Influence of Histidine Incorporation on Buffer Capacity and Gene Transfection Efficiency of HPMA-co-oligolysine Brush Polymers

Abstract

One of the major intracellular barriers to non-viral gene delivery is efficient endosomal escape. The incorporation of histidine residues into polymeric constructs has been found to increase endosomal escape via the proton sponge effect. Statistical and diblock copolymers of N-(2-hydroxypropyl)methacrylamide (HPMA), oligolysine, and oligohistidine were synthesized via reversible-addition fragmentation chain transfer (RAFT) polymerization, and tested for in vitro transfection efficiency, buffering ability, and polyplex uptake mechanism via the use of chemical endocytic inhibitors. Interestingly, histidine-containing statistical and diblock polymers exhibited increased buffer capacity in different endosomal pH ranges. Statistical copolymers transfected better than block copolymers that contained similar amounts of histidine. In addition, only the polymer containing the highest incorporation of oligohistidine residues led to increases in transfection efficiency over the HPMA-oligolysine base polymer. Thus, for these polymer architectures, high histidine incorporation may be required for efficient endosomal escape. Furthermore, uptake studies indicate that non-acidified caveolae-mediated endocytosis may be the primary route of transfection for these copolymers, suggesting that alternative approaches for increasing endosomal escape may be beneficial for enhancing transfection efficiency with these HPMA-oligolysine copolymers.

Heterocyclic amine-modified polyethylenimine as gene carriers for transfection of mammalian cells

“…Histidine 477 has an imidazole group with a pKa of about 6, so it is an effective 478 buffer throughout much of the endosomal pH range [31][32][33][34]. 479 Also histidine could provide a direct fusogenic property on the 480 endosomal membranes at acidic pHs [35,36].…”

Nano‐Sized Sunflower Polycations As Effective Gene Transfer Vehicles

“…[20] The buffering capacities (BCs) of comb and sunflower pDMAEMAs were examined by acid-base titration, and defined as the μmol H + /mg polymer required to decrease the pH value of a polymer solution from 7.4 to 5.0 (Figure 1). [21] The buffering capacity increased with increasing molecular weight of polycation. Intriguingly, sunflower pDMAEMAs showed higher buffering capacity than comb polycations, which may promote endosomal escape of their corresponding polyplexes.…”

Multi-Domain Short Peptide Molecules for in Situ Synthesis and Biofunctionalization of Gold Nanoparticles for Integrin-Targeted Cell Uptake

“…For example, glycine can be replaced with positively charged residues, such as histidine, to serve as carriers for nucleotides in gene delivery applications or as endosome-escaping proton sponges. 25 Immediately after the Au 3+ and MDP solutions were mixed, the container tube was thoroughly vortexed for 10 s. Then, pH of this solution was elevated to ∼10 by 2 M NaOH (aq) , vortexed immediately and vigorously for 10 s, and incubated at room temperature for 1 h by continuous stirring. pH elevation triggered spontaneous oxidation of catechol, which was coupled to the reduction of Au 3+ .…”