Efficacy and Molecular Effects of a Reduced Graphene Oxide/Fe₃O₄ Nanocomposite in Photothermal Therapy Against Cancer

Figures & data

Figure 1 Results of the characterization test for the nanocomposite (A) Raman spectroscopy of rGO-Fe₃O₄, (B) TEM micrographs before and (C) after reduction of graphene oxide and simultaneous attaching of Fe₃O₄ nanoparticles. Red arrows point Fe₃O_4. (D) Visual response of the nanocomposite dissolved in water and exposed to an external permanent magnetic field. (E) Magnetic characterization by VSM at room temperature. The inset shows close view of hysteresis loop.

Table 1 Parameters of the Hysteresis Loop for rGO-Fe₃O₄ and Similar Nanocomposites from Other Studies

Nanocomposite	Production Method	Ms [emu/g]	MR [emu/g]	HC [Oe]	Ref.
rGO-Fe3O4	Chemical reaction with magnetite and reduction with hydrazine	51.1	9.8	88.1	(this work)
M-RGO	Chemical reaction with magnetite and reduction with hydrazine	22.3	0.3	12.0	[^Citation29]
CFO/rGO	Solvothermal method	53.5	12.9	347.5	[^Citation32]
GO–Fe3O4	Inverse chemical co‐precipitation	69.3	13.2	114.0	[^Citation41]
RGO-CoFe2O4	Simple reaction with NaBH4	53.6	25.3	768.0	[^Citation38]
rGO/Fe3O4 NC	Solvothermal method	19.7	1.4	60.4	[^Citation42]
CoOx@C-rGO	Solvothermal method and calcination with glucose	70.6	14.1	399.9	[^Citation39]
RGO/Sr2CuMgFe28O46	Green sol-gel method	18.6	10.9	4803.0	[^Citation43]

Figure 2 Cytotoxicity evaluation. Relative cell viability evaluated by trypan blue, MTT and Calcein AM assays. Statistical analysis was conducted using two-way ANOVA and a Dunnet’s multi comparison test. * is used for p-value < 0.05, ** for p-value < 0.01, *** for p-value < 0.001 and **** for p-value < 0.0001. Error bars depict SD of data.

Figure 3 Interaction between cells and graphene nanocomposite. Fluorescent confocal micrographs of HeLa cells with the nanocomposite pointed with yellow arrows for dark aggregates and rounded in red for intense fluorescent deposits. The area enclosed in white squares is zoomed in the last row. Scale bar is 5 µm.

Table 2 Irradiation Condition, Maximum Temperature Difference, and Cell Viability After Photothermal Therapy with Different Graphene-Based Nanocomposites

Nanocomposite	Concentration [µg/mL]	Irradiation Conditions (Wavelength, Power or Power Density, Duration)	Maximum ΔT[°C]	Cell line	% Cell Viability After Treatment	Ref.
rGO-Fe3O4	50	804 nm, 1 W/cm^2, 5 min	18.0	HeLa	32.6%	(this work)
	100		18.3		23.7%
mGO-CS/SA	50	808 nm, 1 W/cm^2, 5 min	18.0	A549	26.9%	[^Citation56]
	100		23.5		27.4%
N-O-CDs	200	808 nm, 0.8 W/cm^2, 5 min	32.2	HeLa	13.0%	[^Citation57]
GO-IONP-CS/DEX	50	808 nm, 1 W/cm^2, 5 min	17.5	A549	28.1%	[^Citation58]
GO/MnWO4/PEG	200	808 nm, 0.6 W/cm^2, 15 min	26.5	4T1	11.0%*	[^Citation59]
GQDs-Fe/Bi NPs	50	808 nm, 1.7 W/cm^2, 10 min	15.0	HeLa	68.0%	[^Citation55]
	100		28.5		50.0%
Cu2−xSe@rGO	50	980 nm, 1.0 W, 10 min	23.5	HEp-2	70.0%	[^Citation60]
MGBP	100	808 nm, 1 W/cm^2, 10 min	35.0	HeLa	22.0%	[^Citation61]
GO-PEG(TP)	50	980 nm, 0.5 W/cm^2, 10 min	21.5	4T1	16.0%**	[^Citation62]
9T-GQDs	100	1064 nm, 1.0 W/cm^2, 5 min	18.0	4T1	43.0%	[^Citation63]
GO/BaHoF5/PEG	100	808 nm, 0.4 W/cm^2, 10 min	12.8	HeLa	35.0%	[^Citation64]

Notes: *Irradiation duration for cell viability assay was 10 min. **Concentration of nanocomposite for cell viability assay was GO-PEG = 1 mg/mL and TP = 1 mmol/L.

Figure 4 Photothermal behavior of the nanocomposite. (A) Temperature profile of the rGO-Fe₃O₄ dissolved in water and increasing the surroundings temperature while it is irradiated with an 804 nm optical laser (1 W/cm² for 5 minutes). Temperatures in all treatments were normalized to the same initial room temperature (21.1°C). (B) Thermal images of the nanocomposite (100 µg/mL) contained in a petri dish (35 mm) and under the same irradiation conditions. The temperature described is the highest temperature in the dish.

Figure 5 Effect of the photothermal therapy. (A) Efficacy of the therapy evaluated as the relative cell viability. Analysis was conducted using one-way ANOVA and a Dunnet’s multiple comparison test. (B) Relative expression of mRNA of Hsp70 and Bcl-2 genes using β-actin as housekeeping gen. Statistical analysis was conducted using two way ANOVA with Sidak’s multiple comparison test. * refers to a p-value < 0.05. Error bars depict SD of data.

