Ultra-small gadolinium oxide nanocrystal sensitization of non-small-cell lung cancer cells toward X-ray irradiation by promoting cytostatic autophagy

Feifei Li1 Institute of Modern Physics, Chinese Academy of Sciences, Key Laboratory of Heavy Ion Radiation Biology and Medicine of Chinese Academy of Sciences, Key Laboratory of Basic Research on Heavy Ion Radiation Application in Medicine, Gansu Province, Lanzhou 730000, China, [email protected]; [email protected];2 University of Chinese Academy of Sciences, Beijing 100049, China

Zihou Li2 University of Chinese Academy of Sciences, Beijing 100049, China;3 Key Laboratory of Magnetic Materials and Devices, Chinese Academy of Sciences, Division of Functional Materials and Nano Devices, Ningbo Institute of Materials Technology and Engineering, Chinese Academy of Sciences, Ningbo, Zhejiang 315201, China

Xiaodong Jin1 Institute of Modern Physics, Chinese Academy of Sciences, Key Laboratory of Heavy Ion Radiation Biology and Medicine of Chinese Academy of Sciences, Key Laboratory of Basic Research on Heavy Ion Radiation Application in Medicine, Gansu Province, Lanzhou 730000, China, [email protected]; [email protected]

Yan Liu1 Institute of Modern Physics, Chinese Academy of Sciences, Key Laboratory of Heavy Ion Radiation Biology and Medicine of Chinese Academy of Sciences, Key Laboratory of Basic Research on Heavy Ion Radiation Application in Medicine, Gansu Province, Lanzhou 730000, China, [email protected]; [email protected]

Pengcheng Zhang1 Institute of Modern Physics, Chinese Academy of Sciences, Key Laboratory of Heavy Ion Radiation Biology and Medicine of Chinese Academy of Sciences, Key Laboratory of Basic Research on Heavy Ion Radiation Application in Medicine, Gansu Province, Lanzhou 730000, China, [email protected]; [email protected];2 University of Chinese Academy of Sciences, Beijing 100049, China

Ping Li1 Institute of Modern Physics, Chinese Academy of Sciences, Key Laboratory of Heavy Ion Radiation Biology and Medicine of Chinese Academy of Sciences, Key Laboratory of Basic Research on Heavy Ion Radiation Application in Medicine, Gansu Province, Lanzhou 730000, China, [email protected]; [email protected]

Zheyu Shen3 Key Laboratory of Magnetic Materials and Devices, Chinese Academy of Sciences, Division of Functional Materials and Nano Devices, Ningbo Institute of Materials Technology and Engineering, Chinese Academy of Sciences, Ningbo, Zhejiang 315201, China

Aiguo Wu3 Key Laboratory of Magnetic Materials and Devices, Chinese Academy of Sciences, Division of Functional Materials and Nano Devices, Ningbo Institute of Materials Technology and Engineering, Chinese Academy of Sciences, Ningbo, Zhejiang 315201, China

Weiqiang Chen1 Institute of Modern Physics, Chinese Academy of Sciences, Key Laboratory of Heavy Ion Radiation Biology and Medicine of Chinese Academy of Sciences, Key Laboratory of Basic Research on Heavy Ion Radiation Application in Medicine, Gansu Province, Lanzhou 730000, China, [email protected]; [email protected]Correspondence[email protected] [email protected]

Qiang Li1 Institute of Modern Physics, Chinese Academy of Sciences, Key Laboratory of Heavy Ion Radiation Biology and Medicine of Chinese Academy of Sciences, Key Laboratory of Basic Research on Heavy Ion Radiation Application in Medicine, Gansu Province, Lanzhou 730000, China, [email protected]; [email protected]Correspondence[email protected] [email protected]

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Abstract

Background

Gadolinium-based nanoparticles (GdNPs) have been used as theranostic sensitizers in clinical radiotherapy studies; however, the biomechanisms underlying the radio-sensitizing effects of GdNPs have yet to be determined. In this study, ultra-small gadolinium oxide nanocrystals (GONs) were employed to investigate their radiosensitizing effects and biological mechanisms in non-small-cell lung cancer (NSCLC) cells under X-ray irradiation.

Method and materials

GONs were synthesized using polyol method. Hydroxyl radical production, oxidative stress, and clonogenic survival after X-ray irradiation were used to evaluate the radiosensitizing effects of GONs. DNA double-strand breakage, cell cycle phase, and apoptosis and autophagy incidences were investigated in vitro to determine the radiosensitizing biomechanism of GONs under X-ray irradiation.

