The prodrug is subject to enzymatic hydrolysis by phospholipase A2 (PLA2) and non-specific esterases yielding the parent dihydrofuran allocolchicinoid (aC). In circulation, liposomes of pure ePC bind considerable amount of proteins gaining the so-called protein corona and are rapidly removed by mononuclear phagocyte system (also known as reticular endothelial system, RES) [18]. the /-tubulin heterodimers and microtubule elongation, which in turn results in the G2/M cell cycle arrest and ultimately apoptosis [1,2]. Anti-inflammatory effects of colchicine are mainly mediated by inhibition of neutrophil chemotaxis and cytokine release [3]. Low-dose colchicine has been approved to treat familial Mediterranean fever and acute gout flares. It is widely studied as an anti-inflammatory agent in various cardiovascular diseases [4,5,6]. Colchicine is considered as a promising anti-inflammatory drug to treat COVID-19 [7,8,9]: 36 relevant trials are registered at https://clinicaltrials.gov/as of 11 December 2021. Application of colchicine in cancer therapy is limited by its high general toxicity [10]. Heterocyclic allocolchicinoids exhibited lower acute toxicity yet comparable cytotoxicity in vitro and anticancer activity in vivo [11,12,13,14]. Incorporation of drugs in nanosized liposomal carriers allows to decrease systemic toxicity, first of all, due to the decreased volume of distribution [15]. Lipophilic derivatization can be adopted to utilize the bilayer compartment for the drug loading. The approach offers several benefits. (1) High loading capacity can be achieved matching that of DOXIL, and alike, where drug salt crystals are formed within the liposomes. (2) From the technology Tuberculosis inhibitor 1 point of view, it requires no separation of the unencapsulated drug. (3) It offers the opportunity to utilize surface-active enzymes, e.g., phospholipase A2, to release the active drug. Since phospholipase A2 has been reported overexpressed in inflammation sites and in some tumors, this adds to the selectivity of such drug carrier. Earlier, a pair of allocolchicinoids, the carboxymethyl and dihydrofuran congeners, with enhanced cytotoxicity have been synthesized and used to generate phospholipase A2 (PLA2)-responsive phosphatidylcholine prodrugs [16]. The prodrugs were proven to be easily incorporated in egg Tuberculosis inhibitor 1 phosphatidylcholine (ePC) liposomes. In the Igf2r amount of 5 mol. % of the prodrug, no distortion of the bilayer structure was observed according to the surface pressureCarea isotherms [16]. The dihydrofuran allocolchicinoid and its phospholipid prodrug (aC and aC-PC, respectively; Figure 1) have been slightly more cytotoxic (~5- and 2C5-fold, respectively) and produced more stable liposomes compared to the carboxymethyl counterpart as assayed on PANC-1, Colo-357, BxPC-3, and HaCaT cell lines [16]. Assuming there are hydrophobic regions covering up to 20% of the bilayer surface area [17] and taking into account optimized geometry of the allocolchicinoid [16], here we tested whether as Tuberculosis inhibitor 1 much as 25 mol. % of aC-PC could be incorporated in the bilayer of the ePC liposomes. Open in a separate window Figure 1 Schematic representation of a liposome with allocolchicinoid phospholipid prodrug (aC-PC) in the bilayer and matrix lipids used in the work, egg yolk-derived phosphatidylcholine (ePC, a representative structure is shown) and 1-palmitoyl-2-oleoylphosphatidylglycerol (POPG). The prodrug is subject to enzymatic hydrolysis by phospholipase A2 (PLA2) and non-specific esterases yielding the parent dihydrofuran allocolchicinoid (aC). In circulation, liposomes of pure ePC bind considerable amount of proteins gaining the so-called protein corona and are rapidly removed by mononuclear phagocyte system (also known as reticular endothelial system, RES) [18]. The presence of phosphatidylglycerol (PG) and cholesterol (Chol) has been shown to enhance liposome stability toward aggregation in the presence of plasma proteins [19]. Moreover, incorporation of Tuberculosis inhibitor 1 33% Chol in the bilayer (to create the liquid ordered lipid phase) and/or anionic lipids (e.g., GM1) is known to improve the blood-to-RES tissue ratio in mice by 1C2 orders of magnitude [20]. Thus, variation of 1-palmitoyl-2-oleoylphosphatidylglycerol (POPG), a commercially available liquid-phase anionic lipid, and cholesterol content in the ePC bilayer can be used.