Abstract
Enzymatic reductive amination, being a direct, selective and green methodology, has attracted significant interest in a short period of time and is emerging as a powerful tool for the synthesis of chiral alkylated amines. The discovery of an increasing number of imine reductases with reductive aminase (RedAm) activity has enabled mechanistic and substrate profiling studies. However, their potential for commercial applications has not been realized. Here, we report the discovery of RedAm activity in an imine reductase enzyme for the direct reductive amination of a cyclic ketone with methylamine. We also investigate engineering the enzyme to access a cis-cyclobutyl-N-methylamine for the manufacturing of a late-stage drug candidate, Janus kinase 1 (JAK1) inhibitor abrocitinib. The engineered enzyme, SpRedAm-R3-V6, showed >200-fold improvement in performance over the wild-type enzyme and was successfully used to develop a commercial manufacturing process with 73% isolated yield at 99.5% purity and high selectivity (>99:1 cis:trans). This process has been successfully used to manufacture multi-metric tons of the amine, demonstrating the potential of RedAm technology for commercial manufacturing.
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Data availability
Additional data supporting the findings reported in this paper are available as Supplementary Information. All other data are available from the authors upon reasonable request.
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Acknowledgements
We would like to thank Pfizer leadership L. Handanyan, S. Caron, N. Thomson, J. Nelson and C. McWilliams for their support for the work and publication. We would also like to thank R. Lewis and S. France for proof-reading the manuscript.
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R.K., M.J.K., J.S., M.P.B. and C.A.M. contributed to enzyme engineering and enzyme process development work. R.K., E.L.M., N.M.D., C.G.C. and E.C. performed process engineering and process development. D.B., L.A.C., K.M., D.M. and S.H. ran the manufacturing scale reactions. C.A.L., K.M.D., R.P. and J.W. provided analysis, sourcing support and participated in discussion. R.K., M.J.K. and J.S. wrote the paper and the other co-authors provided information and feedback.
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Supplementary Methods, Notes 1 and 2, Figs. 1–4 and Tables 1–4.
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Kumar, R., Karmilowicz, M.J., Burke, D. et al. Biocatalytic reductive amination from discovery to commercial manufacturing applied to abrocitinib JAK1 inhibitor. Nat Catal 4, 775–782 (2021). https://doi.org/10.1038/s41929-021-00671-5
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DOI: https://doi.org/10.1038/s41929-021-00671-5
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