Results

GONs induced hydroxyl radical production and oxidative stress in a dose- and concentration-dependent manner in NSCLC cells after X-ray irradiation. The sensitizer enhancement ratios of GONs ranged between 19.3% and 26.3% for the NSCLC cells under investigation with a 10% survival rate compared with that of the cells treated with irradiation alone. Addition of 3-methyladenine to the cell medium decreased the incidence rate of autophagy and increased cell survival, supporting the idea that the GONs promoted cytostatic autophagy in NSCLC cells under X-ray irradiation.

Conclusion

This study examined the biological mechanisms underlying the radiosensitizing effects of GONs on NSCLC cells and presented the first evidence for the radiosensitizing effects of GONs via activation of cytostatic autophagy pathway following X-ray irradiation.

Keywords:

Supplementary materials

Figure S1 Ultraviolet-visible spectra results confirm that GON solution is very stable in RPMI 1640 medium.

Notes: (A) Absorbance of GON with different concentrations after incubating at 37°C for 24 hours. (B, C) Absorbance of GON at concentrations 0.5 µg/mL (B) and 5.0 µg/mL (C) at various incubation time.

Abbreviation: GONs, gadolinium oxide nanocrystals.

Figure S1 Ultraviolet-visible spectra results confirm that GON solution is very stable in RPMI 1640 medium.Notes: (A) Absorbance of GON with different concentrations after incubating at 37°C for 24 hours. (B, C) Absorbance of GON at concentrations 0.5 µg/mL (B) and 5.0 µg/mL (C) at various incubation time.Abbreviation: GONs, gadolinium oxide nanocrystals.

Figure S2 The influence of GON on cell cycle distribution.

Notes: (A) Cell flow spectra at 24 hours after X-ray irradiation. (B) Cell cycle distributions of three studied cells on time. CK represents the control; CO represents co-treatment with irradiation and GONs.

Abbreviations: GONs, gadolinium oxide nanocrystals; Gd, gadolinium; IR, irradiation.

Figure S2 The influence of GON on cell cycle distribution.Notes: (A) Cell flow spectra at 24 hours after X-ray irradiation. (B) Cell cycle distributions of three studied cells on time. CK represents the control; CO represents co-treatment with irradiation and GONs.Abbreviations: GONs, gadolinium oxide nanocrystals; Gd, gadolinium; IR, irradiation.

Figure S3 Apoptotic rates of A549, NH1299, and NH1650 cells treated with GON and/or radiation for 24-hour and 48-hour posttreatment.

Note: CK represents the control; CO represents co-treatment with irradiation and GONs.

Abbreviations: GONs, gadolinium oxide nanocrystals; Gd, gadolinium; IR, irradiation.

Figure S3 Apoptotic rates of A549, NH1299, and NH1650 cells treated with GON and/or radiation for 24-hour and 48-hour posttreatment.Note: CK represents the control; CO represents co-treatment with irradiation and GONs.Abbreviations: GONs, gadolinium oxide nanocrystals; Gd, gadolinium; IR, irradiation.

Figure S4 Autophagy incidence of A549, NH1299, and NH1650 cells treated with GON and/or X-ray irradiation for 4-hour and 12-hour posttreatment.

Note: CK represents the control; CO represents co-treatment with irradiation and GONs.

Abbreviations: GONs, gadolinium oxide nanocrystals; Gd, gadolinium; IR, irradiation.

Figure S4 Autophagy incidence of A549, NH1299, and NH1650 cells treated with GON and/or X-ray irradiation for 4-hour and 12-hour posttreatment.Note: CK represents the control; CO represents co-treatment with irradiation and GONs.Abbreviations: GONs, gadolinium oxide nanocrystals; Gd, gadolinium; IR, irradiation.

Figure S5 Radiation-induced damages in cytoplasm stimulate the ER stress and mitochondrion dysfunction.

Note: CK represents the control; CO represents co-treatment with irradiation and GONs.

Abbreviations: ER, endoplasmic reticulum; Gd, gadolinium; IR, irradiation.

Figure S5 Radiation-induced damages in cytoplasm stimulate the ER stress and mitochondrion dysfunction.Note: CK represents the control; CO represents co-treatment with irradiation and GONs.Abbreviations: ER, endoplasmic reticulum; Gd, gadolinium; IR, irradiation.

Acknowledgments

The authors acknowledge the support from the National Key Research and Development Program (2017YFC0108500), the National Key Technology Support Program of the Ministry of Science and Technology of China (2015BAI01B11), the National Natural Science Foundation of China (11875299), and the CAS Key Laboratory of Heavy Ion Radiation Biology and Medicine, Institute of Modern Physics (2016-01).

Disclosure

The authors report no conflicts of interest in this work.