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Strategies to Develop Therapeutic N- and O-Hyperglycosylated Proteins

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Recombinant Glycoprotein Production

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1674))

Abstract

Glycoengineering by N- and/or O-hyperglycosylation represents a procedure to introduce potential sites for adding N- and/or O-glycosyl structures to proteins with the aim of producing biotherapeutics with improved pharmacodynamic and pharmacokinetic properties. In this chapter, a detailed description of the steps routinely performed to generate new proteins having high content of N- and/or O-glycosyl moieties is carried out. The rational strategy involves the initial stage of designing N- and/or O-hyperglycosylated muteins to be expressed by mammalian cells and includes the upstream and downstream processing stages necessary to develop hyperglycosylated versions of the proteins of interest with the purpose of beginning the long road toward producing biobetters.

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References

  1. Elliott S, Lorenzini T, Asher S, Aoki K, Brankow D, Buck L, Busse L, Chang D, Fuller J, Grant J, Hernday N, Hokum M, Hu S, Knudten A, Levin N, Komorowski R, Martin F, Navarro R, Osslund T, Rogers G, Rogers N, Trail G, Egrie J (2003) Enhancement of therapeutic protein in vivo activities through glycoengineering. Nat Biotechnol 21(4):414–421. doi:10.1038/nbt799nbt799 [pii]

    Article  CAS  PubMed  Google Scholar 

  2. Sinclair AM, Elliott S (2005) Glycoengineering: the effect of glycosylation on the properties of therapeutic proteins. J Pharm Sci 94(8):1626–1635. doi:10.1002/jps.20319

    Article  CAS  PubMed  Google Scholar 

  3. Egrie JC, Browne JK (2001) Development and characterization of novel erythropoiesis stimulating protein (NESP). Br J Cancer 84(Suppl 1):3–10. doi:10.1054/bjoc.2001.1746S000709200191746X [pii]

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  4. Egrie JC, Dwyer E, Browne JK, Hitz A, Lykos MA (2003) Darbepoetin alfa has a longer circulating half-life and greater in vivo potency than recombinant human erythropoietin. Exp Hematol 31(4):290–299

    Article  CAS  PubMed  Google Scholar 

  5. Perlman S, van den Hazel B, Christiansen J, Gram-Nielsen S, Jeppesen CB, Andersen KV, Halkier T, Okkels S, Schambye HT (2003) Glycosylation of an N-terminal extension prolongs the half-life and increases the in vivo activity of follicle stimulating hormone. J Clin Endocrinol Metab 88(7):3227–3235. doi:10.1210/jc.2002-021201

    Article  CAS  PubMed  Google Scholar 

  6. Fares FA, Suganuma N, Nishimori K, LaPolt PS, Hsueh AJ, Boime I (1992) Design of a long-acting follitropin agonist by fusing the C-terminal sequence of the chorionic gonadotropin beta subunit to the follitropin beta subunit. Proc Natl Acad Sci U S A 89(10):4304–4308

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  7. Ceaglio N, Etcheverrigaray M, Kratje R, Oggero M (2008) Novel long-lasting interferon alpha derivatives designed by glycoengineering. Biochimie 90(3):437–449. doi:S0300-9084(07)00307-0 [pii]10.1016/j.biochi.2007.10.013

    Article  CAS  PubMed  Google Scholar 

  8. Ceaglio N, Etcheverrigaray M, Conradt HS, Grammel N, Kratje R, Oggero M (2010) Highly glycosylated human alpha interferon: an insight into a new therapeutic candidate. J Biotechnol 146(1-2):74–83. doi:S0168-1656(10)00025-8 [pii]10.1016/j.jbiotec.2009.12.020

    Article  CAS  PubMed  Google Scholar 

  9. Ceaglio N, Gugliotta A, Tardivo MB, Cravero D, Etcheverrigaray M, Kratje R, Oggero M (2016) Improvement of in vitro stability and pharmacokinetics of hIFN-alpha by fusing the carboxyl-terminal peptide of hCG beta-subunit. J Biotechnol 221:13–24. doi:S0168-1656(16)30019-0 [pii]10.1016/j.jbiotec.2016.01.018

    Article  CAS  PubMed  Google Scholar 

  10. Samoudi M, Minuchehr Z, Harcum SW, Tabandeh F, Omid Yeganeh N, Khodabandeh M (2017) Rational design of glycoengineered interferon-beta analogs with improved aggregation state: experimental validation. Protein Eng Des Sel 30(1):23–30. doi:gzw058 [pii]10.1093/protein/gzw058

    CAS  PubMed  Google Scholar 

  11. Samoudi M, Tabandeh F, Minuchehr Z, Ahangari Cohan R, Nouri Inanlou D, Khodabandeh M, Sabery Anvar M (2015) Rational design of hyper-glycosylated interferon beta analogs: a computational strategy for glycoengineering. J Mol Graph Model 56:31–42. doi:S1093-3263(14)00204-6 [pii]10.1016/j.jmgm.2014.12.001

    Article  CAS  PubMed  Google Scholar 

  12. Song K, Yoon IS, Kim NA, Kim DH, Lee J, Lee HJ, Lee S, Choi S, Choi MK, Kim HH, Jeong SH, Son WS, Kim DD, Shin YK (2014) Glycoengineering of interferon-beta 1a improves its biophysical and pharmacokinetic properties. PLoS One 9(5):e96967. doi:10.1371/journal.pone.0096967PONE-D-14-02849. [pii]

    Article  PubMed  PubMed Central  Google Scholar 

  13. Croxtall JD, McKeage K (2011) Corifollitropin alfa: a review of its use in controlled ovarian stimulation for assisted reproduction. BioDrugs 25(4):243–254. doi:4 [pii]10.2165/11206890-000000000-00000

