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AAPS PharmSciTech. 2006 December; 7(4): E118–E122.
Published online 2014 March 30. doi:  10.1208/pt070499
PMCID: PMC2750336

Aggregation of recombinant human interferon alpha 2b in solution: Technical note

Summary and Conclusions

Sodium phosphate buffer increased the aggregation of rhIFN-α2b in the range of 1.55 to 1.8103 day−1, as determined by SDS/PAGE under reduced and nonreduced conditions. In contrast, sodium citrate buffer decreased the aggregation rate of this cytokine, as compared with those samples in sodium phosphate buffer. Results from sodium citrate-phosphate buffer were very similar to those obtained with sodium citrate solutions.

On the other hand, EDTA Na2×2H2O reduced the aggregation rate of rhIFN-α2b, showing an aggregation kinetic constant in the range of 0.52 to 0.75×103 day−1. Polysorbates 20 and 80 were less effective than the chelating agent in preventing this degradation pathway.

Additionally, metal ions (Zn2+ and Cu2+) increased the aggregation kinetic constant of rhIFN-α2b, probably through undetermined metal-catalyzing reactions.

Taken together, these data can be useful for the development of new formulations containing rhIFN-α2b as an active ingredient.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.
1. Manning MC, Patel K, Borchardt RT. Stability of protein pharmaceuticals. Pharm Res. 1989;6:903–918. doi: 10.1023/A:1015929109894. [PubMed] [Cross Ref]
2. Bordens R, Grossberg SE, Trotta PP, Nagabhushan TL. Molecular and biologic characterization of recombinant interferon-alpha2b. Semin Oncol. 1997;24:S9-41–S9-51. [PubMed]
3. Sasson A. Biotechnology applied to the production of pharmaceuticals and vaccines. Havana, Cuba: Elfos Scientiae; 1998.
4. Yuen P, Kline D, inventors; Schering Corporation, assignee. Stable aqueous alpha interferon solution formulations. US patent 5 766 582. June 16. 1998.
5. Kirkwood J. Cancer immunotherapy: the interferon-alpha experience. Semin Oncol. 2002;29:18–26. doi: 10.1053/sonc.2002.33078. [PubMed] [Cross Ref]
6. Davis GL, Esteban-Mur R, Rustgi V, et al. Interferon alfa-2b alone or in combination with ribavirin for the treatment of relapse of chronic hepatitis C. International Hepatitis Interventional Therapy Group. N Engl J Med. 1998;339:1493–1499. doi: 10.1056/NEJM199811193392102. [PubMed] [Cross Ref]
7. Ruiz L, Reyes N, Duany L, Franco A, Aroche K, Rando EH. Long-term stabilization of recombinant human interferon α 2b in aqueous solution without serum albumin. Int J Pharm. 2003;264:57–72. doi: 10.1016/S0378-5173(03)00388-0. [PubMed] [Cross Ref]
8. Wang W. Instability, stabilization, and formulation of liquid protein pharmaceuticals. Int J Pharm. 1999;185:129–188. doi: 10.1016/S0378-5173(99)00152-0. [PubMed] [Cross Ref]
9. Beldarrain A, Cruz C, Cruz O, Navarro M, Gil M. Purification and conformational properties of a human interferon alpha 2b produced in Escherichia coli. Biotechnol Appl Biochem. 2001;33:173–182. doi: 10.1042/BA20010001. [PubMed] [Cross Ref]
10. Laemmli UK. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970;227:680–685. doi: 10.1038/227680a0. [PubMed] [Cross Ref]
11. Sigarroa A. Biometry and Experimental Design. Havana, Cuba: Pueblo y Educación; 1985. pp. 383–396.
12. Morehead H, Johnston PD, Wetzel R. Roles of the 29–138 disulfide bond of subtype A of human alpha interferon in its antiviral activity and conformational stability. Biochemistry. 1984;23:2500–2507. doi: 10.1021/bi00306a028. [PubMed] [Cross Ref]
13. Chen BL, Arakawa T, Morris CF, Kenney WC, Wells CM, Pitt CG. Aggregation pathway of recombinant human keratinocyte growth factor and its stabilization. Pharm Res. 1994;11:1581–1587. doi: 10.1023/A:1018905720139. [PubMed] [Cross Ref]
14. Fransson J, Hagman A. Oxidation of human insulin-like growth factor I in formulation studies, II: effects of oxygen, visible light, and phosphate on methionine oxidation in aqueous solution and evaluation of possible mechanisms. Pharm Res. 1996;13:1476–1481. doi: 10.1023/A:1016015226211. [PubMed] [Cross Ref]
15. Fields G, Alonso D, Stiger D, Dill K. Theory for the aggregation of proteins and copolymers. J Phys Chem. 1992;96:3974–3981. doi: 10.1021/j100189a013. [Cross Ref]
16. Speed MA, Wang DIC, King J. Specific aggregation of partially folded polypeptide chains: the molecular basis of inclusion body composition. Nat Biotechnol. 1996;14:1283–1287. doi: 10.1038/nbt1096-1283. [PubMed] [Cross Ref]
17. Cleland JL, Powel MF, Shire SJ. The development of stable protein formulations: a close look at protein aggregation, deamidation, and oxidation. Crit Rev Ther Drug Carrier Syst. 1993;10:307–377. [PubMed]
18. Bam NB, Cleland JL, Yang J, et al. Tween protects recombinant human growth hormone against agitation-induced damage via hydrophobic interactions. J Pharm Sci. 1998;87:1554–1559. doi: 10.1021/js980175v. [PubMed] [Cross Ref]
19. Ha E, Wang W, Wang YJ. Peroxide formation in polysorbate 80 and protein stability. J Pharm Sci. 2002;91:2252–2264. doi: 10.1002/jps.10216. [PubMed] [Cross Ref]
20. Reyes N, Ruiz LI, Aroche K, Gerónimo H, Brito O, Hardy E. Stability of Ala 125 recombinant human interleukin-2 in solution. J Pharm Pharmacol. 2005;57:31–37. doi: 10.1211/0022357055182. [PubMed] [Cross Ref]

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