PEG-polyaminoacid based micelles for controlled release of doxorubicin: Rational design, safety and efficacy study.

A library of amphiphilic monomethoxypolyethylene glycol (mPEG) terminating polyaminoacid co-polymers able to self-assemble into colloidal systems was screened for the delivery and controlled release of doxorubicin (Doxo). mPEG-Glu/Leu random co-polymers were generated by Ring Opening Polymerization from 5 kDa mPEG-NH2 macroinitiator using 16:0:1, 8:8:1, 6:10:1, 4:12:1 γ-benzyl glutamic acid carboxy anhydride monomer/leucine N-carboxy anhydride monomer/PEG molar ratios. Glutamic acid was selected for chemical conjugation of Doxo, while leucine units were introduced in the composition of the polyaminoacid block as spacer between adjacent glutamic repeating units to minimize the steric hindrance that could impede the Doxo conjugation and to promote the polymer self-assembly by virtue of the aminoacid hydrophobicity. The benzyl ester protecting the γ-carboxyl group of glutamic acid was quantitatively displaced with hydrazine to yield mPEG5kDa-b-(hydGlum-r-Leun). Doxo was conjugated to the diblock co-polymers through pH-sensitive hydrazone bond. The Doxo derivatized co-polymers obtained with a 16:0:1, 8:8:1, 6:10:1 Glu/Leu/PEG ratios self-assembled into 30-40 nm spherical nanoparticles with neutral zeta-potential and CMC in the range of 4-7 µM. At pH 5.5, mimicking endosome environment, the carriers containing leucine showed a faster Doxo release than at pH 7.4, mimicking the blood conditions. Doxo-loaded colloidal formulations showed a dose dependent cytotoxicity on two cancer cell lines, CT26 murine colorectal carcinoma and 4T1 murine mammary carcinoma with IC50 slightly higher than those of free Doxo. The carrier assembled with the polymer containing 6:10:1 hydGlu/Leu/PEG molar ratio {mPEG5kDa-b-[(Doxo-hydGlu)6-r-Leu10]} was selected for subsequent in vitro and in vivo investigations. Confocal imaging on CT26 cell line showed that intracellular fate of the carrier involves a lysosomal trafficking pathway. The intratumor or intravenous injection to CT26 and 4T1 subcutaneous tumor bearing mice yielded higher antitumor activity compared to free Doxo. Furthermore, mPEG5kDa-b-[(Doxo-hydGlu)6-r-Leu10] displayed a better safety profile when compared to commercially available Caelyx®.

Thermosensitive "Smart" Surfaces for Biorecognition Based Cell Adhesion and Controlled Detachment.

The biorecognition-based control of attachment/detachment of MCF-7 cancer cells from polymer-coated surfaces is demonstrated. A glass surface is coated with a thermoresponsive statistical copolymer of poly(N-isopropylacrylamide-co-acrylamide) [p(NIPAm-co-Am)], which is end-capped with the Gly-Arg-Gly-Asp-Ser (GRGDS) peptide, and the hydrophilic polymer poly(ethylene glycol) (PEG). Below the lower critical solution temperature (LCST) of p(NIPAm-co-Am) (38 °C), the copolymers are in the extended conformation, allowing for accessibility of the GRGDS peptides to membrane-associated integrins thus enabling cell attachment. Above the LCST, the p(NIPAm-co-Am) polymers collapse into globular conformations, resulting in the shielding of the GRGDS peptides into the PEG brush with consequent inaccessibility to cell-surface integrins, causing cell detachment. The surface coating is carried out by a multi-step procedure that included: glass surface amination with 3-aminopropyltriethoxysilane; reaction of mPEG5kDa -N-hydroxysuccinimide (NHS) and p(NIPam-co-Am)15.1kDa -bis-NHS with the surface aminopropyl groups and conjugation of GRGDS to the carboxylic acid termini of p(NIPam-co-Am)15.1kDa -COOH. A range of spectrophotometric, surface, and microscopy assays confirmed the identity of the polymer-coated substrates. Competition studies prove that MCF-7 cancer cells are attached via peptide recognition at the coated surfaces according to the mPEG5kDa /p(NIPam-co-Am)15.1kDa -GRGDS molar ratio. These data suggest the system can be exploited to modulate cell integrin/GRGDS binding for controlled cell capture and release.