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Using High Content Imaging to Quantify Target Engagement in Adherent Cells
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Using High Content Imaging to Quantify Target Engagement in Adherent Cells

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A high content, high throughput cellular thermal stability assay for measuring drug-target engagement in living cells

Andrew J Massey1

  • 1Vernalis Research, Granta Park, Cambridge, United Kingdom.

Plos One
|April 5, 2018
PubMed

Insights

A new assay, high content imaging-based cellular thermal shift assay (HCIF-CETSA), measures drug target engagement in living cells. This method offers a robust, high-throughput approach for drug discovery research.

Area of Science:

  • Biochemistry
  • Pharmacology
  • Cell Biology

Background:

  • Drug target engagement is crucial for drug discovery but challenging to measure in living cells.
  • Existing methods often lack selectivity or high-throughput capabilities.

Purpose of the Study:

  • To develop and validate a novel, high-throughput method for measuring drug target engagement in living cells.
  • To assess the utility of the assay for drug discovery applications.

Main Methods:

  • Developed the high content imaging-based cellular thermal shift assay (HCIF-CETSA).
  • Utilized immunofluorescent detection of target protein levels after thermal treatment of adherent cells in a 96-well plate format.
  • Validated the assay using Chk1 target engagement by small molecule inhibitors.

Main Results:

  • HCIF-CETSA demonstrated robustness and correlated well with traditional CETSA for a selective Chk1 inhibitor.
  • The assay provides a valuable advancement for high-throughput target engagement determination in adherent cells.
  • Observed differences in EC50 values compared to autophosphorylation IC50 highlight assay-specific characteristics.

Conclusions:

  • HCIF-CETSA is a valuable new method for measuring drug target engagement in living cells.
  • The assay facilitates high-throughput screening and drug discovery efforts.
  • This technique enhances the ability to assess target engagement in a cellular context.

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