(M0930-05-29) Exploring the Effect of Manufacturing Methods in Long-Acting Injectable Formulation Characteristics

Purpose: There are several FDA approved drug product formulations with the long-acting characteristics utilizing poly(lactic-co-glycolic) acid (PLGA). These products include small molecule-based active pharmaceutical ingredients, there has been a growing interest recently for the development of long-acting formulations for biologics. Over the past several years, our group has developed long-acting injectable formulations of PLGA microparticles (MPs) to deliver several different biologics in the inflammatory disease models such as periodontal disease, dry eye disease, and transplant rejection (1-3). These formulations have been produced by utilizing the conventional batch methods which require high energy agitation. However, the use of sonication/homogenization may result in activity loss in the therapeutic agent, especially for protein-based formulations, thus causing a potential barrier to clinical translation (4). Microfluidic-based fabrication methods bring an alternative approach to the development of drug products by reducing the shear stress on the drug-containing droplets and may lead to improve the overall bioactivity of the biologics (5). In this work, we explored the effect of manufacturing methods such as conventional homogenization and microfluidics methods in long-acting injectable PLGA formulation characteristics.

Methods: A model protein, recombinant human CCL22 (rhCCL22), encapsulating PLGA MPs were prepared by a double emulsion solvent evaporation method using two different approaches. For the conventional homogenization method, water droplets of the protein solution were first formed in the PLGA dissolved dichloromethane solution. Then, PLGA droplets were generated via high-speed homogenization of this initial emulsion in the surfactant-containing water solution. Finally, dichloromethane was evaporated to form the PLGA MPs. In the case of the microfluidics method, a hydrodynamic flow focusing chip was used to generate the PLGA droplets after preparing the water droplets of protein solution in the PLGA solution. These droplets were then processed similarly to the conventional homogenization method to remove the solvent and to form the PLGA MPs. Further, the MPs size, loading, release kinetics and bioactivities of encapsulated rhCCL22 were characterized by Coulter Counter volume impedance measurements, scanning electron microscopy imaging, and in vitro release/bioactivity assays.

Results: The conventional homogenization method resulted in rhCCL22 encapsulating PLGA MPs with porous surface morphology. MPs demonstrated a broad range of size distribution in the 10-50 mm range with an average MP size of 23 ± 8.7 mm. The rhCCL22 release kinetics followed a linear trend for the first 21 days of the protein release (Figure 1). However, PLGA MPs that were prepared by the microfluidics method showed a uniform size distribution with an average size of 27 ± 2.5 mm and a similar surface morphology. Interestingly, the rhCCL22 release rate from these MPs was depressed as compared to the release profile of MPs that were prepared by the conventional homogenization method (Figure 2). Additionally, we observed a slight difference in batch-to-batch variations in the context of size distribution of PLGA MPs and the protein release rates (Figure 3) especially in the case of conventional homogenization method. The effect of fabrication method on the bioactivity of rhCCL22 will be discussed.

Conclusion: Microfluidic methods could be an alternative fabrication approach for producing long-acting injectable drug formulations with reduced variation in batches, and potentially improved bioactivity of encapsulated biologics as compared to conventional homogenization methods.

References: (1) A. C. Greene, M. Shehabeldin, J. Gao, S. C. Balmert, M. Ratay, C. Sfeir, S. R. Little, Scientific Reports, 2022, 12, 1.
(2) M. L. Ratay, S. C. Balmert, A. P. Acharya, A. C. Greene, T. Meyyappan, S. R. Little, Scientific Reports, 2017, 7, 1.
(3) J. D. Fisher, W. Zhang, S. C. Balmert, A. M. Aral, A. P. Acharya, Y. Kulahci, J. Li, H. R. Turnquist, A. W. Thomson, M. G. Solari, V. S. Gorantla, S. R. Little, Science Advances, 2020, 6, 11.
(4) S. P. Schwendeman, Crit. Rev. Ther. Drug Carrier Syst. 2002, 19, 73.
(5) D. Liu, H. Zhang, F. Fontana, J. T. Hirvonen, H. A. Santos, Lab Chip, 2017, 17, 1856.

Acknowledgements: Nihan Yonet-Tanyeri and Steven R. Little are funded by NIH grant R01-AR074285. Louis D. Falo is supported by the NIH grants R01-AR074285, and R01-AR071277.


Figure 1. Characterization of protein encapsulating PLGA MPs that are fabricated by conventional homogenization method. (a) Scanning electron microscopy image of protein encapsulating PLGA MPs. (b) Size distribution profile of protein encapsulating MPs. (c) Cumulative protein release curve from PLGA MPs.


Figure 2. Characterization of protein encapsulating PLGA MPs that are fabricated by microfluidics method. (a) Scanning electron microscopy image of protein encapsulating PLGA MPs. (b) Size distribution profile of protein encapsulating MPs. (c) Cumulative protein release curve from PLGA MPs.


Figure 3. A comparative analysis on the batch-to-batch variation of protein encapsulating PLGA MPs as a function of the manufacturing method, conventional homogenization vs microfluidics. (a, b) The change in the size distribution profiles of protein encapsulating MPs. (b) The change in the cumulative protein release from PLGA MPs.