happens next.
00:32:32.13	And this was just three months after the drug was approved.
00:32:35.15	This was in February of 2005.
00:32:37.23	The entire world for Tysabri changed.
00:32:41.03	So, in February of that year, we found our first two cases of PML.
00:32:45.21	And PML is a viral infection in the central nervous system that usually results in death
00:32:50.04	or severe disability.
00:32:51.04	So, it's a very serious infection.
00:32:55.01	Two patients in the MS trial had developed PML.
00:32:58.22	And amazingly, they had developed PML almost within a week of each other.
00:33:02.12	So, after ten years of clinical experience, two patients developed PML within a week.
00:33:06.23	And so this was alarming to everyone, because it... it suggested, well, maybe this is
00:33:11.04	just the tip of the iceberg, and half the patients could have PML.
00:33:14.13	So, dosing of the drug was suspended.
00:33:17.21	And we undertook a comprehensive safety evaluation at that point.
00:33:21.11	This was extremely complicated, because there were 3,000 patients involved or already
00:33:26.05	are still... are still in clinical trials, as well a number of patients who were just beginning
00:33:30.24	to take the drug from its approval.
00:33:34.04	So, what I'll talk about in my second part, in the second lecture, is how the medical
00:33:39.15	and regulatory communities working together with the companies established a path to allow
00:33:44.04	natalizumab to return, as well as the work that we've done since then to better understand
00:33:48.17	the safety of natalizumab and how PML can be monitored.
00:33:51.25	So, at this point, I just want to end by thanking all of the collaborators in the... both the
00:33:58.17	preclinical and clinical studies.
00:34:00.05	I...
00:34:01.05	I will at the end of my second lecture actually give a more detailed list of some of those contributors.
00:34:06.10	But needless to say, there were a ton of very talented people involved in the development
00:34:11.10	of this drug.
00:34:12.11	And also I want to thank the many patients who participated in the clinical trials.
00:34:17.22	Without their participation, it would be impossible to dev... to develop therapies for... for disease.