Quantitative characterization of intact sialylated O-glycans with MALDI-MS for protein biotherapeutics

  • Cheol-Hwan Hwang
  • Hae-Min Park
  • Han-Gyu Park
  • Da-Hee Ahn
  • Seong-Min Kim
  • Byoung Joon Ko
  • Young Hwan Kim
  • Yung-Hun Yang
  • Yun-Gon Kim
Research papers
  • 14 Downloads

Abstract

For validating O-glycosylation of protein biotherapeutics, we presented a quantitative O-glycomics method which is based on the neutralization of sialic acids, the specific release of O-glycans, and the introduction of permanent positive charge followed by quantitative MALDI-MS analysis. This method shows excellent technical reproducibility, linearity and sensitivity. In addition, it enables the quantification of intact O-glycans with minimal degradation or loss of sialic acids on these glycans compared to a conventional HPLC-based method. We then applied this method to quantitatively characterize O-glycans present on Etanercept. The analysis showed the relative abundances of mono- and di-sialylated core 1 O-glycans - were 79.3±0.8% and 17.3±1.4%, respectively. This glycomics technology could allow for the reliable quantitative analysis of intact O-glycans from glycoproteins and may contribute to validation of O-glycosylation protein biotherapeutics in the pharmaceutical industry.

Keywords

Chemical Derivatization MALDI-MS O-glycan Protein Biotherapeutics Quantitative Analysis 

Preview

Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.

References

  1. 1.
    S. Elliott, T. Lorenzini, S. Asher, K. Aoki, D. Brankow, L. Buck, L. Busse, D. Chang, J. Fuller, J. Grant, N. Hernday, M. Hokum, S. Hu, A. Knudten, N. Levin, R. Komorowski, F. Martin, R. Navarro, T. Osslund, G. Rogers, N. Rogers, G. Trail and J. Egrie, Nat. Biotechnol., 21, 414 (2003).CrossRefGoogle Scholar
  2. 2.
    R. J. Sola and K. Griebenow, BioDrugs, 24, 9 (2010).CrossRefGoogle Scholar
  3. 3.
    A. M. Sinclair and S. Elliott, J. Pharm. Sci., 94, 1626 (2005).CrossRefGoogle Scholar
  4. 4.
    R. J. Sola and K. Griebenow, J. Pharm. Sci., 98, 1223 (2009).CrossRefGoogle Scholar
  5. 5.
    P. Kang, Y. Mechref, Z. Kyselova, J. A. Goetz and M. V. Novotny, Anal. Chem., 79, 6064 (2007).CrossRefGoogle Scholar
  6. 6.
    W. A. Tao and R. Aebersold, Curr. Opin. Biotechnol., 14, 110 (2003).CrossRefGoogle Scholar
  7. 7.
    G. Alvarez-Manilla, N. L. Warren, T. Abney, J. Atwood, 3rd, P. Azadi, W. S. York, M. Pierce and R. Orlando, Glycobiology, 17, 677 (2007).CrossRefGoogle Scholar
  8. 8.
    C. Wang, P. Zhang, W. Jin, L. Li, S. Qiang, Y. Zhang, L. Huang and Z. Wang, J. Proteomics, 150, 18 (2017).CrossRefGoogle Scholar
  9. 9.
    M. Toyoda, H. Ito, Y. K. Matsuno, H. Narimatsu and A. Kameyama, Anal. Chem., 80, 5211 (2008).CrossRefGoogle Scholar
  10. 10.
    G.C. Gil, B. Iliff, R. Cerny, W. H. Velander and K. E. Van Cott, Anal. Chem., 82, 6613 (2010).CrossRefGoogle Scholar
  11. 11.
    H. J. Jeong, M. Adhya, H. M. Park, Y. G. Kim and B. G. Kim, Xenotransplantation, 20, 407 (2013).CrossRefGoogle Scholar
  12. 12.
    C. Wang, Z. Wu, J. Yuan, B. Wang, P. Zhang, Y. Zhang, Z. Wang and L. Huang, J. Proteome. Res., 13, 372 (2014).CrossRefGoogle Scholar
  13. 13.
    Y. Shinohara, J. Furukawa, K. Niikura, N. Miura and S. Nishimura, Anal. Chem., 76, 6989 (2004).CrossRefGoogle Scholar
  14. 14.
    Y.W. Kim, C. Sung, S. Lee, K. J. Kim, Y. H. Yang, B. G. Kim, Y. K. Lee, H. W. Ryu and Y. G. Kim, Anal. Chem., 87, 858 (2015).CrossRefGoogle Scholar
  15. 15.
    K. J. Kim, H. J. Kim, H. G. Park, C. H. Hwang, C. Sung, K. S. Jang, S. H. Park, B. G. Kim, Y. K. Lee, Y. H. Yang, J. H. Jeong and Y. G. Kim, Sci. Rep., 6, 24489 (2016).CrossRefGoogle Scholar
  16. 16.
    K. J. Kim, Y. W. Kim, C. H. Hwang, H. G. Park, Y. H. Yang, M. Koo and Y. G. Kim, Biotechnol. Lett., 37, 2019 (2015).CrossRefGoogle Scholar
  17. 17.
    L. Liu, S. Gomathinayagam, L. Hamuro, T. Prueksaritanont, W. Wang, T. A. Stadheim and S. R. Hamilton, Pharm. Res., 30, 803 (2013).CrossRefGoogle Scholar
  18. 18.
    D. Pennica, V. T. Lam, R. F. Weber, W. J. Kohr, L. J. Basa, M. W. Spellman, A. Ashkenazi, S. J. Shire and D. V. Goeddel, Biochemistry, 32, 3131 (1993).CrossRefGoogle Scholar
  19. 19.
    M. DiPaola, J. Li and E. Stephens, J. Bioanal. Biomed., 5, 5 (2013).CrossRefGoogle Scholar
  20. 20.
    S. Houel, M. Hilliard, Y. Q. Yu, N. McLoughlin, S. M. Martin, P. M. Rudd, J. P. Williams and W. Chen, Anal. Chem., 86, 576 (2014).CrossRefGoogle Scholar

Copyright information

© Korean Institute of Chemical Engineers, Seoul, Korea 2018

Authors and Affiliations

  • Cheol-Hwan Hwang
    • 1
  • Hae-Min Park
    • 2
  • Han-Gyu Park
    • 1
  • Da-Hee Ahn
    • 1
  • Seong-Min Kim
    • 1
  • Byoung Joon Ko
    • 3
  • Young Hwan Kim
    • 4
  • Yung-Hun Yang
    • 5
  • Yun-Gon Kim
    • 1
  1. 1.Department of Chemical EngineeringSoongsil UniversitySeoulKorea
  2. 2.Department of ChemistryNorthwestern UniversityEvanston, IllinoisU.S.A.
  3. 3.New Drug Development CenterOsong Medical Innovation FoundationCheongjuKorea
  4. 4.Biomedical Omics TeamKorea Basic Science Institute (KBSI)CheongjuKorea
  5. 5.Department of Biological EngineeringKonkuk UniversitySeoulKorea

Personalised recommendations