Abstract C42: MLKs regulate the activities of the small GTPases, Rho and Rac, to drive both single cell and collective cell migration in breast cancer invasion

Abstract

Abstract Most mortality of breast cancer patients is caused by metastasis to vital organs. One of the key steps of breast cancer metastasis is invasion through the basement membrane, which is controlled by several signaling cascades including small GTPases, Rho and Rac. Mixed lineage kinases (MLKs) are a family of MAP3Ks that activate multiple MAPK pathways, including the c-Jun N-terminal kinase (JNK) pathway. Our lab has determined that MLK3-JNK signaling is critical for migration, invasion and metastasis of triple negative breast cancer cells. Herein, the effect of a small molecule pan-MLK inhibitor on tumor cell migration in 2D wound healing and in 3D dissemination in collagen type I, the most abundant collagen in the body, was determined. Cancer cells can utilize distinct modes of migration. MDA-MB-231 and 4T1 cells were chosen as models for the study of single and collective cell migration, respectively. In addition, the effect of MLK3 on Rho-GTP and Rac-GTP levels was determined. Data from this study demonstrate that an MLK inhibitor is sufficient to block migration of multiple breast cancer cell lines in wound healing assays. Furthermore, this inhibitor dramatically reduces the dissemination of cancer cell lines that utilize single, as well as, collective cell migration. Silencing of MLK3 results in enhanced Rho activity and decreased Rac activity, as well as defective focal adhesion turnover. Inhibition of the activity of MLKs or the downstream MAPK, JNK, also increases Rho activity, which is accompanied by an increase in the number and/or size of focal adhesions and an increase in actin stress fibers. Consistent with the idea that MLK inhibition reduces migration through relieving Rho suppression, addition of a Rho/ROCK inhibitor, Y-27632, partially rescues the migratory defect of MLK inhibition by reducing peripheral focal adhesions and actin stress fibers. Taken together, these data suggest that MLK3 could play a role in controlling the balance between Rac and Rho activity required for cancer cell migration and invasion. Thus, MLK inhibitors could be therapeutically useful in the context of breast cancer metastasis. Citation Format: Chotirat Rattanasinchai, Sean Misek, Jian Chen, Kathleen A. Gallo. MLKs regulate the activities of the small GTPases, Rho and Rac, to drive both single cell and collective cell migration in breast cancer invasion. [abstract]. In: Proceedings of the AACR Special Conference on Tumor Invasion and Metastasis; Jan 20-23, 2013; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2013;73(3 Suppl):Abstract nr C42.