Our “BWH Institute for the Neurosciences seed grant” on ‘A Screening Library to Accelerate Neurology Research’ was funded. This funding will allow us to begin building our collection of CNS drug-like molecules and distributing them to academic researchers.
The LDDN SMA team travels to Kansas City MO for the Annual SMA Research Conference and to meet up with our collaborators from Indiana University; Elliot Androphy, Sarah Custer and Anne Rietz.
Sarah presented ‘Binding to SMN is essential for the role of alpha-COP in supporting neuronal development” and Anne gave both a poster and talk on our ‘Novel small molecules that increase SMN protein and extend survival of SMA mice’.
There was also a little time to taste some Kansas culture.
Our collaborator, Professor Carmela Abraham launches Klogene Therapeutics. Klogene is a start-up company developing novel treatments for neurodegenerative diseases. Its first target is Alzheimer’s disease (AD). The company’s drug development platform is based on small molecule compounds that enhance the expression of the Klotho gene.
The Klotho enhancing compounds were discovered in a collaboration between Carmela Abraham at Boston University School of Medicine (BUSM) and the LDDN at BWH. Klogene’s initial drug discovery platform is based on an exclusive license of the BUSM/BWH intellectual property.
Our R21 grant to discover small molecules to treat Spinal Muscular Atrophy is awarded.
Optimization of a novel series of small molecules for the treatment of SMA with Professors Elliot Androphy at Indiana University and Christian Lorson at Missouri University.
Elliot’s group originally characterized the mechanism of alternative splicing of the SMN2 gene, which leads to skipping of exon 7 and failure to protect from motor neuron loss found in spinal muscular atrophy (SMA). We identified compounds that increase levels of the SMN protein that is deficient in SMA and optimization has led to the discovery of compounds with excellent brain penetration following oral dosing
Compound 1 has excellent potency and efficacy (Panel A). Orthogonal assays confirmed SMN2 activation in SMA patient-derived fibroblasts (~2.0-fold activation, Panel B). Compound 1 was stable in microsomes, and it had acceptable aqueous solubility, which facilitated formulation for PK studies in 40%PEG 400 + 60% (10% HP-β-CD in water). Most notably, following oral dosing at 30 mpk, compound 1 demonstrated excellent plasma (Cmax = 44 uM) and brain (Cmax = 19 uM) levels.
We believe compounds that emerge from the stringent in vitro and in vivo evaluation we are proposing will be of suitable quality for final pre-clinical evaluation and advancement into clinical studies to treat SMA.
Publication of Prof. Glenn Lin’s paper: “Restored Glial Glutamate Transporter EAAT2 Function as a Potential Therapeutic Approach for Alzheimer’s Disease” in The Journal of Experimental Medicine 2015, 212:319-332.
Kou Takahashi,Qiongman Kong, Yuchen Lin,Nathan Stouffer,Delanie A. Schulte, Liching Lai, Qibing Liu, Ling-Chu Chang, Sky Dominguez, Xuechao Xing, Gregory D. Cuny, Kevin J. Hodgetts, Marcie A. Glicksman,and Chien-Liang Glenn Lin
Abstract Glutamatergic systems play a critical role in cognitive functions and are known to be defective in Alzheimer’s Disease (AD) patients. Previous literature has indicated that glial glutamate transporter EAAT2 plays an essential role in cognitive functions, and that loss of EAAT2 protein is a common phenomenon observed in AD patients and animal models. In the current study, we investigated whether restored EAAT2 protein and function could benefit cognitive functions and pathology in APPSw,Ind mice, an animal model of AD. A transgenic mouse approach via crossing EAAT2 transgenic mice with APPSw,Ind. mice (e.g., see Panel C), and a pharmacological approach using a novel EAAT2 translational activator, LDN/OSU-0212320 (e.g., see Panel B), were conducted. Findings from both approaches demonstrated that restored EAAT2 protein function significantly improved cognitive functions, restored synaptic integrity, and reduced amyloid plaques. Importantly, the observed benefits were sustained one month following compound treatment cessation, suggesting that EAAT2 is a potential disease modifier with therapeutic potential for AD.
