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The Effect of Formulation Excipients on Protein Stability and Aerosol Performance of Spray-Dried Powders of a Recombinant Humanized Anti-IgE Monoclonal Antibody1

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Abstract

Purpose. To study the effect of trehalose, lactose, and mannitol on the biochemical stability and aerosol performance of spray-dried powders of an anti-IgE humanized monoclonal antibody.

Methods. Protein aggregation of spray-dried powders stored at various temperature and relative humidity conditions was assayed by size exclusion chromatography and sodium dodecyl sulfate polyacrylamide gel electrophoresis. Protein glycation was determined by isoelectric focusing and affinity chromatography. Crystallization was examined by X-ray powder diffraction. Aerosol performance was assessed as the fine particle fraction (FPF) of the powders blended with coarse carrier lactose, and was determined using a multiple stage liquid impinger.

Results. Soluble protein aggregation consisting of non-covalent and disulfide-linked covalent dimers and trimers occurred during storage. Aggregate was minimized by formulation with trehalose at or above a molar ratio in the range of 300:1 to 500:1 (excipient:protein). However, the powders were excessively cohesive and unsuitable for aerosol administration. Lactose had a similar stabilizing effect, and the powders exhibited acceptable aerosol performance, but protein glycation was observed during storage. The addition of mannitol also reduced aggregation, while maintaining the FPF, but only up to a molar ratio of 200:1. Further increased mannitol resulted in crystallization, which had a detrimental effect on protein stability and aerosol performance.

Conclusions. Protein stability was improved by formulation with carbohydrate. However, a balance must be achieved between the addition of enough stabilizer to improve protein biochemical stability without compromising blended powder aerosol performance.

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REFERENCES

  1. L. G. Presta, S. J. Lahr, R. L. Schields, J. P. Porter, C. M. Gorman, B. M. Fendly, and P. M. Jardieu. Humanization of an antibody directed against IgE. J. Immunol. 151:2623-2632 (1993).

    Google Scholar 

  2. J. Liu, P. Lester, S. Builder, and S. J. Shire. Characterization of complex formation by humanized anti-IgE monoclonal antibody and monoclonal human IgE. Biochemistry 34:10474-10482 (1995).

    Google Scholar 

  3. J. V. Fahy, H. E. Fleming, H. H. Wong, J. T. Liu, J. Q. Su, J. Reimann, R. B. Fick Jr., and D. W. Cockcroft. The effect of an anti-IgE monoclonal antibody on the early-and late-phase responses to allergen inhalation in asthmatic subjects. Am. J. Resp. Critical Care Med. 155:1828-1834 (1997).

    Google Scholar 

  4. L. P. Boulet, K. R. Chapman, J. Cote, S. Kalra, R. Bhagat, V. A. Swystun, M. Laviolette, L. D. Cleland, F. Deschesnes, J. Q. Su, A. DeVault, R. B. Fick Jr., and D. W. Cockcroft. Inhibitory effects of an anti-IgE antibody E25 on allergen-induced early asthmatic response. Am. J. Resp. Critical Care Med. 155:1835-1840 (1997).

    Google Scholar 

  5. R. W. Niven. Delivery of biotherapeutics by inhalation aerosols. Pharm. Technol. 17(7):72-82 (1993).

    Google Scholar 

  6. J. Broadhead, S. K. Rouan Edmond, and C. T. Rhodes. The spray drying of pharmaceuticals. Drug Dev. Ind. Pharm. 18:1169-1206 (1992).

    Google Scholar 

  7. J. Broadhead, S. K. Rouan Edmond, I. Hau, and C. T. Rhodes. The effect of process and formulation variables on the properties of spray-dried β-galactosidase. J. Pharm. Pharmacol. 46:458-467 (1994).

    Google Scholar 

  8. M. Mumenthaler, C. C. Hsu, and R. Pearlman. Feasibility study on spray-drying protein pharmaceuticals. Recombinant human growth hormone and tissue-type plasminogen activator. Pharm. Res. 11:12-20 (1994).

    Google Scholar 

  9. H-K. Chan, A. R. Clark, I. Gonda, M. Mumenthaler, and C. C. Hsu. Spray-dried powders and powder blends of recombinant human deoxyribonuclease (rhDNase) for aerosol delivery. Pharm. Res. 14:431-437 (1997).

    Google Scholar 

  10. Y-F. Maa, H. R. Costantino, P-A. Nguyen, and C. C. Hsu. The effect of operating and formulation variables on the morphology of spray-dried protein particles. Pharm. Dev. Technol. 2:213-223 (1997).

    Google Scholar 

  11. J. L. Cleland, M. F. Powell, and S. J. Shire. The development of stable protein formulations. A close look at protein aggregation, deamidation, and oxidation. Critical Rev. Ther. Drug Carrier Sys. 10:307-377 (1993).

    Google Scholar 

  12. A. R. Clark, N. Dasovich, I. Gonda, and H-K. Chan. The balance between biochemical and physical stability for inhalation protein powders. rhDNase as an example. In R. N Darby, P. R. Byron, and S. J. Farr (eds.), Respiratory Drug Delivery V, Interpharm Press, Buffalo Grove, 1996, pp. 167-174.

    Google Scholar 

  13. J. F. Young. Humidity control in the laboratory using salt solutions. A review. J. Appl. Chem. 17:241-245 (1967).

    Google Scholar 

  14. R. A. Gray, A. Stern, T. A. Bewley, D. McRorie, P. Voelker, J. Liu, and S. J. Shire. Determination of protein concentration by improved optical methods. Pharm. Res. 11:76 (1994).

