Modulation of Doxorubicin-Induced ROS Accumulation in Cardiomyocytes Using Ibuprofen-Conjugated Synthetic Lipids as Carriers.

Abstract

Conjugation of an NSAID such as ibuprofen to the head group of oxanorbornane-based lipids and the use of their aggregates as carriers for doxorubicin (Dox) are discussed here. These conjugates were characterized by various spectroscopic techniques, including 2D-NMR, and insights into their assembly were gathered through PXRD, AFM, SEM, DLS, and qNano techniques. Free lipids as well as their formulations (lipid:cholesterol:Dox in a 3:1.5:2 molar ratio) showed a high tendency to form solid lipid particles, which was verified by TEM analysis. The presence of the ibuprofen unit led to an increase in interlipid spacing and a characteristic change in their packing. Active loading through a pH gradient allowed us to achieve high drug entrapment and a controlled release profile. The formulation AT3.3, prepared by this method, showed a Dox entrapment of ∼90%, with a controlled release of ∼18% by the end of 24 h; only ∼66% of the entrapped Dox was released by the end of 5 days. Cytotoxicity studies in NIH3T3 cells and hemolytic assay results showed that these lipids and their formulations have a good safety profile. Results from flow cytometry experiments in A549 cells revealed that the formulation AT3.3 induces effects similar to free Dox, with cell cycle arrest predominantly at the S phase and G₂/M phase. At the same time, the response from the blank formulation was comparable to that of the control. Confocal microscopy studies in NIH3T3 and A549 cells showed that free Dox gets localized mainly in the nucleus, while the use of the carrier (AT3.3) causes significant localization of the drug on the cytoplasmic side as well. ROS induction due to free Dox and its formulation (AT3.3) in cardiomyocytes and A549 cells was also compared, and the results showed a protective effect in cardiomyocytes when using this formulation.