Recognition, neutralization, and clearance of target peptides in the bloodstream of living mice by molecularly imprinted polymer nanoparticles: a plastic antibody

Abstract

We report that simple, synthetic organic polymer nanoparticles (NPs) can capture and clear a target peptide toxin in the bloodstream of living mice. The protein-sized polymer nanoparticles, with a binding affinity and selectivity comparable to those of natural antibodies, were prepared by combining a functional monomer optimization strategy with molecular-imprinting nanoparticle synthesis. As a result of binding and removal of melittin by NPs in vivo, the mortality and peripheral toxic symptoms due to melittin were significantly diminished. In vivo imaging of the polymer nanoparticles (or "plastic antibodies") established that the NPs accelerate clearance of the peptide from blood and accumulate in the liver. Coupled with their biocompatibility and nontoxic characteristics, plastic antibodies offer the potential for neutralizing a wide range of biomacromolecules in vivo.

Figure 1

Preparation and characterization of polymer NPs. a. Amino acid sequence of target peptide Melittin. Hydrophobic, positive charged and hydrophilic residues are rendered in green, red and black respectively. b. Monomers used for NP synthesis. c. Schematic of the preparation of MIPNPs. d. Solution phase AFM images of MIPNPs. A height profile of cross-section (sky blue line) is shown in insert.

Figure 2

Neutralization of melittin toxicity by NPs. a. Survival rates of mice over a 24 h period after intravenous injection of 4.5 mg kg⁻¹ melittin (green). 30 mg kg⁻¹ of MIPNPs (red), NIPNPs (gray) was systemically administrated via a tail vain 20 seconds after melittin injection. P values are calculated by the Willcoxon test. b. Macroscopic pathology of peritoneal inflammation of mice injected with melittin (4.0 mg kg⁻¹) followed without (left) or with (right) MIPNPs (30 mg kg⁻¹). c. Body weight change of mice injected with melittin (right two columns; 0 mg kg⁻¹, center three columns 4.0 mg kg⁻¹, right three columns 4.5 mg kg) followed with 0 mg kg⁻¹ (white), 9.3 mg kg⁻¹ (gray) or 30 mg kg⁻¹ (black) MIPNPs (48 hours after melittin injection). * No animal was alive. The data represent the mean ± SEM.

Figure 3

Biodistribution of melittin and NPs. a. Fluorescent images of Cy5-melittin after intravenous injection of Cy5-melittin (1 mg kg⁻¹). 27 mg kg⁻¹ of MIPNPs was injected 20 sec after the injection of melittin (right). b. Fluorescent ex vivo images of Cy5-melittin (0.3 mg kg⁻¹, 10 min after injection) of mice followed with and without 10 mg kg⁻¹ MIPNPs. Li, Sp, SI, K, H and Lu indicate liver, spleen, small intestine, kidney, heart and lung respectively. c. Fluorescent images of Cy5-melittin (70 min after injection) in livers from mice with various doses of Cy5-melittin and MIPNPs. d. Biodistribution of ¹⁴C-labeled NPs in a mouse (n = 5 or 4, 10 mg kg⁻¹). e. Fluorescence histology images of the liver shown in c (Cy5-melittin 0.3 mg kg⁻¹ and 10 mg kg⁻¹ of MIPNPs). Green; Cy5-melittin, red; fluoroscein-MIPNPs. The scale bars; 25 μm.