Microfluidic Manufacture of Lipid-Based Nanomedicines

Abstract

Nanoparticulate technologies have revolutionized drug delivery allowing for passive and active targeting, altered biodistribution, controlled drug release (temporospatial or triggered), enhanced stability, improved solubilization capacity, and a reduction in dose and adverse effects. However, their manufacture remains immature, and challenges exist on an industrial scale due to high batch-to-batch variability hindering their clinical translation. Lipid-based nanomedicines remain the most widely approved nanomedicines, and their current manufacturing methods remain discontinuous and face several problems such as high batch-to-batch variability affecting the critical quality attributes (CQAs) of the product, laborious multistep processes, need for an expert workforce, and not being easily amenable to industrial scale-up involving typically a complex process control. Several techniques have emerged in recent years for nanomedicine manufacture, but a paradigm shift occurred when microfluidic strategies able to mix fluids in channels with dimensions of tens of micrometers and small volumes of liquid reagents in a highly controlled manner to form nanoparticles with tunable and reproducible structure were employed. In this review, we summarize the recent advancements in the manufacturing of lipid-based nanomedicines using microfluidics with particular emphasis on the parameters that govern the control of CQAs of final nanomedicines. The impact of microfluidic environments on formation dynamics of nanomaterials, and the application of microdevices as platforms for nanomaterial screening are also discussed.

Keywords: engineering; liposomes; manufacture; microfluidics; nanomedicine; scale-up.

Figure 1

Schematic representation of lipid-based nanomedicines. Liposomes (A–D): Hydrophobic molecules up to few nm in diameter can be entrapped in the phospholipid bilayer (red spheres), while hydrophilic cargo can be loaded in the core (purple pentagon) and their surface can be modified antibodies (hydrophobically anchored (i) or conjugated via a linker or a hydrophilic polymer chain (immunoliposomes (ii)) (A). Liposomes with cavitands able to allow host–guest chemical reactions with molecules of complementary shape or size to allow loading in the bilayer, cavitands, and core (B). Stealth liposomes and targeted stealth liposomes where the liposome surface is decorated with hydrophilic polymer chains such as polyethylene glycol or a stimulus-responsive polymer, and a targeting moiety or diagnostic moiety (blue square) can be conjugated (peptides, cell-penetrating peptides, and antibodies). Drugs, genetic material, or diagnostic agents (gold, silver, or magnetic particles) can be loaded in the bilayer, core, or surface via conjugation, and lipids can be negatively or positively charged (preferred for complexation with DNA/RNA). Micelles or inverse micelles (E) are prepared via self-assembly of amphiphiles such as phospholipids and can load hydrophobic or hydrophilic molecules. Solid lipid nanoparticles (SLNs) (F) are colloidal carriers where liquid lipids have been substituted by a solid lipid, offering unique properties such as small size, large surface area, high drug loading, and the interaction of phases at the interfaces, and they are attractive for their potential to improve performance of pharmaceuticals, nutraceuticals, and other materials, appearing in three forms depending on where drug is loaded (homogeneous matrix (melting point of drug equal to that of lipid), lipid-enriched core (melting point of drug < lipid), and drug-enriched core (melting point of drug > lipid)). Nanostructured lipid carriers (NLCs) (G) are colloidal carriers prepared by blending of solid lipids with oils, but the matrix remains solid at body temperature to overcome problems of SLNs (low payload for drugs, drug expulsion during storage, and high water content of SLN dispersions). Cochleates (H) are phospholipid–calcium precipitates derived from the interaction of anionic lipid vesicles with divalent cations such as calcium with a multilayered structure consisting of large and continuous lipid bilayer sheets rolled up in a spiral structure with no internal aqueous phases.

Figure 2

Summary of schematic designs of microfluidic mixers for lipid nanoparticle development: (A) T-shaped mixer, (B) hydrodynamic flow focusing, (C) bifurcating mixers, (D) chaotic, staggered micromixers, and (E) baffle mixers.

Figure 3

Process of making microfluidic devices using PDMS. Different materials are used to produce the mold, but SU-8 is usually chosen in the production of PDMS-based microfluidic devices. Once the mold has been prepared with the appropriate steps, the next step is casting, followed by hardening and release of PDMS from the mold. The PDMS is deposited on the mold; everything is placed in the oven for 24 h at 65 °C so that the PDMS cures and, once hardened, can be easily removed from the mold. Then, the bonding phase follows, where the surface of the PDMS is generally exposed to oxygen plasma for 10 min and then in contact with a layer of glass or another layer of PDMS to generate a bond. The process ends with the interfacing and integration phase where input and output zones are created with the help of needles, in the case of temporary applications, or with specific structures for longer applications. Reprinted with permission from [207]. Copyright 2022, AIP Publishing LLC.

Figure 4

Microfluidic techniques for liposome and lipid nanoparticle (LNP) formulation. Summary of bulk and microfluidic techniques for production of liposomes (A,B) and lipid nanoparticles (C–F), highlighting advantages (green) and disadvantages (red) for each. Reproduced with permission from [227] and used under the Creative Commons license permission (CC BY 4.0). Copyright 2021, Elsevier Ltd. All rights reserved.

Figure 5

A Schematic diagram depicting a hypothesized LNP formation mechanism. The formation of LNPs in (a) slower and (b) faster mixing conditions. (c) Schematic representation of the formation of LNPs at the interface of ethanol–saline. The process starts with the aggregation of lipids in discs (A). The hydrophobic chains around the edges are stabilized by alcohol molecules and, as the alcohol concentration reduces, these lipid discs bend (B) and rapidly close (C) and form spherical vesicles (D). Thus, the polarity change during the liposome formation process is related to the initial polarity of the organic phase. The figure is adapted from Copyright: © 2017 Maeki et al. [196] under the terms of the Creative Commons Attribution License. All rights reserved.

Figure 6

Schematic of (a) iLiNP (two inlets) and (b) micromixer device. Modified from [299] and used under the Creative Commons license permission (CC BY 4.0). Copyright 2022, American Chemical Society. All rights reserved.