Quantitative Evaluation of Bioorthogonal Chemistries for Surface Functionalization of Nanoparticles

Abstract

We present here a highly efficient and chemoselective liposome functionalization method based on oxime bond formation between a hydroxylamine and an aldehyde-modified lipid component. We have conducted a systematic and quantitative comparison of this new approach with other state-of-the-art conjugation reactions in the field. Targeted liposomes that recognize overexpressed receptors or antigens on diseased cells have great potential in therapeutic and diagnostic applications. However, chemical modifications of nanoparticle surfaces by postfunctionalization approaches are less effective than in solution and often not high-yielding. In addition, the conjugation efficiency is often challenging to characterize and therefore not addressed in many reports. We present here an investigation of PEGylated liposomes functionalized with a neuroendocrine tumor targeting peptide (TATE), synthesized with a variety of functionalities that have been used for surface conjugation of nanoparticles. The reaction kinetics and overall yield were quantified by HPLC. Reactions were conducted in solution as well as by postfunctionalization of liposomes in order to study the effects of steric hindrance and possible affinity between the peptide and the liposome surface. These studies demonstrate the importance of choosing the correct chemistry in order to obtain a quantitative surface functionalization of liposomes.

Topology of Surface Ligands on Liposomes: Characterization Based on the Terms, Incorporation Ratio, Surface Anchor Density, and Reaction Yield

“…The values for incorporation ratios in parenthesis were calculated by us based on following equation: maleimide lipid molar ratio (of the total lipids in the liposomes)×0.55 (ratio facing outward from the membrane) (Eq. 1 in Chart 1) with the assumption of a 100% reaction yield, which means that all of the anchors contained an attached ligand [33][34][35][36][37][38][39][40][41][42][43][44][45][46][47] . mAb: monoclonal antibody, r.t.: room temperature, n.d.; no data.…”

“…33) Most literature reports do not provide these two values or provide sufficiently accurate values to permit the surface topologies to be evaluated. Instead, bioactivities are evaluated with reference to differences in the composition of liposomal lipids [33][34][35][36][37][38][39][40][41][42][43][44][45][46][47] (Table 1). The affinity of a liposome toward binding and its pharmacokinetics are related to its surface topology, 29,30,[48][49][50] which is defined as the ratio of the density of the surface ligand and PEG that are located on the liposomal surface.…”

Liposomes derivatized with multimeric copies of KCCYSL peptide as targeting agents for HER-2-overexpressing tumor cells

“…27 Peptide sequences can be introduced directly during liposome formulation using a peptide amphiphile (PA) or by grafting the liposomal surface after the preparation with a method well-known in literature as the postliposomal modification method. 27 This last method is particularly appealing for functionalization of liposomes with peptides in their dimeric or multimeric form.…”

“…27 Peptide sequences can be introduced directly during liposome formulation using a peptide amphiphile (PA) or by grafting the liposomal surface after the preparation with a method well-known in literature as the postliposomal modification method. 27 This last method is particularly appealing for functionalization of liposomes with peptides in their dimeric or multimeric form. 22 The anchorage of multimeric copies of the peptide on the liposomes can be achieved using activated functional groups available on the peripheral side of the liposome for covalent or noncovalent binding of the peptide.…”

Receptor binding peptides for target-selective delivery of nanoparticles encapsulated drugs

“…Functional groups commonly used are: a) amine for the amine-N-Hydroxysuccinimide coupling method; b) maleimide for Michael addition; c) azide for Cu(I)-catalyzed Huisgen cycloaddition (CuAAC); and d) biotin for non-covalent interaction with avidin or triphosphines for Staudinger ligation, and recently, hydroxylamine for oxime bond. 46 A considerable number of molecular targets for peptides are either exclusively expressed or overexpressed on both cancer vasculature and cancer cells. They can be classified into three wide categories: integrins; growth factor receptors (GFRs); and G-protein coupled receptors (GPCRs).…”