Nano-delivery vehicle based on chlorin E6, photodynamic therapy, doxorubicin chemotherapy provides targeted treatment of HER-2 negative, ανβ3-positive breast cancer

Abstract

The prognosis for patients with HER-2 negative breast cancer is currently poor, largely due to the lack of efficacious targeted therapeutics. Photodynamic nanomaterial technologies have rapidly developed in recent years, but their anti-tumor effects are often limited by poor targeting, low transformation efficiency, toxicity, and other factors. Thus, we prepared a new type of nanoparticles (Ce6/Dox@NPs-cRGD, CDNR) with cyclo(Arg-Gly-Asp-d-Phe-Cys) (c(RGDfC)) that target the ανβ3 receptor. We loaded those nanoparticles (NPs) with a combination of the doxorubicin (Dox) and photosensitizer chlorin E6 (Ce6) to test synergy between chemotherapy and photodynamic therapy (PDT) for the treatment of ανβ3 receptor positive and HER-2 negative breast cancer. Through analysis of the Fourier transform infrared and UV–vis spectra of these NPs, we found that Ce6 and Dox were successfully loaded into the CDNR. According to dynamic light scattering (DLS) analyses, CDNR particles had a diameter of 112.6 nm (polydispersity index 0.11), which was also confirmed via TEM characterization. The zeta potential was about -21.5 mV. Stability studies showed that CDNR particle size was stable in ddH2O, PBS, and DMEM + 5 % FBS for 16 days. The drug loading content of Dox and Ce6 were 5.3 and 6.8 %, respectively. Release studies of CDNR showed that the slow release of Dox was accelerated with increasing GSH concentration, and there was no burst release effect. From studying the absorbance of 9,10-dimethylanthrancene (ABDA), we found that CDNR produces high levels of ROS after excitation with a 670 nm laser, and ROS production increased with increasing radiation time. CDNR was significantly taken up by MCF-7 cells at 6 h because of cRGD targeting. In a CCK8 test, the relative growth rate (RGR) of CDNR +670 nm laser for MCF-7 cells was less than 75 % at 20 μg/mL after 24 h treatment and 15 μg/mL after 48 h treatment. We found that CDNR’s effects on RGR were concentration dependent. Live-cell staining with a DCFH-DA kit and flow cytometry assay further supported that a CDNR +670 nm laser provided the maximum chemotherapy-PDT toxicity and production of intracellular ROS, and that cell death was mainly caused by necrosis and apoptosis. In vivo experiments showed that using the cRGD-targeting strategy, CDNR had a stronger affinity and increased half-life relative to Ce6/Dox@NPs in mice with MCF-7 xenograft tumors. Further, the Cmax of CDNR in the transplanted tumor occurred 8 h post-injection (HPI) and there was still detectable signal at 24 HPI. In addition, MCF-7 bearing mice that were treated with CDNR +670 nm PDT at 8 HPI had a significantly decreased tumor volume (P < 0.05) and prolonged survival time compared to other groups. Thus, CDNR plus 670 nm PDT was associated with favorable anti-tumor activity with no appreciable impact on body weight or the major organs in mice, as determined by immunohistochemistry/immunofluorescence and hematoxylin-eosin staining. In conclusion, CDNR with 670 nm laser irradiation represents a promising new potential treatment paradigm for the management of breast cancers that are ανβ3-receptor positive and HER-2 negative.