Peptide-polymer biotherapeutic synthesis on novel cross-linked beads with "spatially tunable" and "isolated" functional sites.

Solid phase synthesis of polymer biotherapeutics using conventional polymers suffers from many limitations such as low synthetic yield and purity. The conventional polymers prepared by either pre- or post-functionalization strategies have no control over the point of functionalization. Hence we report a novel cross-linked polymer in which the functional groups are spatially tuned to predefined distance with optimal site isolation. This has been achieved by the design and synthesis of a tetra functional PEG, 3,3'-(PEG)bis(1-(4-vinylphenoxy)propan-2-ol) (bis(VPP)PEG). It has been incorporated at cross-linking of 1-12%, into a polystyrene network by free radical suspension polymerization. In this polymer, the distance between hydroxyl functional groups has been spatially tuned in a predefined manner by varying the length of the cross-linker backbone from ethylene glycol to PEG1000 Da and the loading capacity could be varied from 0.1 to 0.9 mmol/g. The polymer has been characterized by SEM, FTIR, and 13C NMR. The polymer exhibits excellent swelling behavior and high chemical stability. The synthetic efficiency of the polymer was demonstrated by the successful synthesis of three structural classes of PEGylated antimicrobial peptide biotherapeutics and the difficult ACP (65-74) fragment. Thus the "spatially defined" and "site isolated" synthesis within the new polymer offers a novel strategy for synthesis of difficult peptide-polymer bioconjugates. The bioassay studies shows that PEGylation of AMPs significantly reduces their hemolytic potential but the retainment of antibacterial property was dependent both on the peptide sequence and the size of PEG used.