Blood-Brain Barrier in Drug Discovery: Optimizing Brain Exposure of CNS Drugs and Minimizing Brain Side Effects. Edited by Kerns and Li Published by Wiley and includes our contribution:
Chapter 19 Case studies of CNS Drug Optimization – Medicinal Chemistry and CNS Biology Perspectives.
Focused on central nervous system (CNS) drug discovery efforts, this book educates drug researchers about the blood-brain barrier (BBB) so they can affect important improvements in one of the most significant – and most challenging – areas of drug discovery.
- Written by world experts to provide practical solutions to increase brain penetration or minimize CNS side-effects
- Reviews state-of-the-art in silico, in vitro, and in vivo tools to assess brain penetration and advanced CNS drug delivery strategies
- Covers BBB physiology, medicinal chemistry design principles, free drug hypothesis for the BBB, and transport mechanisms including passive diffusion, uptake/efflux transporters, and receptor-mediated processes
- Highlights the advances in modelling BBB pharmacokinetics and dynamics relationships (PK/PD) and physiologically-based pharmacokinetics (PBPK)
- Discusses case studies of successful CNS and non-CNS drugs, lessons learned and paths to the market
The LDDN was formed in 2001 as described in this original article by Tom Reynolds.
One of five HCNR cores, the Laboratory for Drug Discovery in Neurodegeneration (LDDN) was launched in mid-2001 and is already running at 90 percent capacity, said HCNR director Adrian Ivinson. The lab grew out of the Partners Program in Neurodegenerative Diseases, which includes scientists at the BWH Center for Neurologic Diseases and the MGH Center for Aging, Genetics, and Neurodegeneration. Peter Lansbury, associate professor of neurology at BWH, chaired the task force for HCNR’s Core D, which evolved into the LDDN. Lansbury now serves as lab chair.
The lab uses high-speed robotic technology to screen FDA-approved drugs and larger collections of druglike molecules to gauge their effects on molecular and cellular processes thought to have causative roles in neurodegenerative diseases. The most active compounds are studied and modified in an attempt to develop compounds that may form the basis of a new drug. “The LDDN is structured like a biotech company, with a biology group that focuses on assay development, screening, and follow-up, and a medicinal chemistry group that focuses on optimizing compounds discovered through the screening efforts of the biologists,” said laboratory director Ross Stein. “So it might be appropriate to think of us as a not-for-profit biotech company.” But with some crucial differences. First, the lab’s personnel structure is designed with both a solid base of expertise in high-throughput screening and drug development (10 staff scientists) and a revolving door giving investigators from across the HCNR community access to the time and resources needed to test their most promising drug targets. In an unusual postdoctoral sabbatical program, postdocs compete for the privilege of spending six to 18 months working alongside the lab’s permanent staff to refine and test new drug ideas that come out of their own research.
Second, because the lab is noncommercial, it can devote substantial effort to research areas typically neglected by industry. “Industry usually follows a pack mentality, working on only a few ‘safe’ targets,” Stein explained. “For example, in Alzheimer’s, everybody and his brother are working on secretases involved in laying the amyloid plaque. But they are not working on enzymes that degrade the plaque–something we are working on. “Also, we work on targets that might not lead directly to therapeutics, but may lead to ‘tool’ compounds that allow more details about the disease to be uncovered.” This exploratory screening is generally seen as a waste of resources in industry, he said, but is highly valued within HCNR’s different scheme of priorities. Finally, he added, “We exist to serve the patient, not the shareholder.”
Ivinson similarly emphasized that HCNR exists to serve its member investigators, not to create a centralized bureaucracy controlling neurodegeneration research at HMS. Besides LDDN, the other cores include the Centers for Translational Neurology Research, Brain Imaging, Molecular Pathology, and Bioinformatics.
