Influence of Formulation Factors on the Aerosol Performance and Stability of Lysozyme Powders: a Systematic Approach
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With the growing interest in developing biologics for pulmonary delivery, systematic fast screening methods are needed for rapid development of formulations. Due to the labile nature of macromolecules, the development of stable, biologically active formulations with desired aerosol performance imposes several challenges both from a formulation and processing perspective. In this study, spray-freeze-drying was used to develop respirable protein powders. In order to systematically map the selected design space, lysozyme aqueous pre-formulations were prepared based on a constrained mixture design of experiment. The physicochemical properties of the resulting powders were characterized and the effects of formulation factors on aerosol performance and protein stability were systematically screened using a logic flow chart. Our results elucidated several relevant formulation attributes (density, total solid content, protein:sugars ratio) required to achieve a stable lysozyme powder with desirable characteristics for pulmonary delivery. A similar logical fast screening strategy could be used to delineate the appropriate design space for different types of proteins and guide the development of powders with pre-determined aerodynamic properties.
KEY WORDSdry powder biologics spray-freeze-drying pulmonary delivery dry powder inhaler
The authors are thankful to the members of Smyth lab (in particular to Dr. Matthew Herpin and Daniel Moraga), to Dr. Abbe Haser, and to the team at Amgen (in particular Nitin Anghan and Michael Schneider) for valuable feedbacks.
This work was supported by Amgen Inc.
Compliance with Ethical Standards
The terms of this arrangement have been reviewed and approved by the University of Texas at Austin in accordance with its policy on objectivity in research.
Conflict of Interest
The author (HDCS) of this paper consults for and has equity ownership in Respira Therapeutics and Nob Hill Therapeutics on inhaled product development. The other authors declare that they have no conflict of interest.
- 1.Morales JO, Fathe KR, Brunaugh A, Ferrati S, Li S, Montenegro-Nicolini M, et al. Challenges and future prospects for the delivery of biologics: oral mucosal, pulmonary, and transdermal routes. AAPS J. 2017:1–17.Google Scholar
- 16.Carpenter JF, Chang BS, Garzon-Rodriguez W, Randolph TW. Rational design of stable lyophilized protein formulations: theory and practice. Rational design of stable protein formulations 2002:109–33.Google Scholar
- 17.Carrasquillo KG, Stanley AM, Aponte-Carro JC, De Jésus P, Costantino HR, Bosques CJ, et al. Non-aqueous encapsulation of excipient-stabilized spray-freeze dried BSA into poly (lactide-co-glycolide) microspheres results in release of native protein. J Control Release. 2001;76(3):199–208.CrossRefGoogle Scholar
- 23.Murugappan S, Patil HP, Kanojia G, ter Veer W, Meijerhof T, Frijlink HW, et al. Physical and immunogenic stability of spray freeze-dried influenza vaccine powder for pulmonary delivery: comparison of inulin, dextran, or a mixture of dextran and trehalose as protectants. Eur J Pharm Biopharm. 2013;85(3):716–25.CrossRefGoogle Scholar
- 24.Amorij J-P, Saluja V, Petersen AH, Hinrichs WL, Huckriede A, Frijlink HW. Pulmonary delivery of an inulin-stabilized influenza subunit vaccine prepared by spray-freeze drying induces systemic, mucosal humoral as well as cell-mediated immune responses in BALB/c mice. Vaccine. 2007;25(52):8707–17.CrossRefGoogle Scholar
- 27.Dailymed. Available from: https://dailymed.nlm.nih.gov/dailymed/.
- 28.Bulk denisty and tap density. The United States Pharmacopeial Convention. 2011;General chapter <616>.Google Scholar
- 29.Physical tests and determinations: inhalation and nasal drug products: aerosols, sprays, and powders—performance quality tests. United States Pharrmacopeia and National Formulary (USP 39-NF 34). Rockville MD: United States Pharmacopeial Convention; 2016. p. 423–49.Google Scholar
- 30.Enzymatic assay of lysozyme (Sigma-Aldrich). Available from: http://www.sigmaaldrich.com/technical-documents/protocols/biology/enzymatic-assay-of-lysozyme.html.
- 38.Ferrati S, Wu T, Fuentes O, Brunaugh AD, Kanapuram SR, Smyth HDC. A systematic analysis of the effect of formulation parameters on the aerosol performance and stability of aerosol performance and stability of spray freeze dried protein powders. Respir Drug Deliv (RDD) Conf Proc. 2018;Google Scholar