Small Molecule Inhibitor Targeting CDT1/Geminin Protein Complex Promotes DNA Damage and Cell Death in Cancer Cells

Area of Science:

• Molecular Biology
• Cell Biology
• Biochemistry

Background:

• DNA replication initiation relies on MCM2-7 complex loading during G1.
• Replication licensing ensures proper DNA replication, preventing genomic instability.
• CDT1 is vital for licensing, with Geminin inhibiting its activity.

Purpose of the Study:

• To develop a high-throughput screening assay to find inhibitors of the CDT1/Geminin complex.
• To characterize the identified inhibitors for their potential in cancer research.

Main Methods:

• Developed an in vitro AlphaScreen™ high-throughput screening assay.
• Biochemical characterization of the lead compound AF615.
• Tested AF615's effects on DNA damage, synthesis, and cancer cell viability.

Main Results:

• Identified AF615 as a potent inhibitor of Geminin binding to CDT1.
• AF615 demonstrated dose-dependent inhibition in vitro and in cells.
• AF615 selectively induced DNA damage, inhibited DNA synthesis, and reduced viability in cancer cell lines, dependent on CDT1.

Conclusions:

• AF615 specifically inhibits the CDT1/Geminin complex.
• AF615 shows potential as a tool to study replication licensing and origin firing.
• AF615 serves as a scaffold for developing novel cancer therapeutics.