Abstract 1240: Runx2 promotes microtubule stability for survival of breast cancer bone metastatic cells
Authors: Othman Ahmad , Winogradzki Marcus , Tandon Manish , Pratap Jitesh 
Published: September 2020

Bone metastasis of breast cancer causes significant patient mortality. During metastasis, breast cancer cells induce autophagy to survive metabolic stress. Although the core components of the autophagy pathway have been characterized, the regulatory mechanism of autophagy in bone metastatic cancer cells is still unknown. To examine autophagy during bone metastasis, we used a bone metastatic isogenic variant of breast cancer MDA-MB-231 cells isolated from a xenograft tumor mouse model of metastasis. Previously, we and others have shown that Runt-related transcription factor-2 (Runx2) promotes cell survival, cell invasion, and metastatic tumor growth associated osteolysis. Therefore, we examined the regulatory mechanisms of Runx2-mediated autophagy for increased cell survival in the bone microenvironment. Our results show that Runx2 promotes microtubule (MTs) stability to enhance autophagy flux. We show that Runx2 silencing significantly decreases acetylated α-Tubulin (Ac-α-Tub) levels. Ac-α-Tub is a marker of stable MTs, which promotes trafficking of autophagosomes. Ac-α-Tub also increases during cellular stress, such as glucose starvation. We found that Runx2 knockdown cells are able to induce Ac-α-Tub upon glucose starvation but are unable to maintain acetylated polymer mass of MTs upon removal of stress. Furthermore, expression of wild type or transcriptionally inactive Runx2 can restore Ac-polymer of MTs in Runx2 knockdown cells, while C-terminal deletion mutant failed to rescue MT polymer mass. Previous studies have shown that the C-terminal of Runx2 serves a scaffolding function by interacting with MTs and multiple proteins including HDAC6, the deacetylase of α-Tubulin. Interestingly, the expression levels of HDAC6 and αTAT1 (α-tubulin acetyltransferase-1) were not altered with Runx2 silencing, suggesting a transcriptionally independent function for Runx2 for maintaining the MTs stability. As MT targeting agents are often used as chemotherapeutics, we found that treatment with vinblastine, a MT disrupting agent, decreases Ac-polymer MTs with Runx2 silencing or expression of C-terminal mutant Runx2, while WT and transcriptionally inactive Runx2 expressing cells exhibit resistance to the treatment. Conversely, treatment with Docetaxel, a MT stabilizing agent, enhances Ac-α-Tub and polymer MTs. Recent reports have also linked high levels of Ac-α-Tub with aggressive breast cancer. We performed immunohistochemistry for Ac-α-Tub levels in bone metastatic patient samples and found significantly strong Ac-α-Tub positive cells in matched and unmatched bone metastatic tumors compared to the primary tumors. Our findings indicate that inhibition of Runx2 may sensitize metastatic tumors to MT targeting agents, and Runx2 and Ac-α-Tub levels may serve as markers for metastatic tumors to help stratify patients for optimal treatment for bone metastatic tumors.

Citation Format: Ahmad Othman, Marcus Winogradzki, Manish Tandon, Jitesh Pratap. Runx2 promotes microtubule stability for survival of breast cancer bone metastatic cells [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 1240.