Fluorophore-tagged Poly-ʟ-lysine Block Copolymers for Biomedical Applications

Abstract

Fluorescent PEGylated poly-ʟ-lysine copolymers were developed through ring-opening polymerization (ROP). To achieve this, ε-Cbz ʟ-Lysine was converted to its corresponding NCA. Subsequently, Poly(ethylene glycol) methyl ether was converted to Poly(ethylene glycol) methyl ether amine, which was then used to initiate the ring-opening polymerization (ROP) of ε-Cbz ʟ-Lysine NCA to yield di-block copolymers. The polymerization conditions such as solvent (only DMF vs DMF-CHCl3), catalyst (with/without IMes-NHC) and [M]/[I] feed ratio (i.e., 25, 50, 75 and 100) were varied to investigate their effect on the PEG polypeptide synthesis. The real-time kinetic studies were performed by FT-IR to understand how the presence of catalyst affects the polymerization rate. Also, α-helical induced enhancement of polymerization was observed. The structure and degree of polymerization (Xn) of these polymers were confirmed by proton NMR spectroscopy. Gel permeation chromatography (GPC) was used to determine the number and weight average molecular weights (Mn and Mw) with DMF as the solvent using PEG standards. The Cbz-protected di-block polymers were deprotected to yield cationic PEG Poly-ʟ-lysine di-block polymers. The coumarin-based fluorescent probes were synthesized by Knoevenagel condensation reaction of 4-(Diethylamino)salicylaldehyde and diethyl malonate. This was then coupled to PEG Poly-ʟ-lysine to yield polymer-probe conjugates. The fluorescent tag on the polymeric architecture provides insight into the polymer's fate, which would otherwise be difficult due to the polymer's non-fluorescent nature. The probe and polymer-probe conjugates show an emission maximum at the wavelengths of 460-480 nm with water and DMSO as a solvent. The dynamic light scattering studies show the size of these nano-assemblies as <160 nm while the zeta potentials were in the range of +19-44 mV. The antimicrobial action of these polymers was tested in E. coli. and the mammalian cell viability was tested in Hela and MCF-7 cancer cell lines. The cellular uptake was tested in MCF-7 cell lines and monitored through confocal microscopy.