Liposomes formulation techniques

Furhman, L., et al. 1995. Evaluation of several liposomal formulations and preparation techniques for the dermal delivery of phosphorothioate antisense oligonucleotides in hairless mouse skin in vitro. In AAPS Annual Meeting, Miami Beach, FL, USA. 

Finally, remote loading and active loading [11] are other methods used to achieve high trapping efficiency in liposome formulations, but unfortunately they can only be applied to a small number of drugs with specific physicochemical properties. This technique will be discussed below in Section 5.3.23. 

The characterization of liposomes is an important task and many analytical techniques have been employed, Thermodynamic, mechanical, chemical, microscopic, spectroscopic, and chromatographic techniques have being used worldwide in order to test the physical and chemical, characteristics of liposomal formulations such as lamellerity, encapsulation efficiency and their stability overtime. 

Dried Reconstituted Vesicles (DRV) (see Note 1), were initially developed in 1984 by Kirby and Gregoriadis (1). They are oligo-or multilamellar liposomes with capability of encapsulating high amounts of aqueous soluble molecules. The fact that the DRV technique involves vesicle formation under mild conditions (e.g., conditions that do not cause decomposition or loss of activity of active substances), makes this technique the method of choice for preparation of liposomal formulations of sensitive active substances as peptides, proteins or enzymes. 

Desai TR, et al. Determination of surface free energy of interactive dry powder liposome formulations using capillary penetration technique. Colloids Surf B Biointerfaces 22(2) 107-113, 2001. 

The second discovery that enabled liposomes to be developed into efficacious anticancer drug delivery systems was the development of an efficient loading procedure that resulted in liposomes with high drug concentrations. This in turn permitted a reduction in the amount of phospholipid to be administered concomitantly with a given drug dose. This advanced technique was developed during formulation studies on doxorubicin that eventually led to a liposomal formulation that is now licensed as Caelyx for the treatment of Kaposi s sarcoma and ovarian cancer. 

All together, these findings have encouraged the development of neutral and cationic lipospheres (CLS) as nonviral DNA-mediated gene transfer techniques because CLS enable the extemporaneous production of pharmaceutical formulations. Like emulsions and liposomes, lipospheres (LS) consist of physiologically well-tolerated ingredients that have often already been approved for pharmaceutical use... 

The procedure chosen for the preparation of lipid complexes of AmB was nanoprecipitation. This procedure has been developed in our laboratory for a number of years and can be applied to the formulation of a number of different colloidal systems liposomes, microemulsions, polymeric nanoparticles (nanospheres and nanocapsules), complexes, and pure drug particles (14-16). Briefly, the substances of interest are dissolved in a solvent A and this solution is poured into a nonsolvent B of the substance that is miscible with the solvent A. As the solvent diffuses, the dissolved material is stranded as small particles, typically 100 to 400 nm in diameter. The solvent is usually an alcohol, acetone, or tetrahydrofuran and the nonsolvent A is usually water or aqueous buffer, with or without a hydrophilic surfactant to improve colloid stability after formation. Solvent A can be removed by evaporation under vacuum, which can also be used to concentrate the suspension. The concentration of the substance of interest in the organic solvent and the proportions of the two solvents are the main parameters influencing the final size of the particles. For liposomes, this method is similar to the ethanol injection technique proposed by Batzii and Korn in 1973 (17), which is however limited to 40 mM of lipids in ethanol and 10% of ethanol in final aqueous suspension. 

In addition to the remote or active loading techniques mentioned above, metal complexation reactions have been demonstrated to achieve accumulation of doxorubicin in liposomes [148,149], Furthermore, copper-topotecan complexation has been recently seen to mediate drug accumulation into liposomes and is proposed as a methodology to prepare liposomal camptothecin formulations [72],... 

However, CED might not always be successful as good technical skills are required (if the catheter is not placed properly, the liposomes will escape within the CSF). Also CED is an invasive technique and could cause inflammation and neurotoxicity and it is determined by many formulation characteristics and diffusional properties of the latter in brain tissue [418]. 

Structural modifications and formulation are two additional approaches proposed for overcoming the enzymatic barrier (and references therein). Using recombinant or synthetic techniques, selective modifications to the protein or peptide sequence can be introduced effectively reducing proteolytic susceptibility, but these changes must not have significant impact on the pharmacological properties of the molecule (e.g., reduced potency or altered selectivity). Moreover, modifications to address a specific enzymatic action will not eliminate vulnerability to others. The formulation approach essentially involves encapsulation systems to protect the protein or peptide from reactions with enzymes, and selected examples include emulsions, liposomes, or enteric-coated capsules (and references therein). 

The solubilization techniques for injectable formulations are similar to those in oral formulations and include pH adjustment, mixed aqueous/organic cosolvents, organic solvent mixtures, cyclodextrin com-plexation, emulsions, liposomes, polymeric gels, and combinations of techniques. " Molecules that are non-ionizable, lipophilic, and non-polar are challenging to formulate owing to their low water solubility and no effect of pH on solubility. Examples include paclitaxel, docetaxel, cyclosporin A, etoposide, loraze-pam, tacrolimus, testosterone enanthate, and halo-peridol decanoate, and they are all solubilized in non-aqueous solutions composed entirely of organic solvent(s), which are usually but not always diluted prior to administration. 

Majority of nanoliposome manufacture techniques either involve utilisation of potentially toxic solvents (e.g. chloroform, methanol, diethyl ether and acetone) or high shear force procedures. It has been postulated that residues of these toxic solvents may remain in the final liposome or nanoliposome preparation and contribute to potential toxicity and influence the stability of the lipid vesicles (35-38). Although there are methods to decrease the concentration of the residual solvents in liposomes (e.g. gel filtration, dialysis and vacuum), these are practically difficult and time-consuming procedures. In addition, the level of these solvents in the final formulations must be assessed to ensure the clinical suitability of the products (39). Therefore, it would be much preferable to avoid utilisation of these solvents in nanoliposome manufacture, which will also bring down the time and cost of preparation especially at the industrial scales. 

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