Zhadan A.A.¹, Kutsenko O.K.¹, Yudintsev A.V.¹, Gorbenko G.P.¹, Deligeorgiev T.², Vasilev A.², Kaloianova S.², Lesev N.².

¹V.N. Karazin Kharkiv National University, 4 Svobody Sq., Kharkiv, 61077, Ukraine

E-mail: malinka9008@mail.ru

²Department of Applied Organic Chemistry, Faculty of Chemistry, University of Sofia, Bulgaria

COMBINED LIPOSOMAL FORMS OF EUROPIUM COMPLEXES AND DOXORUBICIN

During the past decades the growing research efforts are focused on solving the problem of supplying of toxic, poisonous drugs directly to the localization of disease. One of the promising types of drug nanocarriers is represented by liposomes which offer a lot of advantages over the other carriers [1]. These advantages include: resemblance to natural cellular membranes in their composition; biological compatibility; low toxicity; ability for complete decomposition in an organism - liposomal membrane can be easily merged with cellular membrane, thereby ensuring the intracellular delivery of their content; capacity to carry both hydrophilic and lipophilic drugs and protect them from chemical degradation and transformation; increased therapeutic index of the drug; flexibility in coupling with targeting and imaging ligands; improved pharmacokinetic and pharmacodynamic profiles compared to free drugs; gradual uptake of the drug from liposomes, which increases the time of drug activity; reduced side effects, etc [1, 2].

Of great significance in this regard is the elaboration of liposomal forms of new classes of potential antineoplastic drugs with high cytotoxic activity. One class of such compounds is represented by europium coordination complexes [3]. An important parameter characterizing the efficiency of drug loading by the lipid phase of liposomes is the partition coefficient in the system water/lipid [4].

The present work was directed towards evaluating the efficiency of incorporation of two newly synthesized europium complexes (referred to here as V9 and V12) into phosphatidylcholine (PC) bilayer and answering the question of whether these compounds can be included into liposomes together with the known anticancer drug doxorubicin (Dox) .

Multilamellar liposomes were prepared from egg PC by hydration technique: organic solvent was evaporated from ethanol solution of phosphatidylcholine under vacuum to obtain thin lipid film that was further hydrated by 5mM Na- phosphate buffer (pH 7.4). After incubation with V9, V12 and Dox drug-liposome mixtures were put in dialysis bags for separation of free and lipid-bound drug molecules. Further measurements necessary for calculation of partition coefficient were performed with spectrophotometer SF – 46 at the wavelength 322 nm.

A B

C

Fig.1. Chemical structure of the drugs under study (A - V9, B - V12,C – Dox).

Partition coefficient is defined as:

,

where and – molar concentrations of lipid and water, is usually taken as 55.3 M; , are the concentrations of lipid-bound and free drug, respectively:

,, ,

here ε - molar extinction coefficient of the drug; - optical density of the external solution for the liposome-containing cell containing under equilibrium conditions; - optical density of the external solution of the control cell without liposomes; Vout - external volume of buffer; Vin – cell volume filled with the mixture of liposomes and drug. Presented in Table 1 are values derived for europium complexes V9 and V12 in the absence and presence of doxorubicin.

Table 1

Quantitative characteristics for partitioning of europium complexes

into lipid phase

System	Partition coefficient
V9	(8.99±3)×10^3
V9 + Dox	(3.79±1)×10^4
V12	(1.33±0.3)×10^4

The high recovered values of partition coefficient are indicative of efficient incorporation of the lanthanides into PC bilayer. The efficiency of this process depends on chemical structure of the drug. Our findings suggest that the presence of doxorubicin increases the efficiency of partitioning of europium complexes V9, V12 into lipid bilayer. This creates prerequisites for design of the combined liposomal formulations including europium complexes and known water-soluble antineoplastic agent doxorubicin.

References:

1. Mignet N. et al. // Eur Biophys J. 2006.V. 35. P. 155-161.

2. Torchilin V.P. // Nature reviews / Drug Discovery. 2005. V. 4. P. 145–160.

3. Momekov G. et al. // Medicinal Chemistry. 2006. V. 2. P. 439-445.

4. Seydel J.K., Wiese M. // Drug-membrane interactions, Wiley-VCH, 2002, 337 p.