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Synthesis of a New Tri-Branched PEG-IFNα2 and Its Impact on Anti Viral Bioactivity

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Abstract

Efficacy of proteins can be enhanced by using polyethylene glycol (PEG) conjugation (PEGylation) to the protein molecules. Mobile non-toxic PEG chains conjugated to bio-therapeutics increase their hydrodynamic volume and in turn can prolong their plasma retention time and increase their solubility. An important aspect of PEGylation is the selection of PEG molecule with suitable structure and molecular weight. In this study, conceiving the idea that branched PEG-conjugates show superior efficacy over the linear PEG-conjugates, a tri-branched PEG-interferon (mPEG3L2-IFN) was synthesized by reacting a 30 KDa tri-branched mPEG3L2-NHS reagent with IFN to improve its pharmacokinetic properties and reduce the loss of in vitro bioactivity (which is generally exhibited by PEGylation) of the conjugated protein to some extent. The PEGylation procedure was optimized in terms of concentration and molar ratios of reactants, reaction time, temperature and pH conditions of the reaction mix. The conjugate was purified by cation exchange chromatography and characterized by SDS-PAGE and SE-HPLC. Trypsin digestion of mPEG3L2-IFN indicated a single site specificity of PEGylation. Anti viral bioactivity of mPEG3L2-IFN was found to be 2.38 × 107 IU/mg which is approximately 9.52% of native IFNα2 (2.5 × 108 IU/mg) and better than PEGasys from Roche Pharma. Therefore, it is reported that the tri-branched mPEG3L2-NHS reagent has the potential to be used to conjugate proteins for the promising therapeutic results.

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Acknowledgements

This study was supported by Centre for Applied Molecular Biology, Lahore, Pakistan. The authors are grateful to biopharmaceutical lab members of Centre of Excellence in Molecular Biology, Lahore for their cooperation to conduct this study. We also thank Dr. Ahmad Usman Zafar for bioactivity determination and helping in toxicity tests. We are also highly grateful to Prof. Dr. S. Riazuddin for his valuable guidance and providing us the opportunity to conduct this study.

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Authors and Affiliations

  1. Centre of Excellence in Molecular Biology, University of the Punjab, 87-West Canal Bank Road, Thokar Niaz Baig, Lahore, 53700, Pakistan

    Muhammad Farooq Sabar, Mustansara Yaqub, Mohsin Ahmad Khan, Nadeem Ahmad, Muhammad Usman Ghani & Mariam Shahid

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  1. Muhammad Farooq Sabar
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  2. Mustansara Yaqub
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  3. Mohsin Ahmad Khan
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  4. Nadeem Ahmad
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  5. Muhammad Usman Ghani
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  6. Mariam Shahid
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Correspondence to Muhammad Farooq Sabar.

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Sabar, M.F., Yaqub, M., Khan, M.A. et al. Synthesis of a New Tri-Branched PEG-IFNα2 and Its Impact on Anti Viral Bioactivity. Int J Pept Res Ther 16, 239–245 (2010). https://doi.org/10.1007/s10989-010-9219-7

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  • DOI: https://doi.org/10.1007/s10989-010-9219-7

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