An Interview with Ross L. Stein, PhD Director, Laboratory for Drug Discovery in Neurodegeneration, Harvard Center for Neurodegeneration and Repair
Current pharmacological therapies for neurodegenerative diseases and psychiatric disorders leave much to be desired. This is particularly true for Alzheimer’s and Parkinson’s diseases, which are becoming increasingly troublesome for healthcare systems as the proportion of elderly people increases in the populations of developed countries. The prominence of psychiatric disorders in healthcare is perhaps best illustrated by the fact that antipsychotic drugs and antidepressants account for 22% of total sales for the world’s top 10 best-selling drugs. In Potential Breakthroughs in Neurotherapeutics: Alzheimer’s Disease, Parkinson’s Disease, Depression, Bipolar Disorder, and Schizophrenia, a new report from Cambridge Healthtech Associates Advances Reports, author Ken Rubenstein, PhD, provides a comprehensive assessment of ongoing innovative research in neurotherapeutics. In the report, he describes Harvard’s Laboratory for Drug Discovery in Neurodegeneration as a “poster child” for translational research. Following is the transcript of the interview with the center director.
Cambridge Healthtech Associates: Please give a brief description of LDDN, its purpose, history, activities, and your role there?
Dr. Stein: The history goes back to around 1998 when Peter Lansbury, a professor at Brigham, joined forces with some other folks from Brigham and Mass General who were working on neurodegenerative diseases and had grown frustrated with the reception that their ideas about drug discovery were getting from pharma. Essentially when they went to pharma with ideas about possible new ways to look at Huntington’s disease, ALS, or other neurodegenerative diseases, pharma didn’t show much interest primarily because of market size, especially in diseases like ALS or Huntington’s.
So they came up with the idea that if pharma wouldn’t work on their ideas, then perhaps they should consider doing drug discovery in an academic environment. That’s really when LDDN started. Around 1998 they got some funds from Partner’s Healthcare, which is an administrative arm that oversees both Brigham and Mass General; but the LDDN really got going in 2000 when an anonymous donor gave Harvard a large gift with the express purpose that it be used to do research in neurodegeneration in entirely new ways.
With that money Harvard set up what’s called the Harvard Center for Neurodegeneration and Repair. The LDDN was created as one of its core technologies. I was brought on board in May of 2001. Before that I spent about 20 years in pharma, both big and small pharma. I was head of a research group at Merck for about 5 years, and I came to the Boston area about 13 years ago. Essentially I was seduced by biotech, and I went into a company, which came to be called Proscript. We developed inhibitors of the proteasome. You may have heard of the proteasome inhibitor Velcade [bortezomib], a drug Millennium is selling for the treatment of cancer. I’m coinventor of Velcade.
In any event, the Harvard folks had the idea that to get the drug discovery thing really working well they needed to hire people with industrial experience, and that’s why I was brought on board to head it up. I decided to set this up a lot like a biotechnology company with a group of people doing assay development and high-throughput screening. I recruited Marcy Glicksman to head that group. Then I set up another group to do medicinal chemistry. To head that up I hired Greg Cuny. These folks recruited their own staff; they each have about 5 or 6 people reporting to them. All told we have a permanent staff of between 12 and 15 people.
With regard to the projects we work on, it became apparent that with the amount of money we had and the number of potential projects within the Harvard system that we’d be working on, we had to do something creative. We just wouldn’t have enough manpower to do all these projects. So we came up with this idea that we would take advantage of postdocs who are in the labs originating the projects and want to do drug discovery research. The model works this way. A postdoc may have been working in someone’s lab for a year or two. Perhaps he’s discovered a new receptor and thinks that if he can find an antagonist, it’s going to be a cure for Parkinson’s disease, let’s say. Well he comes to us with this idea. He makes the case that this receptor is, in fact, associated with Parkinson’s, and that if he could only get an antagonist, it would have the desired effect. After he convinces us of that, he would then have to give us some kernel of hope that we could develop an assay to actually look at this. It should be the sort of assay that could be miniaturized and made suitable for high-throughput screening. We then work with this person to optimize the assay and, with the optimized assay, do high-throughput screening. We have a compound collection of about 120,000 compounds now and if interesting hits are found from screening this collection, we have the capability to do medicinal chemistry to optimize the compounds so that they’re suitable for testing in vivo models of disease. This model of drug disovery has worked really well. During the past 5 years or so, we’ve probably worked on over 35 projects, and we’ve got a couple of them to the point where we’re doing our first animal testing.
CHA: What would happen to these projects afterward? Would the technology transfer people be looking to license them?
