LipidNanoparticles and PEG: Time Frame of ImmuneCheckpoint Blockade Can Be Controlled by Adjusting the Rate of CellularUptake of Nanoparticles

The engineerability of lipid nanoparticles (LNPs) and their ability to deliver nucleic acids make LNPs attractive tools for cancer immunotherapy. LNP-based gene delivery can be employed for various approaches in cancer immunotherapy, including encoding tumor-associated antigens and silencing of negative immune checkpoint proteins. For example, LNPs carrying small interfering RNAs can offer several advantages, including sustained and durable inhibition of an immune checkpoint protein. Due to their tunable design, modifying the lipid composition of LNPs can regulate the rate of their uptake by immune cells and the rate of gene silencing. Controlling the kinetics of LNP uptake provides additional flexibility and strategies to generate appropriate immunomodulation in the tumor microenvironment. Here, we evaluated the effects of polyethylene glycol (PEG) content ranging from 0.5 to 6 mol % on the cellular uptake of LNPs by immune cells and gene silencing of PD-L1 after intratumoral administration. We evaluated the cellular uptake and PD-L1 blockade in vitro in cell studies and in vivo using the YUMM1.7 melanoma tumor model. Cell studies showed that the rate of cell uptake was inversely correlated to an increasing mol % of PEG in a linear relationship. In the in vivo studies, 0.5% PEG LNP initiated an immediate effect in the tumor with a significant decrease in the PD-L1 expression of immune cells observed within 24 h. In comparison, the gene silencing effect of 6% PEG LNP was delayed, with a significant decrease of PD-L1 expression in immune cell subsets being observed 72 h after administration. Notably, performance of the 6% PEG LNP at 72 h was comparable to that of the 0.5% PEG LNP at 24 h. Overall, this study suggests that PEG modifications and intratumoral administration of LNPs can be a promising strategy for an effective antitumor immune response.