(a: Introduction) Chemotherapy is standard procedure for cancer treatment to eradicate small group of cancer cells that escaped surgical removal, or to control tumor growth. In contrast to radiation therapy and surgery, which treat localized tumors, chemotherapy drugs treat whole body. Since normal cells will be exposed to the drug as well, side effect is major problem of chemotherapy. Patients on chemotherapy commonly experience nausea, vomiting and hair loss. Large dose of drug may leave damage on normal cells, which may result in secondary tumor years later. Clinical studies have demonstrated that treatment of malignant germ cell tumor with topoisomerase poisons such as etoposide increases the rate of secondary leukemia. By developing a method that enhances the effect of chemotherapy, we would require less drug dose and it may help to reduce chemotherapy side effects. (b: experimental procedures) A class of chemotherapy drugs, spindle poisons (e.g. taxol, vinblastine), inhibits mitotic spindle function, activates the mitotic spindle checkpoint, arrests cells in mitosis, and then causes cell death by mechanisms that are poorly understood. To investigate the signal transduction pathways involved in modulating these processes and to isolate factors that serve to sensitize cells to spindle poisons, we developed a cDNA library expression cloning method that identifies genes whose expression increases the substrate attachment of cells arrested in M phase with spindle poisons, due to mitotic exit and/or mitotic apoptosis. (c: summary of results) Here we identify and validate seven cDNA’s from the screen that induce spindle poison sensitivity. We then characterize HeLa cell lines that stably express two of the cDNA’s. pSC3 encodes a fragment of human S8/p45/TRIP1/Sug1, a component of the 19S proteasome regulatory complex. Stable expression of pSC3 decreased the proteasome activity and increased the levels of apoptosis in cells treated with spindle poisons or proteasome inhibitors. Cells stably transfected with pSC3 exhibited decreased expression of BubR1, a kinase required for activation of the mitotic checkpoint in response to treatment with spindle poisons. pSC29 encodes the bromodomain from thyroid hormone receptor coactivator isoform 2. Cells stably transfected with pSC29 also exhibited significantly decreased expression of BubR1. Thus these cDNA fragments both appear to affect stability of a spindle checkpoint component. (d: conclusions) The screening method described provides a general approach for revealing genes and gene fragments whose expression modulates cellular responses to spindle poisons. These genes may be a target for development of drug used in combination with spindle poisons.
[Proc Amer Assoc Cancer Res, Volume 46, 2005]