Abstract 3686: Engineering nanoparticles of polymalic acid for controlled delivery of anticancer drugs
Authors: Portilla-Arias José Antonio , Lanz-Landazuri Alberto , Garcia-Alvarez Montserrat , Martinez de Ilarduia Antxon , Patil Rameshwar , Holler Eggehard , Ljubimova Julia , Muñoz-Guerra Sebastian 
Published: August 2015

Abstract
Objective:
In this work we report new esters obtained by partial or total esterification of Poly(β,L-malic acid) of microbial origin (PMLA) with ethanol or 1-butanol, and the nanoparticles made from them, are explored as drug delivery systems for the anticancer drugs temozolomide (TMZ) and doxorubicin (DOX).
Experimental procedures:
Esterification of PMLA with either ethanol or 1-butanol was performed with the polyacid dissolved in the corresponding alcohol and using controlled amounts of DCC for activation of the carboxylic side group. The conversion degree and purity was ascertained by 1H NMR. Two methods were employed for the formation of nanoparticles depending on the esterification degree of the polymer. For 100% modified polymers, the emulsion-solvent evaporation method and the precipitation-dialysis for partially esterified PMLA. Particle morphology was monitored by SEM and their sizes were determined by DLS. Primary glioma cell line U87MG and invasive breast carcinoma cell line MDA-MB468 (ATCC, USA) were exposed to drug loaded and unloaded nanoparticles and cell viability and uptake was measured using the MTT method and fluorescent microscopy respectively.
Results: Ethyl and butyl PMLA esters with esterification degrees of approximately 50% and fully esterified were obtained. Reaction yields were around 50-70% with higher values attained in the esterification with ethanol. Molecular weights were: 33 KDa for coPMLA-Et50H50; 36 KDa for PMLA-Et100; 34 kDa for coPMLA-Bu50H50 and 53kDa for PMLA-Bu100. Nanoparticles diameters oscillated from 100 to 350 nm, depending on the polymer and on the methodology used for particle formation. DOX and TMZ can be encapsulated in these nanoparticles with efficiencies ranging 20 to 40% w/w. Drug-unloaded nanoparticles were not cytotoxic for the tested cell lines, whereas drug loaded nanoparticles were effective showing significant lower EC50 values when compared with free drug. The most remarkable case was observed for MDA-MB468 cell line treated with free TMZ and TMZ-NPs, in which free TMZ was shown to be ineffective at all concentrations while TMZ loaded nanoparticles reached EC50 at concentrations between 1-2×10-4 M. Two reasons can be invoked to explain such differences. A more sustained presence of encapsulated TMZ in the media and/or the nanoparticle internalization by cells that allows TMZ to be released directly in the cytosol. The most efficient polymer nanoparticles were coPMLA-Et50H50 which showed better internalization of DOX by cells than the free drug.
Conclusions: Esterification of microbial poly(malic acid) with ethanol or 1-butanol rendered easily hydrolysable polyesters and copolyesters, suitable for building nanoparticles useful for anticancer drug encapsulation and controlled delivery.
Citation Format: José Antonio Portilla-Arias, Alberto Lanz-Landazuri, Montserrat Garcia-Alvarez, Antxon Martinez de Ilarduia, Rameshwar Patil, Eggehard Holler, Julia Ljubimova, Sebastian Muñoz-Guerra. Engineering nanoparticles of polymalic acid for controlled delivery of anticancer drugs. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 3686. doi:10.1158/1538-7445.AM2015-3686