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Drug Development

Giving atropisomers another chance

Rising to the challenge, drug discovery chemists are embracing the dynamic molecules once deemed unmanageable

by Tien Nguyen
August 19, 2018 | A version of this story appeared in Volume 96, Issue 33


Structure of two atropisomeric forms of 6,6'-dinitro-2,2'-diphenic acid.

Although they were discovered almost a century ago, atropisomers are only now having their heyday. Uncovered by James Kenner and George Hallatt Christie at the University of Sheffield in 1922, this exotic class of chiral compounds revealed itself to the duo in a deceptively simple molecule.

Assigning atropisomers

When it comes to determining the absolute stereochemistry of an atropisomer, chemists may find themselves out of practice. San Diego State University’s Jeffrey L. Gustafson, who has studied these molecules since graduate school, recalls teaching the technique to a room full of professors during a job talk. Shown is a primer for assigning stereochemistry to 1-1′-bi-2-naphthol (BINOL)—the chiral ligand wunderkind with a rotational half-life of over 5 million years. Gustafson says to first imagine peering down the compound’s axis of chirality. Then prioritize each group using the rules that apply to point-chiral molecules, in which atoms with higher atomic numbers get higher priority (with one being the highest). If pointing from one to two to three takes you counterclockwise, the molecule is an S enantiomer; if clockwise, it’s the R enantiomer.

Source: Adapted from Jeffrey Gustafson

The compound, dubbed 6,6′-dinitro-2,2′-diphenic acid, consists of two aromatic rings lashed together by a carbon-carbon bond that is flanked by a carboxylic acid and nitro group jutting from each ring. In typical molecules, rings linked with a single bond would spin freely. But as the rings in this infamous compound try to rotate, the molecular groups projecting from them clash, jamming the molecule’s rotational motion and creating asymmetry.

Depending on the position in which the molecular rotor locks, two distinct forms of the compound can appear. Kenner and Christie confirmed atropisomers’ existence by crystallizing salts of both forms. The phenomenon, which was later found in macrocycles as well, was formalized in 1933 by biochemist Richard Kuhn, who coined the term “atropisomer” from the Greek word atropos, meaning “without turn.”

But atropisomers are more dynamic than their etymology suggests. And that’s where the trouble lies for chemists. Depending on an atropisomer’s size, shape, and electronic properties—and external factors like solvent and temperature—one chiral form can overcome the rotational energy barrier to morph into the other form. This racemization process can take several seconds, months, or even decades.

For years, atropisomers’ changeability meant they couldn’t be trusted as stable compounds for drug development. As chemists have learned through tragedies like the birth defects caused when the chiral drug thalidomide racemized in pregnant women, losing control over a molecule’s configuration can be fatal.

Yet pharmaceutical chemists, citing the need to explore a larger chemical space for drug candidates and the promise of increased potency, are starting to embrace these molecules—or at least attempting to.

In March, the American Chemical Society national meeting in New Orleans hosted a ballroom-sized session devoted to atropisomers in drug discovery. Soon after, one pharmaceutical company unveiled the large-scale synthesis of a compound boasting dual atropisomeric chiral axes that’s making its way through Phase II clinical trials. And each month seems to bring new publications sharing preclinical results from atropisomers as chemists increasingly turn to these shifty compounds.

Understanding atropisomers

A major catalyst for this reversal in attitude, many say, is the work of Steven R. LaPlante, who worked as a drug discovery expert at Boehringer Ingelheim for more than two decades before founding NMX Research & Solutions and joining the faculty at the University of Quebec.

LaPlante says the dogma at Boehringer, and many other pharmaceutical companies at the time, was that drug candidates wouldn’t be successful if they had any ability to change over time. “If we found that we were stuck with atropisomerism, we would basically cancel the project. And the reason why we’d cancel the project is that nobody understood how to deal with the situation.”

Credit: Sean Toenjes
Chlorine atoms (green balls) keep the rings in this compound from rotating freely, creating two asymmetric forms called atropisomers. It would take over a year for one atropisomeric form of this molecule to overcome the rotational barrier and become a racemic mixture at body temperature.

