A Molecular Approach to Dissecting MEK-Mediated Phosphorylation in Pediatric Leukemia

SUMMARY Acute lymphoblastic leukemia (ALL) is the most common hematological malignancy and the leading cause of cancer-related death in children in the United States. Multi-drug chemotherapy has vastly improved 5-year event- free survival by over 80%; however, the prognosis for children with refractory or relapsed disease is less positive, with complete remission rates dropping to 30%. Mitogen-activated protein kinase (MAPK) signaling pathways are conserved across eukaryotes from yeast to humans. These enzymes play a central role in regulating cellular activities from survival and proliferation to stress response, differentiation, motility, and angiogenesis. MAPK pathways couple diverse extracellular signals (growth factors, hormones, cytokines, and environmental stresses) to distinct intracellular gene programs via a series of activating phosphorylation events. Providing direct input to these kinases are the MAPK/ERK kinase (MEK) family enzymes, also called mitogen-activated protein kinase kinases (MAP2K or MKKs). Currently, there are four FDA-approved MEK inhibitors which block inputs to the ERK1/2 arm of the pathway, all of which inhibit MEK1/2. Loss of MAP2K7 (a.k.a. MEK7) repression and consequent amplification of downstream mitogen-activated protein kinase (MAPK) signaling is thought to contribute to T-cell acute lymphoblastic leukemia (T-ALL) pathology. Currently, the only known function of MEK7 is the phosphorylation of JNK, its downstream signaling partner in the MAP kinase signaling cascade. However, its structurally and functionally homologous isoform MEK4 also functions to phosphorylate this downstream kinase. The relative inputs of MEK4 and MEK7 seem to be variable and context dependent. This gap in the knowledge is currently a major impediment to fully understanding MEK signaling pathways in the context of cancer. Although both MEK4 and MEK7 activate JNK, only MEK7 appears to mediate JNK-dependent inflammatory responses in immune cells. Likewise, it is possible that inhibition of MEK7 in the context of T-ALL may lead to compensatory upregulation of MEK4-promoted JNK phosphorylation. Equipped with highly selective MEK7 inhibitors, we are uniquely enabled to dissect this critical kinase cascade for the first time. Herein, we propose the further development of new molecular tools to attenuate MEK7-mediated JNK phosphorylation. These new probe compounds will then be applied to the cellular dissection of the JNK arm of the MAP kinase signaling cascade in the context of T-ALL. Clinically relevant mouse models will then be employed to identify the optimal drug target(s) in T-ALL.