Dr. Stein: That’s exactly right. In fact, we’ve been actively engaged in business development almost since the point of inception, devising various models by which we could work with pharma and biotech. Only one of the models involves waiting for the stage when we have something to license. There are other business models we work with as well, but one of them involves getting to the point where we have a molecule that is part of a patent, which can then be licensed by pharma.
CHA: With regard to the postdocs in your program, is this looked on as a possible transition to industry for them?
Dr. Stein: That’s a good question, and that wasn’t our intention. The initial intention was to figure out how, with a screening and assay development staff of only 5 or 6 people, we could work on at least a dozen projects a year. As it turns out, though, it’s given these postdocs a lot of training in drug discovery research. It gives them their first glimpse of what an industrial atmosphere might be like. If they don’t like working with us, they certainly won’t like working in industry. I do recommend that this should not be their sole postdoctoral experience. I think that would be a mistake. However, if they’ve done basic research as a postdoc for a year or two, and they want to see if this research has practical value, and they’re thinking about a career in industry, this experience could be of real value to them.
CHA: The NIH has been talking about and taking some action toward translational medicine. They’ve been promoting it, and maybe even trying to shift the academic system for rewards and promotions. I have two questions. First, is the NIH happy with what you’re doing? Do they look to LDDN as a model of some sort? Secondly, how is LDDN playing with the mainstream faculty at Harvard?
Dr. Stein: Well, to address the first question, as soon as I got here, I started to work with the people at NINDS [National Institute of Neurological Disorders and Stroke]. Essentially I called them and said, look, let’s figure out together how I can get some of your money to help support what we’re doing here. In fact, a couple of years ago we were awarded a large grant allowing us to bring in 5 postdocs a year for 5 years. The original money from the gift was supposed to be used only in the Harvard system. So I asked NIH if we could figure out a funding mechanism so that I can open up what we’re doing to the rest of the country. So we have a grant now, where each year we can bring in 5 people from around the country to work with us under our postdoc-driven model of screening followed by medicinal chemistry. This grant is for 5 years at least; that’s 25 new projects. This first year, which will be coming to an end this fall, we’ve had 2 people from the University of California , San Francisco , 1 from Duke, and 1 from the University of Kansas; and our next batch of 5 people will be showing up this coming fall. So that model has worked extremely well. What’s happened too is that a lot of smaller grants were written in to help with medicinal chemistry, the screening effort, etc.
Now, in terms of the faculty, that’s an interesting question too. When I first arrived, they really didn’t know what to make of me. Who’s this guy from industry and what’s he doing at Harvard Medical School? They were skeptical about the whole idea of drug discovery in an academic environment, but now I think they see what we’re doing as really making a contribution.
CHA: Turning now to the science, regarding Alzheimer’s it seems as if a lot of commercial activity now is really centered around a paradigm that relates mainly to amyloid-beta and tau phosphorylation. There are a number of academicians who have some doubts about this, and are suggesting alternating paradigms. Do you have views on this?
Dr. Stein: We intentionally stay agnostic. Basically, we don’t work on a project unless a ‘customer’ comes to us with an idea. We have projects on APP [amyloid precursor protein] processing, on tau, and several cell-based assays related to AD. Understand that it’s not as if we’re firmly in the amyloid camp or the tau camp; we work on any good scientific idea that’s brought to us.
CHA: It sounds as if most of your projects fit into the mainstream paradigm. Two things that have stood out in preparing this report are microtubule dynamics and oxidative stress for neuroprotection. Do these pop up much in your purview?
Dr. Stein: Well, the microtubule dynamics would be in part regulated by tau and tau phosphorylation. We have one project on the dimerization of tau and one on the phosphorylation of tau. In terms of oxidative stress, we have nothing related to that, but I’m not sure there are any good molecules you could go after. It’s almost a black box in terms of drug discovery as far as I can tell. It’s hard for me to get my arms around oxidative stress in terms of real targets that you could go after in a drug discovery sense. But as I said, if someone would come to us with a project that makes scientific sense, and it has to do with oxidative stress, we’d certainly do it.
CHA: Based on your experience at LDDN, what do you consider the prospects for greatly improved new drugs for Alzheimer’s and Parkinson’s diseases entering clinical trials, say, in the next 5 years?
