Assessment of stability, toxicity and immunogenicity of new polymeric nanoreactors for use in enzyme replacement therapy of MNGIE

Abstract

The lack of a crucial metabolic enzyme can lead to accumulating substrate concentrations in the bloodstream and severe human enzyme deficiency diseases. Mitochondrial Neurogastrointestinal Encephalomyopathy (MNGIE) is such a fatal genetic disorder, caused by a thymidine phosphorylase deficiency. Enzyme replacement therapy is a strategy where the deficient enzyme is administered intravenously in order to decrease the toxic substrate concentrations. Such a therapy is however not very efficient due to the fast elimination of the native enzyme from the circulation. In this study we evaluate the potential of using polymeric enzyme-loaded nanoparticles to improve the delivery of therapeutic enzymes. We constructed new 200-nanometer PMOXA-PDMS-PMOXA polymeric nanoparticles that encapsulate the enzyme thymidine phosphorylase. These particles are permeabilised for substrates and products by the reconstitution of the nucleoside-specific porin Tsx in their polymeric wall. We show that the obtained 'nanoreactors' are enzymatically active and stable in blood serum at 37 degrees C. Moreover, they do not provoke cytotoxicity when incubated with hepatocytes for 4 days, nor do they induce a macrophage-mediated inflammatory response ex vivo and in vivo. All data highlight the potential of such nanoreactors for their application in enzyme replacement therapy of MNGIE.

Stimuli-Triggered Activity of Nanoreactors by Biomimetic Engineering Polymer Membranes

“…13 A biomimetic approach was employed to design nanoreactors with tandem operation of enzyme-mediated reactions that acted as artificial organelles inside cells 14 or as simple models of cell compartments. 8 An essential molecular parameter for an in situ reaction inside the cavity of a nanoreactor is membrane permeability to allow transport of the substrates required for the reaction, and the 4 products generated by the reaction.…”

Functionalization of Block Copolymer Vesicle Surfaces

“…Polymersomes that have assembled from amphiphilic block copolymers have proven to be useful tools as drug delivery systems [8,[20][21][22], nanoreactors and sensors [13,17,[23][24][25][26][27][28]. The attachment of targeting ligands or enzymes to the polymersomes, as well as the immobilization of polymer vesicles on surfaces, is of crucial importance in most of the previously mentioned applications.…”

Hematopoietic gene therapy restores thymidine phosphorylase activity in a cell culture and a murine model of MNGIE

“…9,10 Therapeutic approaches have so far focused on reducing the systemic overload of these nucleosides. 7,[11][12][13][14] In recent years, several MNGIE patients have been treated with allogeneic hematopoietic stem cell transplantation (HSCT). 4,12,15,16 After successful HSCT, donorderived nucleated blood cells and platelets provided enough TP activity to reduce the dThd and dUrd concentrations to undetectable or nearly undetectable levels, and subsequent slowing of disease progression or mild clinical improvement.…”