SIGNIFICANCE OF P27 IN GROWTH AND RESPONSE TO THERAPY OF LUMINAL BREAST CANCER

The CDK inhibitor p27kip1 is frequently down modulated in breast cancer (BC) and mutations in its gene, CDKN1B, have been recently identified as driver genetic lesions, almost exclusively in luminal BC (LBC). Clinical studies have recognized p27kip1 expression as predictive of outcome for LBC patients. Resistance to therapy and disease relapse are unfortunate events that occurs frequently, especially in patients carrying the luminal B subtype of BC. We thus hypothesized that elucidating the role of p27kip1 and its mutations in LBC progression and response to treatments could contribute to ameliorate the clinical management of this disease. To this aim, we genetically manipulated the MCF-7 luminal B BC cell line to Knock-Out p27Kip1 gene and Knock-In p27Kip1 mutants, exploiting the Zinc Finger Nucleases technology. We explored the impact of modified expression p27Kip1 on MCF-7 cell behavior in different experimental settings. Tumorigenic and stem-like features were increased by loss of p27Kip1 and by expression of the mutants. Next, we tested the response of these clones to radiotherapy, which represents an integral part of the gold standard treatment for LBC patients. We irradiated MCF-7 clones and evaluated their survival rates by clonogenic assay. Our data clearly demonstrate that loss of p27Kip1 conferred radioresistance to MCF-7 cells. In particular, analyses of H2AX foci and aberrant mitoses highlighted that loss of p27Kip1 affected DNA damage repairing ability and significantly increase the presence of mitotic defects in surviving cells. In conclusion, during my PhD thesis we have generated tools for studying the role of p27Kip1 in LBC, i.e. MCF-7 luminal B BC cells Knock-Out for p27Kip1 and Knock-In for mutants that have been described in human LBC patients. Moreover, we have observed that loss p27Kip1 conferred augmented tumorigenicity, self-renewal potential and radioresistance to luminal B BC cells, suggesting a critical role for p27Kip1 in the maintenance of genomic stability.