    Google Scholar 

  15. J. Liu, J. Ruppel, and S. J. Shire. Interaction of human IgE with soluble forms of IgE high affinity receptors. Pharm. Res. 14:1388-1393 (1997).

    Google Scholar 

  16. Y-F. Maa, P-A. Nguyen, J. D. Andya, N. Dasovich, S. J. Shire, and C. C. Hsu. Effect of spray drying and subsequent processing on residual moisture content and physical/biochemical stability of protein inhalation powders. Pharm. Res. 15:768-775 (1998).

    Google Scholar 

  17. A. Saleki-Gerhardt and G. Zografi. Non-isothermal and isothermal crystallization of sucrose from the amorphous state. Pharm. Res. 11:1166-1173 (1994).

    Google Scholar 

  18. L. M. Crowe, D. S. Reid, and J. H. Crowe. Is trehalose special for preserving dry biomaterials? Biophysical J. 71:2087-2093 (1996).

    Google Scholar 

  19. F. Franks, B. Aldous, and T. Auffret. Using water-soluble glasses as a strategy for bioproduct stabilization. Gen. Eng. News 15:20 (1995).

    Google Scholar 

  20. J. F. Carpenter, M. J. Pikal, B. S. Chang, and T. W. Randolph. Rational design of stable lyophilized protein formulations. Some practical advice. Pharm. Res. 14:969-975 (1997).

    Google Scholar 

  21. D. B. Volkin and C. R. Middaugh. The effect of temperature on protein structure. In T. J. Ahern, and M. C. Manning (Eds.), Stability of Protein Pharmaceuticals, Part A Chemical and Physical Pathways of Protein Degradation, Plenum Press, New York, 1993, pp. 215-247.

    Google Scholar 

  22. H. R. Costantino, R. Langer, and A. M. Klibanov. Solid-phase aggregation of proteins under pharmaceutically relevant conditions. J. Pharm. Sci. 83:1662-1669 (1994).

    Google Scholar 

  23. M. J. Pikal. Freeze-drying of proteins. Process, formulation, and stability. In J. L. Cleland, and R. Langer (Eds.), Formulation and Delivery of Proteins and Peptides. American Chemical Society, Washington, DC, 1994, pp. 120-133.

    Google Scholar 

  24. L. N. Bell, M. J. Hageman, and L. M. Muraoka. Thermally induced denaturation of lyophilized bovine somatotropin and lysozyme as impacted by moisture and excipients. J. Pharm. Sci. 84:707-712 (1995).

    Google Scholar 

  25. H. R. Costantino, K. G. Carrasquillo, R. A. Cordero, M. Mumenthaler, C. C. Hsu, and K. Griebenow. The effect of excipients on the stability and structure of lyophilized recombinant human growth hormone (rhGH). J. Pharm. Sci., in press, 1998.

  26. H. R. Costantino, J. D. Andya, P-A. Nguyen, N. Dasovich, T. D. Sweeney, S. J. Shire, C. C. Hsu and Y-F. Maa. The effect of mannitol crystallization on the stability and aerosol performance of a spray-dried pharmaceutical protein, recombinant humanized anti-IgE monoclonal antibody. J. Pharm. Sci., 18:1406-1411 (1998).

    Google Scholar 

  27. J. F. Carpenter, and J. H. Crowe. An infrared spectroscopic study of the interactions of carbohydrates with dried proteins. Biochemistry 28: 3916-3922 (1989).

    Google Scholar 

  28. S. J. Prestrelski, K. A. Pikal, and T. Arakawa. Optimization of lyophilization conditions for recombinant human interleukin-2 by dried-state conformational analysis using fourier-transform infrared spectroscopy. Pharm Res. 12:1250-1259 (1995).

    Google Scholar 

  29. H. R. Costantino, J. D. Andya, S. J. Shire, and C. C. Hsu. Fouriertransform infrared spectroscopic analysis of the secondary structure of recombinant humanized immunoglobulin G. Pharm. Sci. 3:121-128 (1997).

    Google Scholar 

  30. H. R. Costantino, J. G. Curley, and C. C. Hsu. Determining the water sorption monolayer of lyophilized pharmaceutical proteins. J. Pharm. Sci. 86:1390-1393 (1997).

    Google Scholar 

  31. A. J. Furth. Methods for assaying nonenzymatic glycosylation. Anal. Biochem. 175:347-360 (1988).

    Google Scholar 

  32. F. J. Shen, J. D. Andya, S. J. Shire, and R. J. Harris. The detection of lactosylated sites. Protein Sci. 6(Suppl. 2):152 (1997).

    Google Scholar 

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Author notes

  1. Henry R. Costantino

    Present address: Alkermes, Inc., 64 Sidney St., Cambridge, Massachusetts, 02139

Authors and Affiliations

  1. Pharmaceutical Research and Development, Genentech, Inc., 1 DNA Way, South San Francisco, California, 94080

    James D. Andya, Yuh-Fun Maa, Henry R. Costantino, Phuong-Anh Nguyen, Nancy Dasovich, Theresa D. Sweeney, Chung C. Hsu & Steven J. Shire

  2. Alkermes, Inc., 64 Sidney St., Cambridge, Massachusetts, 02139

    Yuh-Fun Maa

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  1. James D. Andya
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Correspondence to James D. Andya.

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Andya, J.D., Maa, YF., Costantino, H.R. et al. The Effect of Formulation Excipients on Protein Stability and Aerosol Performance of Spray-Dried Powders of a Recombinant Humanized Anti-IgE Monoclonal Antibody1. Pharm Res 16, 350–358 (1999). https://doi.org/10.1023/A:1018805232453

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