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Neuroscience Outtakes Part 2: Vanderbilt’s Jeff Conn on the Role of Academic Labs

On Monday, we highlighted outtakes from our interview with Michael Ehlers, Pfizer’s CSO for neuroscience research, for our story on the state of neuroscience R&D. Today, we wanted to offer a view from academia: Jeff Conn is head of the Vanderbilt Center for Neuroscience Drug Discovery, which in the past several years has generated a number of CNS drug candidates.

While Ehler is focused on the growing body of genetic information that could pave the way for new neuroscience targets, Conn’s lab is taking a somewhat different approach. By scouring the literature for evidence–in humans–of a molecule or target’s activity, the lab then sinks substantial resources into understanding the basic biology driving that activity and designing molecules to exploit it.

In depression, for example, R&D has been stalled by a lack of new targets. But Conn’s lab is intrigued by studies showing that ketamine, an animal tranquilizer (and club drug), swiftly and effectively reduces the symptoms of major depressive disorder. “When I talk to scientists at Vanderbilt, its an approach they’re using for their most refractory patients,” Conn says.

A laundry list of side effects makes wider use of ketamine improbable. As such, Conn’s lab is looking at ways to design molecules that produce the same kind of results on depression without the adverse effects.

Conn, a former Merck researcher, also discussed ways that discovery efforts inside academia can build a scientific case for CNS programs that pharma might otherwise overlook. Vanderbilt scientists spend “twice as much effort in basic science than for the drug discovery itself, and to me, that’s absolutely critical,” Conn says. When the team finds that molecules have different profiles in vitro, they spend a lot of time trying to understand how that will translate into adverse effects in vivo. “In pharma, you have to stay on such a narrow, direct path, that you have to ignore all that,” Conn says. In the academic lab, researchers take a longer, more methodical approach that entails optimizing many different molecules, then putting those in animals to understand what properties a final drug candidate needs to have.

That approach has enabled Vanderbilt scientists to tackle drug targets that have tripped up industry. “mGluR5 is a good example where, early on, we started seeing different properties of molecules in vitro,” he says. “Instead of putting blinders on and moving forward or ignoring it,” an avenue industry scientists are often forced to take, “we deliberately put a lot of effort into optimizing those properties.”

As a result, the Vanderbilt group and its collaborator, J&J, recently moved forward what Conn calls “very safe” schizophrenia drug candidates targeting mGlur5. “I don’t think we ever could have done that in my pharma days because its too far off the critical path,” he says.


Exploring Rational Drug Design

Medicinal chemists strive to optimize molecules that fit snugly into their proposed targets. But in the quest for potency, we often overlook the local physics that govern drugs’ binding to these receptors. What if we could rationally predict which drugs bind well to their targets?

A new review, currently out on J. Med. Chem. ASAP, lays out all the computational backing behind this venture. Three computational chemists (David Huggins, Woody Sherman, and Bruce Tidor) break down five binding events from the point-of-view of the drug target: Shape Complementarity, Electrostatics, Protein Flexibility, Explicit Water Displacement, and Allosteric Modulation….whew!

Selectivity Strategies

Selectivity Strategies for Rational Design | Credit: Huggins, Sherman, Tidor; J. Med. Chem.

Note: Before we dive into this article, let’s clarify a few terms computational drug-hunters use that bench chemists think of differently: ‘decoy’ – a test receptor used to perform virtual screens; ‘ligand’ – the drug docking into the protein; ‘affinity / selectivity’ – a balance of characteristics, or how tightly something binds vs. which proteins it binds to; ‘allosteric’ – binding of a drug molecule to a different site on an enzyme than the normal active site. Regular readers and fans of compu-centric chem blogs such as The Curious Wavefunction and Practical Fragments will feel right at home!

We’ll start at the top. Shape complementarity modeling uses small differences in a binding pocket, such as a methylene spacer in a residue (say, from a Val to Ile swap) to dial-in tighter binding between a target and its decoy. The authors point out that selectivity can often be enhanced by considering a drug that’s literally too big to fit into a related enzymatic cavity. They provide several other examples with a ROCK-1 or MAP kinase flavor, and consider software packages designed to dock drugs into the “biologically active” conformation of the protein.

