How docetaxel entrapment, vesicle size, zeta potential and stability change with liposome composition–A formulation screening study

Limitations of the anticancer drug product Taxotere® have encouraged researchers to entrap the active ingredient docetaxel (DTX) into nanocarriers such as liposomes. However, until now no DTX-liposome formulation has reached the clinic. Hence, in the present study, different Soy-PC based DTX-liposome formulations were screened in an attempt to identify lipid-compositions with promising DTX-entrapment (DTX-EE). Various other quality attributes, such as vesicle size and morphology, poly dispersity index (PDI), zeta potential (ZP), stability and in vitro drug release were also investigated. In an initial study, the inclusion of charged lipids within the liposome bilayer was observed to have a positive effect on DTX-EE. Thus, cationic DOTAP (1,2-Dioleoyl-3-trimethylammonium-propane) and anionic DMPG (1,2-Dimyristoyl-sn-glycero-3-phospho-(1′-rac-glycerol) lipids were selected for further investigations. With anionic DMPG, only a temporary rise in EE was gained with ≥ 20% (w/w) DMPG in Soy-PC lipid-based liposomes, whereas a concentration-dependent increase in EE was observed with cationic DOTAP. A DTX-EE > 95% was obtained with only 5% (w/w) DOTAP in Soy-PC, while neutral liposomes formed from Soy-PC alone, gave 41.5% DTX-EE. In the stability study, a DOTAP concentration > 10% (w/w) in Soy-PC was found to facilitate a stable DTX-EE > 90% after 12 weeks storage. The positive effect of cationic lipids on the EE was confirmed when replacing cholesterol (CHOL), initially shown to suppress DTX-entrapment, with cationic 3ß-[N-(N',N'-dimethylaminoethane)-carbamoyl]Cholesterol (DC-CHOL). Here, DTX-EE was improved from 29.8% to 92.0% (w/w) with 10% (w/w) CHOL and DC-CHOL in Soy-PC, respectively. Finally, PEGylation of DOTAP-liposomes with DSPE-PEG2000 and DSPE-PEG750 reduced the DTX-EE relative to DOTAP-liposome with no PEGylation. As with the DMPG-liposomes, a temporarily raised affinity between DTX and liposomes was obtained with anionic DSPE-PEGylation of Soy-PC liposomes, however, this effect was not maintained after 4 weeks storage. However, in a dialysis set-up, cationic DOTAP-liposomes released DTX to a higher extent than PEGylated liposomes. Thus, the optimal formulation with regard to storage stability and in vivo performance need to be investigated further, applying conditions that are closer to mimic the in vivo-situation. Applying the Dual Asymmetric Centrifugation (DAC) method in liposome production appears favourable due to its good reproducibility. The observed increase in DTX entrapment with cationic lipids or PEGylation appears scalable into pilot manufacturing scale