FRET-Based Investigation of the Dynamics of Peptide-Folded Single Chain Nanoparticles Inside Cancer Cells

Single chain nanoparticles (SCNPs) prepared using diphenylalanine (_LFF) as crosslinker have recently been proposed as protein-inspired drug delivery carriers. The aim of this work is to learn more about the stability of these peptide-based supramolecular systems in biological environments using Förster resonance energy transfer and fluorescence lifetime imaging microscopy (FRET-FLIM). Two self-folding poly(N,N-dimethyl acrylamide) PDMA with 10 mol.% pendant _LFF, labeled at the omega chain end with the cyanine dyes Cy5 and Cy3, respectively, are formed into SCNPs with sizes below 10 nm. The particles are found to be stable in water and do not aggregate in a cellular environment (MCF-7 breast cancer cells). The SCNPs are uptaken by these cells slightly faster than a control nanoparticle of the same polymers (≈100 nm in cell culture media) and with similar total uptake after 1 h. Temperature-dependent studies demonstrate that while the larger nanoparticles are dependent on endocytosis for uptake, the SCNPs can cross the membrane via diffusion. This study demonstrates the value of FRET-FLIM in understanding the behavior of self-assembled nanoparticles in vitro and provides additional evidence for the alternate cell uptake pathways that SCNPs can access over traditional nanoparticle based drug carriers.