Hiding hydrogen bonding groups in large druglike molecules

As most medicinal chemists probably know, one of the major challenges in the field currently is to find out the principles that govern the ability of "large" molecules - especially ones violating the famous and often sacrosanct and overused Lipinski's Rule of 5 - to become drugs. Larger molecules like natural products and macrocycles are thought to be much better modulators of protein-protein interactions and therefore considered to be part of the next generation of important drugs.

In meeting this challenge a widely agreed-upon obstacle is cell permeability. Because of their large size and exposed polar surface area larger molecules find it much harder to get across lipophilic cell membranes. But nature as usual has thrown many exceptions at us. The quintessential example is the immunosuppresant  cyclosporin which is a master of conformational camouflage; in water it opens up to expose its polar groups for optimum solubility while inside a lipophilic membrane it flexes itself and folds up to form intramolecular hydrogen bonds and hide its polar surface area. It thus thrives in both environments.

Forming intramolecular hydrogen bonds to enable compact conformations and hide polar surface area is in fact a dominant strategy used by natural products to enable membrane permeability. Not surprisingly this is a highly sought-after strategy in drug design too, especially for larger molecules which otherwise may not get across membranes. Unfortunately for now, it's much easier to retrospectively identify molecules which adopt intramolecular hydrogen-bonded conformations than engineer such interactions prospectively into a potential druglike compound.

Nonetheless, it is only through such retrospective studies that we can learn the principles which will someday hopefully allow us to design drugs which can fold to form intramolecular hydrogen bonds on demand. That's why I was pleased to see this study in J. Med. Chem. from a joint academic-industrial group in Sweden. The authors were looking at a subset of molecules from a screening collection at the Broad Institute that exhibited activity against T. cruzi, the parasitic agent that causes sleeping sickness. They had a compound with three chiral centers and one double bond which could be cis or trans. They consistently found out that the cis compounds had better cell permeability at increasing pH values as well as better solubility than the trans.

Computational modeling identified a low-energy conformation with a hydrogen bond between an amide NH group and a tertiary amine in its neutral form. This hydrogen bond was present in both conformations but, crucially, it was about 2.6 kcal/mol higher in energy for the cis (8) compound than for the trans (1).

In a nutshell it means that the cis compound would prefer not to form that hydrogen bond and instead expose the polar groups to solvent, accounting for its better solubility. The higher energy for the cis compound probably comes from two extra gauche interactions in the hydrogen bonded conformation compared to the similar one for the trans. I always love it when one can invoke principles of basic college organic chemistry and conformational analysis to explain such observations. The calculations were also supported by solvent-dependent NMR studies.

The cis compound also had a higher pKa, which could account for its higher permeability at pH values approaching physiological conditions. This would be because with its higher pKa value the cis compound would be deprotonated more than the trans as the pH increased; deprotonated, uncharged species can diffuse better through a hydrophobic membrane. The difference in pKa can also be accounted for by the higher energy of the hydrogen bond; remember that pKa is a thermodynamic variable related to equilibrium constants through the Henderson-Hasselbalch equation.

I really like these kinds of studies where an examination of basic chemical properties supported by relatively simple computational calculations can help us rationalize and even predict important druglike properties like solubility and membrane permeability. If nothing else they provide a few more valuable data points on the way to prediction nirvana. Worth a look.

Digital features in the Journal of Medicinal Chemistry

J. Med. Chem. has an editorial describing how the journal plans to make structural information in papers more accessible in a digital format. Most of this would entail having spreadsheets of compounds with SMILES strings supplied along with the paper. I applaud this effort since the ability to view and manipulate structures in 2D and 3D is one which chemists have long coveted but not realized until now.

With this issue, therefore, the Journal of Medicinal Chemistry embarks on an initiative to integrate the chemical information it publishes with digital media and chemical software. The first step in this effort is the linkage of articles with molecular formulas in a format that can be generated by and read into chemical software. The immediate benefit is that readers will be able to transfer compounds from articles into chemical drawing programs, databases, and computational tools. In the near future, this association of articles with computer-readable chemical formulas will provide a foundation for the creation of new journal features that will bring chemistry to life on the Web and in digital readers and tablets. 

