Preparation of EcN-conjugated drug-loaded PSMA/PELA micro-particles and their ability to target hypoxic tumor environment

Abstract

Objective To conjugate Escherichia coli Nissle 1917 (EcN) with drug-loaded poly styrene-co-maleic anhydride(PSMA)/poly (DL-lactic acid) and poly ethyleneglycol block copolymer (PELA) polymer micro-particles and investigate the ability of the micro-particles to target the hypoxic tumor environment. Methods PSMA and PELA were mixed at the ratio of 7 :3 or 5 :5 (m/m) to prepare doxorubicin (Dox)-loaded PSMA/PELA micro-particles through solvent evaporation method. The covalent binding was achieved by EDC-NHS catalytic reaction of the carboxyl groups on the surface of EcN with the amino groups on the surface of PSMA/PELA drug-loaded micro-particles. In an in vitro 3D tumor model constructed using HepG2 cells, we investigated the permeation of EcN-conjugated PSMA/PELA drug-loaded micro-particles into the hypoxic tumor area. We further tested the performance of the drug-loaded micro-particles for targeting the hypoxic tumor environment in a mouse model bearing subcutaneous hepatoma H22 cell xenograft. Results We successfully prepared Dox-loaded micro-particles with different PSMA/PELA ratios (DMP7/3 and DMP5/5) using solvent evaporation method, and both of the micro-particles showed a regular spherical shape with an average size of 700 to 800 nm. DMP7/3 and DMP5/5 micro-particles had comparable Dox-loading efficiency [(4.27±0.19)% vs (3.94±0.11)%] and encapsulation efficiency [(85.4± 2.6)% vs (78.8±3.5)%]. Confocal laser scanning microscopy showed that EcN was successfully conjugated on the surface DMP7/3 micro-particles. Compared with DMP microparticles without EcN conjugation, the EcN-conjugated micro-particles (EcN@DMP7/3) permeated efficiently into the central area in the in vitro 3D tumor model. In the tumor-bearing mice, bio-distribution analyses revealed that at 4 and 24 h after intravenous administration, EcN@DMP7/3 resulted in 3-folder and 6-folder increases in Dox accumulation in the tumors, respectively, as compared with the non-conjugated DMP7/3 micro-particles (P < 0.01). Conclusion EDC-NHS catalytic reaction allows the amino groups on the outer surface of EcN to covalently bind to anhydride on the surface of drug-loaded PSMA/PELA particles without affecting the growth activity of EcN, which effectively drives the drug-loaded particles to target the hypoxic region inside the tumor.

Keywords

• escherichia coli nissle 1917
• active targeting
• hypoxic area
• permeation