br measured at the end of test and
measured at the end of test and the image of excised tumors further illustrated the similar observations (Fig. 8B and Supporting Information Fig. S27). These results are in agreement with the tendency of in vitro cellular uptake, cytotoxicity, cell apoptosis, as well as in vivo drug accumulation in tumors.
In order to explore the mechanism of combined administration, cell apoptosis was detected by TUNEL technique, and the auto-phagy of tumor Solasodine was examined by TEM and immunofluo-rescence. As displayed in Fig. 8C‒E, as well as Supporting Information Fig. S28, compared with 7pep-M-PTX mono-therapy, 7pep-M-RAP used in single induced less cell apoptosis but more autophagic vesicles accumulation in tumor tissue, while the combination of 7pep-M-RAP and 7pep-M-PTX resulted in not
only significantly more apoptosis but also higher level of auto-phagic vesicle accumulation. It is known that RAP is an auto-phagy inducer, and PTX is an apoptosis inducer that acts on tubulin. The combination of the two nanomedicines led to an in-crease in apoptosis and autophagy. Based on the in vitro findings, it could be speculated that the synergistic effect of 7pep-M-RAP and 7pep-M-PTX might be partly due to ACD induced by RAP and apoptosis induced by PTX.
3.7. Toxicity studies after intravenous administration
The changes of body weight, leucocyte, neutrophil and platelet count in nude mice after administration were also examined to
Please cite this article as: Mei D et al., Actively priming autophagic cell death with novel transferrin receptor-targeted nanomedicine for synergistic chemotherapy against breast cancer, Acta Pharmaceutica Sinica B, https://doi.org/10.1016/j.apsb.2019.03.006
Priming autophagic cell death with transferrin receptor-targeted nanomedicine 15
Figure 9 Toxicity studies after intravenous administration. (A) Body weight changes of mice during antitumor efficacy study (mean SD, n Z 6). Arrows indicate the time for injection after tumor cells inoculation. *P < 0.05 vs PBS. (B) Effects of different formulations on white blood cells (WBC) counts at the end of test (mean SD, n Z 5). *P < 0.05 and **P < 0.01 vs PBS; #P < 0.05 vs M-PTX; 66P < 0.01 vs 7pep-M-PTX; ,P < 0.05 vs 7pep-M-Combi. (C) Optical microscopy images of H&E staining livers after treated with different formulations. Magnification, 200 . Red circles indicate the necrotic area in livers.
evaluate the chemotherapy associated toxicity. There was no sig-nificant body weight loss for all treated animals except that free PTX group and free drugs combination group showed a decline trend in body weight (Fig. 9A), indicating the acceptable tolerance of different nanomedicines, especially the combinational therapies. As shown in Fig. 9B, as well as Supporting Information Figs. S29 and S30, compared with control group, the combination group of free PTX and free RAP exhibited a significant decrease in WBC, GRN and PLT counts, while nanomedicines combination group only had a minor adverse effect on the blood cells (P > 0.5), implying reduced hematotoxicity of chemotherapeutic agents due to the encapsulation in nanocarriers. H&E staining results of excised livers showed that free PTX, Free Combi and M-Combi caused obvious liver necrosis, while no apparent liver toxicity was observed in other groups (Fig. 9C). The safety advantages of nanomedicines were likely due to the lack of haemolytic Cremo-phor EL and the modification of 7pep on micelles, resulting in less non-specific distribution and greater accumulation at tumor sites, which was in accordance with the tissue distribution results in vivo.
Therefore, combinational therapies using 7pep-M-RAP with 7pep-M-PTX could achieve better anti-tumor efficacy with low systemic and hematotoxicity.