The Function of Cyclin D1 in DNA Repair: A New Possible Approach to Increase Sensitivity of Ovarian Cancer Cells to Irradiation

Cyclin D1 has been recently investigated as an important protein in homologous recombination-directed repair (HDR) mechanism. Cyclin D1 is recruited to damaged foci by BRCA2 and it enhances RAD51 binding to BRCA2. Furthermore, cyclin D1 is essential for RAD51 gene up-regulation in case of double-stranded break (DSB). Meanwhile molecule 3l, an E-3-(2-chloro-3-indolylmethylene)1,3-dihydroindol-2one, has been showed to reduce cyclin D1 level and, for this reason, 3l treatment could be used to study cyclin D1 role in DNA damage repair. Scope of this thesis is to find a new possible combined approach to increase sensitivity of ovarian cancer IGROV1 cells to irradiation: 3l treatment for 24 hours in order to deplete cyclin D1 expression, with a further UV irradiation to induce DNA damages. In this case, the repair of induced DSBs are analyzed in cells with a small amount of cyclin D1. Results show that UV irradiation induces histone H2AX hyperphosphorylation on Ser139 (γH2AX), marker of DSBs, and down-regulation of cyclin D1 in all the samples. UV-irradiated cells result in an up-regulation of RAD51 gene and formation of repair complex BRCA2-cyclin D1-RAD51 on damaged foci. 3l-treated cells present a depletion of cyclin D1 protein level and irradiation of these cells induces γH2AX. However, 3l-treated and then UV-irradiated cells do not present up-regulation of RAD51 gene and repair proteins colocalization. Anti-CCND1 siRNA silenced cells show comparable results to 3l-treated cells, while scramble siRNA silenced cells present comparable results to control cells. These results confirm the important role of protein cyclin D1 in the repair pathway of DSBs. The combined treatment of a drug leading to a depletion of cyclin D1 and a DNA damage inducer, such as irradiation, could be a potent potential therapy to target cancers with an elevated rate of DNA repair, especially in case of chemo- and radio-resistant tumours.