The Role of PIAS1 in Cancer Radiotherapy Response

Prostate- and breast cancer are the most frequent malignancies and among the leading causes of cancer related deaths in the Western world in men and women, respectively. Besides surgery or anti- hormonal therapy, radiation is frequently applied for patients suffering from either tumor. Radiotherapy aims to induce potentially lethal DNA damage to tumor cells. However, in many cases treatment success is limited by the occurrence of resistances, mostly via optimization of DNA repair mechanisms. Thus, combined therapy approaches designed to target the tumor at multiple sites or to increase the sensitivity to standard therapies may be of tremendous benefit compared to single treatments. PIAS comprise a family of 4 multifunctional proteins which can act as transcriptional co-regulators or SUMO E3 ligases. Importantly, PIAS have been shown to support successful DNA repair, mostly via orchestrated SUMOylation of involved proteins. In previous studies, the applicant demonstrated that PIAS1 expression increases with prostate cancer malignancy and induces cell proliferation via downregulation of the cell cycle inhibitor p21. PIAS1 knockdown leads to p21-mediated cell cycle arrest, thereby attenuating proliferation. Preliminary experiments indicate a similar role for PIAS1 in breast cancer. It is thus hypothesized that (A) PIAS1 knockdown might render cells more sensitive to irradiation, thus being a promising strategy for a combined therapy approach (B) PIAS1 is involved in mechanisms of radioresistance and (C) PIAS1 expression level is predictive for patient survival and therapy outcome. To test these hypotheses, the applicant aims to: (A) Assess the additive effect of PIAS1 knockdown by multiple approaches in combination with radiation in prostate- and breast cancer cells in vitro and in CAM xenografts, and clarify the underlying mechanisms. (B) Evaluate the expression and functional role of PIAS1 radioresistant prostate- and breast cancer cells and assess whether these resistant cells can be re-sensitized to apoptosis by PIAS1 depletion. (C) Clarify whether PIAS1 expression in patient tissue may serve as a prognostic marker for patient survival and response to radiotherapy, employing tissue microarrays from two independent prostate- and breast cancer patient cohorts. Results obtained from these experiments might identify PIAS1 as novel therapeutic target in combination with irradiation and furthermore as a possible marker to improve biopsy diagnostics. This will significantly improve the management of prostate- and breast cancer and may in addition be of value for cancer research in general. The applicant was the first to describe increased PIAS1 expression and an oncogenic role for PIAS1 in prostate cancer and now aims to extend experiments to a translational setting in order to achieve clinical relevance of the findings. The Hertha Firnberg position will be invaluable for the applicant to increase her scientific and personal skills in order to further establish in the field of endocrine related cancers. 1

Radiotherapy aims to kill tumor cells by inducing lethal DNA damage which results in tumor shrinking. Unfortunately, many patients experience disease recurrence after radiotherapy because some tumor cells learn to repair the DNA damage induced by irradiation, survive and consequently build new metastases. Therefore, combined therapies designed to increase the efficiency of irradiation are urgently needed. We previously showed that the protein PIAS1 is increased in tumor tissue of prostate cancer patients and exerts a growth- promoting function. Interestingly however, PIAS1 has also been involved in cellular mechanisms that are required for repair of DNA damage. Within this project we aimed to (a) further extend the knowledge about the function of PIAS1 to breast cancer, another tumor type which shares several characteristics with prostate cancer and (b) test if a knockdown of PIAS1 might increase the efficiency of irradiation in prostate- and breast cancer. First, we assessed the expression of PIAS1 in benign and malignant tissue samples of breast cancer patients. PIAS1 levels were significantly higher in cancerous tissue as well as in lymph node metastases, when compared to benign specimens. A knockdown of PIAS1 diminished growth and viability of breast cancer cells from different molecular subtypes, indicating that PIAS1 exerts a similar function on cell growth in breast cancer as in prostate cancer. Furthermore, several experiments indicated that PIAS1 knockdown impairs DNA repair of tumor cells. Therefore, we tested whether a knockdown of PIAS1 prior to irradiation might increase the effects of radiotherapy. Our studies demonstrated that both single treatments (PIAS1 knockdown; 5 gray of irradiation in a linear accelerator) only slightly diminish the cell number in 2D as well as 3D culturing systems. Strikingly however, combination therapy of PIAS1 knockdown and irradiation led to significantly elevated cell death by induction of severe DNA damage. Taken together, from this project it can be concluded that the growth promoting role of PIAS1 is not limited to prostate cancer but also true for breast cancer. Thus, it will be interesting to analyze the function of PIAS1 also in other tumor entities. Moreover, we could demonstrate that a knockdown of PIAS1 reduces the growth of prostate and breast cancer cells already when applied as single treatment and that this effect is even enhanced when combined with irradiation. It would be thus worth to develop strategies which allow for knockdown of PIAS1 within a patient`s tumor. Finally, findings on the implication of PIAS1 in DNA repair might be of interest for researchers working on other diseases that are characterized by disturbed DNA repair.