Noteworthy Chemistry

January 12, 2015


Automate nanomole scale-up reaction screening. High-throughput experimentation (HTE) is a useful tool for discovering new reactions and for screening scale-up conditions for synthesizing natural products and pharmaceuticals. HTE requires a large number of substrates for multiple trials, but precursors for large libraries of complex drug molecules or natural products are often unavailable.

To solve this supply–demand problem, T. Cernak, S. D. Dreher, and fellow researchers at Merck (Rahway, NJ; West Point, PA; Kenilworth, NJ; and Boston) developed a miniaturized HTE platform for optimizing palladium-catalyzed cross-coupling reactions on the nanomole scale by using biotechnology robotics.

The authors initially examined cross-coupling reactions between a group of electrophilic drug cores (Figure 1) and a set of nucleophilic polar building blocks (Figure 2) with a variety of bases and palladium catalysts. In the structures, Boc is tert-butoxycarbonyl.

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Electrophilic drug cores used to test high-throughput reaction screening
Nucleophilic polar building blocks used to test high-throughput reaction screening

The reactions were run in dimethyl sulfoxide solvent at room temperature. The equipment consisted of high-precision nanoliter robotics and plastic plates with 1536 wells, which allowed 1536 reactions with 20 μg of substrate per reaction to be screened in 2 h. High-throughput liquid chromatography–mass spectrometry was used to analyze the products from all 1536 reactions within 9 h.

Heat maps generated from the analytical data not only helped to identify the optimal reaction conditions, but they also showed the influence of continuous variables on yields. The results could be translated into preparative-scale reactions without loss of yield.

This work is a pioneering effort in the high-throughput screening of reaction conditions with limited substrate resources. With the combination of high-throughput experimentation and analysis, this miniaturized platform allows fast, efficient, accurate identification of the best preparative conditions. Its basic principles can also be applied to other high-value synthetic reactions. (Science DOI: 10.1126/science.1259203; Xin Su)

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Safely run palladium-catalyzed coupling reactions with diazonium salts. F.-X. Felpin and coauthors review a process for safely using diazonium salts in palladium-catalyzed reactions such as Sonogashira, Heck–Matsuda, and Suzuki–Miyaura couplings. This process is safer than its predecessors because there is no need to make large quantities of the diazonium salt or isolate the salt.

The process consists of tandem reactions in which an aniline derivative is added to a solution of the diazotization reagent (typically tert-butyl nitrite), the palladium catalyst, and the coupling partner. Only a very small amount of diazonium salt is present at any time during the reaction.

With this procedure, anilines with electron-donating or electron-withdrawing groups give good-to-excellent yields of coupling products. The conditions may need to be modified slightly, depending on the type of coupling reaction; but within each coupling class, the conditions are uniform. The authors describe a large-scale example of a Heck–Matsuda coupling of 2-aminobenzenesulfonic acid en route to the herbicide prosulfuron. (Org. Process Res. Dev. DOI: 10.1021/op500299t; Will Watson)

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Protein-shelled platinum and graphene form a nanocapacitor. Biosensors and embedded therapeutic devices would be improved if they contained flexible, biocompatible electronic components. These components may incorporate metallic or semiconducting nanoparticles that can be encased in hollow protein shells that control their size and stability, protect the nanoparticles, and reduce their toxicity. To date, there are few reports of electronic devices that incorporate protein-shelled nanoparticles; and little is known about their charge-transport properties.

T. Kim, K. K. Kim, and coauthors at Sungkyunkwan University and Medical School (Suwan), BIO-FD&C (Incheon), and Myongji University (Yongin, all in Korea) encapsulated platinum nanoparticles (PtNPs) inside PepA, a bacterial aminopeptidase molecule with a hollow center. They could vary the size of the PtNPs and thus tune the electronic properties of the protein-coated particles, by varying the ratio of precursor platinum ions to PepA.

The authors assembled a graphene-based field-effect transistor (GFET) with a SiO2/Si substrate, gold electrodes, and an overlayer of graphene topped with PepA-PtNPs. The left side of the figure is a schematic diagram of the GFET. 

GFET (left) and biocapacitor (right)

Although proteins normally increase GFET conductivity, the encapsulated PtNPs reduced the conductivity of the GFET, possibly by acting as electron acceptors and hindering electron transfer between adjacent PepA molecules. Increasing the concentration of PepA-PtNPs reduced the GFET's conductivity further, possibly because surface adsorption of these particles enhances their ability to trap electrons from graphene.

Smaller PtNPs reduced conductivity more than larger ones, which confirms a previous observation that smaller NPs have greater charge-trapping capability. Thus, the size and concentration of NPs can be used to tune the conductivity of the GFET.

