PH-responsive star-shaped poly (ε-carprolactone)-co-poly maleic anhydride micelles for synergistic breast cancer combination chemotherapy

Abstract

Polymeric micelles produced from amphiphilic copolymers, are considered promising drug delivery platforms. Because of their ability to co-deliver hydrophilic (Dox) and hydrophobic (natural sesquiterpene coumarin, Conf) drugs through their shell and core, these nanosized micelles can benefit adjuvant therapy by modulating MDR proteins and increasing cellular uptake. In this study, a novel star-shaped biodegradable pH-sensitive pentaerythritol-grafted anhydride maleic copolymerized with Poly(ε-caprolactone) (PEE-g-PMA-co-PCL) was synthesized and developed as micellar nano-carriers for combination therapy of MDA-MB-231 breast cancer cell line. The physicochemical, characteristics of the synthesized copolymers and micelles were validated using Nuclear Magnetic Resonance Spectroscopy (NMR), Fourier Transform Infrared Spectroscopy (FTIR), CHNS elemental analysis, Dynamic Light Scattering (DLS), Zeta potential, Scanning Electron Microscopy (SEM), and Differential Scanning Calorimetry (DSC). The engineered micelles exhibited nearly 100% encapsulation efficiency for both Dox and Conf. Micelles possess a CMC value of 0.1 μg/mL, an average size of 40.3 nm with uniform distribution; ζ-potential of −11.5 mV in blank form and + 6.82 mV in co-drug-loaded form. Furthermore, micelles' in vitro biodegradation and drug release profiles followed an intensive pH-dependent pattern that benefited targeted delivery. The results of MTT, cellular uptake, cell cycle, and apoptosis by Annexin V assay on MDA-MB-231 cell line cells treated by co-drug loaded micelles (E2D) demonstrated higher cytotoxicity, more cellular uptake, superior inhibitory yield on DNA duplication, and greater apoptosis with a lower necrosis rate compared to monotherapy (ED and EC) and free drugs (Conf, Dox and 2D) which confirmed the synergistic effect of this combination therapy system (CI < 1) in a reduced Dox dosage. In addition, western blot and RT-PCR showed that co-drug-loaded micelles triggered an apoptotic response via the mitochondrial pathway. In conclusion, the novel developed multi-functional pH-sensitive micelles here could open a new perspective for the development of nanomedicine for combination chemo-adjuvant therapy against malignant cancer.