Led by Prof Efstathios Karathanasis, a group at Case Western Reserve University has developed an improved and tunable technology for the delivery of gene silencing signals (siRNA for example) to achieve PD-L1 gene silencing. The vehicle lipid nanoparticles (LNPs) have modified PEG content that adjusts their cellular uptake.
In one series of experiments to show the difference in uptake based on PEG content mouse dendritic cell line DC2.4 cells were stably-transfected with GFP and adhered to plates. These were then exposed to different formulations of LNPs loaded with GFP-specific siRNA cargo. After a specified period of exposure to LNPs the cells were stained with Hoechst for the nucleus and DRAQ9™ (1:500 i.e. 2 µM) to determine the GFP signal, inversely proportional to the degree of silencing delivered by the LNPs.
The three fluorescent components used - Hoechst 33342, GFP and DRAQ9™ are ideally suited to a three-colour experiment of this nature allowing trivial spectral separation and cell compartment segmentation.
Reference:
Lipid Nanoparticles and PEG: Time Frame of Immune Checkpoint Blockade Can Be Controlled by Adjusting the Rate of Cellular Uptake of Nanoparticles.
Andrew S. Choi, Taylor J. Moon, Anubhuti Bhalotia, Aarthi Rajan, Laolu Ogunnaike, Diarmuid W. Hutchinson, Inga Hwang, Aaditya Gokhale, Justin N. Kim, Timothy Ma, and Efstathios Karathanasis.
Molecular Pharmaceutics (2025) Article ASAP
