Interaction of spin-labeled HPMA-based nanoparticles with human blood plasma proteins - the introduction of protein-corona-free polymer nanomedicine.

In this paper, we revised the current understanding of the protein corona that is created on the surface of nanoparticles in blood plasma after an intravenous injection. We have focused on nanoparticles that have a proven therapeutic outcome. These nanoparticles are based on two types of biocompatible amphiphilic copolymers based on N-(2-hydroxypropyl)methacrylamide (HPMA): a block copolymer, poly(ε-caprolactone) (PCL)-b-poly(HPMA), and a statistical HPMA copolymer bearing cholesterol moieties, which have been tested both in vitro and in vivo. We studied the interaction of nanoparticles with blood plasma and selected blood plasma proteins by electron paramagnetic resonance (EPR), isothermal titration calorimetry, dynamic light scattering, and cryo-transmission electron microscopy. The copolymers were labeled with TEMPO radicals at the end of hydrophobic PCL or along the hydrophilic HPMA chains to monitor changes in polymer chain dynamics caused by protein adsorption. By EPR and other methods, we were able to probe specific interactions between nanoparticles and blood proteins, specifically low- and high-density lipoproteins, immunoglobulin G, human serum albumin (HSA), and human plasma. It was found that individual proteins and plasma have very low binding affinity to nanoparticles. We observed no hard corona around HPMA-based nanoparticles; with the exception of HSA the proteins showed no detectable binding to the nanoparticles. Our study confirms that a classical "hard corona-soft corona" paradigm is not valid for all types of nanoparticles and each system has a unique protein corona that is determined by the nature of the NP material.

Acid-labile pHPMA modification of four-arm oligoaminoamide pDNA polyplexes balances shielding and gene transfer activity in vitro and in vivo.

We report novel pH-reversibly surface-shielded polyplexes with enhanced gene transfer activity upon systemic administration. A four-arm-structured sequence-defined cationic oligomer KK[HK[(H-Sph-K)3HC]2]2 was designed and synthesized on solid-phase, containing additional lysine residues not only for improved pDNA polyplex stability, but also providing attachment points for subsequent polyplex functionalization with amine-reactive shielding polymers. Herein, the surface of polyplexes was shielded with hydrophilic polymers, monovalent PEG or monovalent and multivalent pHPMA, optionally attached to the polyplex via the acid-labile linker AzMMMan. Overall, surface modification with PEG or pHPMA resulted in a decrease in the zeta potential of polyplexes, consistent with the degree of surface shielding. At pH 6.0, only polyplexes modified via the acid-labile linkage showed an increase in zeta potential, consistent with a "deshielding" in acidic environment, expected as beneficial for endosomal escape. Shielding was more efficient for multivalent pHPMA (20kDa, 30kDa) as compared to monovalent pHPMA (10kDa, 20kDa, 30kDa) or PEG (5kDa). In vitro transfection studies revealed higher gene expression by the polyplexes with the acid-labile shield as compared to their irreversibly shielded counterparts. Intravenous administration of AzMMMan-pHPMA modified polyplexes in an in vivo tumor mouse model mediated enhanced gene expression in the subcutaneous tumor and reduced undesirable expression in the liver.

Tailoring the physicochemical properties of core-crosslinked polymeric micelles for pharmaceutical applications.

To optimally exploit the potential of (tumor-) targeted nanomedicines, platform technologies are needed in which physicochemical and pharmaceutical properties can be tailored according to specific medical needs and applications. We here systematically customized the properties of core-crosslinked polymeric micelles (CCPM). The micelles were based on mPEG-b-pHPMAmLacn (i.e. methoxy poly(ethylene glycol)-b-poly[N-(2-hydroxypropyl) methacrylamide-lactate]), similar to the block copolymer composition employed in CriPec® docetaxel, which is currently in phase I clinical trials. The CCPM platform was tailored with regard to size (30 to 100nm), nanocarrier degradation (1month to 1year) and drug release kinetics (10 to 90% in 1week). This was achieved by modulating the molecular weight of the block copolymer, the type and density of the crosslinking agent, and the hydrolytic sensitivity of the drug linkage, respectively. The high flexibility of CCPM facilitates the development of nanomedicinal products for specific therapeutic applications.

Nanomedicines for inflammatory arthritis: head-to-head comparison of glucocorticoid-containing polymers, micelles, and liposomes.

As an emerging research direction, nanomedicine has been increasingly utilized to treat inflammatory diseases. In this head-to-head comparison study, four established nanomedicine formulations of dexamethasone, including liposomes (L-Dex), core-cross-linked micelles (M-Dex), slow releasing polymeric prodrugs (P-Dex-slow), and fast releasing polymeric prodrugs (P-Dex-fast), were evaluated in an adjuvant-induced arthritis rat model with an equivalent dose treatment design. It was found that after a single i.v. injection, the formulations with the slower drug release kinetics (i.e., M-Dex and P-Dex-slow) maintained longer duration of therapeutic activity than those with relatively faster drug release kinetics, resulting in better joint protection. This finding will be instructional in the future development and optimization of nanomedicines for the clinical management of rheumatoid arthritis. The outcome of this study also illustrates the value of such head-to-head comparison studies in translational nanomedicine research.

HPMA copolymer conjugates of DOX and mitomycin C for combination therapy: physicochemical characterization, cytotoxic effects, combination index analysis, and anti-tumor efficacy.

The synthesis, characterization and results of evaluation of the biological behavior of HPMA copolymer conjugates bearing anti-cancer drugs doxorubicin and mitomycin C are described. Two HPMA copolymer carrier types were synthesized: the linear copolymer and the biodegradable high-molecular-weight diblock copolymer containing a degradable disulfide bond. The polymer-drug conjugates incubated in buffers modeling the intracellular environment released the drugs more rapidly than those incubated in bloodstream conditions. Significant in vitro and in vivo antitumor synergistic activity of the conjugates in the treatment of EL-4 T-cell demonstrates their high potential for solid tumor treatment.