    Article  CAS  PubMed  Google Scholar 

  14. Aebi M (2013) N-linked protein glycosylation in the ER. Biochim Biophys Acta 1833(11):2430–2437. doi:S0167-4889(13)00132-8 [pii]10.1016/j.bbamcr.2013.04.001

    Article  CAS  PubMed  Google Scholar 

  15. Schwarz F, Aebi M (2011) Mechanisms and principles of N-linked protein glycosylation. Curr Opin Struct Biol 21(5):576–582. S0959-440X(11)00140-0 [pii]10.1016/j.sbi.2011.08.005

    Article  CAS  PubMed  Google Scholar 

  16. Van den Steen P, Rudd PM, Dwek RA, Opdenakker G (1998) Concepts and principles of O-linked glycosylation. Crit Rev Biochem Mol Biol 33(3):151–208. doi:10.1080/10409239891204198

    Article  PubMed  Google Scholar 

  17. Rudd PM, Dwek RA (1997) Glycosylation: heterogeneity and the 3D structure of proteins. Crit Rev Biochem Mol Biol 32(1):1–100. doi:10.3109/10409239709085144

    Article  CAS  PubMed  Google Scholar 

  18. Fares F, Havron A, Fima E (2011) Designing a long acting erythropoietin by fusing three carboxyl-terminal peptides of human chorionic gonadotropin beta subunit to the N-terminal and C-terminal coding sequence. Int J Cell Biol 2011:275063. doi:10.1155/2011/275063

    Article  PubMed  PubMed Central  Google Scholar 

  19. Fares F, Guy R, Bar-Ilan A, Felikman Y, Fima E (2010) Designing a long-acting human growth hormone (hGH) by fusing the carboxyl-terminal peptide of human chorionic gonadotropin beta-subunit to the coding sequence of hGH. Endocrinology 151(9):4410–4417. doi:en.2009-1431 [pii]10.1210/en.2009-1431

    Article  CAS  PubMed  Google Scholar 

  20. Kasturi L, Eshleman JR, Wunner WH, Shakin-Eshleman SH (1995) The hydroxy amino acid in an Asn-X-Ser/Thr sequon can influence N-linked core glycosylation efficiency and the level of expression of a cell surface glycoprotein. J Biol Chem 270(24):14756–14761

    Article  CAS  PubMed  Google Scholar 

  21. Shakin-Eshleman SH, Spitalnik SL, Kasturi L (1996) The amino acid at the X position of an Asn-X-Ser sequon is an important determinant of N-linked core-glycosylation efficiency. J Biol Chem 271(11):6363–6366

    Article  CAS  PubMed  Google Scholar 

  22. Chauhan JS, Rao A, Raghava GP (2013) In silico platform for prediction of N-, O- and C-glycosites in eukaryotic protein sequences. PLoS One 8(6):e67008. doi:10.1371/journal.pone.0067008PONE-D-12-31390. [pii]

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  23. Chuang GY, Boyington JC, Joyce MG, Zhu J, Nabel GJ, Kwong PD, Georgiev I (2012) Computational prediction of N-linked glycosylation incorporating structural properties and patterns. Bioinformatics 28(17):2249–2255. doi:bts426 [pii]10.1093/bioinformatics/bts426

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  24. Chen YZ, Tang YR, Sheng ZY, Zhang Z (2008) Prediction of mucin-type O-glycosylation sites in mammalian proteins using the composition of k-spaced amino acid pairs. BMC Bioinformatics 9:101. doi:1471-2105-9-101 [pii]10.1186/1471-2105-9-101

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  25. Leung M-Y, Cardenas GA, Almeida IC, Gerken TA (2014) Isoform specific O-glycosylation prediction (ISOGlyP)Version 1.2.

    Google Scholar 

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Author information

Authors and Affiliations

  1. UNL, CONICET, FBCB, Cell Culture Laboratory, Ciudad Universitaria UNL-C.C. 242, (S3000ZAA), Santa Fe, Pcia. de Santa Fe, Argentina

    Agustina Gugliotta, Marina Etcheverrigaray, Ricardo Kratje & Marcos Oggero

  2. CONICET, Facultad de Bioquímica y Ciencias Biológicas, Universidad Nacional del Litoral, Santa Fe, Argentina

    Natalia Ceaglio

Authors
  1. Agustina Gugliotta
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  2. Natalia Ceaglio
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  3. Marina Etcheverrigaray
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  4. Ricardo Kratje
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  5. Marcos Oggero
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Corresponding author

Correspondence to Marcos Oggero .

Editor information

Editors and Affiliations

  1. Regional Blood Center of Ribeirão Preto, Ribeirão Preto Medical School, University of São Paulo, Ribeirão Preto, Brazil

    Virgínia Picanço-Castro

  2. School of Pharmaceutical Sciences of Ribeirão Preto, University of São Paulo, Ribeirão Preto, Brazil

    Kamilla Swiech

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Gugliotta, A., Ceaglio, N., Etcheverrigaray, M., Kratje, R., Oggero, M. (2018). Strategies to Develop Therapeutic N- and O-Hyperglycosylated Proteins. In: Picanço-Castro, V., Swiech, K. (eds) Recombinant Glycoprotein Production. Methods in Molecular Biology, vol 1674. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-7312-5_13

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  • DOI: https://doi.org/10.1007/978-1-4939-7312-5_13

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  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-7311-8

  • Online ISBN: 978-1-4939-7312-5

  • eBook Packages: Springer Protocols

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