A. Maiti1, M. Oshi1, N. Hait1, A. Maiti1 1Roswell Park Cancer Institute,Surgical Oncology/ Molecular And Cellular Biology,Buffalo, NY, USA
Introduction: In most malignant solid breast tumors cancer cells are subjected to severe micro-environmental stresses including low nutrients and oxygen. These unfavourable conditions may also enhance cancer metastasis to distant organs. Brain-selective kinase 2 (BRSK2, also known as SAD-A) is a member of the AMP-activated protein kinase subfamily known to express in brain and pancreas. BRSK2 identified as genes essential for neuronal polarization. Recently it was identified to be induced by nutrient deprivation. We identified that BRSK2 expression in ER + PR- HER2- human breast cancer metastasized to Brain tissue compare to triple-negative breast cancer and control breast tissue. To understand the role of BRSK2 in breast cancer metastasis, we overexpressed this gene in different human cancer cells and studied extensively using in vitro culture condition
Methods: We extracted protein from triple-negative and ER+ breast cancer metastasized to brain tissue along with control breast tissue specimens. We overexpressed BRSK2 in BT474, MDA-MB231 and brain metastatic version of MDA-MB231 (231Br), SKBR3. After 24 hours of transfections with BRSK2 plasmid, we allow cells to grow under stress conditions for another 24 hours. Total protein and RNA were extracted and used for further molecular analysis.
Results:Western blot analysis demonstrated that human breast cancer metastasized to brain tissue showed increased expression of BRSK2 compared to control and triple-negative human breast cancer tissue. We implanted these human breasts to brain metastatic tissues in NSG mice breast fat pad and brain and allowed to grow. We observed that human tumors when grown in the brain, have an expression of BRSK2 while the same tissues grown in mice breast have no expression. We, therefore, hypothesize that BRSK2 may be epigenetically downregulated in breast cancer. As expected, our data revealed that HDAC inhibitors TSA and SAHA re-expressed BRSK2 in breast cancer cells compared to the vehicle-treated cells. To uncover further, we overexpressed BRSK2 in MDA-MB231, BT474 cancer cells followed by exposure to glucose and nutrient deprivation condition. Our data revealed that protective autophagy marker LC3 A/B increased when BRSK2 overexpressed in cancer cells. LC3 A/B expression remains elevated under glucose and nutrient stress conditions compared to control vector transfected cells under same condition. Cytokines and growth factors are key players in the breast cancer metastasis induction. Our data also explained that BRSK2 overexpression causes increased expression of several cytokines like CXCL1, IL-1beta, LOXL1, IFNg in breast cancer cells and media compared to control cells
Conclusion:BRSK2 is epigenetically remain silenced in normal conditions. Breast Cancer cells overexpressed BRSK2 to survive the nutrient stress condition through protective autophagy and cytokine signaling, which may also enhance metastasis to the brain.