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Point-of-care production of therapeutic proteins of good-manufacturing-practice quality

Abstract

Manufacturing technologies for biologics rely on large, centralized, good-manufacturing-practice (GMP) production facilities and on a cumbersome product-distribution network. Here, we report the development of an automated and portable medicines-on-demand device that enables consistent, small-scale GMP manufacturing of therapeutic-grade biologics on a timescale of hours. The device couples the in vitro translation of target proteins from ribosomal DNA, using extracts from reconstituted lyophilized Chinese hamster ovary cells, with the continuous purification of the proteins. We used the device to reproducibly manufacture His-tagged granulocyte-colony stimulating factor, erythropoietin, glucose-binding protein and diphtheria toxoid DT5. Medicines-on-demand technology may enable the rapid manufacturing of biologics at the point of care.

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Acknowledgements

We thank DARPA Biologically-derived Medicines on Demand (Bio-MOD) Project Grant (N66001-13-C-4023, ‘CASTing Biologics on Demand’) for financial support. G. Ling (Retd. US Army Colonel and former DARPA Battlefield Medicine Program Director) conceived the vision for Bio-MOD based on his difficulties in securing medicines to treat patients during deployments in Iraq and Afghanistan. This paper is dedicated to him. We thank E. Choi, J. Lewin, A. Bryon, M. Zamisch, T. McQuade, B. Ringeisen, G. Kost, K. Pankratz, R. Cecil, B. Webb, A. Bose, B. Junker, W.-L. Ling, M. McGinnis, P. Latham and R. Gopinath for various discussions, encouragement and support, and I. Shaffer of the Molecular Characterization and Analysis Complex, University of Maryland Baltimore County for sample analysis of leachables and extractables. We thank E. Gutierrez for his assistance with the illustrations. S.V. acknowledges Y. Xia of the MNMR Structural Biology Centre, University of Minnesota for technical help in the NMR spectroscopy. We thank the FDA Emerging Technology Team for helpful guidance and discussions. Disclaimer: This work was conducted while S.V. was employed at the University of Maryland School of Pharmacy. The opinions expressed in the article are the author’s own and do not reflect the view of the National Institutes of Health, the Department of Health and Human Services, or the United States government. No endorsement of this work by the Food and Drug Administration, National Institutes of Health, the Department of Health and Human Services, DARPA, Department of Defense or the United States government is implied.

Author information

M.A., A.A., D.B., X.G., Y.K., M.P., B.P., G.R., D.T., M.T., K.Tr. and B.W. designed, assembled and tested the Bio-MOD system. R.A., A.M., K.M., C.G., C.P., M.P., B.P., D.T. and K.Tr. maintained and reviewed the batch records. D.F., Y.L., H.G., S.V., A.Z. and S.D. did the protein purification. M.C., C.G., P.J., M.P., K.V. and D.W. did the cloning. R.A., S.Bo., S.Br., S.D., X.G., H.G., Y.L., K.M., C.P., M.P., B.P., A.R., P.R., S.S., K.Ta., L.T., K.V., S.V., J.W. and W.L. did the product analysis including gels, activity assays, mass spectrometry, protein concentration, sterility, silver staining, lysate stability, leachables and extractables. S.Br., D.B., M.C., C.G., P.J., M.P., K.Tr., K.V. and S.V. did the protein expression. D.F., X.G., C.G., Y.K., A.M., G.R., L.T., S.V., D.W. and K.V. did the overall and subsystem experimental design, execution and analysis. D.F., X.G., Y.K., C.P., B.P., G.R., L.T., K.V., S.V. and D.W. analysed the data.

Competing interests

G.R., Y.K., L.T., X.G. and D.F. are listed as inventors on the United States patents 9,388,373 ‘Microscale Bioprocessing System and Method for Protein Manufacturing’ and 9,982,227 ‘System and Method for Production of On-Demand Proteins in a Portable Unit for Point of Care Delivery’.

Correspondence to Govind Rao.

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Fig. 1: The current biologically derived medicines on demand, Bio-MOD, system (version 3.0), a miniature system for biologics manufacturing.
Fig. 2: Automated end-to-end expression and purification of G-CSF in Bio-MOD 2.0.
Fig. 3: Operational logic for Bio-MOD 3.0.
Fig. 4: Comparison of G-CSF-His produced in two identical Bio-MODs.
Fig. 5: G-CSF-His produced in the Bio-MOD.
Fig. 6: Real-time analytics from UV sensors.
Fig. 7: Characterization of GBP and EPO produced by Bio-MOD.