Liposomal delivery of hydrophobic RAMBAs provides good bioavailability and significant enhancement of retinoic acid signalling in neuroblastoma tumour cells

Abstract

Retinoid treatment is employed during residual disease treatment in neuroblastoma, where the aim is to induce neural differentiation or death in tumour cells. However, although therapeutically effective, retinoids have only modest benefits and suffer from poor pharmacokinetic properties. In vivo, retinoids induce CYP26 enzyme production in the liver, enhancing their own rapid metabolic clearance, while retinoid resistance in tumour cells themselves is considered to be due in part to increased CYP26 production. Retinoic acid metabolism blocking agents (RAMBAs), which inhibit CYP26 enzymes, can improve retinoic acid (RA) pharmacokinetics in pre-clinical neuroblastoma models. Here, we demonstrate that in cultured neuroblastoma tumour cells, RAMBAs enhance RA action as seen by morphological differentiation, AKT signalling and suppression of MYCN protein. Although active as retinoid enhancers, these RAMBAs are highly hydrophobic and their effective delivery in humans will be very challenging. Here, we demonstrate that such RAMBAs can be loaded efficiently into cationic liposomal particles, where the RAMBAs achieve good bioavailability and activity in cultured tumour cells. This demonstrates the efficacy of RAMBAs in enhancing retinoid signalling in neuroblastoma cells and shows for the first time that liposomal delivery of hydrophobic RAMBAs is a viable approach, providing novel opportunities for their delivery and application in humans.

The liposomal delivery of hydrophobic oxidovanadium complexes imparts highly effective cytotoxicity and differentiating capacity in neuroblastoma tumour cells

“…In this study we used cationic liposomes, having previously found that related cationic formulations are effective in drug delivery in culture and in vivo and are poor activators of the complement system 60,61 . Our previous work on RAMBA chemicals that are similarly hydrophobic to the present oxidovanadium complexes, also showed very effective payload delivery by liposomes in under 24 h, and others describe this occurring in under 6 h 48,62,63 . We demonstrated here in this proof-of-principle study that the liposomal delivery approach for oxidovanadium complexes works well under cell culture conditions.…”

“…AL3 was chosen for this liposomal testing since it was the most hydrophobic and demonstrated the greatest cytotoxicity and differentiative potential. AL3 was incorporated into liposomes derived from DOPC, a neutral lipid used to promote stable lamellar structures, cholesterol to increase stability of the liposomes, and DOTMA as a cationic lipid to improve uptake in mammalian cells 48 . Liposomes containing AL3 were cationic with an average charge of 64 ± 3 mV, a mean diameter of 159 ± 16 nm and polydispersity index of 0.33 ± 0.04.…”

In Vitro Transfection of Up-Regulated Genes Identified in Favorable-Outcome Neuroblastoma into Cell Lines

“…Retinoids induce CYP26 enzyme production in the liver, enhancing their own rapid metabolic clearance, while retinoid resistance in tumor cells occurs in part due to increased CYP26 production. RA-metabolism-blocking agents, which inhibit CYP26 enzymes, can improve RA pharmacokinetics in preclinical NB models [22]. The signaling networks through which RA mediates NB differentiation were inhibited upon MYCN overexpression, which revealed opposing regulation of RA and for a number of differentiation-relevant genes, including LMO4, CYP26A1, ASCL1, RET, FZD7, and DKK1 [23].…”

New luciferin-based probe substrates for human CYP26A1

“…Our findings naturally raise the interesting question whether retinyl esters might also be CYP26A1 substrates. Moreover, retinoic acid metabolism blocking agents (RAMBAs) hold promise for improving the clinical benefit of retinoid treatments, for instance against acute promyelocytic leukemia or neuroblastoma tumors; and interestingly, some of these compounds are esters [ 18 , 19 ]. We expect that the new and convenient proluciferin-based activity assays presented in this study will be useful for the identification of new substrates and inhibitors of CYP26A1.…”