Dr. Stein: My hope for the first 5 years was to establish the place, to open it up to the country at large, and to get a couple of molecules into animal models. We’ve been lucky that we’ve accomplished those goals. In the next 5 years, the goals are to have some major deals with pharma, and to get a molecule ready to go into humans. If we could get that in the next 5 years, I’d really be happy about it. The only way we’re going to get molecules into people is to work with pharma; that’s something we can’t do on our own, of course. We’re not set up to do the work involved in the development phase of R&D. I think we can do discovery pretty well, but for development we’ll have to work with pharma.
You used the word ‘greatly.’ That’s hard to say. We’re definitely going after molecules that are disease-modifying and completely different from molecules on the market now. All the drugs in neurodegeneration that people take today only treat the symptoms. None of our projects is of that sort. They’re all in the realm of disease-modifying agents. So if any of these would work, if any of these get into people, I think it would have a real impact.
CHA: Yet it looks like a lot of the academic work that’s coming out is more in the realm of neuroprotection, which might be considered more symptom-related rather than going after the root cause.
Dr. Stein: I’d say it’s the opposite. If you had a way to protect neurons from degradation, that’s definitely at the root of disease. The problem with such agents is, at what point in the person’s life do they take a neuroprotective agent? That’s the question. When do you give a neuroprotective agent, when a person’s 15 years old, so the neurons are protected for the rest of their life? That’s a really tough thing to do. Again, I’m not sure what you do with neuroprotective agents.
CHA: Perhaps you could identify early Parkinson’s disease patients and neuroprotect before the disease progressed further.
Dr. Stein: That’s a possibility. Then it’s the issue of coupling really good diagnostics with those sorts of agents. You’d need some means to diagnose these diseases well in advance before symptoms show up. That scenario, I think, should be pushed a lot more.
CHA: Do you see any other institutions replicating what LDDN is doing?
Dr. Stein: Yes, I get calls from time to time from people who are trying to set up things like this. It’s tough; it’s really expensive to set up. We were extremely fortunate in getting this gift to kick things off. I think there’s going to be more and more interest in this sort of thing. One thing that makes us unique is that we have medicinal chemistry, screening, and in-house industry expertise; and that’s a good combination of skills to have. I recommend to people that if they decide to set this up, they need more than screening. A lot of people are doing screening in academics; the cost of automation has come down and it’s possible to buy chemical libraries fairly cheaply. There appears to be a lot of screening going on now in academics, for better or worse. What these groups really need is people who’ve done drug discovery before, and they need the capability to do medicinal chemistry.
CHA: Do you see any signs that universities are moving toward training people in some of these things?
Dr. Stein: I think there are some places where this sort of training is going on, but not very many. This is the sort of the thing best done on the job. For example, good medicinal chemists are always those who are excellent synthetic chemists. They go into industry, and there they’re trained during their first 3 to 5 years to transform themselves into medicinal chemists. That’s probably a good way to do it.
United Against ALS
Collaborations can be a bit of a crapshoot. Those involved can never really be sure whether anything will come out of their efforts. Sometimes, though, even if the study falls apart, the experience of working together can be valuable enough. That was the case for Benjamin Wolozin, from the Boston University School of Medicine, and Marcie Glicksman, co-director of the Harvard Neurodiscovery Center’s Laboratory for Drug Discovery in Neurodegeneration. When Wolozin and Glicksman worked together on a study searching for LRRK2 inhibitors, they didn’t get the results they were hoping for, and the project ended. But when Glicksman’s center sent out another request for collaborators on a study to find therapeutics for amyotrophic lateral sclerosis, Wolozin was quick to respond.
Glicksman’s LDDN is a “one-stop shop” for high-throughput drug discovery, Wolozin says. At any given time, the center is in the midst of 15 to 20 projects, all in various stages of development, and all of “The whole idea is that we provide the drug them with collaborators. discovery expertise, having come from industry and been involved in multiple drug discovery projects,” Glicksman says. “Each of our collaborators brings the science and we work with them on how to convert their science into a drug discovery project. It’s very complementary.”