Electrostatic considerations use polar surface maps, the “reds” and “blues” of a receptor’s electronic distribution, to show how

Affinity Optimization

Affinity Optimization - Black dot represents the optimal minimum energy between Coulombic forces (green) and desolvation penalty (blue) | Credit: Huggins, Sherman, Tidor; J. Med. Chem.

molecular contacts can help binding to overcome the desolvation penalty (the energy cost involved in moving water out and the drug molecule in). An extension of this basic tactic, charge optimization screening, can be used to test whole panels of drugs against dummy receptors to determine how mutations might influence drug binding.

Because target proteins move and shift constantly, protein flexibility, the ability of the protein to adapt to a binding event, is another factor worth considering. The authors point out that many kinases possess a “DFG loop” region that can shift and move to reveal a deeper binding cavity in the kinase, which can help when designing binders (for a collection of several receptors with notoriously shifty binding pockets – sialidase, MMPs, cholinesterase – see p. 534 of Teague’s NRDD review).

But these shifting proteins also swim in a sea of water and other cytoplasmic goodies. This means that drug designers, whether they like it or not, must account for explicit water molecules. The authors even suggest a sort of “on-off” switch for including the bound water molecules, but contend that more efforts should be directed to accurate modeling of water in these protein settings.

Finally, the authors weigh the effects of allosteric binding, the potential for a modeled molecule to be highly selective for a site apart from where the protein binds its native ligand. The authors consider the case of a PTP1B ligand that binds 20Å away from the normal active site, at the previously mentioned “DFG loop.” Since this binding hadn’t been seen for related phosphatases, it could then be used to control selectivity for PTP1B.

In each section, the authors provide examples of modeling studies that led to the design of a molecule. Two target classes recur oftenCOX and HIV inhibitors throughout the review: HIV protease inhibitors (saquinavir, lopinavir, darunavir) and COX-2 inhibitors (celecoxib), which have all been extensively modeled.

Two higher-level modeling problems are also introduced: the substrate-envelope hypothesis, which deals with rapidly mutating targets, and tailoring molecules to take rides in and out of the cell using influx and efflux pumps in the membrane. Since different cell types overexpress certain receptors, we can use this feature to our advantage. This strategy has been especially successful in the development of several cancer and CNS drugs.

Overall, the review feels quite thorough, though I suspect regular Haystack readers may experience the same learning curve I did when adapting to the field-specific language that permeates each section. Since pictures are worth a thousand words, I found that glancing through the docking graphics that accompany each section helped me gain a crucial foothold into the text.

Shedding Light on Cancer Cells

Quyen Nguyen is a surgeon at the University of California, San Diego who has worked with chemists to develop molecular beacon type dyes that light up when they come into contact with cancerous tissue or nerve cells. This could give surgeons a sort of chemistry-based augmented reality, showing them exactly where and where not to cut.

Amides: Humble But Useful

A heartfelt thank-you to Chemjobber and See Arr Oh for helpful discussions!

CENtral Science’s benevolent overlord, Rachel Pepling, has organized a blog carnival around the theme of “your favorite chemical reaction”. For the Haystack’s contribution, I thought it would be appropriate to write about a reaction medicinal chemists might find familiar. So I re-read See Arr Oh’s post about which types of reactions were really the most common in the med-chem toolkit. I decided on amide formation, which sits just about at the top of the list. I’m not sure it’s my favorite chemical reaction; I’ve got a special place in my heart for the Heck reaction (or Mizoroki-Heck reaction), though I’ve already blogged extensively about it. But every amide bond formation I ran in grad school worked. That’s justification enough for me!

If this necklace represented a peptide or protein, amide bonds would be the little metal links. (Shutterstock)

Amides are the chemical ties that bind amino acids together to form peptides and proteins. Amides also turn up in a variety of other small molecules that nature makes. So it’s not surprising that amides are frequently found in drugs. Take a look at University of Arizona chemist Jón T. Njarðarson’s poster of top brand name drugs and marvel at the amide-y goodness.