The authors also built a frequency-modulated capacitor by sandwiching an aqueous suspension of PepA-PtNPs between graphene layers coated with poly(methyl methacrylate) (PMMA; see schematic diagram at right in the figure). The specific capacitance decreased with increasing AC signal frequency and with the size of the PtNPs, possibly because of changes in the polarizability of the PepA-PtNP layer. (ACS Nano DOI: 10.1021/nn503178t; Nancy McGuire)

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An improved vaccine protects against nine types of HPV. Human papillomaviruses (HPVs) are a group of related viruses that can spread through skin-to-skin contact during sexual intercourse. HPV is the most common sexually transmitted infection in the United States, affecting more than half of sexually active people. “Low-risk” HPVs can cause genital warts, and “high-risk” HPVs can cause cancer. More than 12 oncogenic HPV types have been identified.

High-risk HPV infection accounts for ≈5% of cancers worldwide. Almost all cervical cancers, most anal cancers, and about half of vaginal, vulvar, penile, and oropharyngeal cancers are caused by high-risk HPVs.

The Gardasil vaccine, approved by the US Food and Drug Administration in 2006, covers four types of HPV that are responsible for 70% of cervical cancers in the United States. On December 10, 2014, FDA approved an enhanced version called Gardasil 9 (Merck, Whitehouse Station, NJ) that covers nine HPV types. The five additional types cause ≈20% of cervical cancers.

Gardasil 9 can potentially prevent ≈90% of cervical, vulvar, vaginal, and anal cancers. Studies showed it to be as effective as original Gardasil for preventing diseases caused by the four shared HPV types (6, 11, 16, and 18) and 97% effective in preventing cancers caused by the five additional HPV types (31, 33, 45, 52, and 58).

Gardasil 9 is approved for use by females age 9–26 and males age 9–15. According to FDA, “Gardasil 9’s full potential for benefit is obtained by those who are vaccinated before becoming infected with the HPV strains covered by the vaccine.” (US Food and Drug Administration. http://www.fda.gov/NewsEvents/Newsroom/PressAnnouncements/ucm426485.htm; Abigail Druck Shudofsky)

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A phthalimide protects protein amino groups in buffered solution. The chemical modification of proteins requires mild, rigorously chemoselective conditions. V. Rai and co-workers at the Indian Institute of Science, Education, and Research (Bhopal) developed a procedure for modifying protein amino groups under biocompatible conditions.

With benzylamine (2 in the figure) as the model substrate, the authors screened several phthalimide protecting groups in a phosphate buffer (pH 7.0) at room temperature. They found that N-hydroxyphthalimide (NHP, 1) gives good yields of the protected amine (3). Under the same conditions, neither benzyl alcohol nor benzyl thiol reacts with NHP, demonstrating that the method is selective for protecting amines. The reactions can be run on a gram scale. 

Reaction of N-hydroxyphthalimide protecting group with model compound benzylamine

When RNase A was protected with 1 equiv NHP, the α-amine was selectively protected with 45–50% conversion. This result was corroborated by the selective protection of the primary amine of the tetrapeptide GGGK-NH2.

Phthalimide-protected amines can be deprotected under buffered conditions by using hydrazine to recover the amine or sodium sulfide or piperidine to recover the aromatic carboxylic acid. The authors are currently developing additional N-hydroxyphthalimides as fluorescent probes or precursors to click chemistry reactions. (Chem. Commun. DOI: 10.1039/C4CC08503E; José C. Barros)

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Choose the right conditions for this drug synthesis scale-up. S. Tortoioli and coauthors at Actelion Pharmaceuticals (Allschwil, Switzerland) and DSM Pharma Chemicals (Regensburg, Germany) compare three scale-up routes to an orexin receptor antagonist. The drug is a potential treatment for disorders ranging from stress to addictions.

The route the authors call 1b starts from N-Boc-proline and requires three steps: amide formation with 3,5-dimethylaniline, Boc (tert-butoxycarbonyl) removal, and sulfonamide formation with 4-methoxybenzenesulfonyl chloride.

Routes 2a and 2b, starting from unprotected proline, also require three steps: sulfonamide, acid chloride, and amide formation; all three steps are fully telescoped. The difference between these routes is the reagent used for acid chloride formation (thionyl chloride for route 2a, pivaloyl chloride for route 2b).

Routes 2a and 2b are superior to 1b according to all of the metrics used for comparison: yield, use of protecting groups and/or class 2 solvents, process mass intensity, reaction mass efficiency, and cost. Route 2a is less costly than 2b, but it is inferior to 2b by four of the other metrics. (Org. Process Res. Dev. DOI: 10.1021/op500277s; Will Watson)

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