Microfluidic-assisted fabrication of phosphatidylcholine-based liposomes for controlled drug delivery of chemotherapeutics

Abstract

Microfluidic enables precise control over the continuous mixing of fluid phases at the micrometre scale, aiming to optimize the processing parameters and to facilitate scale-up feasibility. The optimization of parameters to obtain monodispersed drug-loaded liposomes however is challenging. In this work, two phosphatidylcholines (PC) differing in acyl chain length were selected, and used to control the release of the chemotherapeutic agent doxorubicin hydrochloride, an effective drug used to treat breast cancer. Microfluidics was used to rapidly screen manufacturing parameters and PC formulations to obtain monodispersed unilamellar liposomal formulations with a reproducible size (i.e. < 200 nm). Cholesterol was included in all liposomal formulations; some formulations also contained DMPC(1,2-dimyristoyl-sn-glycero-3-phosphocholine) and/or DSPC(1,2-distearoyl-sn-glycero-3-phosphocholine). Systematic variations in microfluidics total flow rate (TFR) settings were performed, while keeping a constant flow rate ratio (FRR). A total of six PC-based liposomes were fabricated using the optimal manufacturing parameters (TFR 500 μL/min, FRR 0.1) for the production of reproducible, stable liposome formulations with a narrow size distribution. Liposomes actively encapsulating doxorubicin exhibited high encapsulation efficiencies (>80%) for most of the six formulations, and sustained drug release profiles in vitro over 48 h. Drug release profiles varied as a function of the DMPC/DSPC mol content in the lipid bilayer, with DMPC-based liposomes exhibiting a sustained release of doxorubicin when compared to DSPC liposomes. The PC-based liposomes, with a slower release of doxorubicin, were tested in vitro, as to investigate their cytotoxic activity against three human breast cancer cell lines: the non-metastatic ER+/PR + MCF7 cells, the triple-negative aggressive MDA-MB 231 cells, and the metastatic HER2-overexpressing/PR + BT474 cells. Similar cytotoxicity levels to that of free doxorubicin were reported for DMPC5 and DMPC3 binary liposomes (IC50 ~ 1 μM), whereas liposomes composed of a single PC were less cytotoxic (IC50 ~ 3–4 μM). These results highlight that microfluidics is suitable for the manufacture of monodispersed and size-specific PC-based liposomes in a controlled single-step; furthermore, selected PC-based liposome represent promising nanomedicines for the prolonged release of chemotherapeutics, with the aim of improving outcomes for patients.