Amide bond formation isn’t accomplished by a single, archetypical chemical reaction– far from it. I thought I’d provide a brief overview of some classic chemistry in this area and then move into a selection of modern-day additions to the amide-construction toolkit. Continue reading →

LaMattina & Breslow to Talk Future of Pharma

We wanted to point Haystack readers to an upcoming event hosted in conjunction with our parent organization, the American Chemical Society, and Société de Chemie Industrielle: On September 14, our own Rudy Baum will moderate a panel discussion between former Pfizer R&D head John LaMattina and Columbia University chemistry professor Ron Breslow. The topic? “New Business Paradigms for Pharmaceutical Companies.” If the lively discussion today on twitter over the arrival of the “niche blockbuster” (or as Chemjobber coined “nichebuster”) model for pharma is any indication, folks are pretty interested in how drug firms are going to survive in a post-blockbuster era. For those living in the NY/NJ area, you can witness what is sure to be some great banter in person; for everyone else, feel free to sign up for the webcast.

Difficult C. difficile Infections – New Drug, New Targets

Trust your gut . . . scientifically speaking.  From belly-button bacteria to classification of signature microflora (all the various microbes that populate the intestinal tract), it feels like recent popular “culture” grows best in a petri dish. Many scientists now classify humans as superorganisms, meaning our survival depends on a host of “good” internal bacteria that digest fiber, make vitamins, and help the immune system. But what happens when these good bacteria suddenly get wiped out by a non-selective antibiotic? This sets the stage for a Clostridium difficile intestinal conquest.

Simple contact transmits this bacterium between patients in hospitals, causing antibiotic-assisted diarrhea, bloating, and potential colitis. When a patient is treated with a broad-spectrum antibiotic, C. difficile survive by forming spores with tough outer coats, only to thrive again when there are few other bugs in the gut with which to compete.

Two new players have recently entered the fight against the difficult C. difficile: first, Optimer Pharmaceuticals’ new narrow-spectrum antibiotic for C. diff. treatment, Dificid (fidaxomicin), approved in May 2011. This antibiotic macrolide belongs to the tiacumicin class of natural products, members of which have been known since Abbott first isolated compounds from fermentation broths in 1987. Dificid specifically inhibits Clostridium RNA polymerase enzymes; without these enzymes, gene transcription halts, and the cells die.

Clearing the infection is great, but wouldn’t it be nice to ease the intestinal pain while the drug takes hold?

Researchers at UTMB-Galveston might have found a good target for drugs that could do just that. In the August advanced online publications at Nature Medicine, Tor C. Savidge at UTMB-Galveston reports on human metabolites that can inhibit C. difficile toxins TcdA and TcdB, the major agents behind painful antibiotic-assisted diarrhea.  S-nitroso-glutathione, a nitroso (NO)-conjugated version of glutathione found in stool samples of infected patients, can “pass off” its NO group to the sulfur of a specific cystine amino acid residue in the toxins, shutting down their activity. The authors point out that instead of active site binding, the normal mode of action for most enzyme inhibitors, this NO seems to inhibit the toxins via an allosteric site, meaning they bind somewhere else on the toxin but still impair its function. Potency for in vitro inhibition is still in the high micromolar range (43-57 µm), but the study may point the way to the development of more selective NO-transfer drugs.

Sweet Science: Glucose Regulates Stem Cells, Cancer

Most people think of glucose, a humble sugar, as the fuel for several critical body functions, including muscle contraction, brain function, and a host of cellular processes. As it turns out, glucose function might also make a prime target for the development of regenerative medicine and cancer treatments.

Researchers at the Mayo Clinic led by Dr. Andre Terzic report in Cell Metabolism that glucose plays a pivotal role in “re-induction” of pluripotent stem cells (iPSC) from cells called fibroblasts.  These rapidly dividing cells, which normally build connective tissues like collagen, can be convinced to shutter their oxidative, mitochondria-based metabolism in favor of a glycolytic pathway, essentially changing the cells to iPSCs in the process. 

Terzic et al, Cell Metabolism 2011

How does one rewire cellular metabolism?  Expose mouse fibroblasts to a nuclear reprogramming kit (which uses viruses to rewrite nuclear DNA), and then grow them in a high-glucose solution. The scientists used 1H NMR metabolic profiling to monitor changes in cellular metabolism in this sugary environment, finding that in roughly one week they exhibit the same metabolic footprint as embryonic stem cells. No word yet on small molecule drugs that can supplant this process.  
In other glycemic news, Stanford Medical School researchers, led by Dr. Amato J. Giaccia, have reported a small molecule capable of inducing “synthetic lethality” in renal cell carcinomas, a common type of kidney cancer. Treatment of cancer by chemical synthetic lethality combines small-molecule inhibition and genetic mutation to selectively kill cancer cells dependent on these pathways.  Carmen briefly covered this advance in the August 8 issue of C&EN.

