Dual Sensitization Anti-Resistant Nanoparticles for Treating Refractory Breast Cancers via Apoptosis-Inducing

Figures & data

Figure 1 Synthesis and characterization of HA-α-TOS (A and B) and Ce6-TPGS₁₀₀₀ (C and D).

Table 1 Characterization of Prepared Nanoparticles

Nanoparticles	Mean Size (nm)	PDI	Zeta Potential (mV)	EE (%) of Paclitaxel	EE(%) of Ce6-TPGS1000
Paclitaxel nanoparticles	68.1 ± 3.2	0.184 ± 0.012	−15.9 ± 0.4	98.5 ± 0.3	-
Ce6 nanoparticles	92.4 ± 4.3	0.225 ± 0.023	−15.2 ± 0.6	-	82.1 ± 0.4
Dual sensitization anti-resistance nanoparticles	108.5 ± 5.5	0.237 ± 0.019	−14.3 ± 0.5	97.6 ± 0.5	78.5 ± 1.2

Note: Data are presented as the mean ± SD.

Abbreviations: PDI, polydispersity index; EE, encapsulation efficiency.

Figure 2 Characterization of prepared nanoparticles.

Notes: (A) Schematic diagram of dual sensitization anti-resistance nanoparticles; (B) AFM images of paclitaxel nanoparticles (B1), Ce6 nanoparticle (B2), and dual sensitization anti-resistance nanoparticle (B3); (C) the size distribution of prepared nanoparticles; (D) the zeta potentials of prepared nanoparticles; (E) mean particle size of prepared nanoparticles in normal saline and 50% FBS solution; (F) in vitro release of paclitaxel in PBS (pH 7.4). All the data are presented as the mean ± standard deviation (n = 3).

Figure 3 In vitro cytotoxic effect after treating with varying formulations.

Notes: Dual sensitization anti-resistance nanoparticles and other formulations were applied to human breast cancer MCF-7 cells (A and B) or drug-resistant human breast cancer MCF-7/adr cells (C and D) for 48 h. The cells were treated in the dark environment (A and C) or irradiated by a 660nm laser (B and D) for 10 min. Data are presented as the mean ± standard deviation (n = 6).

Figure 4 Survival status of drug-resistant MCF-7/adr cells after treatment with varying nanoparticles in dark conditions (A) and after laser irradiation (B).

Notes: The cells were observed by a fluorescence microscope. The green fluorescence indicates live cells stained by calcein AM, while the red fluorescence was emitted by the dead cells stained by ethidium homodimer. The dashed lines indicate the laser boundary. Scale bar = 200 μm.

Figure 5 Cellular uptake of various coumarin nanoparticles.

Notes: (A) Confocal images of various coumarin nanoparticles in the drug-resistant MCF-7/adr cells after treating with coumarin nanoparticles. Scale bar = 25 μm; (B) mean fluorescence intensity of drug-resistant MCF-7/adr cells after exposure to varying coumarin nanoparticles for 1–8 h.

Table 2 Mean Fluorescence Intensity of MCF-7/Adr Cells After Various Nanoparticles’ Exposure

Time (h)/Groups	Control	Free Coumarin	Coumarin Nanoparticles	Dual Coumarin Nanoparticles
1	5.6 ± 1.2	26.4 ± 3.2	14.7 ± 3.5	23.2 ± 6.7
2	4.8 ± 1.7	56.7 ± 4.6	42.7 ± 4.3	49.8 ± 5.9
4	8.4 ± 1.4	76.3 ± 7.8	145.8 ± 13.8	178.4 ± 26.3
8	7.6 ± 0.7	98.6 ± 14.8	320.4 ± 29.4	350.7 ± 32.2

Figure 6 Apoptosis inducing of various nanoparticles.

Notes: Fluorescence images of the cells and their nuclei (A) were captured by a fluorescence microscope incorporated into the high-content screening system. The drug-resistant MCF-7/adr cells were stained with MitoTracker Red and Hoechst 33342 after treating with varying nanoparticles. The percentages of condensed cells and nuclear fracture (B) were obtained by the Columbus system. The apoptosis of MCF-7/adr cells after treatment was analyzed by flow cytometry after Annexin V/PI staining (C). Scale bar = 100 μm. *, p < 0.05 vs other formulations.

Figure 7 The changed apoptosis-related proteins of drug-resistant MCF-7/adr cells after treatment with various formulations.

Notes: The MCF-7/adr cells were treated with 1) normal saline, 2) free paclitaxel, 3) paclitaxel nanoparticles, 4) Ce6 nanoparticles, and 5) dual sensitization anti-resistant nanoparticles. Data are presented as the mean ± standard deviation (n = 3). *, p < 0.05 vs other formulations.

Figure 8 Anticancer efficacy (A) and body weight (B) of the tumor-bearing mice after intravenous administration of varying formulations.

Notes: Red arrows indicate the drug dosing time points. Data are presented as the mean ± standard deviation (n = 6). a, p < 0.05, versus normal saline; b, p < 0.05, versus free paclitaxel; c, p < 0.05, versus paclitaxel nanoparticles; d, p < 0.05, versus dual sensitization anti-resistance nanoparticles.