Enantioselective synthesis: XRD facilitates anticancer drug preparation

Anticancer Nutlins

Chemists Tyler Davis and Jeffrey Johnston of Vanderbilt University, in Nashville, Tennessee, explain how they have carried out the first highly diastereo- and enantio-selective additions of aryl nitromethane pronucleophiles to aryl aldimines. They then used their approach, which involved identification of an electron-rich chiral bis(amidine) catalyst, in an enantioselective synthesis of (-)-Nutlin-3. The team explains that the configuration was assigned on the basis of analogy to an adduct whose absolute and relative configuration were assigned by X-ray crystallography, carried out for the team by Maren Pink of Indiana University Molecular Structure Center. The Vanderbilt synthesis overcomes the issue of the multiple stereo centres in the Nutlin-3 skeleton as well as offering a straightforward approach to the compound.

Generically speaking, Nutlins are cis-imidazoline analogues that inhibit the interaction of MDM2 and p53, they were identified by screening a chemical library. Nutlin-3 is the more potent of Nutlin-1, 2, and 3, and is the focus of much research. Essentially, it reactivates the necessary biochemical pathway allowing programmed cell death, apoptosis, to restart in cancerous cells. Formally, the compound is 4-[4,5-bis(4-chlorophenyl)-2-(2-isopropoxy-4-methoxy-phenyl)-4,5-dihydro-imidazole-1-carbonyl]-piperazin-2-one. This Hoffmann-La Roche compound is a potent small molecule inhibitor of the negative regulator MDM2 (murine double minute) which modulates activity of the tumour suppressor p53. As such Nutlin-3 is used extensively as a probe in cell biology research and Hoffmann-La Roche is currently developing it as a drug for cancer chemotherapy against neuroblastoma, the most common solid cancer in infants. Nutlin-3 is also being tested in conjunction with other agents, such as dasatinib as a potential treatment for leukaemia.

It is crucial to the further development of this class of compounds that chemists continue to search for novel and efficient synthetic approaches to the parent, which could open up access to derivatives with related properties.

The team confesses to not fully understanding the mechanism at the key stereoinduction step of their reaction scheme, the other parts are relatively straightforward team leader Jeffrey Johnston told SpectroscopyNOW. Nevertheless, the success of their synthesis means that the compound can be fully characterised by spectroscopy and analytical data obtained for its synthetic intermediates. Prior to this work, the precise molecular weight of Nutlin-3 had been determined by high-resolution mass spectrometry. As such, the team turned to an HMBC (600 MHz) experiment to verify that they had produced the desired imidazoline isomer. They saw a cross-peak to the urea carbon from H1 of the imidazoline characteristic of the levorotatory isomer and no cross-peak between H8 and an amide carbonyl carbon, which would have suggested the opposite enantiomer.

"We knew the relative and absolute configuration of our intermediates," Johnston told us, "it was only the chemoselectivity in the dehydration that we needed to establish - that is, among the urea and amide carbonyls, which one supplied the nucleophilic nitrogen to become part of the imidazoline, and which one lost its carbonyl during the dehydration?" The HMBC allowed the team to determine that the amide carbonyl was lost to dehydration, and the urea nitrogen emerged in the imidazoline ring with its carbonyl intact. Since only the mass of Nutlin-3 was provided in the patent (not rotation, not NMR). As such, they couldn't simply compare their analytical data to something published.

Precluding chiral chromatography

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