NanoPULSE : The First Micromixing Method Delivering Consistent Lipid Nanoparticle Formulations Seamlessly Across All Scales

The ongoing development of RNA-based vaccines and therapies employing lipid nanoparticle (LNP) formulations is currently hindered by the absence of a unified formulation technology spanning seamlessly the entire development pipeline – from microliter-scale screening to tens-of-liters manufacturing. This gap forces drug developers to transition between different formulation techniques during scale-up. This poses significant challenges as the manufacturing process critically affects all key LNP quality parameters such as particle size, polydispersity index (PDI), drug loading efficiency, or morphology, leading to prolonged, expensive, and sometimes unsuccessful drug development. Currently, state-of-the-art methods predominantly rely on microfluidic-based mixing, which excels at the microliter scale by providing precise mixing under laminar flow conditions. However, these systems face substantial limitations in scalability and throughput. In contrast, T-mixers and impingement jet mixers (IJM) are better suited for large-scale production, but struggle to accommodate the low-volume formulations essential for early-stage development. Here, we introduce NanoPULSE, a novel, patented active micromixing technology that ensures consistent LNP characteristics across all production volumes, from µL-scale screening to continuous, high-volume, aggregation-free manufacturing. The device employs high-frequency valves with rapid response times to sequentially inject aqueous and organic phases into a T-junction, creating fringes that enhance the diffusion within the mixing channel according to the Taylor-Aris Dispersion principle. Lagrangian numerical simulations, performed using a Python-based code, have been used to predict the impact of different input parameters on particle characteristics and optimize the mixing channel’s dimensions. These simulations have led to the introduction of a novel index to quantify the mixing efficiency of NanoPULSE. Dynamic light scattering (DLS) measurements confirm that both empty and RNA-loaded LNPs maintain constant sizes over a production volume range from 500 µL to 4 L, with polydispersity indexes (PDI) remaining consistent and below 0.2, validating the seamless scalability of the technology. The encapsulation efficiencies/yield and structural morphology of RNA-loaded LNPs produced with NanoPULSE are characterized at different scales and compared to those synthesized using standard microfluidic approaches. Finally, by modulating the injection periods of the solvents, NanoPULSE can produce LNPs of variable sizes using the same formulation. The nanoparticle sizes and PDIs measured at different input parameters are correlated with the simulation results, linking the mixing performance to the nanoparticle characteristics, paving the way for a predictive model of LNP synthesis. These findings underscore the promising repeatability, scalability, and efficiency of LNP synthesis via NanoPULSE.