urmc-099 and Breast-Neoplasms

Mixed lineage kinase 3 (MLK3), also known as MAP3K11, was initially identified in a megakaryocytic cell line and is an emerging therapeutic target in cancer, yet its role in immune cells is not known. Here, we report that loss or pharmacological inhibition of MLK3 promotes activation and cytotoxicity of T cells. MLK3 is abundantly expressed in T cells, and its loss alters serum chemokines, cytokines, and CD28 protein expression on T cells and its subsets. MLK3 loss or pharmacological inhibition induces activation of T cells in in vitro, ex vivo, and in vivo conditions, irrespective of T cell activating agents. Conversely, overexpression of MLK3 decreases T cell activation. Mechanistically, loss or inhibition of MLK3 down-regulates expression of a prolyl-isomerase, Ppia, which is directly phosphorylated by MLK3 to increase its isomerase activity. Moreover, MLK3 also phosphorylates nuclear factor of activated T cells 1 (NFATc1) and regulates its nuclear translocation via interaction with Ppia, and this regulates T cell effector function. In an immune-competent mouse model of breast cancer, MLK3 inhibitor increases Granzyme B-positive CD8

Topics: Animals; Breast Neoplasms; Cell Line, Tumor; Cyclophilin A; Female; Humans; Lymphocyte Activation; Lymphocytes, Tumor-Infiltrating; Mammary Neoplasms, Experimental; MAP Kinase Kinase Kinases; Mice; Mitogen-Activated Protein Kinase Kinase Kinase 11; NFATC Transcription Factors; Phosphorylation; Primary Cell Culture; Protein Kinase Inhibitors; Pyridines; Pyrroles; T-Lymphocytes, Cytotoxic; Tumor Escape

Brain metastasis of breast cancer is an important clinical problem, with few therapeutic options and a poor prognosis. Recent data have implicated mixed lineage kinase 3 (MLK3) in controlling the in vitro migratory capacity of breast cancer cells, as well as the metastasis of MDA-MB-231 breast cancer cells from the mammary fat pad to distant lymph nodes in a mouse xenograft model. We therefore set out to test whether MLK3 plays a role in brain metastasis of breast cancer cells. To address this question, we used a novel, brain penetrant, MLK3 inhibitor, URMC099. URMC099 efficiently inhibited the migration of breast cancer cells in an in vitro cell monolayer wounding assay, and an in vitro transwell migration assay, but had no effect on in vitro cell growth. We also tested the effect of URMC099 on tumor formation in a mouse xenograft model of breast cancer brain metastasis. This analysis showed that URMC099 had no effect on the either the frequency or size of breast cancer brain metastases. We conclude that pharmacologic inhibition of MLK3 by URMC099 can reduce the in vitro migratory capacity of breast cancer cells, but that it has no effect on either the frequency or size of breast cancer brain metastases, in a mouse xenograft model.

Topics: Animals; Brain Neoplasms; Breast Neoplasms; Cell Line, Tumor; Cell Movement; Disease Models, Animal; Female; MAP Kinase Kinase Kinases; Mice; Mice, Nude; Mitogen-Activated Protein Kinase Kinase Kinase 11; Neoplasm Transplantation; Protein Kinase Inhibitors; Pyridines; Pyrroles; Transplantation, Heterologous