Authors are invited to use their existing chemical drawing programs (e.g., ChemDraw, ACD ChemSketch, Marvin Sketch) to generate a computer-readable SMILES formula for each compound presented in their articles. These formulas are then pasted into a simple spreadsheet, along with basic information about each compound. Ideally, this spreadsheet will provide a machine-readable version of the key data presented in the article’s tables. 

Ideally I would like a PDF manuscript to be fully interactive. You should be able to right-click on a protein structure and be able to directly view it in the PDB. Better still would be an ability to rotate and zoom in on the 3D structure inside the PDF format in real time. A similar capability for small molecules would also be immensely useful; being able to look at structural parameters (bond lengths, angles etc.) and conformations extracted from manuscripts in real time would be quite enabling. We aren't there yet but I have little doubt that it's only a matter of time. This is a propitious beginning.

What happens when chemists have nothing better to do on a Wednesday afternoon?

• This. It started with me posting a link on Facebook to an awesome recent paper describing physicists' efforts to reweigh the electron to an accuracy of one part in a trillion. The great Aaron Finke - of the regretfully dead Carbon-Based Curiosities - then weighed in.

   

  Aaron Finke This is what happens when physicists are bored.
  
  "So... uh... whatcha up to?"
  "Nothin, just Facebooking and eating a Hot Pocket"
  "...cool, cool. Wanna... wanna reweigh the electron?"
  "Yes. Yes I do."

  
  
• 

  Ashutosh Jogalekar Computational chemist version:
  
  "So…uh, whatcha up to?
  "Nothin, just twittering and eating a Hungry Man chicken meal."
  "…Cool…Hey, wanna parametrize that aziridine N-C bond with some stretch-bend cross-terms?
  "You bet!"

  
  
• 

  Ashutosh Jogalekar Synthetic chemist version:
  
  "So…uh, whatcha up to?
  "Nothin, just textin and eating an Amy's frozen teriyaki bowl." (clearly the synthetic chemist is more evolved)
  "…Cool…Hey, wanna repeat Woodward's B12 synthesis, but this time using RCM in some of the steps?"
  "Oh God yes!"

  
  
• 

  Aaron Finke Geologists:
  
  "So, uh, whatcha up to?"
  "Not much, just youtubing and eating taco bell"
  "Cool, cool. Hey, wanna get shitfaced?"
  "I already am"
  "Oh ok"

  
  
• 

  Ashutosh Jogalekar Clearly there's a fine line between being bored and being shitfaced.

  
  
• 

  Aaron Finke that's the nice thing about being a geologist
  
  you never have to be in a hurry

  
  
• 

  Ashutosh Jogalekar At least until the continents start coming together again. Then things get truly shitfaced in a hurry.

  
  
• 

  Aaron Finke "So, uh, whatcha up to?"
  "Nothin, just waiting for the continents to collide together again"
  "Cool, cool... how long will that take?"
  "Probably a couple hundred million years."
  "Cool, cool... wanna get shitfaced?"
  "I already am"
  "Okay"

  
  
• 

  Aaron Finke "Well, have fun. I'm gonna go help weigh that electron with the physicists"

  
  
• 

  Ashutosh Jogalekar *Geologist returns after helping physicists reweigh the electron, computational chemists parametrize every single molecule that can be built from 12 heavy atoms and synthetic chemists make vitamin B12 from a simple, four-carbon compound isolated from bat feces*
  
  "So, uh, whatcha up to?"
  "Nothin, just waiting for the continents to collide together again"
  "Cool, cool... how long will that take?"
  "Probably a couple hundred million years."
  "Cool, cool... wanna get shitfaced?"
  "I already am"
  "Okay"

  
  
• 

  Aaron Finke Why am I not a geologist

  
  
• 

  Ashutosh Jogalekar Because you would rather spend your time much more productively on the 67th synthesis of aquabatguanine using hammer and tong chemistry?

