Oxidized graphene nanoparticles as a delivery system for the pro‐apoptotic sphingolipid C₆ ceramide

Abstract

Sphingolipids such as ceramide have attracted much attention as possible anticancer agents due to their potent proapoptotic effects. However, due to their extreme hydrophobicity, there is currently no clinically approved delivery method for in vivo use as a therapeutic agent. To this end, we have developed a novel method for loading the short-chain C 6 ceramide onto oxidized graphene nanoribbons (O-GNRs) and graphene nanoplatelets (GNPs). Mass spectrometry revealed loading efficiencies of 57% and 51.5% for C 6 ceramide onto O-GNRs and GNPs, respectively. The PrestoBlue viability assay revealed that 100 μg/mL of C 6 ceramide-loaded O-GNRs and C 6 ceramide-loaded GNPs reduced HeLa cell viability by approximately 93% and approximately 76%, respectively, compared to untreated HeLa cells, while equal concentrations of these nanoparticles without C 6 ceramide did not significantly reduce HeLa cell viability. We confirmed that this cytotoxicity was apoptotic in nature via capase-3 activity and Hoechst staining. Using live-cell confocal imaging with the fluorescent NBD-ceramide loaded on O-GNRs, we observed robust uptake into HeLa cells within 30 min while NBD-ceramide on its own was uptaken much more rapidly. Transmission electron microscopy confirmed that C 6 ceramide-loaded O-GNRs were actually entering cells. Taken together, these data show that O-GNRs are a promising delivery agent for ceramide. To our knowledge, this study is the first to use such a loading method.

Delivery of long chain C16 and C24 ceramide in HeLa cells using oxidized graphene nanoribbons

“…To verify that O‐GNRs were being synthesized via unzipping carbon nanotubes, TEM characterization was performed using a previously described method (Suhrland et al, ).…”

“…In a previous study, we described a method for loading the short chain C 6 ceramide onto graphene oxide nanoparticles (Suhrland, Truman, Obeid, & Sitharaman, ). We found that oxidized graphene nanoribbons (O‐GNRs) derived from carbon nanotubes were able to load more ceramide and induce higher levels of cytotoxicity and apoptosis than graphene oxide nanoparticles derived via Hummer's method.…”

Bioactive Lipids and Their Derivatives in Biomedical Applications

“…Graphene nanosheets and oxidized graphene nanoribbons have been studied for the delivery of C6 ceramide. Mass spectrometry showed that the loading efficiencies of C6 ceramide onto the graphene nanosheets and oxidized graphene nanoribbons were 51.5% and 57%, respectively, and these formulations were found to reduce the viability of HeLa cells through caspase-3 dependent apoptosis ( Suhrland et al ., 2019 ). Additionally, modified C6 ceramide-loaded graphene nanosheets showed promising anti-cancer effects in a hepatocellular carcinoma model.…”

Bioactive sphingolipids: Advancements and contributions from the laboratory of Dr. Lina M. Obeid

“…To advance ceramide-based therapeutics, her group collaborated with that of Dr. Sitharaman to promote graphene-based nanoparticles of ceramide [ 164 , 165 ].…”