Knock-in reporter constructs and drug screening for compounds to restore RKIP expression in cancer.

RKIP (Raf Kinaes Inhibitory Protein) is ubiquitously expressed in almost all normal tissues of metazoans. Consistent with its negative regulatory role in cell proliferation and survival, RKIP expression is progressively downregulated in cancer accompanied with a worse prognosis and disease outcome. Experiments with cancer cell lines and genetically engineered mice have demonstrated that the expression level of RKIP is a driving factor in determining the disease outcome of cancer, and that restoring RKIP expression is a promising option for therapeutic treatment of low-RKIP-expressing cancers. RKIP expression is mainly regulated at the transcription level such that its expression is tuned down, but not shut off, and is poised to be reactivated. In this study we developed a RKIP gene promoter knock-in reporter breast cancer cell line to screen for compounds that can regulate RKIP transcription in cancer. Using this modified cell line, we were able to identify eight FDA approved compounds that increase RKIP promoter activity by a minimum of 2-fold and can potentially be re-purposed for therapeutic treatment of low-RKIP-expressing breast cancer.