  
  
• 

  Aaron Finke what am I, Chinese?

  
  
• 

  Ashutosh Jogalekar Of course not. The Chinese made aquabatguanosine.

  
  
• 

  Aaron Finke ...again.
  
  well this might be the dumbest conversation on your FB wall ever. You're welcome.

  
  
• 

  Ashutosh Jogalekar Eminently postworthy (with your permission, naturally: I am planning to name it "When chemists have nothing to do on a Wednesday afternoon"). May even help to resurrect the dead CBC.

  
  
• 

  Aaron Finke do it

On making mistakes

In postulating an incorrect structure for DNA, Linus Pauling surprisingly committed an elementary chemical blunder (Image: pauling blog)

In the latest issue of the New York Review of Books, Freeman Dyson has a nice review of Mario Livio's readable book on scientific blunders committed by great scientists. The book is important reading for anyone who wants to understand the true history of science as a process of fits, starts, blind alleys, occasional great successes and of course, many blunders. Livio focuses on five famous scientists - Charles Darwin, Lord Kelvin, Linus Pauling, Fred Hoyle and Albert Einstein - who committed important mistakes. These mistakes sometimes set the field back but they also inspired other scientists to keep on looking and discovering new things. Scientists often build their theories and discoveries on the backs of other failed theories and discoveries. Just as respectable civilizations are often built on the bones of dead ones, respectable science is often built on the bones of scientific failures. And just like the natives are forgotten long after the settlers are celebrated, scientific failures get ignored at the expense of successes even when they are important in explaining the very existence of the successes.

Each of the blunderers in Livio's story blundered in a different manner. Darwin came up with a wrong theory of blending inheritance that he himself realized was acutely lacking in explaining real-world data. Mendel then discovered the right rules for inheritance and initiated a bonafide revolution in science. As Dyson explains, Mendel could improve on Darwin in no small part because he understood statistics and the law of averages better than the self-professed mathematically deficient Darwin. Lord Kelvin made his big blunder when he came up with wrong - and short - ages for both the sun and earth and thereby set up a significant obstacle to Darwin's theory of natural selection which demanded huge tracts of geological time to have passed for the evolution of species. The biological evidence was too overwhelming for Darwin to admit defeat but he clearly could not answer Kelvin's challenge. It was only in the middle part of the twentieth century when the fission and fusion processes powering radioactivity and the sun were worked out that Kelvin's question was posthumously addressed.

Fred Hoyle committed his major blunder and held on to a wrongheaded theory of the origin of the universe until his death. An early reason for Hoyle's recalcitrance in accepting the Big Bang was what he thought was the sheer audacity and fantasy of the theory, with the whole universe seemingly being conjured up from nothing in a flash. This aspect of Hoyle's thinking reminds me of Arthur Eddington's failure to take Subrahmanyan Chandrasekhar's theory of gravitational collapse seriously because he was convinced that there must be a law of nature preventing such a collapse. But the laws of nature are immune to our wishful thinking. The question of what came before the universe is still something that we grapple with, but every important discovery since the 1964 discovery of the cosmic microwave background has validated the Big Bang theory. Hoyle was certainly brilliant enough to have understood this evidence and he demonstrated his great scientific talents when he co-authored a seminal paper on nucleosynthesis with three other scientists. Hoyle thus stands as a curious example of someone who was in equal parts a reactionary and a maverick, not afraid to speculate on everything from extraterrestrial life to artificial intelligence but somehow never warming up to a revolutionary theory of the universe, even when it was supported by copious evidence.

