Complexation with C60 Fullerene Increases Doxorubicin Efficiency against Leukemic Cells In Vitro

Abstract

Conventional anticancer chemotherapy is limited because of severe side effects as well as a quickly evolving multidrug resistance of the tumor cells. To address this problem, we have explored a C₆₀ fullerene-based nanosized system as a carrier for anticancer drugs for an optimized drug delivery to leukemic cells.Here, we studied the physicochemical properties and anticancer activity of C₆₀ fullerene noncovalent complexes with the commonly used anticancer drug doxorubicin. C₆₀-Doxorubicin complexes in a ratio 1:1 and 2:1 were characterized with UV/Vis spectrometry, dynamic light scattering, and high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS). The obtained analytical data indicated that the 140-nm complexes were stable and could be used for biological applications. In leukemic cell lines (CCRF-CEM, Jurkat, THP1 and Molt-16), the nanocomplexes revealed ≤ 3.5 higher cytotoxic potential in comparison with the free drug in a range of nanomolar concentrations. Also, the intracellular drug's level evidenced C₆₀ fullerene considerable nanocarrier function.The results of this study indicated that C₆₀ fullerene-based delivery nanocomplexes had a potential value for optimization of doxorubicin efficiency against leukemic cells.

Keywords: Accumulation; C60 fullerene; Cytotoxicity; Doxorubicin; Leukemic cells; Noncovalent complex.

Fig. 1

Multiple reaction monitoring chromatograms of free Dox (1 μM), C₆₀-Dox 1:1 and C₆₀-Dox 2:1 (1 μM Dox-equivalent concentration) complexes under isocratic flow (acetonitrile, 0.1% formic acid in H₂O, 80:20, v:v), precursor → product ions transition: 544.2 → 130.2 and 361.1 m/z; a.u. arbitrary units

Fig. 2

Optical characterization of complexes. Optical density spectra of free Dox and C₆₀-Dox complexes (a). Fluorescence emission spectra of free Dox and C₆₀-Dox complexes at Dox-equivalent concentration from 3 to 50 μM (b); a.u. arbitrary units

Fig. 3

Hydrodynamic size (diameter, nm) of 1 μM С₆₀-Dox complexes in RPMI cell culture medium supplemented with 10% FBS at 0 (a) and 72-h (b) incubation. Intensity (%) percentage of all scattered light intensity

Fig. 4

Viability of CCRF-CEM, Jurkat, THP1 and Molt16 leukemic cells, treated with equal doses of free Dox or C₆₀-Dox complexes for 24, 48, and 72 h (*p ≤ 0.05 in comparison with the free Dox, **p ≤ 0.05 in comparison with the C₆₀-Dox 1:1 complex, n = 5)

Fig. 5

Intracellular accumulation of the 1 μM free and C₆₀ complexed Dox. Flow cytometry (a) and fluorescent microscopy images (b) of CCRF-CEM cells incubated with Dox and C₆₀-Dox at the ratio 1:1 and 2:1 for 1, 3 and 6 h. Scale bar 20 μM