Process optimization and pre-clinical production of lipid nanoparticles using Sunshine

Significance of the Topic

Lipid nanoparticles (LNPs) are emerging as a critical delivery platform for genetic medicines, including mRNA vaccines and RNA-based therapies. Precise control over nanoparticle size, composition, and encapsulation efficiency is essential to ensure reproducible in vivo performance, reduce batch-to-batch variability, and accelerate translation from lead discovery to clinical production.

Aims and Study Overview

This application note demonstrates how the Sunshine system integrates high-throughput process optimization with scalable continuous manufacturing of LNPs. Using a representative mRNA‐LNP formulation, the study explores key parameters—flow rate ratio (FRR) and total flow rate (TFR)—to achieve target size (~60 nm), low polydispersity index (PDI), and high encapsulation efficiency (EE). The transition from automated screening (“protocol mode”) to liter‐scale continuous production is highlighted.

Methodology and Used Instrumentation

The microfluidic platform employs Unchained Labs’ Sunny 190 XT chip with hydrodynamic flow focusing to assemble LNPs. Automated reagent handling is controlled by the Sunshine system and Sunny Suite Software. Post‐synthesis processing uses Pur‐A‐Lyzer™ Midi Dialysis Kits for solvent exchange. Particle size and PDI are measured by dynamic light scattering (DLS). The RiboGreen assay, read on a FLUOROstar microplate reader, quantifies mRNA encapsulation.

Main Results and Discussion

• Flow Rate Ratio (FRR): Varying the aqueous-to-organic FRR from 2:1 to 6:1 (fixed TFR of 5 mL/min) allowed tuning particle size from 68 nm down to 58 nm post-dialysis, with a consistent average PDI of 0.12. Encapsulation efficiency remained above 90%, peaking at 96% for an FRR of 6:1.
• Total Flow Rate (TFR): At a fixed FRR of 3:1, increasing TFR from 2.5 to 7.5 mL/min reduced particle size from 94 nm to 57 nm and improved PDI (0.089) while maintaining EE above 94%. A slight size increase was observed at 10 mL/min.
• Continuous vs Protocol Mode: Liposomal nanoparticles produced at 3 mL/min and various FRRs in continuous mode matched protocol‐mode performance, with sizes between 19 nm and 57 nm and PDI 0.15–0.20.

Benefits and Practical Applications

The Sunshine system rapidly screens lead formulations at microliter scale, identifies optimal process conditions in minutes, and seamlessly scales to liter‐scale continuous production within the same platform. This dual capability reduces development timelines, material consumption, and process validation burdens, enabling efficient pre‐clinical formulation and manufacturing of nanomedicines.

Future Trends and Opportunities

Advancements may include integrating real‐time analytics (e.g., inline spectroscopy) for closed‐loop process control, exploring new microfluidic chip geometries to further refine particle homogeneity, and expanding the platform to additional nanoparticle types. Process intensification and adaptation for GMP‐compliant workflows will support clinical translation and commercial manufacturing.

Conclusion

The Sunshine microfluidic platform offers an end‐to‐end solution for LNP development, enabling rapid optimization of critical parameters and straightforward scale‐up to continuous manufacturing. By delivering consistent particle characteristics and high encapsulation efficiency, the system accelerates nanomedicine development and streamlines the transition from R&D to pre‐clinical production.

Reference

• Unchained Labs. Process optimization and pre-clinical production of lipid nanoparticles using Sunshine. Application Note, Rev A (2023).