Linus Pauling's mistake was of a different kind and rather hard to understand since it showed an embarrassing lack of knowledge of fundamental chemistry. Coming from someone widely considered to be the greatest chemist of the century this was odd, to say the least. After publishing his groundbreaking papers on the structure of proteins Pauling turned toward DNA and got embroiled in a race to decipher the structure of this all-important molecule with James Watson and Francis Crick. Although the race was perceived much more as such by the duo, Pauling certainly understood the importance of the problem. And then he famously committed an elementary chemical mistake. He published a paper in which the phosphates in DNA pointed inward and were held together by hydrogen bonds. Any good college chemistry student would know that at the pH inside the body (7.4) such hydrogen bonds would not exist and the oxygen atoms would be negatively charged, making them more likely to point outward into the ionic embrace of water. In his memorable book "The Double Helix", James Watson points out how his jaw dropped when he saw the mistake Pauling had made; ironically it was by consulting Pauling's classic "College Chemistry" textbook that he and Crick confirmed the error. As Watson put it, a graduate student under Pauling who made the same mistake would have probably been considered persona non grata at Caltech.

Why did the greatest chemist of the twentieth century miss such an elementary chemical fact about DNA? Even today the reasons are not completely clear. One reason could be that by the early 50s Pauling was much more concerned with nuclear disarmament than serious science, although he kept on publishing prolifically until his death. He could simply have been distracted from pursuing the DNA structure with the kind of full-time zeal that Watson and Crick did. The other reason is that he just missed the obvious. While this may sound surprising, it's a mistake that famous scientists who think out of the box can sometimes make. When it came to cracking the structure of proteins Pauling used a brilliant counter-intuitive approach. When it came to DNA the solution demanded a much more commonsense approach, and Pauling might have been still bogged down in protein structure for his mind to shift to this new kind of thinking. The last possible reason is also the most mundane; Pauling lacked the kind of high-quality structural data from x-ray diffraction that Watson and Crick got (some would say pilfered) from the technically accomplished Rosalind Franklin. When Watson saw the x-ray photographs he recalls feeling his pulse race, convinced that he had clinched it. Sometimes good data is all that separates a brilliant blunder from brilliant glory.

And then there's Albert Einstein whose brilliant blunder seems to indicate a lack of courage rather than a lack of scientific expertise. A lack of courage is another reason why scientists sometimes make important mistakes. In Einstein's case it was his injection of a fudge factor, the cosmological constant, to keep the universe static. Alexander Friedmann and Georges Lemaitre on the other hand had the courage to explore the logical solutions of Einstein's field equations, many of which pointed to an expanding and non-static universe. Einstein who had been a bold revolutionary when he came up with relativity turned out to be a conservative when clearly stating the possible consequences of relativity for the entire universe. In one sense this could be seen as the beginning of Einstein's reactionary streak, marking the time when he started opposing quantum mechanics and the picture of reality it presented. The ultimate irony of the fudge factor, as is now well known, is that it was resurrected by the discovery of the accelerating expansion of the universe and the postulation of dark energy.

As Dyson says, mistakes in science are essential, especially when you are exploring a new field on the cutting edge. No human mind is so all-knowing and perfect that it can cut through the fog of uncertainty and blunder to the solid heart of reality in one fell stroke. Especially at the beginning of a novel direction of research scientists should be liberally allowed to make mistakes. At the end of his review Dyson talks about a blunder he himself made pertaining to the incorrect prediction of the non-existence of charged weak bosons. He will probably agree that he has been so successful in science because he was allowed to make mistakes. Part of making mistakes is simply being able to generate lots of ideas; as one of the blunderers in Livio's book, Linus Pauling, put it, in order to have good ideas one must first have lots of ideas and then throw the bad ones away.

One of the most troubling casualties of the current climate of reduced science funding and flagging interest in science is that young scientists are afraid to make mistakes and therefore to generate lots of ideas. Funding agencies give them only a limited amount of money and ask them to work on "safe" problems; these are both constraints that reduce their appetite for risk-taking. Risk-taking has been one of the most important ingredients in the success of the United States as a leading scientific and technological power. Making mistakes is important not only in science but in business; think of how many computer, aircraft or skyscraper models were tried, tested and discarded before entrepreneurs came up with the correct ones. And it's a process that continues unabated. Once you ask a scientist to stop making mistakes you stop him or her from discovering. The stories of the scientists highlighted by Dyson and Livio as well as countless other episodes from the history of science make this fact clear. We ignore it at the risk of weakening the entire scientific enterprise.