“They have the robotics and the medicinal chemistry in place, so you don’t really have to go too far,” Wolozin adds. “It’s easy to see how to move things forward, instead of trying to figure it out at every step.” Boston University is working on building a setup much like LDDN’s, Wolozin says, but currently doesn’t have the kinds of resources to do the robotic screening and high-content or high-throughput assays for these types of studies, nor the personnel to help carry them out. But beyond that, the two collaborators say, they simply enjoy working with each other. “I find it really easy to work with Marcie,” Wolozin says. “I like it.” Glicksman agrees. “Ben is a great collaborator.”
Stop the Aggregation
The ALS project itself is still in the early stages. The disease is characterized by an abnormal accumulation in motor neurons of insoluble and misshapen proteins, the major component being Tar DNA-binding protein 43. In 2006, researchers at the University of Pennsylvania first identified TDP-43 as the largest component of the inclusions found in the neurons of many ALS patients. It is thought that inhibiting the accumulation of TDP- 43 might also prevent these inclusions from developing — and slow down the process of neuro-degeneration.
According to Wolozin, the researchers are currently using cell-based assays and highthroughput screening to slog through LDDN’s collection of 150,000 compounds to find molecules that inhibit the aggregation of TDP-43 and the formation of inclusions that cause not only ALS, but fronto-temporal dementia as well. Using a neuronal cell line that stably expresses GFP-tagged TDP-43, Wolozin says, the team will test hits to determine whether they inhibit stress granule-formation, which is linked to TDP-43 aggregation.
“We have a couple of compounds that are giving an interesting phenotype, which we’re looking at in more detail,” Glicksman says. The team will most likely take the next three or four months to test about half of the compounds in LDDN’s library and then do characterizations of the hits they see. Glicksman says they’re also performing an assay optimization in tandem with the study. Wolozin says he actually started with the US Food and Drug Administration’s library of compounds — but didn’t get any promising hits — before moving to LDDN’s library. With LDDN’s compounds, he has identified molecules that actually stimulate inclusion formation instead of inhibiting it.
“One of the benefits, and the drawbacks, of the high-content screening is that the compounds we’re looking at might act by directly interfacing with TDP-43 and they could also act on the biochemical processes that regulate TDP-43 inclusion formation, so there’s a lot of ways of interfacing there,” Wolozin says.
But even the identification of the compounds that do the exact opposite of what the researchers want them to do is valuable information, he adds. Those compounds have proven to be “biologically interesting” and could provide clues as to which compounds will work to reduce inclusions. Some of the compounds that they have tested have shown a small reduction in inclusions, some up to 50 percent, but the researchers are looking for “much better than that,” Wolozin says. The researchers are keeping their options open, in case they don’t find what they’re looking for. “We can either mix and match or use medicinal chemistry” to combine compounds with less than 100 percent effectiveness, Wolozin says.
Though they are not far enough along to reach any conclusions, “I really think we’re going to find what we want,” he says. “I would be surprised if we didn’t come up with some hits.”
Glicksman is also hopeful. “Often what we look for is a pretty novel approach, so it’s fairly high risk and there’s going to be a certain drop-off,” she says, much like the first collaboration between Wolozin and LDDN. However, “this one is going well so far,” Glicksman adds.
Stop the Aggregation
The study has already run for longer than the original funding anticipated. The support for this particular project comes from the ALS Therapy Alliance and Project ALS, Glicksman says, and LDDN has support for the study for two years. During the first year, Wolozin’s postdoc Peter Lee’s salary was covered by the money from the ALS foundations. Now, says Wolozin, “I’m funding him myself.” Wolozin and Glicksman are continuing to collaborate outside the lab and are coming up with ideas to find more money to continue the study, which may have an impact beyond the treatment of ALS. Depending on the compounds they find and the mechanism of the compound’s action again TDP-43 aggregation, the research has implications for treatment of multiple neuro-degenerative diseases, a possibility that is “certainly on our minds,” he adds.
“Additional fundraising is continuing to keep this going and we will likely apply for NIH grants with Ben to get additional funding,” Glicksman says. “A project continues if it meets specific criteria to move forward … so a time limit is not imposed as long as the project progresses.”