Synergistic and complete reversal of the multidrug resistance of mitoxantrone hydrochloride by three-in-one multifunctional lipid-sodium glycocholate nanocarriers based on simultaneous BCRP and Bcl-2 inhibition

Figures & data

Table 1 Summary of particle size, zeta potential, and drug encapsulation efficiency of MTO-NLCs and MTO-TMLGNs (mean ± SD, n=3)

Nanoparticles	Particle size (nm)	Zeta potential (mV)	Encapsulation efficiency (%)
Blank NLCs	130.4±4.6	−31.3±2.5	–
MTO-NLCs	136.7±8.6	−19.9±1.4**	35.6±2.9
MTO-TMLGNs	125.0±4.0	−28.7±0.4	97.8±1.9**

Notes: Statistical significant difference compared to blank NLCs or MTO-NLCs;

** P<0.01.

Abbreviations: MTO, mitoxantrone hydrochloride; NLC, nanostructured lipid carrier; SD, standard deviation; TMLGNs, three-in-one multifunctional lipid-GcNa nanocarriers; GcNa, sodium glycocholate.

Figure 1 The proposed schematic illustration of three-in-one multifunctional lipid-GcNa nanocarriers (TMLGNs).

Abbreviations: GcNa, sodium glycocholate; BCRP, breast cancer resistance protein; PEG-PE, 1,2-Distearoyl-sn-glycero-3-phosphoethanolamine-N-[methoxy (polyethyleneglycol)-2000].

Figure 2 Characterization of three-in-one multifunctional lipid-GcNa nanocarriers (TMLGNs).

Notes: (A) TEM. (B) Particle size distribution of TMLGNs. (C) DSC curves. (D) In vitro release profiles of MTO in pH 7.4 PBS containing 0.2% Na₂SO₃ from MTO-Sol, MTO-NLCs, and MTO-TMLGNs determined by the dialysis bag technique (mean ± SD, n=6).

Abbreviations: GcNa, sodium glycocholate; TEM, transmission electron microscopy; MTO, mitoxantrone hydrochloride; NLC, nanostructured lipid carrier; SD, standard deviation; DSC, differential scanning calorimetry; CsA, cyclosporine A; PBS, phosphate buffered saline.

Table 2 Main pharmacokinetic parameters of MTO in rats after the intravenous administration of MTO-Sol and MTO-TMLGNs at dose 1 mg/kg (mean ± SD, n=6)

Parameters	MTO-Sol	MTO-TMLGNs
Cmax (ng/mL)	485.0±27.6	419.1±52.6**
t1/2 (h)	1.52±0.18	10.72±0.89**
AUC0–t (ng/mL·h)	201.9±58.3	928.0±172.6**
AUC0–∞ (ng/mL·h)	203.5±58.3	943.7±174.8**

Notes:

** Statistical significant difference between MTO-Sol and MTO-TMLGNs group; P<0.01.

Abbreviations: MTO, mitoxantrone hydrochloride; TMLGNs, three-in-one multifunctional lipid-GcNa nanocarriers; GcNa, sodium glycocholate; SD, standard deviation.

Figure 3 Distribution of MTO in tissues of mice (B – 0.5 hour; C – 4 hours) and mean plasma concentration–time curves of MTO in rats after the intravenous administration of MTO-Sol and MTO-TMLGNs at a dose of 1 mg/kg (A) (mean ± SD, n=6).

Abbreviations: MTO, mitoxantrone hydrochloride; TMLGNs, three-in-one multifunctional lipid-GcNa nanocarriers; GcNa, sodium glycocholate; SD, standard deviation.

Figure 4 In vitro cytotoxicity of MTO-Sol, MTO-CsA-GcNa-Sol, and MTO-TMLGNs.

Notes: Dose–response curves of MTO in MTO-Sol, MTO-CsA-GcNa-Sol, and MTO-TMLGNs. Curves of (A) MCF-7 cells for 96 hours; (B) MCF-7/MX cells for 48 hours; (C) MCF-7/MX cells for 72 hours; and (D) MCF-7/MX cells for 96 hours. (E) The 50% inhibition concentration (IC₅₀) of MTO-Sol, MTO-CsA-GcNa-Sol and MTO-TMLGNs against MCF-7/MX cells for 48, 72, and 96 hours and MCF-7 cells for 96 hours (mean ± SD, n=3).

Abbreviations: MTO, mitoxantrone hydrochloride; TMLGNs, three-in-one multifunctional lipid-GcNa nanocarriers; GcNa, sodium glycocholate; CsA, cyclosporine A; SD, standard deviation.

Table 3 The reversal factor (RF) of MTO formulations compared to MTO-Sol in MCF-7/MX cells at 48, 72, and 96 hours, and the resistant index (RI) of MTO formulations in MCF-7/MX compared to MCF-7 cells at 96 hours

MTO formulations	RI	RF
	96 hours	48 hours	72 hours	96 hours
MTO-Sol	55.0	–	–	–
MTO-CsA-GcNa-Sol	7.0	15.1	13.0	13.7
MTO-TMLGNs	1.1	226.8	265.6	389.6
MTO-NLDCs^Citation17	9.8	7.8	10.1	11.2

Abbreviations: MTO, mitoxantrone hydrochloride; TMLGNs, three-in-one multifunctional lipid-GcNa nanocarriers; GcNa, sodium glycocholate; CsA, cyclosporine A; NLDCs, nanostructured lipid–dextran sulfate hybrid carriers.

Figure 5 Cell uptake and endocytosis mechanism of MTO-TMLGNs.

Notes: (A) Cell uptake efficiency of MTO in MCF-7/MX cells after incubation with MTO-Sol, MTO-CsA-GcNa-Sol, and MTO-TMLGNs at an MTO concentration of 100 nM for 2 hours. (B) Effects of incubation temperature and endocytosis inhibitors on the uptake efficiency of MTO in MCF-7/MX cells after incubation with MTO-TMLGNs at an MTO concentration of 100 nM for 2 hours. (C) Schematic illustration of the proposed mechanism indicating the increased anticancer activity and reversed multidrug resistance by MTO-TMLGNs – (a) diffusion of released free MTO across the cell membrane and (b) endocytosis of MTO-TMLGNs. **Statistical significant difference compared to uptake efficiency of MTO without inhibitor, P<0.01.

Abbreviations: MTO, mitoxantrone hydrochloride; TMLGNs, three-in-one multifunctional lipid-GcNa nanocarriers; GcNa, sodium glycocholate; CsA, cyclosporine A.

Figure S1 Chemical structures of (A) mitoxantrone hydrochloride, (B) sodium glycocholate (GcNa), and (C) dextran sulfate sodium.

ZhangPLingGPanXNovel nanostructured lipid-dextran sulfate hybrid carriers (NLDCs) overcome tumor multidrug resistance of mitoxantrone hydrochlorideNanomedicine20128218519321704599

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