Folate-targeting annonaceous acetogenins nanosuspensions: significantly enhanced antitumor efficacy in HeLa tumor-bearing mice

Figures & data

Figure 1. The particle size distribution and morphology of ACGs-NSps. (A) The particle size of FA-PEG-ACGs-NSps and PEG-ACGs-NSps measured by DLS. (B) TEM micrograph of FA-PEG-ACGs-NSps. (C) TEM micrograph of PEG-ACGs-NSps.

Figure 2. The stability and in vitro cumulative release profiles of ACGs-NSps. Particle size change of FA-PEG-ACGs-NSps (A) and PEG-ACGs-NSps (B) in 0.9% NaCl, 5% glucose, PBS, and plasma at 37 °C (n = 3, mean ± SD). (C) The in vitro cumulative release profiles of ACGs from FA-PEG-ACGs-NSps at 37 °C in pH 7.4 PBS. Notes: The amount of ACGs released from NSps was estimated by the reduction of quantity inside the dialysis bag with the HPLC method. All data are represented as mean ± SD (n = 3).

Figure 3. In vitro antiproliferative activity and in vivo antitumor experiments on HeLa tumor-bearing mice. (A) Cytotoxicity of ACGs solution, PEG-ACGs-NSps, FA-PEG-ACGs-NSps, FA + PEG-ACGs-NSps, and FA + FA-PEG-ACGs-NSps against HeLa cells (a) and A549 cells (b) for 24 h using MTT assay. All data are represented as mean ± SD (n = 6). (B) (a) The growth of relative tumor volume over time of PTX injections, PEG-ACGs-NSps, and FA-PEG-ACGs-NSps; (b) the average body weight change of mice along with time. All data are represented as mean ± SD. *p < .01, **p < .001 with FA-PEG-ACGs-NSps; &p < .01 with PEG-ACGs-NSps; #p < .01, ##p < .001 with normal saline. (C) The growth of relative tumor volume over time of PTX injections, PEG-ACGs-NSps, and FA-PEG-ACGs-NSps. (D) The average body weight change of mice along with time. All data are represented as mean ± SD. *p < 0.01, **p < 0.001 with FA-PEG-ACGs-NSps; &p < 0.01 with PEG-ACGs-NSps; #p < 0.01, ##p < 0.001with normal saline.

Table 1. IC50 values of ACGs solution, PEG-ACGs-NSps, FA-PEG-ACGs-NSps, FA + PEG-ACGs-NSps, and FA + FA-PEG-ACGs-NSps against HeLa and A549 cell lines after incubation for 24 h.

	IC50 (μg/mL)
Time	Groups	HeLa	A549
24 h	ACGs solution	1.308**	7.494
	PEG-ACGs-NSps	0.915*	7.285
	FA-PEG-ACGs-NSps	0.483	7.201
	FA + PEG-ACGs-NSps	0.936*	7.215
	FA + FA-PEG-ACGs-NSps	1.113*	7.254

Results are presented as mean ± SD; n = 6.

* p < .05.

** p < .01 vs. FA-PEG-ACGs-NSps.

Table 2. In vivo antitumor effects of different groups against HeLa tumors in mice.

Formulation	Tumor weight	Inhibition rate (%)
Normal saline	4.124 ± 0.237	NA
PTX injections (8 mg/kg)	1.927 ± 0.288**,&	53.23%
PEG-ACGs-NSps (0.4 mg/kg)	3.081 ± 0.252*	25.29%
FA-PEG-ACGs-NSps (0.4 mg/kg)	0.970 ± 0.057**^,#,&&	76.45%

Tumor weight results are presented as mean ± SD, n = 8.

* p < .01.

** p < .001 vs. normal saline.

# p < .01 vs. PTX injection.

& p < .01.

&& p < .001 vs. PEG-ACGs-NSps.

Figure 4. The in vivo biodistribution of PEG-ACGs/DiR-NSps, and FA-PEG-ACGs/DiR – NSps in HeLa tumor-bearing mice. (A) Average fluorescence intensity in tumors of two groups at the end of the experiment. (B) The ratio of the fluorescence intensity in tumor to the fluorescence intensity in liver (Te to liver) of two groups at the end of the experiment. All data are represented as mean ± SD. *p < .05, **p < .01 with FA-ACGs-NSps.