LaPlante says this scenario arose often—in at least 60% of projects—and usually during the lead optimization of a drug candidate. In this stage of development, teams engineer every angstrom of a compound, tacking ever-more-specialized substituents onto the core structure in search of the slightest gains in potency. And generally chemists end up making molecules more rigid and compact. This strategy improves activity by locking compounds into a conformation that best fits into a defined pocket in a biological molecule such as a target protein. However, increasing a structure’s rigidity through additional substitutions also increases the potential for atropisomerism.

About a decade ago, LaPlante was working on compounds that would inhibit the enzyme HIV integrase, when atropisomerism struck. But this time, the activity he was seeing in the lead compound was too good to pass up. So he began asking chemists at his company who were involved in drug development further downstream about the requirements for advancing an atropisomer as a drug candidate.

“People started admitting that they didn’t know the real answer, and a lot of people didn’t even know about the phenomenon,” LaPlante says.

Eventually, his inquiries led him to the U.S. Food & Drug Administration, which at the time also didn’t have any documentation on atropisomers. In 2011, LaPlante and colleagues at Boehringer collaborated with FDA to produce a detailed perspective on atropisomers (J. Med. Chem. 2011, DOI: 10.1021/jm200584g). The publication remains the definitive guidance on atropisomers for drug developers, according to an FDA representative.

It offers a number of examples of atropisomerism, some of which were molecules already on the market. These compounds included the X-ray contrast agent iomeprol and the sedative afloqualone, which were developed as racemic mixtures. Although none of the drugs needed to be removed from the market—they were fairly stable—when LaPlante informed drugmakers of their compound’s atropisomerism, he says, “people were not happy to hear about it.”

Structure of iomeprol.

The publication also presents analytical methods for detecting atropisomers. Chiral liquid chromatography is the most common way to confirm their existence, but unusually broad peaks in a nuclear magnetic resonance spectrum can also hint at their presence. Even though atropisomers can complicate drug discovery, the paper makes the case that these compounds don’t have to be discarded. “By being aware of it, now you have choices,” LaPlante says.

The authors provide a figure that divides atropisomers into categories, as well as guidance on how to proceed with each type. The divisions are based on the molecules’ half-lives, or half the time it takes for a pure sample of one atropisomer to overcome its rotational energy barrier and become a one-to-one mixture of both forms. The classifications give scientists across disciplines a way to talk to each other, LaPlante says.

In class I, compounds freely rotate about their chiral axis from one atropisomer to the other in seconds to minutes. Researchers are advised to advance these molecules as a racemic mixture and to not bother trying to isolate either version as long as both forms are safe. Class III atropisomers are stable for years and as such can be developed like common point-chiral molecules that contain an atom bound to four different chemical groups. Class II molecules have intermediate stability, from hours to days, and present the most difficulty.

Image of six structures with increasing rotational half-lives shown on a timeline.
Flipping time
As more substituents surround a molecule’s axis of rotation (shown in red), the time the compound takes to contort itself from one chiral form to the other increases. Racemization half-lives calculated at 25 ͒C. Class I = atropisomers take seconds to minutes to racemize, class II = hours to days, and class III = years.

While compounds in class II could also be developed as a mixture as long as the ratio between the atropisomeric forms stays constant, the article says, chemists may want to consider nudging the molecule into one of the other two classes. For example, adding appendages to a molecule to stiffen it might push the compound to class III, while removing certain groups might loosen it and turn it into a class I compound. Another option is to make the molecules symmetrical by adding matching substituents on the rings, thereby getting rid of atropisomerism altogether.

Since its publication, the FDA-Boehringer article has helped many chemists struggling with atropisomers. But each atropisomer is unique, as are the goals of each team of chemists, so researchers need to approach drug candidates case by case.

Atropisomers in practice

Structure of a chemical probe developed by Bayer for bromodomain inhibition.

Bayer drug discovery chemist Ingo V. Hartung hadn’t had much direct experience with atropisomers until two years ago. “But once you encounter atropisomers, you see them everywhere,” he says. His team came across class II atropisomers that inhibited so-called bromodomain proteins involved in regulating gene transcription. The scientists had been searching for compounds that could serve as chemical probes to bind to and thereby explore the specific roles of these proteins.

