Development of long-circulating nanoparticles for ATP delivery

The antitumoral effect of ATP requires its interaction with membrane receptors in target cells, but due to its short lifetime it is necessary to be kept in high concentrations in the extracellular space. We propose the use of... [ view full abstract ]

The antitumoral effect of ATP requires its interaction with membrane receptors in target cells, but due to its short lifetime it is necessary to be kept in high concentrations in the extracellular space. We propose the use of albumin nanoparticles (ANs) coated with erythrocytes membranes (EMs) to allow extending the time of drug circulation. Our aim was to synthesize and characterize long-circulating ANs, coated with EMs and loaded with ATP. ANs were synthesized by the desolvation method, determining the optimal values of pH (8-10), albumin concentration (10, 20 or 40 mg/ml) and ethanol volume as precipitating agent (2.33 ± 0.04 ml). EMs were derived from erythrocyte lysates and used to coat ANs using an extruder. Size was determined by transmission electron microscopy (ANs = 91.9 ± 4.3 nm and AN-EMs = 98.3 ± 5.1 nm), while hydrodynamic size (AN = 180.5 ± 6.8 nm and AN-EMs = 197.8 ± 3.2 nm) and Ζ potential (AN = + 17.8 ± 3.5 mV and AN-EMs = -13.7 ± 2.9 mV) was determined by dynamic light scattering. The coating of the ANs with the EMs was verified by TEM and confocal microscopy, via the colocalization of hydrophobic (DiD for AN-EMs) and hydrophilic fluorophores (Fluorescein for ANs). The cell uptake of nanoparticles was analyzed by confocal microscopy using HeLa cell cultures treated with nanoparticles stained with the same fluorophores. A smaller number of internalized AN-EMs compared to non-coated ANs was observed. Our current results show that it is possible to obtain nanoparticles from highly biocompatible, biodegradable materials, and that their coating with EMs allows regulating the internalization process in order to promote longer circulation times. These nanoparticles will be used to assess the release of ATP in in vitro and in vivo cancer models.