Monitoring the Activation of the DNA Damage Response Pathway in a 3D Spheroid Model

Area of Science:

• Biochemistry
• Molecular Biology
• Cancer Research

Background:

• Multicellular tumor spheroids (3D models) are crucial for pharmacological evaluation.
• Monitoring the DNA-Damage Response (DDR) pathway in these models presents a significant challenge.

Purpose of the Study:

• To develop a fluorescent biosensor for detecting DDR activation in 3D tumor models.
• To enable monitoring of p53-dependent p21 promoter activation as a readout for DDR.

Main Methods:

• Development and application of the DDR-Act-FP fluorescent biosensor.
• Utilizing ATM kinase inhibitors for pharmacological manipulation of DDR.
• Screening a small compound library to identify DDR activators.
• Live monitoring of DDR activation in multicellular spheroids.

Main Results:

• DDR-Act-FP efficiently reports DDR pathway activation upon DNA damage and drug treatment.
• The biosensor successfully identified activators of the DDR response from a compound library.
• Live monitoring of DDR activation and its modulation was achieved in 3D spheroid models.

Conclusions:

• DDR-Act-FP is a valuable tool for monitoring DDR in 3D tumor models.
• This biosensor facilitates innovative screening and preclinical evaluation assays.
• The study demonstrates the potential for live monitoring of DDR in complex 3D environments.