**6. A case study of the production of cationic liposomes and gene vaccines in scalable top-down processing**

Although top-down strategies are important and high shear processing has various uses, no systematic studies have been carried out on the effects of liposome comminution on the physico-chemical and surface properties of liposomes. The data in the literature are sparse in regards to the kind of impellers, comminution equipment, or type of lipids used.

Aiming to produce the gene vaccine composed of EPC/DOTAP/DOPE liposomes with DNA complexed on their external surface, we initially studied the significance of the process variables for the properties of liposomes composed of egg lecithin. Mechanical forces were used for homogenation and comminution in Caules type stirrer, Ultra-Turrax, and microchannel microfluidizer equipment. A main variable was selected and its effects on the physico-chemical properties of the liposomes characterized. Finally, a scalable discontinuous process was established and EPC/DOTAP/DOPE liposomes produced and complexed with DNA. The physico-chemical and biological properties of the gene vaccine were compared with our previous gene vaccine prepared using Bangham's method.
