Protocol

Cell-Based Assays for High-Throughput Screening

Volume 486 of the series Methods in Molecular Biology pp 139-149

Date:

Assay for Isolation of Inhibitors of Her2-Kinase Expression

  • Gabriela ChiosisAffiliated withProgram in Molecular Pharmacology and Chemistry, Memorial Sloan-Kettering Cancer Center
  • , Adam B. KeetonAffiliated withSouthern Research Molecular Libraries Screening Center, Assay Implementation Group, Southern Research Institute

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Summary

Her2 (ErbB2) protein is overexpressed in breast and other solid tumors, and its expression is associated with progressive disease. Current therapies directed toward Her2 either block dimerization of the receptor or inhibit tyrosine kinase activity to disrupt intracellular signaling. However, little is known about alternative mechanisms for suppressing Her2 expression, possibly by inducing degradation or blocking synthesis. Here, we describe a hybrid western-blotting and enzyme-linked immunosorbent assay (ELISA) designed to identify in low- to medium-throughput format noncytotoxic compounds that reduce expression of Her2 protein.

Key words

Breast cancer Heat-shock protein 90 (Hsp90) Her2 (ErbB2) tyrosine kinase Whole-cell immunoblot