Nanomaterial genotoxicity evaluation using the high-throughput p53-binding protein 1 (53BP1) assay

Area of Science:

• Nanomaterial toxicology
• Genotoxicity testing
• DNA damage response

Background:

• Engineered nanomaterials (NMs) pose challenges for toxicity evaluation due to assay interference and the vast number of materials.
• Robust, high-throughput assays are crucial for assessing NM hazard potential.
• The DNA repair protein 53BP1 (p53-binding protein 1) foci assay is explored for genotoxicity assessment.

Purpose of the Study:

• Evaluate the applicability of a high-throughput 53BP1 foci assay for NM genotoxicity assessment.
• Benchmark the 53BP1 assay against known genotoxic agents and X-ray irradiation.
• Assess the genotoxicity and reactive oxygen species (ROS) production of various NMs and nanobiomaterials (NBMs).

Main Methods:

• Applied a 53BP1 foci immunostaining assay in a high-throughput format.
• Benchmarked the assay with eight known genotoxic agents and X-ray irradiation.
• Evaluated NMs and NBMs for cell viability, genotoxicity (53BP1 and micronucleus assays), and ROS production.

Main Results:

• The 53BP1 assay successfully identified genotoxic compounds in the HCT116 cell line.
• Most tested NMs did not show genotoxicity via the 53BP1 assay, except for (CoO)(NiO) NMs.
• TiO2 NMs were positive in the micronucleus assay, and Fe3O4 NMs elevated ROS without causing DNA damage.

Conclusions:

• The 53BP1 assay demonstrates adequate predictivity for genotoxicity of benchmark substances.
• The 53BP1 assay is suitable for high-throughput screening of NM genotoxicity.
• The 53BP1 assay can serve as a complementary tool in New Approach Methodologies for NM safety assessment.