Two transporters for glucose, GLUT1 and GLUT2, control how kidney cells use sugar; however, genetic mutations cause cancerous cells to favor GLUT1. STF-31, a sulfonamide which targets GLUT1, selectively shuts down glucose transport to those cancerous cells lacking functional VHL tumor suppressor genes, a common mutation in renal cell carcinomas. But tumor cells are notoriously tricky. The Stanford scientists wondered if in the absence of the GLUT1 activity, cancer cells: might simply use an alternative pathway called oxidative phosphorylation to stay alive.  They found that adding excess pyruvate (fuel for the oxidative phosphorylation engine) could not compensate for glucose starvation, and RCC cells still died. Other non-specific glucose transporter inhibitors (fasentin or phloretin, which hit GLUT1 and GLUT2 indiscriminately) killed normal kidney cells as well as cancerous cells, which confirms overexpressed GLUT1 as STF-31’s RCC target.

Remedium Technologies Gets A Grip On Severe Bleeding

Maryland is serious about nurturing startups. Drahl/C&EN

In the last year we’ve covered many up-and-coming drugs for controlling the delicate balance between clotting and bleeding. But what happens when something—an injury or a major surgical procedure—overwhelms that system?

Controlling big bleeds is big business, from the battlefield to the operating room. This Monday, at the American Chemical Society’s Middle Atlantic Regional Meeting (MARM) in College Park, Maryland, I heard from Matthew Dowling, CEO of a startup looking to make its mark in that space. The company is called Remedium Technologies, and it’s developing chemically modified versions of a natural biopolymer to make improved materials for stanching blood flow.

Remedium is one of several companies getting on its feet with help from technology incubation programs the University of Maryland. Representatives from several of those companies, including Dowling, gave talks at a MARM symposium on the science of startups. Look here for the MARM session’s program- it includes other companies in the drug and vaccine space, including Azevan Pharmaceuticals (which C&EN wrote about in 2001 when it was called Serenix), Leukosight, and SD Nanosciences.

The biochemical pathway that regulates clotting can’t support severe injuries that lead to profuse bleeding, Dowling said Monday. While several treatments exist for this kind of severe injury, where sutures might not work to close a wound, they have drawbacks that Dowling thinks Remedium’s technology can address.

The company’s material of choice is chitosan, a biopolymer that can be scavenged from waste shells of shrimp or crabs. Chitosan wound dressings are already on the market, but they become saturated with blood and quit sticking to tissue after about 30 minutes, which can lead to more bleeding. As a bioengineering graduate student at Maryland, Dowling developed an alternative chitosan modified with hydrophobic groups that help it stick to tissues longer. This modified biomolecule is the basis of Remedium’s technology. The company likens the material to Velcro because it is the sum total of weak interactions between hydrophobic groups and tissue that help the material stick around, Dowling explains. Once the wound has had time to heal, the material can be gently peeled away.  The chemical structure of Remedium’s hydrophobic groups is proprietary; Dowling used benzene n-octadecyl tails in graduate school.

The company has two products in development- a modified chitosan “sponge” and a spray-on blood clotting foam. Neither of those products is yet available for purchase. In College Park, Dowling showed a video demonstrating how the modified chitosan makes blood congeal quickly, and how the effect can be reversed by applying alpha-cyclodextrin. In a second video, the sponge is tested on a bleeding pig that’s had a major blood vessel cut open. This presentation is similar to what Dowling gave Monday.

Dowling has been running Remedium full-time since he obtained his doctorate from Maryland in 2010—the company was founded while he was still in graduate school, and several classmates are also in the company’s management. The company has an exclusive license for the chitosan technology from the university, and has four patents pending. It has also won several business competitions, including Oak Ridge National Laboratory’s (ORNL) 2010 Global Venture Challenge. Dowling says the university’s technology incubation resources are what made it possible for him to start a company while still in grad school, from providing office space in a building just off campus, to regular meetings with staffers knowledgeable about navigating the regulatory and funding process.