Effects of Doxorubicin Delivery by Nitrogen-Doped Graphene Quantum Dots on Cancer Cell Growth: Experimental Study and Mathematical Modeling

Abstract

With 18 million new cases diagnosed each year worldwide, cancer strongly impacts both science and society. Current models of cancer cell growth and therapeutic efficacy in vitro are time-dependent and often do not consider the E_max value (the maximum reduction in the growth rate), leading to inconsistencies in the obtained IC₅₀ (concentration of the drug at half maximum effect). In this work, we introduce a new dual experimental/modeling approach to model HeLa and MCF-7 cancer cell growth and assess the efficacy of doxorubicin chemotherapeutics, whether alone or delivered by novel nitrogen-doped graphene quantum dots (N-GQDs). These biocompatible/biodegradable nanoparticles were used for the first time in this work for the delivery and fluorescence tracking of doxorubicin, ultimately decreasing its IC₅₀ by over 1.5 and allowing for the use of up to 10 times lower doses of the drug to achieve the same therapeutic effect. Based on the experimental in vitro studies with nanomaterial-delivered chemotherapy, we also developed a method of cancer cell growth modeling that (1) includes an E_max value, which is often not characterized, and (2), most importantly, is measurement time-independent. This will allow for the more consistent assessment of the efficiency of anti-cancer drugs and nanomaterial-delivered formulations, as well as efficacy improvements of nanomaterial delivery.

Keywords: IC50; cancer; doxorubicin; drug delivery; fluorescence; graphene quantum dots; imaging; mathematical modeling; nanoparticles.

Figure 1

(a) Growth of HeLa (red), MCF-7 (orange), and HEK293 (yellow) cells over 14 days. (b) HEK 293, (c) HeLa, and (d) MCF-7 growth curves after treatment with doxorubicin at 0 mg/mL control (red), 0.00005 μg/mL (orange), 0.0005 μg/mL (yellow), 0.005 μg/mL (green), and 0.05 μg/mL (blue) concentrations. (e) Cytotoxicity of doxorubicin in MCF-7 (orange), HeLa (red), and HEK 293 (yellow) cells, evaluated via a thiazolyl blue tetrazolium bromide (MTT) cell viability assay.

Figure 2

Nitrogen-doped graphene quantum dot (N-GQD) (black), doxorubicin (DOX) (red), and DOX-N-GQD (blue) complex absorption (a) and fluorescence (b) spectra.

Figure 3

(a) Confocal 3D z-stack fluorescence images of DOX-N-GQD complex in MCF-7 cells at (a) 3 h, (b) 6 h, (c) 9 h, (d) 12 h, and (e) 24 h time points. Emission mainly originated from cell nuclei. (f) Variation of average fluorescence of doxorubicin per cell unit area in MCF-7 cells with time reflecting internalization dynamics.

Figure 3

(a) Confocal 3D z-stack fluorescence images of DOX-N-GQD complex in MCF-7 cells at (a) 3 h, (b) 6 h, (c) 9 h, (d) 12 h, and (e) 24 h time points. Emission mainly originated from cell nuclei. (f) Variation of average fluorescence of doxorubicin per cell unit area in MCF-7 cells with time reflecting internalization dynamics.

Figure 4

(a) Cytotoxicity of DOX (yellow), N-GQDs (orange), and DOX-N-GQDs (red) in MCF-7, as evaluated via an MTT cell viability assay. (b) MCF-7 cell growth curve up to 14 days after treatment with DOX-GQDs with DOX concentrations of 0 μg/mL (N-GQDs alone) (red), 0.00005 μg/mL (orange), 0.0005 μg/mL (yellow), 0.005 μg/mL (green), and 0.05 μg/mL (blue).

Figure 5

(a) Mathematical model fit to growth data of MCF-7 cells treated with DOX-N-GQDs. (b) Mathematical model fit to growth data of MCF-7 treated with DOX alone. (c) Best fit parameters with 95% confidence interval for MCF-7 cell growth.

Figure 6

Rate of MCF-7 survival after (a) DOX, (b) DOX-N-GQD treatment at day 2 (red), day 4 (orange), day 6 (yellow), and day 8 (green).

Figure 7

(a) IC₅₀ determined from the simulated MTT assays. (b) IC₅₀ (concentration of the drug at half maximum effect) for DOX (red) or DOX-N-GQD (blue) treatment in MCF-7 cells. (c) Simulation of the MTT assay for doxorubicin alone (circles) and DOX-N-GQDs (squares) using the fit parameters in Figure 5c.

Figure 7

(a) IC₅₀ determined from the simulated MTT assays. (b) IC₅₀ (concentration of the drug at half maximum effect) for DOX (red) or DOX-N-GQD (blue) treatment in MCF-7 cells. (c) Simulation of the MTT assay for doxorubicin alone (circles) and DOX-N-GQDs (squares) using the fit parameters in Figure 5c.