Evaluation of PolyMPC-Dox Prodrugs in a Human Ovarian Tumor Model.

A polymer prodrug, composed of doxorubicin (Dox) conjugated covalently to poly(methacryloyloxyethyl phosphorylcholine) (polyMPC), was evaluated for the treatment of human ovarian tumors in animals. PolyMPC-Dox prodrugs were prepared using facile conjugation chemistry to yield conjugates soluble in water and injectable saline, with a Dox loading of ∼19 weight percent. Toxicity evaluation showed that polyMPC was well-tolerated in mice at doses up to 800 mg/kg, confirming the biocompatibility of the polymer carrier at a high concentration. Additionally, the polyMPC-Dox prodrug was well-tolerated in animals at a Dox equivalent dose of 10 mg/kg, greater than twice the maximum tolerated dose of free Dox (∼4 mg/kg) in the same mouse strain. In a human ovarian tumor model (SKOV-3), polyMPC-Dox accumulated in tumors at twice the level of free Dox, with no additional off-target organ uptake, a result of improved pharmacokinetics afforded by the prodrug and passive targeting attributed to an enhanced permeability and retention effect. When administered to human ovarian tumor-bearing mice using a recurring dosing regimen comparable to that used clinically, polyMPC-Dox significantly retarded tumor growth relative to treatment with free Dox. Moreover, animals treated with multiple doses of polyMPC-Dox (eight total doses) exhibited enhanced survival, with a notably reduced incidence of toxicity or adverse events relative to mice treated with free Dox. These in vivo results demonstrate advantages of treating human ovarian tumors with polyMPC-Dox, including reduced systemic toxicity, improved drug accumulation in tumors, and enhanced therapeutic efficacy.

Efficacy of polyMPC-DOX prodrugs in 4T1 tumor-bearing mice.

We report the in vivo efficacy, in tumor-bearing mice, of cancer prodrugs consisting of poly(methacryloyloxyethyl phosphorylcholine) (polyMPC) conjugated to doxorubicin (DOX). Our synthesis of polyMPC-DOX conjugates established prodrugs with tunable drug loading, pH sensitive release kinetics, and a maximum tolerated dose in the range of 30-50 mg/kg (DOX equivalent) in healthy mice. Here we show prolonged circulation of polyMPC-DOX, with a measured in vivo half-life (t1/2) 8 times greater than that of the free drug. We observed reduced drug uptake in healthy tissue, and 2-3 times enhanced drug accumulation in tumors for polyMPC-DOX prodrugs compared to free DOX, using BALB/c mice bearing 4T1 tumors. Prolonged survival and reduced tumor growth were observed in mice receiving the polyMPC-DOX prodrug treatment. Moreover, we evaluated immunogenicity of polyMPC-DOX prodrugs by examining complete blood count (CBC) and characteristic cytokine responses, demonstrating no apparent innate or adaptive immune system response.

Promoting cell adhesion on slippery phosphorylcholine hydrogel surfaces.

The growing interest in regenerative medicine has created a need for superior polymer matrices that suit multiple physical, mechanical, and biological requirements. While the phospholipid bilayer of a cell membrane is considered optimal for interacting with biologics, polymeric materials composed of 2-methacryloyloxyethyl phosphorylcholine (MPC) offer a cell membrane-like synthetic alternative. In this work, thiol-containing phosphorylcholine polymers were synthesized by radical copolymerization of a lipoic acid-functionalized methacrylate with MPC. The canonical cell adhesion oligopeptide (GRGDS) was incorporated into the polymers by copolymerization of a GRGDS-containing methacrylamide prepared by solid phase peptide synthesis. The relative amounts of phosphorylcholine, lipoic acid and oligopeptide were controlled by the monomer feed ratios, and the polymers were characterized by NMR spectroscopy and aqueous gel permeation chromatography (GPC). These multifunctional polymers formed hydrogels rapidly (<10 minutes) by Michael addition when poly(ethylene glycol)diacrylate (PEGDA) was added at pH 9 - an initiator-free gelation performed in a completely aqueous environment. Two cell lines, live mouse skeletal muscle myoblasts (C2C12) and human ovarian cancer (SKOV3) cells, were observed to specifically attach, spread and proliferate only on hydrogels containing the GRGDS peptide sequence, with a notable dependence on peptide concentration. The remarkable hydrophilicity and biocompatibility attributed to polyMPC combined with the facile gelation conditions of these polymers affords a platform of new bio-cooperative materials suitable for cell studies.

Disulfide cross-linked phosphorylcholine micelles for triggered release of camptothecin.

A series of block copolymers based on 2-methacryloyloxyethyl phosphorylcholine (MPC) were synthesized by reversible addition-fragmentation chain transfer (RAFT) polymerization. Incorporation of dihydrolipoic acid (DHLA) into the hydrophobic block led to formation of block copolymer micelles in water. The micelles were between 15 and 30 nm in diameter, as characterized by dynamic light scattering (DLS), with some size control achieved by adjusting the hydrophobic/hydrophilic balance. Cross-linked micelles were prepared by disulfide formation, and observed to be stable in solution for weeks. The micelles proved amenable to disassembly when treated with a reducing agent, such as dithiothreitol (DTT), and represent a potential delivery platform for chemotherapeutic agents. As a proof-of-concept, camptothecin (CPT) was conjugated to the polymer scaffold through a disulfide linkage, and release of the drug from the micelle was monitored by fluorescence spectroscopy. These CPT-loaded prodrug micelles showed a reduction in release rate compared to physically encapsulated CPT. The use of disulfide conjugation facilitated drug release under reducing conditions, with a half-life (t1/2) of 5.5 h in the presence of 3 mM DTT, compared to 28 h in PBS. The toxicity of the micellar prodrugs was evaluated in cell culture against human breast (MCF7) and colorectal (COLO205) cancer cell lines.