Chandra V, Park J, Chun Y, Lee JW, Hwang I, Kim KS. Water-dispersible magnetite-reduced graphene oxide composites for arsenic removal. ACS Nano. 2010;4(7):3979–3986. doi:10.1021/nn100889720552997

 PubMed Web of Science ®Google Scholar

Liu Y, Chen Z, Zhang Y, et al. Broadband and lightweight microwave absorber constructed by in situ growth of hierarchical CoFe 2 O 4/reduced graphene oxide porous nanocomposites. ACS Appl Mater Interfaces. 2018;10(16):13860–13868. doi:10.1021/acsami.8b0213729589899

 PubMed Web of Science ®Google Scholar

Szymczyk A, Paszkiewicz S, Typek J, et al. Magnetic properties of poly(trimethylene terephthalate‐block‐poly(tetramethylene oxide) copolymer nanocomposites reinforced by graphene oxide–Fe3O4 hybrid nanoparticles. Phys Status Solidi. 2019;216(23):1900402. doi:10.1002/pssa.201900402

 Google Scholar

Zong M, Huang Y, Zhang N. Reduced graphene oxide-CoFe2O4 composite: synthesis and electromagnetic absorption properties. Appl Surf Sci. 2015;345:272–278. doi:10.1016/j.apsusc.2015.03.203

 Web of Science ®Google Scholar

Vinodhkumar G, Wilson J, Inbanathan SSR, Potheher IV, Ashokkumar M, Peter AC. Solvothermal synthesis of magnetically separable reduced graphene oxide/Fe3O4 hybrid nanocomposites with enhanced photocatalytic properties. Phys Rev B Condens Matter. 2020;580:411752. doi:10.1016/j.physb.2019.411752

 Web of Science ®Google Scholar

Liu Y, Guo H, Sun K, Jiang J. Magnetic CoOx@C-reduced graphene oxide composite with catalytic activity towards hydrogen generation. Int J Hydrogen Energy. 2019;44(52):28163–28172. doi:10.1016/j.ijhydene.2019.09.034

 Web of Science ®Google Scholar

Hakimi M, Alimard P, Yousefi M. Green synthesis of reduced graphene oxide/Sr2CuMgFe 28O46 nanocomposite with tunable magnetic properties. Ceram Int. 2014;40(8PART A):11957–11961. doi:10.1016/j.ceramint.2014.04.032

 Google Scholar

Xie M, Zhang F, Peng H, et al. Layer-by-layer modification of magnetic graphene oxide by chitosan and sodium alginate with enhanced dispersibility for targeted drug delivery and photothermal therapy. Colloids Surf B Biointerfaces. 2019;176:462–470. doi:10.1016/j.colsurfb.2019.01.02830682619

 PubMed Web of Science ®Google Scholar

Geng B, Yang D, Pan D, et al. NIR-responsive carbon dots for efficient photothermal cancer therapy at low power densities. Carbon. 2018;134:153–162. doi:10.1016/j.carbon.2018.03.084

 Web of Science ®Google Scholar

Zhang F, Xie M, Zhao Y, et al. Chitosan and dextran stabilized GO-iron oxide nanosheets with high dispersibility for chemotherapy and photothermal ablation. Ceram Int. 2019;45(5):5996–6003. doi:10.1016/j.ceramint.2018.12.070

 Web of Science ®Google Scholar

Chang X, Zhang Y, Xu P, Zhang M, Wu H, Yang S. Graphene oxide/MnWO 4 nanocomposite for magnetic resonance/photoacoustic dual-model imaging and tumor photothermo-chemotherapy. Carbon. 2018;138:397–409. doi:10.1016/j.carbon.2018.07.058

 Web of Science ®Google Scholar

Badrigilan S, Shaabani B, Gharehaghaji N, Mesbahi A. Iron oxide/bismuth oxide nanocomposites coated by graphene quantum dots: “three-in-one” theranostic agents for simultaneous CT/MR imaging-guided in vitro photothermal therapy. Photodiagnosis Photodyn Ther. 2018;25:504–514. doi:10.1016/j.pdpdt.2018.10.02130385298

 PubMed Web of Science ®Google Scholar

Zhen SJ, Wang TT, Liu YX, Wu ZL, Zou HY, Huang CZ. Reduced graphene oxide coated Cu2−xSe nanoparticles for targeted chemo-photothermal therapy. J Photochem Photobiol B Biol. 2018;180:9–16. doi:10.1016/j.jphotobiol.2018.01.020

 PubMed Web of Science ®Google Scholar

Su Y, Wang N, Liu B, et al. A phototheranostic nanoparticle for cancer therapy fabricated by BODIPY and graphene to realize photo-chemo synergistic therapy and fluorescence/photothermal imaging. Dyes Pigm. 2020;177:108262. doi:10.1016/j.dyepig.2020.108262

 Web of Science ®Google Scholar

Liu J, Yuan X, Deng L, et al. Graphene oxide activated by 980 nm laser for cascading two-photon photodynamic therapy and photothermal therapy against breast cancer. Appl Mater Today. 2020;20:100665. doi:10.1016/j.apmt.2020.100665

 Web of Science ®Google Scholar

Liu H, Li C, Qian Y, et al. Magnetic-induced graphene quantum dots for imaging-guided photothermal therapy in the second near-infrared window. Biomaterials. 2020;232:119700. doi:10.1016/j.biomaterials.2019.11970031881379

 PubMed Web of Science ®Google Scholar

Chang X, Zhang M, Wang C, Zhang J, Wu H, Yang S. Graphene oxide/BaHoF5/PEG nanocomposite for dual-modal imaging and heat shock protein inhibitor-sensitized tumor photothermal therapy. Carbon. 2020;158:372–385. doi:10.1016/j.carbon.2019.10.105

 Web of Science ®Google Scholar