Breast cancer is a complex and multifactorial disease, making their categorization an extremely demanding task, especially with regard to therapeutic response and disease progression. Understanding the biological basis of formation and tumor progression of breast cancer is very important for the development of new, more specific and effective therapeutic strategies. Experimental evidences indicate that the pro-apoptotic gene PAWR (PKC apoptosis WT1 regulator, also known as Par-4, Prostate Apoptosis Response-4) plays an important role in the selective induction of apoptosis in a wide variety of cancer cells. Par-4 acts both in intrinsic and extrinsic pathway of apoptosis and studies have shown that systemic expression of Par-4 confers resistance to the development of spontaneous tumors in mice. Furthermore, the recombinant Par-4 injected intravenously in mice inhibited tumor growth and metastasis. Previous data from our group demonstrated that reduced nuclear expression of Par-4 is associated with poor prognosis in breast cancer. However, little is known regarding the Par-4's role in tumorigenesis of breast cancer. In the present study, we sought to evaluate the effects of suppression of Par-4 in proliferation, survival, and chemosensitivity of breast cancer cells in vitro and in vivo. MDA-MB-231 breast cancer cells were transfected with different sequence of the pRS vector containing the shRNA to silencing the Par-4. Western Blot technique was used to characterize the clones silenced for Par-4. Using fluorescence-based microplate assay and flow cytometry we evaluated the effect of reduced expression of Par-4 in the rate of cell proliferation and cell cycle. In our results, no difference was observed in the proliferation rates and in the phases of cell cycle between MDA-MB-231 cells with reduced expression of Par-4 in comparison with the control cells. On the other hand, by flow cytometry we have found that decreased expression of Par-4 reduces the sensitivity of the MDA-MB-231 cells to treatment with chemotherapeutic agents as docetaxel and 5-Fluorouracil. Interestingly, by flow cytometry with Annexin and PI, we have observed a significant reduction in the percentage of late apoptosis in MDA-MB-231 cells with reduced expression of Par-4 in comparison with the control cells after 24 hours of treatment with docetaxel (19.50% and 32.64%, p<0.05). Furthermore, MDA-MB-231 cells with reduced expression of Par-4 also showed reduced activation of caspase-7. To confirm if the effect of reduced expression of Par-4 also extends to in vivo experiments, MDA-MB-231 cells transfected with silencing vector for Par-4 or the empty vector were injected into the left flank of Balb/c NUDE animals. Although preliminary, our data indicate that mice injected with breast cancer cells with reduced expression of Par-4 develop tumours twice as fast as mice injected with the control cells. Our findings could be of particular interest in to highlight the important role of Par-4 in breast tumour formation and in the modulation of chemiosensitivity.

Supported by FAPESP and CNPq.

Citation Format: Lourival A. Oliveira Filho, Maria José Carlini, Maria Aparecida Nagai. Role of cellular and extracellular Par-4 expression in tumor formation and drug sensitivity. [abstract]. In: Proceedings of the AACR Special Conference on Translational Control of Cancer: A New Frontier in Cancer Biology and Therapy; 2016 Oct 27-30; San Francisco, CA. Philadelphia (PA): AACR; Cancer Res 2017;77(6 Suppl):Abstract nr A42.