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Imparting Bone Affinity to Glycoproteins Through the Conjugation of Bisphosphonates

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Abstract

Purpose. To develop a novel means of conjugating bisphosphonates onto the carbohydrate moieties of glycoproteins to enhance protein affinity to bone.

Methods. 1-Amino-1,1-diphosphonate methane (aminoBP) was conjugated onto the carbohydrate moietites of oxidized fetuin by using 4-(maleimidomethyl)cyclohexane-1-carboxyl-hydrazide (MMCCH). Bone affinity of the resulting conjugates was compared to proteins obtained from another means of conjugation, whereby aminoBP was conjugated onto fetuin's lysine moieties by using succinimidyl-4-(N-maleimidomethyl)-cyclohexane-1-carboxylate (SMCC).

Results. The use of the MMCCH resulted in the conjugation of up to seven aminoBPs per molecule of fetuin. These conjugates gave a 2.6-, 2.0-, 30.5-, and 1.84-fold increased affinity for untreated, ashed, demineralized bone and hydroxyapatite, respectively, as compared to conjugates from the SMCC reaction. Both conjugates exhibited a pH-independent, equally slow degradation in adult bovine serum-containing media.

Conclusion. The use of the MMCCH chemistry to conjugate aminoBP onto fetuin was feasible. Furthermore, the described processes of conjugation resulted in amino-BP-dependent increase in the glycoprotein's affinity to various bone matrices in a manner that exceeds the affinity produced by the previously established method, which used SMCC.

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Authors and Affiliations

  1. Faculty of Pharmacy & Pharmaceutical Sciences, University of Alberta, Edmonton, Alberta, T6G 2G6, Canada

    Sébastien A. Gittens & Hasan Uludağ

  2. Department of Biomedical Engineering, Faculty of Medicine, University of Alberta, Edmonton, Alberta, T6G 2G6, Canada

    Sébastien A. Gittens

  3. Department of Cell Biology & Anatomy, Faculty of Medicine, University of Calgary, Calgary, Alberta, Canada

    John R. Matyas

  4. Department of Surgery, Faculty of Medicine, University of Calgary, Calgary, Alberta, Canada

    Ronald F. Zernicke

  5. Faculty of Kinesiology, University of Calgary, Calgary, Alberta, Canada

    Ronald F. Zernicke

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  1. Sébastien A. Gittens
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  2. John R. Matyas
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  3. Ronald F. Zernicke
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  4. Hasan Uludağ
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Correspondence to Hasan Uludağ.

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Gittens, S.A., Matyas, J.R., Zernicke, R.F. et al. Imparting Bone Affinity to Glycoproteins Through the Conjugation of Bisphosphonates. Pharm Res 20, 978–987 (2003). https://doi.org/10.1023/A:1024445903306

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