The crucial question for the researchers was whether the atropisomeric forms of the compound they hit upon had similar or different bioactivity and if they interconvert under their assay conditions, Hartung says. The researchers found that both atropisomeric forms displayed about the same potency and slight racemization. Because Hartung and his team wanted others to be able to use the probe without worrying about its stereochemical integrity, they decided that a racemic mixture of the class II atropisomers would suit their needs (J. Med. Chem. 2017, DOI: 10.1021/acs.jmedchem.7b00306).

Structure of a clinical candidate kinase inhibitor.

Atropisomers also appear regularly among kinase inhibitors, with 80% of FDA-approved kinase inhibitors containing an atropisomeric axis, according to a recent review by San Diego State University’s Jeffrey L. Gustafson (Future Med. Chem. 2018, DOI: 10.4155/fmc-2017-0152). His lab has shown that favoring one atropisomeric form over the other can increase kinase selectivity, as one form tends to bind the desired target while the other causes off-target effects (Angew. Chem. Int. Ed. 2015, DOI: 10.1002/anie.201506085).

Stephane Perreault and colleagues at Gilead Sciences recently published work on an atropisomeric kinase inhibitor for the treatment of cancer. The team used a computational model to intentionally design a rigid, class III atropisomer to inhibit phosphoinositide 3-kinase (J. Med. Chem. 2018, DOI: 10.1021/acs.jmedchem.8b00797). The researchers found that one chiral form is 500-fold as active as the other. The active atropisomer also exhibits much better selectivity for the target kinase as well as metabolic stability. “It was like night and day,” Perreault says.


Yet another atropisomeric kinase inhibitor, of Bruton’s tyrosine kinase (BTK), currently being evaluated in Phase II clinical trials for rheumatoid arthritis, comes from Bristol Myers-Squibb. BMS-986142 contains one point-chiral center and two atropisomeric chiral axes, making it a diastereomeric compound with eight possible isomers. The less stable atropisomeric axis has a half-life on the order of hours to days, which means it can’t be heated above about 45 °C without the compound morphing. To keep the molecule from racemizing, the team had to design its synthetic routes and analysis with a close eye on temperature.

Structure of BMS-986142.

During the discovery stage, BMS analytical chemist Jun Dai and the team developed methods to analyze the compounds’ isomers. She estimates that the researchers screened at least twice as many separation methods for atropisomers as they would have for normal chiral compounds because of the atropisomers’ potential for temperature-dependent conversion. “It was challenging but rewarding,” she says.

To determine the proportion of early atropisomers with half-lives of minutes to hours, the team ran high-performance liquid chromatography analysis at low temperature, chilling the column with ice or cooling equipment. Isolating some atropisomeric compounds required researchers to use ice-bath cooling during fraction collection and even solvent evaporation. The medicinal chemistry route to BMS-986142 required three chiral column purifications to obtain a single diastereomer with the best binding properties (J. Chromatogr. A 2017, DOI: 10.1016/j.chroma.2017.01.016).

Process synthesis, however, generally isn’t amenable to column chromatography steps, which can take weeks to months on a large scale. “To be honest, when I first saw it, I really wasn’t sure how we were going to make it,” says BMS chemist Thomas Razler, who led the process chemistry efforts to scale-up BMS-986142.

The researchers say extensive knowledge sharing between medicinal, analytical, and process teams about the atropisomeric compound was key to the program’s success. The process team took advantage of the fact that the diastereomeric forms of BMS-986142 had very different solubility profiles, enabling the chemists to replace all chiral chromatography with simpler crystallization steps and produce more than 200 kg of a single enantiomer and diastereomer (Org. Lett. 2018, DOI: 10.1021/acs.orglett.8b01218).

Although the final molecule is stable as a solid, the team says that in solution, the risk of racemization is higher. Citing ongoing work in that area of development, Razler declined to elaborate on how the molecule behaves in its formulation but notes the team hopes to publish that information next year. The atropisomerism is still an issue, he says, but a fascinating one.

UPDATE: This story was adjusted on Aug. 21, 2018, to display the stereochemistry of the more active atropisomeric kinase inhibitor developed by Gilead rather than the less active one. The spelling of phosphoinositide in the compound’s label was also corrected.
This story was additionally corrected on Sept. 15, 2018, to properly attribute descriptions of Bristol-Myers Squibb’s analytical and separation techniques to atropisomeric compounds early in drug development rather than to BMS-986142.


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