First published on the Scientific American Blog Network.

Free online medicinal chemistry course at Davidson College

I wanted to alert interested readers to an introductory course on medicinal chemistry by Prof. Erland Stevens at Davidson College that seems to cover pretty much every basic and important aspect of drug discovery and medicinal chemistry that I can recall. 

As Prof. Stevens mentioned in an email, "The overall goal of the course is to get a student up to speed to watch a medicinal chemistry lecture (and maybe even ask a question at the end)." While the material may be a little too basic for readers of this (or Derek's) blog, it seems quite valuable for high-school or college students, and even for more experienced professionals who might want to brush up on their drug science.

The best part is that the course is free and available online.

Here's the website: https://www.edx.org/course/davidsonx/davidsonx-001x-medicinal-chemistry-1220

And here's the list of topics:

Week 1 – brief history of medicinal chemistry, introduction to drug development process and regulatory approval

Week 2 – proteins (enzymes and receptors) as drug targets, enzyme inhibition, ligand-receptor binding theory

Week 3 – pharmacokinetics (compartment models, Vd, clearance)

Week 4 – metabolism, phase I, phase II, prodrugs, genetic variability

Week 5 – drug-target complementarity, drugs as part of chemical space, chemical libraries

Week 6 – lead discovery, screening, filtering hits by metrics/structural alerts/predicted PK, SOSA, natural products

Week 7 – lead optimization, functional group replacements, isosteres, directed libraries, peptidomimetics

Syngenta, atrazine and keeping the science separate from the policy

The New Yorker has an excellent piece of reporting on the efforts of Tyrone Hayes, a UC Berkeley biology professor and his efforts to investigate potentially very important and deleterious effects of the herbicide atrazine on sexual dimorphism in frogs. In some of his experiments male frogs seemed to develop female genitalia. The major part of the piece is about how Syngenta - the maker of the multibillion dollar herbicide - tried to discredit Hayes. Ample supporting evidence is provided in internal memos and emails released as part of a law suit.

Many aspects of the story are worth thinking about but one of the most important ones is how such stories always risk the danger of conflating unethical behavior by companies with the underlying science. Syngenta shenanigans reported in the article are clearly unacceptable and stifling, but the message about atrazine is far more ambiguous. The piece points out several questions that the EPA raised about Hayes's studies (as well as Syngenta's), including proper statistical analysis and the extrapolation of amphibian studies to humans.

The important point is that these are valid and critical questions, even if Syngenta was using them to discredit Hayes (at one point one scientist dismisses statistical concerns as "routine", as if routine meant trivial). The motives of those wanting to use science to their own ends does not automatically affect the validity or lack therefore of the science itself. This is something that few environmentalists, in my experience, appreciate. Fortunately some do; for instance I have had commenters on my posts on GMOs explicitly saying that while they do support the science showing the safety of GMOs, they are much more concerned about the bullying and muzzling tactics used by companies like Monsanto. Sadly such commenters are precious and few.

The Syngenta/atrazine story falls in the same category. The company clearly used muzzling and shady tactics on Hayes but the verdict of atrazine's effects on human populations is clearly out there. In 2010 the EPA ruled out banning the herbicide for want of better evidence, and its decision only shows you how complicated it is to link the effect of any chemical to environmental or human damage. Personally - and I can't say I have reviewed the evidence in detail - I think Hayes is on to something but it's not certain exactly what.

I don't doubt that this article will spark furious allegations against Syngenta. But those who want to participate in this debate should keep something very simple in mind; science kowtows to no policy, even one designed to denigrate it. In your zeal to prosecute human being or corporations for unacceptable or criminal behavior, make sure that science does not become a casualty.